MATERIALS AND METHODS Materials2,4,6-tripyridyltriazine (TPTZ), DPPH, gallic acid, quercetin, beta-carotene were purchased from Sigma-Aldrich (MO, USA), ferric chloride, shells from four species of legumes, methanol, ethanol. All other reagents were analytical grades. Preparation of sampleShells from four species of legumes were: Soybean (G. max) namely as KDL collected from Bogor, red kidney bean (P vulgaris) as sample MRH from Garut, Bogor peanut (V. subterranea) as sample BGR from Sukabumi, peanut (A. hypogaea) as sample TNH from Kendal-Semarang, were thoroughly washed with tap water, wet sortation, cut, dried, and grinded into powder.ExtractionA total of 300 g of powdered samples were extracted by reflux using increasing gradient polarity solvents. The n-hexane extract was repeated three times. The remaining residue was then extracted three times with ethyl acetate. Finally, the remaining residue was extracted three times with ethanol. Hence, there were four n-hexane extracts (namely KDL1, MRH1, BGR1, and TNH1), four ethyl acetate extracts (KDL2, MRH2, BGR2, and TNH2) and four ethanolic extracts (KDL3, MRH3, BGR3, and TNH3).
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