Fermentative hydrogen and methane productions using membrane bioreactors Julius Gbenga Akinbomi

Fermentative hydrogen and methane productions using membrane bioreactors Julius Gbenga Akinbomi
Thesis for the Degree of Doctor of Philosophy
Fermentative hydrogen and methane productions using membrane
bioreactors
Julius Gbenga Akinbomi
Copyright© Julius Akinbomi
Swedish Centre for Resource Recovery
University of Borås (Sweden)
ISBN 978-91-87525-73-5 (printed)
ISBN 978-91-87525–74-2 (pdf)
ISSN 0280-381X Skrifter från Högskolan i Borås, nr. 72
Digital version:
http://urn.kb.se/resolve?urn=urn:nbn:se:hb:diva-671
Printed in Sweden by Responstryck AB,
Borås, 2015
ii
Abstract
The role of energy as a stimulant for economic growth and environmental sustainability
of any nation has made the focus on green fuels, including fermentative hydrogen (bioH2) and
methane (bioCH4), to be a priority for the World’s policy makers. Nigeria, as the most populous
African country, with worsening energy crisis, can benefit from the introduction of the bioH2 and
bioCH4 technologies into the country’s energy mix, since such technologies have the potential of
generating energy from organic wastes such as fruit waste.
Fruit waste was studied in detail in this work because of its great economic and
environmental potential, as large quantities of the wastes (10–65% of raw fruit) are generated
from fruit consumption and processing. Meanwhile, bioH2 and bioCH4 productions involving
anaerobic microorganisms in direct contact with organic wastes have been observed to result in
substrate and product inhibitions, which reduce the gas yields and limit the application of the
technologies on an industrial scale. For example, in this study, the first experimental work to
determine the effects of hydraulic retention times and fruit mixing on bioH2 production from
single and mixed fruits revealed the highest cumulative bioH2 yield to be equivalent to 30% of
the theoretical yield. However, combining the fermentation process with the application of
membrane encapsulated cells and membrane separation techniques, respectively, could reduce
substrate and product inhibitions of the microorganisms. This study, therefore, focused on the
application of membrane techniques to enhance the yields of bioH2 and bioCH4 productions from
the organic wastes.
The second experimental work which focused on reduction of substrate inhibition,
involved the investigation of the effects of the PVDF membrane encapsulation techniques on the
bioH2 and bioCH4 productions from nutrient media with limonene, myrcene, octanol and hexanal
as fruit flavours. The results showed that membrane encapsulated cells produced bioCH4 faster
and lasted longer, compared to free cells in limonene. Also, about 60% membrane protective
effect against myrcene, octanol and hexanal inhibitions was obtained. Regarding bioH2
production, membrane encapsulated cells, compared to free cells, produced higher average daily
yields of 94, 30 and 77% with hexanal, myrcene and octanol as flavours, respectively. The final
part of the study, which was aimed at reducing product inhibition, involved the study of the
effects of membrane permeation of volatile fatty acids (VFAs) on the bioreactor hydrodynamics
in relation to bioH2 production. The investigation revealed that low transmembrane pressure of
104Pa was required to achieve a 3L h-1m-2 critical flux with reversible fouling mainly due to cake
layer formation, and bioH2 production was also observed to restart after VFAs removal.
The results from this study suggest that membrane-based techniques could improve bioH2
and bioCH4 productions from fermentation media with substrate and product inhibitions.
.
Keywords: Encapsulation, Inhibition, hydrodynamics, hydrogen, methane, fruit flavour,
Membrane bioreactor
iii
List of Publications
The thesis is founded on the results presented in the following articles:
Paper I:
Youngsukkasem, S., Akinbomi, J., Rakshit, S.K., Taherzadeh, M.J. (2013).
Biogas production by encased bacteria in synthetic membranes: protective effects
in toxic media and high loading rates. Environmental Technology 34, 2077-2084.
Paper II:
Akinbomi, J and Taherzadeh, M.J. (2015). Evaluation of Fermentative Hydrogen
Production from Single and Mixed Fruit Wastes. Energies 8(5), 4253-4272.
Paper III:
Trad, Z., Akinbomi, J., Vial, C., Larroche, C., Taherzadeh, M.J., Fontaine, J-P.
(2015). Development of a submerged anaerobic membrane bioreactor for
concurrent extraction of volatile fatty acids and biohydrogen production.
Bioresource technology, 196, 290-300
Paper IV:
Akinbomi, J., Wikandari, R., Taherzadeh, M.J. Enhanced fermentative hydrogen
and methane productions from inhibitory-fruit flavour medium with membraneencapsulated cells (submitted)
Paper V:
Akinbomi, J., Brandberg, T., Sanni, S.A., Taherzadeh, M.J. (2014). Development
and dissemination strategies for accelerating biogas production in Nigeria.
BioResources 9, 5707-5737.
Paper VI:
Ylitervo, P., Akinbomi, J and Taherzadeh, M.J. (2013). Membrane bioreactors
potential for ethanol and biogas production: A review. Environmental
Technology 34, 1711-1723
Statement of Contributions
Julius Akinbomi’s contributions to each of the above publications are:
Paper I:
Responsible for part of the experimental work, data analyses and manuscript writing
Paper II:
Responsible for the experimental work, data analyses and manuscript writing
Paper III: Responsible for part of the experimental work, data analyses and manuscript writing
Paper IV: Conceived the idea together with the co-authors as well as responsible for the
experimental work, data analyses and manuscript writing
Paper V:
Responsible for the literature survey, data collection and manuscript writing
Paper VI: Responsible for part of the literature survey, data collection and manuscript writing
iv
Reflection on My PhD Journey
Having gone through a challenging, but exciting and rewarding PhD experience, I thought it
was good to reflect on my journey and write briefly on it.
My motivation for coming to Sweden for PhD study in Resource Recovery
The journey of the PhD programme began in Nigeria with the aim of working at the frontier
of biogas technology in Nigeria. During the period, Nigeria was really in need of affordable
technology that could guarantee stable power supply to her population. Besides, the country was
also seeking strategies to efficiently manage the huge amount of waste that was inevitably being
turned out on a daily basis through the activities of her teeming population. Having read about
how electricity could be generated through biogas from organic wastes, and how by adopting the
green fuel technology, the increasing energy demands of the growing world population could be
met without exhausting the natural resources and polluting the environment, I was motivated to
choose biogas as a course of study.
My dream as a teacher in Lagos State University (LASU), Lagos, Nigeria
As a teacher in one of the Universities in Nigeria, Lagos State University (LASU), I aspired
to be among the professional teachers that would help in improving the quality of teaching and
learning in Nigeria by influencing innovative and entrepreneurial students who would be able to
put Nigeria and the World at large on a sustainable footing. Knowing fully well that teachers are
employed not just to teach but to teach thoroughly at a high professional standard and that good
teaching is often informed by good research, I made plans to start my PhD study immediately
after my Master’s degree programme in Chemical Engineering. Few months before the start of
my PhD programme in Faculty of Engineering, LASU, Nigeria, Professor Mohammad
Taherzadeh and Dr Kayode Adekunle came to LASU to enlighten us on the resource recovery
programme going on at University of Borås, Sweden, and share with us how Borås as a city uses
biogas for vehicle fuel and combined heat and power (CHP). Not long after the seminar
presentation, four PhD students including me were informed of the opportunity of going to
Sweden for PhD study in Resource Recovery through student exchange programme between
Nigeria and Sweden. When I heard about it, - ‘it was like a dream come true’
v
The first year of my PhD study
The journey from Nigeria to Sweden was such an eventful one, as I was not alone but with
three other PhD students. We got to Sweden during winter period; it was my first time of seeing
snow falling as I had never experienced it in Nigeria. I was not used to the very cold weather at
the beginning so it was really tough for me to adapt to the cold weather. But afterwards, I got
acclimated to the varying weather conditions.
My PhD programme in Sweden, which focused on biohydrogen and biogas production from
organic wastes and agricultural residues, was supervised by a full-fledged Professor in Resource
Recovery, Mohammad Taherzadeh. My first year was quite busy as the course load was intense,
and I also had to conduct some experiments in the laboratory in order to get acquainted with the
equipment as well as the safety measures involved while using the equipment. However, it was
worthwhile to devote much time to the various lectures, presentations and laboratory work.
The second year of my PhD study
During the second year of the PhD programme, I went to Nigeria with the aim of setting-up a
mini biogas laboratory unit in my home country University for easy cross-border green fuel
technology. However, due to the reorganisation that was going on during the period, the project
was not feasible. Nonetheless, it is anticipated that in the nearest future, the project will be
feasible as it could be a launch pad for the development and dissemination of green fuel
technology in Nigeria. Meanwhile, my stay in Nigeria was not an idle period as I was busy
writing some articles together with some PhD students on the experiments carried out the
previous year.
The third year of my PhD study
During the third year of the PhD studies, I was in France at Blaise Pascal University,
Clermont Ferrand, for a six-month student exchange programme. The research in France was
carried out in collaboration with a PhD student, Zaineb Trad, under the supervision of Professor
Christophe Vial and Professor Christian Larroche. The aim of the research was to develop an
innovative anaerobic membrane bioreactor (AnMBR) with combined benefits of external and
immersed AnMBRs for simultaneous production of biohydrogen and volatile fatty acids (VFAs).
The bioreactor did not only allow VFAs to be removed for further applications with minimal
vi
modification to the hydrodynamics, but also prevented the inhibition of biohydrogen production
by total VFAs.
Beyond the boundary of academic activities, my short duration in France gave me the
opportunity to learn about the French culture, including its geography, history, religion, food,
among others. One thing I find common to most countries in Europe is the importance they
attach to their indigenous languages. Unlike in Sweden where people are not reluctant to speak
English to you, French people are not enthusiastic about speaking English with foreigners.
Although I know how difficult it is for students to have the mastery of foreign languages during
the beginning of their carriers in foreign countries, the language barrier actually motivates
foreign students to be determined to learn the languages of their host countries so that it will help
them in interpersonal interactions.
The final period of my PhD study
The remaining period of the PhD programme was spent in Sweden for the completion of
other relevant research work.
Looking back, the PhD experience has been, though challenging, a fulfilling one for me! It is
true that PhD journey is not a bed of roses and never a straight forward one. There is hardly any
research without something going wrong at one stage or another. However, according to an
adage that says ‘Life is 10% of what happened to us and 90% of how we react to it’, it is the
attitude of the researcher that determines the eventual success or failure of the research work.
Consequently, the PhD experience has taught me many lessons, including:
•
being able to bring creative ideas into realisation
•
ability to learn new skills and expertise required for the research and cope with
difficulties encountered during the research, and
•
being able to take criticism and turn research failure into success
-
One thing is certain; the PhD study has been a pleasant and exciting one for me!
vii
Acknowledgements
First of all, I want to express my profound gratitude to my main supervisor, Professor
Mohammad Taherzadeh for his guidance, thorough supervision, constructive criticisms, expert
advice, encouragement and support. He took time out of his extremely busy schedule to read my
manuscripts and dissertation as well as bringing to my attention the details that needed to be
addressed. In fact, the success of this research is due, in no small measure, to the support I
received from him. Words cannot really express how grateful I am to you Sir. I am also grateful
to Dr. Tomas Brandberg for his assistance during my PhD studies.
I am thankful to Lagos State University, Nigeria and University of Borås, Sweden for
giving me the opportunity and support to have my PhD studies in Sweden. I also want to show
my appreciation to Professor S.A. Sanni and Dr. Kayode Adekunle who initiated the
collaboration between the two Universities. My special thanks go to my examiner, Professor
Kim Bolton and Dr Päivi Ylitervo for reading and making useful suggestions for the final draft
of this thesis. Regarding my colleagues in Swedish Centre for Resource Recovery and other
departments at the University of Borås as well as in France (Blaise Pascal University), I have had
the privilege on different occasions of meeting many intelligent Post-doc, PhD and Master’s
students since the inception of my PhD programme in Resource Recovery in 2011. Although
they are too numerous to name here, I appreciate their contributions to the success of my study.
To all the members of staff at University of Borås, I am grateful for the conducive research
environment that was made available to me. I am also indebted to my two supervisors when I
was in France, Professor C. Larroche and Professor C. Vial, as well as the PhD student, Zaineb.
My special thanks go to my darling wife, Dayo, for her patience and support, and also
for taking good care of our princess, Feyi, during my absence. I am also grateful to my in-laws.
And to my precious mother and late father of blessed memory; you are my role models. You
taught me from my childhood to be good, responsible, strong, determined and prepared to pursue
and achieve my lifetime goals. According to one of your sayings, ‘if there is a will there will be
a way’- I will forever be grateful to you. To all my big sisters and brothers, I appreciate you all
for your endless love, prayers, supports and encouragements. I am indeed grateful.
---To my God-the source of my strength- THANK YOU LORD!
viii
Nomenclature
ADP
Adenosine diphosphate
AnMBRs
Anaerobic membrane bioreactors
ATP
Adenosine triphosphate
bioH2
Fermentative hydrogen
bioCH4
Fermentative methane
C
Concentration
C/N
Carbon to Nitrogen ratio
C:N:P:S
Proportion of carbon, nitrogen, phosphorus and sulphur
CHP
Combined heat and power
CH3COOH
Acetate
CH3CH2COOH
Propionate
CH3CH2CH2COOH Butyrate
CH3CHOHCOOH
Lactate
CH3CH2OH
Ethanol
C6H12O6
Glucose
CH3COCOOH
Pyruvate
CoA
Coenzyme A
CO2
Carbon dioxide
COD
Chemical oxygen demand
D
Diffusion coefficient
E (t)
Exit age distribution
∆E
Change in internal energy
ECMBRs
External cross-flow membrane bioreactors
ESMBRs
Externally submerged membrane bioreactors
Fe
Iron
fd (ox)
Oxidised ferredoxin
fd (red)
Reduced ferredoxin
∆G
Free energy change
HRT
Hydraulic retention time
ix
∆H
Enthalpy change
H2
Hydrogen
+
H
Proton
H2S
Hydrogen sulphide
ISMBRs
Internally submerged membrane bioreactors
J
Filtrate flux
K
Potassium
m
Maximum acceptable absolute value of mix relative deviation
N
Nitrogen
+
NAD
Oxidised Nicotiamide adenine dinucleotide
NADH
Reduced Nicotiamide adenine dinucleotide
Ni
Nickel
N2
Nitrogen
NH3
Ammonia
O2
Oxygen
OLR
Organic loading rate
∆P
Applied pressure (Transmembrane pressure)
PFRO
Pyruvate-ferredoxin oxidoreductase
PFL
Pyruvate-formate lyase
PVDF
Poly (vinylidene fluoride)
PTFE
Poly (tetrafluoroethylene)
PE
Polyethylene
PP
Polypropelene
P
Phosphorus
Pf
Feed pressure
Pp
Permeate pressure
Pr
Retentate pressure
P2G
Power-to-Gas
q
Heat
LCFA
Long chain fatty acids
R
Gas constant
x
Re
Cake layer or external fouling
Ri
Irreversible adsorption and pore plugging
Rm
Intrinsic membrane resistance
RT
Total resistance
RTD
Residence Time Distribution
∆S
Entropy change
Se
Selenium
t
Time
T
Temperature
TMP
Transmembrane pressure
∆V
Change in volume
x
Thickness
w
Total work done
W
Tungsten
µ
Viscosity
VFA(s)
Volatile fatty acid(s)
xi
Table of Contents
Abstract
iii
List of Publications
iv
Reflection on my PhD Journey
v
Acknowledgements
viii
Nomenclature
ix
Table of Contents
xii
Chapter 1. Introduction
1
1.1. Background
1
1.2. Objectives and Scope
2
1.3. Thesis Structure
4
1.4. Contribution of the Thesis
5
1.5. Research ethics and social aspects
5
Chapter 2. Fermentative process for hydrogen and methane productions
7
2.1. Basics of fermentation process
7
2.2. Dark fermentation: a pathway to effective biomethane production
9
2.2.1. Hydrogen production methods
9
2.2.2. Microbiology of hydrogen, volatile fatty acids and methane productions
12
2.2.2.1. Hydrogen
13
2.2.2.2. Volatile fatty acids
14
2.2.2.3. Methane
14
2.2.3. Thermodynamics of fermentative hydrogen and methane productions
2.3. Factors influencing fermentative hydrogen and methane productions
15
18
2.3.1. Nature of feedstock
19
2.3.2. Medium pH and alkalinity
19
2.3.3. Inoculum pretreatment
20
2.3.4. Complexity of the seed cultures
21
2.3.5. Temperature
21
xii
2.3.6. Retention times and organic loading rates
22
2.3.7. Inhibitors
22
2.3.8. Mixing
23
2.3.9. Hydrogen partial pressure
24
2.3.10. Nutrient supplementation
24
2.4. End-use technologies for fermentative hydrogen and methane
2.4.1. Attractive qualities of hydrogen and methane as energy carriers
2.5. Implications of fermentative hydrogen and methane for technological applications
25
26
26
2.5.1. Process limitations
27
2.5.2. Infrastructure barriers
28
Chapter 3. Feedstocks for fermentative hydrogen and methane productions
3.1. Feedstocks suitability for hydrogen and methane productions
31
31
3.1.1. Non-lignocellulosic feedstocks
31
3.1.2. Lignocellulosic feedstocks
32
3.2. Types of feedstocks for hydrogen and methane productions
33
3.2.1. Agriculture crop wastes
33
3.2.2. Livestock manure
34
3.2.3. Municipal solid waste
34
3.2.4. Industrial wastes and municipal wastewater
34
3.3. Inhibitory effects of fruit flavours and volatile fatty acids
3.3.1. Inhibitory effects of fruit flavours
36
36
3.3.1.1. Proposed mechanism of flavour toxicity to bacteria
36
3.3.1.2. Adaptation of bacteria to toxic environment
37
3.3.2. Inhibition of volatile fatty acids
38
3.4. Limiting the inhibitory effects of fruit flavours and volatile fatty acids
38
3.4.1. Control measure to limit fruit flavour inhibition during fermentation
38
3.4.2. Control measure to limit volatile fatty acids inhibition during fermentation
39
xiii
Chapter 4. Membrane processes for improvement of fermentative
hydrogen and methane productions
4.1. Membrane classification
43
43
4.1.1. Application of PVDF membrane in fermentative hydrogen and
methane production s
45
4.2. Influence of membrane permeability on membrane performance
45
4.3. Encapsulation technology for cell retention and inhibition control
46
4.4. Application of hollow fibre membrane configuration for VFA permeation
48
4.5. Limitations of membrane technology: Membrane fouling and cost
48
4.6. Implications of membrane applications in this study
49
Chapter 5. Bioreactor hydrodynamics for fermentative hydrogen
and methane productions
51
5.1. Ideal and real reactors
51
5.2. Mixing in bioreactors
52
5.2.1. Mixing and mean circulation times
52
5.3. Residence time distribution measurement
53
5.4. Membrane filtration
54
6. Conclusions and Future Work
57
6.1. Conclusions
57
6.2. Future Work
59
References
61
xiv
CHAPTER 1
Introduction
1.1.
Background
Humanity is endowed with diverse resources in form of materials, energy, services and
knowledge, which could be utilised for maximal benefits. Meanwhile, the usage of some of these
resources such as fossil fuels has resulted in negative consequences including resource depletion,
global climate change and environmental pollution, which could ultimately threaten human
existence. In contrast, resources such as wastes, which are inevitably generated in large amounts
from daily human activities such as food consumption, farming activities and industrial
processing, could be a source of huge wealth for a nation, without any negative consequences, if
the wastes are properly and efficiently utilised. For instance, Nigeria is the most populous
country in Africa with over 165 million people and an annual growth rate of about 2.8% (Paper
V) but the country faces worsening energy crisis with 60% of the population having no access to
the national power supply while those that have access to the power supply experience frequent
power outages. Besides, Nigeria also has the challenge of inefficient waste management system
for the huge amount of wastes inevitably generated daily by the teeming population of the
country. Therefore, Nigeria could benefit immensely from a technology that could effectively
turn wastes into affordable energy for the people (Paper V). Although there are various waste
management techniques, such as recycling, composting, landfill and incineration, anaerobic
digestion offers numerous benefits which include minimal environmental impact and waste
valorisation for production of energy carriers (hydrogen, methane and ethanol, among others),
organic fertilizers and other valuable products (1, 2).
During anaerobic digestion, the initial step of producing hydrogen from organic wastes
before using the metabolites (mainly volatile fatty acids) as building blocks of valuable
compounds (biomethane, biolipids and microbial fuel cells), enables efficient valorisation of the
organic wastes. The traditional single stage of anaerobic digestion to generate only methane for
energy usage does not allow for efficient and optimal utilization of the feedstock for energy
production. It has been observed that only 30% of the methane production during anaerobic
1
digestion is produced from carbon dioxide reduction using hydrogen while more than 70% of the
methane production comes from acetic acid conversion by heterotrophic methanogenic archaea
(3-5). Consequently, high concentration of hydrogen and carbon dioxide is left unconverted in
the digester and only a small portion of the hydrogen produced ends up being consumed by the
hydrogen consuming microorganisms. In this regard, anaerobic digestion process could be better
utilised if more energy in the form of hydrogen, in addition to methane, could be obtained from
the process (6). However, for efficient anaerobic digestion and high productivity, especially
during continuous process, it is often necessary to retain bacterial cells for a long time to obtain
high cell density and protect the microorganisms from substrate and product inhibitions (Papers I
and IV). In Papers I and IV, hydrophilic poly (vinylidene fluoride) (PVDF) membranes with pore
size of 0.1μm were used to hold and restrict the movement of the fermentative bacteria and
archaea for efficient performance. It is also often required to constantly remove some portion of
the fermentation broth to prevent product inhibition of the process (Paper III). In the study
carried out in Paper III, hydrophilic PVDF hollow fibre membrane module operated in the crossflow ‘outside-in’ mode and placed in a recirculation loop while coupled to a 5-L mechanically
stirred tank reactor, was used to extract volatile fatty acids (VFAs) from the fermentation broth.
The integration of bacterial cell retention and product recovery during continuous fermentation
processes could be effectively achieved by using membrane techniques, which have the benefits
of increasing cell concentration and reducing substrate and product inhibitions.
1.2.
Objectives and Scope
Industrial production of combined hydrogen and methane via dark fermentation process is
still extremely limited due to low hydrogen and methane yields obtained from various laboratory
research works. The low yield has been attributed to the unfavourable energetics of the hydrogen
and methane productions as well as the tendency of the fermentation process to naturally produce
cell biomass (7). Consequently, most fermentative organisms only produce a relatively small
amount of hydrogen along with other fermentation products including acetate, butyrate, butanol
and acetone, resulting in suboptimal methane production. Acetate production allows the
formation of adenosine triphosphate (ATP), while formation of other reduced products allows
the oxidation of nicotinamide adenine dinucleotide (NADH), which is necessary to maintain
redox balance in the fermentation process. Other factors including environmental and process
2
parameters, inefficient substrate conversion, substrate and product inhibition also contribute to
the low hydrogen yield from the fermentation process (8).
Considering the yield-related challenges associated with fermentative hydrogen production
and subsequent methane production during fermentation process, this research, therefore,
intended to improve the yields of fermentative hydrogen and methane through membrane control
of substrate (Papers I and IV) and product (Paper III) inhibitions (Figure 1.1) as well as using
varying operational parameters (Paper II). Moreover, in order to determine the feasibility and
sustainability of future commercial production of hydrogen and biogas productions in Nigeria as
well as other parts of the world, in terms of feedstock availability and bioreactors suitability,
reviews were conducted on biogas development in Nigeria (Paper V) and also on various types
of membrane bioreactors that could be used for ethanol and biogas production (Paper VI).
Figure 1.1. Schematic diagram for the scope of the research
The aim of this research was achieved through the investigation of the following activities:
(i) Demonstration of the protective effects of hydrophilic PVDF membranes on fermentative
bacteria against inhibitory effects of fruit flavour media during fermentation process
(Papers I and IV),
3
(ii) Evaluation of the potential of hydrogen yield enhancement from fruit fermentation through
varying hydraulic retention times and fruit mixing (Paper II),
(iii) Investigation of the feasibility of employing membrane VFA permeation to improve
fermentative hydrogen production without any major modification of the hydrodynamics in
the anaerobic membrane bioreactor system (Paper III),
(iv) Assessment of feedstock availability for commercial production of biogas production in
Nigeria (Paper V),
(v) Study of membrane bioreactors suitable for biogas production (Paper VI).
Generally, hydrogen and methane productions can be enhanced using a suitable microbial
species, process modification, efficient bioreactor design and genetic techniques. The scope of
this research was, however, limited to the application of process modification and bioreactor
design for the improvement of fermentative hydrogen and methane productions.
1.3.
Thesis Structure
The thesis is organised into two parts: the first part provides information on the basic
principles that the research work was based on, while the second part comprises of the six
articles from the research. The experimental work mainly focused on two subject areas with
regard to fermentative hydrogen and methane productions. The first area was on the application
of membrane encapsulation techniques to protect bacteria from substrate inhibition and thereby
enhance the hydrogen and methane production potential of the bacteria. The second area of the
research focused on reducing the effect of product inhibition on bacteria through process
parameters and membrane permeation of volatile fatty acids with consequent improvement on
the fermentative hydrogen production.
The chapters included in the first part of the thesis are as follows:
Chapter 1 introduces the main reasons for conducting research on the investigated subject as
well as the intended objectives of the research.
Chapter 2 lays the foundation for the research problem with literature review on fermentation
process and thermodynamics for hydrogen and methane productions including
determining factors, end-use technologies as well as the implications of the
technology applications.
4
Chapter 3
provides information on potential feedstock for fermentative hydrogen and
methane productions along with the inhibitory effects of fruit flavours and VFAs
Chapter 4
describes membrane processes and the underlying principles of membrane
encapsulation and VFAs permeation
Chapter 5
relates fluid hydrodynamics in the bioreactors to the bioreactor performance and
the effectiveness of the fermentation process
Chapter 6
1.4.
summarises the main research conclusions and provides directions for future work
Contribution of the Thesis
Generation of hydrogen and methane through anaerobic fermentation process has been
established as an environmentally-friendly and non-energy intensive technique that could play a
significant role in the future green and zero-emission world. Fermentative methane technology
on an industrial scale has been around for decades in most advanced countries, while
fermentative hydrogen production is not yet commercialised due to the challenge of low
hydrogen yield from the process. Research efforts have therefore been intensified to find ways
of improving, not only hydrogen yields, but also methane yields from fermentation of diverse
organic compounds. The results from this research are significant in the area of reducing the
effects of substrate and product inhibitions on the fermentative hydrogen and methane
productions, thereby, enhancing the yields of the two energy carriers. Moreover, the study
provides insight into the antimicrobial effects of fruit flavours on both fermentative hydrogen
and methane and can be used as a guide for improving the gas yields during commercial
applications.
1.5.
Research ethics and social aspects
In view of the growing global threats of energy insecurity and climate change due to
greenhouse gas emissions, coupled with the inefficient waste management system, especially, in
most third world countries, the research was focused on how to efficiently recover resources in
terms of energy or useful products from waste materials while simultaneously reducing
environmental pollution. It is anticipated that efficient production of fermentative hydrogen and
5
methane could be used as a tool in tackling global challenges including energy insecurity,
climate crises and inefficient waste management system. Energy carriers, such as fermentative
hydrogen and methane, provide utility in terms of energy, and its effective demand by consumers
will depend on factors including cost effectiveness, appropriateness of the technology,
availability, reliability, efficiency and technical potentials. Therefore, the effects of these
variables, especially, membrane techniques and process enhancement, were, therefore,
investigated during this study (Papers I - IV) for the improvement of fermentative hydrogen and
methane productions.
Nevertheless, there are some considerable ethical problems related to commercial application
of biofuels such as fermentative hydrogen and methane. It is believed that increasing demand for
biofuel production may simultaneously cause the rise in demand for arable lands used for
growing food crops, thereby, leading to food shortage. Growing biofuel crops in arable lands
may compete with food production for arable land, water and plant nutrients, which may create
more problems including increase in global market price for food making it beyond the reach of
poor people. In addition, there is a risk of environmental pollution due to application of fertilizer
and pesticides in the production of biofuel crops. In other words, ethical dilemmas often arise in
the process of tackling some of the global crises including management of natural resources,
energy, climate changes and food crises. Consequently, it is necessary to consider the effects of
the development of any green technology on various actors (people, animals and the natural
world) in such a way that the green technology being developed will have the least possible
damage, if any, to the various actors.
Regarding ethical issues relating to my research, the food-versus-biofuel problem is not a
barrier, as organic waste materials that are produced from daily human activities, are the
potential feedstocks for the production of the biofuels. Furthermore, during the research work,
ethical norms, including honesty, objectivity, integrity, carefulness, openness, respect for
intellectual property, confidentiality, reliable publication and competence, which govern research
conduct in my fields were strictly adhered to in order to make the results of the investigations
acceptable to the general public (9, 10).
6
CHAPTER 2
Fermentative process for hydrogen and methane productions
The fermentation process for the production of hydrogen and methane is an anaerobic
digestion process in which complex organic feedstock is broken down microbially into simpler
substances in the absence of oxygen.
2.1. Basics of fermentation process
Anaerobic fermentation process is carried out by different species of anaerobic
microorganisms in several successive steps, with each step depending on the preceding one. The
fermentation process consists of four steps, which include hydrolysis, acidogenesis, acetogenesis
and methanogenesis (11) (Figure 2.1). In hydrolysis, the complex components in the feedstock
including carbohydrates, proteins and fats, are initially broken down by extracellular enzymes
into their respective monomeric units- glucose (sugars), amino acids and fatty acids. The
extracellular enzymes including amylases, proteases and lipases, are secreted by various strains
of hydrolytic bacteria to break down the complex compounds into soluble compounds that could
easily be transported across the bacterial cell membrane. The enzymes production as well as
feedstock particle size, duration of enzyme-particle contact, pH, among others, determine the rate
of the hydrolysis process (12, 13). The products of the hydrolysis are converted by acidifying
bacteria into volatile fatty acids (VFAs), alcohols, carbon dioxide (CO2), hydrogen (H2) and
ammonia (NH3) during acidogenesis (14). The acidifying bacteria, which are a mixture of
facultative and obligatory bacteria, are important in creating an anaerobic condition during the
fermentation process, as the facultative anaerobes have the potential of using up the oxygen that
might have been mistakenly introduced into the process along with the feed.
During acetogenesis, low molecular weight VFAs are converted by acetogenic bacteria
into acetic acid (CH3COOH), CO2 and H2, which can be easily utilised by methanogenic archaea
(15). Acetogenesis requires efficient and continuous removal of hydrogen formed from the
fermentation process, as the process can only be favoured thermodynamically at low partial
pressure of hydrogen (11). This was one of the reasons why recovery of fermentative hydrogen
7
during dark fermentation process was given a priority during this research study (Papers II, III
and IV) as it could make the fermentation process cost effective in terms of the energy recovery.
At the final stage, methanogenic archaeal group uses three biochemical pathways, namely,
acetotrophic, hydrogenotrophic and methylotrophic pathways, to produce methane from the
products of previous stages including CH3COOH, CO2, H2., formate, methanol and methylamine.
It has however be shown that more than 70% of the methane production in anaerobic digestion
comes from acetate conversion (3, 16, 17).
Figure 2.1. Microbial degradation process for fermentative hydrogen and methane productions
The four phases during fermentation process, namely, hydrolysis, acidification,
acetogenesis and methanogenesis can all take place in a bioreactor resulting in one-stage
fermentation. Alternatively, the four phases may be divided into two parts called two-stage
8
fermentation process. The first part including hydrolysis, acidogenesis and acetogenesis, takes
place in the first bioreactor, while the second part called methanogenesis takes place in the
second reactor (18). The two-stage fermentation is appropriate for combined hydrogen and
methane productions as it allows for the optimisation of process parameters including pH and
temperature, which differ for both hydrogen and methanogenic archaea (19-21). The recovery of
hydrogen as energy carrier during anaerobic fermentation studies is usually done in a two-stage
fermentation process in which VFAs, produced as the by-products of the dark fermentation
process in the first reactor, are further degraded to produced other valuable products including
hydrogen, methane, biodiesel and bioplastics.
Fermentation process for hydrogen and methane productions can be operated in batch,
fed-batch or continuous mode (22, 23). Batch reactor is operated as a closed culture system, in
which the bioreactor is filled with nutrients and other additives at the beginning of the process;
thereafter, the reactor is sealed for the digestion process to complete. Then the fermentation
products are recovered at the end of the process. In this study, fermentative methane productions
from feedstocks containing flavour compounds were investigated using batch fermentation
processes (Papers I and IV). In fed-batch fermentation, critical elements of the nutrient solutions
are added to the bioreactor, while the fermentation broth remains in the bioreactor until the end
of the fermentation process (24, 25). Regarding continuous fermentation process, the bioreactor
is operated as an open system in which substrates are added continuously to the bioreactor with
simultaneous withdrawal of the digested sludge. Fermentative hydrogen production in this study
was investigated using continuous fermentation process, as the process was more effective in
preventing the growth of hydrogen consuming microorganisms such as methanogenic archaea.
However, the mode of operation was not completely a continuous operation as the substrate
feeding and effluent withdrawals were done only once each day during the experiment.
2.2. Dark fermentation: a pathway to effective biomethane production
2.2.1. Hydrogen production methods
Various industrial methods for hydrogen production exist including steam reforming of
methane, thermo-chemical water splitting and biomass gasification, pyrolysis and ‘Power-toGas’ (P2G) techniques (Figure 2.2). The P2G is a new technique of hydrogen production that is
9
now attracting researchers’ attention because of its potential to store excess electricity in form of
gas fuel by the conversion of electrical power to gas fuels. In the P2G techniques, different
pathways exist including ‘Power-to-Hydrogen’, ‘Power-to-Methane’ and ‘Power-to-Syngas’. In
the Power-to-Hydrogen, the hydrogen produced from the electrolysis of water is used directly as
transport fuel or for other purposes (26). Regarding Power-to-Methane’, the hydrogen produced
from the electrolysis is combined with CO2 to form methane using a methanation reaction
(Sabatier or biological methanation) (27-31). In the Power-to-Syngas’, the hydrogen formed
from the electrolysis of water is combined with CO2 in a conversion reactor to produce a mixture
of gases, including hydrogen, carbon monoxide and water, which is called syngas (32).
Figure 2.2. Hydrogen production techniques
10
Meanwhile, most of the above techniques employed in the industrial hydrogen production,
are energy intensive and non-environmentally friendly techniques. Consequently, low-energy
techniques that are also non-polluting are currently being focused on as alternative hydrogen
production techniques. The low-energy techniques include dark fermentation (33, 34),
biophotolysis, photo-fermentation (35, 36) microbial electrolysis and enzymatic techniques.
Biophotolysis is a water splitting process using green algae or cynaobacteria via direct and
indirect routes with light as the energy source. Molecular oxygen and hydrogen are produced by
utilising inorganic CO2 in the presence of sunlight and water. In photo-fermentation, hydrogen is
produced through the activities of photosynthetic bacteria, which have the ability to utilise
diverse substrates, ranging from inorganic to organic acids with light as the energy source.
Microbial electrolysis involves the application of external electric potential to enhance hydrogen
production from microbial cells using various organic substrates. Among the biological
techniques, dark fermentation seems like a promising alternative as future commercial hydrogen
production process because, unlike other biological methods, it does not require light energy and
has lower energy demands as well as having higher hydrogen production rate (37). Moreover, it
is simple and robust and has the potential for small footprint (7, 38, 39).
Dark fermentation is different from other biological processes in the sense that it uses
organic substrates as both energy and carbon sources. It is a process that involves the microbial
conversion of organic substrates to biohydrogen in an oxygen-free environment. It is called dark
fermentation because the fermentation takes place in the absence of light, unlike photofermentation, that is a light-dependent process. In normal anaerobic digestion process, dark
fermentation usually occurs together with methanogenesis, as hydrogen and acetate produced
during dark fermentation process are used as substrates for methanogenic archaea (40). However,
when the two processes occur together in a single-stage system, energy recovery is usually
inefficient as hydrogen produced is consumed by hydrogen consuming microorganisms leaving
only methane as the only gaseous energy carrier that could be recovered. Moreover, the two
processes differ in terms of the required nutrients, optimal environmental conditions, growth
kinetics, among others; hence, any disturbance in the process optimal conditions can affect the
efficiency of the microorganisms community. If the dark fermentation is, however, separated
from the methane forming process, for example, in a two-stage system, overall energy extraction
11
from the substrate conversion could be increased with the additional energy obtained through
methane generation from the by-products of the hydrogen fermentation.
2.2.2. Microbiology of hydrogen, volatile fatty acids and methane productions
Anaerobic fermentation of organic compounds for energy production and cell growth
usually involves electron generation, which must be disposed off to electron acceptors. In dark
fermentation reactions, which take place in an environment that lacks terminal electron acceptors
such as oxygen, sulphate, nitrate and ferric iron, redox balance is maintained by the production
of molecular hydrogen (H2) with protons (H+) from water serving as electron acceptor. Electrons
released during the conversion of organic compounds into series of degraded and oxidised
intermediate compounds, are utilised to convert coenzymes such as nicotiamide adenine
dinucleotide (NAD+) to their reduced form. The reduced coenzyme returns back to its oxidised
form by reducing intermediate compounds including pyruvate (Figure 2.3). Reduction
equivalents including formate, reduced ferredoxin and nicotiamide adenine dinucleotide
(NADH) function as electron donors to hydrogen (41).
Figure 2.3. Metabolic process during dark fermentative hydrogen production
12
2.2.2.1. Hydrogen
During anaerobic fermentation process, the bacteria break down organic compounds into
pyruvate, which is further degraded with the aid of either of two enzymes, namely, pyruvateformate lyase (PFL) and pyruvate-ferredoxin oxidoreductase (PFRO) (Equations 2.1 & 2.2). The
most commonly used enzyme during fermentative hydrogen production or fermentation
involving obligate or thermophilic bacteria, is PFRO.
PFL
Pyruvate + CoA 
→ Acetyl − CoA + formate
2.1
Pyruvate + CoA 
→ Acetyl − CoA + CO 2 + 2 fd (red )
2.2
PFOR
The pyruvate generated from fermented sugars is cleaved by pyruvate ferredoxin oxidoreductase
in the presence of coenzyme A (CoA) to generate acetyl CoA, reduced ferredoxin and CO2
(Equation 2.3), while the reduced ferredoxin generated, catalyses hydrogen formation (Equations
2.4 & 2.5).
C 6 H 12O 6 + 2 NAD + → 2CH 3COCOOH ( pyruvate) + 2 NADH + 2 H +
2.3
Pyruvate + CoA + 2 fd (ox) → Acetyl − CoA + 2 fd (red ) + CO 2
2.4
+
2 H + fd (red ) → H 2 + fd (ox)
2.5
Hydrogen production is related to the activity of an iron-sulphur protein called
ferredoxin, an electron carrier of low redox potential. The metabolic process for hydrogen
production is dependent on the reduction of the metabolite ferredoxin, which in turn depends on
the recycling of ferredoxin through oxidation. The transfer of electrons from NADH to reduced
ferredoxin ensures the continuation of the recycling process of ferredoxin (42). The fermentative
bacteria need to regenerate the cytoplasmic electron carrier NAD to maintain the glycolysis.
Three main classes of hydrogen forming enzymes including [FeFe]-hydrogenase, [NiFe]hydrogenase and nitrogenase, catalyse the recycling processes of ferredoxin (42). In clostridium
bacteria, the hydrogen production is mostly due to [FeFe]-hydrogenase with activity that is
hundred times higher than [NiFe]-hydrogenase and a thousand times higher than nitrogenase (43,
44).
13
2.2.2.2. Volatile fatty acids
During dark fermentation, the main aqueous products are acetate, propionate and
butyrate, while formate, lactate, valerate and caproate are also produced as minor acidogenic
products (45). The acetyl-CoA produced during the cleavage of PFRO, is the essential
intermediate in the production of both volatile fatty acids and solvents. When volatile fatty acids
are generated, there are no reductions that could prevent the reduced ferredoxin from transferring
electrons to a hydrogenase that permits the use of protons as a final acceptor; thus, the ferredoxin
is re-oxidised and molecular hydrogen is released from the cell. However, under certain
unfavourable conditions such as high hydrogen partial pressure, the formation of hydrogen is
limited, and carbon flow to acid production pathway is switched to the solvent production
pathway, which involves reduction. As a result, ferredoxin is unable to transfer electrons to a
hydrogenase for hydrogen production, thereby the cell is forced to channel electrons through
NADH:ferredoxin oxidoreductase (NADH consumption) to form some reduced compounds such
as lactate, ethanol and butanol, resulting in a lowered hydrogen yield (42, 46). This is why it is
better to have two-stage fermentation process, which allows continuous removal of hydrogen
from the process, while the metabolic products from the first stage are used for methane
production, thereby leading to efficient energy recovery.
2.2.2.3. Methane
High proportion of methane production in an anaerobic digester occurs from the use of
acetate and hydrogen by methane-forming bacteria. Aceticlastic cleavage of acetate and
reduction of CO2 are the two major pathways to methane production. The pathways involving
propionic and butyric acids fermentation only have minor contribution to methane production.
There are three principal groups of methane-forming bacteria including hydrogenotrophic,
acetotrophic and methylotrophic methanogens. Hydrogenotrophic methanogens use hydrogen to
convert CO2 into methane, acetotrophic methanogens split acetate into methane and CO2, while
methylotrophic methanogens grow on substrates that contain methyl group including methanol
and methylamines. The acetotrophic methanogens reproduce more slowly than the
hydrogenotrophic methanogens and are adversely affected by the accumulation of hydrogen.
The maintenance of low partial hydrogen pressure in an anaerobic digester is, therefore,
favourable for the activity of acetotrophic methanogens (47).
14
2.2.3. Thermodynamics of fermentative hydrogen and methane productions
The thermodynamics of fermentation process determines the bioH2 and bioCH4 yields
from the process. As every chemical reaction involves loss or gain of electrons, the bacteria
involved in the fermentation process conserve their energy through the coupling of ATP for the
breakdown of organic compounds in their environment. The amount of energy released during
the process depends on the distance between the electron donor and electron acceptor, as the
coupling of two reactions cannot occur if they are separated from each other. The energy
released is necessary for various bacterial activities including mass transport of molecules across
bacterial cell membrane. In essence, bacterial metabolism involves energy transformation, and
the energy transfer mechanism is based on thermodynamics principles (first and second laws of
thermodynamics) (40, 48). The total energy involved in bacterial metabolism (oxidation and
reduction reactions) must be conserved to maintain the integrity of the bacteria as stated in the
first laws of thermodynamics, which states that total amount of energy in nature, is constant. In
other words, heat (q) added to a system of given energy content must appear as a change in the
internal energy (∆E) of the system or in the total work carried out by the system on the
surrounding (w) (Equations.2.6 & 2.7).
q = ∆E + w
2.6
∆E = q − w
2.7
When heat addition to a system also results in volume change (∆V) at a constant pressure (P), the
change in internal energy can be referred to as enthalpy (∆H) (Equations 2.8 & 2.9).
∆H = ∆E + P ∆V
2.8
or ∆H = q − w
2.9
The second law of thermodynamics expresses that all reactions that occur proceed in a direction
that the degree of randomness, commonly referred to as entropy (S) of the universe, increases to
the maximum possible towards an equilibrium position (Equations 2.10 & 2.11).
15
∆S =
q
T
2.10
or q = T ∆S
2.11
where q, T and ∆S represent heat, temperature and entropy change.
The combination of first and second law of thermodynamics (Equation 2.12) indicates that the
tendency to attain position of maximum entropy is the driving force of all processes including
the biological processes, and heat is either given up or absorbed by the system and its
environment to enable them to reach a state of maximum entropy.
∆H = T ∆ S − w
2.12
The change in heat (enthalpy) and entropy are related by the free energy (Equations 2.13 &
2.14), which is the energy released to perform useful work.
∆G = ∆H − T ∆S
2.13
∆G = − w (since ∆H = T ∆S − w )
2.14
The values of free energy change (∆G) determine the spontaneity of a reaction. Free
energy values that are greater than zero (∆G > 0), equal to zero (∆G = 0) or less than zero (∆G <
0) represent that the reactions are not spontaneous, at equilibrium or spontaneous, respectively.
As given in the equation, increase in temperature of a reaction process could make the reaction
spontaneous depending on the enthalpy change of the system. As a consequence, most of the
experiments investigated in this study were carried out at thermophilic temperatures (55°C)
(Papers I, II & IV). Regarding the transfer of substances through cell membranes and other
surfaces, the exchange free energy (∆G) for the transport of a mole of substance of
concentration, C1, from one place to another where it is present at C2 is given as (Equation 2.15)
∆G = RT log
C2
C1
2.15
16
The reaction is favourable when ∆G is negative, that is, when C2 is less than C1. In the absence
of intervening factors, an equilibrium will be reached where C2= C1, resulting in ∆G being equal
to zero.
In a dark fermentation process, 12 mol H2 per mol glucose could theoretically be obtained
from the complete conversion of glucose to H2 and carbon dioxide (Equation 2.16). But, the
reaction is not thermodynamically favourable due to the production of a large quantity of
metabolic products (VFAs, alcohols and lactate) associated with hydrogen production. The
thermodynamic constraints make the maximum attainable hydrogen yields to be 4 and 2 mol/mol
glucose if the associated metabolism products are acetate and butyrate, respectively (Equations
2.17 & 2.18). However, the fermentation with only acetate as the main organic acid (Equation
2.18) has higher theoretical yield than with other organic acids as products (Figure 2.4) under
equilibrium conditions (7). Acetate and hydrogen are not the only fermentation products formed
during the process, other secondary fermentation products such as ethanol, butyrate and lactate
are also formed, thereby reducing the molar yield of the hydrogen production (49).
C 6 H 12O 6 + 6 H 2O → 12 H 2 + 6CO 2
(∆G° = +3.2 kJ)
2.16
C 6 H 12O 6 + 2 H 2O → 4 H 2 + 2CO 2 + 2CH 2COOH ( acetate)
(∆G° = -206 kJ)
2.17
C 6 H 12O 6 → 2 H 2 + 2CO 2 + CH 3CH 2CH 2COOH (butyrate)
(∆G° = -254 kJ)
2.18
The actual yield during the real experiment is often less than the maximum theoretical yield.
For example, in this study (Paper II), experiment was conducted to explore means of increasing
hydrogen production from fruit wastes including orange, apple, banana, grape and melon. The
highest yield obtained was from the fruit mixture with equal weight proportion at an operating
temperature of 55°C and HRT of 5 days, and the yield was just 30% of the theoretical yield
(Paper II). The low yield might be attributed to the tendency of anaerobic fermentation processes
to form other secondary fermentation products including ethanol, propionate and lactate, which
consume hydrogen by uptake hydrogenases (7).
17
Figure 2.4. Comparison of theoretical hydrogen yield of glucose fermentation pathways
2.3. Factors influencing fermentative hydrogen and methane productions
The production of hydrogen and methane during fermentation process is facilitated by the
concerted action of various anaerobic microorganisms. The microorganism efficiency regarding
the gas production depends on several factors: nature of feedstock, inoculum pretreatment,
medium pH and alkalinity, temperature, solid and hydraulic retention times, organic loading
rates, hydrogen partial pressure, mixing, inhibitors, carbon to nitrogen ratio (C/N) and inoculum
to substrate ratio (ISR) (50, 51). The factors are known to influence the microbial metabolism
processes and thereby determine the processing time, production rate, yield and relative
composition of hydrogen and methane generated from the fermentation process.
18
2.3.1 Nature of feedstock
The characteristics of feedstock including composition, C/N ratio and particle size affect
the feedstock biodegradability, yield and rate of hydrogen and methane productions during
anaerobic process. The microorganisms use the feedstock as a source of energy, electron
acceptors and building blocks for new cell growth. The amount of the major components of
feedstock including proteins, lipids, carbohydrates (monosaccharides, disaccharides and
polysaccharides) and lignin, influences the ease or difficulty of the biodegradation of the
feedstock (52). Readily degradable feedstocks, such as low molecular sugars, food waste, among
others, degrade faster than fats, proteins and lignocellulosic materials. Lignocellulosic materials
usually require pretreatment prior to their digestion due to the presence of lignin that tightly
binds cellulose and hemicelluloses together. Various pretreatment methods that could enhance
the biodegradation of biomass include physical, chemical or biological methods (53). Feedstock
composition also affects the C/N of the feedstock with the optimum C/N ratio for anaerobic
digestion reported to be in the range of 20 - 30: 1 (54). Very high C/N ratio leads to low biogas
production due to low protein formation that affects the energy and structural metabolism of the
microorganisms in terms of the substrate degradation efficiency. On the other hand, low C/N
ratio increases ammonia concentration which could possibly result in ammonia/ammonium
inhibition of the fermentation process (52). Furthermore, particle size of feedstock also plays a
significant role in the biodegradation of the feedstock as small particle size provides high surface
area for microorganism activities (55). A particle size of 2 mm was reported to be optimum for
biodegradation of some feedstock (56). In Paper III, ground straw was sieved to a particle size of
2mm before the particles were used as substrate in the investigation of the effect of VFA
permeation on biohydrogen production (Paper III).
2.3.2. Medium pH and alkalinity
The pH of the fermentation broth plays a significant role in the effectiveness of the
fermentation process as it directly affects the activities of the bacteria involved in the
fermentation process. Generally, methanogens are more sensitive to acidic conditions than other
anaerobic bacteria involved in the fermentation process. Hydrogen and methane productions
during fermentation process require different pH values of 5.5 - 6.5 (57) and 6.5 - 8.2 (1, 58, 59),
respectively. Anaerobic fermentation has a natural way of controlling the pH of the medium
19
through the buffering system of the dissolution of carbon dioxide and ammonia (alkalinity) to
control the high and low pH fermentation media respectively (60, 61). However, the process
buffering system is often overwhelmed by the nature and loading rate of the feedstock, thereby
requiring external measures for the pH regulation. The initial pH values of the substrate media
used for hydrogen (Papers II, III and IV) and methane productions (Papers I and IV) during this
study were on average 5.5 and 6.8, respectively.
In Paper IV, the initial pH range for all the reactors for the continuous hydrogen
fermentation was from 5.2 to 5.9. However, gradual reduction in the pH values of the fermenting
media below 5.0 was observed at the beginning of the experiment, which could be attributed to
the production of organic acids associated with the hydrogen formation during the fermentation
process (62). The pH profile indicated that the pH values for all the reactors did not vary
significantly but were nearly constant throughout the experiment, with an average value of 4.40
± 0.04. This could possibly imply that the daily effluent withdrawal from the reactor system
could have prevented the accumulation of organic acids that could have led to drastic reduction
in the pH value of the fermentation media. Moreover, it could also be due to the adaptation
potential of fermentative microorganisms to the inhibitory fermentative media.
2.3.3. Inoculum pretreatment
Several methods of inoculum pretreatment, including heat shock treatment, acid/base
treatment, as well as using chemical inhibitors such as 2-bromoethanesulfonic acid (BESA),
acetylene and chloroform, have been used to improve hydrogen and methane productions during
fermentation process. Effective hydrogen production often requires initial pretreatment of the
seed inoculum in order to suppress the activities of the hydrogen consuming bacteria, since they
are usually in syntrophic association with hydrogen producing bacteria. However, it has been
observed that inoculum pretreatment alone could not sustain the inhibition of the hydrogen
consuming processes for a long period of time. Operation of a fermentation process with
pretreated inoculum in the medium, at initial pH of 5.5, has been proved to be effective in the
inhibition of the hydrogen consuming processes such as methanogenesis and homoacetogenesis
(63). In this study, hydrogen producing bacteria was heat-pretreated at 100°C for 15 min. The
initial pH of the mixture of the heat-pretreated inoculum and the substrate was then adjusted to
around 5.5 before they were used for the continuous hydrogen production from the medium
20
containing inhibitory flavour compounds. This was to ensure that the growth of the hydrogen
consuming microorganisms such as methanogenic archaea, were effectively inhibited during the
process (Papers II, III and IV)
2.3.4. Complexity of the seed cultures
Hydrogen production can also be influenced by the nature of the seed cultures, which can
either be pure cultures or mixed cultures. Pure culture fermentation involves a single species of
seed culture throughout the fermentation process, while mixed culture fermentation is carried out
by multiple strains of seed culture. In pure culture, there is limited interference by the hydrogen
consuming bacteria, such as sulphate-reducing, homoacetogenic and methane producing bacteria.
However, special care needs to be taken when pure culture is used as it can easily be
contaminated by hydrogen consuming bacteria. One important benefit of mixed cultures is their
ability to utilise a variety of substrates due to the presence of diverse microorganisms.
Consequently, mixed cultures were used in all the anaerobic fermentation processes (batch and
continuous) involved in this study (Papers I, II, III and IV)
2.3.5 Temperature
Anaerobic fermentation processes for hydrogen and methane productions can be run at
different temperatures including psychrophilic temperature (less than 20°C), mesophilic
temperature (30 - 42°C, usually 35°C), thermophilic (50 - 60°C, usually 55°C) and
hyperthermophilic (>80°C) (60, 64-66). It is, however, important that temperature should be kept
constant when operating fermentation process in any of the temperature range, as temperature
fluctuations can reduce the gas production. Compared with mesophilic process, thermophilic
process is faster and more efficient with higher gas production as it increases compounds
solubility and enhances reaction rates. However, thermophilic process is more energy intensive
and sensitive to any disturbance in terms of environmental and operational parameters (67).
Temperature influences the physicochemical characteristics of the fluid medium as well as the
growth rate of the bacteria in the anaerobic bioreactor (68). In this study, most of the experiments
were operated at thermophilic temperature 55°C (Papers I, II and IV), except the experiment
conducted to investigate the effects of membrane permeation of VFAs on bioreactor
hydrodynamics and hydrogen production, which was operated at mesophilic temperature (Paper
21
III). The reactor used for the study in Paper III was, however, equipped with a two-stage impeller
to ensure uniformity in temperature and other process parameters.
2.3.6. Retention times and organic loading rates
Retention times, including solids and hydraulic retention times (SRT and HRT), are the
average times that the solid and liquid part of feedstocks, respectively, spend in the digester
before they are removed. They are often dependent on the nature of the feedstock, the
temperature of the process, digester volume and organic loading rate. Feedstocks that are easily
degraded will require shorter times than those that are not easily degraded. The SRT and HRT of
feedstock in a completely mixed bioreactor without recycling are generally the same. The range
of HRT of anaerobic digester for solid waste treatment can vary from 3 to 55 days (Paper II)
according to the nature of the feedstock, process temperature and bioreactor configuration (52),
while SRT, especially for high rate digesters, can range from 10 to 20 days (69). The technique
of low HRT and pH is often employed during continuous hydrogen production for effective
elimination of hydrogen consuming bacteria such as methanogens (70, 71). In Paper III, effects
of mixing and varying HRTs of 3, 5 and 8.6 days of fruit wastes (orange, apple, banana, grape
and melon) on bioH2 in a continuous process at 55°C for 47 days were investigated. Although it
was observed that there was no statistically significant effect of the interaction of HRT and fruit
mixing on bioH2, there was an improvement in cumulative bioH2 yields from all the feedstocks
when HRT was 5 days while fruit mixture with equal fruit proportion produced the highest
cumulative bioH2 yield of 513mL/g VS (30% of the theoretical yield).
The organic loading rate (OLR), which is the quantity of feedstock volatile solids fed into
a fixed digester volume within a period of time, is related to HRT value. For a constant volatile
solid (VS) of a feedstock, low HRT is coupled with high OLR, while for a varied VS, OLR value
can vary at the same HRT rate. The OLR of a continuous fermentation process is an important
parameter that should be managed effectively, as very low or high OLR could result in lower gas
production or VFA accumulation, respectively (11).
.
2.3.7. Inhibitors
Several substances including antibiotics, disinfectants and detergents, food preservatives,
organic substances, ammonia, sulphide, oxygen, heavy metals, among others, can act as
22
inhibitors during fermentation process since they could cause inhibition of bacterial growth and
performance (11, 72). The sources of the inhibitors are either associated with fresh feedstocks
such as fruit flavours in fruit wastes (Papers I, II and IV) or as by-products of the bacterial
metabolism activities such as VFAs (Paper III). The effects of these inhibitors are influenced by
factors including the adaptation ability of the bacteria to the inhibition (acclimation), the absence
or presence of other inhibitors (antagonism or synergism), and variation in process parameters
(11).
2.3.8. Mixing
During fermentation processes, three phase reactions including liquid-solid, gas-solid and
gas-liquid, exist inside the bioreactors. Mixing of bioreactors facilitates the transfer of energy,
nutrients and metabolites within the bioreactors thereby ensuring efficient reactions among the
three phases. In the absence of mixing, slurry inside the digester tends to accumulate to form
scum layer on the surface, thereby preventing the upward movement of the gas. Mixing enhances
the fermentation process by providing adequate contact among the fresh feedstock, bacteria and
nutrients as well as reducing scum build-up (73, 74). Mixing also facilitates physical and
chemical uniformity of digested sludge by preventing the formation of dead zones due to particle
deposition. However, optimisation of the mixing conditions is necessary, as gentle mixing is
good to avoid bacterial shearing and inhibition while vigorous agitation is needed to achieve
biomass or solid suspension in a bioreactor (75).
In Paper III, a two-stage impeller consisting of a bottom impeller with four-blade disk
turbine, and the top impeller with a three-bladed pitched turbine, was employed to promote
uniform flow discharge in all directions within the membrane bioreactor during the experiment
to investigate the effects of VFA permeation on the bioreactor hydrodynamics and hydrogen
production. The stirred tank reactor, which was coupled to the hollow fibre-membrane module,
was observed to be instantaneously mixed, in comparison to the fluid in the recirculation loop.
The results not only showed that the properties of the fluid in the stirred tank were unaffected by
the presence of the recirculation loop, but also showed that the stirring in the tank had no
apparent effect on the mixing properties of the loop. Besides, the loop exhibited nearly the same
behaviour with and without permeate extraction, provided the recirculation flow rate was more
than ten times as high as the withdrawal flow rate (Paper III).
23
2.3.9. Hydrogen partial pressure
Hydrogen production pathways are very sensitive to hydrogen partial pressure as
accumulation of hydrogen in the reactor headspace may increase the partial pressure of hydrogen
in the reactor system. During dark fermentation process, the disposal of electrons via pyruvateferredoxin oxidoreductase or NADH-ferredoxin oxidoreductase and hydrogenase might be
affected by the corresponding NADH and acetyl-CoA levels as well as environmental conditions
such as high hydrogen partial pressure. When hydrogen concentration increases, the law of mass
action limits the formation of hydrogen and the cell is forced to channel electrons through
NADH: ferredoxin oxidoreductase, shifting the metabolic to produce more reduced metabolites
and solvents including ethanol, lactate, propionic acid, and butanol, resulting in reduction in the
hydrogen production (76-78). As solvent production involves reductions, ferredoxin is unable to
transfer electrons to a hydrogenase for H2 evolution.
In the presence of high concentration of hydrogen, the formation of reduced metabolites
and solvents including propionate, lactate and ethanol are more thermodynamically favourable
than acetate and butyrate formations (Equations 2.17 – 2.21) (7, 41, 43, 49, 79-81). Although
ethanol formation is less thermodynamically favourable, its formation under some conditions
yields no hydrogen (Equation 2.21). For this reason, it is necessary to avoid high partial pressure
of hydrogen that can force the cell to switch from acidogenic to solventogenic fermentation. In
this study, hydrogen partial pressure was controlled by the continuous removal of the generated
hydrogen from the bioreactor system, and also by daily mixing of the bioreactors (Papers II, III
and IV)
C 6 H 12O 6 + 2 H 2 → 2CH 3CH 2COOH + 2 H 2O ( propionate)
(−279.4 kJ )
2.19
C 6 H 12O 6 → 2CH 3CHOHCOOH + H +
( Lactate)
(−225.4 kJ )
2.20
C 6 H 12O 6 → 2CH 3CH 2OH + 2CO 2
( Ethanol )
(−164.8 kJ )
2.21
2.3.10. Nutrient supplementation
In addition to the feedstock supplied during fermentation process, microorganisms also
require some nutrients including nitrogen, phosphorus (P), potassium (K), iron (Fe), copper,
cobalt, manganese, calcium, molybdenum, vanadium, magnesium, sodium, nickel (Ni), selenium
24
(Se), tungsten (W) and zinc in certain proportions, for effective bacterial metabolism and growth.
Nitrogen and phosphorus are macronutrients that are required in large quantities by anaerobic
microorganisms. For example, nitrogen is an essential component of amino acids and is required
for optimal growth of the microorganisms. For high-strength and low-loading organic wastes, the
ratio of chemical oxygen demand (COD), nitrogen and phosphorus that are mostly used are
1000:7:1 and 350:7:1, respectively (11). Micronutrients such as cobalt, iron and nickel, among
others, are necessary as they are incorporated in enzyme systems for proper substrate degradation
and conversion into hydrogen and methane. Cobalt, for example, is required as an activator of
enzymes during hydrogen and methane productions. Iron is an important component of
hydrogenases, the enzymes involved in the production of hydrogen.
Metal ions such as magnesium and sodium are also required for transport across cell
membrane and as cofactors of other enzymes. Yeast extract is often used to supply some of the
nutrients to the microorganism as it contains amino acids, minerals and vitamins including, the B
vitamins, biotin and folic acid. However, for these nutrients to be biologically available to
anaerobic microorganisms, they must be present in the nutrient media in soluble form and not as
precipitates. Besides, the available nutrient must not be in excess as excess of these nutrients can
inhibit the efficiency of the anaerobic microorganisms (11, 82). For instance, for effective
fermentation of municipal solid wastes, the average optimal values (based on dry basis) of
nutrient supplementation for C/N, C/P and C/K have been given as 20 - 30,150 - 300 and 40 100, respectively while the concentrations of Fe, Ni, Se, and W, have been reported to be 100 5000, 5 - 20, 0 - 0.05 and 0.05 - 1 mg/kg, respectively (83).
