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Following an initial cleanup step on the AB-8 macroporous resin, highspeed countercurrent chromatography (HSCCC) was successfully applied for the first time to the isolation and purification of four flavonoids from Flos Gossypii. HSCCC was performed with a two-phase solvent system composed of chloroformmethanol-isopropanol-water (5:5:1:3, v/v) adding 0.4% phosphoric acid in the aqueous stationary phase. The separation yielded quercetin (5.6 mg), quercetin-3 0 -O-D-glucoside (7.2 mg), quercetin-3-O-b-D-glucoside (16 mg), quercetin-7-O-b-Dglucoside (8.3 mg), from 100 mg of the crude extract in a one step separation. The structure identification was made by 1 H NMR, 13 C NMR.

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