ISOLATION OF FIVE BIOACTIVE COMPONENTS FROM EUPATORIUM ADENOPHORUM SPRENG USING STEPWISE ELUTION BY HIGH-SPEED COUNTER-CURRENT CHROMATOGRAPHY.

Semi-preparative high-speed counter-current chromatography (HSCCC) was successfully performed for isolation and purification of caffeic acid 4 bioactive flavonoids from Eupatorium adenophorum Spreng using stepwise elution with a pair of two-phase solvent systems composed of ethyl acetate-methanol-water at the volume ratios of 10: 1: 10 and 5: 1: 5 (v/v). From 378.5 mg of crude extract 24.1 mg of caffeic acid, 6.7 mg of 4'-methyl quercetagetin 7-O-(6"-O-E-caffeoylglucopyranoside), 6.5 mg of quercetagetin 7-O-(6"-O-acetyl-β-D-glucopyranoside), 31.8 mg of eupalitin 3-O-β-D-galactopyranoside and 36.7 mg of eupalitin were obtained with the purities of 96.0%, 91.2%, 82.3%, 95.1% and 85.6%, respectively. The structures of the separated compounds were identified by EI-MS, (1)HNMR and (13)CNMR.