2.4. End-use technologies for fermentative hydrogen and methane
The huge dependency of world’s energy needs on fossil fuels has resulted in vast
depletion of fossil fuel reserves, coupled with the increased carbon dioxide emission from the
combustion of the fossil fuels. The negative impact of fossil fuel usage could be reduced if
renewable energy carriers including biohydrogen and biomethane are used as replacement or
complement to fossil fuels (84, 85).
25
2.4.1. Attractive qualities of hydrogen and methane as energy carriers
Biohydrogen and biomethane are biofuels that are formed from recently living organisms
called biomass, or their metabolites, as opposed to materials enclosed in geological formation for
a long time before their transformation to fossil fuels. Other biofuels include biodiesel and
bioethanol, among others. Biogas is a colourless gas produced through anaerobic decomposition
of organic material by microorganisms. Depending on the nature of the organic materials and
operating conditions, the gas composition includes methane (bioCH4), carbon dioxide (CO2),
nitrogen (N2), Oxygen (O2), hydrogen sulphide (H2S), and ammonia (NH3) with composition of
40 - 75%, 25 - 40%, 0.5 - 2.5%, 0.1 - 1%, 0.1 - 0.5% and 0.1 - 0.5%, respectively (1, 86).
Upgrading of biogas through cleaning and removal of trace components including water (H2O),
H2S, NH3 and CO2, results in bioCH4 with properties close to natural gas, and hence can be used
for exactly the same end uses as that of natural gas.
Biomethane has various end-uses including heating, cooking, electricity and automotive
fuels as well as serving as feedstock for the production of various products in chemical process
industries (87). The gas could be used for combined heat and power (CHP) on site or be fed-in
into the existing natural gas grid as a substitute or supplement to natural gas. Biomethane can
also be transported by trucks as compressed gas or in liquid form. Regarding hydrogen, it has
high energy content per unit mass (88) as well as having a non-greenhouse gas, water vapour, as
its only combustion product (84, 89-96). Hydrogen has also been shown to be a versatile fuel as
it can be used directly as gas in hydrogen fuel cells (84) and in adapted internal combustion
engines, or stored as liquid or a metal hydride (97) for future purposes. Hydrogen gas can also be
transmitted alone through natural gas pipelines (98) or be mixed with methane to form a more
efficient fuel called hythane. Furthermore, hydrogen can be used in the production of syngas for
electricity generation or diesel production. Besides its application as energy source, hydrogen
can also be used as reactant in hydrogenation process, ammonia, methanol and syngas
production, among others (99).
2.5. Implications of fermentative hydrogen and methane for technological applications
Despite the numerous benefits of application of dark fermentation for hydrogen and
subsequent methane generation, the actualisation of the technology for commercial use is still
limited by process and infrastructural challenges. These barriers must be overcome before the
26
industrial production of fermentative hydrogen and methane can be economical. For example, in
the study carried out in Paper V regarding the feasibility of the development and dissemination
of biogas production in Nigeria, it was found that despite the huge potential for electricity
generation from organic biomass feedstock (Table 2.1), more than 60% of the population does
not have access to the national power supply because they are not connected to the grid system.
People that are even connected to the grid system often experience frequent power outages
(Paper V). The non-existence of green fuels in the energy mix of the country has been attributed
to lack of favourable policy formulation and implementation. The role of the government in
stimulating the market penetration of green fuel technology cannot be overrated. Government
intervention, regulatory mechanisms for green fuel technology, increased awareness level and
capacity building are necessary for the development of green fuel technology in Nigeria (Paper
V).
2.5.1. Process limitations
Although biomethane from biogas has been used in Europe and US on a large scale for various
end-user technologies, including, vehicle fuel and CHP, the gas supply is very low, compared to
the increasing demand for heat, electricity and transport fuels, however, in most developing
countries, the technology is nearly non-existent.
Table 2.1. Theoretical electricity generation from available biomass feedstock in Nigeria (Paper V)
Biomass feedstock
Agriculture crop wastes
Livestock manure
Livestock abattoir waste
Organic MSW
Human waste
Total
Total
potential
biomass
feedstock
(tonnes * 106)
Quantity of
available
biomass
feedstock
(tonnes * 106)
171.86
32.40
0.83
33.12
86.12
324.33
51.56
9.79
0.83
33.12
86.12
181.42
27
BMP of biogas
produced based
on 0.7m3/kgVS
at 35°C
(m3 *109)
20.77
3.69
0.34
13.27
34.29
72.36
Potential electricity
production based
on 3.73 kWh /
m3CH4
kWh x 109)
77.47
13.76
1.27
49.50
127.90
270
Electricity
production
(Terawatt hour,
TWh)
77.47
13.76
1.27
49.50
127.90
270
Regarding fermentative hydrogen, it is still at the research phase due to the process
challenges including low production rates and yields, inefficient substrate conversion, as well as
substrate and product inhibitions. During dark fermentation, more reduced compounds including
ethanol, propionic acid and lactic acids are often produced as fermentation products in place of
hydrogen. Moreover, hydrogen production during fermentation process is associated with the
production of volatile fatty acids, which at very high concentrations could permeate through the
bacterial cells causing cell lyses (100).
2.5.2. Infrastructure barriers
The widespread application of fermentative hydrogen and methane as major energy
carriers in transportation, combined heat and power (CHP), and domestic activities, among
others, will depend on the availability of infrastructure required for its production, storage and
transportation. The production, distribution and storage of fermentative hydrogen and methane
require good infrastructure. Concerning fermentative methane, the infrastructure is available, as
it can be conditioned for transportation through the existing natural gas grid. Alternatively,
stand-alone systems can be used where fermentative methane is used at the site of production for
the generation of heat and electricity. With regard to the infrastructures for hydrogen transport,
distribution and storage are nearly non-existent; the existing infrastructures could also be adapted
for their utilisation in the short-term, prior to the development of the actual infrastructure for
hydrogen distribution (99). Hydrogen and methane could be mixed together to form an improved
gas mixture referred to as bio-hythane, which can then be transported in the existing natural gas
distribution system. However, hydrogen is a light gas with poor specific volume energy density,
compared to other fuels. Its low volumetric energy content limits its storage efficiency. The
current hydrogen storage technique including liquefaction and compression at 350 to 700 bar in
steel tanks are not efficient as it results in energy loss of 10% H2 during compression and 30 to
40% in liquefaction. Alternative storage methods for hydrogen are being developed whereby H2
is bound in storage materials through absorption, adsorption or chemical reactions. The
feasibility of application of fermentative hydrogen in stand-alone systems is, therefore, limited
for distant locations due to challenges involved in storing hydrogen (99).
In view of the increasing greenhouse gas emissions and the challenges of meeting the
increasing energy demand without resource depletion, efforts should be intensified in the
28
establishment of a good and more coherent framework including stable political government,
effective policy, investment programmes, financial support schemes and utilisation strategies,
among others, to accelerate fermentative hydrogen and methane development and ensure its
sustainability. The government needs to create a platform for fermentative hydrogen and
methane technology to have competitive benefits including lower costs, flexibility, excellent
performances, and sustainability, among others, over fossil fuels energy. The development of the
technology could make the energy supply available, adequate, affordable, convenient and
reliable especially for people in developing countries (101).
29
30
CHAPTER 3
Feedstocks for fermentative hydrogen and methane productions
The interesting fact about fermentative hydrogen and methane productions is that all
organic feedstocks could be used as potential feedstock for their production (102); however, the
gas production and quality depend on the composition of the feedstocks.
3.1. Feedstocks suitability for hydrogen and methane productions
Feedstock composition is an important factor that influences the performance of
microorganisms and subsequent gas production and quality, during fermentation process for
hydrogen and methane productions. The proportion of carbon, nitrogen, phosphorus and sulphur
(C:N:P:S) in the feedstock must be such that it provides for the needs of the microorganisms in
terms of energy sources and development of new cells, as the mass ratio of C:N:P:S in
microorganism biomass is approximately 100:10:1:1 (60). The optimal nutrient ratio for carbon,
nitrogen, sulphur and phosphorus (C: N: P: S) has been reported as 500: 15:5:3 and 600:15:5:3
for acidogenesis and methanogenesis, respectively (60, 103). The C:N ratio must not be too high
or too low, or else, the fermentation process can suffer from nitrogen deficiency or ammonia
inhibition, respectively (104, 105). Thus, based on biodegradation, feedstock can be classified
into two major groups, namely, non- lignocellulosic and lignocellulosic feedstock.
3.1.1. Non-lignocellulosic feedstocks
Non-lignocellulosic feedstocks are biomass material that are not lignified and therefore
can be easily degraded by microorganisms. The main compositions of non-lignocellulosic
feedstocks include carbohydrates, proteins and lipids. The hydrolysis rate of carbohydrate is
faster, when compared to lipid and protein (106). Carbohydrate-rich feedstocks contain diverse
sugars including monosaccharides (glucose, fructose, etc), disaccharides (sucrose and lactose)
and polysaccharides (cellulose, hemicelluloses, starch and glycogen). Monosaccharides and
disaccharides are easily and rapidly broken down into VFAs, H2, CO2 and alcohols by anaerobic
microorganisms and could lead to VFA accumulation if it is not controlled (18, 107-109). Most
31
polysaccharides, except starch, are slowly degraded during fermentation process due to their
structure. Starch consists of straight and branched chains of D-glucose units that form a structure
that could be easily degraded by microorganisms. D-glucose isomer occurs naturally in higher
living organisms unlike L-glucose isomer, which is synthesised in the laboratory. Although both
starch and cellulose are polymeric forms of D-glucose units, they differ in the orientation of the
1, 4-glycosidic linkages of the glucose units. In starch, the orientation of all the glucose repeating
units are the same; in other words, the glucose units are connected by alpha linkages (alpha 1, 4
linkages). On the contrary, each repeating glucose unit in cellulose is rotated 180 degrees around
the axis of the polymer backbone chain, forming glycosidic linkages called beta linkages (beta 1,
4 linkages). In addition, cellulose is mostly linear chains of glucose molecules unlike starch with
chains of glucose units that can either be linear, branched or a mix. Consequently, the differences
in the orientations and nature of branching of the glycosidic linkages in starch and cellulose
influence their chemical and physical properties, especially their biodegradation (110).
Regarding protein rich feedstocks, they consist of long chains of amino acids, which are
broken down during fermentation process to release ammonia depending on the temperature or
pH of the medium. High concentration of ammonia could become inhibitory to the fermentation
process, as the dissolution of the ammonia in the fermentation medium raises the pH of the
medium, thereby affecting the activities of the microorganism (16, 106, 111-113). The hydrolysis
rate of protein is slower than carbohydrate and lipid degradation (114). Lipid - rich feedstocks
consist of different forms of fatty acids and glycerol including milk, cheese, meat, palm oil and
vegetable oils (115). During fermentation process, lipids are hydrolysed into long chain fatty
acids (LCFA) which form foam at high concentration due to their surface-active properties (116,
117). Moreover, fats can easily bind to microorganism cells, thereby making the attached cells
prone to wash-out (116). Hence, high concentration of lipids, though very energy-rich, can be
inhibitory to microorganisms during fermentation process (118-121).
3.1.2. Lignocellulosic feedstocks
Lignocellulosic biomass consists of three major components, namely, cellulose,
hemicelluloses and lignin as well as small fraction of ash and soluble substances called
extractives. Cellulose is a polymer of glucose with a crystalline structure of long chains of
glucose forming microfibrils that cannot be easily degraded. In contrast, hemicelluloses could be
32
easily degraded into the component sugars consisting of several five- and six-carbon sugars. In
plants, lignin acts as glue that holds cellulose and hemicelluloses together. Lignocellulosic
feedstocks are often subjected to various physical, thermal, chemical and biological
pretreatments to improve their digestibility (53). During hydrolysis of lignocelluloses, only
cellulosic and hemicellulosic components are converted into fermentable sugars, while lignin
remains unconverted since it is a polymer of recalcitrant aromatic lignin units including phydroxyphenyl , guaiacyl , and syringyl units (122, 123).
3.2. Types of feedstocks for hydrogen and methane productions
Nutritional requirements as well as energy required for several anaerobic bacteria
responsible for hydrogen and methane production could be obtained from organic feedstocks
including agricultural crop wastes, livestock manure, sewage sludge, municipal solid wastes,
organic industrial wastes and wastewater, among others. The feedstocks contain high organic
contents that make them suitable and fermentable (124). In this study, both real and synthetic
substrates supplemented by other nutrients were used for the fermentation processes. In Paper I,
synthetic substrate, which includes acetic acid, butyric acid, propionic acid, methanol and
glucose, mixed at a mass ratio of 3:1:1:1:1, as well as the tested fruit flavour (limonene), were
used. Similarly, in Paper IV, synthetic substrate which contained 20g/L each of glucose, yeast
extract and nutrient broth, as well as synthetic fruit flavours including hexanal, myrcene and
octanol were used. In contrast, real substrates including fruit wastes (orange, apple, banana,
grape and melon) and wheat straw were used in Papers II and III, respectively.
3.2.1. Agricultural crop wastes
Agricultural crop wastes including straws, stalks, and bark generated during farming
operation and processing of crops such as corn, rice, potatoes, and fruits, among others, have the
potential to generate hydrogen and methane (125). Some agricultural crop wastes such as fruit
and potatoes are easily degradable (18), while others such as straws are lignocellulosic with high
contents of cellulose, hemicelluloses and lignin; therefore, they are difficult to digest by
microorganism without some forms of pretreatments to break down their complex structure (20,
53, 126). Lignocellulosic crop wastes with high C/N ratio, such as, apple pomace, corn cobs,
fruit wastes and rice hulls, with average C/N ratios of 48, 98, 40 and 121 (127), respectively;
33
could be mixed with livestock manure, including poultry and pig manure, for effective gas
production during fermentation process (128).
3.2.2. Livestock manure
Animal wastes contain diverse nutrients suitable for the growth of anaerobic
microorganisms; however, the nutrient compositions vary for different animal wastes. Poultry
and pig manures, for example, contain higher concentration of protein than cattle manure, while
cattle manure contain less organic content than manures from pigs and poultry (111, 129, 130).
Most livestock manures have low C/N ratios because of the high nitrogen content. For example,
broiler litter, cattle, laying hens, sheep, swine and turkey litter, have average C/N ratios of 14,
19, 6, 16, 14 and 16, respectively (127). If livestock manure is used as sole feedstock during
fermentation process, there could, therefore, be problems with ammonia inhibition or low gas
production. As a result, co-digestion of livestock manures with other feedstock is usually
employed for efficient fermentation process (131).
3.2.3. Municipal solid waste
Municipal solid waste (MSW) consists of solid wastes from residential, commercial,
institutional and industrial sources with the exception of construction waste. The composition of
the waste depends on factors such as the source of the waste, the standard of living and habits of
residents, as well as sorting method and the climatic conditions. The generated MSW usually has
both organic and inorganic components, and therefore, requires sorting and removal of the
inorganic components including plastics, glass, and metals, as well as construction wastes before
the waste is used as feedstock for anaerobic digestion. The C/N ratios of MSW vary depending
on the sources of the wastes, for instance, the average range of C/N ratios of food wastes, night
soil and sewage sludge are reportedly given as 14 - 16, 6 - 10 and 5 - 16, respectively (127).
3.2.4. Industrial wastes and municipal wastewater
Industrial wastes and municipal wastewater including organic wastewater from
industries, schools, hospitals, government parastatal, sewage sludge from septic tanks or sewers
and slaughterhouse wastes are potential feedstocks for hydrogen and methane productions during
fermentation process. Slaughterhouse wastes contain high concentration of protein and fat, which
34
include suspended organic solids such as grease, hair, feathers, manure and undigested feed.
Although the gas production potential of slaughter wastes is higher than animal manure, the high
protein and fat contents often lead to inhibition due to ammonia and LCFA during fermentation
process (132-134). Therefore, slaughterhouse wastes are often co-digested with feedstock with
lower protein and fat contents to improve the gas production during the fermentation process.
Application of sewage sludge as feedstock for fermentation process is an effective waste
management system as it helps to get rid of pathogenic organisms that could be harmful to
people if the sludge is discarded in the environment. However, the gas production potential of
sewage sludge is lower than that of livestock manure as it contains lower organic content.
Moreover, the sludge could contain substances such as heavy metals and organic pollutants that
could be inhibitory to anaerobic microorganisms since the sludge comes from wastewater that
contains faeces, urine and laundry waste. The average values of typical composition of untreated
wastewater, including total solids, suspended solids, biochemical oxygen demand (BOD5 at
20°C), chemical oxygen demand (COD), total organic carbon (TOC), nitrogen, free ammonia,
phosphorus and alkalinity (as CaCO3), among others, have been reported to be 720, 220, 220,
500, 160, 40, 25, 8 and 100 mg/L, respectively (135).
Generally, feedstocks with high or low C/N ratios could cause thermodynamics
imbalance during fermentation process. When high C/N ratio feedstocks, such as agricultural
crop wastes, are solely used as substrates during fermentation process, the low nitrogen content
is rapidly consumed thereby affecting the natural potential of the process to control the acidity of
the process. In the same vein, when low C/N ratio feedstocks, including livestock manure, are
used exclusively as substrates during fermentation process, excess nitrogen is released causing
increase in the process alkalinity and subsequent thermodynamics shift towards the production of
reduced metabolites including propionate, lactate and ethanol (Equations 2.19 -.2.21) Therefore,
to optimise the fermentation process, mixture of feedstocks with high and low C/N ratios are
often used as substrates.
35
3.3. Inhibitory effect of fruit flavours and volatile fatty acids
3.3.1. Inhibitory effects of fruit flavours
The availability of fruit waste in large quantities with 10-65% of the raw fruits generated
as wastes from the fruit consumption and processing (136); as well as the high content of the
fruit organic matter content; makes it a potential feedstock for anaerobic fermentation process.
Naturally, plants fend off parasitic attack using several defence mechanisms including the
production of a range of volatile organic compounds which often show antimicrobial activity
against a wide range of bacteria, yeasts and moulds. The antimicrobial characteristic of plants
has been known for a long period of time with Chinese using plants in medicinal therapies as far
back as 5000 years ago and Egyptians using plants for food preservation and in mummification
in 1550 BC (137, 138). In flowering plants, fruits produce various volatile organic compounds
including esters, alcohols, aldehydes, ketones, lactones and terpenoids, among others, which
make up their characteristic aroma and flavour characteristics (139). Although the fruit flavours
in plants are present in limited quantity with a range of 0.001 - 0.01% of the fresh fruit weight
(140, 141), their toxicity against microorganism depends on their threshold concentration and
interaction with other compounds. Previous research activities on the effects of fruit flavour have
confirmed the toxicity of fruit flavours against microorganisms (142-147).
The various factors that contribute to the flavour quality of fruits include genetics, harvest
maturity, environmental conditions, post-harvest handling and storage (148, 149). The
antimicrobial effect of fruit flavour compounds can be beneficial regarding the improvement of
food shelf life; the effect can also be detrimental especially during anaerobic digestion of
feedstock containing the flavour compounds since they reduce the bacterial effectiveness (150).
3.3.1.1 Proposed mechanism of flavour toxicity to bacteria
Flavour compounds initiate their antimicrobial activity against bacteria through their
interaction with bacterial cell membrane. The toxicity of a flavour compound during
fermentation process depends on its cell membrane permeability. The hydrophobic nature of
most flavour compounds allows them to interact with the cell membrane and accumulate within
the phospholipid bilayer of the bacteria. The integrity of the cell is lost if the concentration of the
accumulated flavour compound exceeds a tolerable limit (151-159).
36
The proposed mechanism through which a flavour compound develops its toxicity
against bacteria includes: degradation of bacterial cell wall, disruption of cell membrane, leakage
of cell contents, and depletion of proton motive force as well as cytoplasm coagulation. The
effects eventually lead to loss of cell viability and death (159) (Figure 3.1).
Figure 3.1. Proposed Antimicrobial mechanisms of flavour compounds
3.3.1.2. Adaptation of bacterial to toxic environment
Bacteria can adapt to an inhibitory medium for a while as long as the inhibitor
concentration is kept constant. The flavour inhibition could be acute or chronic; it is acute if the
bacterial is exposed to a relatively high concentration within a short time, but it is chronic if the
toxicity results from gradual and long exposure of the bacteria to the toxic compound. The
duration of the chronic toxicity during fermentation process depends on the contact time and the
ratio of the toxic compound to the bacterial population (153, 160).
As bacterial resistance cannot be sustained by itself for a long period of time (153); one
of the objectives of using membrane encapsulated bacteria during the fermentation process in
this research (Papers I and IV) was to reduce the exposure period of bacteria to the flavour
compounds. During the research, the bacteria was protected against the flavour compounds by
using a hydrophilic polyvinylidene fluoride (PVDF) membrane, which allowed water soluble
nutrients to diffuse while repelling the flavour compounds.
37
3.3.2. Inhibition of volatile fatty acids
During fermentation process, different microbial activities take place simultaneously,
with the fermentative bacteria depending on one another activities. Slight process imbalances
due to variation in operational parameters including temperature, retention time, OLR, among
others (161), could result in accumulation of VFAs, the most important intermediates during
fermentation process (162). However, above certain threshold, VFAs could be toxic to the
activities of the fermentative bacteria, as it has been shown that glucose fermentation could be
inhibited at total VFA concentration above 4g L-1 (163, 164). Propionic and butyric acids
inhibition are more toxic than acetic acid inhibition with propionic acid concentrations over 3 g
L-1 having the potential to cause digester failure (165). VFA toxicity occurs as a result of the
increase in the undissociated form of the VFAs, which could diffuse into the bacterial membrane
with consequent dissociation, leading to reduction in the cytoplasmic pH inside the bacteria. In a
single stage digester, there is likelihood of VFA accumulation especially during the digestion of
easily degradable feedstock. The reason for this is that methanogens in low concentration are not
able to metabolise acetate until their number increases exponentially.
3.4. Limiting the inhibitory effects of fruit flavours and volatile fatty acids
Hydrogen and methane productions from organic feedstock during fermentation process
are often not optimized due to substrate and product inhibitions, which affect the efficiency of
the fermentative bacteria.
3.4.1. Control measure to limit the fruit flavour inhibition during fermentation
The hydrophobic nature of most flavour compounds enables them to create partition in
the phospholipids of the bacterial cell membrane, thereby rendering them permeable and leading
to the leakage of cell contents. Hence, it is expected that enclosing bacteria in hydrophilic
polymers may help to reduce the penetration of the flavour compounds since flavour components
have lower hydrophilicity and larger molecule sizes than water. Therefore, in this work (Papers I
and IV), synthetic encapsulating sachets made of hydrophilic PVDF membranes were used to
enclose the fermentative bacteria. Each flat sheet membrane with pore size, thickness and
diameter of 0.1 µm, 125 µm and 90 mm, respectively; was cut and folded into rectangular
dimensions with width and length of 3 and 6 cm, respectively. The membranes were heat-sealed
38
with heating and cooling times of 5.0 and 5.5s, respectively. One side of the membrane sachet
was left open for cell insertion after which the opening was sealed to form a membrane capsule.
The substrates used during the experiments were synthetic media, while the flavour compounds
including limonene, hexanal, myrcene and octanol, were used as the inhibitory substances
(Papers I and IV). The experimental set-ups for the study are shown in Figures 3.2 and 3.3. The
protective effects of the membrane in the media containing the flavour compounds were
investigated by comparing the gas production potentials (bioH2 and bioCH4) of both
encapsulated and free cells. In the limonene containing medium, membrane-encapsulated cells,
compared with the free cells, produced methane faster and were able to survive the effects of the
inhibitory flavour medium at a loading rate of 15g COD L-1d-1 for a longer period. The free cells
failed completely at an OLR of 7.5 COD L-1d-1. In fermentation broth containing myrcene,
octanol and hexanal, methane cumulative yields of 182 ± 15, 111 ± 81 and 150 ± 24 mL/ COD
were obtained from encapsulated cells while no methane production was observed from the free
cells. Regarding hydrogen production, average daily yields of 179 ± 26, 198 ± 16 and 189 ± 17
mL/ COD were produced from encapsulated cells in medium containing myrcene, octanol and
hexanal, respectively, while average yields of 133 ± 77, 88 ± 71 and 68 ± 76 mL/ COD were
produced from the free cells. It was observed that though free cells of bioH2 producing bacteria
were able to produce reasonable amounts of bioH2 regardless of the flavour inhibitors, the
amount of bioH2 produced was less, compared to that of encapsulated cells.
3.4.2. Control measure to limit the volatile fatty acids inhibition during fermentation
In recent times, the VFAs produced during anaerobic fermentation process have been
utilised as a suitable carbon substrate for denitrification and the production of biodegradable
plastics, electricity via microbial fuel cell, biogas, hydrogen as well as lipids for biodiesel (166).
Hence, VFAs are valuable substrates, and their production during fermentation process can be
regulated through filtration for effective fermentation process. Various strategies have been
employed to limit VFA inhibition, including varying operational parameters such as temperature,
pH, organic loading rate, and mixing, with filtration being increasingly used because of its
benefits (167). In this study, VFA inhibition on anaerobic microorganisms during fermentation
was limited by extracting the VFA using PVDF hollow fibre membrane (Paper III). The VFA
permeation was carried out using an externally submerged hollow-fibre membrane module
39
coupled with a mechanically-stirred tank reactor (Figure 3.4). The effects of the membrane
permeation of the VFA on the bioreactor hydrodynamics and hydrogen productions were
investigated. Furthermore, mixing and transmembrane pressure (TMP) across the membrane
bioreactor system were also studied as a function of the operating conditions, while focusing on
the cleaning procedure to assess the feasibility of the filtration process. The results showed an
improved biohydrogen production with a defined VFA permeate extraction, as the biohydrogen
production was observed to restart after VFA extraction. While low recirculation flow rate and
TMP values maximised the permeate flux, the fouling mechanism due to cake layer was
observed to be reversible. The cleaning procedure based on gas scouring and backwashing with
the substrate was defined, while low TMP of 104 Pa was required to achieve a 3 L h-1m-2 critical
flux. Additionally, it was observed that the hydrodynamic properties in the bioreactor were not
significantly modified.
40
Figure. 3.2. Experimental setup for investigating the effect of membrane-encapsulated cells on
fermentative methane production from inhibitory-fruit flavour medium (Paper 1 and IV)
(a) Empty membrane; (b) Encapsulated cells (width position); (c) Encapsulated cells (length position);
(d) Batch process (bioreactors with encapsulated cells); (e) Batch process (bioreactors with free cells)
(f) Semi-continuous process setup:
1- Membrane bioreactor with encapsulated cells, 2- Warm water jacket, 3 – Water heater,
4- Volumetric gas analyser,
5- Purge container, 6 – Feed container,
7 - Peristaltic pump
8 -Data acquisition system
41
Figure 3.3. Experimental setup for investigating the effect of membrane-encapsulated cells
on fermentative hydrogen production from inhibitory-fruit flavour medium (Paper IV)
Figure 3.4. Set-up for concurrent extraction of volatile fatty acids and biohydrogen production (Paper III)
42
CHAPTER 4
Membrane processes for improvement of fermentative hydrogen
and methane productions
Membrane is a thin barrier that controls the permeation of a chemical species using
differences in concentration, pressure and/or electrical potential gradient between the two
compartments they separate (168). The benefits of membrane application include the prevention
of washout of slow-growing bacteria (169) and the protection of bacteria from toxic effects of
substrate and product inhibitions.
4.1. Membrane classification
The distinctiveness of membrane depends on its properties including pore size and shape,
symmetry, and the membrane materials. Regarding membrane materials, membranes can consist
of organic or inorganic materials (170). Organic membranes are produced using various
polymers such as cellulose acetate, polysulfone, polyamide, polyethylene (PE), polypropylene
(PP), polytetrafluoroethylene (Teflon PTFE), polyethersulfone and polyvinylidene fluoride
(PVDF); however, inorganic membranes refer to membranes made of materials such as ceramic,
carbon, silica, zeolite, oxides (alumina and zirconia) and metals such as palladium, silver, among
others (171). Most membranes are usually made of hydrophobic polymers owing to their
qualities including chemical resistance, low swelling, low fouling tendency and good separation
performance. Common hydrophobic membranes that are often employed are PVDF, PTFE, PE,
PP and zeolites (172). However, hydrophobic membranes are usually hydrophilised to allow
water permeation during fermentation process.
Membranes are also classified according to their filtration techniques including
microfiltration, ultrafiltration, nanofiltration and reverse osmosis, depending on their pore size
distribution (Papers I, III and IV). Membranes used for microfiltration generally have pore size
of approximately 0.03 to 10 microns, with an operating pressure between 100 and 400 kPa while
the range of membrane pore size for ultrafiltration membranes is approximately 0.002 to 0.1
microns, with an operating pressure between 200 and 700 kPa. Nanofiltration membranes have
43
pore size between 0.0001 to 0.01 microns with operating pressure between 600 kPa and 1000
kPa. Reverse osmosis filtration technique is able to retain almost all molecules, except water, due
to size of the pore which ranges from 0.0001 to 0.001 microns (173, 174).
There are several types of membrane module configurations, which are either planar or
cylindrical in geometry. The planar or flat sheet configurations include plate-and-frame and
spiral-wound modules while cylindrical configurations include hollow fibre and tubular modules.
Hollow fibre and tubular modules have shell and tube configurations, but hollow fibre modules
have greater number of tubes with smaller diameters than the tubes in tubular modules. The
tubular is often applied to enhance turbulent flow. The fibres in hollow fibre modules may
consist of several hundreds to over 10,000 fibres with the fibres bundled together longitudinally,
potted in a resin on both ends and encapsulated in a pressure vessel. The mode of operation of
the hollow fibre module could be inside-out or outside-in. In inside-out the feed flow enters
through the centre of the fibre (lumen) and is filtered radially through the fibre wall, while
outside-in operation, the feed flow enters from outside the fibre into the inside of the fibre. The
fibre modules have the benefit of high filtration surface area per unit volume; however, flux
maintenance and membrane fouling are key issues during filtration process (175). The spiralwound module consists of flat sheet membrane wound around a central perforated tube. It is
often used for cross-flow filtration. The flat sheet modules consist of flat and thin-film composite
sheets with a thin layer being supported on a thicker layer that has wider pore. They are cheap
and usually disposable.
As reviewed in Paper VI, anaerobic membrane bioreactors (AnMBRs) could be
configured in three different ways: internally submerged membrane (ISMBR), externally
submerged membrane (ESMBR) (Paper VI) and external cross-flow membrane (ECMBR)
bioreactors (172, 176). In ISMBR, the membrane is placed directly inside the fermentor,
however, in ESMBR, the membrane is placed in a separate container different from the
fermentor. The benefit of ESMBR is that it is easier to clean than ISMBR, but it often requires
pump for sludge recirculation during the fermentation process (177). Unlike the submerged
membrane bioreactors, the ECMBR is operated in a cross-flow mode in which the fermented
broth is pumped parallel to the membrane surface. Although ECMBR has the benefit of reducing
the frequency of fouling formation, sustaining the continuous operation is energy intensive (177).
44
4.1.1. Application of PVDF membrane in fermentative hydrogen and methane productions
PVDF is a commonly used membrane in microfiltration and ultra-filtration membrane
processes because of its low price and excellent qualities including high hydrophobicity,
mechanical strength, thermal stability, as well as, chemical and ultra violet resistance (178). The
membrane is chemically stable to a wide range of chemical compounds such as inorganic acids,
oxidants, halogens, aromatic, aliphatic and chlorinated solvents. The PVDF membrane is a semicrystalline polymeric membrane consisting of both crystalline and amorphous phases with
crystalline part responsible for its excellent thermal stability, while the amorphous part is
responsible for the flexibility of the membrane (172). The macromolecular straight chain of
PVDF is enveloped by fluorine and hydrogen atoms (-CH2-CF2-), with fluorine atoms, being a
high electronegative element, providing high dissociation energy of the C-F bonds, which makes
the membrane to have higher thermal stability compared to other polymeric membranes (179).
PVDF has a glass transition temperature (Tg) of around -39⁰C, a melting point temperature of
around 160⁰C and a thermal decomposition temperature of above 316⁰C.
PVDF also exhibits piezoelectric properties which make it to be valuable as sensor and
actuator materials. However, the intrinsic hydrophobic nature of PVDF makes it prone to organic
fouling and low wettability with high resistance to water flow. As a result, several membrane
modification techniques including blending, surface coating, irradiation grafting and plasma
modification, are used to incorporate hydrophilicity into hydrophobic PVDF membranes to
enhance their performances (180). Considering the excellent quality of PVDF membranes, this
study, therefore, used hydrophilic PVDF membranes in all the experiments investigated. Two
different membrane configurations, including flat sheet and hollow fibre modules were employed
in the membrane encapsulation and VFA extraction, respectively.
4.2 Influence of membrane permeability on membrane performance
The effectiveness of membrane for VFA permeation and protection against inhibitory
effects of substrate and product inhibition during fermentation process, which was the focus of
this research, depends on the permeability of the membrane. The membrane permeation of
molecules
is
influenced
by
the
nature
of
the
membrane
materials
(pore
size,
hydrophobicity/hydrophilicity, free volume and filler particles) and solubility of the permeants
45
(molecular size, concentration, among others) (181-183). The ability of a membrane to regulate
the permeation of various molecules through it is an important feature that is employed in
separation processes. The permeation process can either follow solution-diffusion model where
the permeants dissolve and diffuse through the membrane, or pore flow model where the
permeants pass through the membrane pores. In this study, water permeability of the PVDF
membrane was determined using distilled water. An average value of 0.048 mL/min of pure
water permeability was obtained for the PVDF membrane.
4.3. Encapsulation technology for cell retention and inhibition control
Encapsulation of bacterial cell is a technique used for restricting bacterial mobility within
a polymeric semi-permeable membrane. In encapsulation, the microorganisms are restricted by
the membrane walls, however, the cells move around within the membrane (184, 185). In this
technique, the membrane is semi-permeable as it allows substrates and nutrient to pass through it
depending on the membrane pore size, while retaining the microorganism. The benefits of
encapsulation technique is that anaerobic microorganism can be retained in the digester longer
without being washed out during continuous operation, thereby giving the cells sufficient time to
grow and degrade the substrate supplied (186). Additionally, the microorganisms could be easily
recovered at the end of the fermentation process, thereby reducing the purification cost (187).
Encapsulation also encourages the use of small reactor system since the concentration of the
microorganism is higher than in free cells. It also acts as a protective barrier for the
microorganism against the effect of substrate and product inhibitions (188). In this study (Papers
I and IV), encapsulation technique was used to confine the movement of the bacteria within the
walls of flat sheet PVDF membrane (Figure 4.1). The technique, however, has some limitations
which include occasional cell leakage and inefficient diffusion of substrate to the microorganism
in the membrane as well as the diffusion of product away from the microorganism and out to the
external medium (187, 189).
46
Figure. 4.1. Cell Encapsulation strategy used during the research (Papers I and IV)
Besides encapsulation, other methods of restricting bacterial movement either completely
or to a small limited region include adsorption, covalent binding, entrapment and crosslinking.
Adsorption is a reversible immobilisation technique as it is based on weak binding forces
between the support and the microorganisms. Due to the weak binding forces, which may
include van der Waals forces, ionic or hydrogen bonds, high rate leakage and unstable binding
are common occurrences (189). Adsorption is prone to changes in environmental conditions, for
example, pH, temperature and ionic strength, among others. Unlike encapsulation, entrapment is
an irreversible immobilisation technique in which microorganisms are entrapped in a support
matrix or inside a membrane. Cross-linking is also an irreversible method of immobilisation that
is support-free but it involves the joining of microorganism to one another to form a large
structure (190). The method is cost effective, and it increases the volumetric activity of the
microorganisms. Covalent binding involves the formation of a covalent bond between the
microorganisms and the support material. In this technique, microorganism leakage is minimized
as a result of the strong linkage between the support and the organisms (186, 187).
47
4.4. Application of hollow fibre membrane configuration for VFA permeation
Accumulation of volatile fatty acids (VFA) during dark fermentation is a major challenge
as it has been attributed, among other factors, to inhibition of microbial cell growth and substrate
consumption, leading to low hydrogen yield (191). Nonetheless, different kinds of separation
techniques have been applied to separate VFAs from the fermentation broth, such as ion
exchange (192), electrodialysis (193, 194), adsorption (195), liquid-liquid extraction (196), and
membrane separation (197), but most of the separation techniques are expensive and inefficient.
However, with improvement in membrane structure and qualities, membrane costs have become
inexpensive and affordable (198). Moreover, membranes have the advantage of coupling VFA
separation with biomass retention during anaerobic fermentation process, thereby maximizing
bioenergy production.
In hollow fibre membrane bioreactor, optimising module packing density is crucial as it
reduces the number of fibre modules as well as the size of the membrane tank, hence, reducing
the capital and operation costs of the reactor system. When submerged hollow fibre membrane is
used during anaerobic digestion process, the filtration is often in outside-inside mode. In outsideinside filtration mode, an uneven distribution of the transmembrane pressure (TMP) may develop
along the module fibres, as increased fibre length and ratio of outside diameter to inside diameter
along with reduced fibre inner diameter, could increase the non-uniformity of the flux
distribution. The calculated TMP values based on the suction pressure at the outlet could be
different from the actual TMP along the fibres, thereby affecting the accuracy of the membrane
permeability (199).
4.5. Limitations of membrane technology: Membrane fouling and cost
The resistance to fluid flow through the membrane during filtration process is often due
to membrane fouling, which is a term that is used to describe the loss of membrane throughput.
Generally, fouling occurs when particulate, colloidal or soluble materials are deposited inside
membrane pores or surface. Membrane fouling is a major barrier to membrane application in
fermentation processes as it is associated with flux or permeate flow reduction, low permeate
quality and increased operational costs due to increased energy consumption (200). Normally,
membrane bioreactors (MBRs) are designed for the maintenance of constant flux but when
membrane fouling sets in, it causes increase in TMP and decrease in flux to the point that
48
membrane cleaning or replacement is required. Membrane fouling is influenced by factors such
as sludge characteristics, operational parameters and membrane qualities (201). Although,
membrane fouling cannot be entirely avoided during membrane filtration process, the frequency
of its occurrence could be reduced through physical cleaning such as relaxation and backwashing
or chemical cleaning. Chemical cleaning of membrane is more effective in removing membrane
fouling than physical cleaning, but frequent use of chemical cleaning can damage the membrane
and shorten the membrane life-time (202-204). Previously, membrane cost was part of the
barrier to the application of membrane technology, but extensive research on membrane
improvement has resulted in cheap and affordable membrane in recent times (170, 198).
However, operating costs associated with membrane fouling abatement is still the main barrier to
the application of membrane technology (201).
4.6. Implications of membrane applications in this study
The applications of membranes in this study was done in two parts; the first part, which
focused on reduction of substrate inhibition, involved the investigation of the effects of the
PVDF membrane encapsulation techniques on the bioH2 and bioCH4 production from fruit waste
media. The results of methane production from the thermophilic batch and continuous
fermentation of nutrient media with limonene, myrcene, octanol and hexanal as fruit flavours
showed that membrane encapsulated, compared to free cells, produced methane faster with
limonene as flavour. The encapsulated cells were able to survive the effects of the inhibitory
activity of limonene flavour medium at a loading rate of 15g COD L-1d-1 for a longer period even
after the free cells had completely failed at an OLR of 7.5g COD L-1d-1 (Paper I). Likewise, with
myrcene, octanol and hexanal as fruit flavours, bioCH4 cumulative yields of 182 ± 15, 111± 81
and 150 ± 24 mL/g COD, respectively, were obtained from encapsulated cells while no CH4
production was observed from free cells during batch fermentation process. About 60%
membrane protective effect against myrcene, octanol and hexanol inhibitions was obtained
(Paper IV).
Regarding bioH2 production, average daily yields of 68 ± 76, 133 ± 77, 88 ± 71 mL/g
COD were produced from the free cells with hexanal, myrcene and octanol, respectively, while
average yields of 189 ± 16, 179 ± 26 and 198 ± 17 mL/g COD were produced from encapsulated
cells containing hexanal, myrcene and octanol respectively (Paper IV). This indicated that
49
membrane encapsulated cells, compared to free cells, produced higher daily yields of 94, 30 and
77% for hexanal, myrcene and octanol as flavours, respectively. It was observed that though free
cells of bioH2 producing bacteria were able to produce reasonable amount of bioH2 regardless of
the flavour inhibitors, the amount of bioH2 produced was less compared to that of encapsulated
cells (Paper IV).
The final part of this study, which was aimed at reducing product inhibition, involved the
study of the effects of membrane permeation of volatile fatty acids (VFAs) on the bioreactor
hydrodynamics in relation to bioH2 production. The investigation revealed that low
transmembrane pressure of 104 Pa was required to achieve a 3 L.h-1m-2 critical flux with
reversible fouling mainly due to cake layer formation, while bioH2 production was observed to
restart after VFAs removal (Paper III). The results from this study suggest that membrane-based
techniques could actually improve bioH2 and bioCH4 productions from fermentation media with
substrate and product inhibitions.
50
CHAPTER 5
Bioreactor hydrodynamics for fermentative hydrogen and methane
productions
5.1. Ideal and real reactors
Ideal reactors are model systems analogous to effective reactors for which transport and
mixing processes can be exactly described mathematically. On the contrary, the transport and
mixing processes in real reactors can only be roughly known due to different flow effects that
could occur, causing dead zones (stagnant region) and channelling (short-circuits or bypassing)
(Figure 5.1). Dead zone is a region with low or no mixing in a mixed reactor, while channelling
occurs when a portion of the reactants is transported to the output in a shortest time possible
without taking part in the reaction process (205). The occurrence of dead zone and channelling in
a reactor affects the reactor performance and reduces the product yield.
Figure 5.1. Fluid flow in non-ideal reactor
51
5.2. Mixing in bioreactors
Mixing is a physical process, which is designed to reduce non-uniformities in fluids
through the prevention of gradients in the fluid properties including temperature and
concentration, among others. Mixing also facilitates physical and chemical uniformity of
digested sludge by preventing the formation of dead zones due to particle deposition. It prevents
gas bubbles being trapped in the fermentation broth. However, mixing should not be strong as it
can cause shearing and wash-out of the microorganisms, thereby, leading to loss of
microorganism activity (75, 206, 207). For effective performance during fermentation process,
bioreactors are often equipped with impellers to facilitate uniform mixing and good contact
between the substrate and microorganisms, as well as to prevent sedimentation of the substrate
(73, 74). Apart from mechanical mixers, mixing can also be carried out through gas injection or
recirculation of digested slurry with the aid of pressure pumps. However, for effective mixing to
occur, the fluid distributed by impeller or other techniques must pass over the entire bioreactor in
a good time. Moreover, the velocity of the fluid leaving the mixer must be able to take the fluid
to the extreme part of the reactor. Effectiveness of mixing could be evaluated using two
parameters, including mixing time and mean circulation time (208, 209).
5.2.1 Mixing and mean circulation times
Mixing time (tm) is the time required to attain homogenisation to the molecular scale.
However, since molecular scale measurement could not be determined experimentally, terminal
mixing time is often determined. Terminal mixing time is the time required to achieve a given
degree of homogeneity on the scale of observation, starting from the completely non-integrated
state. Experimentally, measurement of mixing time is done through tracer technique, whereby
the tracer (usually a pulse of electrolyte solution) is injected into the reactor. The mixing time is
taken as the time at which the tracer concentration (Ci) at the measurement location has almost
attained the expected final mean tracer concentration (Cf). Mathematically, the mixing time is
defined as the time from tracer addition to the time when the tracer concentration is given as
(Equation 5.1):
Cf − Ci
=m
Cf
5.1
52
where m is the maximum acceptable absolute value of the relative deviation of the mix. At the
beginning of the experiment, m = 1.0, while at the end of the experiment when complete
homogeneity has been attained, m = 0. The lowest value of m at the end of the experiment which
can be accurately measured is 0.05 (5%). Mixing time can be identified from the response curve
as the time after which the concentration of the tracer (Ci) differs from the final concentration
(Cf) by less than 5% of the total concentration difference (Cf - Ci). The mean circulation time is
the main time interval between two consecutive peaks of maximum concentration of tracer on
the response curve (Figure 5.2) during the measurement of residence time distribution.
Figure 5.2. A typical response curve of Mixing and mean recirculation times
5.3. Residence time distribution measurement
In real reactors, fluid molecules spend different times before their exit from the reactor,
with some molecules finding their way to the exit immediately as they enter the reactor, while
other molecules stay for a long time before their exit from the reactors. Residence time
53
distribution (RTD) of a reactor is a probability distribution function that describes the duration
that a fluid molecule spends inside the reactor (210, 211). It is usually expressed as an exit age
distribution, E(t) (Equation 5.1).
∫
∞
0
E (t ) dt =1
5.1
The average residence time for all the fluid molecules is given as (eq 5.2):
∞
t = ∫ t.E (t ) dt
5.2
0
In the absence of dead zones and channelling, the average residence time calculated from RTD
will be equal to the hydraulic retention time (HRT) calculated from the total reactor volume and
the volumetric flow rate of the fluid. Measurement of residence time distribution of fluid
molecules in a reactor provides insight regarding the mixing performance of the reactor and
consequently, facilitates the effective reactor design or retrofit that allows for a high and
sustainable organic loading rate, a short hydraulic retention time and maximal hydrogen and
methane yields (212).
5.4. Membrane filtration
Membrane filtration can be carried out using one or two techniques which include deadend and cross-flow filtration depending on the direction of the flow of the feed suspension that is
being filtered. In dead-end filtration, the direction of the feed suspension to be filtered is
perpendicular to the membrane surface, while, in cross-flow filtration, the direction of feed
suspension is tangential or parallel to the membrane surface. Dead-end filtration is effective for
separating out particles with low concentration from feed suspension, and it is often carried out
in batch mode where the mass of the deposited material or retentate increases until the material
can no longer be deposited again. Hydraulic resistance of the retentate increases with filtration
time, which results in the decrease of the filtration rate across the membrane. Cross-flow
filtration is usually operated using multi–sheet, hollow fibre or tubular membrane configurations
(213, 214). Darcy’s law of filtration expresses membrane performance during filtration in terms
of the filtrate flux (J), which is the volume of the filtrate that passes through the unit membrane
54
area in unit time (Equation 5.3), where ∆P, µ and RT represent the applied pressure
(transmembrane pressure), filtrate viscosity and total resistance to the fluid flow (215). The total
resistance, based on the resistance-in-series model, is a combined resistance of three resistances,
including intrinsic membrane resistance (Rm), reversible resistance due to cake layer or external
fouling (Re), and internal fouling resistance due to irreversible adsorption and pore plugging (Ri)
(Equation 5.4). Darcy’s law highlights that the permeate flux through a porous membrane is
directly proportional to the transmembrane pressure (TMP) and the membrane area, but is
inversely proportional to the membrane resistance due to fouling and to feed viscosity (216). The
expression for the TMP is given by Equation 5.5, where Pf, Pr and Pp are feed, retentate and
permeate pressures, respectively.
J=
∆P
µ RT
=
TMP
µ RT
5.3
RT = Rm + Re + Ri
TMP =
5.4
Pf + Pr
− Pp
2
5.5
The filtrate flux through the membrane materials decreases with the occurrence of
membrane fouling. Hence, flux or filtration efficiency, is directly affected by membrane fouling
with a consequent decrease in system productivity and increase in operating cost. The rate of
membrane fouling could be reduced by carrying out filtration process below the critical flux and
by simultaneously maintaining high shear rate through velocity gradient or gas sparging close to
the membrane. Membrane fouling can also be reduced by using appropriate membrane
configuration and modules, as in the case of hollow fibre membrane modules (217). In this
study; during the investigation of the effects of membrane permeation of VFAs on bioreactor
hydrodynamics and bioH2 production (Paper III), the experimental set-up was designed to enable
both gas scouring (bubbling of CO2 through membrane surface) and back-washing with permeate
to control membrane fouling.
55
In the first study of the cleaning procedures in Paper III, CO2 was injected for 5 minutes
after a filtration step of 15 minutes using the digestate; then, the same cycle was repeated. In the
second study (Paper III), the same procedure was applied, but CO2 cleaning procedure was
replaced by permeate backwashing. Backwashing was obtained using the reversible flow of the
withdrawal peristaltic pump, which, however, presented the drawback of using a fraction of the
filtrate for backwashing. The fouling was studied by following the permeate flow rate over time
as a function of the rotation speed of the withdrawal pump. The effectiveness of cleaning was
assessed through the evolution of the permeate flow rate over several filtration-cleaning cycles.
The results showed that backwashing was slightly more efficient for recovering the initial
density flux (4.45 L h-1m-2), since the permeate flux after backwashing remained above 4.40 L h1
m-2, while it was about 4.30 L h-1m-2after CO2 injections. This implied that fouling was probably
reversible, but the cleaning remained incomplete using only CO2 injections. Moreover, the
results also revealed that low transmembrane pressure of 104Pa was required to achieve a 3 L h-1
m-2 critical flux with reversible fouling mainly due to cake layer formation, while bioH2
production was observed to restart after VFAs removal (Paper III).
56
CHAPTER 6
Conclusions and Future Work
6.1. Conclusions
Fermentative hydrogen and methane are energy carriers that are neutral with regard to CO2
emissions and they could, therefore, play a significant role in the development of a sustainable
global climate. Although utilization of the fermentative hydrogen and methane as separate
energy carriers offers numerous benefits including low fossil-fuel dependency, effective waste
management system, reduction of greenhouse gas emission and energy accessibility, the usage of
the mixture of hydrogen and methane has the potential of providing more benefits than
individual utilization. The mixture of hydrogen and methane as fuel offers synergistic benefits of
the two energy carriers, in the sense that, hydrogen complements the drawbacks of methane that
affect its combustion efficiency, including narrow flammability range, high ignition temperature,
and slow burning speed. Presently, the market of fermentative hydrogen as a fuel is non-existent,
while that of fermentative methane is still at the beginning stages even with the commercial
applications in some advanced countries. The major barriers associated with the widespread
applications of fermentative hydrogen and methane as fuels include low gas yields and
availability of required infrastructure for gas production (Paper V), distribution and storage. This
research has focused on how to contribute to the facilitation of widespread application of
fermentative hydrogen and methane through yield improvement.
The low yields of hydrogen and methane productions during fermentation process are partly
due to substrate and product inhibitions. This study, therefore, investigated the potential of
employing membrane technology and varying operational parameters in yield enhancement of
fermentative hydrogen and methane. The major results from this study are summarized as
follows:
•
Enclosing fermentative bacteria in hydrophilic PVDF membrane prevented bacterial
wash-out as well as protected the bacteria against harsh environment such as inhibitory
flavour medium. Compared with the free cells, membrane-encapsulated cells produced
methane faster and were able to survive the effects of the inhibitory flavour medium at a
57
loading rate of 15 g COD L-1d-1 for a longer period even after the free cells had
completely failed at an OLR of 7.5 g COD L-1d-1.
•
With myrcene, octanol and hexanal as fruit flavours, CH4 cumulative yields of 182 ± 15,
111± 81 and 150 ± 24 mL/g COD, respectively, were obtained from encapsulated cells,
while no CH4 production was observed from free cells during batch fermentation.
Regarding bioH2 production, average daily yield of 68 ± 76, 133 ± 77, 88 ± 71 mL/g
COD were produced from free cells with hexanal, myrcene and octanol, respectively,
while average yield of 189 ± 16, 179 ± 26 and 198 ± 17 mL/g COD were produced from
encapsulated cells containing hexanal, myrcene and octanol, respectively. It was
observed that though free cells of bioH2 producing bacteria were able to produce
reasonable amounts of bioH2 regardless of the flavour inhibitors, the amount of bioH2
produced was less, compared to that of encapsulated cells.
•
Meanwhile, a study conducted to evaluate effects of hydraulic retention time (HRT) and
fruit mixing on bioH2 production from single and mixed fruits indicated that there was no
statistically significant effect of the interaction of HRT and fruit mixing on bioH2 yield.
However, it was observed that there was an improvement in cumulative bioH2 yield from
all the feedstocks when HRT was 5 days, while fruit mixture with equal fruit proportion
produced the highest cumulative bioH2 yield of 513mL/g VS (30% of the theoretical
yield).
•
The investigation of the effect of membrane permeation of VFAs on the hydrodynamics
in the bioreactor in relation to bioH2 production from glucose and straw as separate
feedstock in a submerged anaerobic bioreactor revealed that low transmembrane pressure
of 104Pa was required to achieve a 3 Lh-1m-2 critical flux with reversible fouling mainly
due to cake layer formation, while bioH2production was observed to restart after VFAs
removal.
The results from this research suggest that membrane-based techniques and varying
operational parameters could improve the yields of hydrogen and methane productions from
fermentation media with substrate and product inhibitions. It is, therefore, expected that
application of the knowledge from this research along with the provision of government support
58
scheme in various countries will facilitate the widespread commercial production and utilisation
of fermentative hydrogen and methane in terms of the economic viability of the technology.
6.2. Future Work
Based on the results from this research work, research activities stated below will be the
direction of future work for further development of fermentative hydrogen and methane
technology.
•
During the research, hydrogen and methane fermentation were carried out separately.
However, to obtain higher yield and determine the overall substrate conversion efficiency
for energy utilisation, the partially digested sludge consisting of VFAs still need to be
utilised to extract additional energy from it. The future work will therefore involve
combining hydrogen and methane productions in a two-stage system where VFAs
extracted or the digested sludge from stage one will be used as substrates for the second
stage. It will therefore be possible to evaluate accurately the economic feasibility of the
process.
•
Part of the research work in this thesis involved investigation of the hydrodynamic and
yield effects of VFA permeation using external immersed membrane module, it will be
interesting to study the effects using internally submerged membrane module
•
In the experiment conducted to investigate the effect of fruit flavour inhibition on the
bacterial activities as linked to hydrogen production, it was observed that hydrogen
production from encapsulated cells remained relatively constant for three days, after feed
input and effluent withdrawal had stopped. The hydrogen production, however, declined
after the three days. Therefore, it may be interesting to investigate if intermittent loading
of feedstock with inhibitors can improve the microorganism adaptation to the inhibitory
compounds, possibly giving the microorganisms enough time to neutralise the effect of
the inhibitors without affecting the hydrogen yield.
•
Extensive studies with modelling should be done on the membrane permeability of fruit
flavour on bacteria cell membrane, with the aim of determining the direct relationship
59
between the concentration of the inhibitory compounds and the survival duration as well
as the surviving tactics of the microorganisms. Furthermore, since the production of
hydrogen is associated with other reduced metabolites, other than acetic acid, which has
the highest theoretical hydrogen yields; modelling approach will also be applied to
establish the direct relationship among the secondary metabolites and feedstock
compositions as well as process parameters.
•
Further studies will also be conducted on how to improve the efficiency of the microbial
activities. From previous research studies, it has been found that sole pretreatment of seed
sludge could not sustain the flourish and proliferation of the hydrogen producing
microorganisms for a long period of time, as the hydrogen production decreased after
running the experiment for a while. It will, therefore, be interesting to find the optimal
factor parameters that could be combined for effective long-term maintenance of
hydrogen producing bacteria during fermentation process.
•
During the experiment on the effects of fruit flavour inhibition on hydrogen and methane
production, synthetic fruit flavour compounds were used. It will be interesting in the
future to use the flavour compounds extracted directly from the fruit wastes for the
investigation.
•
Although this study showed that membrane encapsulated cells could improve the yields
of fermentative hydrogen and methane productions, the membrane could, however,
restrict the growth of the anaerobic microorganism during the doubling period. Therefore,
it will be interesting to study the effects of the membrane wall restriction on the growth
of the microorganisms in relation to the yields of the fermentative hydrogen and methane
production.
60
References
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
Weiland, P., Biogas production: current state and perspectives. Applied Microbiology and
Biotechnology 2010, 85, 849-60.
De-Baere, L., Anaerobic digestion of solid waste: state-of-the-art. Water Science and
Technology 2000, 41, 283-290.
Mountfort, D. O.; Asher, R. A., Changes in proportions of acetate and carbon dioxide
used as methane precursors during the anaerobic digestion of bovine waste. Applied and
Environmental Microbiology 1978, 35, 648-654.
Griffin, M. E.; McMahon, K. D.; Mackie, R. I.; Raskin, L., Methanogenic population
dynamics during start-up of anaerobic digesters treating municipal solid waste and
biosolids. Biotechnology and Bioengineering 1998, 57, 342-355.
Karakashev, D.; Batstone, D. J.; Angelidaki, I., Influence of environmental conditions on
methanogenic compositions in anaerobic biogas reactors. Applied and Environmental
Microbiology 2005, 7, 331-338.
Han, S. K.; Kim, S. H.; Kim, H. W.; H.S, S., Pilot-scale two-stage process: a combination
of acidogenic hydrogenesis and methanogenesis. Water Science and Technology 2005,
52, 131-138.
Hallenbeck, P. C.; Benemann, J. R., Biological hydrogen production: Fundamentals and
limiting processes. International Journal of Hydrogen Energy 2002, 27, 1185-1193.
Sinha, P.; Pandey, A., An evaluative report and challenges for fermentative biohydrogen
production. International Journal of Hydrogen Energy 2011, 36, 7460-7478.
Stern, J. E.; Elliot, D., The Ethics of Scientific Research. University Press of New
England: London, 1977.
Harris, J., Scientific research is a moral duty. Journal of Medical Ethics 2005, 31, 242248.
Gerardi, M. H., The microbiology of anaerobic digesters. John Wiley and Sons, Inc. New
Jersey, 2003.
Bryant, M., Microbial methane production: theoretical aspects. Journal of Animal Science
1979, 48, 193-201.
Smith, P., The microbial ecology of sludge methanogenesis. Developments in Industrial
Microbiology 1966, 1966, 156-161.
McInerney, M. J., Anaerobic hydrolysis and fermentation of fats and proteins. In Biology
of Anaerobic Microorganisms, Zehnder, J. B., Ed. John Wiley and Sons: USA, 1988; pp
373-415.
Schink, B., Energetics of syntrophic cooperation in methanogenic degradation.
Microbiology and Molecular Biology Reviews 1997, 61, 262-280.
Schurer, A. A.; Nordberg, Ammonia, a selective agent for methane production
bysyntrophic acetate oxiation at mesophilic temperature. Water Science and Technology
2008, 57, 735-740.
Karakashev, D.; Batstone, D. J.; Trably, E.; Angelidaki, I., Acetate oxiation is the
dominant methanogenic pathway from acetate in the absence of Methanosaetaceae.
Applied and Environmental Microbiology 2006, 72, 5138-5141.
61
18.
19.
20.
21.
22.
23.
24.
25.
26.
27.
28.
29.
30.
31.
32.
33.
34.
35.
Boullaguii, H.; Torrijos, M.; Gordon, J. J.; Moletta, R.; Cheik, R. B.; Touhami, Y.;
Delgenes, J. P.; Hamid, M., Two-phases anaerobic digestion of fruit and vegetable
wastes: Bioreactor performance. Biochemical Engineering Journal 2004, 21, 193-197.
Pohland, F. G.; Gosh, S., Developments in anaerobic stabilization of organic wastes-the
two phase concept. Environmental Letters 1971, 1, 255-266.
Parawira, W.; Read, J. S.; Mattiasson, B.; Bjornsson, L., Energy production from
agricultural residues: high methane yields in pilot-scale two-stage anaerobic digestion.
Biomass and Bioenergy 2008, 32, 44-50.
Schober, G.; Schäfer, J.; Schmid-Staiger, U.; Trösch, W., One and two-stage digestion of
solid organic waste. Water Research 1999, 33, 854-860.
Fannin, F. F.; Biljetina, R., Reactor design. In Anaerobic digestion of biomass,
Chynoweth, D. P.; Isaacson, R., Eds. Elsevier Applied sciences: London, 1984.
Sakar, S.; Yetilmezsoy, K.; Kocak, E., Anaerobic digestion technology in poultry and
livestock treatment- a literature review. Waste Management and Research 2009, 27, 3-18.
Yamane, T.; Shimizu, S., Fed-batch techniques in microbial processes. Advances in
Biochemical Engineering/Biotechnology 1984, 30, 147-194.
Longobardi, G. P., Fed-batch versus batch fermentatioon. Bioprocess Engineering 1994,
10, 183-194.
Eberle, U.; Muller, B.; von-Helmolt, R., Fuel cell electric vehicles and hydrogen
infrastructure status. Energy and Environmental Science 2012, 5, 8780-8798.
Deutzmann, J. S.; Sahin, M.; Spormann, A. M., Extracellular enzymes facilitate electron
uptake in biocorrosion and bioelectrosynthesis and bioelectrosynthesis. MBiosphere
2015, 6, e00496-15.
Yates, M. D.; Siegert, M.; Logan, B. E., Hydrogen evolution catalyzed by viable and
non-viable cells on biocathodes. International Journal of Hydrogen Energy 2014, 39,
16841-16851.
Marshall, C. W.; Ross, D. E.; Fichot, E. B.; Norman, R. S.; May, H. D., Electrosynthesis
of commodity chemicals by an autotrophic microbial community. Applied and
Environmental Microbiology 2012, 78, 8412-20.
Siegert, M.; Yates, M. D.; Call, D. F.; Zhu, X.; Spormann, A.; Logan, B. E., Comparison
of nonprecious metal cathode materials for methane production by
electromethanogenesis. ACS Sustainable Chemistry and Engineering 2014, 2, 910-917.
Cheng, S.; Xing, D.; Call, D. F.; Logan, B. E., Direct biological conversion of electric
current into methane by electromethanogenesis. Environmental Science and Technology
2009, 2009, 3953-3958.
Chen, X.; Guan, C.; Xiao, G.; Du, X.; Wang, J.-Q., Syngas production by high
temperature steam/CO2 coelectrolysis using solid oxide electrolysis cells. Faraday
Discussions 2015.
Joyner, A. E.; Winter, W. T.; Godbout, D. M., Studies on some characteristics of
hydrogen production by cell-free extracts of rumen anaerobic bacteria. Canadian Journal
of Microbiology 1977, 23, 346-353.
Nandi, R.; Sengupta, S., Microbial production of hydrogen: An overview. Critical
Reviews in Microbiology 1998, 24, 61-84.
