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

Abstract

Semi-preparative high-speed countercurrent chromatography (HSCCC) was successfully performed for isolation and purification of caffeic acid and 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 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, FAB-MS, ¹HNMR, and ¹³CNMR.

Keywords:

• eupalitin
• eupalitin 3-O-β-D-galactopyranoside
• Eupatorium adenophorum Spreng
• 4′-methyl quercetagetin 7-O-(6″-O-E-caffeoylglucopyranoside)
• quercetagetin 7-O-(6″-O-acetyl-β-D-glucopyranoside)
• stepwise high-speed countercurrent chromatography

ACKNOWLEDGMENTS

Financial support by Special Fund for Agro-Scientific Research in the Public Interest (project 200803022 & 201103027) and the Fundamental Research Fund for Safety Control Analysis of Food, Organisms and Environment from Beijing Education Committee and the State Key Laboratory of Chemical Resource Engineering (Project CRE-B-2008104) are gratefully acknowledged.