Preparative Isolation of Antioxidant Flavonoids from Small Black Soybeans by Centrifugal Partition Chromatography and Sequential Solid-Phase Extraction

초록

Unlike other members of the leguminous family that are generally used as food, Rhynchosia volubilis, a small soybean with a black seed coat, is used as a folk medicine in Korea. Using an on-line high performance liquid chromatography-2,2MODIFIER LETTER PRIME-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) radical cation screening system, one peak in the ethyl acetate fraction exhibited outstanding antioxidant capacity. To purify and determine the antioxidant compounds associated with this peak, we developed a centrifugal partition chromatography (CPC) method. The ethyl acetate fraction (500 mg) was subjected to CPC using a two-phase solvent system of n-hexane-ethyl acetate-ethanol-water (0.1:8:2:7, v/v) with a dual-mode CPC (descending-ascending order) in a bid to separate glycitin (3.1 mg), daidzin (14.4 mg), 6MODIFIER LETTER PRIMEMODIFIER LETTER PRIME-O-acetyl genistin (6.9 mg), 6MODIFIER LETTER PRIMEMODIFIER LETTER PRIME-O-acetyl daidzin (5.3 mg), and a mixture of epicatechin and genistin. The mixture was further separated through solid-phase extraction to yield epicatechin (6.8 mg) and genistin (23.5 mg), respectively. Structural identification of the R. volubilis flavonoids was performed by NMR and ESI-MS.

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