Ike, A.; Toda, N.; Tsuji, N.; Hirata, K.; Miyamoto, K., Hydrogen photoproduction from
CO2-fixing microalgal biomass: Application of halotolerant photosynthetic bacteria.
journal of fermentation and Bioengineering 1997, 84, 606-609.
62
36.
37.
38.
39.
40.
41.
42.
43.
44.
45.
46.
47.
48.
49.
50.
51.
52.
53.
Melis, A.; Happe, T., Hydrogen production. Green algae as a source of energy. Plant
Physiology 2001, 127, 740-748.
Benemann, J., Hydrogen biotechnology: Progress and prospects. Nature Biotechnology
1996, 14, 1101-1103.
Kraemer, J. T.; Bagley, D. M., Improving the yield from fermentative hydrogen
production. Biotechnology Letters 2007, 29.
Venkata Mohan, S.; Chiranjeevi, P.; Mohanakrisha, G., A rapid and simple protocol for
evaluating biohydrogen production potential (BHP) of waste water with simultaneous
process optimization. International Journal of Hydrogen Energy 2012, 37, 3130-3141.
Conrad, R., Contribution of hydrogen to methane production and control of hydrogen
concentrations in methanogenic soils and sediments. FEMS Microbiology Ecology 1999,
28, 193-202.
Nath, K.; Das, D., Improvement of fermentative hydrgen production. Various
approaches. Applied Microbiology 2004, 65, 520-529.
Mathews, J.; Wang, G., Metabolic pathway engineering for enhanced biohydrogen
production. International Journal of Hydrogen Energy 2009, 34, 7404-7416.
Li, C.; Fang, H. H. P., Fermentative hydrogen production from wastewater and solid
wastes by mixed culture. Critical. Reviews in Environmental Science and Technology.
2007, 37, 1-39.
Das, D.; Veziroǧlu, T. N., Hydrogen production by biological processes: a survey of
literature. International Journal of Hydrogen Energy 2001, 26, 13-28.
Fang, H. H.; Yu, H., Mesophilic acidification of gelatinaceous wastewater. Journal of
Biotechnology 2002, 93, 99-108.
Hallenbeck, P. C., Fundamentals of the fermentative production of hydrogen. Water
science andTechnology 2005, 52, 21-9.
Saady, N. M. C., Homoacetogenesis during hydrogen production by mixed cultures dark
fermentation: Unresolved challenge. International Journal of Hydrogen Energy 2013, 38,
13172-13191.
Conrad, R.; Schink, B.; Phelps, T. J., Thermodynamics of H2-consuming and H2producing metabolic reactions in diverse methanogenic environments under in situ
conditions. FEMS Microbiology Letters 1986, 38, 353-360.
Khanal, S. K.; Chen, W. H.; Li, L.; Sung, S., Biological hydrogen production: effects of
pH and intermediate products. International Jounal of Hydrogen Energy 2004, 29, 11231131.
Chang, J. S.; Lee, K. S.; Lin, P. J., Biohydrogen production with fixed-bed bioreactors.
International Journal of Hydrogen Energy 2002, 27, 1167-1174.
Nath, K.; Kumar, A.; Das, D., Effect of some environmental parameters on fermentative
hydrogen production by Enterobacter cloacae DM11. Canadian Journal of Microbiology
2006, 52, 525-532.
Hartmann, H.; Ahring, B. K., Strategies for the anaerobic digestion of the organic
fraction of municipal solid waste: an overview. Water Science and Technology 2006, 53,
7-22.
Taherzadeh, M. J.; Karimi, K., Pretreatment of lignocellulosic wastes to improve ethanol
and biogas production: a review. International Journal of Molecular Science 2008, 9,
1621-1651.
63
54.
55.
56.
57.
58.
59.
60.
61.
62.
63.
64.
65.
66.
67.
68.
69.
70.
Weiland, P., State of the art of solid-state digestion-recent developments. In Solid-State
Digestion-State of the Art and Further R & D Requirements, Rohstoffe, F. N., Ed.
Gulzower Fachgespräche: 2006; Vol. 24, pp 22-38.
Palmowski, L. M.; Muller, J. A., Influence of the size reduction of organic waste on their
anaerobic digestion. Water Science and Technology 2000, 41, 155-162.
Mshandete, A.; Björnsson, L.; Kivaisi, A. K.; Rubindamayugi, M. S. T.; Mattiasson, B.,
Effect of particle size on biogas yield from sisal fibre waste. Renewable Energy 2006, 31,
2385-2392.
Khanal, S. K., Overview of anaerobic biotechnology. In Anaerobic biotechnology for
bioenergy production: Principles and Applications, John Wiley & Sons, Ltd.: USA,
2008.
Kim, J. K.; Park, C.; Kim, T. H.; Lee, M.; Kim, S. H.; Kim, S. W.; Lee, J. P., Effects of
various pretreatments for enhanced anaerobic digestion with waste activated sludge.
Journal of Booscience and Bioengineering. 2003, 95, 271-275.
Lee, D. H.; Behara, S. K.; Kim, J.; Park, H. S., Methane production potential of leachate
generated from korean food waste recycling facilities> a lab scale study. Waste
Management and Research 2009, 29, 876-882.
Zupancic, G. D.; Grilc, V., Anaerobic treatment and biogas production from organic
waste. In Management of organic waste, Kumar, S., Ed. In Tech: Croatia, 2012.
Sung, S.; Liu, T., Ammonia inhibition on thermophilic anaerobic digestion. Chemosphere
2003, 53, 43-52.
Hwang, M. H.; Jang, N. J.; Hyun, S. H.; Kim, I. S., Anaerobic bio-hydrogen production
from ethanol fermentation: the role of pH. Journal of Biotechnology 2004, 111, 297-309.
Luo, G.; Karakashev, D.; Xie, L.; Zhou, Q.; Angelidaki, I., Long-term effect of inoculum
pretreatment on fermentative hydrogen production by repeated batch cultivations:
homoacetogenesis and methanogenesis as competitors to hydrogen production.
Biotechnology and Bioengineering 2011, 108, 1816-27.
Noah, M. M.; Wiegel, J., Life at extreme limits. The anaerobic halophilic
alkalithermophiles. Annual New York Academy of Sciences 2008, 1125, 44-57.
Wagner, I. D.; Wiegel, J., Diversity of thermophilic anaerobes. Annals of New York
Academy of Sciences 2008, 1125, 1-43.
Kothari, R.; Pandey, A. K.; kumar, S.; Tyagi, V. V.; Tyagi, S. K., Different aspects of dry
anaerobic digestion for bio-energy: An overview. Renewable and Sustainable Energy
Reviews 2014, 39, 174-195.
Ward, A. J.; Hobbs, P. J.; Holliman, P. J.; Jones, D. L., Optimization of the anaerobic
digestion of agricultural resources. Bioresource Technology 2008, 99, 7928-7940.
Dela-Rubia, M. A.; Perez, M.; Romero, L. I.; Sales, D., Anaerobic mesophilic and
thermophilic municipal sludge digestion. Chemical and Biochemical Engineering. 2002,
16, 119-124.
Tchobanoglous, G.; Burton, F. L.; Stensel, H. D.; Metcalf; Eddy, Wastewater
Engineering: Treatment and Reuse. McGraw-Hill Education: 2003.
Chen, C. C.; Lin, C. Y.; Chang, J. S., Kinetics of hydrogen production with continuous
anaerobic cultures utilizing sucrose as the limiting substrate. Applied Microbiology and
Biotechnology 2001, 57, 56-64.
64
71.
72.
73.
74.
75.
76.
77.
78.
79.
80.
81.
82.
83.
84.
85.
86.
87.
Oh, Y. K.; Kim, S. H.; Kim, M. S.; Park, S. C., Thermophilic biohydrogen production
from glucose with trickling biofilter. Biotechnology and Bioengineering 2004, 88, 690698.
Kroeker, E. J.; Schulte, D. D.; Sparling, A. B.; Lapp, H. M., Anaerobic treatment process
stability. Journal of Water Pollution Control Federation. 1979, 51, 718-727.
Karim, K.; Klasson, T.; Hoffmann, R.; Drescher, S. R.; DePaoli, D. W.; Al-Dahhan, M.
H., Anaerobic digestion of animal waste: Effect of mixing. Bioresource Technology
2005, 96, 1607-1612.
Meroney, R. N.; Colorado, P. E., CFD simulation of mechanical draft tube mixing in
anaerobic digester tanks. Water Research 2009, 43, 1040-1050.
Sroot, P. G.; McMahon, K. D.; Mackie, R. I.; Raskin, L., Anaerobic codigestion of
municipal solid waste and biosolids under various mixing conditions: I. Digester
performance. Water Research 2001, 24, 1804-1816.
Hawkes, F. R.; hussy, I.; Kyazze, G.; Dinsdale, R.; Hawkes, D. L., Continuous dark
fermentative hydrogen production by mesophilic microflora: Principles and progress.
International Journal of Hydrogen Energy 2007, 32, 172-184.
Hawkes, F. R.; Dinsdale, R.; Hawkes, D. L.; Hussy, I., Sustainable fermentative
hydrogen production: challenges for process optimisation. International Journal of
Hydrogen Energy 2002, 27, 1339-1347.
Levin, D. B.; Pitt, L.; Love, M., Biohydrogen production production: prospects and
limitations to practical application. International Journal of Hydrogen Energy 2004, 29,
173-185.
Angenent, L. T.; Karim, K.; Al-Dahhan, M. H.; Wrenn, B. A.; Domiguez-Espnosa, R.,
Production of bioenergy and biochemicals from industrial and agricultural wastewater.
Trends in Biotechnology 2004, 22, 477- 485.
Carver, S. M.; Nelson, M. C.; Lepistö, R.; Yu, Z.; Tuovinen, O. H., Hydrogen and
volatile fatty acids production during fermentation of cellulosic substrates by a
thermophilic consortium at 50 and 60°C. Bioresource Technology 2011, 104, 424-431.
Ueno, Y.; Haruta, S.; Ishii, M.; Igarashi, Y., Microbial community in anaerobic
hydrogen-producing microflora enriched from sludge compost. Applied Microbiology
and Biotechnology 2001, 57, 555-562.
Ye, C.; Cheng, J. J.; Creamer, K. S., Inhibition of anaerobic digestion process: A review.
Bioresource Technology 2008, 99, 4404-4064.
Goswami, D. Y.; Kreith, F., Handbook of Energy Efficiency and Renewable Energy.
CRC Press: 2007.
Reith, J. H.; Wijffels, R. H.; Barten, H., Biomethane and Biohydrogen: Status and
perspectives of biological methane and hydrogen production. Dutch Biological Hydrogen
Foundation-NOVEM: Netherland, 2003.
Barbir, F.; Veziroglu, T. N.; Plass, H. J., Environmental damage due to fossil- fuels use.
International Journal of Hydrogen Energy 1990, 15, 739-749.
Salomon, K. R.; Silva Lora, E. E., Estimate of the electric energy generating potential for
different sources of biogas in Brazil. Biomass and Bioenergy 2009, 33, 1101-1107.
Claassen, P. A. M.; Lopez-Contreras, A. M.; Sijtsma, L., Utilisation of biomass for the
supply of energy carriers Applied Microbiology and Biotechnology 1999, 52, 741-755.
65
88.
89.
90.
91.
92.
93.
94.
95.
96.
97.
98.
99.
100.
101.
102.
103.
104.
105.
106.
Rachman, M. A.; Nakashimada, Y.; Kakizono, T.; Nishio, N., Hydrogen production with
high yield and high evolution rate by self-flocculated cells of Enterobacter aerogenes in a
packed-bed reactor. Applied Microbiology and Biotechnology 1998, 49, 450-454.
Armor, J. N., Catalysis and the hydrogen economy. Catalysis Letters 2005, 101, 131-135.
Fyfe, W. S., Clean energy for 10 billion humans in the 21st century: is it possible?
International Journal of Coal Geology 1999, 40, 85-90.
Lenssen, N.; Flavin, C., Sustainable energy for tommorrow's world-The case for an
optimistic view of the future. Energy Policy 1996, 24, 769-781.
Valdez-Vazquez, I.; Sparling, R.; Risbey, D.; Rinderknecht-Seijas, N.; Poggi-Varaldo, H.
M., Hydrogen generation via anaerobic fermentation of paper mill wastes. Bioresource
Technology 2005, 96, 1907-1913.
Yamin, J. A. A., Comparative study using hydrogen and gasoline as fuels: Combustion
duration effect. International Journal of Energy Research 2006, 30, 1175-1187.
Yamin, J. A. A.; Gupta, H. N.; Bansal, B. B.; Srivastava, O. N., Effect of combustion
duration on the performance and emission characteristics of a spark ignition engine using
hydrogen as a fuel. International Journal of Hydrogen Energy 2000, 25, 581-589.
Bockris, J. O. M., The economics of hydrogen as a fuel. International Journal of
Hydrogen Energy 1981, 6, 223-241.
Mormirlan, M.; Veziroglu, T. N., Current status of hydrogen energy. Renewable and
Sustainable Energy Reviews 2002, 6, 141-179.
Dong, G. X.; Wu, B. R.; Zhu, L.; Du, J., Microstructure and electrochemical properties of
low-temperature hydrogen storage alloy used in Ni/MH batteries. Transactions of
Nonferrous Metals Society of China 2007, 17, S941-S944.
Kloeppel, J.; Rogerson, S., The Hydrogen Economy. Electronics World and Wireless
World 1991, 97, 668-671.
Reith, J. H.; Wijffels, R. H.; Barten, H., Biomethane and Biohydrogen. Status and
perspectives of biological methane and hydrogen production. Dutch Biological Hydrogen
Foundation -NOVEM: The Nerthelands, 2003.
Wang, J.; Wann, W., Factors influencing fermentative hydrogen production: A review.
International Journal of Hydrogen Energy 2009, 34, 799-811.
Winkler, H.; Simões, A. F.; Rovere, E. L. l.; Alam, M.; Rahman, A.; Mwakasonda, S.,
Access and Affordability of Electricity in Developing Countries. World Development
2011, 39, 1037-1050.
Qiao, W.; Yan, X.; Ye, J.; Sun, Y.; Wang, W.; Zhang, Z., Evaluation of biogas
production from different biomass wastes with/without hydrothermal pretreatment.
Renewable Energy 2011, 36, 3313-3318.
Deublein, D.; Steinhauser, A., Biogas from Waste and Renewable Resources: An
Introduction Wiley-VCH: Weinheim, 2008.
Speece, R. E., Nutrient Requirements In Anaerobic Digestion of Biomass, Chynoweth, D.
P.; Isaacson, R., Eds. Elsevier Applied Science: London, 1984.
Yen, H.-W.; Brune, D., Anaerobic co-digestion of algael sludge and waste paper to
produce methane. Bioresource Technology 2007, 98, 130-134.
Lay, J.-J.; Fan, K.-S.; Chang, J.; Ku, C.-H., Influence of chemical nature of organic
wastes on their conversion to hydrogen by heat-shock digested sludge. International
Journal of Hydrogen Energy 2003, 28, 1361-1367.
66
107.
108.
109.
110.
111.
112.
113.
114.
115.
116.
117.
118.
119.
120.
121.
122.
123.
124.
125.
126.
Gunaseelan, V. N., Anaerobic digestion of biomass for methane production: a review.
Biomass and Bioenergy 1997, 13, 83-114.
Lee, J. P.; Lee, J. S.; Park, S. C., Two-phase methanisation of food wastes in pilot scale.
Applied Biochemistry and Biotechnology 1999, 77-79, 585-593.
Demirel, B.; Scherer, P. A., Production of methane from sugar beet silage without
manure addition by a single -stage anaerobic digestion process. Biomass and Engineering
2008, 32, 203-209.
Erglyst, H. N.; Quigley, M. E.; Hudson, G. J., Definition and measurement of dietary
fibre. European Journal of Clinical Nutrition 1995, 49, S48-S62.
Hashimoto, A. G., Ammonia inhibition of methanogenesis from cattle waste.
Agricultural Wastes 1986, 17, 241-261
Warren, K. S., Ammonia toxicity and pH. Nature 1962, 195, 47-49.
Sprott, G. D.; Patel, G. B., Ammonium toxicity in pure cultures of methanogenic
bacteria. Systematic and Applied Microbiology 1986, 7, 758-363.
Vidal, G.; Carvalho, A.; Méndez, R.; Lema, J. M., Influence of the content in fats and
proteins on the anaerobic biodegradability of dairy wastewaters. Bioresource Technology
2000, 74, 231-239.
Quéméneur, M.; Marty, Y., Fatty acids and sterols in domestic wastewaters. Water
Research 1994, 28, 1217-1226.
Pereira, M. A.; Sousa, D. Z.; Mota, M.; Alves, M. M., Mineralisation of LCFA
Associated with anaerobic sludge: kinetics, enhancement of methanogenic activity, and
effect of VFA. Biotechnology and Bioengineering 2004, 88, 502-511.
Fernandez, A.; Sanches, A.; Font, X., Anaerobic co-digestion of a simulated organic
fraction of municipal solid wastes and fats of animal and vegetable origin. Biochemical
Engineering Journal 2005, 26, 22-28.
Koster, I. W.; Cramer, A., Inhibition of methanogenesis from acetate in granular sludge
by long-chain fatty acids. Applied and Environmental Microbiology 1987, 53, 403-409.
Angelidaki, I.; Ahring, B. K., Effects of free long-chain fatty acids on thermophilic
anaerobic digestion. Applied Microbiology and Biotechnology 1992, 37, 808-812.
Lalman, J. A.; Bagley, D. M., Anaerobic degradation and methanogenic inhibitory effects
of oleic and stearic acid. Water Resources 2001, 35, 2975-2983.
Chen, Y.; Cheng, J. J.; creamer, K. S., Inhibition of anaerobic digestion process: A
review. Bioresource Technology 2008, 99, 4044-4064.
Ralph, J.; Landucci, L. L., NMR of lignins. In Lignin and Lignans: Advances in
Chemistry, Heitner, C.; Dimmel, D. R.; Schmidt, J. A., Eds. CRC Press: Boca Raton, FL,
2010; pp 137–234.
Boerjan, W.; Ralph, J.; Baucher, M., Lignin biosynthesis Annual Review of Plant Biology
2003, 54, 519–546
Saratable, G. D.; Chen, S.; Lo, Y.; Saratale, J. L. G.; Chang, J. S., Outlook of
biohydrogen production from lignocellulosic feedstock using dark fermentation. A
review. Journal of Science and Industrial Research 2008, 67, 962-979.
Kalra, M. S.; Panwar, J. S., Anaerobic digestion of rice crop residues. Agricultural
Wastes 1986, 17, 263-269.
Yadvika, S.; Sreekrishnan, T. R.; Kohli, S.; Rana, V., Enhancement of biogas production
from solid substrates using different techniques-a review. Bioresource Technology 2004,
95, 1-10.
67
127.
128.
129.
130.
131.
132.
133.
134.
135.
136.
137.
138.
139.
140.
141.
142.
143.
144.
145.
Rynk, R., On-Farm Composting Handbook. lthaca; NRAES: New York, 1992.
Lethomäki, A.; Huttunen, S.; Lethinen, T. M.; Rintala, J. A., Anaerobic digestion of grass
silage in batch leach bed processes for methane production. Bioresource Technology
2008, 99, 3267-3278.
Chynoweth, D. P.; Wilkie, A. C.; Owens, J. M., Anaerobic treatment of piggery slurryReview. Asian-Australian Journal of Animal Sciences 1999, 12, 607-628.
Zeeman, G.; Wiegant, W. M.; Koster-Treffers, M. E.; Lettinga, G., The influence of the
total ammonia concentration on the thermophilic digestion of cow manure. Agricultural
Wastes 1985, 14, 19-35.
Moller, H. B.; Sommer, S. G.; Ahring, B. K., Methane productivity of manure, straw and
solid fraction of manure. Biomass and Bioenergy 2004, 26, 485-495.
Cuetos, M. J.; Golmez, X.; Oterbo, M.; Moran, A., Anaerobic influence of co-digestion
with the organic fraction of municipal solid waste (OFMSW). Biochemical Engineering
Journal 2008, 40, 99-106.
Rosenwinkel, K. L.; Meyer, H., Anaerobic treatment of slaughterhouse residues in
municipal digesters. Water Science and Technology 1999, 40, 101-111.
Salminen, E.; Rintala, J., Anaerobic digestion of organic solid poultry slaughterhouse
waste-a review. Bioresource Technology 2002, 83.
Tchobanoglous, G.; Burton, F. L.; Metcalf; Eddy, Wastewater engineering : treatment,
disposal, and reuse. McGraw-Hill: New York, 1991.
Ros, M.; Franke-Whittle, I. H.; Morales, A. B.; Insam, H.; Ayuso, M.; Pascual, J. A.,
Archaeal community dynamics and abiotic characteristics in a mesophilic anaerobic codigestion process treating fruit and vegetable processing waste sludge with chopped fresh
artichoke waste. Bioresource Technology 2013, 136, 1-7.
Davidson, P. M.; Naidu, A. S., Phytophenols. In Natural food antimicrobial systems,
Naidu, A. S., Ed. CRC Press: Boca Raton, FL, 2000; pp 265-294.
Cowan, M. M., Plant products as antimicrobial agents. Clinical microbiology reviews
1999, 12, 564-82.
Jiang, Y.; Song, J., Fruits and Fruit Flavor: Classification and Biological
Characterization. In Handbook of Fruit and Vegetable Flavors, John Wiley and Sons,
Inc. 2010; pp 1-23.
Goff, S. A.; Klee, H. J., Plant volatile compounds: Sensory cues for health and nutritional
value Science 2006, 311, 815-819.
Hui, Y. H.; Chen, F.; Nollet, L. M. L.; Guiné, R. P. F.; Martín-Belloso, O.; MínguezMosquera, I.; Paliyath, G.; Pessoa, F. L. P.; Quéré, J. L. L.; Sidhu, J. S., Handbook of
Fruit and Vegetable Flavors. Wiley: 2010.
Youngsukkasem, S.; Akinbomi, J.; Rakshit, S.; Taherzadeh, M. J., Biogas production by
encased bacteria in a synthetic membranes: Protective effects in toxic media and high
loading rates. Environ. Technol. 2013, 34, 2077-2084.
Wikandari, R.; Youngsukkasem, S.; Millati, R.; Taherzadeh, M. J., Performance of semicontinuous membrane biotreactors in biogas production from toxic feedstock containing
D-limonene Bioresource Technology 2014, 170, 350-355.
Martin, M. A.; Siles, J. A.; China, A. F.; Martin, A., Biomethanization of orange peel
waste. Bioresource Technology 2010, 101, 8993-8999.
Mizuki, E.; Akao, T.; Saruwatari, T., Inhibitory effect of citrus Unshu peel on anaerobic
digestion. Biological Wastes 1990, 33, 161-168.
68
146.
147.
148.
149.
105.
151.
152.
153.
154.
155.
156.
157.
158.
159.
160.
161.
162.
163.
Grohmann, K.; Baldwin, E.; Buslig, B., Production of ethanol from enzymatically
hydrolyzed orange peel by the yeast Saccharomyces cerevisiae. Applied Biochemistry
and Biotecnology 1994, 45-46, 315-327.
Winniczuk, P. P.; Parish, M. F., Minimum inhibitory concentrations of antimicrobials
against micro-organisms related to citrus juice. Food Microbiology 1997, 14, 373-381.
Muna, A.; Zhang, F. J.; Wu, F. F.; Zhou, C. H.; Tao, J., Advances in fruit aroma volatile
research. Molecules 2013, 18, 8200-8229.
Kader, A. A., Perspective flavour quality of fruits and vegetables. Journal of the Science
of Food and Agriculture 2008, 88, 1863-1869.
Speece, R. E., Anaerobic biotechnology for industrial waste treatment. Environmental
Scence andTechnology 1983, 17, A416-A427.
Sikkema, J.; Bont, J. A. M.; Poolman, B., Interactions of cyclic hydrocarbons with
biological membranes. Journal of Biological Chemistry 1994, 269, 8022-8028.
Griffin, S. G.; Wyllie, S. G.; Markham, J. L.; Leach, D. N., The role of structure and
molecular properties of terpenoids in determining their antimicrobial activity. Flavour
and Fragrance Journal 1999, 14, 322-332.
Burt, S., Essential oils: their antibacterial properties and potential applications in foods-a
review. International Journal of Food Microbiology 2004, 94, 223-253.
Gutierrez, M. E.; Garcia, A. F.; de-Madariaga, M. A.; Sagrista, M. L.; Casado, F. J.;
Mora, M., Interaction of tocophenols and phenolic compounds with membrane lipid
components. Evaluation of their antioxidant activity in a liposomal model system. Life
Science 2003, 72, 2337-2360.
Lee, S. E.; Hwang, H. J.; Ha, J. S.; Jeong, H. S.; Kim, J. H., Screening of medicinal plant
extracts for antioxidant activity. Life Science 2003, 73, 167-179.
Dorman, H. J. D.; Deans, S. G., Antimicrobial agents from plants: Antibacterial activity
of plant volatile oils. Journal of Applied Microbioiogy. 2000, 88, 308-316.
Cardozo, P. W.; Calsamiglia, S.; Ferret, A.; Kamel, C., Effects of natural plant extracts
on protein degradation and fermentation profiles in continuous culture. Journal of Aninal
Science 2004, 82, 3230-3236.
Molero, R.; Ibars, M.; Calsamiglia, S.; Ferret, A.; Losa, R., Effect of a specific blend of
essential oil compounds on dry matter and crude protein degradability in heifers fed diets
with different forage to concentrate rations. Animal Feed Science Technology 2004, 114,
91-104.
Ruiz, B.; Flotats, X., Citrus essential oils and their influence on the anaerobic digestion
process: an overview. Waste management (New York, N.Y.) 2014, 34, 2063-79.
Castillejos, L.; Calsamiglia, S.; Ferret, A.; Losa, R., Effects of dose and adaptation time
of a specific blend of essential oils compounds on rumen fermentation. Animal Feed
Science and Technology. 2007, 132, 186-201.
Hong, C.; Haiyun, W., Optimization of volatile fatty acid production with co-substrate of
food wastes and dewatered excess sludge using response surface methodology.
Bioresource Technology 2010, 101, 5487-5493.
Mechichi, T.; Sayadi, S., Evaluating process imbalance of anaerobic digestion of swine
manure slurry in an sequencing batch reactor. Water Research 2005, 34, 3087-3106.
Ahring, B. K.; Sandberg, M.; Angelidaki, I., Volatile fatty acids as indicators of process
imbalance in anaerobic digesters. Applied Microbiology and Biotechnology 1995, 43,
559-565.
69
154.
165.
166.
167.
168.
169.
170.
171.
172.
173.
174.
175.
176.
177.
178.
179.
180.
181.
Siegert, I.; Banks, C., The effect of volatile fatty acid additions on the anaerobic digestion
of cellulose and glucose in batch reactors. Process Biochemistry 2005, 40, 3412-3418.
Boone, D. R.; Xun, L. Y., Effects of pH, temperature, and nutrients on propionate
degradation by a methanogenic enrichment culture. Applied and Environmental
Microbiology 1987, 53, 1589-1592.
Lee, W. S.; Chua, A. S. M.; Yeoh, H. K.; Ngoh, G. C., A review of the production and
applications of waste-derived volatile fatty acids. Chemical Engineering Journal 2014,
235, 83-99.
Choi, J. H.; Fukushi, K.; Yanamoto, K., A study on the removal of organic acids from
wastewaters using nanofiltration membranes. Separation and Purification Technology
2008, 59, 17-25.
Noble, R. D.; Stern, S. A., Membrane Separation Technology: Principles and
Applications. Elsevier: The Nertherlands, 1995.
Khanal, S. K., Anaerobic reactor configurations for bioenergy production. In Anaerobic
biotechnology for bioenergy production: Principles and Applications John Wiley &
Sons, Ltd.: USA, 2008.
Judd, S., The MBR Book-Principles and applications of membrane bioreactors in water
and wastewater treatment. Elsevier: London, 2006.
Baker, R. W., Membrane Technology and Applications 2nd ed.; John Wiley and Sons,
Ltd.: Chichester, 2004.
Singhania, R. R.; Christophe, G.; Perchet, G.; Troquet, J.; Larroche, C., Immersed
membrane bioreactors: An overview with special emphasis on anaerobic bioprocesses.
Bioresource Technology 2012, 122, 171-180.
Baker, R., Microfiltration in membrane technology and applications. 3rd ed.; John Wiley
& Sons Ltd: California, 2012.
Crittenden, J.; Trussell, R.; Hand, D.; Howe, k.; Tchobanoglous, G., Principles of water
treatment. 2nd ed.; John Wiley and Sons: New Jersey, 2012.
Lebegue, J.; Heran, M.; Grasmick, A., Membrane biofreactor: distribution of critical flux
throughout an immersed HF bundle. Desalination 2008, 231, 245-252.
Ylitervo, P.; Akinbomi, J.; Taherzadeh, M. J., Membrane bioreactors' potential for
ethanol and biogas production: a review. Environmental technology 2013, 34, 1711-23.
Liao, B. Q.; Kraemer, J. T.; Bagley, D. M., Anaerobic membrane bioreactors:
applications and research directions. Critical Reviews in Environmental Science and
Technology 2006, 36, 489-530.
Oshima, K. H.; Evans-Strickfaden, T. T.; Highsmith, A. K.; Ades, E. W., The use of a
microporous polyvinylidene fluoride (PVDF) membrane filter to separate contaminating
viral particles from biologically important proteins. Biologicals 1996, 24, 137-145.
Morihama, A. c. D.; Mierzwa, J. C., Clay nanoparticles effect on performance and
morphology of poly(vinylidene fluoride) membranes. Brazilian Journal of Chemical
Engineering 2014, 31, 79-93.
Zhao, Y.; Qian, Y.; Zhu, B.; Xu, Y., Modification of porous poly(vinylidene fluoride)
membrane using amphiphilic polymers with different structures in phase inversion
process. Journal of Membrane Science 2008, 310, 567-576.
Pourbafrani, M.; Talebnia, F.; Niklasson, C.; Taherzadeh, M. J., Protective effect of
encapsulation in fermentation of limonene-contained media and orange peel hydrolyzate.
International Journal of Molecular Sciences 2007, 8, 777-787.
70
182.
183.
184.
185.
186.
187.
188.
189.
190.
191.
192.
193.
194.
195.
196.
197.
198.
199.
Crank, J.; Park, G. S., Diffusion in polymers. Academic Press: New York, 1968.
Nicholson, J. W., The chemistry of polymers. The Royal Society of Chemistry:
Cambridge, 1997.
Bickerstaff, G. F., Immmobilization of enzymes and cells. In Methods in Biotechnology,
1st ed.; Bickerstaff, G. F., Ed. Humana Press: Totowa, USA, 1997.
Cao, L., Carrier-bound immobilized enzymes: Principles, Applications and Design. John
Wiley and Sons: New York, USA, 2005.
Webb, C.; Dervakos, G. A., Studies in viable cell immobilization. Academic Press: San
Diego, 1996.
Kourkoulas, Y.; Bekatorou, A.; Banat, I. M.; Merchant, R.; Koutinas, A. A.,
Immobilization technologies and support materials suitable in alcohol beverages
production: a review. Food Microbiology 2004, 21, 377-397.
Fukui, S.; Tanaka, A., Immobilized microbial cells. Annual Reiew of Microbiology 1982,
36, 145-172.
Guisan, J. M., Immobilization of enzymes and cells. In Methods in Biotechnology,
Walker, J. M., Ed. Humana Press: Totowa, USA, 2006; Vol. 22.
Xie, T.; Wang, A.; Huang, L.; Li, H.; Chen, Z.; Wang, Q.; Yin, X., Review: Recent
advance in the support and technology used in enzyme immobilization. African Journal
of Biotechnology 2009, 8, 4724-4733.
Singhania, R. R.; Patel, A. K.; Christophe, G.; Fontanille, P.; Larroche, C., Biological
upgrading of volatile fatty acids, key intermediates for the valorization of biowaste
through dark anaerobic fermentation. Bioresource Technology 2013, 145, 166-174.
Gluszcz, P.; Jamroz, T.; Sencio, B.; Ledakowicz, S., Equilibrium and dynamic
investigations of organic acids adsorption onto ion-exchange resins. Bioprocess and
Biosystems Engineering 2004, 26, 185-90.
Huang, C.; Xu, T.; Zhang, Y.; Xue, Y.; Chen, G., Application of electrodialysis to the
production of organic acids: State-of-the-art and recent developments. Journal of
Membrane Science 2007, 288, 1-12.
Wang, Z.; Luo, Y.; Yu, P., Recovery of organic acids from waste salt solutions derived
from the manufacture of cyclohexanone by electrodialysis. Journal of Membrane Science
2006, 280, 134-137.
Joglekar, H. G.; Rahman, I.; Babu, S.; Kulkarni, B. D.; Joshi, A., Comparative
assessment of downstream processing options for lactic acid. Separation and Purification
Technology 2006, 52, 1-17.
Alkaya, E.; Kaptan, S.; Ozkan, L.; Uludag-Demirer, S.; Demirer, G. N., Recovery of
acids from anaerobic acidification broth by liquid–liquid extraction. Chemosphere 2009,
77, 1137-1142.
Kertész, R.; Schlosser, Š., Design and simulation of two phase hollow fiber contactors for
simultaneous membrane based solvent extraction and stripping of organic acids and
bases. Separation and Purification Technology 2005, 41, 275-287.
Visvanathan, C.; Ben-Aim, R.; Parameshwaran, K., Membrane separation bioreactors for
wastewater treatment. Critical Reiews in Environmental Science and Technology 2000,
30, 1-48.
Chang, S.; Fane, A. G., The effect of fibre diameter on filtration and flux distribution —
relevance to submerged hollow fibre modules. Journal of Membrane Science 2001, 184,
221-231.
71
200.
201.
202.
203.
204.
205.
206.
207.
208.
209.
210.
211.
212.
213.
214.
215.
216.
217.
Yamamura, H.; Kimura, K.; Watanabe, Y., Mechanism involved in the evolution of
physically irreversible fouling in microfiltration and ultrafiltration membranes used for
drinking water treatment. Environmental Science and Technolnology 2007, 41, 67896794.
Drews, A., Membrane fouling in membrane bioreactors-characterization, contradictions,
cause and cures. Journal of Membrane Science 2010, 363, 1-28.
Wu, Z.; Wang, Z.; ma, Y.; Zhou, Q.; Yang, D., Membrane fouling properties under
different filtration modes in a submerged membrane bioreactor. Process Biochemistry
2010, 45, 1699-1706.
Tian, J. Y.; Xu, Y.; Chen, Z.; Nan, J.; Li, G., Air bubbling for alleviating membrane
fouling of immersed hollow-fiber for ultrafiltration of river water. Desalination 2010,
260, 225-230.
Cheryan, M., Fouling and cleaning in ultrafiltration and microfiltration handbook. 2nd
ed.; CRC press: Florida, 1998.
Macmillin, R. B.; Weber, M., The theory of short-circuiting in continuous flow mixing
vessels in series and kinetics of chemical reactions in such systems. Transactions of
American Institute of Chemical Engineers 1935, 31, 409-458.
Karim, K.; Hoffmann, R.; Klasson, K. T.; Al/Dahhan, M. H., Anaerobic digestion of
animal waste: effect of mode of mixing. Water Research 2005, 39, 3597-3606.
Stroot, P. G.; McMahon, K. D.; Mackie, R. I.; Raskin, L., Anaerobic codigestion of
municipal solid waste and biosolids under various mixing conditions-I. Digester
performance. Water Research 2001, 35, 1804-1816.
Russeaux, J. M.; Muhr, H.; Plasari, E., Mixing and micromixing times in the forced
vortex region of unbaffled mixing devices The Canadian Journal of Chemical
Engineering 2001, 79, 697-707.
Rahman-Al-Ezzi, A.; Najmuldeen, G. F., Gas Hold-Up, Mixing Time and Circulation
Time in Internal Loop Airlift Bubble Column. International Journal of Engineering
Research and Applications 2014, 4, 286 - 294
Nauman, E. B., Residence Time Distributions. In Handbook of Industrial Mixing, John
Wiley and Sons, Inc.: 2004; pp 1-17.
Danckwerts, P. V., Continuous flow systems. Distribution of residence times. Chemical
Engineering Science 1953, 2, 1-13.
Levenspiel, O., Chemical Reaction Engineering. 3rd ed.; Wiley: New York, 1999.
Seadler, J.; Henley, E., Separation process principles. 2nd ed.; John Wiley and Sons:
New Jersey, 2006.
Perry, R. H.; Green, D. W., Perry's Chemical Engineers' Handbook. 8th ed.; McGrawHill Professional: New York, 2007.
Naessens, W.; Maere, T.; Nopens, I., Critical review of membrane bioreactor models-Part
1: Biokinetic and filtration models. Bioresource Technology 2012, 122, 95-106.
Field, R. W.; Wu, D.; Howell, J. A.; Guptha, B. B., Critical flux concept for
microfiltration fouling. Journal of Membrane Science 1995, 100, 259-272.
Song, L., Flux deline in crossflow microfiltration and ultrafiltration: mechanisms and
modeling of membrane fouling. Journal of membrane science 1998, 139, 183-200.
72
Paper I
This article was downloaded by: [Julius Akinbomi]
On: 21 February 2013, At: 23:03
Publisher: Taylor & Francis
Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House,
37-41 Mortimer Street, London W1T 3JH, UK
Environmental Technology
Publication details, including instructions for authors and subscription information:
http://www.tandfonline.com/loi/tent20
Biogas production by encased bacteria in synthetic
membranes: protective effects in toxic media and high
loading rates
Supansa Youngsukkasem
Taherzadeh
a
a b
, Julius Akinbomi
a c
b
, Sudip K. Rakshit & Mohammad J.
a
School of Engineering, University of Borås, Borås, Sweden
b
School of Environment, Resources and Development, Asian Institute of Technology,
Pathumthani, Thailand
c
Department of Chemical and Polymer Engineering, Lagos State University, Lagos, Nigeria
Accepted author version posted online: 28 Jan 2013.Version of record first published: 18 Feb
2013.
To cite this article: Supansa Youngsukkasem , Julius Akinbomi , Sudip K. Rakshit & Mohammad J. Taherzadeh (2013):
Biogas production by encased bacteria in synthetic membranes: protective effects in toxic media and high loading rates,
Environmental Technology, DOI:10.1080/09593330.2013.770555
To link to this article: http://dx.doi.org/10.1080/09593330.2013.770555
PLEASE SCROLL DOWN FOR ARTICLE
Full terms and conditions of use: http://www.tandfonline.com/page/terms-and-conditions
This article may be used for research, teaching, and private study purposes. Any substantial or systematic
reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any form to
anyone is expressly forbidden.
The publisher does not give any warranty express or implied or make any representation that the contents
will be complete or accurate or up to date. The accuracy of any instructions, formulae, and drug doses should
be independently verified with primary sources. The publisher shall not be liable for any loss, actions, claims,
proceedings, demand, or costs or damages whatsoever or howsoever caused arising directly or indirectly in
connection with or arising out of the use of this material.
Environmental Technology, 2013
http://dx.doi.org/10.1080/09593330.2013.770555
Biogas production by encased bacteria in synthetic membranes: protective effects in toxic media
and high loading rates
Supansa Youngsukkasema,b , Julius Akinbomia,c , Sudip K. Rakshitb and Mohammad J. Taherzadeha∗
of Engineering, University of Borås, Borås, Sweden; b School of Environment, Resources and Development, Asian Institute of
Technology, Pathumthani, Thailand; c Department of Chemical and Polymer Engineering, Lagos State University, Lagos, Nigeria
a School
Downloaded by [Julius Akinbomi] at 23:03 21 February 2013
(Received 26 September 2012; final version received 22 January 2013 )
A bioreactor including encased digesting bacteria for biogas production was developed, and its performance in toxic media
and under high organic loading rates (OLRs) was examined and compared with traditional digestion reactors. The bacteria
(3 g) were encased and sealed in 3 × 6 cm2 PVDF (polyvinylidene fluoride) membranes with a pore size of 0.1 μm, and
then several sachets were placed in the reactors. They were then examined in toxic medium containing up to 3% limonene
as a model inhibitor in batch reactors, and OLRs of up to 20 g COD/L.day in semi-continuous digestions. The free and
encased cells with an identical total bacterial concentration of 9 g in a medium containing 2% limonene produced at most
6.56 and 23.06 mL biogas per day, respectively. In addition, the digestion with free cells completely failed at an OLR of
7.5 g COD/L.day, while the encased cells were still fully active with a loading of 15 g COD/L.day.
Keywords: biogas; synthetic membrane; encapsulation; cell containment; inhibitor
Introduction
Biogas or biomethane is a renewable energy source with
several applications, for example, car fuel, heating, cooking, or electricity production. Biogas consists mainly of
methane and carbon dioxide, but may also contain minor
impurities of other components [1]. The anaerobic digestion
process and production of methane consists of hydrolysis,
acidogenesis, acetogenesis, and methanogenesis. Typically,
methanogenesis is more sensitive to the environment than
the earlier stages [2]. Moreover, the doubling time of hydrolysis and acidogenesis in bacteria is about 1.0–1.5 days,
while acetogens and methanogens need about 1–4 and
5–15 days doubling time, respectively [3]. Consequently,
the methane-forming bacteria need a longer retention time
and are very sensitive to the process conditions; hence, they
can easily be washed out. Therefore, if the dilution rate of
methane-forming bacteria in the digester is too high, or if the
withdrawal of digester sludge is premature, the population
size of methane-forming bacteria is greatly reduced.
Other process challenges, such as low methane yield
and an unstable process, can often occur during the anaerobic digestion process. An important factor is toxicity and the
presence of inhibitor compounds in the substrates or wastes.
The impact of the toxic substance is that it mostly inhibits
the performance of bacteria in the digester by disturbing
bacterial growth, resulting in anaerobic reactor upset or failure [4]. There are many inhibitors that play an important role
∗ Corresponding
author. Email: [email protected]
© 2013 Taylor & Francis
in the digestion process, including: ammonia, light, heavy
metals, and organic compounds such as long-chain fatty
acids and phenol [4–6]. In the anaerobic digestion of fruit
waste, D-limonene is an example of an important inhibitor
of bacterial activities [7]. Limonene is a hydrocarbon classified as a cyclic terpene, which is a major component of peel
oil in citrus fruits. Not only does limonene cause odours
in citrus fruits, it is also well known as an antimicrobial
agent [8–10]. In the field of ethanol production, limonene
was reported as being an inhibitor of ethanol productivity [11,12]. Furthermore, it also inhibits the digestion of
bacteria and inhibits biogas production [13]. It is common
to pre-treat the substrate or use cell protection methods in
order to reduce or prevent these inhibitory effects and avoid
washing out of the cells [7,14,15].
Retaining the bacteria inside the digester by immobilization could be a solution to these problems. Cell
immobilization is an attractive method for maintaining a
high cell concentration in the reactor. In addition, immobilizing microbial cells at a high density not only improves the
productivity of a bioreactor, but the microbial cells immobilized in a polymeric membrane can also be protected
from harsh environmental conditions such as pH, temperature, organic solvents, and toxic components. Immobilized
microbial cells can also be handled more easily and recovered from the solution without difficulty [16]. Continuous
processes can be operated at a high cell density without loss
Downloaded by [Julius Akinbomi] at 23:03 21 February 2013
2
S. Youngsukkasem et al.
of microbial cells even at high dilution rates, which results
in a higher bioreactor volumetric productivity [3]. Among
various cell immobilization methods, encapsulation, or cell
containment, is highly attractive. Upon encapsulation, the
cells are retained in a capsule made by a membrane permeable to nutrients and metabolites [17]. The necessary
properties of the membrane include having a good substrate
and easier product transfer, protection of the cells, and no
leakage from the capsules. Encapsulation has been applied
to various bioprocesses such as whole-cell biocatalysts, artificial cells, and biosorbents [16,18]. Another type of cell
encapsulation has cells contained behind a barrier, and can
be achieved by using microporous membrane filters, by
entrapment of cells in a microcapsule, or by cell immobilization onto an interaction surface of two immiscible liquids
[16,19–22]. This method could also be an interesting way
to retain the cells by using a synthetic polymeric membrane
for continuous processes, and to protect the cells in order to
enhance biogas productivity. However, we have found no
report in the literature on using encapsulation or cell containment technology with digesting bacteria in continuous
process and cell protection testing for biogas production.
The aims of this work were to investigate the performance of methane-producing bacteria encased in synthetic
polymeric membranes in a long-term digester with different
organic loading rates (OLRs) for rapid biogas production,
and to investigate its protective effect against limonene. The
total volume of biogas and methane, and the total amounts
and composition of the volatile fatty acids (VFAs) were also
determined regularly.
Materials and methods
Anaerobic culture preparation
Anaerobic cultivations were prepared following a method
described elsewhere [23,24]. The inoculum was obtained
from a 3000 m3 municipal solid waste digester operating at
thermophilic (55◦ C) conditions (Borås Energy & Environment AB, Sweden). It was then degassed by pre-incubating
at 55◦ C for 3 days. The active anaerobic culture was homogenized and passed through a sieve with a pore size of
1.0 mm in order to separate any remaining large particles.
The sludge was then centrifuged at 14, 000 × g for 10 min
to separate the supernatant and bacteria, which were thereafter used as an inoculum for cell containment in different
experiments.
Synthetic medium
The nutrients for methanogenic bacteria were mimicked
from the normal products of the first stage in digestion, i.e.
the hydrolysis. The composition of this synthetic medium
was the carbon source, including acetic acid, propionic
acid, butyric acid, methanol, and glucose mixed at a mass
ratio of 3:1:1:1:1; a basal medium [25] was used for
addition of nutrients necessary for the anaerobic cultures.
Table 1. Composition of the basal medium used
to supply nutrients in the synthetic medium.
Components
NH4 Cl
MgSO4 .7H2 O
KCl
Na2 S .9H2 O
CaCl2 .2H2 O
CaCl2 .2H2 O
(NH4 )2HPO4
FeCl2 .4H2 O
CoCl2 .6H2 O
KI
MnCl2 .4H2 O
CuCl2 .2H2 O
ZnCl2
AlCl3 .6H2 O
Na2 MoO4 .2H2 O
H3 BO3
NiCl2 .6H2 O
Na2 WO4 .2H2 O
Na2 SeO3
Cysteine
Amount (mg/L)
1200
400
400
300
50
50
80
40
10
10
0.5
0.5
0.5
0.5
0.5
0.5
0.5
0.5
0.5
10
The composition of the medium is shown in Table 1. The
pH of the mixture of synthetic substrates was adjusted to
7.0 ± 0.5.
Membrane sachet preparations and cell containment
procedure
A cell containment technique was employed following
the method previously described [26]. Flat plain PVDF
(polyvinylidene fluoride, Durapore® ) membranes supplied
by Thermo Fisher Scientific Inc. (Sweden) were used as a
synthetic membrane supporting material. The membranes
were cut into rectangular shapes of 6 × 6 cm and folded to
create membrane pockets of 3 × 6 cm2 . These membrane
sheets were then heat-sealed (HPL 450 AS, Hawo, Germany) on two sides with heating and cooling times of 4.5
and 4.5 s, leaving one side open for the insertion of the
inoculums. Bacterial inoculums (3 g) were then injected
carefully into the synthetic membrane pockets, and sealed
accordingly. The sachets containing the inoculum were used
immediately for biogas production.
Protective effect of sachets in batch anaerobic digestion
process
The batch digestion processes were performed in order to
examine the protective effect of the sachets on the bacteria
against limonene as the inhibitor. Limonene (Fluka, Sweden) was mixed with a synthetic medium in concentrations
of 0 (control), 1, 2, and 3% (v/v). In each digesting reactor,
three sachets were placed in a synthetic medium. The reactors used were serum glass bottles with 150 mL working
volume, closed with butyl rubber seals and plastic caps.
Environmental Technology
Downloaded by [Julius Akinbomi] at 23:03 21 February 2013
Anaerobic digestions with free cells were performed in parallel with identical conditions as reference. The headspace
of each bottle was flushed with 80% nitrogen and 20% carbon dioxide gas mix to attain the anaerobic conditions and
help to keep the pH neutral at the beginning of the process
[23]. Digestions were carried out under thermophilic conditions at 55 ± 1◦ C in an incubator for 10 days. All digesters
were shaken two times per day in order to have better contact
between the inoculums and substrate.
Effect of high loading rates in semi-continuous
anaerobic digestion process
A long retention time of, for example, 30 days is usually
a challenge in anaerobic digestion processes, resulting in
large digestion reactors and challenging their economical
feasibility. In this work, the efficiency of encased bacteria
at different feeding rates of synthetic wastewater [27] was
investigated, and compared with free cells in a long-term,
semi-continuous process.
In order to investigate the performance of cells encased
in synthetic sachets from a long-term perspective, a semicontinuous process mode was employed (Figure 1). In fact,
the reactor was fed continuously by a recirculating pump.
However, considering the system boundary around both
the digester and the recirculating pump, the system can
be defined as semi-continuous. The reactors were made inhouse from Plexiglas® , had a total inner volume of 1.5 L and
were fitted with rubber seals and an outlet for the biogas. In
this experiment, 37 sachets containing a total of 111 g bacteria were added into 1.1 L medium in each reactor. In parallel,
the same amount of the bacteria was used as inoculum in
the reference reactor using free cells. Thermophilic conditions were maintained at 55 ± 1◦ C throughout the process
by passing warm water from a water bath through the reactor
Figure 1. Schematic diagram of semi-continuous digestion
process. The numbering indicates (1) sachets containing
methane-forming bacteria, (2) warm water jacket, (3) water heater,
(4) gas analyser, (5) purge container, (6) feed container, (7)
peristaltic pump, and (8) controller.
3
Table 2. Experimental set-up of organic loading rates condition during the semi-continuous process of testing free
and encased methane-producing bacteria during anaerobic
digestion.
Organic loading
rate (OLR)
(gCOD/L.day)
2.5
5.0
7.5
10.0
15.0
20.0
Working
volume
of reactor
(mL)
Synthetic
medium
strength
(g COD/L)
Volume of
medium
(mL)
1100
1100
1100
1100
1100
1100
9.33
9.33
9.33
9.33
93.3
93.3
294
588
884
1100
177
354
jacket (Figure 1). The OLR was then gradually increased by
increasing the volume of medium fed (Table 2). Each loading rate was maintained for 7 days, giving a total digestion
process of 42 days. During the digestion, the medium was
circulated continuously with a flow rate of 150 mL/min.
The key parameters to monitor anaerobic digestion processes are normally biogas production and volatile fatty
acid accumulation, which gives an indication of how well
the digesting bacteria can handle the degradation of a
substrate [24,28].
Analytical methods
The volume of biogas was automatically recorded using
a data acquisition system (AMPTS, Bioprocess Control
Sweden AB, Sweden). The VFAs were analysed using a
HPLC system (Waters, Milford, MA) equipped with an
ion-exchange column (Aminex HPX-87H, Bio-Rad, Hercules, CA) working at 60◦ C using 5 mM sulphuric acid as
an eluent with a flow rate of 0.6 mL/min, and a UV detector
(Waters 2414) for detection of the VFAs. The methane was
quantified using a gas chromatograph (Auto system, PerkinElmer, USA) on a packed column (Perkin-Elmer, 6’ × 1.8”
OD, 80/100 Mesh, USA) and a thermal conductivity detector (Perkin-Elmer, USA) with an injection temperature of
150◦ C. Nitrogen at a flow rate of 20 mL/min at 60◦ C was
used as carrier gas. A 0.25 mL syringe (VICI, precisions
sampling Inc., USA) was employed for the gas sampling.
The obtained peak area was compared with a standard
methane gas analysed at the same condition (STP: 273.15 K,
101.325 kPa). All experiments were operated in duplicate
and the results are shown as means ± standard deviation.
Results and discussion
Protective effect of the sachet membranes
The presence of toxic compounds is a challenge for digestion processes. A variety of inorganic and organic materials,
which are commonly present in industrial biogas processes,
can be toxic in anaerobic digesters. D-limonene, a major
component of peel oil in citrus fruits, is an antibacterial
4
S. Youngsukkasem et al.
production began immediately on the first day of digestion
and continued until the last day. The maximum daily
methane production occurred on the fifth day, with methane
volumes at 0, 1, 2, and 3% limonene being 23.65, 20.03,
23.06, and 19.00 mL, respectively. It should be noted that
the additional barrier of the membrane cases did not affect
the diffusion of substrate into the encased cell and products
out of it. These results reveal that methane-producing bacteria encased in a synthetic polymeric membrane (PVDF) are
better protected against the inhibitor as opposed to free cells.
(a) 180
CH4 (NmL/g COD)
160
0% Limonene
1% Limonene
2% Limonene
3% Limonene
140
120
100
80
60
40
20
Biogas production at different organic loading rates
0
(b) 140
0% Limonene
1% Limonene
2% Limonene
3% Limonene
CH4 (NmL/g COD)
Downloaded by [Julius Akinbomi] at 23:03 21 February 2013
120
100
80
60
40
20
0
1
2
3
4
5
6
7
8
9
10
Time (Days)
Figure 2. Accumulated methane productions with different concentrations of limonene in synthetic medium. (a) using free cells;
(b) using encased methane-producing bacteria.
agent, which is effective in killing bacteria such as Mycobacterium tuberculosis and Streptococcus mutans and a variety
of fungi at very low concentrations [8,29–31]. In this work,
the efficiency of encased methane-producing bacteria in
membrane sachets was investigated, using 0–3% limonene
as a model inhibitor. Methane production was determined
regularly, with results shown in Figure 2.
Daily methane production by the encased bacteria
increased dramatically up until the fifth day of digestion for
most treatments, regardless of the limonene concentration.
Then, it decreased until the last day of digestion, limited
by the amount of substrate given. With both encased cells
and free cells, a limonene concentration of 3% tended to
impact on the lowest methane production compared with
2%, 1% and 0%. However, methane production from free
cells with different concentrations of limonene was substantial. The maximum methane production from free cells
was seen on the fifth day of digestion, with methane volumes of 31.22, 27.53, 6.56, and 8.43 mL, from free cells
with 0, 1, 2, and 3% limonene, respectively. In contrast,
the maximum methane volume from encased cells was
practically unaffected by the presence of limonene. Methane
Anaerobic digestion in a semi-continuous digester was performed with encased cells and free cells as a reference. It
was performed under thermophilic conditions for 42 days
while at the same time increasing the OLR until no gas
production occurred.
At the beginning of the process, the suspended sludge
containing methane-producing bacteria sedimented at the
bottom of the reactor even though the substrate was always
circulated through the reactor. Biogas was produced and
the bubbles rose to the top of the reactor. As part of the
semi-continuous process, a certain volume of the medium
(Table 1) in the reactor was withdrawn and replaced with
fresh medium, causing some free cells to be washed out,
which was not the case for the encased cells in the sachets.
The sachets containing bacteria swelled immediately after
the first day of digestion, resulting in an increase of the
sachet volume inside the reactor (Figure 3). Moreover, gas
bubbles were clearly observed in the medium throughout
the digestion process, which indicates that gas was produced
inside the sachets and subsequently released. Most of the
sachets stayed intact and had no cell leakage throughout the
process.
The results of digestion are presented in Figure 4(a).
With low OLR (2.5 g COD/L.day), biogas was produced
continuously by both the free and the encased cells, although
there was a substantial difference in their biogas production rates. The average daily biogas productions from
the encased and free cells were 916 and 137 mL, respectively. After 7 days, the organic load was doubled to
5.0 g COD/L.day. This resulted in increasing biogas production by both the free and encased methane-producing
bacteria. The same trend of more biogas from encased cells
than free cells was observed, which was 1837 vs. 729 mL per
day, respectively. On the other hand, when the loading rate
was increased to 7.5 g COD/L.day, the biogas production
by the encased cells still increased to an average of 2889 mL
per day. In contrast, for the reactor with free cells, the biogas production declined as the loading rate was increased
to 7.5 g COD/L.day, and digestion completely failed by the
18th day.
With encased cells in the semi-continuous processes,
the loading rate was further increased to 10, 15, and
20 g COD/L.day. Biogas was still produced continuously,
Downloaded by [Julius Akinbomi] at 23:03 21 February 2013
Environmental Technology
5
Figure 3. Semi-continuous anaerobic digestion systems of membrane bioreactor including encased bacteria. (a) The 1st day of digestion
and (b) the 2nd day of digestion.
despite these very high OLRs. The average biogas produced
at loading rates of 10 and 15 g COD/L.day were 3552 and
5441 mL per day, respectively. On the other hand, at an OLR
of 20 g COD/L.day biogas production decreased dramatically. The average biogas volume produced at this OLR was
3606 mL. The maximum substrate tolerance level refers to
the highest organic load tolerated by the microbial community without any decrease in their activity. In a similar
study for a traditional biogas process, Liu et al. [32] reported
a maximum and stable biogas production rate of 4.25 m3
(m3 d)−1 at an OLR of 6.0 kg VS/m3 .day by thermophilic
anaerobic digestion of municipal biomass waste. However,
a higher OLR of 8.0 kg VS/m3 .day resulted in a maximum
methane production rate of 2.94 m3 /m3 .day [33]. In our
study, the highest load introduced to the encased reactors
was 20 g COD/L.day, after which daily biogas production
declined.
These results reveal that the encased methane-forming
bacteria were able to tolerate higher OLR than the free cells,
and were able to produce more biogas in a semi-continuous
process. This means that anaerobic digestion systems with
encased methane-forming bacteria have a better system
capacity to prevent washing out of cells, and to produce
biogas faster than the free cells.
Degradation of volatile fatty acids by free cells and
encased cells
VFAs serve as substrate for methane-forming bacteria,
resulting in the production of methane. Their production and
consumption can give an accurate idea of the balance, in the
sense that the different bacteria can have sufficient production and consumption rates. Moreover, high concentrations
of VFAs can inhibit the anaerobic process [4]; therefore, it
is important to monitor these compounds. In this work, the
degradation and composition of total VFAs was regularly
analysed in the culture of both free and encased cells during
the semi-continuous processes. The results are summarized
in Figure 4(b).
At low OLR of 2.5 g COD/L.day, the total VFAs in the
reactors of both the free and the encased cells decreased
continuously from 5 g/L to 2.34 and 0.92 g/L, respectively,
in accordance with the increasing biogas production. As
the loading rate increased to 5 g COD/L.day, the total VFA
concentrations in both the encased and free cell cultures
were stable between the 8th and 14th day of digestion.
However, the values started to increase for the free cells,
from 2.47 to 5.70 g/L, when the OLR was increased to
7.5 g COD/L.day. This is an indication that the anaerobic
process had failed, since no biogas was produced from the
VFAs and washing out of cells could be observed. On the
other hand, the encased cells were able to keep the levels of total VFAs low as the loading rate was increased
from 5 to both 7.5 and 10 g COD/L.day. The variation
in the measured level ranged only from 1.45–0.25 g/L.
As the OLR was increased to 15 g COD/L.day, the deterioration of total VFAs slightly decreased, meaning that
higher levels of VFAs were detected. Nevertheless, the
concentrations increased continuously during the last period
Downloaded by [Julius Akinbomi] at 23:03 21 February 2013
6
S. Youngsukkasem et al.
Figure 4. The digestion performance of free and encased methane-forming bacteria in a semi-continuous anaerobic process. (a) daily
biogas productions and (b) accumulated total volatile fatty acids (VFAs).
(20 g COD/L.day), meaning that the highest tolerable concentration of the cells had been reached. From the 35th day
until the end of the experiment, the total VFAs concentration in the bioreactor increased from 10 to 40 g/L. With
increasing OLRs, the anaerobic reactor suffered from an
increase of VFA concentration, resulting in inhibition of
the methanogenic bacteria [33]. Forbes et al. [34] reported
that an OLR of 9 g COD/L.day at a hydraulic retention time
(HRT) of 1 day made the thermophilic bioreactor unstable, shown by a high concentration of VFAs in the effluent.
Wijekoon et al. [35] showed the total VFA concentration
to increase from 2.5 to 4.7 and 7.0 g/L with increased OLR
of 5.1 to 8.1 and 12 kg COD/L.day, and the process was
not stable at an OLR of 12 kg COD/L.day due to high
accumulated VFAs.
The composition of VFAs, including acetate, propionate, and butyrate, was also investigated. For the encased
bacterial system, the results (Figure 5) show that acetate,
propionate and butyrate concentrations decreased dramatically, even though the organic loading was increased continuously from 2.5 to 10.0 g COD/L.day. However, when an
OLR of 15 or 20 g COD/L.day was introduced to the reactor, acetate, propionate, and butyrate accumulated, starting
with propionate (Figure 5(b)). In contrast, Figure 5(a)
shows the trend of acetate, propionate, and butyrate being
consumed by free cells in the semi-continuous anaerobic
reactor. Acetate, propionate, and butyrate concentrations
decreased slightly during the 7 days of 2.5 g COD/L.day.
It then increased when 7.5 g COD/L.day was added to the
reactor.
Finally, it is worth noting the lower VFA values in
the reactor with the encased cells compared with the free
cells (Figure 5). This means higher degradation of VFAs
by encased bacteria than that of the suspended cells. This
Downloaded by [Julius Akinbomi] at 23:03 21 February 2013
Environmental Technology
Figure 5.
7
Profile of acetate, propionate, and butyrate during semi-continuous anaerobic digestion by (a) free cells and (b) encased cells.
is probably a consequence of the higher concentration or
higher activity of the encased bacteria, resulting in a higher
substrate degradation rate.
Conclusion
Encasing methane-producing bacteria in semi-permeable
membrane (PVDF) is a successful technique for enhancing
biogas production. PVDF had the ability to retain the cells
in the reactor. A protective effect of the PVDF membranes
against limonene, a potent inhibitor of microorganisms, was
also obtained. It was shown that the encased bacteria could
produce methane faster than free cells from substrates containing limonene concentrations of 0, 1, 2, and 3%. Encased
bacteria were able to produce more biogas than free cells
during the process, lasting for 42 days with the maximum
OLR of 20 g COD/L.day, while the free cells had already
failed at OLR 7.5 g COD/L.day. However, further research
is required to investigate the potential of different kinds
of membrane for cell encasement, in order to improve the
industrial development of biogas production, and anaerobic
digestion processes longer than 42 days should be studied in order to investigate the life span of the membrane.
