METHOD FOR DETERMINATION OF CARBOXYLIC

METHOD FOR DETERMINATION OF CARBOXYLIC ACIDS IN GREEN COFFEE BEANS BY
RP-HPLC
Cíntia S. G. Kitzberger, M. Brígida S. Scholz, Marta T. Benassi. Instituto Agronômico do
Paraná, Área de Ecofisiologia Vegetal, Rodovia Celso Garcia Cid, km 375 CEP 86047-902, +55
43 3376 2396, Londrina, PR, Brazil, and Universidade Estadual de Londrina, Departamento de
Pós Graduação em Ciência de alimentos, Londrina, PR, Brazil.
The aim of this study was to validate the methodology to determine carboxylic acids contents in
green coffee beans using RP-HPLC/Diode array detection. Coffee was extracted by heating in
water (80°C, 30 min), cooling, pH adjustment to 7.0 and filtration. Solid phase extraction was
carried out in cartridge strongly anion exchange resin (Dowex 1X4 200 mesh). The resin
conditioning and sample cleaning was conducted in water. The acids were eluted with 1 M HCl,
submitted to cleaning in water and filtered through a nylon membrane (0.45 M). A C18 ACE 5
column (250 x 4.6 mm id, 5 µm), detection at 210 nm and isocratic elution of 0.005 N H2SO4 (pH
2.5) were used. The flow rate gradient was: 0-2 min at 0.7 mL min-1, 2-15 min at 0.4 mL min-1
and 0.7 mL min-1 to the end. The oven temperature and sample tray were 30°C and 5°C,
respectively. Identification and quantification was based on comparison with standards. The
determinations were performed in the linear range of 125-2000 mg 100g-1 for quinic and citric
acids, and 50-800 mg 100g-1 for malic acid. The detection limits were 0.03, 0.11 e 0.03 mg
100g-1 and quantification limits were 0.10, 0.32 e 0.10 mg 100g-1 to quinic, malic and citric acids,
respectively. The recoveries ranged from 84, 77.5 e 84%, respectively, for quinic, malic and
citric. Good agreement of results with literature values was verified and the method was
considered efficient to determine carboxylic acids in green coffee.
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