Furthermore, the protective effect of sachets towards other
inhibitors of the anaerobic digestion process would be
interesting to study.
Acknowledgements
This work was financially supported by the Swedish Research
Council, European Commission program EM-Euro Asia, Rajamangala University of Technology Isan (Thailand), and Lagos
State University (Nigeria).
References
[1] Deublein D, Steinhauser A. Biogas from waste and renewable resources. Germany: Wiley-VCH Verlag GmbH & Co.
KGaA; 2008.
[2] Griffin ME, McMahon KD, Mackie RI, Raskin L.
Methanogenic population dynamics during start-up of
8
[3]
[4]
[5]
[6]
[7]
Downloaded by [Julius Akinbomi] at 23:03 21 February 2013
[8]
[9]
[10]
[11]
[12]
[13]
[14]
[15]
[16]
[17]
[18]
[19]
S. Youngsukkasem et al.
anaerobic digesters treating municipal solid waste and
biosolids. Biotechnol Bioeng. 1997;57:251–379.
Gerardi MH. The microbiology of anaerobic digesters.
Hoboken (NJ): Wiley; 2003.
Chen Y, Cheng JJ, Creamer KS. Inhibition of anaerobic digestion process: a review. Bioresour Technol.
2008;99:4044–4064.
Appels L, Baeyens J, Degrève J, Dewil R. Principles
and potential of the anaerobic digestion of waste-activated
sludge. Progr Energy Combust Sci. 2008;34:755–781.
Long JH, Aziz TN, Reyes Iii FLdl, Ducoste JJ. Anaerobic co-digestion of fat, oil, and grease (FOG): a review of
gas production and process limitations. Process Saf Environ
Protect. 2012;90:231–245.
Martín MA, Siles JA, Chica AF, Martín A. Biomethanization
of orange peel waste. Bioresour Technol. 2010;101:8993–
8999.
Singh P, Shukla R, Prakash B, Kumar A, Singh S, Mishra
PK, Dubey NK. Chemical profile, antifungal, antiaflatoxigenic and antioxidant activity of Citrus maxima Burm.
and Citrus sinensis (L.) Osbeck essential oils and their
cyclic monoterpene, dl-limonene. Food Chem Toxicol.
2010;48:1734–1740.
Winniczuk PP, Parish ME. Minimum inhibitory concentrations of antimicrobials against micro-organisms related to
citrus juice. Food Microbiol. 1997;14:373–381.
Lappas CM, Lappas NT. d-Limonene modulates T lymphocyte activity and viability. Cell Immunol. 2012;279:30–41.
Wilkins MR. Effect of orange peel oil on ethanol production
by Zymomonas mobilis. Biomass Bioenergy 2009;33:538–
541.
Oberoi HS, Vadlani PV, Nanjundaswamy A, Bansal S, Singh
S, Kaur S, Babbar N. Enhanced ethanol production from
Kinnow mandarin (Citrus reticulata) waste via a statistically
optimized simultaneous saccharification and fermentation
process. Bioresour Technol. 2011;102:1593–1601.
Mizuki E, Akao T, Saruwatari T. Inhibitory effect of
Citrus unshu peel on anaerobic digestion. Biol Wastes
1990;33:161–168.
Forgács G, Pourbafrani M, Niklasson C, Taherzadeh MJ,
Hováth IS. Methane production from citrus wastes: process development and cost estimation. J Chem Technol
Biotechnol. 2012;87:250–255.
Pourbafrani M, Talebnia F, Niklasson C, Taherzadeh
MJ. Protective effect of encapsulation in fermentation of
limonene-contained media and orange peel hydrolyzate. Int
J Mol Sci. 2007;8:777–787.
Park JK, Chang HN. Microencapsulation of microbial cells.
Biotechnol Adv. 2000;18:303–319.
Talebnia F. Ethanol production from cellulosic biomass
by encapsulated Saccharomyces cerevisiae [PhD thesis].
Chalmers University of Technology; 2008.
Nedovic V, Kalusevic A, Manojlovic V, Levic S, Bugarski
B. An overview of encapsulation technologies for food
applications. Procedia Food Sci. 2011;1:1806–1815.
Kourkoutas Y, Bekatorou A, Banat IM, Marchant R, Koutinas AA. Immobilization technologies and support materials
suitable in alcohol beverages production: a review. Food
Microbiol. 2004;21:377–397.
[20] Nafea EH, L.A P-WAM, Martens PJ. Immunoisolating
semi-permeable membranes for cell encapsulation: focus on
hydrogels. J Controlled Release. 2011;154:110–122.
[21] Uludag H, De Vos P, Tresco PA. Technology of mammalian
cell encapsulation. Adv Drug Delivery Rev. 2000;42:29–64.
[22] Randall CL, Kalinin YV, Jamal M, Shah A, Gracias DH.
Self-folding immunoprotective cell encapsulation devices.
Nanomed Nanotechnol Biol Med. 2011;7:686–689.
[23] Angelidaki I, Alves M, Bolzonella D, Borzacconi L, Campos
JL, Guwy AJ, Kalyuzhnyi S, Jenicek P, van Lier JB. Defining
the biomethane potential (BMP) of solid organic wastes and
energy crops: a proposed protocol for batch assays. Water
Sci Technol. 2009;59:927–934.
[24] Hansen TL, Schmidt JE, Angelidaki I, Marca E, Jansen JlC,
Mosbæk H, Christensen TH. Method for determination of
methane potentials of solid organic waste. Waste Manage.
2004;24:393–400.
[25] Isci A, Demirer GN. Biogas production potential from cotton
wastes. Renewable Energy. 2007;32:750–757.
[26] Youngsukkasem S, Rakshit KS, Taherzadeh MJ. Biogas
production by encapsulated methane producing bacteria.
Bioresources 2012;7:56–65.
[27] Richards BK, Cummings RJ, Jewell WJ, Herndon FG.
High solids anaerobic methane fermentation of sorghum and
cellulose. Biomass Bioenergy. 1991;1:47–53.
[28] Salminen EA, Rintala JA. Semi-continuous anaerobic digestion of solid poultry slaughterhouse waste: effect of hydraulic
retention time and loading. Water Res. 2002;36:3175–3182.
[29] Caccioni DRL, Guizzardi M, Biondi DM, Agatino R,
Ruberto G. Relationship between volatile components of citrus fruit essential oils and antimicrobial action on Penicillium
digitatum and Penicillium italicum. Int J Food Microbiol.
1998;43:73–79.
[30] Settanni L, Palazzolo E, Guarrasi V, Aleo A, Mammina
C, Moschetti G, Germanà MA. Inhibition of foodborne
pathogen bacteria by essential oils extracted from citrus fruits
cultivated in Sicily. Food Control. 2012;26:326–330.
[31] Espina L, Somolinos M, Lorán S, Conchello P, García D,
Pagán R. Chemical composition of commercial citrus fruit
essential oils and evaluation of their antimicrobial activity acting alone or in combined processes. Food Control.
2011;22:896–902.
[32] Liu X, Gao X, Wang W, Zheng L, Zhou Y, Sun Y. Pilot-scale
anaerobic co-digestion of municipal biomass waste: focusing on biogas production and GHG reduction. Renewable
Energy. 2012;44:463–468.
[33] Liu X, Wang W, Shi Y, Zheng L, Gao X, Qiao W, Zhou
Y. Pilot-scale anaerobic co-digestion of municipal biomass
waste and waste activated sludge in China: effect of organic
loading rate. Waste Manage. 2012;32:2056–2060.
[34] Forbes C, O’Reilly C, McLaughlin L, Gilleran G, Tuohy
M, Colleran E. Application of high rate, high temperature
anaerobic digestion to fungal thermozyme hydrolysates from
carbohydrate wastes. Water Res. 2009;43:2531–2539.
[35] Wijekoon KC, Visvanathan C, Abeynayaka A. Effect
of organic loading rate on VFA production, organic
matter removal and microbial activity of a two-stage
thermophilic anaerobic membrane bioreactor. Bioresour
Technol. 2011;102:5353–5360.
Paper II
Energies 2015, 8, 4253-4272; doi:10.3390/en8054253
OPEN ACCESS
energies
ISSN 1996-1073
www.mdpi.com/journal/energies
Article
Evaluation of Fermentative Hydrogen Production from Single
and Mixed Fruit Wastes
Julius Akinbomi * and Mohammad J. Taherzadeh
Swedish Centre for Resource Recovery, University of Borås, 501 90 Borås, Sweden;
E-Mail: [email protected]
* Author to whom correspondence should be addressed; E-Mail: [email protected];
Tel.: +46-334-354-585.
Academic Editor: Calliope Panoutsou
Received: 5 February 2015 / Accepted: 5 May 2015 / Published: 12 May 2015
Abstract: The economic viability of employing dark fermentative hydrogen from whole
fruit wastes as a green alternative to fossil fuels is limited by low hydrogen yield due to the
inhibitory effect of some metabolites in the fermentation medium. In exploring means of
increasing hydrogen production from fruit wastes, including orange, apple, banana, grape
and melon, the present study assessed the hydrogen production potential of singly-fermented
fruits as compared to the fermentation of mixed fruits. The fruit feedstock was subjected to
varying hydraulic retention times (HRTs) in a continuous fermentation process at 55 °C for
47 days. The weight distributions of the first, second and third fruit mixtures were 70%,
50% and 20% orange share, respectively, while the residual weight was shared equally by
the other fruits. The results indicated that there was an improvement in cumulative
hydrogen yield from all of the feedstock when the HRT was five days. Based on the results
obtained, apple as a single fruit and a fruit mixture with 20% orange share have the most
improved cumulative hydrogen yields of 504 (29.5% of theoretical yield) and 513 mL/g
volatile solid (VS) (30% of theoretical yield ), respectively, when compared to other fruits.
Keywords: whole fruit wastes; singly-digested fruits; mixing proportion; biohydrogen;
retention time; significant effect
Energies 2015, 8
4254
1. Introduction
The quest for renewable, efficient and environmentally-friendly alternative energy sources to fossil
fuels has stimulated intense research studies on fermentative hydrogen (H2) production from biomass.
Among other renewable energy sources (solar, hydro-power, wind and geothermal), fermentative H2
production has received remarkable interest due to its striking properties, including having very high
energy content per unit mass and being a clean energy carrier, as it forms only water vapor
during combustion [1–3]. Fermentative H2 production can facilitate the quick transition of the
hydrocarbon-based economy to a hydrogen-based economy, especially in the transport sector. Fuel cell
electric vehicles powered by fermentative H2 are zero emission vehicles that could be used as green
energy technology to tackle the challenge of depleted fossil fuel reserves and pollution associated with
conventional transport fuels. Furthermore, the considerable attention on fermentative H2 is also due to
the reliability of the continuous supply of feedstock, which is inevitably generated from daily human
and animal activities. Among the feedstock available for H2 production, fruit wastes have relative great
economic and environmental potential due to the large quantities of wastes generated from fruit
consumption and industrial processing (10%–65% of raw fruit) [4]. The application of fruit wastes as
feedstock for H2 production is an eco-friendly process, since littered fruit wastes could constitute a health
nuisance to people and the environment. Meanwhile, due to low H2 yield from the fermentation
process, most of the hydrogen currently used in various industrial applications is obtained from
non-green sources, including steam reforming of natural gas, water electrolysis and coal gasification [5–7].
Low H2 yield during the fermentation process is attributed to, among other factors, the natural tendency of
the fermentation process to be optimized to produce cell biomass instead of H2. In a dark fermentation
process, 12 mol H2/mol glucose could be theoretically obtained from complete conversion of glucose to
H2 and carbon dioxide (Equation (1)), but the reaction is thermodynamically impossible due to the
production of a large quantity of by-products (volatile fatty acids (VFAs), alcohols and lactate)
associated with H2 production. The thermodynamic constraints make the maximum attainable H2
yields to be 4 and 2 mol/mol glucose if the associated by-products are acetate and butyrate,
respectively (Equations (2) and (3)) [8,9]:
C6H12O6 + 6H20 o 12H2 + 6CO2 (¨G° = +3.2 kJ)
(1)
C6H12O6 + 2H20 o 4H2 + 2CO2 + 2CH3COOH(acetate) (¨G° = í206 kJ)
(2)
C6H12O6 o 2H2 + 2CO2 CH3CH2CH2COOH(butyrate) (¨G° = í254 kJ)
(3)
The low H2 from the fermentation process could also be attributed to feedstock inhibition including
the antimicrobial inhibition of flavor compounds in fruit. During the ripening process, fruits usually
produce flavor compounds (esters, alcohols, aldehydes, ketones, lactones and terpenoids), which are
used as natural defense mechanisms in plants against microbial invasion (Table 1) [10–24]. Although
the amount of fruit flavors in fruits is not high (usually 0.001%–0.01% of the fruit’s fresh weight),
the antimicrobial effect of the flavor compounds cannot be ignored. The positive effect of the flavor
compounds in fruit is responsible for fruit freshness and the long shelf life of fruits, while a negative
effect has been observed to be responsible for the inhibition of bacterial activities during the fermentation
process. For instance, D-limonene, which is a citrus flavor belonging to a class of terpenoids, was found
Energies 2015, 8
4255
to have an antimicrobial effect at a very low concentration of 0.01% w/v [11,25–27] and to cause the
failure of the anaerobic digestion process, even at a very low concentration of 400 µL/L [26]. As a
result, some strategic approaches have been developed to mitigate the antimicrobial effect of limonene
and, consequently, to improve bioenergy production during anaerobic digestion. In a study by
Youngsukkasem et al. [28], an encapsulated membrane was effectively used to reduce the inhibitory
effect of D-limonene on the fermentative bacteria. Membrane-encased bacteria were observed to
tolerate up to 3% limonene in the synthetic medium with consequent higher methane production than
free cells. Wikandiri et al. [29] also developed a membrane bioreactor that could tolerate between 5 and
10 g/L of limonene in the feedstock for biomethane generation. Similarly, various techniques, including
feedstock and inoculum pretreatment, optimal inoculum-to-substrate ratio, gas sparging, bioreactor
design and a two-stage fermentation system, among others, have been employed for improving the
recovery efficiency of fermentative H2 from food wastes, especially fruit wastes [30–33].
Table 1. Fruit flavors in some fruits.
Fruit
Orange
Banana
Apple
Grape
Melon
Fruit Flavor
Flavor compound
Hexanal and nonanal
Octanol, 3-methyl butanol
Į-pinene, car-3-ene, myrcene and limonene
Pentanone, heptanone, undecanone
Butanal, hexanal and E-2-hexanal
1-butanol, 2-pentanol, 3-methyl-1-butanol, 1-hexanol and eugenol
Ethyl acetate, butyl acetate, 2-methyl propyl acetate, hexyl acetate,
hexyl butanoate and butyl butanoate
n-Hexanal, E-2-hexenal, nonanal, acetaldehyde
Hexanol and butanol
Car-3-ene
Ethyl butanoate, ethyl -2-methylbutanoate, hexyl acetate, etc.
Epicatechin
Hexanal
Octanol and hexanol
Hexyl acetate, ethyl acetate and ethyl hexanoate
Quercetin and epicatechin
Nonanal, benzaldehyde and E-2-nonenal
Ethyl 2-methyl propyl acetate and 2-methyl butyl acetate
References
Flavor group
Aldehydes
Alcohols
Terpenoid
Ketones
Aldehydes
Alcohol
[10]
[10,23]
[10,11]
[12,13]
[12,13]
[12,13]
Esters
[12,13]
Aldehydes
Alcohols
Terpenoid
Ester
Polyphenol
Aldehyde
Alcohols
Esters
Polyphenol
Aldehydes
Esters
[10,14]
[14]
[10]
[14,15,23]
[16,17]
[10]
[14,18]
[10,18,19]
[16]
[20–22]
[20–22]
Meanwhile, in most research work on fermentative H2 from fruit wastes, fruit peels are often used
as feedstock, since they are discarded during consumption and industrial processing. However,
research should also be focused on the application of whole fruit wastes as feedstock, since large
quantities of whole fruit wastes are generated during harvest, transportation and storage, due to microbial
or pest attack. The objective of this study was therefore to investigate the effect of combined varying
hydraulic retention time (HRT) and fruit mixing ratio on fermentative H2 yield from whole fruit wastes in
order to have an idea of how to manage whole fruit wastes feedstock for optimal fermentative hydrogen
production. The potential of fruit biodegradability and fermentative hydrogen production could be
enhanced or lessened by the characteristics of fruit mixtures (nutrient composition, carbon to nitrogen
Energies 2015, 8
4256
ratio, toxicity) obtained when two or more fruits were combined. Besides nutrient composition, the
inherent antimicrobial flavor compounds in fruits could also be a decisive factor in the overall effect.
Unlike single fruit waste, mixed fruit wastes have the advantage of mutual interaction between various
fruits in the fruit mixture, which might result in varying degrees of inhibitory and enhanced effects due
to factors, such as additive (a combined effect equal to the sum of the individual effects), synergistic
(a combined effect greater than the sum of the individual effects) and antagonistic effects (a combined
effect less than the individual effects).
2. Results and Discussion
The performance of the fermentative H2 production process from the whole fruit wastes of orange,
banana, apple, grape and melon was evaluated by measuring the hydrogen yield and volatile fatty acid
(VFA) productions. The hydraulic retention times (HRTs) were reduced as the organic loading rates
(OLR) were subsequently increased in the experiments. Although the mass of the different fruit wastes
fed to the reactors were equal, their OLRs were not the same, because of different percentages of volatile
solids in each fruit waste. The fermentation process was in three phases, Phase 1, 2 and 3, which
corresponded to HRT of 8.6, 5 and 3 days, respectively. The continuous stirred tank reactors (CSTRs)
were started up with an initial HRT of 8.6 days, which was the first phase of the fermentation process
spanning a period of 15 days. During the second phase, which covered the period between the 16th and
30th days, the HRT was decreased from 8.6 down to five days. The HRT was furthered reduced to
three days during the third phase, which spanned the period between the 31st and 47th days.
2.1. Hydrogen Production Yields from Singly-Digested Fruits
The hydrogen production from the singly-digested fruits was monitored throughout the three phases
of the 47-day period. During the first phase (HRT of 8.6 days), hydrogen yields from the fermentation
of individual fruits increased, except the yield from grape, which decreased sharply (Figure 1) due to
the decrease in pH values. The highest (493 mL/gVSadded; VS, volatile solid) and lowest
(216 mL/gVSadded) average hydrogen yields during the first phase were obtained from apple and
melon, respectively. The hydrogen yields of all of the fruits increased during the second phase
(HRT of five days), with apple and melon still producing the highest and lowest average hydrogen
yields of 635 and 352 mL/gVSadded, respectively. The average hydrogen yields during the third phase
(HRT of three days), however, decreased with apple and grape, producing the highest and lowest
yields of 440 and 182 mL/gVS, respectively.
Energies 2015, 8
4257
Figure 1. Hydrogen yields from the fermentation of single fruits.
2.2. Hydrogen Production Yields from Mixed Fruits
Hydrogen yields from the fermentation of the three fruit mixtures, including 70% (Mix 1),
50% (Mix 2) and 20% (Mix 3) orange share, also followed a similar pattern, but with better
performance than the fermentation of individual fruits, except apple (Figure 2). During the first phase,
hydrogen yields from Mix 1 and Mix 3 increased, but there was a slight decrease in hydrogen yield
from Mix 2. The highest average hydrogen yield of 523 mL/gVS obtained during the first phase was
from the fermentation of Mix 3. In the second phase, hydrogen yields from Mix 1 and Mix 2 increased,
while there was a slight reduction in hydrogen yield from Mix 3. However, Mix 3 produced the highest
average hydrogen yield of 553 mL/gVS at the end of the second phase. The hydrogen yields from the
three fruit mixtures decreased during the last phase, with Mix 3 still producing the highest yield of
491 mL/gVS. Mix 3 seemed to perform averagely better than other mixed fruits in terms of hydrogen
yield. This might be due to the ineffectiveness of the toxicity of the individual inhibitors as a
consequent effect of the mutual interaction of the different flavor compounds in the fruit mixture.
Besides, since limonene constitutes the major component of citrus essential oils, the reduction of orange
percentage in Mix 3 could also impact the reduced antimicrobial effect of limonene in the mixture.
Moreover, the use of an appropriate amount of nutrient combination in any mixture of fruit waste is
necessary for optimal hydrogen yields from the fermentation process, as excess or insufficient nutrients
may affect the stability and gas productivity of the process [34]. The macronutrients (Na, K, Ca and Mg),
micronutrients (Fe, Co, Ni and Mo) and some vitamins are necessary for the cell growth and metabolic
activities of fermentative microorganism [34–37].
Energies 2015, 8
4258
Figure 2. Hydrogen yields from the fermentation of mixed fruits.
2.3. VFAs Production
The knowledge of the distribution of VFA compositions formed during the fermentation process is
important, as it provides information about the metabolic pathways involved in the process. The
distribution of VFA compositions produced during the fermentation of the fruit wastes showed that
acetic and butyric acids were the dominant VFAs, while propionic, iso-butyric and iso-valeric were
produced in very low amounts (Figure 3). At the end of the first phase of the initial OLR, the acetic
acids produced from orange, banana, apple, grape, melon, Mix 1, Mix 2 and Mix 3 were 0.37, 0.62,
0.73, 0.75, 0.48, 0.56, 0.74 and 1.05 g/L, respectively. As the HRT decreased, the values of the acetic
acid increased, except for grape and melon, which decreased during the second phase. At the end of
the fermentation process, apple and Mix 3 produced significant amounts of acetic acids, which might
be connected to their high cumulative hydrogen yields, as acetic acid production is often associated
with hydrogen production [38,39].
Energies 2015, 8
4259
Figure 3. Volatile fatty acid (VFA) composition of the fermentation process.
2.4. Comparison of Hydrogen Yields and Acetic Acid Productions with Theoretical Values
In a biological in vivo system, the theoretical maximum hydrogen yield that could be obtained from
glucose at standard temperature and pressure is 4 mol H2/mol glucose when acetic acid is the only
soluble metabolite [40,41]. However, during dark fermentation, hydrogen production is often produced
along with reduced metabolites, including alcohols, lactic, propionic and valeric acids, which are
involved in the hydrogen consuming pathway, thereby leading to actual hydrogen yields being
significantly lower than the theoretical values [42–46]. On the other hand, high hydrogen yields are
usually linked with the moderate accumulation of a mixture of acetic and butyric acids though
accumulation of butyric acids, and its branched isomer could be an indication of process
instability [47]. In the present study, the hydrogen yields from the fermentation of all of the fruit
substrates, except grape and melon, increased with the increase in acetic acid production as the HRT
decreased from 8.6 down to five days (Table 2). This was expected as the concentration of volatile
fatty acids usually increased with the increase in substrate loading, which correlated with the decrease
in HRT from 8.6 down to five days [39]. Meanwhile, the hydrogen yields decreased with the increase
in acetic acid production, as the HRT was further decreased from five down to three days. In the
comparison of actual yields to theoretical yields, the relative yield was calculated as given in
Equation (4).
 Actual yield 

Re lativeYield 
  100
 Theoretical yield 
(4)
Energies 2015, 8
4260
2.4.1. Relative Yield of Hydrogen Production
The theoretical yield of hydrogen was based on Equation (2) with the assumption that at standard
temperature and pressure (STP), a maximum of four moles of hydrogen (H2) could be produced from
one mole of glucose when acetic acid was produced as the only reduced metabolite. The chemical
oxygen demand (COD) equivalent of four moles of hydrogen is 1.4 LH2/g COD of glucose at STP.
In other words, 1.4 L of hydrogen gas could be generated through complete anaerobic degradation of
1 g COD of glucose at STP. The total COD of the individual fruit waste was calculated, and the
equivalent theoretical yield was determined. The theoretical yield estimated and the actual yields
obtained experimentally were then used to calculate the relative yield using Equation (4).
Table 2. Comparison of hydrogen yields and acetic acid production with theoretical values.
Parameter
Average hydrogen yields
Average acetic acid production
Period (d)
1–15
16–30
31–47
1–15
16–30
HRT (d)
8.6
5.0
3.0
8.6
5.0
31–47
3.0
Fruit
AY
(mL/gVS)
RY
(%)
AY
(mL/gVS)
RY
(%)
AY
(mL/gVS)
RY
(%)
AY
(g/L)
RY
(%)
AY
(g/L)
RY
(%)
AY
(g/L)
RY
(%)
Orange
Banana
Apple
Grape
Melon
Mix 1
Mix 2
Mix 3
279
389
493
347
216
268
270
523
16.1
22.7
28.9
20.5
12.6
15.5
15.7
30.5
403
403
635
384
352
456
479
553
23.3
23.5
37.3
22.6
20.5
26.4
28.0
32.3
204
268
440
182
347
271
377
491
11.8
15.7
25.8
10.7
20.2
15.7
22.0
28.6
0.37
0.62
0.73
0.75
0.48
0.56
0.74
1.05
55
93
109
112
72
84
110
157
0.71
1.05
0.91
0.71
0.37
1.03
1.03
1.15
118
157
136
106
55
154
154
172
1.21
1.04
1.30
1.07
0.86
1.20
1.18
1.62
181
155
194
160
128
179
176
242
AY, actual yield; RY, relative yield.
Comparing the actual yields to theoretical yields, maximum relative yields of hydrogen production
obtained during the three phases were 37.3% and 32.3% from apple (single fruit fermentation) and Mix 3
(mixed fruit fermentation), respectively. For the whole fermentation period of 47 days, the maximum
relative yields of hydrogen production were 29.5% and 30.0% from apple and Mix 3, respectively.
On average, this indicates that only 30% of the chemical oxygen demand in the fruit mixture with Mix 3
could be converted into hydrogen (assuming the fruit wastes could be utilized as glucose). This low
value might be due to the effects of the toxicity of the fruit flavors and soluble metabolites.
2.4.2. Relative Yield of Volatile Fatty Acids
The theoretical yield of acetic acid production was also based on Equation 2 with the assumption
that at standard temperature and pressure (STP), a maximum of two moles of acetic acid (CH3COOH)
could be produced from one mole of glucose as the only reduced metabolite. The chemical oxygen
demand (COD) equivalent of two moles of acetic acid is 0.666 g-O2/L. In the case of acetic acid, the
theoretical yield was based on 1 g COD of the fruit waste. The estimated values of the theoretical acetic
acid production and the actual production of acetic acid were then used to calculate the relative yield.
Energies 2015, 8
4261
Throughout the whole fermentation period of 47 days, the maximum acetic acid concentration was
1.62 g/L from Mix 3. Although the maximum acetic acid concentration was less than the inhibiting
acetic concentration (greater than 2 g/L) [48], the decrease in hydrogen yield during the last
fermentation period could have been due to the organic loading exceeding its threshold limit. At a high
loading rate, acetic acid might accumulate and permeate the cell membrane of the hydrogen-producing
bacteria with subsequent disruption in the activities of the bacteria.
The relative acetic acid yields corresponding to the maximum hydrogen yields were 136% and 172%
for apple and 20% orange share, respectively. It appeared that the fermentative bacteria involved in the
fermentation of apple and 20% orange share were able to maintain their cell physiological balance in
the presence of extra acetic acids. The reason that some relative yields of acetic acid production were
higher than 100% could be due to the fact that the theoretical yield was based on 1 g COD of glucose
as feedstock, whereas in the real fruit feedstock, the nutrient composition included carbohydrate,
protein and lipids that could have been converted into acetic acids.
2.5. Significant Effects of Varying Hydraulic Retention Times, Fruit Mixing and Their Interaction on
Hydrogen Yield and Acetic Acid Production
The experiment was conducted to investigate the effects of hydraulic retention time (HRT) and fruit
mixing, as well as their interactions on hydrogen yield and acetic acid production. The results of the
two-way analysis of variance (ANOVA) are summarized in Tables 3 and 4. Based on the results
obtained, the hypothesis tests on the effects of hydraulic retention time (HRT) and fruit mixing, as well
as their interactions (HRT and fruit mixing interaction) on hydrogen yield indicated that the factors did
not have significant effects since their p-values, 0.061, 0.259 and 0.763 (Table 3) for HRT, fruit
mixing and HRT and fruit mixing interaction, respectively, were all greater than the chosen Į-level
(0.05). The results were further corroborated with the Turkey pairwise comparisons for the difference in
means (Table 4), which showed that the difference in means of hydrogen yield due to all of the factor
levels were not significantly different, since their adjusted p-values were all greater than the chosen
Į-level (0.05). Regarding the effects of the factors on acetic acid production, the hypothesis tests
(Table 3) showed that HRT and fruit mixing really had significant effects on the production of acetic
acids, since their p-values of 0.000 and 0.009 for HRT and fruit mixing, respectively, were lower than
the chosen Į-level (0.05). On the contrary, the effect of the interaction of HRT and fruit mixing on
acetic acid production was not statistically significant, since its p-value (0.830) was greater than the
chosen Į-level (0.05).
The Tukey pairwise comparisons for the difference in means due to HRT (Table 4) showed that the
adjusted p-values for the differences between the mean for HRT of three days and the means for HRT of
five days (0.024) and 8.6 days (0.000) were all lower than the chosen Į-level (0.05), which indicated that
these differences were significant. In the comparison of the difference in means for acetic acid production
due to fruit mixing, the adjusted p-value (0.009) was lower than the chosen Į-level (0.05), indicating
that the difference was statistically significant.
For the effect of the interaction of HRT and fruit mixing on acetic acid production, the adjusted
p-values for the difference between the mean for mixed fruit operated at HRT of three days and the
means for single fruits operated at HRT of five days (0.010) and 8.6 days (0.001) were lower than the
Energies 2015, 8
4262
chosen Į-level (0.05). Similarly, the adjusted p-values for single fruits (0.010) and mixed fruits
(0.037), when operated separately at HRT of three and 8.6 days, were lower than the chosen Į-level
(0.05). Furthermore, the adjusted p-value (0.044) for the difference between the mean for mixed fruit
operated at HRT of five days and the mean for single fruit operated at HRT of 8.6 days was lower than
the chosen Į-level (0.05), which indicated that the difference was statistically significant.
Meanwhile, as a consequence of the insignificant effect of fruit mixing on hydrogen yield, the fruit
feedstock was further considered as individual substrates for analysis of variance (Table 5). Based on
the results obtained, the hypothesis tests showed that both HRT and individual substrates had significant
effects on hydrogen yields, since their p-values (0.001) were lower than the chosen Į-level (0.05).
Similarly, the effects of HRT and individual substrates on acetic acid production were statistically
significant, since their p-values, 0.000 and 0.001 for HRT and individual substrate, respectively, were
lower than the chosen Į-level (0.05).
Table 3. Summary of two-way analysis of variance (ANOVA) for the effects of hydraulic
retention time (HRT) and fruit mixing (Mix) on hydrogen yield and acetic acid production.
Response
Factor
Factor Type
Factor Levels
Factor Values
dF
Adj SS
Adj MS
F-Value
HRT
Fixed
3
3.0; 5.0; 8.6
2
3,8951
3.28
3.28
0.061
Mix
Fixed
2
N; Y
1
16,187
1.36
1.36
0.259
2
3,266
0.27
0.27
0.763
Error
18
11,890
Total
23
Hydrogen
HRT and mix
yield
interaction
p-Value
HRT
Fixed
3
3.0; 5.0: 8.6
2
1.05371
0.526854
12.82
0.000
Mix
Fixed
2
N, Y
1
0.35219
0.352188
8.57
0.009
2
0.01551
0.007754
0.19
0.830
Error
18
0.73985
0.041103
Total
23
Acetic acid
HRT and mix
production
interaction
Table 4. Tukey pairwise comparisons: response = H2 yield, acetic acid production;
factor = HRT, mix and mix and HRT interaction.
Response
H2 yield
Factor
HRT
Mix
Mix and HRT
interaction
Difference of
Difference
SE of
Simultaneous
factor levels
of means
difference
95% CI
5.0–3.0
131.8
56.3
(í12.0; 275.5)
2.34
8.6–3.0
15.3
56.3
(í128.4; 159.0)
0.27
0.960
8.6–5.0
í116.5
56.3
(í260.2; 27.3)
í2.07
0.125
YES–NO
53.6
46.0
(í42.9; 150.2)
1.17
0.259
(NO 5.0)–(NO 3.0)
147.2
69.0
(í71.8; 366.2)
2.13
0.314
(NO 8.6)–(NO 3.0)
56.6
69.0
(í162.4; 275.6)
0.82
0.960
t-value
Adjusted
p-value
0.076
Energies 2015, 8
4263
Table 4. Cont.
Response
Factor
HRT
Acetic acid
Mix
Mix and HRT
interaction
Difference of
Difference
SE of
Simultaneous
factor levels
of means
difference
95% CI
(YES 3.0)–(NO 3.0)
91.5
79.6
(í161.4; 344.3)
1.15
0.855
(YES 5.0)–(NO 3.0)
207.8
79.6
(í45.0; 460.6)
2.61
0.145
(YES 8.6)–(NO 3.0)
65.5
79.6
(í187.4; 318.3)
0.82
0.960
(NO 8.6)–(NO 5.0)
í90.6
69.0
(í309.6; 128.4)
í1.31
0.774
(YES 3.0)–(NO 5.0)
í55.7
79.6
(í308.6; 197.1)
í0.70
0.980
(YES 5.0)–(NO 5.0)
60.6
79.6
(í192.2; 313.4)
0.76
0.971
(YES 8.6)–(NO 5.0)
í81.7
79.6
(í334.6; 171.1)
í1.03
0.903
(YES 3.0)–(NO 8.6)
34.9
79.6
(í218.0; 287.7)
0.44
0.998
(YES 5.0)–(NO 8.6)
151.2
79.6
(í101.6; 404.0)
1.90
0.434
(YES 8.6)–(NO 8.6)
8.9
79.6
(í244.0; 261.7)
0.11
1.000
(YES 5.0)–(YES 3.0)
116.3
89.0
(í166.3; 399.0)
1.31
0.778
(YES 8.6)–(YES 3.0)
í26.0
89.0
(í308.7; 256.7)
í0.29
1.000
(YES 8.6)–(YES 5.0)
í142.3
89.0
(í425.0; 140.3)
í1.60
0.610
t-value
Adjusted
p-value
5.0–3.0
í0.305
0.105
(í0.572; í0.037)
í2.91
0.024
8.6–3.0
í0.528
0.105
(í0.795; í0.261)
í5.04
0.000
8.6–5.0
í0.223
0.105
(í0.491; 0.044)
í2.13
0.111
YES–NO
0.2502
0.0855
(0.0706; 0.4298)
2.93
0.124
(NO 5.0)–(NO 3.0)
í0.346
0.128
(í0.753; 0.061)
í2.70
0.124
0.010
(NO 8.6)–(NO 3.0)
í0.506
0.128
(í0.913; í0.099)
í3.95
(YES 3.0)–(NO 3.0)
0.237
0.148
(í0.233; 0.707)
1.60
0.607
(YES 5.0)–(NO 3.0)
í0.026
0.148
(í0.496; 0.444)
í0.18
1.000
(YES 8.6)–(NO 3.0)
í0.313
0.148
(í0.783; 0.157)
í2.11
0.325
(NO 8.6)–(NO 5.0)
í0.160
0.128
(í0.567; 0.247)
í1.25
0.808
(YES 3.0)–(NO 5.0)
0.583
0.148
(0.113; 1.053)
3.94
0.010
(YES 5.0)–(NO 5.0)
0.320
0.148
(í0.150; 0.790)
2.16
0.302
(YES 8.6)–(NO 5.0)
0.033
0.148
(í0.437; 0.503)
0.23
1.000
(YES 3.0)–(NO 8.6)
0.743
0.148
(0.273; 1.213)
5.02
0.001
(YES 5.0)–(NO 8.6)
0.480
0.148
(0.010; 0.950)
3.24
0.044
(YES 8.6)–(NO 8.6)
0.193
0.148
(í0.227; 0.663)
1.31
0.778
(YES 5.0)–(YES 3.0)
í0.263
0.166
(í0.789; 0.262)
í1.59
0.614
(YES 8.6)–(YES 3.0)
í0.550
0.166
(í1.076; í0.024)
í3.32
0.037
(YES 8.6)–(YES 5.0)
í0.287
0.166
(í0.812; 0.239)
í1.73
0.530
Energies 2015, 8
4264
Table 5. Summary of two-way analysis of variance (ANOVA) for the effects of hydraulic
retention time (HRT) and substrate.
Response
Factor
Factor Type
Factor Levels
Factor Values
dF
Adj SS
Adj MS
F-Value
p-Value
HRT
Fixed
3
3.0; 5.0; 8.6
2
83,069
41,534
11.52
0.001
7
186,264
26,609
7.38
0.001
14
50,472
3,605
23
319,805
2
1.1074
0.55372
30.39
0.000
7
0.8524
0.12178
6.68
0.001
Error
14
0.2551
0.01822
Total
23
Apple; Banana; Grape; Melon;
Hydrogen
Substrate
Fixed
8
Mix1; Mix 2; Mix 3; Orange
yield
Error
Total
HRT
Fixed
3
Acetic
Apple; Banana; Grape; Melon;
Mix
acid
production
3.0; 5.0; 8.6
Fixed
8
Mix1; Mix 2; Mix 3; Orange
The Tukey pairwise comparisons for the difference in means of hydrogen yield due to HRT (Table 6)
showed that the adjusted p-values for the differences between the mean for HRT of five days and the
means for HRT of three days (0.001) and 8.6 days (0.007) were lower than the chosen Į-level (0.05),
which indicated that these differences were significant. For the effects of individual substrates on
hydrogen yield, the Turkey pairwise comparisons showed that the adjusted p-values for the differences
between the mean for apple and the means for grape (0.010), melon (0.010), Mix 1 (0.026) and orange
(0.007) were all lower than the chosen Į-level (0.05). In the same vein, the adjusted p-values for the
difference between the mean for Mix 3 and the means for grape (0.010), melon (0.010), Mix 1 (0.026)
and orange (0.007) were lower than the chosen Į-level (0.05), which indicated that the mean differences
were significant. Meanwhile, the Tukey comparison for the difference in means of acetic acid production
due to HRT showed that the adjusted p-values for the differences between the mean for HRT of three
days and the means for HRT of five days (0.001) and 8.6 days (0.0022) were lower than the chosen
Į-level (0.05). Furthermore, the adjusted p-value (0.021) for the difference between the mean for HRT
of five days and 8.6 days was lower than the chosen Į-level (0.05), which indicated that the difference
was significant. In the case of the effects of individual substrates on acetic acid production, the
adjusted p-values for the differences between the mean for Mix 3 and the means for grape (0.026),
melon (0.000) and orange (0.007) were below the chosen Į-level (0.05). Similarly, the adjusted
p-values for the difference between the mean for melon and the means for apple (0.036) and Mix 2
(0.034) were below the chosen Į-level (0.05), which showed that the mean differences
were significant.
Energies 2015, 8
4265
Table 6. Tukey pairwise comparisons: Response = H2 yield, acetic acid production;
factor = HRT, substrate.
Response
H2 yield
Factor
HRT
Substrate
Acetic acid
Difference of
Difference
SE of
Simultaneous
Factor Levels
of Means
Difference
95% CI
5.0–3.0
135.6
30.0
(57.1; 214.2)
4.52
0.001
8.6–3.0
25.6
30.0
(í52.9; 104.2)
0.85
0.677
8.6–5.0
í110.0
30.0
(í188.5; í31.5)
í3.66
0.007
t-Value
Adjusted
p-Value
Banana-Apple
í169.3
49.0
(í342.3; 3.6)
í3.45
0.057
Grape-apple
í218.3
49.0
(í391.3; í45.4)
í4.45
0.010
Melon-Apple
í217.7
49.0
(í390.6; í44.7)
í4.44
0.010
Mix 1-Apple
í191.0
49.0
(í364.0; í18.0)
í3.90
0.026
Mix 2-Apple
í147.3
49.0
(í320.3; 25.6)
í3.01
0.123
Mix 3-Apple
í0.3
49.0
(í173.3; 172.6)
í0.01
1.000
Orange-Apple
í227.3
49.0
(í400.3; í54.4)
í4.64
0.007
Grape-Banana
í49.0
49.0
(í222.0; 124.6)
í1.00
0.967
Melon-Banana
í48.3
49.0
(í221.3; 124.6)
í0.99
0.969
Mix 1-Banana
í21.7
49.0
(í194.6; 151.3)
í0.44
1.000
Mix 2-Banana
22.0
49.0
(í151.0; 195.0)
0.45
1.000
Mix 3-Banana
169.0
49.0
(í4.0; 342.0)
3.45
0.058
Orange-Banana
í58.0
49.0
(í231.0; 115.0)
í1.18
0.924
1.000
Melon-Grape
0.7
49.0
(í172.3; 173.6)
0.01
Mix 1-Grape
27.3
49.0
(í145.6; 200.3)
0.56
0.999
Mix 2-Grape
71.0
49.0
(í102.0; 244.0)
1.45
0.821
Mix 3-Grape
218.0
49.0
(45.0; 391.0)
4.45
0.010
Orangeí Grape
í9.0
49.0
(í182.0; 164.0)
í0.18
1.000
Mix 1-Melon
26.7
49.0
(í146.3; 199.6)
0.54
0.999
Mix 2-Melon
70.3
49.0
(í102.6; 243.3)
1.43
0.827
Mix 3-Melon
217.3
49.0
(44.4; 390.3)
4.43
0.010
Orange-Melon
í9.7
49.0
(í182.6; 163.3)
í0.20
1.000
Mix 2-Mix 1
43.7
49.0
(í129.3; 216.6)
0.89
0.982
Mix 3-Mix 1
190.7
49.0
(17.7; 363.6)
3.89
0.026
Orange-Mix 1
í36.3
49.0
(í209.3; 136.6)
í0.74
0.994
Mix 3-Mix 2
147.0
49.0
(í26.0; 320.0)
3.00
0.124
Orange-Mix 2
í80.0
49.0
(í253.0; 93.0)
í1.63
0.726
Orange-Mix 3
í227.0
49.0
(í400.0; í54.0)
í4.63
0.007
5.0–3.0
í0.3150
0.0675
(í0.4916; í0.1384)
í4.67
0.001
HRT
8.6–3.0
í0.5225
0.0675
(í0.6991; í0.3459)
í7.74
0.002
8.6–5.0
í0.2075
0.0675
(í0.3841; í0.0309)
í3.07
0.021
Substrate
Banana-Apple
í0.077
0.110
(í0.466; 0.312)
í0.70
0.996
Grape-apple
í0.137
0.110
(í0.526; 0.252)
í1.24
0.906
Melon-Apple
í0.410
0.110
(í0.799; í0.021)
í3.72
0.036
Mix 1-Apple
í0.050
0.110
(í0.439; 0.339)
í0.45
1.000
Mix 2-Apple
0.003
0.110
(í0.386; 0.392)
0.03
1.000
Mix 3-Apple
0.293
0.110
(í0.096; 0.682)
2.66
0.213
Orange-Apple
í0.217
0.110
(í0.606; 0.172)
í1.97
0.534
Grape-Banana
í0.060
0.110
(í0.449; 0.329)
í0.54
0.999
Energies 2015, 8
4266
Table 6. Cont.
Response
Factor
Difference of
Difference
SE of
Simultaneous
Factor Levels
of Means
Difference
95% CI
Melon-Banana
í0.333
0.110
(í0.722; 0.056)
í3.02
Mix 1-Banana
0.027
0.110
(í0.362; 0.416)
0.24
1.000
Mix 2-Banana
0.080
0.110
(í0.309; 0.469)
0.73
0.995
t-Value
Adjusted
p-Value
0.119
Mix 3-Banana
0.370
0.110
(í0.019; 0.759)
3.36
0.068
Orange-Banana
í0.140
0.110
(í0.529; 0.249)
í1.27
0.896
Melon-Grape
í0.273
0.110
(í0.662; 0.116)
í2.48
0.278
Mix 1-Grape
0.087
0.110
(í0.302; 0.476)
0.79
0.991
Mix 2-Grape
0.140
0.110
(í0.249; 0.529)
1.27
0.896
Mix 3-Grape
0.430
0.110
(0.041; 0.819)
3.90
0.026
Orangeí Grape
í0.080
0.110
(í0.469; 0.309)
í0.73
0.995
Mix 1-Melon
0.360
0.110
(í0.029; 0.749)
3.27
0.079
Mix 2-Melon
0.413
0.110
(0.024; 0.802)
3.75
0.034
Mix 3-Melon
0.703
0.110
(0.314; 1.092)
6.38
0.000
Orange-Melon
0.193
0.110
(í0.196; 0.582)
1.75
0.657
Mix 2-Mix 1
0.053
0.110
(í0.336; 0.422)
0.48
1.000
Mix 3-Mix 1
0.343
0.110
(í0.046; 0.732)
3.12
0.102
Orange-Mix 1
í0.167
0.110
(í0.556; 0.222)
í1.51
0.790
Mix 3-Mix 2
0.290
0.110
(í0.099; 0.679)
2.63
0.223
Orange-Mix 2
í0.220
0.110
(í0.609; 0.169)
í2.00
0.516
Orange-Mix 3
í0.510
0.110
(í0.899; í0.121)
í4.63
0.007
3. Experimental Section
3.1. Experimental Materials (Feedstock, Seed Sludge and Nutrient)
Whole fruit wastes, including apple (Malus sp.), banana (Musa sp.), grape (Vitis sp.), melon
(Cucumis sp.) and orange (Citrus sp.), obtained from a local shop (Borås, Sweden), were used as
feedstock for the anaerobic digestion process. The fruits as a whole (rind and inner part of the fruits)
were ground separately in a kitchen blender (Waring Commercial, Torrington, CT, USA) before they were
stored in a cold room (5 °C) to reduce their deterioration prior to the fermentation process.
The hydrogen-producing microorganism (HPM) was prepared from sludge obtained from a
thermophilic (55 °C) biogas plant (Borås Energi & Miljö AB, Borås, Sweden) that utilized the organic
fraction of municipal solid waste as feedstock for biogas production. The HPM was enriched by
adjusting the pH to 5.0 ± 0.1 coupled with heat treatment at 105 °C for 1 h. The nutrients used for the
growth of HPM were a mixture of macro- and micronutrients, including (g/L): FeCl2. 4H2O, 11.401;
KH2PO4, 4.681; NH4Cl, 0.814; NaHCO3, 3.000; MgSO4.7H2O, 0.320; NiSO4.6H2O, 0.032; CaCl2,
0.050; Na2B4O7.10H2O, 0.007; (NH4)6Mo7O24.4H2O, 0.014: ZnCl2, 0.023; CoCl6.H2O, 0.021;
CuCl2.2H2O, 0.010.
Energies 2015, 8
4267
3.2. Experimental Setup and Procedures
The fermentative production of hydrogen was carried out using eight continuous stirred tank
bioreactors (CSTR) in order to evaluate the performance, in terms of hydrogen yields and VFA
production, of the fermentative hydrogen production process from the whole fruit wastes. Each CSTR
reactor (active volume of 3 L and headspace of 1 L) contained fermentation medium at 55 °C and a pH
of 5.0 (Figure 4). Eight different fruit substrates were used, including single fruits (banana, apple,
grape, melon, orange) and mixed fruits, which differed in the amount of orange in the mixtures. The
weight distribution of the first, second and third fruit mixtures were 70%, 50% and 20% orange share,
respectively, while the residual weight was shared equally by banana, apple, grape, melon and orange.
A thermostatic water bath (GD 100, Grant instruments Ltd., Cambridgeshire, UK) was employed to
provide the heating energy needed to maintain the required temperatures in the CSTRs. Prior to the start
of the fermentation process, the fruit slurries were left to attain room temperature (22 °C) before they
were fed into the bioreactors. A mixture of HPM, feedstock, distilled water, macro- and micro-nutrients
in solution at a volumetric ratio of 8:2:6:4:1, respectively, was added to the CSTRs and left for three
days without any daily feedstock feeding in order to adapt or acclimatize the inoculum to the
fermentation environment. The fermentation process was started, after the third day, with the feeding of
the fruit feedstock mixed with distilled water, buffer solution (1 M NaHCO3), macro- and
micro-nutrients at a ratio of 10:60:3:20:7, respectively, into the CSTR. The whole experiment was run
for 47 days with an increase in the organic loading rate (OLR).
Volumetric gas
measurement
Influent feed (fruit waste,
nutrient, distilled water)
Manual effluent
withdrawal
Data acquisition
system
Inoculum
Water-jacketed continuous
stirred tank reactor (CSTR)
Sedimentation tank
Figure 4. Schematic diagram of the fermentative hydrogen production from single and mixed fruits.
3.3. Analytical Procedures
The characteristics of the feedstock for the two-stage fermentation process, including total solid
(TS), volatile solid (VS) and pH (Table 7), were determined according to standard methods [49]. The
main nutrient compositions of the fruit wastes and their mixtures are presented in Table 8 according to
the nutrient database of the U.S. Department of Agriculture [50]. The gas produced was measured
Energies 2015, 8
4268
using the Automatic Methane Potential Testing System (AMPTS, Bioprocess Control AB, Lund,
Sweden), which is based on the principle of water displacement and buoyancy, while the hydrogen
composition of the gas produced was sampled with the aid of a 0.25-ȝL pressure-tight gas syringe
(VICI, Baton Rouge, LA, USA) and analyzed using a gas chromatograph (GC, Perkin-Elmer, 710
Bridgeport Avenue, Shelton, CT, USA) equipped with a packed column (Perkin-Elmer, 6' × 1.8" OD,
80/100, Mesh, 710 Bridgeport Avenue, Shelton, CT, USA) and a thermal conductivity detector (TCD,
PerkinElmer, 710 Bridgeport Avenue, Shelton, CT, USA) set at 200 °C. The temperatures of the oven
and injector were set at 75 °C and 150 °C, respectively, while nitrogen gas at a flow rate, temperature
and pressure of 20 mL/min, 60 °C and 1 bar, respectively, was used as the carrier gas. The volatile
fatty acids (VFA) in the effluent samples were measured using a high-performance liquid
chromatograph (HPLC, Waters 2695, Waters Corporation, Milford, MA, USA) equipped with an RI
detector (Waters 2414, Waters Corporation, Milford, MA, USA) and a biohydrogen-ion exchange
column (Aminex HPX-87H, Bio-Rad, Hercules, CA, USA) operating at 60 °C and 0.6 mL/min and
with 5 mM sulfuric acid as the effluent.
Table 7. Measured characteristics of fruit wastes. TS, total solid.
Parameters
TS (%)
VS (%)
pH
Apple
11.72 ± 0.02
11.53 ± 0.35
3.90 ± 0.01
Banana
16.36 ± 0.24
15.5 ± 0.05
5.06 ± 0.02
Grape
19.32 ± 0.24
18.68 ± 0.06
3.67 ± 0.02
Melon
8.93 ± 0.26
8.39 ± 0.16
4.88 ± 0.01
Orange
16.85 ± 0.30
16.26 ± 0.20
4.04 ± 0.02
Table 8. Nutrient composition in 100 g of fruit.
Fruit
Apple
Melon
Banana
Orange
Grape
Mix 1 *
Mix 2 *
Mix 3 *
Carbohydrate
g
%
14.06
97
9.09
93
22.84
94
15.50
90
18.10
95
15.71
92
15.77
95
15.92
94
Protein
g
%
0.27
2
0.54
6
1.09
5
1.30
8
0.72
4
1.10
6
0.59
3
0.78
5
Lipid
g
%
0.20 1
0.14 1
0.33 1
0.30 2
0.16 1
0.28 2
0.26 2
0.23 1
C:N Ratio *
52:1
17:1
21:1
12:1
25:1
14:1
27:1
20:1
* Estimated value.
4. Conclusions
The present study assessed the effect of hydraulic retention times and fruit mixing on biohydrogen
production from fruit wastes in continuous stirred tank bioreactors. The results of the two-way analysis
of variance indicated that there was no statistically-significant effect of the interaction of hydraulic
retention time and fruit mixing on hydrogen yields and acetic acid production. However, the results
established that significant improvement in hydrogen yields could be obtained when apple and Mix 3
were used as individual substrates. The results also indicated that operating the fermentative hydrogen
production at a hydraulic retention time of 5 days could greatly increase hydrogen yield, as it reduced
Energies 2015, 8
4269
the amount of acetic acid accumulated during the fermentation process. It could therefore be inferred
that fermentation of apple or Mix 3 at hydraulic retention time of 5 days could be used to reduce the
effect of bacterial inhibition due to flavor compounds in the fruits and thereby enhance hydrogen
production from the fruit wastes.
Acknowledgments
The authors wish to express their gratitude to the Swedish Research Council (Sweden) and Lagos
State University (Nigeria) for providing financial support during the research work. We are also
grateful for the assistance given by Magnus Lundin on statistical analysis of the data and to Khamdan
Cahyari for his support during the study.
Acronyms
AY
COD
HPM
MS
Mix 1
Mix 3
PY
SV
TVFA
VS
Actual yield
Chemical oxygen demand
Hydrogen producing microorganism
Mean square
70% orange mixed fruit
20% orange mixed fruit
Percent yield
Source of variation
Total volatile fatty acids
Volatile solid
CIs
dF
HRT
Mix
Mix 2
OLR
SS
TS
VFA
Confidence intervals
degree of freedom
Hydraulic retention time
Mixing
50% orange mixed fruit
Organic loading rate
Sum of squares
Total solid
Volatile fatty acids
Conflicts of Interest
The authors declare no conflicts of interest
References
1.
2.
3.
4.
5.
6.
Benemann, J. Hydrogen biotechnology: Progress and prospects. Nat. Biotechnol. 1996, 14,
1101–1103.
Bockris, J.O.M. The economics of hydrogen as a fuel. Int. J. Hydrog. Energy 1981, 6, 223–241.
Mormirlan, M.; Veziroglu, T.N. Current status of hydrogen energy. Renew. Sustain. Energy Rev.
2002, 6, 141–179.
Ros, M.; Franke-Whittle, I.H.; Morales, A.B.; Insam, H.; Ayuso, M.; Pascual, J.A. Archael
community dynamics and abiotic characteristics in a mesophilic anaerobic co-digestion process
treating fruit and vegetable processing waste sludge with chopped fresh artichoke waste.
Bioresour. Technol. 2013, 136, 1–7.
Ueno, Y.; Haruta, S.; Ishii, M.; Igarashi, Y. Microbial community in anaerobic hydrogen-producing
microflora enriched from sludge compost. Appl. Microbiol. Biotechnol. 2001, 57, 555–562.
Kapdan, I.K.; Kargi, F. Biohydrogen production from waste materialk. Enzyme Microb. Technol.
2006, 38, 569–582.
Energies 2015, 8
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
23.
24.
4270
Levin, D.B.; Chahine, R. Challenges for renewable hydrogen production processes. Int. J.
Hydrog. Energy 2008, 33, 279–286.
Westermann, P.; Jørgensen, B.; Lange, L.; Ahring, B.K.; Christensen, C.H. Maximizing renewable
hydrogen production from biomass in a bio/catalytic refinery. Int. J. Hydrog. Energy 2007, 32,
4135–4141.
Nandi, R.; Sengupta, S. Microbial production of hydrogen: An Overview. Crit. Rev. Microbiol.
1998, 24, 61–84.
Nursten, H.E.; Williams, A.A. Fruit aromas: A survey of compounds identified. Chem. Ind. 1967,
486–497.
Winniczuk, P.P.; Parish, M.E. Minimum inhibitory concentrations of antimicrobials against
micro-organisms related to citrus juice. Food Microbiol. 1997, 14, 373–381.
Jordan, M.J.; Tandon, K.; Shaw, P.E.; Goodner, K.I. Aromatic profile aqueous banana essence
and banana fruit by gas-chromatography-mass spectrometry (GC-MS) and gas-chromatographyolfactometry. J. Agric. Food Chem. 2001, 49, 4813–4817.
Nogueira, J.M.F.; Fernandes, P.J.P.; Nascimento, A.M.D.C. Composition of volatiles of banana
cultivars from Madeira island. Phytochem. Anal. 2003, 14, 82–90.
Rizzolo, J.; Polesello, A.; Teleky-Vamossy, G. CGC/Sensory analysis of volatile compounds
developed from ripening apple fruit. J. High Resolut. Chrom. 1989, 12, 824–827.
Holland, D.; Larkov, O.; Bar-Yaákov, I.; Bar, E.; Zax, A.; Brandeis, E. Developmental and varietal
differences in volatile ester formation and acetyl-CoA: Alcohol acetyl transferase activities in apple
(Malus domestica Borkh.) fruit. J. Agric. Food Chem. 2005, 53, 7198–7203.
Tsanova-Savova, S.; Fany, R.; Maria, G. (+)- Catechin and (-) Epicatechin in Bulgarian Fruits.
J. Food Comp. Anal. 2005, 18, 691–698.
Schieber, A.; Petra, K.; Reinhold, C. Determination of phenolic acids and flavonoids of apple and
pear by high-performance liquid chromatography. J. Chromatogr. 2000, 910, 265–273.
Dieguez, S.C.; Lois, L.C.; Gomez, E.F.; de Ia Pena, M.L.G. Aromatic composition of the Vitis
vinifera grape Albariño. Lebensm. Wiss. Und. Technol. 2003, 36, 585–590.
Aubert, C.; Baumann, S.; Arguel, H. Optimisation of the analysis of flavour volatile compounds
by liquid-liquid microextraction (LLME). Apllication to the aroma analysis of melons, peaches,
grapes, strawberries, and tomatoes. J. Agric. Food Chem. 2005, 53, 8881–8895.
Perry, P.L.; Wang, Y.; Lin, J.M. Analysis of honeydew melon (Cucumis melo var. Inodorus)
flavor and GC/MS identification of (E,Z)-2,6-nonadienyl acetate. Flav. Frag. J. 2009, 24, 341–347.
Portnoy, V.; Benyamini, Y.; Bar, E. The molecular and biochemical basis for varietal variation in
sesquiterpene content in melon (Cucumis melo L.) rinds. Plant Mol. Biol. 2008, 66, 647–661.
Aubert, C.; Pitrat, M. Volatile compounds in the skin and pulp of Queen Anne’s pocket melon.
J. Agric. Food Chem. 2006, 54, 8177–8182.
Hui, Y.H. Handbook of Fruit and Vegetable Flavours; John Wiley & Sons: Hoboken, NJ, USA,
2010.
Utama, I.; Made, S.; Wills, R.B.H.; Ben-yehoshua, S.; Kuek, C. In vitro efficacy of plant volatiles
for inhibiting the growth of fruit and vegetable decay microorganisms J. Agric. Food Chem. 2002,
50, 6371–6377.
Energies 2015, 8
4271
25. Grohmann, K.; Baldwin, E.; Buslig, B. Production of ethanol from enzymatically hydrolyzed
orange peel by the yeast Saccharomyces cerevisiae. Appl. Biochem. Biotechnol. 1994, 45–46,
315–327.
26. Mizuki, E.; Akao, T.; Saruwatari, T. Inhibitory effect of citrus Unshu peel om anaerobic digestion.
Biol. Wastes 1990, 33, 161–168.
27. Martin, M.A.; Siles, J.A.; China, A.F.; Martin, A. Biomethanization of orange peel waste.
Bioresour. Technol. 2010, 101, 8993–8999.
28. Youngsukkasem, S.; Akinbomi, J.; Rakshit, S.; Taherzadeh, M.J. Biogas production by encased
bacteria in synthetic membranes: Protective effects in toxic media and high loading rates.
Environ. Technol. 2013, 34, 2077–2084.
29. Wikandari, R.; Youngsukkasem, S.; Millati, R.; Taherzadeh, M.J. Performance of semi-continuous
membrane bioreactors in biogas production from toxic feedstock containing D-limonene.
Bioresour. Technol. 2014, 170, 350–355.
30. Pan, J.; Zhang, R.; el-Mashad, H.M.; Sun, H.; Yimg, Y. Effect of food to microorganism ratio on
biohydrogen production from food waste via anaerobic fermentation. Int. J. Hydrog. Energy 2008,
33, 6968–6975.
31. Liu, D.; Zeng, R.J.; Angelidaki, I. Hydrogen and methane production from household solid waste
in the two-stage fermentation process. Water Res. 2006, 40, 2230–2236.
32. Kim, S.H.; Han, S.K.; Shin, H.S. Optimization of continuous hydrogen fermentation of food waste
as a function of solids retention time independent of hydraulic retention time. Process Biochem.
2008, 43, 213–218.
33. Chu, C.F.; Xu, K.Q.; Li, Y.Y.; Inamori, Y. Hydrogen and methane potential based on the nature
of food waste materials in a two-stage thermophilic fermentation process. Int. J. Hydrog. Energy
2012, 37, 10611–10618.
34. Lin, C.; Lay, C. A nutrient formulation for fermentative hydrogen production using anaerobic
sewage sludge microflora. Int. J. Hydrog. Energy 2005, 30, 285–292.
35. Demirel, B.; Scherer, P. Trace element requirements of agricultural biogas digesters during
biological conversion of renewable biomass to methane. Biomass Bioenergy 2011, 35, 992–998.
36. Demirel, B.; Scherer, P. Production of methane from sugar beet sludge without manure addition
by a single stage anaerobic digestion process. Biomass Bioenergy 2008, 32, 203–209.
37. Pobeheim, H.; Munk, B.; Johansson, J.; Guebitz, G.M. Influence of trace elements on methane
formation from a synthetic model substrate for maize silage. Bioresour. Technol. 2010, 101, 836–839.
38. Thauer, R.K.; Jungermann, K.; Decker, K. Energy conservation in chemotrophic anaerobic
bacteria. Microbiol.Mol. Biol. Rev. 1977, 41, 100–180.
39. Dohanyos, M.; Kosova, B.; Zabranska, J.; Grau, P. Production and utilization of volatile fatty
acids in various types of anaerobic reactors. Water Sci. Technol. 1985, 17, 191–205.
40. Vardar, S.G.; Maeda, T.; Wood, T.K. Metabolically engineered bacteria for producing hydrogen
via fermentation. Microb. Biotechnol. 2008, 1, 107–125.
41. Thauer, R.K. Limitation of microbial H2-formation via fermentation In Microbial Energy
Conversion; Schlegel, H.G., Barnea, J., Eds.; Pergamon Press: New York, NY, USA, 1977.
Energies 2015, 8
4272
42. Hawkes, F.R.; Hussy, I.; Kyazze, G.; Dinsdale, R.; Hawkes, D.L. Continuous dark fermentative
hydrogen production by mesophilic microflora: Principles and progress. Int. J. Hydrog. Energy
2007, 32, 172–184.
43. Hawkes, F.R.; Dinsdale, R.; Hawkes, D.L.; Hussy, I. Sustainable fermentative hydrogen
production: Challenges for process optimisation. Int. J. Hydrog. Energy 2002, 27, 1339–1347.
44. Ren, N.; Li, J.; Li, B.; Wang, Y.; Liu, S. Biohydrogen production from molasses by anaerobic
fermentation with a pilot-scale bioreactor system. Int. J. Hydrog. Energy 2006, 31, 2147–2157.
45. Buyukkamaci, N.; Filibeli, A. Volatile fatty acid formation in an anaerobic hybrid reactor.
Process Biochem. 2004, 39, 1491–1494.
46. Hallenbeck, P.C.; Benemann, J.R. Biological hydrogen production: Fundamentals and limiting
processes. Int. J. Hydrog. Energy 2002, 27, 1185–1193.
47. Levin, D.B.; Pitt, L.; Love, M. Biohydrogen production: Prospects and limitations to practical
application. Int. J. Hydrog. Energy 2004, 29, 173–185.
48. Polprasert, C. Organic Waste Recycling: Technology and Management; John Wiley & Sons:
Chichester, UK, 1996.
49. American Public Health Association Inc. (APHA). Standard Methods for the Examination of Water
and Wastewater, 20th ed.; APHA: Washington, DC, USA, 1998.
50. USDA National Nutrient Database for Standard Reference, Release 27; US Department of
Agriculture, Agricultural Research Service, Nutrient Data Laboratory, Beltsville, MD, USA,
August 2014.
© 2015 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article
distributed under the terms and conditions of the Creative Commons Attribution license
(http://creativecommons.org/licenses/by/4.0/).
Paper III
Bioresource Technology 196 (2015) 290–300
Contents lists available at ScienceDirect
Bioresource Technology
journal homepage: www.elsevier.com/locate/biortech
Development of a submerged anaerobic membrane bioreactor for
concurrent extraction of volatile fatty acids and biohydrogen production
Zaineb Trad a,b,c, Julius Akimbomi d, Christophe Vial b,c,⇑, Christian Larroche b,c,
Mohammad J. Taherzadeh d, Jean-Pierre Fontaine b,c
a
Université Clermont Auvergne, Université Blaise Pascal, LABEX IMobS3, BP 10448, F-63000, F-63171 Clermont-Ferrand, France
Université Clermont Auvergne, Université Blaise Pascal, Institut Pascal, BP 20206, F-63174 Aubière cedex, France
c
CNRS, UMR 6602, IP, F-63178 Aubière, France
d
Swedish Centre for Resource Recovery, University of Borås, S-50190, Sweden
b
h i g h l i g h t s
g r a p h i c a l a b s t r a c t
An externally-submerged anaerobic
membrane reactor was used to
produce BioH2.
Mixing, transmembrane pressure
(TMP) and fouling were investigated.
TMP was low (10 kPa) and fouling
was reversible, mainly due to cake
layer formation.
Gas scouring and backwashing with
the substrate were used as a cleaning
procedure.
Biohydrogen production was shown
to restart after removing VFA in the
permeate.
a r t i c l e
i n f o
Article history:
Received 3 June 2015
Received in revised form 24 July 2015
Accepted 25 July 2015
Available online 30 July 2015
Keywords:
Anaerobic membrane bioreactor
Fouling
Submerged membrane
Volatile fatty acids
a b s t r a c t
The aim of this work was to study an externally-submerged membrane bioreactor for the cyclic extraction of volatile fatty acids (VFAs) during anaerobic fermentation, combining the advantages of submerged
and external technologies for enhancing biohydrogen (BioH2) production from agrowaste. Mixing and
transmembrane pressure (TMP) across a hollow fiber membrane placed in a recirculation loop coupled
to a stirred tank were investigated, so that the loop did not significantly modify the hydrodynamic
properties in the tank. The fouling mechanism, due to cake layer formation, was reversible. A cleaning
procedure based on gas scouring and backwashing with the substrate was defined. Low TMP, 104 Pa,
was required to achieve a 3 L h1 m2 critical flux. During fermentation, BioH2 production was shown
to restart after removing VFAs with the permeate, so as to enhance simultaneously BioH2 production
and the recovery of VFAs as platform molecules.
Ó 2015 Elsevier Ltd. All rights reserved.
1. Introduction
⇑ Corresponding author at: Université Clermont Auvergne, Université Blaise
Pascal, Institut Pascal, BP 20206, F-63174 Aubière cedex, France. Tel.: +33
(0)473405266; fax: +33 (0)473407829.
E-mail address: [email protected] (C. Vial).
http://dx.doi.org/10.1016/j.biortech.2015.07.095
0960-8524/Ó 2015 Elsevier Ltd. All rights reserved.
The benefits of biohydrogen (BioH2) production through dark
fermentation process regarding reduction in greenhouse gas
emission and valorization of organic waste materials cannot be
downplayed. However, low yield and production rate of BioH2 have
been the major barriers to the commercial-scale application of the
Z. Trad et al. / Bioresource Technology 196 (2015) 290–300
291
List of abbreviations
AnSMBR
CSTR
RTD
TMP
VFAs
A
CðtÞ
EðtÞ
EðhÞ
g
h
hin
hout
J
J pred
J0
k
ka
kb
kc
kd
Lp0
anaerobic submerged membrane reactor
continuous stirred tank reactor
residence time distribution
transmembrane pressure (Pa)
volatile fatty acids
membrane surface area (m2)
normalized concentration (-)
residence time function (-)
normalized residence function (-)
gravitational acceleration (m s2)
height (m)
inlet fiber height (m)
outlet fiber height (m)
permeate flux (L h1 m2)
predicted permeate flux (L h1 m2)
initial value of the flux (L h1 m2)
kinetic parameter of fouling (model-dependent unit)
complete blocking parameter (s1)
standard blocking parameter (s1/2)
intermediate blocking parameter (m1]
cake formation parameter (s m2)
water permeability (L h1 m2 bar1)
dark fermentation process. Among the contributing factors to low
BioH2 yield is the accumulation of volatile fatty acids (VFAs), which
results in a decrease in pH and consequent inhibition of the fermentation process. VFAs, including acetic, propionic and butyric
acids, are usually produced through the main mechanism of the
dark fermentation process, while other organic acids, such as
succinic, lactic and fumaric acids are produced when there is
imbalance during the digestion process. Therefore, regulation of
VFAs production by continuously removing them from the fermentation medium is crucial for the process stability and efficiency.
Meanwhile, extracted VFAs from fermentation medium can be
used for the production of fuels and energy through biochemical
of thermochemical pathways. The VFAs can also find a direct usage
as food additives in the food and the pharmaceutical industries
(Venkata-Mohan and Pandey, 2013). Different techniques including ion exchange (Gluszcz et al., 2004), adsorption (Joglekar
et al., 2006), electrodialysis (Huang et al., 2007; Wang et al.,
2006), liquid-liquid extraction (Mostafa, 1999; Senol and Dramur,
2004), distillation (Mumtaz et al., 2008), esterification (Pereira
et al., 2011), reactive extraction (Hong et al., 2001) and membrane
process (Wodaki and Nowaczyk, 1997) have been employed to
recover organic acids from fermentation broth. Among the various
techniques for VFAs recovery, membrane processes seem to be the
most efficient, eco-friendly and economic method. Membrane
filtration exhibits numerous benefits including small footprint,
and enhanced retention which enables sludge retention time to
be independently controlled from hydraulic retention time. It also
reduces the additional cost of disinfectant since it allows the
removal of microorganisms from VFAs to a certain degree for
subsequent treatment.
Generally, the efficiency and economics of membrane filtration
depend on membrane module design (tubular, plate and frame,
rotary disk or hollow fiber), pore size (microfiltration, ultrafiltration, nanofiltration or reverse osmosis), membrane material
(organic, inorganic, metallic, hydrophobicity or hydrophilicity),
filtration mode (dead end or cross mode), operating conditions
(flux, hydraulic retention time and sludge retention time) and
sludge characteristics (biomass concentration, pH, extracellular
polymeric substances and soluble microbial product). As a result,
many challenges are associated with the application of membrane
n
Pin
Pdiff
Pf
Pout
Pp
Pr
Ref
Rif
Rm
RT
t
tc
tm
blocking index
inlet pressure (Pa)
differential pressure (Pa)
feed pressure (Pa)
outlet pressure (Pa)
permeate pressure (Pa)
retentate pressure (Pa]
external fouling resistance (m1)
internal fouling resistance (m1)
clean membrane resistance (m1)
total membrane resistance (m1)
time (s)
circulation time (s)
mixing time (s)
fluid velocity (m s1)
filtrate volume (m3)
normalized time (–)
water viscosity (Pa s)
liquid density (kg m3)
variance of the error of the flux model (L2 h2 m4)
space time in the membrane module (s)
v
V
h
l
q
r2
s
filtration process, among which the most important is membrane
fouling. Membrane fouling is caused by particles deposition, plugging and narrowing of membrane pores and surfaces (Bae and Tak,
2005; Defrance et al., 2000). Flux and, hence, filtration efficiency is
directly affected by membrane fouling with a consequent decrease
in system productivity and increase in operating cost. Darcy’s law
highlights that the permeate flux through a porous membrane is
directly proportional to the transmembrane pressure (TMP) and
the membrane area, but is inversely proportional to the membrane
resistance due to fouling and to feed viscosity, as shown in Eq. (1)
(Field et al., 1995). In this equation, J is the permeate flux, l the viscosity of the liquid feed; TMP and RT, the transmembrane pressure
and the total resistance, are given by Eqs. (2) and (3).
J¼
TMP
l RT
TMP ¼
Pf þ Pr
Pp
2
RT ¼ Rm þ Ref þ Rif
ð1Þ
ð2Þ
ð3Þ
In the above equations, Pf, Pr, Pp, Rm, Ref and Rif are the feed,
retentate and permeate pressures, the clean membrane resistance,
and the external and internal fouling resistance, respectively. As
illustrated in Eq. (1), the rate of membrane fouling could be
reduced by carrying out filtration process below the critical flux,
and by maintaining simultaneously high shear rate through velocity gradient or gas sparging close to the membrane. Membrane
fouling can also be reduced by using appropriate membrane configuration and modules, as in the case of hollow fiber membrane
modules which constitute a common membrane configuration
employed in many industrial membrane processes owing to its
excellent mass transfer qualities and high membrane surface area.
Among the two main membrane configurations including
submerged and side-stream membrane bioreactors (MBRs),
internally-submerged MBR is usually preferred to side-stream
MBRs owing to its advantages which include smaller footprint
and less energy requirement (Cote et al., 1997; Singhania et al.,
2012). However, in some commercial applications, side-stream
MBRs were selected when a higher frequency for membrane
292
Z. Trad et al. / Bioresource Technology 196 (2015) 290–300
replacement is required at a given flow rate. An attractive alternative that combines the advantages of submerged and side-stream
technologies is the externally-submerged membrane bioreactor:
it can be operated at low pressure, while membrane cleaning and
replacement are easier (Singhania et al., 2012). However, this has
been disregarded in comparison to internally-submerged
devices. Up to now, most of the previous research work on VFAs
recovery from fermentation broths focused on the comparison of
separation technology used as a downstream treatment. The cyclic
extraction of VFAs during acidogenic fermentation using an
externally-submerged membrane coupled to the anaerobic digester
has not been investigated in detail. Consequently, the aim of this
work was to develop an original anaerobic externally-submerged
membrane bioreactor (AnSMBR) for the concurrent cyclic extraction of VFAs and bioH2 production. In addition, transmembrane
pressure (TMP) was studied as a function of the operating
conditions, while focusing on the cleaning procedure to assess the
feasibility of the filtration process.
rotation speed of the impeller was varied from 20 to 150 rpm.
The top impeller promotes equal flow discharge in the axial and
the radial directions due to the 45° angle, while the bottom impeller promotes mainly a radial flow discharge. A peristaltic pump,
denoted circulation pump, was used to impose the flow rate in
the external membrane module. The permeate was recovered
using another peristaltic pump, denoted withdrawal pump, that
sucked the liquid phase, causing a negative gauge pressure inside
the fibers. As a result, both the stirred tank and the loop could be
operated under atmospheric pressure, which prevented fermentation inhibition by H2 gas. The recirculation flow rate could be varied from 50 to 395.3 mL/min. This corresponded to space time
values in the tank between 12 and 100 min, and to space time values in the membrane module between 1.2 and 10 min. For
permeate recovery, the rotation speed of the withdrawal pump
could be varied from 200 to 1000 rpm, which was equivalent to
flow rates between 3 and 16 mL/min for water at atmospheric
pressure. The permeate flow rate was estimated by recording the
mass of permeate over time using an electronic balance.
2. Methods
2.2. Experimental procedures
2.1. Experimental set-up
The anaerobic membrane reactor system employed for
the membrane filtration experiments was composed of an
externally-submerged hollow fiber microfiltration (MF) membrane
module that was operated in the cross-flow ‘‘outside-in’’ mode and
placed in a recirculation loop coupled to a 5-L mechanically stirred
tank reactor (Fig. 1) from GPC (France). The cylindrical membrane
module was made up of 142 free polyvinylidene fluoride (PVDF)
fibers placed in 32.5 cm length housing with a surface area
A = 0.155 m2 and a 0.2 lm cut-off diameter. The volume of the
recirculation loop was measured, about 0.5 L. The stirred tank
(internal diameter of 17 cm and vessel height 35 cm) was equipped
with a two-stage impeller: the bottom impeller was a four-blade
disk (Rushton) turbine of 5.6 cm diameter and the top impeller
was a three-bladed 45° pitched turbine of 8.8 cm diameter. The
The first task was to validate the design of the bioreactor including the potential application of an externally-submerged membrane module, the volume ratio between the tank and the
module, and the range of recirculation and withdrawal flow rates.
Mixing time and residence time distribution (RTD) analysis as well
as TMP and permeate flow rate measurements were first carried
out using water. The results from the experiments were useful
for describing not only the hydrodynamic behavior of the
membrane module for fouling prevention and the influence of
the recirculation loop on the mixing properties of the stirred tank,
but also for the optimization of the inlet and outlet positions of the
recirculation loop in the tank. Experiments were later conducted to
measure TMP and flow rates using the actual fermentation broth as
it was done when water was used. All the experiments were done
in triplicate.
Fig. 1. Experimental setup: anaerobic SMBR with an external hollow fiber membrane module. (1) Tank reactor (5 L), (2) control unit, (3) impeller, (4) peristaltic pump, (5)
hollow fiber membrane module, (6) permeate withdrawal pump; Pin, Pout and Pdiff are pressure sensors.
Z. Trad et al. / Bioresource Technology 196 (2015) 290–300
2.2.1. Mixing and RTD analysis
A conductivity tracer technique was applied, in which distilled
water and sodium chloride (NaCl) were used as the working fluid
and the tracer, respectively. The conductivity tracer technique
involved the manual injection of an approximated d-Dirac pulse
of 1.0 M NaCl solution in the reactor system and the subsequent
detection of the tracer response signal using conductivity probes,
such as the CDC 241-9 (cell constant, 0.913 cm1) and the CDC
749 (cell constant, 1.45 cm1), connected to a CDM210
conductivity-meter (Radiometer Analytical, France). Both mixing
time and RTD (Residence Time Distribution) experiments were carried out in the stirred tank reactor, while only RTD was measured
in the membrane module with and without permeate extraction.
Data was recorded at 1 Hz frequency.
First, batch experiments were carried out to estimate mixing
time as a function of the rotation speed of the impeller. These were
compared to mixing time measurements in the presence of the
recirculation loop. The mixing time (tm) and mean circulation (tc)
times in the stirred tank were extracted from the experimental
curves of tracer concentration vs. time as a function of recirculation
flow rate and impeller rotation speed, as in Liu (2012). Then, RTD
analysis was carried out in the membrane fiber bundle under continuous flow conditions, without and with permeate extraction, as
a function of the impeller rotation speed in the tank, the circulation
flow rate and the speed of the withdrawal pump.
2.2.2. Transmembrane pressure (TMP) and permeate flow rate
measurements
Membrane filtration was driven in the cross-flow ‘‘outside-in’’
mode, i.e. with the filtration proceeding from outside to inside of
the fibers, as the feed solution was being pumped from the membrane module. As a result, the flow rate of permeate, J, was
deduced by weighing the mass of permeate withdrawn over time.
TMP was determined by measuring the pressure at the inlet and
the outlet of the fiber module and on the permeate side of the
membrane, using two absolute and one differential pressure sensors (Keller A.G, Germany). The differential pressure sensor and
one absolute pressure sensor were placed close to the membrane
inlet, while the other absolute pressure sensor was placed close
to the outlet of the membrane unit. TMP was deduced from experimental data using Eqs. (4)–(8).
TMP ¼
ðPin þ qghin Þ þ ðP out þ qghout Þ
Pp
2
ð4Þ
where Pin is the inlet pressure (measured), Pout the outlet pressure
(measured) and Pp the permeate pressure, while Pdiff is the differential pressure (measured), defined as:
Pdiff ¼ Pin P p
ð5Þ
As the fibers are vertical, gravitational potential energy must be
accounted for and the difference of height, Dh = hout hin, between
the outlet (top) and the inlet (bottom) of the fiber module emerges
from Eq. (5). As a result, TMP can be expressed as:
Pout Pin
þ Pdiff
TMP ¼ qg Dh þ
2
When there is no flow (fluid velocity
brane, Eq. (7) is obtained:
qg Dh ¼
Pout Pin
þ Pdiff
2
ð6Þ
v = 0) through the mem-
m¼0
ð7Þ
Finally, one deduces that:
TMP ¼
ðPout Pin Þ
Pout Pin
þ Pdiff þ Pdiff
2
2
m¼0
ð8Þ
293
which combines measurements when recirculation is stopped
(m ¼ 0) to data obtained during filtration.
2.2.3. Anaerobic digestion experiments
To obtain fermentation broth for filtration experiments, acidogenic fermentation was carried out using mixed cultures with a
mesophilic bacterial consortium. Glucose (20 g/L) was initially
used as the carbon source in order to enhance the reproducibility
of BioH2 production using batch experiments. Other complementary nutrients included K2HPO43H2O (6 g/L), KH2PO4 (6 g/L),
(NH4)2SO4 (12 g/L) as the nitrogen source, CaCl2, 2H2O (0.15 g/L),
FeSO4, 7H2O (0.4 g/L), MnSO4, 6H2O (0.15 g/L), NaCl (12 g/L),
MgSO4 (1.2 g/L), ZnSO4 (8.2 g/L). pH was controlled at 6, through
minute addition of KOH solution (8 M). Cultures were carried out
at 35 °C, starting with a 100% CO2 atmosphere and redox potential
about 350 mV. The mesophilic consortium promoted only BioH2
production because methanogenic bacteria had been removed.
The broth from the batch experiments was used, first, for testing
fouling and cleaning procedures in the membrane module. Then,
fed-batch fermentation experiments were carried out, with a cyclic
extraction of the liquid phase using the membrane module during
anaerobic digestion, so as to validate the design of the AnSMBR.
Fermentation experiments involving wheat straw as the substrate
(with the same complementary nutrients as with glucose) were
also conducted to ensure that mixing conditions in the tank were
compatible with the suspension of straw particles, while avoiding
their circulation in the membrane module. Substrate addition
(either 20 g/L glucose powder or 20 g/L grinded straw with particles of about 2 mm size after sieving) was carried out using a funnel; rapid addition coupled to anaerobic and low redox conditions
prevented contamination and did not disturb significantly the
fermentation.
Glucose and the VFAs profile, both in the tank and in the permeate, were measured after sampling, using an HPLC device (Agilent
Technologies 1100 series, USAs). The HPLC was fitted with two columns (Rezex ROA 300 7.8 nm, Phenomenex, USA) mounted in a
serial assembly inside an oven (50 °C) equipped with a refractometer as detector. The mobile phase was 2 mM sulfuric acid solution
prepared in ultra-pure water with a flow rate of 0.7 mL/min (isocratic mode). For the analysis, 2 mL of sample was mixed with
250 mL of Ba(OH)28H2O (0.3 M) and 250 mL of ZnSO47H2O (5%
w/v). The mixture was centrifuged at 10,000g for 5 min. The supernatant was filtered through a 0.45 mm cellulose acetate filter and
injected for analysis. The gas phase (mainly BioH2 and CO2) was
analyzed using an Agilent 3000 micro-GC gas analyser equipped
with two capillary columns and two thermal conductivity detectors. Volumes of gas produced were measured using a volumetric
flowmeter (Ritter, GC-1, Germany).
2.2.4. Analysis of fouling and membrane cleaning
The experimental set-up was designed to enable both gas
scouring (bubbling of gas through membrane surface) and backwashing with permeate. In this work, CO2 was used as the gas
phase, even though the final objective was to use the gas phase
produced during the fermentation process, i.e. a mixture of CO2
and H2, for gas scouring. Backwashing was obtained using the
reversible flow of the withdrawal peristaltic pump, which presented the drawback to use a fraction of the filtrate for backwashing. Fouling was studied by following the permeate flow rate over
time as a function of the rotation speed of the withdrawal pump.
The effectiveness of cleaning was assessed through the evolution
of the permeate flow rate J over several filtration-cleaning cycles.
294
Z. Trad et al. / Bioresource Technology 196 (2015) 290–300
3. Results and discussion
3.1. Validation of reactor design: hydrodynamics and mixing
3.1.1. Characterization of the stirred tank reactor
Mixing in the batch stirred tank bioreactor was studied, first, as
a reference in order to determine the influence of the additional
external membrane module on the behavior of the reactor. The
evolution of the normalized salt concentration was measured near
the impeller after tracer injection close to the free surface. The
response curves always exhibited a similar pattern for impeller
rotation speed between 20 and 150 rpm, as illustrated in Fig. 2
for 30 rpm. The mixing time (tm) in this case was about 128 s,
which was far smaller than the minimum residence time of the
fluid in the tank when the recirculation pump was operated at
its maximum flow rate, about 12.5 min. The curve showed that
the tracer quickly reached the sensor with a circulation time (tc)
about 15 s. As expected, tm and tc decreased when higher impeller
speed was applied (Fig. 2). Above 100 rpm, tc became so small, so
that it could not be measured any more, as it was about
one-tenth of the mixing time, which was, roughly, inversely proportional to rotation speed, as the product between rotation speed
and tm was about 70 (Table 1). Complementary experiments with
straw particles showed that 100 rpm was the minimum speed to
obtain a fully suspended solid phase. Similarly, additional experiments in which the recirculation loop was fed showed that the
mixing behavior of the stirred tank remained unaffected by the
recirculation. Consequently, the stirred tank appeared to be ‘‘instantaneously’’ mixed in comparison to the fluid in the recirculation loop. Experiments also validated the position of the
aspiration point of the loop near the surface in order to prevent
the presence of solids in the loop. In addition, Fig. 2 highlighted
the typical mixing pattern of turbulent flow, which was in agreement with Reynolds number that reached 1200 for rotation speed
at 30 rpm in the stirred tank. The Reynolds number of the stirred
tank varied between 103 and 2 104 for both impellers, considered
separately, which covers the region from the transitional flow to
fully-developed turbulent flow conditions. However, all the
response curves presented a similar shape, which was in agreement with a gradual transition between the flow regimes usually
observed in stirred tanks.
3.1.2. Characterization of the membrane module
As underlined above, the stirred tank appeared to be perfectly
mixed in less than 155 s at the lowest rotation speed, 20 rpm,
Fig. 2. Illustration of normalized NaCl concentration-time curve at various impeller
rotation speeds in the batch stirred tank.
Table 1
Mixing time and circulation times as a function of rotation speed in the tank.
Speed (rpm)
20
30
40
50
60
70
80
90
100
150
tm (s)
tc (s)
155
19
128
15
110
8
90
6
75
5
63
5
50
5
48
4
42
–
27
–
and in less than 30 s at the highest one, 150 rpm. This was far
lower than the residence time in the tank in which the minimum
value was 12.5 min for the maximum recirculation flow rate. As
the volume of the membrane module was one-tenth of that of
the tank, the minimum residence time in the module was 65 s,
indicating that the mixing time in the tank might also be smaller
than the residence time in the recirculation loop, for example at
a flow rate at which the straw was fully suspended, i.e. 100 rpm
(Table 1). This was the key information that confirmed that rotation speed in the stirred tank and recirculation flow rate could be
chosen so that the membrane module did not affect the mixing
properties and, therefore, the culture conditions in the AnSMBR.
As a result, mixing in the recirculation loop could be studied independently from the tank.
The normalized tracer response curves E(t) were measured by
injecting the tracer at the inlet of the recirculation loop. Typical
examples obtained without permeate extraction are shown in
Fig. 3a. RTD curves were obtained when the tank was not mechanically stirred. However, similar results were also obtained when
different rotation speeds were applied, which confirmed experimentally that the tank and the loop could be treated as independent systems. As expected, peak time became shorter with
increasing flow rate in Fig. 3a. Meanwhile, when the curves were
plotted vs. the normalized time h (i.e. time divided by the residence
time in the loop), all the curves presented a similar shape. Without
permeate extraction (Fig. 3b), all the typical curves exhibited a
peak emerging at h = 0.5, followed by a long tail. A key point was
that the experimental curves exhibited a shape rather close to
the theoretical RTD observed for an empty pipe in the laminar flow
regime (dashed curve in Fig. 3d), which could be expressed as a
function of the Heaviside function H(t) as follows:
EðtÞ ¼
s2 2t
3
H t
s
2
¼ EðhÞ ¼
1
H h
2
2sh
1
3
ð9Þ
This revealed that laminar flow conditions prevailed in the
membrane module, indicating that the bundle mainly behaved as
an empty tube, despite the presence of the fibers. As a matter of
fact, the Reynolds number based on the diameter of the bundle
was lower than 1000 at the highest flow rate. This showed that,
even without the fibers, laminar flow conditions would prevail.
As a result, it seemed that the surface area occupied by the fibers,
which implies higher speed and higher viscosity in comparison to
an empty pipe, did not change significantly the hydrodynamics.
Similarly, as shown also in Fig. 3c, permeate extraction did not
affect the trends, even when the recirculation flow rate was low.
With water, the withdrawal flow rate was about 15 mL/min in
Fig. 3c, which indicated that only 15% of the inlet flow was
extracted when it was 100 mL/min while for higher recirculation
flow rate, the permeate was less than 10% of the inlet flow. As a
result, this explained why the RTD curves were unaffected by permeate extraction.
As a conclusion, the results not only showed that the properties
of the fluid in the stirred tank were unaffected by the presence of
the recirculation loop, but also showed that the stirring speed in
the tank had no apparent effect on the mixing properties of the
loop. In addition, the loop exhibited nearly the same behavior with
and without permeate extraction, provided the recirculation flow
rate was more than ten times as high as the withdrawal flow rate.
Z. Trad et al. / Bioresource Technology 196 (2015) 290–300
295
Fig. 3. RTD curves in the membrane module: as a function of time and recirculation flow rate without permeate extraction (a); normalized curve as a function of normalized
time and recirculation flow rate without (b) and with permeate extraction (permeate withdrawal pump at 1000 rpm and impeller rotational speed rotation of 60 rpm) (c);
normalized curve as a function of normalized time (flow rate 300 mL/min, without permeate extraction) and compared with a laminar dispersion model (d).
As a consequent of these results, the AnSMBR could, therefore, be
described as a perfectly mixed tank coupled to a recirculation loop
which could be assimilated to a tubular laminar flow region and
does not affect significantly the hydrodynamics of the liquid phase
in the tank.
3.2. Validation of reactor design: filtration
The reactor design was validated using the results obtained
from filtration and cleaning measurements of the microfiltration
(MF) membrane using water and fermentation broth.
3.2.1. Characterization of membrane properties using water and broth
To quantify the hydraulic performance of the membrane module, different key parameters were investigated. First, a master
TMP curve was established from measurements using water as a
function of the flow rates imposed simultaneously by the withdrawal pump and the recirculation pump (Fig. 4a). This showed
that an increase in TPM was observed when the permeate flow rate
was increased, while TMP decreased when the recirculation flow
rate in the loop was increased. The increase of TMP vs. J was
expected (Eq. (1)); TMP should be proportional to J, as illustrated
qualitatively in Fig. 4a and quantitatively in Fig. 4b with a circulation flow rate of water at 50 mL/min. The slope for water in Fig. 4b
enabled the membrane permeability to water (Lp0 ) to be calculated
using Eq. (10):
J 0 ¼ Lp0 TMP
ð10Þ
Since the slope of the adjusted straight line was equal to
2.5 103, (Lp0 ) could be calculated and was equal to
250 L h1 m2 bar1, which was in agreement with the order of
magnitude of (Lp0 ) on ultrafiltration membranes. Consequently,
Rm was 1.45 1012 m1. It must be mentioned that this value was
obtained on a membrane that had already been used for filtration
with the digestate and that had only been subjected to physical
cleaning procedures. This highlighted the adequate properties of
the PVDF membrane in terms of material and pore size, as these
(Lp0 ) and Rm values were close to literature data for perfectly clean
membranes, between 50 and 500 L h1 m2 bar1 (El-Rayess et al.,
2012; Puspitasari et al., 2010; Yang et al., 2011).
Conversely, the decrease in TMP when the circulation flow rate
of water, as the fluid phase, was increased seemed
counter-intuitive. When the circulation flow rate was increased,
pressure drop increased in the module; consequently, both the
inlet pressure Pin and the differential pressure Pdiff increased at
constant permeate flow rate, while the outlet pressure remained
nearly constant, close to atmospheric pressure. Meanwhile, an
increase in TMP could be expected. As illustrated in Eq. (8); Pin
and Pdiff actually had opposite effects on TMP. Pin Pout was equal
296
Z. Trad et al. / Bioresource Technology 196 (2015) 290–300
Fig. 4. Analysis of membrane properties: influence of recirculation flow and permeate flow rates on TMP with water (a); Evolution of J vs. TMP at constant recirculation flow
rate: clean membrane (water) at 50 mL/min; j membrane filtration with culture medium on glucose at 50 mL/min and N for membrane filtration with culture medium on
glucose at 100 mL/min (b).
to pressure drop which was proportional to qv2/2 in the bundle.
While Pin was increased by the same pressure drop, it was also
decreased by the kinetic term qv2/2 in the inlet tube where the
fluid velocity was far higher than in the bundle. This effect seemed
to be predominant and might explain why a lower TMP was
required at constant J when the recirculation flow rate was
increased. Consequently, the TMP value required to achieve the
desired permeate flow rate could be reduced by increasing the circulation flow rate, indicating that increasing the circulation flow
rate decreased the apparent membrane resistance RT when water
was used as the fluid phase in the AnSMBR.
Based on the result, experiments were carried out to determine
the membrane properties using the fermentation broth. Two values of the recirculation flow rate were studied: 50 mL/min and
100 mL/min. As usually observed in tangential filtration due to
fouling, the curves initially presented an increase in J as a function
of TMP, followed by a plateau value above a TMP of 104 Pa which
corresponded to the maximum permeation flow rate that could
be achieved due to membrane fouling (Fig. 4b). As a result, any
further increase in TMP did not increase J, which was due to the
accumulation of foulants on the surface and/or in the pores of
the membrane. This figure also showed that the maximum J values
were dependent on the recirculation flow rate, 2.7 L h1 m2 for
100 mL/min and about 4 L h1 m2 for 50 mL/min. In addition, for
the same J value, TMP values of, at least, one order of magnitude
higher were required with the fermentation broth than with distilled water. These results could be due to the higher viscosity of
the broth (about twice as high as water) and from fouling that
resulted in an increase of RT in the linear region, roughly by a factor
5. It was also important to point out that the maximum J value
decreased when the recirculation flow rate was decreased, which
highlighted that higher shear rate in the module did not favor
membrane cleaning; conversely, higher fluid velocity accelerated
the accumulation of foulants.
As a conclusion, the hydraulic properties were determined both
with distilled water and the digestate. The results demonstrated
the feasibility of the filtration, provided irreversible fouling did
not occur and the membrane module could be operated during
anaerobic digestion. A key point was that TMP values required
for permeate recovery remained low, about 104 Pa, which confirmed the opportunity to use an externally-submerged membrane
reactor: contrary to side-stream membrane bioreactors, high TMP
and shear were not needed for filtration.
3.2.2. Analysis of cleaning procedures
In order to evaluate the cleaning methods used for membrane
after fouling, gas scouring was studied, first. CO2 was injected for
5 minutes after a filtration step of 15 minutes using the digestate;
then, the same cycle was repeated. In a second experiment, the
same procedure was applied, but using backwashing instead of
CO2 injection. The results showed that backwashing was
slightly more efficient for recovering the initial density flux
(4.45 L h1 m2), since the permeate flux after backwashing always
remained above 4.4 L h1 m2, while it was about 4.3 L h1 m2
after three CO2 injections (Fig. 5a). This indicated that fouling
was probably reversible, but that cleaning remained incomplete
using only gas scouring. The lower effectiveness of gas scouring
was demonstrated by an additional experiment in which a first filtration step of 145 min was applied, and then stopped for 70 min,
with the aim to simulate a longer filtration cycle. Physical cleaning
using CO2 injection was carried out just before starting a new filtration cycle. In Fig. 5b, the evolution of the permeate flux J vs. time
using the digestate showed that the initial permeate flux in phase
A, 4.5 L h1 m2, was almost recovered in phase C (4.2 L h1 m2)
after CO2 gas injection, but not totally, as expected. This, however,
confirmed that fouling was mainly reversible, but the permeate
flux declined rapidly in phase C in Fig. 5b, which highlighted that
the kinetics of fouling became more rapid when the membrane
was fouled the first time. A similar, but slower trend was observed
with backwashing, which highlighted that cleaning should be
applied frequently. In this case, using permeate for backwashing
was not acceptable from a techno-economic point of view. A more
attractive solution, therefore, involved using the substrate instead
of the permeate in a ‘‘cyclic’’ process.
As a result, a methodology of membrane cleaning was defined
which included flushing with CO2 for 3 min every 15 min during
permeate extraction and backwashing with fresh culture medium
when cyclic extraction was stopped. The consequence was that
many filtration/physical cleaning cycles could be operated before
there was a need for chemical methods. This procedure could also
be applied in the presence of straw, provided a strainer was placed
at the inlet and the outlet of the recirculation loop to avoid clogging the membrane, and that filtration was carried out with an
inlet point close to the top of the reactor at a reduced rotation
speed. These results also confirmed that PVDF membranes could
be used in this process, which was in accordance with literature
data (Nguyen et al., 2011) for low surface energy of PVDF; a
297
Z. Trad et al. / Bioresource Technology 196 (2015) 290–300
Fig. 5. Permeate flux J vs. time for filtration cycles with cleaning: comparison of scouring and backwashing cleaning methods (a); analysis of a filtration cycle: three phases –
A. Filtration – B. Filtration is stopped, gas scouring is applied 5 min before phase C – C. Filtration (b); Modelling of fouling mechanism for data from phase A in (b) (c);
modelling of fouling mechanism for data from phase C in (b) (d).
property that would promote the detachment of deposits and,
thus, cleaning and unclogging of the membrane.
3.2.3. Analysis of fouling mechanisms
In the literature, many studies have tried to identify the mechanisms of membrane fouling in anaerobic membrane bioreactors
(Charfi et al., 2012; Choo and Lee, 1998; He et al., 2005; Wang
and Tarabara, 2008) in order to prevent particle fouling on the
membrane surface or blocking in the membrane pores that caused
a rapid decline in filtration flux, as well as a major difficulty for
keeping a high performance. Different models were commonly
used to describe the fouling mechanisms and their relationship
with the biotic parameters (microorganisms can be attached, grow
on the membrane surface and produce enough of extracellular
polymeric substances to facilitate the development of a biofilm
on the surface of the membrane) and the abiotic parameters (deposition of material within the pores of the membrane and/or on its
surface). A first class of models was aimed at defining the relationship between the biological materials in the fluid and the effectiveness of separation techniques based on MF and ultrafiltration (UF).
However, an alternative approach to identify the mechanisms of
colloidal fouling based on Hermia’s analysis (Hermia, 1982) could
be used to define a general equation for different fouling mechanisms causing the flux decline as illustrated in Eq. (11).
2
d t
dt
¼k
2
dV
dV
n
ð11Þ
where k is constant and n is the blocking index equal to 2, 3/2, 1 or 0
for complete blocking, intermediate blocking, standard blocking,
and cake formation, respectively. The filtration time and the filtrate
volume V recovered at time t per membrane unit area A could then
be related to the permeate flux (Eq. (12)).
J¼
1 dV
A dt
ð12Þ
Combining Eqs. (12) and (11) gives:
dJ
¼ kJðAJÞ2n
dt
ð13Þ
This method is advantageous (Wang and Tarabara, 2008), as it
provides the opportunity to use a single plot to identify all blocking
mechanisms, and also determine how the data can be interpreted
in terms of blocking laws. A significant disadvantage of the direct
application of Eq. (13) is the high sensitivity of the value of n with
respect to the noise in the flux data, and therefore, its influence on
the choice of the fouling mechanism. The mathematical form of the
four conventional fouling mechanisms are summarized in Eqs.
(14)–(17) and expressed as a function of J0 (the initial flow rate)
and k, a kinetic parameter.
J ¼ J 0 eðka tÞ
J¼
ðComplete blocking; n ¼ 2Þ
J0
kb J 0 t þ 1
ðIntermediate blocking; n ¼ 1Þ
4J
J ¼ 1 0 2
kc J 20 t þ 1
J0
J¼
1=2
2kd J 20 t þ 1
ð14Þ
ð15Þ
ðStandard blocking; n ¼ 3=2Þ
ð16Þ
ðCake formation; n ¼ 0Þ
ð17Þ
298
Z. Trad et al. / Bioresource Technology 196 (2015) 290–300
Table 2
Optimized parameters and variance of the error of classical fouling models.
Fouling mechanisms
(a) Complete blocking
ka
Optimized ki
Phase A of Fig. 5
7.11 102 s1
Phase C of Fig. 5
3.41 102 s1
P
2
J pred J Þ
r2 ¼ ðN2
calculated on N experimental points
Phase A of Fig. 5
2.94 101
Phase C of Fig. 5
4.6 101
(b) Intermediate blocking
(c) Standard blocking
(d) Cake formation
kb
3.98 103 s1/2
9.44 102 s1/2
kc
1.14 102 m1
2.81 102 m1
kd
2.10 101 s m2
2.20 101 s m2
8.75 103
2.78 101
1.87 102
1.72 101
1.06 103
3.96 102
Fig. 6. Typical example of experimental data from dark fermentation including biogas volume and composition, pH, and VFAs concentrations in the liquid phase.
These models have been related to experimental data
(Fig. 5c and d) which corresponded to a clean membrane and to
a membrane just after gas scouring in Fig. 5b, respectively.
Experimental data was fitted by optimizing simultaneously J0
and k using a method of least squares based on the Levenberg–
Marquardt algorithm that minimizes the difference between
experimental J and predicted Jpred permeate flux values. The
adjusted parameters of the predicted profiles in Fig. 5c and d were
presented in Table 2 together with the variance of the error (r2) for
each model. The analysis of the results obtained from the
experiments on a clean membrane (Fig. 5c) showed that the cake
formation model (n = 0) with an optimized kinetic parameter
kd ¼ 2:1 102 s m2 exhibited a better fit of the experimental data
in comparison to the other models. Conversely, complete blocking
presented the highest error in Table 2. The same models were used
to predict fouling after CO2 gas scouring (Fig. 5d). Although fouling
was more rapid in this case, the plot displayed the same trends and
fouling corresponded to the cake formation model. As a conclusion,
these results clearly confirmed the reversibility of fouling, suggested in the previous section, and justified from a theoretical
point of view that chemical cleaning was not often required and
that a combination of gas scouring and backwashing constituted
an adequate cleaning procedure.
3.3. Validation of reactor design: coupling filtration and fermentation
The validation of the AnSMBR design and operating conditions
was carried out during anaerobic digestion experiments. Fig. 6
summarizes the evolution of the biogas volume and composition,
pH, and VFAs concentrations in the liquid phase over time. This figure also showed that pH control was effective and that the biogas
contained only CO2 and BioH2. Experiments were initially carried
out as in a conventional batch anaerobic bioreactor, with periodic
addition of glucose and withdrawal of digestate, mainly for
chemical analysis. After a latent period, gas production started,
mainly with CO2. Hydrogen content in the biogas was significant
after 400 h. For VFAs analysis, formate, acetate and butyrate were
shown to be the main VFA compounds, while lactate could also
be observed. On the contrary, propionate and succinate, isobutyrate, valerate and ethanol played a secondary role. Butyrate
was always the most abundant VFA when BioH2 production was
efficient, which indicated that the reaction followed mainly the
butyrate fermentation pathway to BioH2. This was confirmed by
the maximum BioH2 yield of 1.1 mol H2/mol glucose with a maximum production rate between 4 and 5 L H2/day L for 20 g/L substrate with a maximum of 58% BioH2 in the biogas during the
experiments, the maximum being 42% in Fig. 6. However, a rapid
accumulation of lactate emerged when the BioH2 productivity
decreased, which occurred after 1000 and 1500 h, respectively. A
comparison with literature data (Table 3) shows that the
AnSMBR of this study outperforms batch and fed-batch conventional bioreactors in terms of hydrogen production rate, which
was expected due to the inhibition by VFAs that emerges in conventional reactors, as mentioned by Kargi and Pamukoglu (2009).
The comparison of the gas phase composition also agrees with literature data, both in conventional and membrane bioreactors.
Conversely, a comparison of the VFA profiles cannot be drawn, as
this differs too strongly in the data of Table 3, which confirms that
it depends primarily on the combined effects of the substrate and
the microorganisms, rather than on the bioreactor configuration.
Finally, Table 3 also shows that without optimization and in unfavourable fed-batch conditions, the BioH2 production rate is close to
the values reported in the literature on other external submerged
membrane bioreactors, which confirms that the technology developed in this work is promising.
In this experiment, the reactor was operated as an AnSMBR
after 1400 h while permeate withdrawal using the process defined
in the previous sections was applied twice. The first filtration cycle
Anaerobic sludge
Cow dung
compost
150 mL – batch
Wheat straw
Wheat starch
5–35 g/L
20 g/L
11.1 g/L
16 g/L
30 g COD/
L
43.4 g
COD/L
52.7 g
COD/L
20 g/L
Feedstock
content
EtOH, ethanol; HAc, acetic acid; HBu, butyric acid; HPr, propionic acid; HLa, lactic acid; HCa, caproic acid.
Mixed microflora
Sucrose
Glucose
Mixed
mesophilic
microflora
Seed sludge
2 L – fed batch
Pore size: 0.04 lm
Effective surface
area: 0.047 m2
Pore size: 0.45 lm
Effective filtration
area: 0.1 m2
Food waste
Tofu
processing
waste
Synthetic
wastewater
Seed sludge
Pore size: 0.45 lm
Surface area: 0.1 m2
Size:
240 340 10 mm
Surface area:
0.025 m2
Glucose or
straw
Feedstock
(substrate)
Mixed microflora
Mixed
mesophilic
microflora
Micro-organism
Pore size: 0.2 lm
Effective surface
area: 0.155 m2
Membrane module
characteristics
Reactor without membrane module
1.9 L – CSTR
Submerged plate frame membrane with
5-L CSTR
Submerged hollow fiber module with 4LCSTR
Submerged hollow fiber module with 5-L
CSTR
Submerged flat sheet module with 5-L
CSTR
Reactor coupled to the membrane module
Submerged external hollow fiber
membrane with 5-L fed-batch tank
Membrane configuration
36
35
35
35
23
60
55
35
T
(°C)
–
–
4
9
8
8
10.5
–
HRT
(h)
(l) HAc, HBu, HLa
(g) 51–60% H2,
40–49% CO2
(il) HAc, HBu,
HLa, HPr
(g) 30–58% H2,
32–70% CO2
(l) HAc, HBu, HLa,
HCa, EtOH
(g) 42–50% H2,
40–51% CO2
(l) HAc, HBu, HPr,
HLa
(g) H2, CO2
(g) CO2, H2
Effluent
composition
(l) HAc, HBu, HPr
(g) H2, CO2
(l) EtOH, VFAs (C2–C6)
(g) 21.3–57.6% H2, 42–78% CO2
(l) HAc, HBu, HPr
(g) H2, CO2
4–5 L/m2 h
11.1 L/m2 min
4.32 L/m2 h
0.8–1.0 L/m2 d
3 L/m2 h
Permeate flux
0.25 L/d g
dried straw
1.3 L/d g
substrate
0.01 L/d g
0.36 L/d g
substrate
0.2 L/d g COD
0.3 L/d g COD
0.2 L/d g COD
0.2–0.25 L/d g
substrate
H2 production
rate
Table 3
Comparison of hydrogen production by anaerobic cultures using different kinds of conventional bioreactors (batch, fed-batch, CSTR) and submerged membrane bioreactors.
Kargi and
Pamukoglu
(2009)
Fan et al. (2006)
68.1 mL H2/g dried
straw
Han et al. (2014)
3.1
Lee et al. (2010)
Shen et al. (2010)
Kim et al. (2011)
Lee et al. (2014)
This study
References
3.31
1.19
0.004–0.008 mol/g
COD
1.87
2.2
1.1
H2 yield (mol
H2/mol hexose)
Z. Trad et al. / Bioresource Technology 196 (2015) 290–300
299
300
Z. Trad et al. / Bioresource Technology 196 (2015) 290–300
was operated at about 1400 h. In this period, the composition of
the biogas still exhibited a high BioH2 content, but the volume of
biogas produced was declining and the lactate concentration was
increasing. The first consequence of membrane filtration was that
the concentration of VFA decreased because the VFA profile of
the permeate was similar to that of the broth, which was partially
replaced by fresh substrate. This was followed by a peak of BioH2
content in the biogas and the rapid consumption of lactate anions.
These trends were similar to those observed after a conventional
addition of fresh substrate and withdrawal of digestate, for example, after the addition of glucose around 600 h. This result highlighted that the AnSMBR behaved as a conventional fed-batch
anaerobic bioreactor in this case, although it ensured that biomass
was maintained in the tank.
The second filtration cycle was operated around 1600 h. In this
case, the amount of biogas produced had decreased and this
contained almost exclusively CO2. The lactate concentration had
increased sharply with the total amount of VFAs (Fig. 6).
Filtration was used to remove and replace about one fourth of
the reactor volume and the effect of dilution appeared clearly in
the figure. Contrary to the previous cycle, bioH2 production did
not restart immediately due to the inhibition by the high VFAs content. However, the increase of lactate and VFAs contents was
delayed and by maintaining biomass in the bioreactor, a change
of metabolic pathway was achieved, so that lactate could be consumed and used as a substrate for bioH2 production, with butyrate
as the main by-product. As a result, bioH2 production restarted,
which highlighted the versatility of the AnSMBR for BioH2 production. By comparing these results with those of the previous
filtration cycle, one can conclude that bioH2 could be effectively
produced by dark fermentation using the externally-submerged
AnSMBR which seemed to be more versatile than a conventional
fed-batch bioreactor. Besides, lactate concentration could be a
good indicator of the time when a filtration cycle had to be started
so as to optimize BioH2 productivity with the microbial consortium
of this work.
4. Conclusions
In this work, the applicability of an original AnSMBR with an
externally-submerged MF module for BioH2 production was
assessed. Its geometry and operating conditions were chosen, so
that the hydrodynamic properties in the fermenter were not significantly modified. Low recirculation flow rate and TMP values maximized permeate flux. The fouling mechanism, due to cake layer
formation, was reversible. A cleaning procedure based on physical
methods was defined. During fermentation, BioH2 production was
shown to restart after removing VFAs with the permeate, so as to
enhance simultaneously BioH2 production and the recovery of
VFAs as platform molecules.
Acknowledgements
LABEX IMobS3 Innovative Mobility: Smart and Sustainable
Solutions, the French National Centre for Scientific Research
(CNRS), Auvergne Regional Council and the European Funds of
Regional Development (ERDF/FEDER) are gratefully acknowledged.
References
Bae, T.H., Tak, T.M., 2005. Interpretation of fouling characteristics of ulrafiltration
membranes during the filtration of membrane bioreactor mixed liquor. J.
Membr. Sci. 264, 151–160.
Charfi, A., Ben-Amar, N., Harmand, J., 2012. Analysis of fouling mechanisms in
anaerobic membrane bioreactors. Water Res. 46, 2637–2650.
Choo, K.H., Lee, C.H., 1998. Hydrodynamic behavior of anaerobic biosolids during
crossflow filtration in the membrane anaerobic bioreactor. Water Res. 32,
33387–33397.
Cote, P., Buisson, H., Pound, C., Arakaki, G., 1997. Immersed membrane activated
sludge for the reuse of municipal wastewater. Desalination 113, 189–196.
Defrance, L., Jaffrin, M.Y., Guptha, B., Paullier, P., Geaugery, V., 2000. Contribution of
various constituents of activated sludge to membrane bioreactor fouling.
Bioresour. Technol. 73, 105–112.
El-Rayess, Y., Albasi, C., Bacchin, P., Taillandier, P., Mietton-Peuchot, M., Devatine, A.,
2012. Analysis of membrane fouling during cross-flow microfiltration of wine.
Innov. Food Sci. Emerg. Technol. 16, 398–408.
Fan, Y.-T., Zhang, Y.-H., Zhang, S.-F., Hou, H.-W., Ren, B.-Z., 2006. Efficient conversion
of wheat straw wastes into biohydrogen gas by cow dung compost. Bioresour.
Technol. 97, 500–505.
Field, R.W., Wu, D., Howell, J.A., Guptha, B.B., 1995. Critical flux concept for
microfiltration fouling. J. Membr. Sci. 100, 259–272.
Gluszcz, P., Jamroz, T., Sencio, B., Ledakowicz, S., 2004. Equilibrium and dynamic
investigations of organic acids asorption onto ion-exchange resins. Bioproc.
Biosyst. Eng. 26, 185–190.
Han, H., Jia, Q., Wei, L., Shen, J., 2014. Influence of Cu2+ concentration on the
biohydrogen production of continuous stirred tank reactor. Int. J. Hydrogen
Energy 39, 13437–13442.
He, Y., Xu, P., Li, C., Zhang, B., 2005. High-concentration food wastewater treatment
by an anaerobic membrane bioreactor. Water Res. 39, 4110–4118.
Hermia, J., 1982. Constant pressure blocking filtration laws-Application to power
law non-Newtonian fluids. Trans. IChemE 60, 183–187.
Hong, Y., Hong, W., Han, D., 2001. Application of reactive extraction to recovery of
carboxylic acids. Biotechnol. Bioproc. Eng. 6, 386–394.
Huang, C., Xu, T., Zhang, Y., Chen, G., 2007. Application of electrodialysis to the
production of organic acids: state-of-the-art and recent developments. J.
Membr. Sci. 288, 1–12.
Joglekar, H.G., Rahman, I., Babu, S., Kulkarni, B.D., Joshi, A., 2006. Comparative
assessment of downstream processing options for lactic acid. Sep. Purif.
Technol. 52, 1–17.
Kargi, F., Pamukoglu, M.Y., 2009. Dark fermentation of ground wheat starch for biohydrogen production by fed-batch operation. Int. J. Hydrogen Energy 34, 2940–
2946.
Kim, M.-S., Lee, D.-Y., Kim, D.-H., 2011. Continuous hydrogen production from tofu
processing waste using anaerobic mixed microflora under thermophilic
conditions. Int. J. Hydrogen Energy 36, 8712–8718.
Lee, D.-Y., Li, Y.-Y., Noike, T., 2010. Influence of solids retention time on continuous
H2 production using membrane bioreactor. Int. J. Hydrogen Energy 35, 52–60.
Lee, D.-Y., Xu, K.-Q., Kobayashi, T., Li, Y.-Y., Inamori, Y., 2014. Effect of organic
loading rate on continuous hydrogen production from food waste in submerged
anaerobic membrane bioreactor. Int. J. Hydrogen Energy 39, 16863–16871.
Liu, M., 2012. Age distribution and the degree of mixing in continuous flow stirred
tank reactors. Chem. Eng. Sci. 69, 382–393.
Mostafa, N.A., 1999. Production and recovery of volatile fatty acids from
fermentation broth. Energy Convers. Manage. 40, 1543–1553.
Mumtaz, T., Abd-Aziz, S., Rahman, N.A.A., Tee, P.L., Shirai, Y., Hassan, M.A., 2008.
Pilot-scale recovery of flow molecular weight organic acids from anaerobically
treated palm oil mill effluent (POME) with energy integrated system. Afr. J.
Biotechnol. 7, 3900–3905.
Nguyen, P.T., Lasseuguette, E., Medina-Gonzalez, Y., Remigy, J.C., Roizard, D., 2011. A
dense membrane contactor for intensified CO2 gas/liquid absorption in postcombustion capture. J. Membr. Sci. 377, 261–272.
Pereira, C.S.M., Silva, V.M.T.M., Rodrigues, A.E., 2011. Ethyl lactate as a solvent;
properties, applications and production processes-a review. Green Chem. 13,
2658–2671.
Puspitasari, V., Granville, A., Le-Clech, P., Chen, V., 2010. Cleaning and ageing effect
of sodium hypochlorite on polyvinylidene fluoride (PVDF) membrane. Sep.
Purif. Technol. 72, 301–308.
Senol, A., Dramur, U., 2004. Predicting liquid-liquid equilibria of amine extraction of
carboxylic acid through solvation energy relation. Solvent Extr. Ion Exc. 25,
865–883.
Shen, L., Zhou, L., Mahendran, B., Bagley, D.M., Liss, N.L., 2010. Membrane fouling in
a fermentative hydrogen producing membrane bioreactor at different organic
loading rates. J. Membr. Sci. 360, 226–233.
Singhania, R.R., Christophe, G., Perchet, G., Troquet, J., Larroche, C., 2012. Immersed
membrane bioreactors: an overview with special emphasis on anaerobic
bioprocesses. Biores. Technol. 122, 171–180.
Venkata-Mohan, S., Pandey, A., 2013. Biohydrogen production: an introduction. In:
Pandey, A., Chang, J.-S., Hallenbeck, P.C., Larroche, C. (Eds.), Biohydrogen.
Elsevier, USA, p. 358.
Wang, F., Tarabara, V.V., 2008. Pore blocking mechanisms during early stages of
membrane fouling by colloids. J. Colloid Interface Sci. 328, 464–469.
Wang, Z., Luo, Y., Yu, P., 2006. Recovery of organic acids from waste salt solutions
derived from the manufacture of cyclohexanone by electrodialysis. J. Membr.
Sci 280, 134–137.
Wodaki, R., Nowaczyk, J., 1997. Extraction and separation of propionic and acetic
acid by permeation in a hybrid membrane system composed of liquid and ion
exchange polymer membrane. Solvent Extr. Ion Exc. 15, 1085–1106.
Yang, X., Wang, R., Shi, L., Fane, A.G., Debowski, M., 2011. Performance improvement
of PVDF hollow fiber-based membrane distillation process. J. Membr. Sci. 369,
437–447.
Paper IV
Enhanced fermentative hydrogen and methane productions
from inhibitory-fruit flavour medium with membraneencapsulated cells
Julius Akinbomi*, Rachman Wikandari, Mohammad, J. Taherzadeh
Swedish Centre for Resource Recovery, University of Borås, 501 90 Borås, Sweden;
* Author to whom correspondence should be addressed; E-Mail: [email protected];
Tel.: +46-334-354-487
Abstract: This study focused on the possibility of improving fermentative hydrogen and
methane productions from inhibitory-fruit flavour medium using PVDF membrane
encapsulated cells. Hexanal, myrcene and octanol that are naturally produced in fruits such
as apple, grape, mango, orange, straw berry and plum, were investigated. Batch and semicontinuous fermentation processes at 55°C were carried out. Presence of 5g/L of myrcene,
octanol and hexanal resulted in no methane formation by fermenting bacteria, while
encapsulated cells in the membranes resulted in successful fermentation with 182, 111 and
150 mL/gCOD of methane respectively. These inhibitors were not so serious inhibitors on
hydrogen producing bacteria. With free cells in the presence of 5g/L (final concentration)
of hexanal, myrcene and octanol flavour media, average daily yields of 68, 133, 88
mL/gCOD of hydrogen were obtained. However, cell encapsulation further improved these
hydrogen yields to 189, 179 and 198 mL/gCOD. The results from this study indicate that
the yields of fermentative hydrogen and methane productions from inhibitory medium
could be improved using encapsulated cells.
Keywords: Encapsulated bacteria; fruit flavours; membrane; hydrogen; methane;
inhibition;
1. Introduction
The increasing energy demand and depleting fossil fuel reserves coupled with the global warming
have stimulated a rapid growth in developing alternative energy sources that are sustainable, renewable
and environmentally friendly. Energy carriers such as hydrogen and methane have been suggested as
good substitutes for fossil fuels. Accordingly, various production pathways have been explored for
hydrogen and methane productions including water electrolysis (power-to-gas), thermo-chemical
processing, photo-chemical processing, photo-catalytic processing and photo-electrochemical
processing [1], as well as biological methods including photo-fermentation [2, 3] and anaerobic
2
fermentation [4, 5]. Among the diverse production pathways, anaerobic fermentation via dark
fermentation for hydrogen and methane production seems to be a promising option because of its low
energy requirement, renewable and non-polluting qualities as well as being able to utilize organic
residuals as carbon and energy sources. However, dark fermentation process is characterized by low
hydrogen yield, which consequently affects methane production. This phenomenon has been attributed
to factors; such as, substrate and product inhibitions, environmental and operating parameters as well
as the tendency of the fermentation process to result in biomass production. The challenge with most
of the hydrogen production pathways during dark fermentation is the problem of underutilization of
substrate with only one-third of the substrate having potential to be converted to hydrogen while the
remaining two-thirds forming organic acids and reduced compounds [6]. Moreover, some fermentative
feedstocks often contain inhibitory compounds that tend to inhibit the feedstock degradability by
anaerobic microorganisms.
Fruit waste has been widely utilized as feedstock during anaerobic fermentation due to its
degradability and availability from the huge turnout of the wastes from human consumption and
processing. However, the yields from the fruit fermentation processes are often low, which have been
attributed, among other factors, to the flavour compounds inherently present in the fruits. Flavours are
complex mixture of various organic compounds, including aldehydes, terpenoids, alcohols, ketones,
lactones and esters; with antimicrobial activity against a wide range of bacteria, yeasts and molds [713]. The antimicrobial natures of fruit flavours are evident in their various applications including as
food preservatives [14, 15] and alternative medicines [11, 16-21]. Previous research activities on the
effect of fruit flavour also confirmed the toxicity of fruit flavour compounds [22-28]. The toxicity of
flavour compounds against bacteria probably comes from the hydrophobic quality of flavour
compounds that allows them to penetrate and bind with phospholipids of bacterial cell membrane as
well as other cell organelles; thereby making them water permeable [29-34]. The cell integrity is lost if
the concentration of the accumulated flavour compound exceeds a tolerable limit. Although the
adaptive potential of bacterial against flavour compounds had been reported [35-37]; the hold-up time
of the bacteria depends on the concentration of the flavour compounds and the exposure period of the
bacteria to the flavour compounds, as the bacterial resistance cannot by itself be sustained for a long
period of time [38]. Considering the hydrophobic nature of flavour compounds, a hydrophilic barrier
can be created around the bacterial cells during fermentation process to prevent direct bacterial contact
with the flavour compounds as well as to reduce bacteria exposure time to the flavour compounds. The
technique of employing hydrophilic poly (vinylidene) fluoride barrier around anaerobic microorganism
in a medium containing flavour compounds in order to reduce the antimicrobial effects of the flavour
compounds during fermentation process; formed the basis of this study. Polyvinylidene fluoride
3
(PVDF) membrane is a semi-crystalline polymeric membrane consisting of both crystalline and
amorphous phases with crystalline part responsible for its excellent thermal stability while the
amorphous part is responsible for the flexibility of the membrane [39]. The membrane is chemically
stable to a wide range of chemical compounds including inorganic acids, oxidants, halogens, aromatic,
aliphatic and chlorinated solvents. However, the intrinsic hydrophobic nature of PVDF makes it to be
prone to organic fouling and low wettability with high resistance to water flow. Consequently, several
membrane modification techniques including blending, surface coating, irradiation grafting and plasma
modification, are used to incorporate hydrophilicity into hydrophobic PVDF membranes to enhance
their performances [40].
In several studies involving fermentation processes, cell encapsulation has been proved to be an
effective technique for cell stability, high biomass concentration, and enhanced fermentative hydrogen
and methane production [41, 42]. Cell encapsulation is vital for cell survival and increased tolerance
to toxic medium such as industrial wastewaters which contain toxic compounds including phenols,
benzenes and halogenated aliphatics, among others [43]. Application of membrane in cell
encapsulation has the potential of enhancing the total energy value of fermentation process which is
among the main objectives of producing hydrogen and methane from the process [44, 45]. Meanwhile,
there has been no previous report on the effects of using hydrophilic PVDF membranes for cell
encapsulation on fermentative hydrogen from media containing hexanal, myrcene and octanol flavours
though there were some reports on protective effects of membrane encapsulation on fermentative
methane production from limonene contained media [22, 23]. The objective of this study was therefore
to investigate the potential of using membrane-encapsulated cells to improve hydrogen and methane
productions from media containing hexanal, myrcene and octanol during batch and semi-continuous
fermentation processes. Enclosing fermentative microorganisms inside a hydrophilic membrane during
fermentation process could reduce the bacterial exposure to the antimicrobial effects of fruit flavours,
minimize the penetration of the fruit flavours, and thereby improve the yields of hydrogen and
methane productions. Besides, information from further studies on the direct correlation between the
concentration of flavour compounds and their corresponding antimicrobial effects could be applied in
the health sector to combat the menace of malaria and dengue fever epidemic in tropical regions of the
world.
2. Results and Discussion
Effective fermentative hydrogen and methane productions from fruit wastes during anaerobic
digestion is often limited by the inherent fruit flavours, which act as fruit defense mechanism against
microbial invasion. Hexanal, myrcene and octanol are fruit flavours that are naturally produced in
4
fruits such as apple, grape, mango, orange, straw berry and plum, which are essential parts of human
diet. Consequently, large quantities of the slowly digestible fruit wastes are generated from their
production, processing and consumption, thereby, constituting environmental pollution and health
hazards to people. It would therefore be necessary to devise a technique for improving degradation of
the fruit wastes and thereby increase hydrogen and methane production potential of the fruit wastes.
2.1. Effects of fruit flavours on methane production during batch fermentation process
The batch fermentation process for methane production of encapsulated and free cells from
medium containing 0.5% w/v (5g/L) concentration of fruit flavours including hexanal, myrcene and
octanol was carried out at 55°C for 11 days with manual mixing of the reactors twice a day. The results
indicated no methane production from free cells directly in contact with all the fruit flavours at
concentration of 0.5% w/v (Figure 1). On the contrary, cumulative methane yield of 182 ± 15, 111 ±
81 and 150 ± 24 mL/g COD were obtained from the encapsulated bacteria immersed in medium with
myrcene, octanol and hexanal, respectively. The lowest methane yield was from octanol indicating that
the inhibitory effect of octanol seemed to be stronger than that of hexanal and myrcene. Although this
could be related to the solubility, size and chemical structure of the flavour compounds, which
influence the flavour permeability; the mechanisms of inhibition during fermentation process are
sometimes difficult to understand partly due to the various adaptive abilities of fermentative
microorganisms.
Comparison of the accumulative methane production from encapsulated cells without flavour
compounds (membrane control) and encapsulated cells with flavour compounds (membrane hexanal,
membrane myrcene and membrane octanol); the results indicate that membrane protective effect
against the flavour compounds could be given approximately as 60%. This implied that the membrane
could protect more than half of the hydrogen production from being affected by the inhibitory effect of
the flavour compounds. The results, therefore suggest that it is possible to improve biomethane
production from medium with high concentration of fruit flavours using the technique of membranecell encapsulation technique.
5
Figure 1. Batch fermentation process for accumulated methane production from substrate
with fruit flavours in comparison to control experiment
2.2. Effects of fruit flavours on hydrogen production during semi- semi-continuous fermentation
process
The inhibitory effects of three flavour compounds (hexanal, myrcene and octanol) on hydrogen
production potentials of fermentative microorganisms were investigated during the semi-semicontinuous fermentation operated for 18 days at 55°C. The concentrations of the fruit flavour
compounds were increased at interval of 5 days starting with 0.05g/L through 0.5 and finally 5g/L.
After 15 days of fermentation process, feed supply and effluent withdrawal from the reactor were
stopped in order to observe for three days how the system adjust to the previous loading of the
inhibitory flavour medium. The average daily yields (Figure 2) and accumulated volumes of hydrogen
(Figure 3) obtained from the fermentation process clearly showed the protective effects of employing
encapsulated cells during anaerobic fermentation process. The average hydrogen productions from the
encapsulated cells were higher than the productions from the free cells. Meanwhile, none of the flavour
compounds used during the semi-semi-continuous fermentation process could be said to have the most
inhibitory effects, as the degree of the inhibitory effects varied among the fruit flavour. For example,
in batch fermentation process, octanol was found to have the most inhibitory effect while during the
semi-semi-continuous process; both hexanal and myrcene were observed to have greater inhibitory
effects than octanol. When the flavour compounds were exposed to the free cells, hexanal showed the
6
most inhibitory effect as indicated by the low average daily hydrogen yield of 68mL/gCOD, while
among the encapsulated cells, myrcene showed the lowest average daily hydrogen yield of 179
mL/gCOD. The variation could be due to the complexity of antimicrobial mechanisms of flavour
compounds coupled with the adaptive potential of the fermentative microorganisms.
At flavour concentration of 0.05 g/L, the inhibitory effects of the flavour compounds were not
significant as the average hydrogen yields from the encapsulated and free cells were almost the same
(Table 1). However, the inhibitory effects of the flavour compounds, especially among the free cells,
were considerably significant when the concentration was increased to 0.5 g/L. The percentage
reduction in average daily hydrogen yields from hexanal, myrcene and octanol were 77, 45 and 35%
respectively (Table 2). The increase in the flavour concentration did not have much effect on
encapsulated cells except when myrcene was used as the flavour which resulted in the reduction of the
average daily hydrogen yield by 23%. When the flavour concentration was increased from 0.5 to 5 g/L,
most of the cells including both encapsulated and free cells experienced improved activity regarding
the increase in average daily hydrogen yields (Table 2). The reason might probably be due to the
adaptive ability of the anaerobic microorganisms to the inhibitory medium as well as the potential of
the microorganisms to degrade some of the flavour compounds. Meanwhile, when the supply of
nutrient and withdrawal of effluent stopped, the average daily hydrogen production from the free cells
dropped significantly except for free cells in hexanal medium which experienced yield increase.
However, the average daily yields from the encapsulated cells did not experience much change after
three days of ending the feed supply and effluent withdrawal.
Throughout the experiment, it might be worthwhile to state that the increase in the concentration of
flavour compound in the fermentation medium did not significantly affect the average daily hydrogen
yield from the encapsulated cells as the hydrogen production was nearly constant. It was also observed
that, though free cells of hydrogen producing bacteria were able to produce reasonable amounts of
hydrogen regardless of the flavour inhibitors, the amount of hydrogen produced was less compared
with encapsulated cells. Based on the results, it could therefore be concluded that fermentative
hydrogen and methane productions from inhibitory fruit flavour medium could be improved using the
technique of membrane-cell encapsulation.
7
Figure 2. Semi-continuous fermentation process for daily hydrogen yield from substrate
with fruit flavours in comparison to control experiment
Figure 3. Semi-continuous fermentation process for cumulative hydrogen volume from
substrate with flavours in comparison to control experiment
8
Table 1. Average hydrogen yields at the three flavour concentrations
Flavour
compound
Free cells
Hexanal
Myrcene
Octanol
Membrane
Hexanal
Myrcene
Octanol
(A)
(B)
0.05g/L flavour
concentration
Average hydrogen yield (ml / gCOD)
0.5g/L flavour
5g/L flavour
No feeding and withdrawal
concentration
concentration
179.6
183.9
126.2
42.3
101.2
81.5
7.55
138.7
100.9
27.23
91.53
14.4
176.5
197.9
183.8
193.2
152.5
202.7
196.4
187.9
200.9
192.7
175.6
210.8
Table 2. Effects of change in flavour concentration on average hydrogen yield
Flavour
compound
(A)
(B)
Free cells
Hexanal
Myrcene
Octanol
Membrane
Hexanal
Myrcene
Octanol
Increase of flavour
concentration from
0.05 to 0.5g/L
Change in average hydrogen yield (%)
Increase of flavour
Reduction of flavour concentration
concentration from
from 5 to 0 g/L (no feeding and
0.5 to 5g/L
withdrawal)
(-) 77
(-) 45
(-) 35
(-) 82
(+) 27
(+)19
(+) 72
(-) 34
(-) 85
(+) 9
(-) 23
(+) 9
(+) 2
(+) 19
(-) 1
(-) 2
(-) 7
(+) 5
- reduction
+ increase
2.3. Digestate pH values during the semi-semi-semi-continuous fermentation process
The pH plays an important role during fermentative hydrogen production as it affects the metabolic
pathways in hydrogen production as well as limiting hydrogen consumption by hydrogenotrophic
methanogens [46-49].
Hydrogen and methane productions during fermentation process require
different pH values of 5.5 – 6.5 and 6.5 – 8.2, respectively. In this study, batch fermentation was used
for methane production with the pH range of 6.8 - 7.2 while semi-continuous fermentation was used
for hydrogen production with the range of initial pH range values of 5.2 to 5.9 [50]. During the semicontinuous fermentation, gradual reduction in the pH values of the fermenting media below 5.0 was
observed at the beginning of the experiment, which could be attributed to the production of organic
acids associated with the hydrogen formation during fermentation process [51]. The pH profile (Figure
9
4) indicated that the pH values for all the reactors did not vary significantly but were nearly constant
throughout the experiment with an average value of 4.40 ± 0.04. This could possibly imply that there
the daily effluent withdrawal from the reactor system could have prevented the accumulation of
organic acids that could have led to drastic reduction in the pH value of the fermentation media.
Moreover, it could also be probably due to the adaptation potential of fermentative microorganisms to
the inhibitory fermentative media.
Figure 4. Daily digestate pH values during the semi-continuous fermentation process
2.4. Implication of membrane applications for cell encapsulation
Encapsulation techniques could have some limitations including the inefficient diffusion of
nutrients to the microorganisms in the membrane as well as membrane fouling. It is often necessary to
determine the water permeability of the membranes to be employed during fermentation process. The
permeability results can also be useful in the determination of loss in membrane efficiency during after
the fermentation process. In this study, an average value of 0.048 mL/min of pure water permeability
was obtained for the PVDF membrane. This indicated that in a time period of one minute, the
membrane could allow an approximate value of 0.048mL of distilled water to pass through it..
Membrane permeability is influenced by various factors including the membrane materials (pore size,
hydrophobicity/hydrophilicity, free volume and filler particles) and solubility of the permeates [52-54].
The resistance to fluid flow through the membrane during filtration process is often due to
membrane fouling, which is a term that is used to describe the loss of membrane throughput.
10
Generally, fouling occurs when particulate, colloidal or soluble materials are deposited inside
membrane pores or surface. Membrane fouling is a major barrier to membrane application in
fermentation processes as it is associated with flux or permeate flow reduction, low permeate quality
and increased operational costs due to increased energy consumption. Membrane fouling is influenced
by factors such as sludge characteristics, operational parameters and membrane qualities. Although,
membrane fouling cannot be entirely avoided during membrane filtration process, the frequency of its
occurrence could be reduced through physical cleaning such as relaxation and backwashing or
chemical cleaning. Chemical cleaning of membrane is more effective in removing membrane fouling
than physical cleaning, but frequent use of chemical cleaning can damage the membrane and shorten
the membrane life-time. Previously, membrane cost was part of the barrier to the application of
membrane technology, but extensive research on membrane improvement has resulted in cheap and
affordable membrane in recent times. However, operating costs associated with membrane fouling
abatement is still the main barrier to the application of membrane technology.
3. Experimental Section
3.1. Materials
3.1.1. Anaerobic sludge,
Anaerobic sludge used for the digestion was obtained from a 3000-m3 municipal solid waste
thermophilic (55°C) digester at Borås Energy and Environment AB (Borås, Sweden). Prior to the start
of the experiment, the sludge was incubated at 55°C for 3 days to allow the bacteria to be activated and
digest the left-over carbon source. After the incubation, the sludge was thoroughly mixed and filtered
with a screen of 1mm pore size to remove particles bigger than the pore size of the screen. For
encapsulation purposes, the sludge was centrifuged at 14,000 x g for 10 min to separate the solid
inoculum from the supernatant [22].
3.1.2. Membrane-encapsulation procedure
The synthetic encapsulating sachets for holding the bacteria were made of flat sheet hydrophilic
polyvinylidene fluoride (PVDF) membranes (Durapore®, Thermo Fisher Scientific Inc., Sweden) with
pore size, thickness and diameter of 0.1µm, 125 µm and 90 mm, respectively. The encapsulating
sachets were prepared as described in previous report [22]. Each membrane was cut and folded into
rectangular dimensions with width and length of 3 and 6cm respectively. The membranes were heatsealed (HPL 450 AS, Hawo, Germany) with heating and cooling times of 5.5 s while leaving one side
11
left open for cell insertion after which the opening was sealed to form a membrane capsule. The
sealing and cooling times for the membranes were 5.0 and 5.5 s, respectively. The fermentation
process was carried out immediately the membrane encapsulation procedure was completed.
3.1.3. Nutrient medium and flavour compounds
The nutrient medium used during the fermentation process was a synthetic medium consisting of
20 g/L glucose (supplied by Merck), 20 g/L yeast extract (supplied by Merck) and 20 g/L nutrient
broth (supplied by Sigma-Aldrich). The nutrient broth contained D (+) -glucose (1 g/L), peptone (15
g/L), sodium chloride (6 g/L) and yeast extract (3 g/L). The medium was sterilized by filtration
through a 0.2 µm membrane before it was used for the fermentation process. The flavour compounds
(supplied by Sigma-Aldrich), consisting of hexanal, myrcene and octanol, were used as inhibitors
during the fermentation process.
3.2. Experimental set-up and procedures
The experiment was separated into two parts. The first part was batch fermentation for methane
production while the second part was the semi-continuous fermentation process for hydrogen
production. Both fermentation processes were carried out under thermophilic condition (55°C) and the
same flavour compounds including hexanal, myrcene and octanol were used. The seed inoculum was
incubated at 55°C for three days before it was employed for both batch and semi-continuous
fermentation process digestion process [55].
3.2.1. Batch fermentation process for methane production
The reactors used for the batch fermentation of methane were 118 mL serum glass bottles with
active volume of 53.5 mL and headspace of 65.5 mL. Each reactor was filled with 1.0 mL of filtered
nutrient medium containing 20 g/L each of nutrient broth, yeast extract and D (+) -glucose. Three fruit
flavour compounds, including hexanal, myrcene and octanol, were used with each having 0.5% w/v
concentration prepared by dissolving 5 g of the inhibitor in 1 liter of distilled water. Fifty milliliters of
the anaerobic sludge was measured and centrifuged from which 3 g pellet was used for the
encapsulation. For each flavour investigated, the batch fermentation reactors were grouped into two
categories, including, encapsulated and free cells. For encapsulated cells with inhibitor; the reactor
bottle contained 3 g of the inoculum pellet encapsulated in the membrane, 47 ml of distilled water, 2.5
mL of the flavour compound (0.5% w/v) and 1 mL of nutrient medium. Regarding the free cells with
inhibitor; the reactor bottle contained 50 mL of uncentrifuged inoculum and 2.5 mL of the flavour
compound and 1 mL of nutrient medium. Besides the two groups of reactor bottles, other groups
12
included membrane and free cells controls; both of which differed from the first two groups by the
replacement of the fruit flavour with 2.5 mL of distilled water. Blank reactors containing 50 mL of
non-centrifuged inoculum and 3.5 mL of distilled water were also prepared. After filling the serum
glass bottles with appropriate medium of pH between 6.8 and 7.2, they were closed with rubber seals
and plastic caps. The bottle headspace was flushed with 80% nitrogen and 20% carbon dioxide to
create anaerobic environment [56]. All the experiments were carried out in triplicates and incubated at
55°C in a water bath. During the course of the experiment, the reactor bottles were shaken manually
two times everyday to enhance the fermentation activities.
3.2.2. Semi-continuous fermentation process for hydrogen production
The semi-continuous experiments were carried out using parallel 500 mL bioreactors and
automatically gas volume recording system (AMPTS, Bioprocess Control Sweden AB, Sweden). Prior
to the start of the semi-continuous experiment, the sludge for the fermentative hydrogen production
was heat-pretreated at 100°C for 15 min and the initial pH adjusted to values between 5.2 and 5.9, as
hydrogen production has been observed to be enhanced at the pH range [57]. An average of 32 g pellet
of the inoculum sludge (equivalent to 5.6 g VSS/L) obtained from the centrifuged sludge was used
separately for each reactor with free and encapsulated cells. Regarding the reactors with free cells, the
inoculum pellet (5.6 g VSS/L) was added into each 500 mL glass reactor bottle (liquid volume of 450
mL) containing 300 mL of filtered nutrient medium and 97 mL of distilled water. The nutrient medium
was composed of 20 g/L each of nutrient broth, yeast extract and D-glucose. The resulting mixture was
thoroughly mixed so that the inoculum pellet could dissolve completely to form homogeneous mixture.
The flavour compounds (myrcene, octanol and hexanal) were prepared in three different
concentrations including 0.05, 0.5 and 5 g/L, after which, 21 mL of the prepared flavour solutions was
added into each reactor. For encapsulated cell-reactors, the inoculum pellet (32g) was divided into
eight equal portions (4 g each), which were inserted into eight membrane sachets. Each reactor bottle
contained eight membrane sachets with each sachet enclosing 4 g of inoculum pellet. The whole
experiment was started with the addition of lowest flavour concentration (0.05 g/L) while the gradual
increase in concentration was done at an interval of 5 days. With the constant active volume of 450
mL and daily flow of 50 mL/d, the hydraulic retention time (HRT) throughout the experiment was 9
days. Throughout the experiment, the reactor bottles were shaken twice a day to ensure adequate
contact among the nutrients, anaerobic cells and flavour compounds. The pH of the effluent withdrawn
each day of the experiment was measured in order to gain insight into the state of the fermentation
process during the experiment.
13
3.3. Analytical method
The volumes of biogas and hydrogen generated during the anaerobic fermentation processes were
measured using a data acquisition system (AMPTS, Bioprocess Control Sweden AB, Sweden). The
individual gas compositions were determined by using a 0.25 mL syringe (VICI, precious sampling
Inc., USA) for the gas sampling while the gas quantification was done using a gas chromatograph
(Perkin-Elmer, USA). The gas chromatograph was equipped with a packed column (Perkin-Elmer, 6′ x
1.8″ OD, 80/100, Mesh, USA) and a thermal conductivity detector (Perkin-Elmer, USA) with an inject
temperature of 150°C. Nitrogen at a flow rate of 20 mL/min at 60°C was used as carrier gas.
3.4. Membrane performance measurement
The ability of a membrane to regulate the permeation of various molecules through it is an
important feature that is employed in separation processes.. The driving forces producing movement of
permeants, which could be concentration, pressure, temperature and electromotive force; are connected
in such a way that overall driving force is the chemical potential. Membrane permeability determines
the rates of movement of nutrients and inhibitors into the cells of the fermentative microorganisms as
well as the discharge of the cell metabolism products. In this study, distilled water was used to
determine the pure water permeability (PWP) parameter of the hydrophilic PVDF membranes used
during the experimental work in this study. The time required for a definite quantity of distilled water
to pass through the membranes was observed and recorded. The water flow rate through the membrane
was calculated by dividing the volume of permeated water by the time required for the permeation.
Since the experiment was carried out at room temperature (22°C), the temperature correction of 0.794
was used to adjust the values obtained from the permeability test.
4. Conclusions
The major barrier associated with the widespread applications of fermentative hydrogen and
methane as fuels include, among others, the low yields of the gas production. The low yields have been
partly attributed to substrate inhibition. This study, therefore, investigated the inhibitory effect of some
flavour compounds in fruits, which is one of the important factors contributing to low hydrogen and
methane yields during fermentation process of fruit wastes. The potential of employing membrane
technology to improve the yields of hydrogen and methane from such process was explored. The
results suggest that the membrane-based techniques could actually improve hydrogen and methane
production from fermentation media with substrate inhibition. Compared with the free cells,
14
membrane-encapsulated cells produced methane faster and were able to survive the effects of the
inhibitory flavour medium. Higher gas productions were also observed from encapsulated cells, when
compared to free cells, in the inhibitory fruit flavour. However, it could be observed from the results
obtained that the membrane could not completely protect the fermentative organism against the
inhibitory effects of flavour compounds. Therefore, further membrane improvement is necessary to
effectively protect the microorganism from the inhibitory fruit flavour medium
Acknowledgments
This study was financially supported by the Swedish Research Council (Sweden).
Author Contributions
Julius Akinbomi is the first author and responsible for designing, performing and writing of the
article while Rachma Wikandari assisted in the designing of the experiment. Professor Mohammad
Taherzadeh is the main supervisor of the first author responsible for guidance and supervision of the
project from experimental designing to the revision of the manuscript.
Conflicts of Interest
The authors declare no conflict of interest.
References
1.
2.
3.
4.
5.
6.
7.
8.
9.
Mormirlan, M.; Veziroglu, T. N., Current status of hydrogen energy. Renew. Sustain. Energy
Rev. 2002, 6, 141-179.
Ike, A.; Toda, N.; Tsuji, N.; Hirata, K.; Miyamoto, K., Hydrogen photoproduction from CO2fixing microalgal biomass: Application of halotolerant photosynthetic bacteria. journal of
fermentation and Bioengineering 1997, 84, 606-609.
Melis, A.; Happe, T., Hydrogen production. Green algae as a source of energy. Plant
Physiology 2001, 127, 740-748.
Joyner, A. E.; Winter, W. T.; Godbout, D. M., Studies on some characteristics of hydrogen
production by cell-free extracts of rumen anaerobic bacteria. Canadian Journal of
Microbiology 1977, 23, 346-353.
Nandi, R.; Sengupta, S., Microbial production of hydrogen: An overview. Critical Reviews in
Microbiology 1998, 24, 61-84.
Hallenbeck, P. C., Fermentative hydrogen production: Principles, progress, and prognosis.
International Journal of Hydrogen Energy 2009, 34, 7379-7389.
Ouattara, B.; Simard, R. E.; Holley, R. A.; Piette, G. J.; Begin, A., Antibacterial activity of
selected fatty acids and essential oils against six meat spoilage organisms. International journal
of food microbiology 1997, 37, 155-62.
Upadhyay, R. K., Essential oils: Anti-microbial, antihelminthic, antiviral, anticancer and antiinsect properties. J. Appl. Biosci. 2010, 36, 1-22.
Karaman, S.; Digrak, M.; Ravid, U.; Ilcim, A., Antibacterial and antifungal activity of the
essential oils of Thymus revolutus Celak from Turkey. J. Ethnopharmacol. 2001, 76, 183-186.
15
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
23.
24.
25.
26.
27.
28.
29.
Friedman, M.; Henika, P. R.; Levin, C. E.; Mandrell, R. E., Antibacteria activities of plant
essential oils and their components against Escherichia coli O157 H7 and Salmonella enterica
in apple juice. J. Agric Food Chem. 2004, 52, 6042-6048.
Singh, A. K.; Tripathi, A. K.; Bindra, R. L.; Kumar, S., Essential oil and isolates for controlling
household insects, housefly, cockroach and mosquito. J. Med. Arom. Plant Sci. 2000, 22, 2526.
Vimal, M.; Vijaya, P. P.; Mumtaj, P.; Seema-Farhath, M. S., Antibacterial activity of selected
compounds of essential oils from indigenous plants. Jornal of Chemical and Pharmaceutical
Research 2013, 5, 248-253.
Smith-Palmer, A.; Stewart, J.; Fyfe, L., Antimicrobial properties of plant essential oils and
essences against five important food-borne pathogens. Lett. Appl. Microbiol. 1998, 26, 118122.
Mishra, A. K.; Dubey, N. K., Evaluation of essential oils for their toxicity against fungi causing
deterioration of stored food commodities. Appl. Environ. Microbial. 1994, 60, 1101-1105.
Faid, M.; Bakhy, K.; Anchad, M.; Tantaoui-Elaraki, A.; 1995;58:547-550., A.,
Physicochemical and microbiological characterizations and preservation with sorbic acid and
cinnamon. J. Food. Prod. 1995, 58, 541-550.
Reynolds, J. E. F., Martindale-the extra pharmacopoeia. 31st edition ed.; Pharmaceutical
Society of Great Britain: London, 1996.
Hammer, K. A.; Carson, C. F.; Riley, T. V., Susceptibility of transient and commercial skin
flora to the essential oil of Melaleuca alternifolia (Tea Tree oil). Ame. J. Infection Control
1996, 24, 186-189.
Shubina, L. P.; Siurin, S. A.; Savchenko, M., Inhalations of essential oils in the combined
treatment of patients with chronic bronchitis. Vrachebnoe-Delo. Kiev Part 1990, 5, 66-67.
Moris, J. A.; Khettry, A.; Seitz, E. W., Antimicrobial activity of aroma chemicals and essential
oils. J. Amer. Oil. Chemist Society 1979, 56, 595-603.
Hammer, K. A.; Caarson, C.; Riley, T. V., Antimicrobial activity of essential oils and other
plant extracts. J. Appl. Microbiol. 1999, 86, 985-990.
Singh, A. K.; Srivastava, M.; Singh, A. B.; Srivastava, A. K., Cinnamon bark oil, a potent
fungitoxicant against fungi causing respiratory tract myoses. Allergy 1995, 50, 995-999.
Youngsukkasem, S.; Akinbomi, J.; Rakshit, S.; Taherzadeh, M. J., Biogas production by
encapsulated bacteria in a synthetic membranes: Protective effects in toxic media and high
loading rates. Environ. Technol. 2013, 34, 2077-2084.
Wikandari, R.; youngsukkasem, S.; Millati, R.; Taherzadeh, M. J., Performance of semi-semicontinuous membrane biotreactors in biogas production from toxic feedstock containing Dlimonene Bioresource Technology 2014, 170, 350-355.
Martin, M. A.; Siles, J. A.; China, A. F.; Martin, A., Biomethanization of orange peel waste.
Bioresource Technology 2010, 101, 8993-8999.
Mizuki, E.; Akao, T.; Saruwatari, T., Inhibitory effect of citrus Unshu peel on anaerobic
digestion. Biol. Wastes 1990, 33, 161-168.
Grohmann, K.; Baldwin, E.; Buslig, B., Production of ethanol from enzymatically hydrolyzed
orange peel by the yeast Saccharomyces cerevisiae. Appl. Biochem. Biotecnol. 1994, 45-46,
315-327.
Winniczuk, P. P.; Parish, M. F., Minimum inhibitory concentrations of antimicrobials against
micro-organisms related to citrus juice. Food Microbiol 1997, 14, 373-381.
Talebnia, F.; Niklasson, C.; Taherzadeh, M. J., Ethanol production from glucose and diluteacid hydrolyzates by encapsulated S. cerevisiae. Biotechnol. Bioeng 2004, 90, 345-353.
Griffin, S. G.; Wyllie, S. G.; Markham, J. L.; Leach, D. N., The role of structure and molecular
properties of terpenoids in determining their antimicrobial activity. Flavour. Fragr. J. 1999, 14,
322-332.
16
30.
31.
32.
33.
34.
35.
36.
37.
38.
39.
40.
41
42
43
44
45
46.
47.
48.
Gutierrez, M. E.; Garcia, A. F.; de-Madariaga, M. A.; Sagrista, M. L.; Casado, F. J.; Mora, M.,
Interaction of tocophenols and phenolic compounds with membrane lipid components.
Evaluatiuon of their antioxidant activity in a liposomal model system. Life Sci. 2003, 72, 23372360.
Lee, S. E.; Hwang, H. J.; Ha, J. S.; Jeong, H. S.; Kim, J. H., Screening of medicinal plant
extracts for antioxidant activity. Life Sci. 2003, 73, 167-179.
Davidson, P. M.; Naidu, A. S., Phytophenols. In Natural food antimicrobial systems, Naidu, A.
S., Ed. CRC Press: Boca Raton, FL, 2000; pp 265-294.
Dorman, H. J. D.; Deans, S. G., Antimicrobial agengts from plants: Antibacterial activity of
plant volatile oils. J. Appl. Microbiol. 2000, 88, 308-316.
Sikkema, J.; Bont, J. A. M.; Poolman, B., Interactions of cyclic hydrocarbons with biological
membranes. J. Biol. Chem. 1994, 269, 8022-8028.
Cardozo, P. W.; Calsamiglia, S.; Ferret, A.; Kamel, C., Effects of natural plant extracts on
protein degradation and fermentation profiles in semi-continuous culture. J. Anin. Sci. 2004,
82, 3230-3236.
Molero, R.; Ibars, M.; Calsamiglia, S.; Ferret, A.; Losa, R., Effect of a specific blend of
essential oil compounds on dry matter and crude protein degradability in heifers fed diets with
different forage to concentrate rations. Anim. Feed Sci. Tecnol. 2004, 114, 91-104.
Castillejos, L.; Calsamiglia, S.; Ferret, A.; Losa, R., Effects of dose and adaptation time of a
specific blend of essential oils compounds on rumen fermentation. Feed Sci. Technol. 2007,
132, 186-201.
Burt, S., Essential oils: their antibacterial properties and potential applications in foods-a
review. Int. J. Food Microbiol 2004, 94, 223-253.
Singhania, R. R.; Christophe, G.; Perchet, G.; Troquet, J.; Larroche, C., Immersed membrane
bioreactors: An overview with special emphasis on anaerobic bioprocesses. Bioresource
Technology 2012, 122, 171-180.
Zhao, Y.-H.; Qian, Y.-L.; Zhu, B.-K.; Xu, Y.-Y., Modification of porous poly(vinylidene
fluoride) mebrane using amphiphilic polymers with different structures in phase inversion
process. Journal of Membrane Science 2008, 310, 567-576.
Sivagurunathan, P.; Gopalakrishnan, K..; Lin, C-Y., Hydrogen and ethanol fermentation of
various carbon sources by immobilized Escherichia coli (XL1 –Blue). International Journal of
Hydrogen Energy 2014, 39, 6881 – 6888.
Sivagurunathan, P.; Gopalakrishnan, K.; Sen, B.; Lin, C-Y., Development of a novel hybrid
immobilization material (HY-IM) for fermentative biohydrogen production from beverage
wastewater. Journal of the Chinese Chemical Society 2014, 61, 827 - -830.
Moreno-Andrade, I.; Moreno, G.; Kumar, G.; Buitron, G., Biohydrogen production from
industrial wastewaters. Water Science and Technology 2015, 71, 105 – 110.
Buitron, G.; Kumar, G.; Martinez-Arce, A., Moreno, G., Hydrogen and methane production via
a two-stage processes (H2-SBR + CH4-UASB) using tequila vinasses. International Journal of
Hydrogen Energy 2014, 39, 19249 – 19255.
Bakonyi, P.; Kumar, G.; Nemestothy, N.; Lin, C.Y.; Belafi-Bako, K., Biohydrogen purification
using a commercial polyimide membrane module: Studying the effects of some process
variables. International Journal of Hydrogen Energy, 2013, 38, 15092 – 15099.
Lay, J. J., Modelling and optimization of anaerobic digested sludge converting starch to
hydrogen. Biotechnol Bioeng. 2000, 68, 269-278.
Zhang, T.; Liu, H.; Fang, H. H. P., Biohydrogen production from starch in wastewater under
thermophilic condition. Journal of Environmental Management 2003, 69, 149-156.
Mah, R. A.; Ward, D. M.; Baresi, L.; Glass, T. L., Biogenesis of methane. Ann. Rev. Microbiol
1977, 31, 309-341.
17
49.
50.
51.
52.
53.
54.
55.
56.
57.
Oh, S. E.; Van-Ginkel, S.; Logan, B. E., The relative effectiveness of pH control and heat
treatment for enhancing biohydrogen gas production. Environ Sci Technol 2003, 37, 51865190.
Khanal, S. K.; Chen, W. H.; Li, L.; Sung, S., Biological hydrogen production: effects of pH
and intermediate products. International Jounal of Hydrogen Energy 2004, 29, 1123-1131.
Hwang, M. H.; Jang, N. J.; Hyun, S. H.; Kim, I. S., Anaerobic bio-hydrogen production from
ethanol fermentation: the role of pH Journal of biotechnology 2004, 111, 297-309.
Pourbafrani, M.; Talebnia, F.; niklasson, C.; Taherzadeh, M. J., Protective effect of
encapsulation in fermentation of limonene-contained media and orange peel hydrolyzate. Int J
Mol Sci 2007, 8, 777-787.
Crank, J.; Park, G. S., Diffusion in polymers. Academic Press: New York, 1968.
Nicholson, J. W., The chemistry of polymers. The Royal Society of Chemistry: Cambridge,
1997.
Hansen, T. L.; Schmidt, J. E.; Angelidaki, I.; Marca, E.; Jansen, J. l. C.; Mosbæk, H.;
Christensen, T. H., Method for determination of methane potentials of solid organic waste.
Waste Management 2004, 24, 393-400.
Angelidaki, I.; Alves, M. M.; Bolzonella, D.; Borzacconi, L.; Campos, J. L.; Guwy, A. J.;
Kalyuzhnyi, S.; Jenicek, P.; van-Lier, J. B., Defining the biomethane potential (BMP) of solid
organic wastes and energy crops: a proposed protocol for batch assays. Water Science and
Technology 2009, 59, 927-934.
Lin, C-Y.;Lay, C-H.; Sen, B.; Chu, C-Y.; Kumar, G.; Chen, C-C.; Chang, J-S., Fermentative
hydrogen production from wastewaters: A review and prognosis. International Journal of
Hydrogen Energy 2012, 37, 15632 - 15642
Paper V
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
Development and Dissemination Strategies for
Accelerating Biogas Production in Nigeria
Julius Akinbomi,a,b Tomas Brandberg,a Sikiru A. Sanni,b and
Mohammad J. Taherzadeh a
Following the worsening energy crisis of unreliable electricity and
unaffordable petroleum products coupled with the increase number of
poverty-stricken people in Nigeria, the populace is desperately in need of
cheap alternative energy supplies that will replace or complement the
existing energy sources. Previous efforts by the government in tackling
the challenge by citizenship sensitization of the need for introduction of
biofuel into the countryʼs energy mix have not yielded the expected
results because of a lack of sustained government effort. In light of the
shortcomings, this study assesses the current potential of available
biomass feedstock for biogas production in Nigeria, and further proposes
appropriate biogas plants, depending on feedstock type and quantity, for
the six geopolitical zones in Nigeria. Besides, the study proposes
government-driven biogas development systems that could be effectively
used to harness, using biogas technology, the estimated 270 TWh of
potential electrical energy from 181 million tonnes of available biomass,
in the advancement of electricity generation and consequent
improvement of welfare in Nigeria.
Keywords: Biogas; available feedstock; Nigerian’s prospect; Biogas-consultancy; Electricity
Contact information: a: Swedish Centre for Resource Recovery, University of Borås, 501 90 Borås,
Sweden; b: Department of Chemical and Polymer Engineering, Lagos State University, Lagos, Nigeria;
* Corresponding author: [email protected]
INTRODUCTION
Energy accessibility is the catalyst for economic growth, development, and
poverty alleviation, and it determines the level of social development in a country. Over
the years, Nigeria has been facing numerous challenges including a severe electricity
shortage, an inefficient waste management system, and environmental degradation. More
than 60% of the population does not have access to the national power supply because
they are not connected to the grid system; and even for those that are connected to the
grid system, power outages are a common challenge (Kennedy-Darling et al. 2008;
Okoye 2007). As a result of an unstable power supply, most people currently rely on
generators for their supply of off-grid electricity. Inadequate and inaccessible energy
services have compelled most industries and businesses that could not afford the high
cost of running their business operations, to close down shop, a situation that has led to a
surge in the number of impoverished or unemployed people. Also, owing to insufficient
refining capacity to cope with the domestic demand, the Nigerian economy heavily relies
on imported petroleum products. The heavy reliance of Nigerian economy on the fossil
fuel market makes it vulnerable to any little instability in global oil market. For example,
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5707
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
following the recent halt in the importation of Nigerian crude oil by the United States of
America due to the shale oil revolution, the Nigerian minister of petroleum had urged the
country to adopt sustainable economic policies, as a matter of urgency, for fear of
impending economic stress that the development might have in the future (TheScoopNG
2014).
Although the usage of fossil fuels products has contributed immensely to the
global economic growth and development, the negative effects of its application in the
area of health and environment are gradually overshadowing the economic benefits,
coupled with the facts that fossil fuels are finite in supply and consequently the prices of
their products are vulnerable to frequent increase. The frequent increase in the price of
fossil fuel products has brought untold hardship to people in developing countries, not the
least in Nigeria. Because of the increase in poverty, most people who could not afford
the expensive fossil fuel products have resorted to the environmentally unfriendly
practice of felling wood for cooking, causing dwindling forest reserves. Besides the
challenge of electricity shortage, Nigeria also faces the problem of an inefficient
management system of wastes, including agricultural, municipal solid waste (MSW), and
sewage, among others. The wastes are generated daily in large quantities but are disposed
in unhygienic and unsustainable ways such as burning, unsanitary land filling, or
indiscriminate dumping of waste on the streets and drains. Landfilling, for example, has
the potential of causing further water and air pollution through leachate and gases, which
are the two main products generated from a landfill. An inefficient waste management
system due to lack of technical expertise, regulatory setup, and adequate funds, has
contributed to various environmental challenges currently being experienced in Nigeria.
Consequently, environmental pollution, flooding, and disease epidemics from
indiscriminate waste dumping on the streets and drains are common occurrences in the
country (Amori et al. 2013; Leton and Omotosho 2004).
Nigeria has an estimated population of over 165 million people and an annual
growth rate of about 2.8% (Factbook 2014; FAOSTAT 2014; Shaaban and Petinrin
2014). The country has a total area of 924,000 km2, out of which 33.0 % is arable land
replanted after each harvest, while 3.1% is cultivated with permanent crops. The tropical
climatic conditions in the country, which are characterized by high humidity in the south
and high temperatures in the north with an average temperature of 27 °C, encourage
large-scale agriculture. Because of the high population, huge amounts of waste are
inevitably produced daily without an effective waste management system, and moreover
more energy is required to satisfy the increasing energy demand. Meanwhile, the
abundant waste generated daily can be utilized as energy resources for provision of
adequate energy for the citizenry by the adoption of biogas technology. The technology
offers numerous benefits, including provision of energy for cooking, heating, lighting,
and as a vehicle fuel, job creation, income revenue generation, reduction of workload or
drudgery for women, agricultural development, and air pollution reduction (Nyns 1986).
Besides, the country also needs an effective waste management system to manage the
huge amount of waste being generated daily in Nigeria. Application of biogas
technology has the potential of maintaining a balance between production and
consumption of waste and energy, since the technology is based on conversion of organic
waste materials into energy in form of biogas. The warm climatic conditions are adequate
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5708
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
for anaerobic digestion process of organic wastes without the need for extra heating.
Channeling wastes into biogas production could therefore be one of the most efficient
ways of waste disposal, energy production, and environmental protection.
The news from Nigerian Finance Minister, on April 2014, that Nigeria is currently
Africa’s largest economy and 26th largest in the World, comes with mixed feelings for
many Nigerians. The positive aspect of the news is that the growth of the economy has
placed Nigeria within reach of its vision 20:2020 to become one of the world’s top 20
economies by the year 2020. And this will definitely increase investment opportunities in
the country. The negative aspect of the news is that the growth impact has not benefitted
poorer members of society, as 60% of the population does not have access to energy and
as such many people have become impoverished. Previous efforts by the government in
finding solutions to the problems by citizenship sensitization of the need to introduce
biofuel energy into the country’s energy mix have not yielded the expected results
because of lack of sustained government effort. Little attention has been paid to the
development of biogas technology in Nigeria, with only few units of biogas pilot plants
developed by different research centres (Sambo 2005). The development and application
of biogas technology have been hampered by a number of factors including storage
difficulty of biomass residues, technical barriers, poor financial support from the
government, and low levels of public awareness of the benefits of using biogas as an
energy source. This study therefore aims at examining current biogas production
potentials of Nigerian biomass resources, and proposing strategies for an accelerated
biogas development in Nigeria.
Potential Assessment of Nigerian Biomass Feedstock for Biogas
Production
Biogas is a colourless and odourless mixture of gases produced through anaerobic
decomposition of organic materials by microorganisms, and depending on the nature of
the organic materials and operating conditions, the gas composition includes methane,
carbon dioxide, nitrogen, oxygen, hydrogen sulphide, and ammonia with compositions of
40-75%, 25-40, 0.5-2.5%, 0.1-1% 0.1-0.5%, and 0.1-0.5%, respectively (Salomon and
Lora 2009; Weiland 2010). Biogas can be used to augment conventional energy sources
for various purposes including cooking, heating, vehicle fuel, and electricity generation,
while the sludge from the anaerobic process can be used as organic fertilizer. Potential
biogas feedstocks that are available in Nigeria include agricultural crop and residues,
livestock wastes, municipal solid wastes and sewage.
Biogas production from agricultural crop wastes
Agricultural crop wastes are potential sources of biogas energy, especially in
Nigerian rural areas where nearly everyone practices farming. Nigeria produces a wide
range of agricultural crops in large quantities for consumption and exportation, and
consequently huge amount of residues are generated from the crops. Agricultural crop
wastes may consist of rotten crops due to inadequate storage facilities. There are infected
crops due to diseases and also residues produced from crop processing after harvest.
Table 1 shows the average quantity of agricultural crop wastes from the production
between year 2003 and 2012 in Nigeria.
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5709
bioresources.com
7.2
3.2
6.1
8.4
Maize
Groundnut
with shell
Millet
Oil palm
fruit
6
10.90
0.40
0.40
0.13
0.08
16.56
1.94
2.16
1.34
7.36
12.20
18.30
2.02
1.03
0.40
1.01
0.25
1
1
0.65
0.419
2.30
0.27
0.30
0.42
2.3
2
3
0.24
0.122
0.047
0.26
Cassava peelings
Cocoa husk
Cocoa pods
Coconut shell
Coconut husk
Maize stalk
Maize cob
Maize husks
Groundnut
husk/shell
Groundnut straw
Millet straw
Millet stalk
Empty fruit bunch
Oil palm fibre
Oil palm shell
Rice husk
(Tonnes x 10 )
2.70
Estimated
Quantity of
Potential
Residues
0.062
Residue
-to Product
Ratio
b-l
(RPR)
Cassava stalk
Residue Type
0.94
5.15
8.54
12.81
1.41
0.72
0.28
0.71
1.89
7.63
0.28
0.28
0.09
0.06
11.59
1.36
1.51
Estimated
Quantity of
Unavailable Crop
residues (70% of
Potential Crop
Residues)
6
(Tonnes x 10 )
0.40
2.21
3.66
5.49
0.61
0.31
0.12
0.30
0.81
3.27
0.12
0.12
0.04
0.02
4.97
0.58
0.65
6
(Tonnes x 10 )
Estimated
quantity of
available crop
residues
0.21
1.13
1.87
2.80
0.31
0.16
0.06
0.15
0.41
1.67
0.06
0.06
0.02
0.01
2.53
0.30
0.33
6 m-o
(Tones x 10 )
Actual vs.
converted to
biogas (51% of
crop residue)
0.14
0.79
1.31
1.96
0.22
0.11
0.04
0.11
0.29
1.17
0.04
0.04
0.01
0.01
1.77
0.21
0.23
BMP of
biogas
produced
based on
3
0.7m /kg VS
at STP
3
9
(m x 10 )
0.16
0.89
1.47
2.21
0.24
0.12
0.05
0.12
0.33
1.32
0.05
0.05
0.02
0.01
2.00
0.23
0.26
BMP of
biogas
produced
based on
3
0.7m /kg
VS at 35 ˚C
3
9
(m x 10 )
b-l
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5710
Rice, paddy 3.9
a
FAOSTAT, 2014;
AIT-EEC 1983; Bernard and Kristoferson 1985; Bhattacharya et al. 1993; Iye and Bilsborrow 2013; Jekayinfa et al. 2012; Jölli and Giljum 2005; Kristoferson and
Bokhalders 1991; Ryan and Openshaw, 1991; Smeets et al., 2004; Webb, 1979a; Webb, 1979b
m-o(Burke 2001; Deublein and Steinhauser 2008; Parkin and Owen 1986)
0.2
0.4
Cocoa
beans
Coconut
(Tonnes x 10 )
43.6
Cassava
6
Average
Production
Agricultural
Crops
Table 1. Biochemical Methane Potential (BMP) of Biogas from Average Crop Production between Year 2003 and 2012
in Nigeria a
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
0.9
8.0
1.2
0.5
3.2
4.3
0.02
1.7
Potatoes
Sorghum
Sugarcane
Soybeans
Sweet
potatoes
Cocoyam
Tobacco
Tomatoes
Potatoes stem
and leaves
Sorghum straw
Sorghum stalk
Sugar cane
bagasse
Sugar cane straw
Soybeans straw
and pods
Sweet potatoes
residues
Cocoyam
residues
Tobacco
stem/stalks
Tomato stem
Tomato leaves
Vegetable
residues
residues
Residue Type
6
0.3
0.3
0.45
2.0
0.36
0.51
0.51
2.43
0.04
1.55
1.15
1.80
1.75
1.5
3.5
0.36
16.00
20.96
0.36
(Tones x 10 )
0.36
Estimated
Quantity of
Potential
Residues
2
2.62
0.3
0.4
Residueto Product
Ratio
b-l
(RPR)
0.03
0.36
0.36
1.09
0.81
1.23
0.25
1.26
0.25
11.20
14.67
Estimated
Quantity of
Unavailable Crop
Residues (70% Of
Potential Crop
Residues)
6
(Tones X 10 )
6
0.01
0.15
0.15
0.47
0.35
0.53
0.11
0.54
0.11
4.80
6.29
(Tones x 10 )
Estimated
Quantity of
Available
Crop
Residues
0.01
0.08
0.08
0.24
0.18
0.27
0.06
0.28
0.06
2.45
3.21
6 m-o
(Tones x 10 )
Actual vs.
Converted to
Biogas (51% of
Crop Residue)
0.0043
0.05
0.05
0.17
0.12
0.19
0.04
0.19
0.04
1.71
2.24
BMP of
Biogas
Produced
Based on
3
0.7m /KgVs
At STP
3
9
(m x 10 )
3
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5711
0.19
0.14
0.21
0.04
0.22
0.04
1.93
2.53
0.17
0.08
0.03
0.29
0.0048
0.06
0.06
9
(m x 10 )
BMP of Biogas
Produced
Based on
3
0.7m /kgVS at
0
35 C
a
Fresh
5.4
vegetables
1.70
0.73
0.37
0.26
Mangoes,
0.8
1.8
1.44
1.01
0.43
0.22
0.15
mangosteens
Melon seed
0.5
residues
1.26
0.63
0.44
0.19
0.10
0.07
Ginger
0.2
Ginger residue
1.15
0.23
0.16
0.07
0.04
0.02
a
FAOSTAT, 2014;
b-l
AIT-EEC 1983; Bernard and Kristoferson 1985; Bhattacharya et al. 1993; Iye and Bilsborrow 2013; Jekayinfa et al. 2012; Jölli and Giljum, 2005; Kristoferson and
Bokhalders 1991; Ryan and Openshaw 1991; Smeets et al., 2004; Webb 1979a; Webb 1979b
m-o
(Burke 2001; Deublein and Steinhauser 2008; Parkin and Owen 1986)
Average
Production
(Tonnes x
6
10
Agricultural
Crops
Table 1. (contʼd). Biochemical Methane Potential (BMP) of Biogas from Average Crop Production between 2003 and 2012 in Nigeria
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
6
Coffee husk
Cowpea husk
residue
residues
residues
residues
residues
residues
residues
Plantain
residues
Cotton stalks
Straw
Yam peelings
0.004
2.7
3.7
0.1
1.0
1.2
0.8
1.1
0.07
2.7
0.45
0.09
1.47
0.32
0.06
1.03
6
(Tonnes x 10 )
0.22
Estimated Quantity
of Unavailable Crop
Residues (70% Of
Potential Crop
Residues)
0.03
0.02
2.35
2.00
0.05
0.14
0.16
0.43
0.59
0.04
0.44
0.10
Estimated
Quantity of
Available
Crop
Residues
6
(tones x 10 )
0.01
0.01
1.20
1.02
0.03
0.07
0.08
0.22
0.30
0.02
0.22
0.05
6 m-o
(Tones x 10 )
Actual vs.
Converted to
Biogas (51% of
Crop Residue)
0.16
0.03
BMP of
Biogas
Produced
Based on
3
0.7m /kgVS
at STP
3
9
(m x 10 )
20.77
0.18
0.04
BMP of
Biogas
Produced
Based on
3
0.7m /kgVS
0
at 35 C
3
9
(m x 10 )
Akinbomi et al. (2014). “Nigerian biogas technology,”
21
2.9
1.8
1.8
0.45
0.45
1.8
1.8
1.9
1.8
0.06
5.48
4.66
0.13
0.32
0.38
1.01
1.39
0.09
3.40
BioResources 9(3), 5707-5737.
0.08
7.83
6.66
0.18
0.45
0.54
1.44
1.98
0.13
4.86
5712
0.59
0.21
0.02
1.02
residues
0.2
2.10
6
(Tonnes x 10 )
Estimated
Quantity of
Potential
Residues
1.46
0.74
0.52
Seed cotton
0.5
3.52
1.76 1.23
0.53
0.27
0.19
Wheat
0.08
1.750
0.14 0.10
0.04
0.02
0.01
Yams
33.7
0.25
8.43 5.90
2.53
1.29
0.90
Total
171.86
120.30
51.56
26.29
18.41
a
FAOSTAT, 2014;
b-l
AIT-EEC 1983; Bernard and Kristoferson 1985; Bhattacharya et al. 1993; Iye and Bilsborrow 2013; Jekayinfa et al. 2012; Jölli and Giljum 2005;
Kristoferson and Bokhalders 1991; Ryan and Openshaw 1991; Smeets et al. 2004; Webb 1979a; Webb 1979b
m-o
(Burke 2001; Deublein and Steinhauser 2008; Parkin and Owen 1986)
Cashew shell
0.7
0.45
Residue
-to Product
Ratio
b-l
(RPR)
0.01
0.01
0.95
0.80
0.02
0.05
0.07
0.17
0.24
0.02
residues
Residue type
0.7
(Tonnes x 10 )
Average
Production
0.01
0.01
0.84
0.71
0.02
0.05
0.06
0.15
0.21
0.01
Chili & pepper
(green)
Cashews with
shells
Carrot and
turnings
Coffee, green
Cowpea, dry
Fruit, Citrus
Kolanuts
Okra
Onions, dry
Pawpaw
Pineapples
Pulses
Plantain
Agricultural
crops
Table 1. (contʼd). Biochemical Methane Potential (BMP) of Biogas from Average Crop Production between 2003 and 2012 in Nigeria
PEER-REVIEWED REVIEW ARTICLE
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
The method used for calculating the average quantity of crop wastes is based on a
residue-to-product ratio (RPR) method in which the RPR for different crops are used to
multiply annual production of each crop. The RPR ratio, which represents the amount of
residues that could be obtained from a unit amount of crop harvested, were selected from
different literature sources, since each source covered only some of the crops studied.
Meanwhile, the available quantities of residues using the RPR ratios might not be the
actual values in practice due to climatic variations coupled with the facts that different
studies indicated varying RPR’s for the same crop; the quantity obtained could still be
used as the best guide for policy makers to get a picture of the amount of residues that
could be generated from each crop, since the RPR ratios made provisions for variations in
crop, variety, climate and different farming activities. VS ratios were taken from
literature sources different from those that the RPR ratios were taken from because
information on VS ratios was not given in the literature containing RPRs ratios.
Biochemical methane potential (BMP) was calculated based on the assumption that the
waste could be taken as primary solids, and that a cubic metre of BMP could be obtained
from one kg VS of the primary solids as given in Khanal (2008). The average quantity of
crop residues obtained annually from the harvesting and processing of the agricultural
crops was estimated to 172 million tonnes.
Meanwhile, about 70% of the residues generated during crop harvesting and
processing are often used for other purposes such as soil mulch, fuel, building materials,
and animal fodder (Dayo 2007; Jibrin et al. 2013). As regards animal fodder, the most
commonly fed crop residues include cassava and yam peels, cowpea husk, and groundnut
husks, brans, oilcakes, maize, millet, and sorghum stovers (DE-Leew 1997; Onwuka et
al. 1997; Singh et al. 2011). Leguminous crop residues are often preferred to cereal
residues as animal fodder because of their higher nutritive value, digestibility, crude
protein content, and minerals (Owen 1994). The quantity of crop residues available for
biogas production could therefore be reduced. In fact, it has been observed that during the
rainy season, agricultural crop residues supply 58% of animal fodder (Jibrin et al. 2013).
Taking the crop residues used for other purposes into consideration, the quantity of
available crop residues for biogas production was estimated at approximately 52 million
tonnes, from which 21 billion cubic metres of methane gas could be generated at 35 oC
(Table 1).
Biogas production from livestock waste: livestock manure and abattoir waste
Livestock waste includes dead livestock due to diseases, livestock manure,
slaughterhouse wastes such as hair, feather, bones, blood, undigested food, and meat
from animal and poultry processing industries. Among the livestock reared in Nigeria,
only cattle, goats, sheep, pigs, and chicken are produced in large quantities, as shown in
Table 2a. The amount of animal dung that could be obtained from the average annual
population of the livestock was estimated to be approximately 32 million tonnes, from
which 3.7 billion cubic metres of methane gas could be produced. However, the available
animal manure for biogas production may in reality be lower, since the considerable
amounts of animal dung is often left on the grazing field to improve the soil quality.
Regarding abattoir waste, a huge amount is usually generated daily in Nigeria
because of high consumption of meat by people. Often these wastes are not treated before
being discharged into nearby streams and rivers, thereby constituting an environmental
and health hazard to the people living in the neighbourhood. Compositions of abattoir
wastes generally include animal blood, intestinal content, waste tissue, and bone. From
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5713
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
the common reared livestock in Nigeria, an estimated amount of 0.83 million tonnes
(Table 2b) abattoir waste could be generated annually, which could be harnessed using
biogas technology to produce about 0.34 billion cubic metres of methane gas.
Biogas production from municipal solid waste (MSW)
The quantity and composition of MSW generated in any particular region depends
most importantly on factors such as people’s lifestyles, standard of living, consumption
patterns, local climate, as well as cultural and educational differences. The waste
generation rate in low-income countries (developing countries) has been found to be
within the range of 0.4 to 0.6 kg/person/day (Blight and Mbande 1996; Chandrappa and
Das 2012; Cointrea 1982). This is similar to the waste generated rate of 0.44 to 0.66
kg/capita/day generated in some urban region in Nigeria (Ogwueleka 2009). The
moisture and organic content of the waste generated in developing countries are
reportedly reasonably high, which makes them to be suitable for anaerobic digestion
(Babayemi and Dauda 2009).
In this study, the average waste generated rate of 0.62 kg/capita/day was used as a
representative value for each person in Nigeria. To estimate the average quantity of MSW
generated in Nigeria, the average waste generated rate of the six Nigerian Geo-Political
zones including North-central, North-East, North-West, South-East, South-West, SouthSouth was obtained from their six respective cities namely, Abuja (0.66 kg/capita/day),
Bauchi (0.86), Kano (0.56), Aba (0.40), Lagos (0.63), and Port-Harcourt (0.6) (all
kg/capita/day) (Adewunmi et al. 2005; Babayemi and Dauda 2009; Ogwueleka 2009;
Usman and Mohammed 2012). An estimated value of 37 million tonnes organic MSW
residues could be available for biogas production with BMP of 13 billion cubic metres
(Table 3).
Biogas production from human wastes
Human waste, often called black water, consists of faeces and urine and forms
part of sewage generated from a community. The other part of the sewage is called grey
water, which represents wastewater from all sources including bathroom, kitchen, and
laundry without human wastes (Katukiza et al. 2012; Uwidia and Ademoroti 2011).
Unlike human wastes, grey water is often highly contaminated with different substances
including domestic wastes such as soaps/detergents, shampoo, pharmaceuticals, and
industrial wastes, which make them unsuitable as feedstock for biogas production without
adequate pre-treatment, as they may cause failure of biogas digesters. Within Nigerian
urban communities, pit latrines are common in low-income households (Chaggu et al.
2002; Howard et al. 2003; Kulabako et al. 2010; WHO and UNICEF 2010), while water
closet toilets are common in middle and high-income households. In Nigerian rural
communities, soil pit and open defection are still the common forms of human waste
disposal, since many rural dwellers do not have any form of toilets (Esrey et al. 1998). Pit
latrines and water closet toilets are usually connected to septic tanks, which collect and
transports human wastes into a soak away pit.
However, most septic and soakaway systems in Nigeria are not properly designed,
located, operated, and maintained with consequent pollution of soil, surface water, and
groundwater. Lack of good sanitation systems for disposing human wastes have been a
major concern to many Nigerians and often facilitate the spread of diseases among
people. Therefore, proper treatment of human waste before disposal is required, and this
could be best achieved by anaerobic digestion.
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5714
bioresources.com
Stocks
(Heads )
Annual Dry
Dung from
Each Animal
p
(Tonnes)
1.0
0.3
0.15
0.15
0.005
16.60
2.04
8.00
5.11
0.90
Total Annual
Animal Dung
6
(Tonnesx10 )
16.60
2.04
8.00
5.11
0.90
32.4
Total Annual
Animal Dung
9
(kg x 10 )
Available Animal
Dung (30% of
Production)
9
(kg x 10 )
4.98
0.61
2.40
1.53
0.27
9.79
Actual vs.
Converted to
Biogas (% of
Production)
48
45
48
48
42
Actual vs.
Converted
to Biogas
9
(kg X10 )
2.39
0.28
1.15
0.73
0.11
BMP of Biogas
Produced Based
3
on 0.7 m / kgVS
3
9
at STP(m x 10 )
1.67
0.20
0.81
0.51
0.08
Average
Weight of
Slaughtered
q
animal (kg)
353
60
23
33
1.7
Amount of Waste from
each slaughtered
Animal (35% of Animal
r
Body Weight) (kg)
123.6
21.0
8.1
11.6
0.6
Actual vs.
Converted to
Biogas (% of the
Manure Production)
51
51
51
51
51
Actual vs.
Converted
to Biogas
9
(kg x10 )
0.158
0.051
0.056
0.082
0.077
BMP of Biogas
Produced Based
3
on 0.7 m /Kgvs
3
9
at STP(m x 10 )
0.111
0.036
0.039
0.057
0.054
Waste
Generated Rate
s
(kg/capita/year)
226.30
375.95
146.00
Population
6
( x10 )
165
165
165
Actual vs. Converted
to Biogas (% of the
Manure Production)
45
50.4
50.4
5715
Actual vs.
Converted to
9
Biogas (kg x10 )
16.80
31.26
12.14
43.40
BioResources 9(3), 5707-5737.
Annual Total
Waste(kg x
9
10 )
37.34
62.03
24.09
86.12
Akinbomi et al. (2014). “Nigerian biogas technology,”
Organic MSW
Urine
Total
Human
Feces
wastes
Total
s
(World-Bank 1992).
Waste
BMP of Biogas Produced
3
Based on 0.7 m /Kgvs at
3
9
STP (m x 10 )
11.76
21.88
8.50
30.38
BMP of Biogas Produced
3
Based on 0.7 m /kg vs. at
0
3
9
35 C (m x 10 )
13.27
24.70
9.59
34.29
BMP of Biogas
Produced Based
3
on 0.7m /kgVS at
0
3
9
35 C (m x 10 )
0.125
0.040
0.044
0.065
0.061
0.335
BMP of Biogas
Produced Based
3
on 0.7 m / kgVS
0
3
9
at 35 C(m x 10 )
1.89
0.22
0.91
0.58
0.09
3.69
Table 3. Biochemical Methane Potential (BMP) of Biogas from Average MSW and Human Wastes Generated in Nigeria
Annual
Slaughtered
Livestock
9
Waste(kg x 10 )
Cattle
2,541,377
0.31
Pigs
4,752,865
0.10
Goats
21,461,956
0.11
Sheep
13,628,125
0.16
Chicken
257,127,778
0.15
Total Waste
0.83
q
(Achoja, 2013; Adeshinwa et al., 2003; Akinfolarin and Okubanjo, 2010; r(World-Bank 1998)
Annual
Slaughtered
Livestock
Table 2b. Methane Potential of Biogas from Average Annual Abattoir Wastes in Nigeria Generated (2003 to 2012)
Cattle
16,571,801
Pigs
6,792,244
Goats
53,027,703
Sheep
34,030,382
Chicken
171,331,000
Total
p
(World-Bank 1977)
Livestock
Table 2a. Methane Potential of Biogas from Average Livestock Population & Manure Production in Nigeria (2003 to 2012)
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
PEER-REVIEWED REVIEW ARTICLE
From the Nigerian population of about 165 million, it is estimated that 86 million
tonnes human wastes (faeces and urine) could be obtained annually from which 128
billion cubic metres of methane gas could be produced (Table 3).
Electricity Production Potential of Nigerian Biomass Feedstock
Various studies have shown the existence of a strong relationship between human
development and annual per capita energy consumption (Meisen and Akin 2008; NBS
2009). This indicates that the level of social development in a country is reflected in the
level of electricity consumption. The potential for electricity energy generation from the
biomass feedstock studied was estimated as 270 TWh for all the available biomass
feedstock (Ostrem 2004), as given in Table 4.
Table 4. Theoretical Electricity Generation from Available Biomass Feedstock in
Nigeria
Biomass Feedstock
Total
Potential
Biomass
Feedstock
6
(tonnes x10 )
Agricultural crop wastes
171.86
Livestock manure
32.40
Livestock abattoir waste
0.83
Organic MSW
33.12
Human waste
86.12
Total
324.33
t
(Ostrem 2004)
Quantity of
Available
Biomass
Feedstock
6
(tonnes x10 )
51.56
9.79
0.83
33.12
86.12
181.42
BMP of biogas
produced based
3
on 0.7 m / kgVS
0
at 35 C
3
9
(m x 10 )
20.77
3.69
0.34
13.27
34.29
72.36
Potential Electricity
Production based
on 3.73
3
kWh/m CH4
9 t
(kWh x10 )
77.47
13.76
1.27
49.50
127.90
270
Electricity production
(Terawatt hour, TWh)
77.47
13.76
1.27
49.50
127.90
270
When considering Nigerian energy needs, average cooking energy demand per
capita per day had been estimated at 0.26 m3 of biogas (Adeoti et al. 2000), which is
equivalent to 0.97 kWh of electricity per capita per day. In other words, annually, each
person will need an average of 354 kWh of electricity, which could be satisfactorily
obtained from the biomass feedstock studied. In fact, the estimated 270 TWh of
electricity energy from all the available biomass feedstock could be used to satisfy the
energy needs of about 763 million people, which are far greater than the Nigerian
population. Large scale electricity generation from biogas powered generator will be a
cheaper, easier, and more affordable source of cooking energy, as it will eliminate
challenges including biogas storage, explosion risks, adaptability of other cooking stoves,
among others, involved in using biogas cooking stoves.
Biogas Energy Market in Nigeria: Current and Future
The current sources of electricity in Nigeria are gas, hydropower, oil, coal with
cooking, lighting, and running of electrical appliances, in line with the domestic activities
that usually consume energy in most Nigerian households. A majority of people living in
rural areas rely mostly on firewood, dried animal dung, crop residues, and charcoal for
cooking because they could not afford the high cost of kerosene and LPG, while
electricity is usually unreliable and inaccessible (IEA 2006). Even in the urban areas
where electricity, LPG, and kerosene are available to many households, the usage of the
energy sources for cooking depends on the household income, with people often giving
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5716
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
preference to low-cost energy source (Arthur et al. 2010; Davis 1998; Howells et al.
2005). Most people with little or no access to electricity rely majorly on fuel wood and
charcoal (Abila 2012), while most low-income households in urban areas often prefer
using charcoal to firewood because of its durability, availability, and less polluting nature
(Sebokah 2009).
Despite the fact that biogas technology is a proven and established technology in
many parts of the World such as Germany, United Kingdom, Switzerland, France,
Austria, Netherlands, Sweden, Denmark, Norway, Republic of Korea, Finland, Republic
of Ireland, Brazil, China, and India (Table 5); the rate of development of biogas
technology in most African countries is still at a low ebb. The rapid development of
biogas technology in most European countries could be linked to various strategies
employed by the respective countries, and most especially by the Renewable Energy
Directive (RES) proposed by the European Union, which sets a binding target for all
Members States to reach a 20% share of renewable energies in the total energy
consumption by 2020. Biogas technologies in Europe, United States, and Latin America
are often on a large scale with biogas produced used for various applications such as
electricity generation, district heating, injection into natural gas pipelines, and as
transportation fuel in buses, cars and trains. However, in Asian and some African
countries, biogas technologies are on a small or household scale with the produced biogas
being used for domestic purposes such as cooking and lighting, among others (Peters and
Thielmann 2008; Sorda et al. 2010).
In Nigeria, some biogas projects have been executed, including construction of
biogas plants at Zaria prison in Kaduna, Ojokoro in Lagos, Mayflower School Ikene in
Ogun State, and a biogas plant at Usman Danfodiyo University in Sokoto with capacity
of the digesters ranges between 10 and 20 m3 (Abubakar 1990; Adeyanju 2008; Atuanya
and Aigbirior 2002; Dangogo and Fernando 1986; Igoni et al. 2008; Ilori et al. 2007;
Lawal et al. 1995; Odeyemi 1983; Ojolo et al. 2007; Sambo 2005). However, the biogas
projects are yet to be commercialized, since most of them are either non-operational or
still at the research stage. The failure of various pilot biogas programmes and a low level
of biogas development and dissemination in Nigeria have been attributed to a number of
factors including lack of policy formulation, ineffective implementation of existing
biofuel policies, lack of government commitment, technical inadequacy (inaccessibility
to spare parts, unskilled operators), ineffective waste management system, poor storage
facility and transportation system, lack of continuity of previous biogas programme
initiatives by the successive governments, inadequate structural facilities, and a low level
of awareness of benefits accrued from biogas technology. The current energy situation in
Nigeria shows that biogas energy is not yet part of Nigeria’s energy mix as the mix is
currently dominated by fuel wood, petroleum products, and hydroelectricity.
Meanwhile, all hope is not lost, as this is a common experience with the
introduction of new technologies, which often require fostering for a period of time
before achieving their stable implementation in terms of ample social, environmental, and
economic benefits. However, lessons should be drawn from the failed biogas projects and
used in the future design and operation of biogas plants. Effort must be geared towards
preventing failure of biogas plants, as this can do a great damage to market penetration of
the technology since prospective users or customers of the technology can lose interest in
making any investment in the technology. Furthermore, strategies that are being
employed in developed countries to advance biogas technology could also be adapted in
Nigeria too.
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5717
bioresources.com
336
1273
1023
10494
BioResources 9(3), 5707-5737.
5718
Strategies that contribute to Germany being the largest biogas producer in Europe include:
(i) setting of target to use 30% renewable energy in electricity production and 14% in heat production by 2020.
(ii) Provision of electricity feed-in tariff (FIT) system that gives primary access to the grid and ensures producers a
premium or tariff for the electricity produced. The FIT system was facilitated by the Gas Entry/Energy Management
Act, which required natural grid operators to connect biomethane suppliers to the grid.
(iv) Setting of Renewable Heat Act which provides financial support for new buildings that use at least 30% biogas
derived heat as their source of heat. This led to an increase in combined heat and power(CHP).
(iii) Exemption of production and use of biogas from Eco-tax.
(iv) Granting of investment subsidies and favourable loans to investor in biogas applications.
Strategies include:
(i) Provision of FIT for electricity generation from August 2011. The FIT includes: € 0.17 for up to 250kW, 0.13
€ 0.16 for >251 kWh up to 500kWh and € 0.11 for >500kWh.
(ii) Awarding of double Renewable Obligation Certificates (ROCS) plants involved in anaerobic digestion (AD).
(iii) Provision of renewable heat incentive in form of tariff of €0.08 for biomethane injected into the natural gas grid
and combusted downstream from April 2011.
(iv) Provision of Renewable Transport Fuel Obligation Certificates worth €0.13 in 2010.
Strategies include:
(i) Provision of FIT system for electricity.
(ii) Fund provision from Swiss Gas Association, a voluntary support program, for biomethane injection in order to
achieve the set target of injecting 300 GWh biomethane annually within 6 years.
(iii) Financial support for projects on reduction of greenhouse gas emissions.
Strategies include:
(i) Provision of FIT system for electricity produced from biogas with energy efficiency bonus and manure bonus
included. The FIT incentives include 0.8580 to 0.14521 EUR/kWhe for landfills, 0.1182 to 0.2110 EUR/kWhe for AD
plants, 45 to 95 EUR/MWh for biomethane from landfills, 69 to 125 EUR/MWh for upgrading the biogas to
biomethane from AD plants.
Strategies
Akinbomi et al. (2014). “Nigerian biogas technology,”
France
4
600
610
IEA Bioenergy (2014)
Switzerland
3
t
United
Kingdom
Energy
Number of
production
biogas
plants at the (GWh/
end of 2013 year
(A) Medium to Large scale Biogas Plant
Germany
9945
40970
Table 5. Comparison of Biogas Production Strategies in Different Countries
2
1
S/N
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
Norway
Republic of
Korea
Finland
9
10
11
73
78
129
167
252
242
569
1925
500
1218
n.a
1589
BioResources 9(3), 5707-5737.
5719
Strategies include:
(i) Provision of FIT system which is supported by Green Electricity Law. The incentives include 0.1950 EUR/kWh
up to 250 kWhe, 0.1693 EUR/kWh from 250 - 500 kWhe, 0.1334 EUR/kWh from 500 - 750 kWhe, 0.1293EUR /kWh
for higher than 750 kW + 0.02 EUR/kWh if biogas is upgraded + 0.02 EUR/kWh if heat is used efficiently.
There is a support scheme that favours large scale biogas applications.
Although Sweden has no feed-in tariff system, other support systems exist which include target setting of zero
emission of greenhouse gases by 2050, provision of economic incentives including tax-free policy on emission of
carbon dioxide, nitrous oxide and sulphur taxes during biogas production, green certificate system, free parking
charge for biogas fueled vehicles and introduction of climate investment programme.
The strategies employed include a bottom-up approach, access to investment grants, implementation of energy
taxes and introduction of various financial incentives for both upgraded biogas supplied to the natural gas grid and
u
to purified biogas entering a town gas grid
Strategies include
(i) Banning of landfilling biodegradables of since 2009 which led to increase in available biogas feedstock.
(ii) Provision of delivery support system that gives 3.5 EUR per ton of manure delivered to biogas plants.
(iii) Provision of biogas investment aid.
(iv) Tax-exemption and investment aid for infrastructure related on biogas fueled vehicles.
Although there are no tariffs or subsidies for biogas, there is Renewable Portfolio Standard (RPS) system
implemented since 2012 which mandates 2% of the total power generation to be supplied using the appropriate
kind of renewable energy. Moreover, 10% Value Added Tax (VAT) and 2% tariffs are charged when the mixture of
CNG and biogas is sold.
Strategies include:
(i) Establishment of Electric Market Authority to support new biogas plants, which produce more than 100 kVA, with
a feed-in tariff which guarantees a minimum price of 83.50 EUR/MWh electricity and 50 EUR/MWh heat premium
on top of basic subsidy if the generated heat is utilized, provided that the total efficiency is at least 50%.
(ii) Financial support by the Ministry of Agriculture and Forestry for biogas plants built on farms aiming at producing
their own energy and heat.
(iii) Exemption of production and use of biogas from excise tax.
Strategies
Akinbomi et al. (2014). “Nigerian biogas technology,”
Raven and Gregersen (2007)
Denmark
8
u
Netherland
Sweden
Number of
Energy
biogas
production
plants at the (GWh/
end 2013
year
(A) Medium to Large scale Biogas Plant
Austria
336
585
Table 5. (contʼd). Comparison of Biogas Production Strategies in Different Countries
6
7
5
S/N
PEER-REVIEWED REVIEW ARTICLE
14
15
16
17
18
19
20
21
13
bioresources.com
22
697
v
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5720
(B) Small-scale or household digesters
Number of small Strategies
scale digesters
as at 2007
w,x
China
26.5 million )
In most Asian countries, biogas programs developed rapidly because of significant financial and technical support provided
y,z
by their governments and various aid agencies . In Nepal, the support is through various policy instruments such as
India
4 million
aa
biogas
support
programme
(BSP)
which
is
technologically
standardized and free from political interference. Demand for
Nepal
268,464
biogas digesters was stimulated through subsidy and quality control mechanisms. When government financial support
Vietnam
152,349
declined, Nepal generated financial support by developing biogas plants across the country as clean development
Bangladesh 26,311
mechanism (CDM) projects, and all these strategies have contributed to Nepalʼs achievement of having the highest per
Cambodia
19,173
v,ab,ac
capita biogas plant in the World
. Bangladeshi government has been actively involved in the development of biogas
Indonesia
7835
x
technology
in
the
country.
In
China
and India, drivers to rapid biogas development include strong government support,
Pakistan
5357
x
technical knowledge, availability of fermentation materials ,
Chinaʼs principle of adaptability of materials of construction of biogas plants to locality contributed the rapid development of
biogas technology in the country
v
w
x
y
z
Surendra et al. (2014); Chen et al. (2010); Bond and Templeton (2011); Kristoferson and Bokhalders (1991); Gunnerson and Stuckey (1986);
aa
ab
ac
Amigun and Blottnitz (2010); Desai (1992); BSP (2012)
Brazil
Strategies include:
(i) Implementation of landfill levy of 75 EUR/ton of waste landfilled to discourage landfilling activities.
(ii) Requirement of population centres with an excess of 25 000 persons to provide collection of source segregated
food waste in order to encourage digestion of organic fraction of municipal solid waste.
(iii) There is FIT system which includes 0.15 EUR/kWhe for AD CHP equal to or less than 500 kW; 0.13EUR/kWhe
for AD CHP greater than 500 kW; 0.11 EUR/kWhe for AD (non CHP) equal to or less than 500 kW and 0.10
EUR/kWhe for AD (non CHP) greater than 500 kW.
Strategies include:
(i) Provision of credit to producers of biogas technologies.
(ii) Provision of rural technical assistance to promote improvement of infrastructure associated with biogas
technology.
Strategies
Table 5. (contʼd). Comparison of Biogas Production Strategies in Different Countries
Number of
Energy
biogas
production
plants at the (GWh/
end 2013
year
(A) Medium to Large scale Biogas Plant
12
Republic of 30
n.a
Ireland
S/N
PEER-REVIEWED REVIEW ARTICLE
22
23
24
25
26
27
28
29
bioresources.com
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5721
(B) Small-scale or household digesters
Number of
Strategies
small scale
digesters as at
2007
Kenya
6749
Biogas technology in most African countries has recorded little success due to less availability of technical and operational
x
support among other factors. Only few of the installed biogas plants are still operational . Few operational biogas plants
Ethiopia
5011
often experience numerous downtimes
Tanzania
4980
Uganda
3083
Burkina Faso
2013
Senegal
334
Cameroon
159
Benin
42
v
Surendra et al. (2014)
v
Table 5. (contʼd). Comparison of Biogas Production Strategies in Different Countries
PEER-REVIEWED REVIEW ARTICLE
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
Obviously, adequate preparation is needed for the pre-design, design, operation
and post–design of the biogas plants in order to accelerate the development of biogas
technology in Nigeria.
Appropriateness of Digesters for Biogas Production in Nigeria
Digester design is an important factor in the sustainability of biogas technology.
Although a digester can be adapted to suit a purpose different from that for which it was
made, for effective performance the type of digesters to be selected depends on several
factors including the feedstock type and availability, purpose, operational factors, scale,
bacterial growth system, temperature, and population, among others. Table 6 shows
examples of digesters commonly used in different applications of biogas technology.
Digesters in most developed countries are usually medium to large digesters, while
digesters in most developing countries are mostly household or small-scale digesters.
There seems to be, therefore, a correlation between the scale of digester and biogas
utilization; with gas utilization in most developing countries specifically for cooking and
lighting, while gas utilization in most developed countries is for large scale electricity
generation, heat, and vehicle fuels. The three common types of digesters used in most
developing countries include Chinese fixed dome digester, Indian floating drum digester,
and flexible balloon digester. Of these, the floating drum and fixed dome digesters
installations are more robust and expensive than flexible balloon installations, which are
cheap but subject to damage. Often, a trade-off needs to be made between choosing
between expensive but robust, and cheap but non-durable designs.
In Nigeria, digester suitability could be based on feedstock type and availability,
geopolitical zones, population, and climatic vulnerability (i.e. rainfall decline, coastal
flooding, and erosion). The six geopolitical zones and the year 2011 zone-based
population percentage of the states in Nigerian are shown in Fig. 1. According to
feedstock type and availability, Table 7 indicates the potential agricultural feedstock that
could be used for large-scale biogas in the six zones. In the North West, the major
agricultural crops that could generate large quantity of residues for large-scale production
of biogas include guinea corn, maize, millet, beans, rice, cotton and groundnut, cassava,
and yam. According to the climatic vulnerability (Table 8), the zone is extremely
vulnerable, so adequate storage facility is needed to ensure continuous supply of the
feedstock, though there is significant irrigation system spread across the zone. For
effective costs, time, and labour management, a very large biogas plant dedicated to
electricity provision for the whole zone can be located at Gombe, which is a state at the
centre of the North East zone. Moreover, since Northern Nigeria is notable for
commercial livestock farming, residues from the major crops stated above and livestock
manure could be co-digested in the proposed biogas plant. Meanwhile, the availability of
livestock manure for biogas production will depend on government support for provision
of ranches to prevent nomadic farming, which is the common livestock farming system in
the northern Nigeria. Besides the proposed large biogas plant, each state in the region
could also support installation of household and community biogas plants that could use
municipal wastes, sewage, and household wastes as feedstock.
The major agricultural crops that could be produced in the other five zones and
from which large quantities of residues could be generated are also given in Table 8.
There is no significant difference in terms of the available feedstock for biogas
production in the different zones in Northern Nigeria. There is, however, significant
difference between the feedstock available in the Southern and Northern zones.
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5722
1
S/N
ai
Ferrer et al. (2011)
Plug flow
ai
digester
11-13%
5723
(i) It does not require incorporated mixer as the inputs move in a slug
(ii) Capital and operating costs are not extremely high
(iii) The digester requires occasional cleaning due to down times
(iv) The digester prefers non-fibrous feedstock such as animal excrements.
(i) It is good for co-digestion of substrates with easy and difficult digestible
components.
(ii) it does not require much control.
(iii) There is often a volume variation of the culture media which can affect its
productivity.
(i) It is suitable for households where there is daily routine of activities.
(ii) The gas output is constant.
(iii) It often requires that substrate be fluid and homogeneous especially if it is
automatic.
(i) It has low capital cost.
(ii) It is designed for emission and odour control.
(iii) it is not designed to optimize biogas production.
(iv) it has a poor bacteria to substrate contact leading to long retention time
and low digestion time.
(i) It can accommodate wide range of solids.
(ii) There is high potential of optimized biogas production.
(iii) It has high capital and operational cost.
(i) It is easy and does not require daily routine of feeding.
(ii) The major demerit is the unsteady nature of the gas-output.
BioResources 9(3), 5707-5737.
It is usually cylindrical with
low height to diameter ratio
with an incorporated mixer
to ensure thorough mixing.
It is a linearly arranged
reactors with inputs
entering from one end, and
effluents exiting on the
other end with a retention
time between 20 to 30
days.
It is a covered pond where
anaerobic digestion of
organic material takes
place.
It is fed and emptied often
automatically at regular
intervals.
It is filled completely and
emptied completely after a
fixed retention time.
A portion of the culture
media is withdrawn at
intervals and fresh medium
is added to the system.
Merits and demerits
bioresources.com
Description
Akinbomi et al. (2014). “Nigerian biogas technology,”
Tomori (2012);
ah
Continuous
stirred tank
reactor
2-10%
Continuous
mode
digester
Continuous
Anaerobic
lagoon
digester
Semicontinuous
mode
digester
Semicontinuous
< 2%
Batch mode
digester
Batch
Mode of
operation
Total Solids
Digester
Basis of Classification
Table 6. Classification of Digestersah
PEER-REVIEWED REVIEW ARTICLE
Thermophilic
digester
50-60 C
aj
Mesophilic
digester
30-35 C
ak
It is a digester which is
operated at a low
temperature range of 30o
35 C.
It is a digester which is
operated at a low
temperature range of 50o
60 C.
This is a digester with a
single reactor where, all
anaerobic processes take
place.
This digester consists of two
or more reactors where
different anaerobic processes
take place.
It is a digester which is
operated at a low
o
temperature range of 5-20 C.
The digester has a fixed, nonmovable gasholder that sits
on top of the digester.
Description
The digester has a movable
gasholder that floats either
directly on the slurry or in a
water jacket of its own.
BioResources 9(3), 5707-5737.
5724
(i) It is often used for large scale operation.
(ii) Biogas production is faster than other temperature range.
(iii) It is expensive to operate due to high heat energy requirement.
(iv) Bacteria operating at this temperature range are few and sensitive to
temperature fluctuations.
(i) It is more efficient than single stage system since it allows specialization of
acid and methane producing bacteria.
(ii) Biogas production is optimized.
(iii) It is more expensive to run than single stage system.
(I) It does not require extra energy for heating as the operating temperature is
often the temperature of the digester environment.
(ii) It is often used for small scale operation.
(iii) The solid retention time (SRT) is over 100 days.
(i) Most existing anaerobic bacteria are at this temperature range.
(ii) It is less expensive to operate than thermophilic digester.
Merits and demerits
(i) It has the advantage of constant gas pressure as this depends on the
weight of gasholder.
(ii) Its construction is easy.
(iii) Material costs of the steel gas holder are high.
(iv) It has a short life span since the steel parts are susceptible to corrosion.
(i) It has low construction costs.
(ii) It has a long life span if it is well-constructed since it is often constructed
using non-rusting steel parts.
(iii)The gas is often ineffectively utilized as the gas pressure fluctuates
significantly.
(iv) If the digester is not gas-tight, gas leakage may occur.
(i) It is less expensive to operate than multiple stage digester.
(ii) It is subjected to frequent disruptions due to occurrence of many reactions
in the same reactor.
bioresources.com
Singh and Sooch (2004) ; Santerre and Smith (1982)
o
o
Psychrophilic
digester
Multiple
stage
digester
One-stage
digester
5-20 C
o
Onestage
digestion
process
Multiple
stage
processes
Fixed-dome
ak
digester
Digester
Floating drum
aj
digester
Akinbomi et al. (2014). “Nigerian biogas technology,”
Tomori (2012);
Temperature
4
ah
Stages of
anaerobic
digestion
process
Fixed gas
holder
Basis of Classification
Gas
holder Floating
structure
Gasholder
3
S/N
2
Table 6. (Contʼd). Classification of Digestersah
PEER-REVIEWED REVIEW ARTICLE
Large scale
Small or
medium
scale
Family scale or
household
biogas plants
that cannot
produce more
than 100 kW
Farm-scale
biogas plants
that can
produce
between 100
and 500 kW
Centralized codigestion
biogas plants
that can
produce more
than 500 kW
Fixed film
growth digester
Digester
Suspended
growth digester
5725
(i) The digester is more profitable to generate higher agricultural,
environmental and economic benefits for the society due to its
economies of scale, higher capacity utilization and adequate
professional management.
(ii) This type of digesters is common in most developed countries
including Denmark, USA, Germany, France, Sweden, among others.
(iii) The demerit of large scale biogas plant is that if there is biological
process inhibition there will be a total breakdown in the gas production
for a period of time, since there is only one large digester. Furthermore,
the installation cost is higher than small scale digester
(iv) High investment costs are often required because of the complex
structure which include large reactor volumes, additional components
including pumps, temperature regulators, pre-storage tank among
others, included in their designs
The digester is designed for codigestion of many suitable
feedstocks, and it is often centrally
located to reduce costs, time and
manpower required to transport
feedstock and digestate to and from
the digester, respectively.
BioResources 9(3), 5707-5737.
(i) The digester can be simple or complex with various sizes, designs
and technologies.
(ii) Many farm scale biogas plants are operated in European countries
such as Germany, France, Austria, Denmark, and Sweden, among
others.
It is designed to produce biogas at
the community level for electricity,
heat production and gas engine fuel.
The feedstock is often from one or
two neighbouring farms.
(i) It is simple, cheap, robust and easy to operate and maintain, and can
be constructed with local materials.
(ii) Millions of family scale digesters are operated in countries like China,
India or Nepal.
(i) It allows long SRTs and short hydraulic retention times (HRTs).
(ii) Biogas production is optimized.
Merits and demerits
There is high potential of wash out of bacteria if the system is used for
continuous operation.
bioresources.com
Description
In this digester, the bacteria are
suspended and distributed
throughout the digester.
In this system, the bacteria are held
by a media (e.g. membrane) for a
long period.
The digester is designed to produce
biogas for family cooking and lighting
needs. The feedstock for the
digester is often from the household
or family small farms.
Akinbomi et al. (2014). “Nigerian biogas technology,”
Tomori (2012)
ah
Scale
6
Very small
scale
Basis of Classification
Bacterial
Suspended
growth
bacteria
system
Attached
bacteria
S/N
5
Table 6. (Contʼd). Classification of Digestersah
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5726
Fig. 1. Geopolitical zones and the zone-based population percentage of the states in Nigeria
PEER-REVIEWED REVIEW ARTICLE
Total
Total
North
Central
Total
North
West
Geopolitical
zones
North
East
bioresources.com
33.15
428.48
176.23
110.24
115.06
286.66
1,149.82
204.30
115.00
408.45
278.58
99.70
218.37
254.96
1579.36
31.02
340.78
54.27
12.70
78.53
91.29
4.40
612.99
10,168.22
Beans
2,187.96
18.38
14.19
13.94
2,234.47
2,535.64
43.04
3.71
4.16
54.77
2,641.32
2,879.39
999.76
3,791.45
1,012.16
294.54
1,157.30
33.62
Cassava
109.23
1,917
231.54
478.14
BioResources 9(3), 5707-5737.
26.37
5727
1,405.35
1,147.82
1,396.8
Major Agricultural crops production in 2010 (production in thousand metric tons)
Cocoyam Cotton
Maize
Melon
Millet
Rice
Groundnut Guinea
corn
492.51
184.20
401.99
197.86
269.05
10.36
740.24
398.08
293.42
357.79
720.40
8.13
0.61
267.31
330.96
54.03
262.38
571.46
92.64
188.85
7.12
187.86
121.86
192.36
7.38
58.02
253.84
0.69
250.18
105.08
62.92
280.68
3.23
22.52
0.70
473.04
84.76
109.31
288.95
15.51
164.86 1,965.27
1.39
1,643.58
1,127.14
112.12
2,322.90
10.75
24.41
1,006.06
485.38
732.42
297.51
603.95
2.33
17.98
43.86
288.66
69.64
86.77
256.06
155.19 77.40
390.81
41.56
195.57
702.13
3.67
21.02
18.03
714.16
71.66
154.60
270.60
0.94
59.62
140.55
323.11
422.05
526.94
745.72
20.90
11.09
535.33
68.66
82.09
200.45
128.92 324.54
309.26
36.47
183.42
608.42
14.02
410.69 1,624.52
18.03
3,046.71
1,442.46
1,526.9
3,387.33
6.36
0.23
261.06
3.15
20.54
104.02
28.00
54.28
26.14
628.85
177.99 199.17
636.67
546.62
677.91
170.33
0.40
70.96
341.48
309.00
171.86
164.03
8.77
14.51
78.47
51.39
45.97
102.87
477.78
117.49
124.74
67.76
317.51
175.34
36.22
43.45
112.79
127.57
123.40
39.61
5.01
12.02
7.18
17.48
5.87
Akinbomi et al. (2014). “Nigerian biogas technology,”
NBS (2012).
ad
Kogi
Niger
Benue
Kwara
Plateau
Nasarawa
FCT,
Abuja
Kaduna
Kebbi
Zamfara
Sokoto
Kano
Jigawa
Katsina
Taraba
Borno
Bauchi
Adamawa
Gombe
Yobe
Member
States
Table 7. Geopolitical Zones and their Major Agricultural Crops Production in 2010ad
PEER-REVIEWED REVIEW ARTICLE
141
1.96
10.78
12.74
75.97
10.31
3.37
31.46
10.20
131.31
119.24
17.65
1.62
2.49
Soya bean
8,391.84
2,854.95
2,85495
2,408.72
1.06
2,409.78
1,203.31
3,914.17
522.61
608.28
2,143.47
213.96
Yam
Total
Total
South
East
0.78
1.95
0.78
1.22
1.34
3.34
Ebonyi
Enugu
Imo
Abia
Anambra
102.25
99.09
730.21
247.80
228.98
142.61
142.38
137.80
899.57
-
-
-
23.51
4.57
102.14
12.09
3.96
58.40
20.86
6.04
101.35
47.03
4.59
BioResources 9(3), 5707-5737.
203.94
56.10
700.89
93.48
97.34
87.98
73.84
65.93
418.57
31.36
151.69
181.31
5728
330.96
-
330.96
-
-
1,810.60
2,004.19
10,035.10
1,058.59
2,831.63
2,181.93
415.74
1,786.72
8,274.61
48.38
146.19
141.87
13.74
22.02
17.13
52.89
22.44
1.17
-
431.14
134.78
43.20
193.36
115.13
132.99
1050.60
76.49
33.04
504.43
3,302.47
AkwaIbom
Bayelsa
Edo
CrossRiver
Delta
Rivers
0.12
0.17
0.17
0.06
0.52
-
20.02
0.14
0.63
20.79
-
2,920.01
1,519.79
322.58
2,205.09
834.34
1,377.65
9,179.46
2,380.37
Oyo
Ogun
Lagos
Ondo
Osun
Ekiti
107.11
124.70
605.71
189.34
161.69
1188.55
192.43
Major Agricultural crops production in 2010 (production in thousand metric tons)
Cassava
Beans Cocoyam
Cotton
Maize
Melon
Millet
Member
States
Akinbomi et al. (2014). “Nigerian biogas technology,”
Geopolitical
zones
South
West
Total
South
South
bioresources.com
75.84
334.85
46.40
4.21
385.46
54.03
18.62
3.19
0.29
85.73
86.02
-
Rice
0.35
1.8
6.79
2.43
0.17
9.39
0.06
1.39
-
0.49
0.72
1.21
-
Groundnut
Table 7. (Contʼd). Geopolitical Zones and their Major Agricultural Crops Production in 2010ad
PEER-REVIEWED REVIEW ARTICLE
0.26
0.57
0.65
0.08
7.82
8.55
0.31
-
Guinea
corn
24.44
0.38
24.82
-
-
-
Soya
bean
-
1,639.52
960.63
6520.05
1,039.93
2,319.05
994.87
798.69
1,417.43
6,569.97
233.26
563.56
2,387.28
2,507.71
284.66
17.24
2,278.69
428.12
1,685.09
7,201.51
735.80
Yam
bioresources.com
State
%
Kaduna
16.9
Kebbi
9.1
Zamfara
9.2
Sokoto
10.3
Kano
26.4
Jigawa
12.0
Katsina
16.1
Mean maximum
temperature ranged
from 31.1 to 42.6
˚C while monthly
minimum
temperature ranged
from 11.1 to19.2 ˚C
Total rainfall varied
from 2000-3000mm
Guinea corn, millet,
Maize, beans,
groundnut, rice,
cotton, soya beans,
and commercial
livestock wastes
State
Taraba
Borno
Bauchi
Adamawa
Gombe
Yobe
%
11.9
22.1
24.7
16.5
12.4
12.4
North West
ϰϭ͕ϴϮϲ͕ϰϯϬ
North East
ϮϮ͕ϮϳϬ͕ϭϮϬ
North Central
Ϯϰ͕Ϯϭϯ͕ϲϱϰ
ϯϮ͕ϰϴϯ͕ϯϭϬ
South West
Ϯϰ͕ϱϲϴ͕ϲϴϳ
South South
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5729
State
%
State
%
State
%
Kogi
15.9
Oyo
20.3
Akwa-Ibom
18.8
Niger
19.3
Ogun
13.6
Bayelsa
8.0
Benue
20.4
Lagos
32.8
Edo
15.0
Kwara
11.3
Ondo
12.4
Cross-River 13.6
Plateau
15.1
Osun
12.3
Delta
19.6
Nasarawa
8.9
Ekiti
8.6
Rivers
25.0
FCT, Abuja 9.1
Temperature Mean maximum
Mean maximum
Climatic
Mean maximum
Mean maximum
af
temperature ranged
ranged between
condition
ranged between
ranged between
from 31.1 to 42.6
31.1- 42.6 ˚C
31.1- 42.6 ˚C
31.1- 42.6 ˚C
˚C while monthly
temperature while
temperature while
temperature while
minimum
monthly minimum
monthly minimum
monthly minimum
temperature ranged
temperature ranged temperature ranged
temperature ranged
from 11.1 to19.2 ˚C
from 11.1-19.2 ˚C
from 20.0-24.1 ˚C
from 20.0-24.1 ˚C
Rainfall
Total rainfall varied
Total rainfall varied Total rainfall varied
Total rainfall varied
from 300-1000mm
from 2000-3000mm from 2000-3000mm
from 2000-3000mm
Cassava, yam,
Cassava, yam,
Yam, melon,
Feedstock in high quantity
Guinea corn,
cocoyam, maize,
melon, cocoyam,
cassava, maize,
for biogas production
maize, millet,
melon, rice and
maize, rice and
groundnuts, soya
beans, rice, cotton
and groundnut.
domestic livestock
domestic livestock
bean, rice, beans,
wastes
wastes
guinea corn and
cassava, yam and
commercial
commercial
livestock wastes
livestock wastes
ag
Climate vulnerability Index
1
3
4
6
5
ae
af
ag
:NBS (2012); Nigeria Climate Review Bulletin (2010); Federal Ministry of Environment (Special Climate Unit) (2010)
1,2,3,4,5,6 – degree of relative climate vulnerability( 1: extremely vulnerable; 6: least vulnerable)
Geopolitical Zones
Projected Population in 2011
Member states and their
zone-based population
ae
percentage
Table 8. Geopolitical Zones, Climatic Patterns, and Vulnerability of Major Agricultural Crops Production in 2010ad
PEER-REVIEWED REVIEW ARTICLE
%
13.2
20.0
24.3
17.2
25.3
2
Mean maximum
ranged between
31.1- 42.6 ˚C
temperature while
monthly minimum
temperature ranged
from 20.0-24.1 ˚C
Total rainfall varied
from 2000-3000mm
Cassava, Yam, rice,
cocoyam, maize,
melon and domestic
livestock wastes
State
Ebonyi
Enugu
Imo
Abia
Anambra
ϭϴ͕ϵϯϮ͕ϯϭϱ
South East
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
While cereals and legumes are produced in large quantity in the Northern zones,
root and tuber crops such as yam, cocoyam, and cassava are produced in large quantity in
the Southern zones. The variation is due to the climatic variations in the different zones.
The central states for which the proposed large scale biogas plants could be located in the
North West, North Central, South West, South East, and South South are Katsina, Abuja,
Oyo, Rivers, and Anambra. The areas chosen for the potential large scale biogas plants
should be protected from flooding and other destructive events. Furthermore, livestock
farming in all the southern zones are mainly domestic, unlike in the North where it is
practiced on a large scale, so the livestock manure may be used for household or
community biogas digesters. The digesters could also use sewage, household and
municipal wastes as feedstock to biogas production for domestic purposes.
Biogas Technology Development and Dissemination Strategies
Development of a viable biogas market is a prerequisite to attracting biogas
investors. For a biogas market to be viable, people or potential biogas users must be
informed of the benefits accrued unto them by using biogas as energy source. The
following strategies are necessary for development of viable biogas market.
The need for government intervention
The role of government in stimulating the market penetration of biogas
technology cannot be overrated, and thus, for easy penetration of biogas energy into
energy market, the government needs to play an active role in ensuring that the biogas
energy is sufficient, efficient, affordable, steady, and dependable (Winkler et al. 2011).
Government interventions through subsidy provisions and tax holidays are needed to
reduce the initial cost of investing in biogas technology. Uninterrupted development of
biogas technology and dissemination requires unwavering and long-term government
support in many areas, including financial support, legislative support, and technical
support. The high level of biogas technology in most developed countries as discussed in
Table 5 has been attributed to favourable policy formulation and implementation (Palvas
et al. 2010; Stehlik 2010). It is therefore obvious that government support and
development of biogas technology are inseparable. Government has an important role to
play in the creation of an enabling environment for private sector participation in biogas
technology in such a way that the produced biogas will be affordable to meet energy
needs of the citizenry. At the present oil price, the initial capital cost of biogas production
investment is higher than that of fossil fuel products, which tends to make fossil fuel
products more affordable than biogas. However, the biogas sector can be made more
affordable than any other energy sources if appropriate government measures are put in
place. Favourable policies including a ban on landfilling of organic waste, setting of a
target for inclusion of a specific percentage of biogas energy in the Nigerian energy mix,
financial remuneration in form of feed-in system incentives, tax exemption/holiday,
tipping fees on treated wastes, green certificates, affordable connection fees to the biogas
based grid system, low biogas price, subsidy, among others, should be promulgated and
implemented so that demand for biogas as a product would be encouraged. Markets for
the two main products from anaerobic digesters including biogas and organic fertilizer,
should be developed so that the biogas technology would be financially profitable and
economically attractive to would-be investors. The economic viability of biogas
technology will depend on income generation ability of its two products including biogas
for cooking, lighting and power and digestate for organic fertilizer, or for fishpond or
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5730
bioresources.com
PEER-REVIEWED REVIEW ARTICLE
animal feed. Nigeria has about 36 states, a federal capital territory, and 760 local
governments with 12, 19, and 5 states having between 6-17, 18-29, and 30-44 local
governments respectively. In order to effectively accelerate the development and
dissemination of biogas technology, government at all levels including all the three tiers
of Nigerian government namely, federal, state, and local governments should bear the
bulk of responsibility involved in the development and dissemination of biogas
technology.
Increased awareness level and capacity building development
The level of awareness of the benefits of biogas technology needs to be raised, as
many people are not acquainted with benefits associated with biogas technology. In the
rural area, for example, some people still have the notion that food cooked using fuel
wood tastes better than food cooked using other energy sources. There should also be a
feedback mechanism whereby biogas credibility as perceived by users could be easily
communicated. This speeds up dissemination of the technology. This could be achieved
by setting up a monitoring system based on cell phone technology, since most people
both in urban and rural areas now use cell phones. Furthermore, many people lack the
technical know-how in operating and maintaining biogas plants. People should be trained
to construct, operate, and maintain biogas plants for efficient and optimum production. It
would also be wise to make use of locally available materials in Nigeria for biogas
projects in order to reduce the difficulty involved in getting spare parts of plants and
thereby ensure the sustainability of the biogas programme. Capacity building through
technical training to enhance local capability in the operation and maintenance of biogas
plants could be achieved through the establishment of biogas research institutes or
consultancy centres where biogas operators and users can find answers to their various
questions and most importantly obtain any urgent assistance they may need from
seasoned biogas experts and consultants at the centres..
Regulatory mechanism for biogas market in Nigeria
In 2007, the Nigerian government, upon realizing the urgent need to incorporate
biogas energy into her national energy mix, set up a national biofuels initiative under the
Renewable Energy Division of Nigerian National Petroleum Corporation (NNPC), to
coordinate the development of biofuel technology in the country. A ministry such as
NNPC should therefore be organized and equipped to fill in the potential supply-demand
gap by buying back excess biogas energy or supplying biogas energy in case of
inadequacy as depicted in Fig. 2.
NNPC
Biogas feedstock
Provider
Biogas consumers
Biogas producer
Fig. 2. Proposed biogas market regulatory mechanism in Nigeria
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5731
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
This is necessary since one of the main challenges to biogas dissemination in
most countries where biogas technology is well developed is the cost of fuelling
appliances using biogas as compared to fuelling it using other conventional energy
sources such as petroleum products. Customer’s demand of biogas as a commodity or
service that provides utility in terms of energy will depend on their willingness, attitude,
and ability to pay for the commodity and the satisfaction derived from its usage. In other
words, effective demand of biogas technology as a commodity will depend on factors that
include its cost effectiveness, appropriateness, availability, reliability, efficiency, and
technical potential. All of these factors need to be put into consideration for successful
development and dissemination of biogas technology in Nigeria.
CONCLUSIONS
Assessment of different types of wastes in this study has shown that there is huge
potential of biomass feedstock for commercial biogas production in Nigeria. The
available Nigerian biomass feedstock includes agricultural crop wastes, livestock manure,
abattoir waste, organic MSW, and human waste with potential quantity of 52, 10, 0.8, 33,
and 86 million tonnes respectively. Total biochemical potential of biogas that could be
generated from the biomass feedstock is 72 billion cubic metres, from which 270 TWh of
electricity could be generated, which is enough to satisfy the annual electricity need of
the Nigerian population. Furthermore, according to the geopolitical zoning of biomass
and manure, northern zones have the potential for high production of cereal and legumes
crop residues, while the southern zones have the potential for high production of root and
tuber crops. It could be a wise idea to have a large centrally located biogas plants in each
geopolitical zone that could be used to generate electrical energy to power each zone.
Presently, biogas energy has not been incorporated into the Nigerian energy mix
since the current level of biogas technology in Nigeria is very low. Most of the few
existing pilot scale digesters are currently non-operational, while the few biogas plants
that are operational have frequent downtimes. The problem has been attributed to
technical, economic, and social impediments including poor digester designs,
management, maintenance, planning, monitoring, lack of awareness, and inadequate
dissemination strategy. Most importantly among the barriers to the dissemination of
biogas technology in Nigeria is the lack of support from the government in the area of
policy promulgation (legislative framework) and implementation, provision of subsidies,
soft loans, and tax incentives in addition to good structural facilities. Meanwhile, in order
for Nigeria to meet and surpass the lowest threshold of energy accessibility of 100 kWh
of electricity and 1200 kWh of modern fuels per person per year proposed by
International Energy Agency (IEA) (AGECC 2010; GEA 2012; IEA 2011), concerted
effort and shared responsibility from various stakeholders including policy makers
(government), researchers, industries, educators, and end-users must be geared towards
introduction of a successful and sustainable biogas technology to provide alignment
between economic, social, environmental, and regulatory variables needed for the
technology. The target of 80% electricity coverage by 2015 in the roadmap to power
sector reforms may not be realizable if urgent measures on accelerating biogas
development and dissemination are not put in place (Jonathan 2010). If barriers to the
development of biogas technology could be surmounted, opportunities such as huge
availability of biogas feedstock, favourable climate which promotes large scale
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5732
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
agriculture, huge population, among others, abounds for accelerated development of
biogas technology in Nigeria. Strategies including creation of biogas research or
consultancy centres and re-organization of a ministry such as NNPC to fill in the
potential supply-demand gap by buying back excess biogas energy or supplying biogas
energy in case of inadequacy; provisions of soft loans, promotion of specialized
programmes focusing on technology and knowledge transfer from countries with well
developed biogas technology, among others, will help in the acceleration and
dissemination of biogas technology in Nigeria. The need for Nigeria to diversify her
energy sources and toe the line of biogas technology by harmonizing biogas into the
existing energy supply-chain cannot be over-emphasized in view of the country’s vision
to be among the top 20 economies of the world by 2020. Given political will and
government unwavering support coupled with the effective management of technical,
social, political, legal, technical, human, cultural, and environmental factors, biogas
technology could be evolved and disseminated to meet the daily energy needs of the
Nigerian citizenry.
ACKNOWLEDGMENTS
This work was supported financially by the Swedish Energy Agency and Lagos
State University. The authors are indeed very grateful for the support provided.
REFERENCES
Abila, N. (2012). "Biofuels development and adoption in Nigeria: Synthesis of drivers,
incentives and enablers," Energy Policy 43, 387-395.
Abubakar, M. (1990). "Biogas generation from animal wastes," Nigeria Journal of
Renewable Energy 1(1), 69-73.
Adeoti, O., Ilori, M. O., Oyebisi, T. O., and Adekoya, L. O. (2000). "Engineering design
and economic evaluation of a family-sized biogas project in Nigeria," Technovation
20(2), 103-108.
Adewunmi, I. K., Ogedengbe, M. O., Adepetu, J. A., and Fabiyi, Y. L. (2005). "Planning
Organic fertilizer industries for municipal solid wastes management," Journal of
Applied Sciences Research 1(3), 285-291.
Adeyanju, A. A. (2008). "Effect of seeding of wood-ash on biogas production using pig
waste and cassava peels," J. Eng. Appl. Sci. 3, 242-245.
AGECC. (2010). Energy for a Sustainable Future. Summary Report and
Recommendations, AGEEC-The Secretary General's Advisory Group on Energy and
Climate Change. United Nations, New York.
Akpan, U.S.’ and Ishak, S.R.(2012). “Electricity Access in Nigeria: is off-grid
electrification using solar photovoltaic panels economically viable?” A
sustainability, Policy, and Innovative Development Research (SPIDER) Solutions
Nigeria Project. (INTERIM REPORT).
Amori, A. A., Fatile, B. O., Ihuoma, S. O., and Omoregbee, H. O. (2013). "Waste
generation and management practices in residential areas of Nigerian tertiary
institutions," Journal of Educational and Social Research 3(4).
Amigun, B., von Blottnitz, H. (2010). Capacity-cost and location-cost analyses for biogas
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5733
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
plants in Africa. Resources, Conservation and Recycling 55, 63-73
Arthur, M., Zahran, S., and Bucini, G. (2010). "On the adoption of electricity as a
domestic source by Mozambican households," Energy Policy 38, 7235-7249.
Atuanya, E. I., and Aigbirior, M. (2002). "Mesophilic biomethanation and treatment of
poultry wastewater using a pilot scale UASB reactor," Environ. Monitor. Assess. 77,
139-147.
Babayemi, J. O., and Dauda, K. T. (2009). "Evaluation of solid waste generation
categories and disposal options in developing countries: A case study of Nigeria," J.
Appl. Sci. Environ. Management 13, 83-88.
Blight, G. E., and Mbande, C. M. (1996). "Some problems of waste management in
developing countries," Journal of Solid Waste Technology Management 23(1), 19-27.
Bond, T., and Templeton, M. R. (2011). “History and future of domestic biogas plants in
the developing world,” Energy for Sustainable Development 15, 347-354
Biogas Support Programme (BSP) (2012). BSP Year Book, BSP, Nepal.
Burke, D. (2001). Dairy Waste Anaerobic Digestion Handbook, Environmental energy
company, Olympia WA 98516, 1-57.
Chaggu, E. J., Mashauri, A., VanBuren, J., Sanders, W., and Lettinga, J. (2002).
"PROFILE: Excreta disposal in Dar es Salaam," Environ. Manag. 30(5), 609-620.
Chandrappa, R., and Das, D. B. (2012). "Waste quantities and characteristics," in: Solid
Waste Management, Environmental Science and Engineering, Springer-Verlag,
Heidelberg, Berlin.
Chen, Y., Yang, G., Sweeney, S., and Feng, Y. (2010). “Household biogas use in rural
China: a study of opportunities and constraints,” Renewable and Sustainable Energy
Reviews 14(1), 545-9.
Cointrea. (1982). "Environmental management of urban solid wastes in developing
countries: A project guide," W. B. Urban Development Department, (ed.), City,
Washington.
Dangogo, S. M., and Fernando, C. E. C. (1986). "A simple biogas plant with additional
gas storage system," Nigerian Journal of Solar Energy 5, 138-141.
Davis, M. (1998). "Rural household energy consumption: The effects of access to
electricity-evidence from South Africa," Energy Policy 26, 207-217.
Dayo, F. B. (2007). "Nigerian Energy Balancesö 1990-2005 Technical paper, Triple
'E'Systems Inc.".
DE-Leew, P. N. (1997). "Crop residue in tropical Africa: Trends in supply, demand and
use," in C. Renard, (ed.), Crop Residues in Sustainable Mixed Crop-Livestock
Farming Systems, UK: International Livestock Research Institute (ILRI) and CAB
International Wallingford.
Desai, A. V. (1992). “Alternative energy in the third world- a reappraisal of subsidies,”
World Dev 20, 959-965.
Deublein, D., and Steinhauser, A. (2008). Biogas from Waste and Renewable Resources,
Wiley-VCH Verlag GmbH & Co. KGaA, 115-128.
Esrey, S. A., Gough, J., Rapapor, D., Sawyer, R., Simpson-Herbert, M., and Vargas, J.
(1998). "Ecological sanitation," S. I. D. C. Agency, (ed.), Stockholm.
Factbook, C. W. (2014). "Nigeria population growth rate[accessed on 10.02.14],"
http://www.indexmundi.com/nigeria/population_growth_rate.html.
FAOSTAT. (2014). "Crop Production[accessed on 10.02.14]," Statistics Division, Food
and Agriculture Organization of the UN. Available at
http://faostat.fao.org/site/567/DesktopDefault.aspx?PageID=567#ancor: Rome, Italy.
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5734
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
Federal Ministry of Environment (Special Climate Unit) (2010). “National environmental
economic and development study (NEEDS) for climate change in Nigeria (final draft)
Ferrer, I., Gaefi, M., Ugetti, E., Ferrer-Marti, L., and Velo, E. (2011). “Biogas production
in low-cost household digesters at the Peruvian Andes,” Biomass Bioenergy 35, 16681674.
GEA. (2012). Global Energy Assessment-Towards a Sustainable Energy Future:
Cambridge University Press, Cambridge UK and New York, USA and the
International Institute for Applied Systems Analysis, Laxemburg, Austria.
Gunnerson, C. G., and Stuckey, D. C. (1986). “Anaerobic digestion-principles and
practices for biogas systems,” World Bank Technical Paper Number 49, Washington,
DC
Howard, G., Pedley, S., Barrett, M., Nalubega, M., and Johal, K. (2003). "Risk factors
contributing to microbiological contamination of shallow groundwater in Kampala,
Uganda," Water Res. 37(3421-3429).
Howells, M. I., Alfstad, T., Victor, D. G., Goldstein, G., and Remme, U. (2005). "A
model of household energy services in a low-income rural African village," Energy
Policy 33(1833-1851).
IEA. (2011). "International Energy Agency. World Energy Statistics," Paris.
IEA. (2006). "Energy for cooking in developing countries," in: IEA, World Energy
Outlook 2006, OECD Publishing, 419-446.
IEA Bioenergy (2014). “Task 37 biogas country overview (Country reports),” IEA
Bioenergy Task 37.
Igoni, A. H., Ayotamuno, M. J., Eze, C. L., Ogaji, S. O. T., and Probert, S. D. (2008).
"Designs of anaerobic digesters for producing biogas from municipal solid-waste,"
Applied Energy 85 430-438.
Ilori, M. O., Adebusoye, A., Lawal, A. K., and Awotiwon, O. A. (2007). "Production of
biogas from banana and plantain peels," Adv. Environ. Biol. 1, 33-38.
Jibrin, M. U., Amonye, M. C., Akonyi, N. S., and Oyeleran, O. A. (2013). "Design and
development of a crop residue crushing machine," International Journal of
Engineering Inventions 2(8), 28-341.
Katukiza, A. Y., Ronteltap, M., Niwagaba, C. B., Foppen, J. W. A., Kansiime, F., and
Lens, P. N. L. (2012). "Sustainable sanitation technology options for urban slums,"
Biotechnology Advances 30(5), 964-978.
Kennedy-Darling, J., Hoyt, N., Murao, K., and Ross, A. (2008). The Energy Crisis of
Nigeria: An Overview and Implications for the Future, The University of Chicago.
Khanal, S. K. (2008). Anaerobic Biotechnology for Bioenergy Production. Principles and
Applications, John Wiley & Sons, Ltd. Publication, USA.
Kristoferson, L.A., and Bokhalders, V. (1991). Renewable Energy Technologies – Their
Applications in Developing Countries, Intermediate Technology Publications,
London, United Kingdom.
Kulabako, N. R., Nalubega, M., Wozei, E., and Thunvik, R. (2010). "Environmental
health practices, constraints and possible interventions in peri-urban settlements in
developing countries - A review of Kampala, Uganda," Int. J. Environ. Health Res.
20(4), 231-57.
Lawal, A. K., Ayanleye, T. A., and Kuboye, A. O. (1995). "Biogas production from some
animal wastes," Niger. J. Microb. 10, 124-130.
Leton, T. G., and Omotosho, O. (2004). "Landfill operations in the Niger delta region of
Nigeria," Engineering Geology 73(1-2), 171-177.
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5735
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
Meisen, P., and Akin, I. (2008). "The case for meeting the millenium development goals
through access to clean electricity," Global Energy Network Institute (GENI), San
Diego, CA.
NBS. (2009). "Social statistics in Nigeria," National Bureau of Statistics, Nigeria, Abuja.
NBS. (2012). National Bureau of statistics/Federal Ministry of Agriculture and Rural
development. “Collaborative survey on national agriculture sample survey (NASS),”
2010/2011 Draft Report.
National Bureau of Statistics (2012). “Social statistics in Nigeria. Part III: Health,
employment, public safety, population and vital registration in Nigeria
Nigeria Climate Review Bulletin (2010). Nigerian Meteorological Agency, Abuja,
Nigeria.
Nyns, E. J. (1986). "Biomethanation Process," in H. J. Rehn and G. Reed (eds.),
Microbial Degradations, VCH, Weinheim, Germany.
Odeyemi, O. (1983). "Resource assessment for biogas production in Nigeria," Nigerian J.
Microbiol. 3 59-64.
Ogwueleka, T. C. (2009). "Municipal solid waste characteristics and management in
Nigeria," Iran. J. Environ. Health. Sci. Eng. 6(3), 173-180.
Ojolo, S. J., Oke, S. A., Animasahun, O. K., and Adesuyi, B. K. U. A. (2007).
"Utilisation of poultry, cow and kitchen wastes for biogas production: A comparative
analysis " Iranian J. Environ. Health Sci. Eng, 4, 223-228.
Okoye, J. K. (2007). "Background study on water and energy issues in Nigeria to inform
the national consultative conference on dams and development," The Federal
Ministry of Agriculture and Water Resources & Society for Water and Public Health
Protection, Nigeria.
Onwuka, C. F. I., Adetiloye, P. O., and Afolami, C. A. (1997). "Use of household wastes
and crop residues in small ruminant feeding in Nigeria," Small Ruminant Research,
24, 233-237.
Ostrem, K. (2004). "Greening waste: Anaerobic digestion for treating the organic fraction
of municipal solid wastes," Columbia University, Columbia.
Owen, E. (1994). "Cereal crop residues as feed for goats and sheep," Livestock Res. Rural
Devel. 6(1).
Palvas, M., Tous, M., Bebar, L., and Stehlik, P. (2010). "Waste to energy - An evaluation
of environmental impact," Applied Thermal Engineering 30, 2326-2332.
Parkin, G. F., and Owen, W. F. (1986). "Fundamentals of anaerobic digestion of
wastewater sludges," J. Environ. Eng. ASCE 112(5), 867-920.
Peters, J., and Thielmann, S. (2008). “Promoting biofuels: Implications for developing
countries,” Energy Policy 36, 1538-1544.
Raven, R. P. J. M., and Gregersen, K. H. (2007). “Biogas plants in Denmark: Successes
and setback,” Renewable and Sustainable Energy Reviews 11, 116-132.
Salomon, K. R., and Lora, E. E. S. (2009). "Estimate of the electric energy generating
potential for different sources of biogas in Brazil," Biomass and Bioenergy 33(9),
1101-1107.
Sambo, A. S. (2005). "Renewable energy for rural development : The Nigerian
perspective," ISESCO Science and Technology Vision 1, 12-22.
Santerre, M.T., and Smith, K. R. (1982). “Measures of appropriateness: The resource
requirements of anaerobic digestion (biogas) systems,” World Dev. 10, 239-261
Sebokah, Y. (2009). "Charcoal production: Opportunities and barriers for improving
efficiency and sustainability," In: Bio-carbon Opportunities in Eastern and Southern
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5736
PEER-REVIEWED REVIEW ARTICLE
bioresources.com
Africa Harnessing Carbon Finance to Promote Sustainable Forestry, Agroforestry
and Bio-energy, UNDP (United Nations Development programme, New York, USA,
102-26.
Shaaban, M., and Petinrin, J. O. (2014). "Renewable energy potentials in Nigeria:
Meeting rural energy needs," Renewable and Sustainable Energy Reviews 29, 72-84.
Singh, B. B., Musa, A., Ajeigbe, H. A., and Tarawali, S. A. (2011). "Effect of feeding
crop residues of different cereals and legumes on weight gain of Yankassa rams,"
International Journal of Livestock Production 2(2), 17-23.
Singh, K. J., and Sooch, S. S. (2004). “Comparative study of economics of different
models of family size biogas plants for state of Punjab, India. Energy Convers.
Manag. 45, 1329-1341
Sorda, G., Banse, M., and Kemfert, C. (2010). “An overview of biofuel policies across
the world,” Energy Policy 38, 6977-6988.
Stehlik, P. (2010). "Contribution to advances in waste-to-energy technologies," Journal
of Cleaner Production 17, 919-31.
Surendra, K. C., Takara, D., Hashimoto, A. G., and Khanal, S. K. (2014). “Biogas as a
sustainable energy source for developing countries: Opportunities and challenges,”
Renewable and Sustainable Energy Reviews 31, 846-859
TheScoopNG (2014). ‘US Stops Importation of Nigeria’s Crude oil’ (June 6, 2014). The
Scoop. Retrieved from http://www.thescoopng.com/us-stops-importing-nigeriancrude-oil/. June 7, 2014.Tomori, O. (2012). “Feasibility study of a large scale biogas
plant in Lagos, Nigeria,”
M.Sc. Thesis, Murdoch University, Australia
Usman, H. A., and Mohammed, B. U. (2012). "Solid waste management, Bauchi, Nigeria
(obstacies and prospects)," Journal of Environmental Science and Resources
Management, 4.
Uwidia, I. E., and Ademoroti, C. M. A. (2011). "Characterisation of domestic sewage
from an estate in Warri, Nigeria," International Journal of Chemistry 3(3).
Weiland, P. (2010). "Biogas production: Current state and perspectives," Appl. Microbiol.
Biotechnol. 85, 849-860.
WHO and UNICEF. (2010). "Progress on drinking water and sanitation," Joint
Monitoring Program Report (JMP).
Winkler, H., Simões, A. F., Rovere, E. L. l., Alam, M., Rahman, A., and Mwakasonda, S.
(2011). "Access and affordability of electricity in developing countries," World
Development 39(6), 1037-1050.
World-Bank. (1977). Production for Tradiitional and Non-conventional Energy Sources
in Developing Countries, World Bank, Washington, DC.
World-Bank. (1992). Introduction to Wastewater: Nature, Sources of Generation and
Treatment with a Focus on Sewage, World Health Organization, WHO.
World-Bank. (1998). Meat Processing and Rendering: Pollution Prevention and
Abatement Handbook, Environmental Department, World Bank, Washington, DC.
Article submitted: April 12, 2014; Peer review completed: May 14, 2014; Revised
version received and accepted: June 8, 2014; Published: June 20, 2014.
Akinbomi et al. (2014). “Nigerian biogas technology,”
BioResources 9(3), 5707-5737.
5737
Paper VI
This article was downloaded by: [Julius Akinbomi]
On: 09 July 2013, At: 23:34
Publisher: Taylor & Francis
Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House,
37-41 Mortimer Street, London W1T 3JH, UK
Environmental Technology
Publication details, including instructions for authors and subscription information:
http://www.tandfonline.com/loi/tent20
Membrane bioreactors’ potential for ethanol and biogas
production: a review
Päivi Ylitervo
a b
, Julius Akinbomi
a c
& Mohammad J. Taherzadeh
a
a
School of Engineering, University of Borås , Borås , Sweden
b
Industrial Biotechnology, Chalmers University of Technology , Gothenburg , Sweden
c
Department of Chemical and Polymer Engineering , Lagos State University , Lagos , Nigeria
Accepted author version posted online: 19 Jun 2013.Published online: 08 Jul 2013.
To cite this article: Environmental Technology (2013): Membrane bioreactors’ potential for ethanol and biogas production: a
review, Environmental Technology, DOI: 10.1080/09593330.2013.813559
To link to this article: http://dx.doi.org/10.1080/09593330.2013.813559
PLEASE SCROLL DOWN FOR ARTICLE
Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained
in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no
representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the
Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and
are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and
should be independently verified with primary sources of information. Taylor and Francis shall not be liable for
any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever
or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of
the Content.
This article may be used for research, teaching, and private study purposes. Any substantial or systematic
reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any
form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http://
www.tandfonline.com/page/terms-and-conditions
Environmental Technology, 2013
http://dx.doi.org/10.1080/09593330.2013.813559
Membrane bioreactors’ potential for ethanol and biogas production: a review
Päivi Ylitervoa,b , Julius Akinbomia,c and Mohammad J. Taherzadeha∗
a School
of Engineering, University of Borås, Borås, Sweden; b Industrial Biotechnology, Chalmers University of Technology,
Gothenburg, Sweden; c Department of Chemical and Polymer Engineering, Lagos State University, Lagos, Nigeria
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
(Received 19 February 2013; final version received 5 June 2013 )
Companies developing and producing membranes for different separation purposes, as well as the market for these, have
markedly increased in numbers over the last decade. Membrane and separation technology might well contribute to making
fuel ethanol and biogas production from lignocellulosic materials more economically viable and productive. Combining
biological processes with membrane separation techniques in a membrane bioreactor (MBR) increases cell concentrations
extensively in the bioreactor. Such a combination furthermore reduces product inhibition during the biological process,
increases product concentration and productivity, and simplifies the separation of product and/or cells. Various MBRs have
been studied over the years, where the membrane is either submerged inside the liquid to be filtered, or placed in an external
loop outside the bioreactor. All configurations have advantages and drawbacks, as reviewed in this paper. The current review
presents an account of the membrane separation technologies, and the research performed on MBRs, focusing on ethanol
and biogas production. The advantages and potentials of the technology are elucidated.
Keywords: membrane bioreactor; ethanol; biogas; filtration
Introduction
Industrial applications of membrane technology for biological applications have in the last decade flourished over the
world. Several large companies, such as Kubota, MerckMillipore, Pall, GE Healthcare, and Sartorius, are now
providing membrane products aimed for highly different
separation processes. Many membranes are also used in
biological processes in the form of membrane bioreactors
(MBRs). At present, most MBRs are employed in water
or wastewater treatment, in order to achieve a quality of
the effluent, sufficiently suitable for reuse or recycling.[1,2]
MBRs for wastewater treatment are well developed, and
are today used commercially in many countries for mostly
dilute wastewater streams, while MBRs for ethanol production is still a new concept and need development for larger
scale utilization.
Although there is widespread industrial application of
membrane technology in wastewater treatment, the treatment process has been limited to aerobic biological process
with only few industries employing anaerobic processes
owing to the feed types, biological process and operational
conditions of anaerobic MBR (AnMBR).[3,4] The complexity of the biological reactions and different types of
microorganisms involved in anaerobic processes presents
unique effects on the membrane fouling characteristics,
when compared with the aerobic processes. Moreover,
there is high probability of process failure due to the
∗ Corresponding
author. Email: [email protected]
© 2013 Taylor & Francis
presence of inhibitory substances, such as heavy metals,
chlorinated hydrocarbons, and cyanides, often present in
feeding wastewaters or sludge. There is also additional
cost of heating the AnMBRs to mesophilic or thermophilic
temperatures especially during cold climates.[5] These
challenges result in the limited industrial application of
AnMBRs for wastewater treatment and biogas/ethanol production. However, the interest in application of anaerobic
process in membrane reactors for wastewater treatment is
increasing because the process unlike the aerobic process
has the potential of energy recovery from the waste and
reduction of greenhouse gas emissions.[3] Furthermore,
AnMBR, unlike aerobic MBR (AeMBR) produces very low
amount of residual sludge since it has low biomass yields
and growth rates. With more exhaustive studies to address
the challenges of anaerobic processes, full-scale industrial
application of AnMBRs will be widespread. MBRs for
water and wastewater treatment have been extensively covered in numerous other reviews,[6–9] and are therefore not
in focus in this paper. Some MBRs producing biogas from
wastewater is, however, mentioned, as some MBRs treating
wastewater can be utilized for anaerobic biogas production.
One of the major complications in biotechnology and in
biological processes, hampering a successful commercial
process, is downstream processing, such as separation of
products or biocatalysts from the spent medium or product
purification.[10] Furthermore, product streams formed by
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
2
P. Ylitervo et al.
biological processes are often very dilute, and consist of a
complex mixture of components. Downstream separation
processes can therefore be both expensive and technically
challenging.[10] As a consequence, bioprocesses in general, and downstream processing in particular, need to have
their process steps enhanced and integrated, in order to
improve yields, cut process time, and reduce operation and
capital costs.[11]
In MBRs, a biological process is integrated with a
permselective membrane. In the process of ethanol or
biogas production, the membrane can either be used for
separating cells from the medium, thereby increasing the
biomass concentration in the bioreactor, to aid removal
of the inhibitor, or to recover product in situ. Membrane
separation techniques can also easily be coupled with continuous processes.[12] Continuous processes have several
advantages compared with traditional batch operations,
especially since they use smaller reactors or employ higher
dilution rates, lowering capital as well as maintenance
costs. The continuous process demands an overall low
level of inhibitors in the culture medium and/or high cell
concentrations,[13] which is accomplished with the MBR
technology. Furthermore, slow-growing microorganisms in
anaerobic biogas reactors, such as methanogenic bacteria,
can benefit from being retained inside the bioreactor.
The aim of the present review was to introduce the
MBR technology in ethanol and biogas processes, and to
summarize the development of MBRs and the membrane
technologies for these biofuels, along with their advantages
and future potential.
Ethanol and biogas production
Ethanol and biogas (methane) are renewable and environmentally friendly fuels which can be used as an alternative
to the traditional fossil fuels. Optimistically, the development of ethanol and biogas production can decrease the
worlds’ dependence on fossil fuel, thereby reducing the
excessive emissions of greenhouse gases, which accelerate global warming. As both ethanol and biogas can be
produced from renewable feedstocks, such as agricultural,
municipal and forest residues, their production does not
contribute to the net emissions of carbon dioxide. The basic
problem with ethanol and biogas production is the question of process economy and product yield. One interesting
and promising approach is to use membrane technology and
MBR processes, which, by assisting in achieving high cell
densities, separating cells, products, or residual compounds
in the process, greatly would improve the ethanol and biogas
production economy.
In anaerobic digestion of waste materials, the amount
of biogas (mainly methane and carbon dioxide) produced is
mainly dependent on the interactions of different consortia
of degrading microorganisms. The conversion of organic
material to methane is immensely complex, and consists of
four major stages: hydrolysis, acidogenesis, acetogenesis,
and methanogenesis. The last stage, performed by methaneforming bacteria, is commonly the most sensitive step in
the process, since methane-forming bacteria have a very
slow growth rate and are sensitive to inhibitors, pH, and
other process conditions. It is therefore important to prevent bacteria from being washed out from the reactor,
and to reduce inhibitor levels. AnMBRs have proven to
provide a successful technique for avoiding cell washout,
longer retention times, and presence/accumulation of toxic
compounds.[14]
Ethanol is currently produced mainly from sugar and
starch-rich materials, which are also used as food and feed.
The current global debate on food vs. fuel renders these
types of raw materials little appeal for ethanol production.
Being omnipresent, wastes and lignocellulosic residuals
from municipalities, agriculture, and forest industries are
deemed more suitable as raw materials.[15] Nevertheless,
lignocelluloses are usually very recalcitrant, and need pretreatment or complete hydrolysis into fermentable sugars
prior to being utilized for production of, e.g. ethanol or
biogas.
Various different sugar containing hydrolysates produced from lignocellulosic materials have been applied for
ethanol production in MBRs. Lignocellulosic materials are
abundant and thus of interest as feedstock for producing
fermentable sugars for ethanol production. However, the
material has to be hydrolysed by, e.g. acid or enzymatic
hydrolysis, prior to fermentation. Dilute acid hydrolysis
is performed at high temperature, with relatively low concentrations of, e.g. sulphuric acid. During the degradation
process, several toxic compounds are formed, which could
affect the fermentation process negatively. Numerous studies have therefore been carried out, striving to find ways to
overcome the toxicity of hydrolysates, e.g. by using high
cell densities,[16] MBR,[17,18] or detoxification.[19,20]
In the production of bulk fuels, such as ethanol and
biogas, every step has to be optimized in order to obtain
an economically viable large-scale production. It is therefore crucial to maximize productivity without cell cultures
being destabilized due to, e.g. inhibition problems or
energy-intensive product recovery. Continued development
of MBRs will hopefully aid in resolving these issues.
Membrane concept
In order to separate components or cells in a liquid mixture by means of filtration, membranes are coupled with
the bioreactors. The porous membrane is manufactured to
contain ceramic, metallic, or polymeric material. The separation of the mixed compounds in the liquid is usually
brought about by applying pressure or vacuum across the
porous permselective membrane, but can also be prompted
by a concentration gradient.[21,22] The membrane forms
a barrier, allowing some components to pass the membrane more readily than others, and this selectivity is mostly
determined by the pore size of the membranes,[1] but
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
Environmental Technology
other characteristics of the membranes, such as hydrophilicity, can also affect the selection, hindering hydrophilic (or
hydrophobic) components to permeate the membrane.[23]
Membranes are typically defined according to their separation mode, i.e. microfiltration, ultrafiltration, nanofiltration,
and reverse osmosis. In MBRs, aimed at ethanol and biogas
production, microfiltration and ultrafiltration membranes
are employed.
Presently, there are two principally different MBR
designs, where the membrane is either operated under
direct pressure or vacuum. In the first configuration, with
direct pressure, the membrane is placed in an external loop
separated from the bioreactor (external cross-flow membrane) and a pump forces the bioreactor broth into the
membrane module and to permeate through the membrane
(Figure 1(a)).[22] The external cross-flow MBRs are operated in a cross-flow mode, where the liquid to be filtered
flows with high velocity parallel to the membrane surface,
which thereby also hinder cake formation on the membrane
surface. This mode of operation reduces the fouling tendency of external cross-flow membranes,[24] and increases
the flux through the membrane. Among the disadvantages
of external cross-flow MBRs, are the significant energy
amounts required for sustaining a continuous flow through
the membrane,[1] and the complex reactor design can be
mentioned.
In contrast, when the membrane is operated at lower than
atmospheric pressure or vacuum, a pump can be used to pull
permeate through the membrane. This configuration is usually named submerged or immersed, since the membrane is
placed directly in the liquid (Figure 1). The advantages of
submerged MBRs are that they usually require less energy
to run compared with external cross-flow MBR.[1,22] The
submerged MBR can, however, be problematic to operate
at high particulate or cell concentrations, due to fouling.
Usually a larger membrane surface area has to be applied
in submerged than cross-flow MBRs.[1] A way to disrupt
fouling and cake formation is to vigorously purge gas across
the submerged membrane surface.[22]
There are two configurations in which the vacuumdriven submerged membranes can be designed. The membrane can either be immersed directly into the bioreactor
(Figure 1(b)), or submerged in a separate container which
is connected to the bioreactor (Figure 1(c)). The external
chamber configuration with submerged membrane has the
advantage of being easier to clean,[22] but require energy
to pump the retentate back to the bioreactor.
When membranes do not allow components to pass
through, the particles or cells in the concentrated liquid tend
to attach and accumulate on the membrane surface, curbing the filtering process. Upon component accumulation,
the flow through the membrane is reduced by different phenomena. In combination, these phenomena are referred to as
fouling, where compounds deposit on the membrane surface
or inside the membrane. Fouling is also the main obstacle for
optimal membrane function.[1] MBRs operation is largely
3
Figure 1. Schemes of MBR configurations (a) external
cross-flow membrane, (b) internal submerged membrane and (c)
external submerged membrane.
dependent on the membrane flux and usually overtime
membrane fouling will result in a lowered permeate flux or
an increased transmembrane pressure (TMP). If the membrane in the MBR is operated at a flux below a critical value,
the TMP can be kept constant with no fouling of the membrane. If the flux is increased over a critical flux value, the
TMP will increase rapidly together with membrane fouling.
However, there is never any perfect non-fouling operation,
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
4
P. Ylitervo et al.
but by keeping the flux at a value lower than the critical
flux (sustainable flux) can give a slower linear increase in
TMP.[2,22] By monitoring the TMP overtime, the membrane module can also be cleaned prior to extensive fouling,
which makes long cycles of continuous processes possible.
If the fluid to be filtered contains a large amount
of solids and soluble compounds, as is the case during
fermentation processes, the membrane flux tends to fall
sharply with increasing time and cell growth.[12] Different
techniques have been explored to overcome this problem, e.g. turbulent promoters,[25] rotating filters,[26,27]
unsteady flows,[28] intermittent jets,[29] periodical inversion of flows,[30] ultrasound,[31] gas injection,[12,32]
anti-fouling membranes,[2] and dynamic membranes.[33]
Most of these methods increase the turbulence of the liquid near the membrane, which destabilizes the cake layer
on the membrane surface, thereby improving the filtration
flux. The most suitable method to maintain a high permeate
flux is dependent on which membrane module is applied.
For example, gas sparging has been shown to be effective to
reduce concentration polarization and/or increase permeate
flux in tubular, hollow-fibre, plate and frame (PF) and spiral
wound (SW) membrane modules.[34]
Ultrafiltration membranes used for cell separation are
reported to have a lower fouling tendency than microfiltration membranes, which may be attributed to differences
in pore size. Since the pores of microfiltration membranes
are in the same order of magnitude as the microorganisms,
smaller cells can lodge themselves inside the larger pores,
causing physical blockage of the pores.[35] However, if the
pores are much smaller than the cells, as in ultrafiltration
membranes, the shear force of the flow will force the cells
to ‘roll off’ the surface.[35] Besides the cell concentration,
other components in the media, such as proteins, carbohydrates, and particles, hold the potential to increase fouling
rates.[35,36] In the present paper, the phenomena of fouling
are not covered in further detail, as it is discussed immensely
elsewhere.[2,8]
Membrane processes: development trends and
perspectives
Membrane filtration is presently used within many different areas, such as biopharmaceutical processes, desalination, water treatment,[8] food and beverage manufacturing, industrial production of paints, adhesives, chemicals,
etc.[35,37] The demands on the filtration processes and the
filter membranes largely depend on the application. Regulations for filters and filtration processes are very strict in
drug and other biopharmaceutical applications, whereas filters used in, e.g. paint, adhesive, and chemical industries are
much less refined, and considerably cheaper. In the beverage industry, where it is essential to remove microorganisms
to avoid contamination and consumer illness as well as
microbial spoilage, microfiltration may be an alternative
to pasteurization, given that an adequate membrane is used
to filter the product.[37]
In bioprocesses, membrane separation is used for different upstream and downstream applications.[21,38] The
interest in membranes for biotechnological applications
is mainly driven by the demand for higher productivity
and reduced production costs. Over the years, MBRs have
proven to hold many advantages; they offer for instance high
product yields, ascertained sterility, high biological activity,
and superior separation efficiency. In addition, the process
usually has a low energy consumption, enables a continuous operation, and is simple to operate and scale up. At
present, membranes are applied in biotechnology for sterile
filtration, liquid clarification, cell harvesting, virus removal,
protein concentration, etc.[21]
Configuration of membranes in MBRs
The membranes for MBRs are produced in various configurations, such as PF, hollow-fibre, SW, and tubular
geometries.[21,39] Hollow-fibre membranes have been
applied at a laboratory scale, as they provide a large surface area per packing density.[21] In ethanol and biogas
production processes, many different configurations have
been tested at a small scale, several of these are described
in this paper.
Hollow-fibre and tubular membrane reactors
Yeast has been successfully immobilized inside hollowfibre membranes, which normally are used for ultrafiltration. Hollow-fibres membranes have hence been employed
for the cultivation of yeast, bacteria, mammalian cells,[39]
and enzymes.[40] Hollow-fibre membranes are generally
used in bundles, joined and sealed with a cylindrical housing at each end. The housing enables separation of the
extracapillary space from the fibre lumen. Cells have been
cultivated in the extracapillary space of the module, while
the medium was pumped through the lumen space, and
nutrient molecules diffused through the fibre membrane.
The porosity of the membrane needs to be selected in accordance with what the membrane must retain, i.e. the cells and
the product molecules, or only the cells.[39]
Very high cell densities have been acquired in hollowfibre reactors, since they offer very large surface area per
volume.[41] The housing of the hollow-fibre membrane
module allows the cells to escape shear forces and contamination, since the porosity of the membrane facilitates
selective nutrients to permeate the extracapillary space.[39]
A previous study [40] describes how Saccharomyces
cerevisiae was grown on the shell side of asymmetricwalled polysulfone membranes as well as on the surface of
isotropic-walled polypropylene hollow-fibre membranes.
The medium was continuously pumped through the hollowfibre lumen in order to supply the cells with nutrients,
and to remove products by diffusion. In the asymmetric
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
Environmental Technology
membranes, the yeast reached very high, tissue-like
densities of more than 1010 cells/mL, and in some regions
the cells accounted for almost 100% of the volume. However, the cell packing showed a radial distribution across
the fibre wall, indicating that the cells located further than
100 μm from the lumen surface did not receive sufficient
amount of glucose. In the asymmetric membranes, the yeast
density reached approximately 3.5 × 109 cells/mL.[40]
Notwithstanding that very high cell densities were reached,
low nutrient and product transport rates curtailed the system
performance, and over time, accumulation of CO2 inside the
reactor reduced the ethanol production as well. The highest
ethanol productivity reported was 26 g/(L h).[40] However,
much higher productivities have been achieved by, e.g. fermenting lactose in an MBR, using a cross-flow membrane.
Cheryan and Mehaia [13] reached an ethanol productivity
of 240 g/(L h) at a cell concentration of 90 g/L.
There are nevertheless some drawbacks with the use of
hollow-fibres, such as fouling, clogging fibres, problems in
accessing the cell mass, difficulty to sustain a well-defined
fibre spacing, and rupturing fibres when cells grow and produce gas.[39] In external hollow-fibre modules, the high
recirculation speed will furthermore result in high pumping
costs, and may also damage the cells.[24]
Tubular membranes may be preferable to hollow-fibres,
in order to avoid the risk of clogging fibres.[42,43] In a trial
conducted by Escobar et al.,[43] a set of ceramic tubular
membranes was tested at pilot scale in a 7000 L MBR.
Ceramic tubular membranes were applied as they can easily be backwashed, have high flux rates, and can be cleaned
with aggressive chemicals if needed. The cell concentration
inside the MBR was regulated and kept at below 120 g/L.
By applying low fluxes (below 70 L/m2 h), the membranes
could be operated successfully for approximately 4.5 days
before cleaning. However, operating the membranes at
higher fluxes required cleaning more often.[43]
Plate and frame and spiral wound membrane reactors
Even though PF membrane reactors have a lower surface
area per volume compared with hollow-fibres, they hold
most of the advantages of hollow-fibres. PF MBRs have
been applied for many purposes in order to produce, e.g.
antibodies or enzymes or to apply enzymes by themselves.
These flat membranes can be used, e.g. in frames that are
separated with a spacer creating a space between the membranes where cells or enzymes can be added.[44] In these
MBRs, the space between the membranes is accessible and
the cells can hence be replaced if necessary. Furthermore,
the distance between the cells and the medium is easily controlled by changing the spacers thickness between the two
flat membranes.[39] PF modules which do not have any
spacer material between the membranes also exist.[45]
The PF membrane can also serve as a separation unit
to separate, e.g. pectin, antibodies, cells, enzymes from
5
a liquid in the same way as hollow-fibres [44] or to
concentrate, e.g. fermentation broths.[46] For example,
Pyle et al. [46] used both flat sheet and tubular membrane
modules to concentrate a bakers’ yeast suspension up to
20% dry weight. In a study performed by Thuvander,[47] a
PF module from Alfa Laval was used to retain cells in the
bioreactor during continuous fermentation of molasses. The
PF module was both employed internally and externally to
the bioreactor. During the cultivation, the membrane flux
rapidly decreased down to 6.3–4.6 L/m2 during the first
hours, were it stayed during the main part of the cultivation.
The yeast concentration increased steadily during cultivation and reached almost 15 g/L after around 60 h when using
the external PF module.[47]
As in PF modules, SW modules also use flat sheet membranes. However, in SW modules the membrane sheets
are tightly wound together around a central collector tube,
usually with a mesh-like spacer between each sheet of
membrane. The design of the SW modules makes them
only suitable for use on feed streams that only contain fine
suspended solids.[48] In ethanol production, SW modules
have, e.g. been used to combine two separation processes,
extraction and membrane permeation in a single module in
order to lower the energy cost during ethanol separation.
Offeman and Robertson [49] successfully developed a SW
module where ethanol in the fermentation broth could be
separated. In the module, one membrane contains solvent
and extracts the ethanol from the broth. A second membrane
can then by the help of a vacuum removed the ethanol from
the solvent.[49] SW modules have also been applied during enzymatic hydrolysis of corn starch to produce clarified
glucose syrup. The formed sugars can thereafter be utilized
for ethanol production.[50]
Membrane technology in ethanol and biogas processes
Ethanol production process
Attaining a high ethanol productivity in the bioreactors is
crucial for keeping the bulk chemical costs low. Keeping
the cell density high is one way of achieving high productivity, but dilution rate and cell growth rate undermine this
in conventional continuous cultivations. Today, centrifugation or filtration is the most preferred mode of separation of
microbial cells at the industrial scale.[51] However, other
cell retention methods, such as cell immobilization,[52–55]
encapsulation,[53,56–59] or cross-flow membranes,[13,18]
have also been utilized to maintain high biomass concentrations in the bioreactor. MBRs have been applied in several
studies on cell recycling, with the purpose of gaining higher
productivity.
MBRs have been examined for ethanol production, with
the membranes coupled either internally or externally to the
bioreactor,[18,60] and different types of membrane technologies and MBRs were tested with the aim of improving
the fermentative production of ethanol. MBRs have also
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
6
P. Ylitervo et al.
been used for cell retention/recycling [13] to obtain high
biomass concentrations in the bioreactors. Furthermore,
extractive MBRs have been applied to remove inhibitors
such as ethanol,[61,62] and pervaporation MBRs have been
used to eliminate volatile inhibiting compounds from the
cultivation broth.[63]
Mercier et al. [12] succeeded to reach a final yeast
concentration of 150 g/L by performing a 100 h continuous fermentation, using cross-flow filtration to recycle cells
back to the bioreactor. To avoid severe fouling and declining membrane flux at higher biomass concentrations, air
was injected into the feed stream.[12] In contrast, Lafforgue
et al. [42] were able to obtain as much as 345 g/L of yeast
biomass in a bioreactor coupled to a microfiltration membrane for cell recycling. Within 100 h, the total biomass
level had reached 300 g/L with a viability of 75%, and the
system was stable for more than 50 h. While performing fermentation at very high yeast concentrations, both cell size
and morphology were altered, and the viscosity of the broth
was increased.[42]
Several studies on ethanol fermentation in MBRs have
been conducted, using, e.g. glucose as the carbon source
[17,60,61,64]; these are summarized in Table 1. Chang
et al. [60] used an internal stainless steel filter, and could
successfully produce ethanol from glucose at an average
yield of 92.7% of the theoretical yield, with a productivity
of 20 g/(L h) ethanol, much higher than achieved in traditional continuous systems.[70] The MBR was furthermore
operated for 10 days without complications, and showed
a stable ethanol production at an average cell concentration of approximately 55 g/L. The cell concentration was
maintained at the same level by controlling dilution rate
and bleed ratio. Several other substrates such as tapioca
hydrolysate,[17] wood hydrolysate,[16,18] and lactose [13]
have also been successfully used in MBRs for fermentation
of ethanol.
Fermentation of the liquid part of enzymatically hydrolysed oak wood was successful in a continuous cultivation,
using an MBR with a submerged membrane, and yielded an
ethanol concentration of more than 70 g/L.[65] Prior to fermentation, the hydrolysate was concentrated up to 180 g/L
glucose by vacuum evaporation, and sterilized at 60◦ C for
120 min. The low temperature applied was to avoid the
formation of large amounts of toxic materials during the
sterilization process. The maximum ethanol productivity
during the continuous fermentation was 16.9 g/(L h).[65]
Removal of inhibitor from the fermentation processes
Fermentation can be inhibited by a high concentration of
the product (e.g. ethanol), carbon sources (such as sugars), and other compounds that follow the process, such as
furans or phenolic compounds present in the lignocellulosic
hydrolysates. Product inhibition can be reduced by removing or degrading the toxic compounds from the broth in situ,
using different methods. Continuous product recovery of
volatile compounds can be carried out by creating a vacuum
in the bioreactor, or in a separate chamber, where the volatile
inhibitory compounds (e.g. ethanol) subsequently can be
distilled from the broth.[71,72] Nonvolatile inhibitors can
be removed by using conventional liquid extraction methods (the most commonly used method),[11,73,74] adsorption on ion-exchange resin,[75] activated carbon,[45,76,77]
or polymeric adsorbents.[78]
Inhibitor levels are also possible to reduce by using
MBRs. Since very high cell densities can be achieved in
MBRs, the cells’ own capacity of performing in situ detoxification of some compounds can be utilized. For example,
one study showed that a continuous yeast cultivation in an
MBR was able to ferment a sugar solution containing up to
17.0 g/L furfural, without drastic changes of ethanol productivity. Furfural is known to severely affect the growth of
yeast, even at low concentrations. However, by effectively
keeping the yeast at a high density in the MBR enabled rapid
degradation of the incoming furfural, leaving the permeated
fluid from the MBR with merely low levels of furfural.[79]
The fermenting yeast can also be protected from toxic
compounds in the medium by means of encapsulation.
By enclosing the yeast inside spherical hydrophilic alginate membranes, hydrophobic inhibitors, such as limonene
present in citrus waste, can be hindered to enter and negatively affect the cells. For example, Pourbafrani et al. [23]
encapsulated yeast in this manner and were able to ferment
a medium containing as much as 1.5% limonene.
A high concentration of ethanol tends to have inhibition effects on the process, and the toxicity of ethanol
is hence directly related to the sugar concentration in the
medium.[80] As an extreme example, an ethanol-tolerant
yeast strain of Saccharomyces diastaticus was able to yield
a final ethanol concentration of 17.5% (v/v).[81] High
ethanol concentrations result in lower batch yields, thus
reducing the production capacity. On the other hand, recovering ethanol from a dilute fermentation broth requires a
large amount of energy, resulting in increased processing costs.
Membrane-based extraction
Ethanol fermentation by means of membrane-based extraction has previously been reported.[61,82] The membrane
separates the aqueous and solvent phases, thus eliminating
formation of emulsions and a need to separate the aqueous and solvent phases in downstream processes. Both
SW, PF and hollow-fibre membranes have been used to
separate the culture medium from the extracting solvent.
For instance, multi-membranes, composed of three separate flat sheet membranes, were employed for extractive
ethanol fermentation by Cho and Shuler.[62] Applying tributyl phosphate as the extracting solvent, they succeeded
to completely ferment 200 g/L glucose, using a 6:4 volume ratio of tributyl phosphate to medium. Frank and
Sirkar [83] performed hollow-fibre membrane extractive
150,000 MWC
50,000 MWC
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
Dry bakers’ yeast (Oriental
yeast)
Dry bakers’ yeast (Oriental
yeast)
–
–
Submerged
Note: MWC, molecular weight cutoff.
Silicalite-silicon
rubber
Saccharomyces cerevisiae
NRRL-Y-132
–
10,000 MWC
10,000 MWC
Hollow-fiber
–
fermentor
Hollow-fiber
Polysulfone,
fermentor
polypropylene
Hollow-fiber
Polyamide
fermentor
(B) Extractive fermentor
Hollow-fiber
Celgard® X-20
fermentor
(polypropylene)
(c) Pervaporation fermentation
Submerged
Silicalite
–
0.14 μm
External cross-flow Ceramic tubular
0.45 μm
Saccharomyces cerevisiae
140 L
(Bakers’yeast)
Saccharomyces cerevisiae
–
ATCC 96581
Saccharomyces cerevisiae
7000 L
Saccharomyces cerevisiae
2.4 L
7013
Kluyveromyces fragilis
0.5–3 L
NRRL 2415
Saccharomyces cerevisiae R1: 4.5 L R2: 1.5 L
CBS 8066
Saccharomyces cerevisiae
625 mL
NRRL-Y-132
Saccharomyces cerevisiae
–
ATCC 4126
Saccharomyces cerevisiae
40 mL
ATCC 24858
2 μm
150 mL
500 mL
75 mL
1.5 L
Saccharomyces cerevisiae
0.3 μm
1.5 L
1.8 L
1.5 L
Working volume/
reactor size
Saccharomyces cerevisiae
Saccharomyces cerevisiae
ATCC 24858
Saccharomyces cerevisiae
Microorganism
0.3 μm
2 μm
2/10 μm
50,000 MWC
Ceramic cylindrical
tubes
Ceramic cylindrical
tubes
Fluoro polymer
Porous stainless steel
cylindrical tubes
Porous stainless steel
cylindrical tubes
Pore
size/MWC
External cross-flow Durapore filter
(Millipore)
External cross-flow Ceramic tubular
External cross-flow Carbon/zirconium
oxide tubular
External cross-flow Hollow-fiber
Submerged
Submerged
Submerged
Submerged
(A) Cell retention
Submerged
Membrane
material
Summary of studies on various MRBs for ethanol production.
Membrane
configuration
Table 1.
10
–
–
5.0 × 108 cells/mL
10
17
17
31.6
6
cells/mL
260
41
240
–
33
–
100–300
1010
R1: 59 R2: 156.8
90
60–100
300
12
15
13
28.4
17.7
16.9
118
42
1.5 × 109 cells/mL
58
14.7
20
Productivity
(g/(L h))
208
50–150
Biomass
(g/L)
Glucose 200 g/L
Glucose 100–120 g/L
Glucose 300 g/L
Glucose 100 g/L
Glucose 10–89 g/L
Glucose 100 g/L
Glucose 500 g/L
Lactose 50 g/L
Dextrose 93–95 g/L
Glucose 150 g/L
Wood hydrolysate
Molasses
Glucose 100 g/L
Glucose 200 g/L
Tapioca hydrolysates
Wood hydrolysate
Glucose 100 g/L
Glucose 100 g/L
Medium
[69]
[68]
[61]
[41]
[40]
[67]
[64]
[13]
[43]
[42]
[18]
[47]
[66]
[65]
[17]
[60]
Reference
Environmental Technology
7
8
P. Ylitervo et al.
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
fermentation, with dibutyl phthalate as solvent. The advantage of extracting the inhibitors was however limited due
to low glucose levels during the fermentation process. The
full potential of extractive fermentation is only attained at
high glucose concentrations. Kang et al. [61] used a feed
medium holding a glucose concentration of 300 g/L in their
extractive fermentation with hollow-fibre membranes, and
the derived ethanol productivity increased significantly during the in situ solvent extraction, particularly when the
solvent/substrate ratio was increased.
Improving the membrane separation technology of fermentation and distillation would greatly increase the potential for making ethanol production technically appealing
and more economical. Ethanol separation need to be more
cost-effective and less energy-intensive.[77] Online solvent extraction and pervaporation are techniques that use
membranes to facilitate the separation of ethanol from the
fermentation broth.
Pervaporation of ethanol
In the pervaporation process using hydrophobic membranes, components are transformed from liquid to vapour
and transferred through a hydrophobic membrane. Selectivity is not based on the thermodynamic evaporation
equilibrium, but on the differences in membrane sorption, and diffusion properties of the different components.
Using hydrophobic pervaporation membranes for separating volatile organic compounds (e.g. ethanol) has become
a fascinating alternative to conventional processes, such as
distillation, extraction, adsorption, and stripping.[63] Pervaporation membranes can be connected either externally
in a sub-stream, or be integrated into the bioreactor.
One of the virtues of coupling hydrophobic pervaporation membranes with the fermentation process is the
elimination of volatile compounds exerting product inhibition, by selectively removing these compounds from its
dilute aqueous medium. By keeping the inhibitory compounds below a critical level, the overall productivity is
enhanced. Another advantage of the pervaporation technique is the ethanol concentration attained during the
process. Since the permeate from the hydrophobic pervaporation membrane holds a high concentration of ethanol,
the energy needed for further concentration and purification
is low. Although ethanol and water can be effectively separated by distillation, this procedure constitutes an energy
sink in the process.[77] As much as 60% of the energy
obtained from combustion of ethanol may be required for
the distillation separation process.[78]
Different membranes made of e.g. polyvinyl alcohol,
polyurethane,[84] and silicalite,[68] have been utilized for
pervaporation of ethanol during fermentation. Hydrophobic pervaporation membranes are usually made of silicones, and are hence impermeable to electrolytes. As a
consequence, most organic acids and salts are enriched in
the fermenter.[68]
By using a silicalite pervaporation membrane, Nomura
et al. [68] succeeded to extract a permeate from fermentation
broth, with a high concentration of ethanol. The permselectivity of the hydrophobic pervaporation membrane enabled
a permeate containing 81.0% ethanol to be extracted from a
fermentation broth, where the concentration of ethanol was
4.73%.[68] By-products, such as acidic compounds in the
fermentation broth, can however adsorb onto the membrane
silicalite surface, and negatively influence the pervaporation performance. This can be prevented by coating the
surface with a thin layer of hydrophobic silicon rubber.[69]
Application of membrane technology in biogas production
All aerobic membrane technologies (AeMBRs) utilized so
far for wastewater treatment can be used for biogas production in an anaerobic process with slight design modification
of the AeMBR. AnMBR technology for wastewater treatment still needs to scale the hurdle of operational parameters
including low temperatures and hydraulic retention times
comparable to AeMBR technology.[85] However, AnMBR
technology has received some degree of acceptance in the
recent years as a substitute in place of AeMBR for wastewater and other waste streams. It is because of AnMBRs
potential of energy recovery in addition to providing the
same benefits as AeMBR with reduced energy, while AeMBRs require high energy input for aeration.[86] In AnMBR,
emphasis is on biogas recovery along with waste treatment. The absence of oxygen as an electron acceptor in
anaerobic process prompts the microbial systems to dispose electrons into methane (biogas).[14] Unlike AeMBRs
which emits greenhouse gases such as carbon dioxide and
nitrous oxide (if nitrifying/denitrifying), AnMBRs generate useful gases such as biohydrogen and biogas that could
be used for energy production.[87] AnMBR technology is
being applied for biohydrogen production since hydrogen
has high energy content (142 kJ/g) and devoid of harmful emissions during utilization.[88] The advantage of the
AnMBR technology in this area is that both biohydrogen
and biogas could be produced using a single membrane
system since in anaerobic digestion, the hydrogen production phase occurs briefly before the methanogenic phase
occurs. The methanogenesis could be inhibited using various means including chemical inhibition, pH control, promotion of ferric-reducing conditions, control of hydrogen
partial pressure, among others.[89,90]
Furthermore, membrane technology has been developed
for biogas purification, which eliminates major problems
commonly encountered in conventional methods of purification. For instance, high concentration of carbon dioxide in
biogas makes its desulphurization difficult in conventional
method that is based on a chemical reaction of hydrogen sulphide (H2 S) with quicklime or slaked lime in a solid or liquid
form. This is because the carbon dioxide also reacts quickly
with the quick and slaked lime making it insufficient for biogas desulphurization.[91] Moreover, conventional biogas
Environmental Technology
Table 2.
Application of membrane technology for hydrogen and biogas production.
Membrane
configuration
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
9
Membrane
characteristics
Medium
(A) Laboratory scale AnMBR for hydrogen production
Submerged
Kubota Co, plate flame (FS) type of
Glucose
0.45 μm pore size, 0.1 m2 effective
area and 240 × 340 × 10 mm module
size
External
US Filter C., tubular, ceramic, pore
Glucose
size:0.2–0.8 μm; membrane surface
area:0.005 m2 and membrane
dimension of ID = 7 mm, OD =
10 mm; length = 250 mm
External
Deposition Sciences Inc., hollow fibre, Glucose
MF, pore size: 0.2 μm
Submerged
MF, hollow fibre
Tofu processing waste
Glucose
Submerged
GE Water and Process Technologies,
ZW-1 hollow-fibre UF membrane
module, pore size, 0.04 μm
(B) Laboratory and pilot scale AnMBR for methane production
External (P)
Stork, WFFX 0281, UF, MWCO:
Domestic wastewater
100 kDa
External (L)
Weir Envig. Tubular, UF. PES, MWCO: Swine manure
20 kDa
External (P)
Stork, WFFX 0281, UF, MWCO:
Domestic wastewater
100 kDa
External (L)
Weir Envig. Tubular, UF. PES, MWCO: Swine manure
20 kDa
External (P)
Stork, WFFX 0281, UF, MWCO:
Slaughterhouse
100 kDa
wastewater
Submerged (L)
Zenon, capillary, UF, pore size: 0.1 μm Landfill leachate
Submerged (L)
Kubota, PE, FS, pore size: 0.4 μm
Municipal solid
Submerged (L)
FS, PVDF, pore size:0.3 μm
Kraft evaporator
condensate plus
methanol
Permeate
Hydrogen
Temp
flux
production
(◦ C) (L/m2 h) rate (m3 /m3 h) Reference
35
5
2.43–2.56a
[94]
–
57–60
0.64
[95]
35
–
1.02–1.48
[96]
60
23
4.32
11.1
12.81–19.86a
0.02–0.2b
[97]
[98]
37
–
0–55
[99]
–
5–10
2.26
[100]
37
3.5–13
< 30
[101]
37
5–10
2–3
[102]
37
2.22/2.46
50–102
[103]
35
35
37
–
0.5–9
5.6–12.5
–
–
–
[104]
[105]
[106]
Note: L = laboratory; P = Plot.
a Units are L/L/d.
b Unit is mol/L/d.
upgrading techniques such as absorption or adsorption need
high amount of energy and great space. Membrane system,
on the other hand, can perform three separation steps of
carbon dioxide (CO2 ), and H2 S removal and dehydration in
one step, thereby reducing the space needed. In a laboratory
study by Harasimowicz et al., [91] CO2 and methane gases
were efficiently separated with hollow-fibre module membrane, A-2 type (UBE) supplied by UBE Europe GmbH.
Two gas mixtures were used in the experiment with the first
mixture consisting of 50% of both methane and CO2 , while
the second mixture consisted of 68% of CH4 , 30% of CO2 ,
and 2% of H2 S. The result was 89.5% and 93.5% methane
recovery in first and second mixtures, respectively. Polyimides are polymeric membranes that are commonly used
for biogas purification because of its low manufacturing cost
and chemical resistant to gases present.[92] Biogas has also
been produced by encasing the methane-producing bacteria in synthetic and natural membranes.[93] This method
enhanced the rate of biogas production by maintaining a
high cell density in the reactor, and by protecting the cells
from inhibition factors in the medium.[93] Table 2 shows
the application of membrane technology for biogas and
hydrogen production.
Market penetration of membrane technology
AeMBR technology had been commercialized since 1970s
as external loop and later as immersed process for wastewater treatment, desalination, and water purification in countries such as UK/Ireland, France, Germany, Japan, China,
South Korea, Iberia, Benelux, Italy, USA, Canada, and
Mexico, making great contributions in the market. Companies including Ge-Zenon, Kubota, Mitsubishi Rayon,
and Memcor have dominated the membrane market.[107–
110] As of 2009, about 4400 AeMBRs had been installed
in over 200 countries by Kubota, Mitsubishi Rayon, and
Ge-Zenon.[110] The shared percentage of the number of
installed membrane-based biogas plants installed between
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
10
P. Ylitervo et al.
1990 and 2012 in USA, the Netherlands, Austria, Norway
and Germany were 66%, 13%, 13%, 4%, and 4%, respectively. As at 2010, it was reported that 14 AnMBRs that
treated different types of wastes including organic wastes,
food residues, stillage, and wastewaters, were in operation
in Japan. Two AnMBRs were also in operation in the USA
for treating food industry effluents and acid cheese whey,
respectively.[111] For gas permeation using membrane, the
main suppliers are Air Liquid Medal, Air products, GMT
Membrantechnik, Evonik, IGS Generon Membrane technology, MTR Inc., Parker, UBE membranes, and UOP
former Grace.[112]
Membrane materials are usually made of organic polymers, metallic and inorganic materials, e.g. ceramics.
Although metallic and inorganic membranes have properties such as high tolerance to corrosion, abrasion, oxidation,
and better fouling recovery when compared with organic
polymers, polymeric membranes are more frequently used
since they have lower costs. The choice of polymer, however, is often a compromise among the membrane properties
which include hydrophilicity, ease of fabrication, cost, and
robustness. Although, hydrophobic polymers have good
properties including chemical resistance, biocompatibility, low swelling, among others, hydrophobic membranes
were found to be more prone to fouling than hydrophilic
membranes since most reactions between foulants and
membranes are hydrophobic in nature. Therefore, most
hydrophobic membranes are hydrophilized to obtain some
desirable qualities. Generally, polyolefins, polyethersulfone
(PES), and polyvinylidene difluoride (PVDF) are polymers
that are usually favoured.[85,86,104]
Challenges and possible remedies for sustainability of
the MBRs’ future
The many benefits associated with the MBR technology
makes it a reliable and valuable option, favourable over
other waste management techniques. Some of the many
advantages are footprint efficiency (reducing capital costs),
high effluent quality, low-pressure system, high capacity,
and the technology is easy to control. An important feature of the MBR technology is its ability to retain biomass
when using high dilution rates, preventing biomass washout
during the continuous process.[43]
In spite of the many advantages and the fast pace
of development of MBR technology, commercial application is still confronted with some limitations, particularly
for ethanol productions, membrane fouling constituting
the major one. Membrane fouling is a barrier to any
process involving membrane application since it causes
decline in permeate flux and thereby increases energy
input along with operating costs.[113] High particulate
and cell concentrations, along with several antifoaming agents, such as polyoxyethylene, polyoxypropylene,
oleyl ether, polyglycols, and silicon oils severely foul the
membranes.[13,43]
Although anti-fouling measures such as biogas
sparging,[114] agitation and aeration, and membrane vibration are used to reduce fouling rate in membrane operation,
they cannot eliminate membrane fouling completely.[106]
Unrelenting efforts are being made by various researchers
to find ways of keeping membrane fouling to the minimum
to avoid frequent cleaning thereby lengthening membrane
lifespan.[115–119] MBRs techniques producing ethanol
have rarely been scaled up and utilized on industrial scale,
and are not as developed as MBRs in water and wastewater
treatments. Biogas production in MBRs has in contrast been
more successful by the combination of anaerobic digesters
and MBRs utilizing different wastewaters, and has also been
successfully installed on larger scale at several places. It is,
therefore, important to continue transferring the knowledge
from the successful wastewater MBR concepts and evaluate and solve the present problems in ethanol and biogas
in order to improve the performance of the MBRs. Much
work still remains before MBRs for ethanol and biogas
production can be operated at high cell and/or particulate
concentrations.
Conclusion
In MBRs, a biological process, such as, e.g. ethanol fermentation or anaerobic biogas production, is combined with a
membrane separation technique. The membrane can be coupled with the bioreactor, either by submerging it inside the
bioreactor culture, or by placing it in an external loop outside the bioreactor. The MBR allows the cell concentrations
in the bioreactor to be significantly increased. Furthermore,
product inhibition is diminished, product concentration is
amplified, and the separation of product and/or cells after
the process can be simplified. In addition, the productivity
of ethanol fermentation can be increased profoundly when
using an MBR instead of the traditional batch or continuous
processes. Accordingly, MBR holds a recognizable potential for the development of faster and more economically
viable processes to be applied in the production of ethanol
and biogas.
Acknowledgement
This work was financially supported by the Swedish Energy
Agency and the Swedish Research Council.
References
[1] Judd S, Judd C. The MBR book, principles and applications of membrane bioreactors for water and wastewater
treatment. 2nd ed. Oxford: Butterworth-Heinemann; 2011.
[2] Meng F, Chae SR, Drews A, Kraume M, Shin HS, Yang
F. Recent advances in membrane bioreactors (MBRs):
membrane fouling and membrane material. Water Resour.
2009;43:1489–1512.
[3] Visvanathan C, Abeynayaka A. Developments and future
potentials of anaerobic membrane bioreactors (AnMBRs).
Membr Water Treat. 2012;3:31–23.
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
Environmental Technology
[4] Fukura DH. A global perspective of low pressure membranes. Fountain Valley (CA): National Water Research
Institute; 2008. p. 1–8.
[5] Kraume M, Drews A. Review: membrane bioreactors
in wastewater treatment – status and trends. Chem Eng
Technol. 2010;33:1251–1259.
[6] Marrot B, Barrios-Martinez A, Moulin P, Roche N. Industrial wastewater treatment in a membrane bioreactor: a
review. Environ Prog. 2004;23:59–68.
[7] Gander M, Jefferson B, Judd S. Aerobic MBRs for domestic
wastewater treatment: a review with cost considerations.
Sep Purif Technol. 2000;18:119–130.
[8] Judd S. The status of membrane bioreactor technology.
Trends Biotechnol. 2008;26:109–116.
[9] Wisniewski C. Membrane bioreactor for water reuse.
Desalination. 2007;203:15–19.
[10] Woodley JM, Bisschops M, Straathof AJJ, Ottens M. Future
directions for in-situ product removal (ISPR). J Chem
Technol Biotechnol. 2008;83:121–123.
[11] Schügerl K, Hubbuch J. Integrated bioprocesses. Curr Opin
Microbiol. 2005;8:294–300.
[12] Mercier M, Maranges C, Fonade C, Lafforgue-Delorme
C. Yeast suspension filtration: flux enhancement using
an upward gas/liquid slug flow – application to continuous alcoholic fermentation with cell recycle. Biotechnol
Bioeng. 1998;58:47–57.
[13] Cheryan M, Mehaia MA. A high-performance membrane
bioreactor for continuous fermentation of lactose to ethanol.
Biotechnol Lett. 1983;5:519–524.
[14] Visvanathan C, Abeynayaka A. Developments and future
potentials of anaerobic membrane bioreactors. Water Treat.
2012;3:1–23.
[15] Lin H, Liao BQ, Chen J, Wang L, Wang F, Lu X.
New insights into membrane fouling in a submerged
anaerobic membrane bioreactor based on characterization of cake sludge and bulk sludge. Bioresour Technol.
2011;102:2373–2379.
[16] Chung IS, Lee YY. Ethanol fermentation of crude acid
hydrolyzate of cellulose using high-level yeast inocula.
Biotechnol Bioeng. 1985;27:308–315.
[17] Lee W, Lee Y-S, Chang H, Chang Y. A cell retention
internal filter reactor for ethanol production using tapioca
hydrolysates. Biotechnol Tech. 1994;8:817–820.
[18] Brandberg T, Sanandaji N, Gustafsson L, Johan Franzén
C. Continuous fermentation of undetoxified dilute acid
lignocellulose hydrolysate by Saccharomyces cerevisiae
ATCC 96581 using cell recirculation. Biotechnol Prog.
2005;21:1093–1101.
[19] Martinez A, Rodriguez ME, Wells ML, York SW, Preston
JF, Ingram LO. Detoxification of dilute acid hydrolysates
of lignocellulose with lime. Biotechnol Prog. 2001;17:287–
293.
[20] Rivard C, Engel R, Hayward T, Nagle N, Hatzis C, Philippidis G. Measurement of the inhibitory potential and detoxification of biomass pretreatment hydrolysate for ethanol
production. Appl Biochem Biotechnol. 1996;57–58:183–
191.
[21] Carstensen F, Apel A, Wessling M. In situ product recovery: submerged membranes vs. external loop membranes.
J Membr Sci. 2012;394–395:1–36.
[22] Liao BQ, Kraemer J, Bagley D. Anaerobic membrane
bioreactors: applications and research directions. Crit Rev
Environ Sci Technol. 2006;36:489–530.
[23] Pourbafrani M, Talebnia F, Niklasson C, Taherzadeh
M. Protective effect of encapsulation in fermentation of
limonene-contained media and orange peel hydrolyzate. Int
J Mol Sci. 2007;8:777–787.
11
[24] Chang HN, Yoo IK, Kim BS. High density cell culture by membrane-based cell recycle. Biotechnol Adv.
1994;12:467–487.
[25] Finnigan SM, Howell JA. The effect of pulsed flow on
ultrafiltration fluxes in a baffled tubular membrane system.
Desalination. 1990;79:181–202.
[26] Kroner KH, Nissinen V. Dynamic filtration of microbial
suspensions using an axially rotating filter. J Membr Sci.
1988;36:85–100.
[27] Wereley ST, Akonur A, Lueptow RM. Particle–fluid velocities and fouling in rotating filtration of a suspension. J
Membr Sci. 2002;209:469–484.
[28] Howell JA, Field RW, Wu D. Yeast cell microfiltration:
flux enhancement in baffled and pulsatile flow systems. J
Membr Sci. 1993;80:59–71.
[29] Maranges C, Casasnovas C, Lafforgue-Delorme C. Crossflow filtration of Saccharomyces cerevisiae using an
unsteady jet. Biotechnol Tech. 1995;9:649–654.
[30] Redkar SG, Davis RH. Cross-flow microfiltration with
high-frequency reverse filtration. AlChE J. 1995;41:501–
508.
[31] Kobayashi T, Chai X, Fujii N. Ultrasound enhanced crossflow membrane filtration. Sep Purif Technol. 1999;17:
31–40.
[32] Mercier M, Fonade C, Lafforgue-Delorme C. Influence
of the flow regime on the efficiency of a gas-liquid
two-phase medium filtration. Biotechnol Tech. 1995;9:
853–858.
[33] Zhang X, Wang Z, Wu Z, Lu F, Tong J, Zang L. Formation
of dynamic membrane in an anaerobic membrane bioreactor for municipal wastewater treatment. Chem Eng J.
2010;165:175–183.
[34] Li QY, Ghosh R, Bellara SR, Cui ZF, Pepper DS. Enhancement of ultrafiltration by gas sparging with flat sheet
membrane modules. Sep Purif Technol. 1998;14:79–83.
[35] Patel PN, Mehaia MA, Cheryan M. Cross-flow membrane
filtration of yeast suspensions. J Biotechnol. 1987;5:1–16.
[36] Fillaudeau L, Carrère H. Yeast cells, beer composition and
mean pore diameter impacts on fouling and retention during cross-flow filtration of beer with ceramic membranes.
J Membr Sci. 2002;196:39–57.
[37] Starbard N. Beverage industry microfiltration. Hoboken
(NJ): Blackwell Publishing; 2008.
[38] van Reis R, Zydney A. Bioprocess membrane technology.
J Membr Sci. 2007;297:16–50.
[39] Belfort G. Membranes and bioreactors: a technical challenge in biotechnology. Biotechnol Bioeng. 1989;33:1047–
1066.
[40] Inloes DS, Taylor DP, Cohen SN, Michaels AS, Robertson CR. Ethanol production by Saccharomyces cerevisiae
immobilized in hollow-fiber membrane bioreactors. Appl
Environ Microbiol. 1983;46:264–278.
[41] Park TH, Kim IH. Hollow-fibre fermenter using ultrafiltration. Appl Microbiol Biotechnol. 1985;22:190–194.
[42] Lafforgue C, Malinowski J, Goma G. High yeast concentration in continuous fermentation with cell recycle obtained
by tangential microfiltration. Biotechnol Lett. 1987;9:
347–352.
[43] Escobar J, Rane K, Cheryan M. Ethanol production in a
membrane bioreactor. Appl Biochem Biotechnol. 2001;91–
93:283–296.
[44] Giorno L, Drioli E. Biocatalytic membrane reactors: applications and perspectives. Trends Biotechnol. 2000;18:339–
349.
[45] Lee SS, Wang HY. Repeated fed-batch rapid fermentation
using yeast cells and activated carbon extraction system.
Biotechnol Bioeng Symp. 1982;12:221–231.
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
12
P. Ylitervo et al.
[46] Pyle DL, Pritchard M, Scott JA, Howell JA. The concentration of yeast suspensions by crossflow filtration. In: Pyle D,
editor. Separations for biotechnology 2. The Netherlands:
Springer; 1990. p. 63–73.
[47] Thuvander J. Continuous ethanol fermentation using membrane bioreactors (MBR) [Thesis report]. Department of
Chemical Engineering, Lund University, 2012.
[48] Wang LK, Shammas NK, Cheryan M, Zheng Y-M, Zou
S-W. Treatment of food industry foods and wastes by
membrane filtration. In: Wang L Kea, editor. Handbook
of environmental engineering, Volume 13: membrane and
desalination technologies. London: Springer Science and
Business Media; 2008. p. 237–270.
[49] Offeman RD, Robertson GH. Spiral-wound liquid membrane module for separation of fluids and gases. The United
States of America As Represented by The Secretary Of
Agriculture. 2004.
[50] Cheryan M, Escobar J, Shalhevet R. Increasing the efficiency of ethanol production through the use of a membrane
technology. Darby: DIANE Publishing Company; 1994.
[51] Axelsson H. Cell separation, centrifugation. In: Flickinger
MC, editor. Encyclopedia of industrial biotechnology: bioprocess, bioseparation, and cell technology. Hoboken (NJ):
John Wiley & Sons;2009. p. 1–21.
[52] Rakin M, Mojovic L, Nikolic S, Vukasinovic M, Nodovic
V. Bioethanol production by immobilized Sacharomyces
cerevisiae var. ellipsoideus cells. Afr J Biotechnol. 2009;8:
464–471.
[53] Kourkoutas Y, Bekatorou A, Banat IM, Marchant R, Koutinas AA. Immobilization technologies and support materials
suitable in alcohol beverages production: a review. Food
Microbiol. 2004;21:377–397.
[54] Sakurai A, Nishida Y, Saito H, Sakakibara M. Ethanol production by repeated batch culture using yeast cells immobilized within porous cellulose carriers. J Biosci Bioeng.
2000;90:526–529.
[55] Williams D, Munnecke DM. The production of ethanol by
immobilized yeast cells. Biotechnol Bioeng. 1981;23:1813–
1825.
[56] Talebnia F, Taherzadeh MJ. In situ detoxification and continuous cultivation of dilute-acid hydrolyzate to ethanol by
encapsulated S. cerevisiae. J Biotechnol. 2006;125:377–
384.
[57] Najafpour G, Younesi H, Syahidah Ku Ismail K. Ethanol
fermentation in an immobilized cell reactor using Saccharomyces cerevisiae. Bioresour Technol. 2004;92:251–260.
[58] Talebnia F, Niklasson C, Taherzadeh MJ. Ethanol production from glucose and dilute-acid hydrolyzates by encapsulated S. cerevisiae. Biotechnol Bioeng. 2005;90:345–353.
[59] Park JK, Chang HN. Microencapsulation of microbial cells.
Biotechnol Adv. 2000;18:303–319.
[60] Chang HN, Lee WG, Kim BS. Cell retention culture with
an internal filter module: continuous ethanol fermentation.
Biotechnol Bioeng. 1993;41:677–681.
[61] Kang W, Shukla R, Sirkar KK. Ethanol production in a
microporous hollow-fiber-based extractive fermentor with
immobilized yeast. Biotechnol Bioeng. 1990;36:826–833.
[62] Cho T, Shuler M. Multimembrane bioreactor for extractive
fermentation. Biotechnol Prog. 1986;2:53–60.
[63] Lipnizki F, Hausmanns S, Laufenberg G, Field R, Kunz
B. Use of pervaporation-bioreactor hybrid processes in
biotechnology. Chem Eng Technol. 2000;23:569–577.
[64] Ben Chaabane F, Aldiguier AS, Alfenore S, Cameleyre X,
Blanc P, Bideaux C, Guillouet SE, Roux G, Molina-Jouve
C. Very high ethanol productivity in an innovative continuous two-stage bioreactor with cell recycle. Bioprocess
Biosyst Eng. 2006;29:49–57.
[65] Lee WG, Park BG, Chang YK, Chang HN, Lee JS, Park SC.
Continuous ethanol production from concentrated wood
hydrolysates in an internal membrane-filtration bioreactor.
Biotechnol Prog. 2000;16:302–304.
[66] Park BG, Lee WG, Chang YK, Chang HN. Long-term
operation of continuous high cell density culture of
Saccharomyces cerevisiae with membrane filtration and
on-line cell concentration monitoring. Bioprocess Eng.
1999;21:97–100.
[67] Mehaia M, Cheryan M. Ethanol production in a hollow fiber bioreactor using Saccharomyces cerevisiae. Appl
Microbiol Biotechnol. 1984;20:100–104.
[68] Nomura M, Bin T, Nakao SI. Selective ethanol extraction
from fermentation broth using a silicalite membrane. Sep
Purif Technol. 2002;27:59–66.
[69] Ikegami T, Yanagishita H, Kitamoto D, Negishi H, Haraya
K, Sano T. Concentration of fermented ethanol by pervaporation using silicalite membranes coated with silicone
rubber. Desalination. 2002;149:49–54.
[70] Grosz R, Stephanopoulos G. Physiological, biochemical, and mathematical studies of micro-aerobic continuous ethanol fermentation by Saccharomyces cerevisiae.
I: hysteresis, oscillations, and maximum specific ethanol
productivities in chemostat culture. Biotechnol Bioeng.
1990;36:1006–1019.
[71] Cysewski GR, Wilke CR. Rapid ethanol fermentations
using vacuum and cell recycle. Biotechnol Bioeng.
1977;19: 1125–1143.
[72] Maiorella B, Blanch HW, Wilke CR. By-product inhibition effects on ethanolic fermentation by Saccharomyces
cerevisiae. Biotechnol Bioeng. 1983;25:103–121.
[73] Roffler SR, Blanch HW, Wilke CR. In situ extractive
fermentation of acetone and butanol. Biotechnol Bioeng.
1988;31:135–143.
[74] Barton WE, Daugulis AJ. Evaluation of solvents for extractive butanol fermentation with Clostridium acetobutylicum
and the use of poly(propylene glycol) 1200. Appl Microbiol
Biotechnol. 1992;36:632–639.
[75] Barboza M, Almeida R, Hokka C. Kinetic studies of
clavulanic acid recovery by ion exchange chromatography.
Bioseparation. 2001;10:221–227.
[76] Cartón A, Benito GG, Rey JA, de la Fuente M. Selection of
adsorbents to be used in an ethanol fermentation process.
Adsorption isotherms and kinetics. Bioresour Technol.
1998;66:75–78.
[77] Jones RA, Tezel FH, Thibault J, Tolan JS. Bio-ethanol
production to be blended with gasoline: improvements in
energy use by adsorption. Int J Energy Res. 2007;31:1517–
1531.
[78] Pitt WW, Haag GL, Lee DD. Recovery of ethanol from
fermentation broths using selective sorption–desorption.
Biotechnol Bioeng. 1983;25:123–131.
[79] Ylitervo P, Franzén C, Taherzadeh MJ. Impact of furfural
on rapid ethanol production using a membrane bioreactor.
Energies. 2013;6:1604–1617.
[80] Letourneau F, Villa P. Saccharomyces yeast growth on
beet molasses effects of substrate concentration on alcohol
toxicity. Biotechnol Lett. 1987;9:53–58.
[81] Wang FS, Sheu JW. Multiobjective parameter estimation problems of fermentation processes using a
high ethanol tolerance yeast. Chem Eng Sci. 2000;55:
3685–3695.
[82] Grobben NG, Eggink G, Petrus Cuperus F, Huizing HJ.
Production of acetone, butanol, and ethanol (ABE) from
potato wastes: fermentation with integrated membrane
extraction. Appl Microbiol Biotechnol. 1993;39:494–
498.
Downloaded by [Julius Akinbomi] at 23:34 09 July 2013
Environmental Technology
[83] Frank GT, Sirkar KK. An integrated bioreactor-separator:
in situ recovery of fermentation products by a novel
dispersion-free solvent extraction technique. Biotechnol
Bioeng Symp Ser. 1986;17:303–316.
[84] Lee KR, Teng MY, Hsu TN, Lai JY. A study on pervaporation of aqueous ethanol solution by modified polyurethane
membrane. J Membr Sci. 1999;162:173–180.
[85] Smith AL, Stadler LB, Love NG. Perspectives on anaerobic membrane bioreactor treatment of domestic wastewater: a critical review. Bioresour Technol. 2012;122:
149–159.
[86] Lin H, Peng W, Zhang M, Chen J, Hong H, Zhang Y. A
review on anaerobic membrane bioreactors: applications,
membrane fouling and future perspectives. Desalination.
2013;314:169–188.
[87] Stuckey DC. Recent developments in anaerobic membrane
reactors. Bioresour Technol. 2012;122:137–148.
[88] Das D. Advances in biohydrogen production processes:
an approach towards commercialization. Int J Hydrogen
Energy. 2009;34:7349–7357.
[89] Angenent LT, Karim K, Al-Dahhan MH, Wrenn BA,
Domiguez-Espinosa R. Production of bioenergy and biochemicals from industrial and agricultural wastewater.
Trends Biotechnol. 2004;22:477–485.
[90] Kim IS, Hwang MH, Jang NJ, Hyun SH, Lee ST.
Effect of low pH on the activity of hydrogen utilizing methanogen in biohydrogen process. Int J Hydrogen
Energy. 2004;29:1133–1140.
[91] Harasimowicz M, Orluk P, Zarkrzewska-Trznadel G,
Chimielewski AG. Application of polyimide membranes
for biogas purification and enrichment. J Hazard Mater.
2007;144:698–702.
[92] Baker R, Okhandwala K. Natural gas processing with membranes: an overview. Ind Eng Chem Res. 2008;47:2109–
2121.
[93] Youngsukkasem S, Rakshit SK, Taherzadeh MJ. Biogas
production by encapsulated methane-producing bacteria.
Bioresources. 2012;7:56–65.
[94] Lee DY, Li YY, Noike T. Continuous hydrogen production
by anaerobic mixed microflora in membrane bioreactor.
Bioresour Technol. 2009;100:690–695.
[95] Oh S, Iyer P, Bruns MA, Logan E. Biological hydrogen production using a membrane bioreactor. Biotechnol Bioeng.
2004;87:119–127.
[96] Lee KS, Lin PJ, Fangchiang K, Chang JS. Continuous
hydrogen production by anaerobic mixed microflora using
a hollow-fiber microfiltration membrane bioreactor. Int J
Hydrogen Energy. 2007;32:950–957.
[97] Kim MS, Lee DY. Continuous hydrogen production from
tofu processing waste using anaerobic mixed microflora
under thermophilic conditions. Int J Hydrogen Energy.
2011;36:8712–8718.
[98] Shen L, Bagley DM, Liss SN. Effect of organic loading
rate on fermentative hydrogen production from continuous stirred tank and membrane bioreactors. Int J Hydrogen.
2009;34:3689–3696.
[99] Saddoud A, Ellouze M, Dhouib A, Sayadi S. A comparative
study on the anaerobic membrane bioreactor performance
during the treatment of domestic wastewaters of various
origins. Environ Technol. 2006;27:991–999.
[100] Zhang J, Padmasiri SI, Fitch M, Norddahl B, Raskin L, Morgenroth E. Influence of cleaning frequency and membrane
history on fouling in an anaerobic membrane bioreactor.
Desalination. 2007;207:153–166.
13
[101] Saddoud A, Ellouze M, Dhouib A, Sayadi S. Anaerobic
membrane bioreactor treatment of domestic wastewater in
Tunisia. Desalination. 2007;207:205–215.
[102] Padmasiri SI, Zhang J, Fitch M, Norddahl B, Morgenroth
E, Raskin L. Methanogenic population dynamics and performance of an anaerobic membrane bioreactor (AnMBR)
treating swine manure under high shear conditions. Water
Resour. 2007;41:134–144.
[103] Saddoud A, Sayadi S. Application of acidogenic fixed-bed
reactor prior to anaerobic membrane bioreactor for sustainable slaughterhouse wastewater treatment. J Hazard Mater.
2007;149:700–706.
[104] Bohdziewicz J, Neczaj E, Kwarciak A. Landfill leachate
treatment by means of anaerobic membrane bioreactor.
Desalination. 2008;221:559–565.
[105] Trzcinski AP, Stuckey DC. Treatment of municipal solid
waste leachate using a sumerged anaerobic membrane
bioreactor at mesophilic and psychrophilic temperatures:
analysis of recalcitrants in the permeate using GC-MS.
Water Resour. 2010;44:671–680.
[106] Xie K, Lin HJ, Mahendran B, Bagley DM, Leung KT,
Liss SN, Liao BQ. Performance and fouling characteristics of a submerged anaerobic membrane bioreactor for
kraft evaporator condensate treatment. Environ Technol.
2010;31:511–21.
[107] Wang Z, Wu Z, Mai S, Yang C, Wang X, An Y,
Zhou Z. Research and applications of membrane bioreactors in China: progress and prospect. Sep Purif Technol.
2008;62:249–263.
[108] Frost and Sullivan. Strategic analysis of the European
membrane bioreactor markets. Market Research; 2005.
[109] Frost and Sullivan. US and Canada membrane bioreactor
markets. Market Research; 2004.
[110] Judd S, Judd C. The MBR book. Amsterdam: Elsevier;
2010.
[111] Kanai M, Ferre V, Wakahara S, Yamamoto T, and Moro
M. A novel combination of methane fermentation and
MBR/kubota submerged anaerobic membrane bioreactor
process. Desalination. 2010;250:860–967.
[112] Scholz M, Melin T, Wessling M. Transforming biogas into
biomethane using membrane technology. Renew Sustain
Energy Rev. 2013;17:199–212.
[113] He Y, Bagley DM, Leung KT, Liss SN, Liao B-Q. Recent
advances in membrane technologies for biorefining and
bioenergy production. Biotechnol Adv. 2012;30:817–858.
[114] Wang Z, Yu H, Ma J, Zheng X, Wu Z. Recent advances
in membrane bio-technologies for sludge reduction and
treatment. Biotechnol Adv. (in press).
[115] Howell JA. Future research and developments in the
membrane field. Desalination. 2002;144:127–131.
[116] Howell JA. Future of membranes and membrane reactors
in green technologies and for water reuse. Desalination.
2004;162:1–11.
[117] Lesjean B, Rosenberger S, Schrotter J-C, Recherche
A. Membrane-aided biological wastewater treatment
– an overview of applied systems. Membr Technol.
2004;2004:5–10.
[118] Yang Q, Chen J, Zhang F. Membrane fouling control in
a submerged membrane bioreactor with porous, flexible
suspended carriers. Desalination. 2006;189:292–302.
[119] Melin T, Jefferson B, Bixio D, Thoeye C, De Wilde W,
De Koning J, van der Graaf J, Wintgens T. Membrane
bioreactor technology for wastewater treatment and reuse.
Desalination. 2006;187:271–282.
Was this manual useful for you? yes no
Thank you for your participation!

* Your assessment is very important for improving the work of artificial intelligence, which forms the content of this project

Related manuals

Download PDF

advertisement