High-speed counter-current chromatography: an effective method for the isolation of flavonoids and profisetinidin from fustic (Cotinus coggygria Scop.)

Abstract

While chromatographic methods such as column chromatography and semi-preparative HPLC are widely employed during purifications of plant extracts, high-speed counter-current chromatography (HSCCC) is only occasionally utilized. This method based on liquid-liquid partition is advantageous as it avoids the loss of analytes due to irreversible adsorption. The present research focuses on the analysis of the diethyl ether-soluble fraction of a crude extract from Cotinus coggygria (Anacardiaceae) wood. After preliminary separation through vacuum liquid chromatography over RP-18 material (solvent: acetonitrile/water gradient), two of the resulted fractions (A-3, CH3CN/H2O 10/90, v/v and A-8, CH3CN/H2O 36/64, v/v) were purified on Sephadex LH-20 (solvent: methanol), yielding enriched fractions; four thereof were selected for HSCCC separations. An optimized solvent system was developed, comprising a mixture of hexane, ethyl acetate, methanol and water (1/2.5/1/1; all v/v). For separations, a P.C. Inc. (Potomac, USA; HSCCC multilayer coil, series 690) instrument was used. The system was operated in the „tail to head“ mode, with the upper solvent layer as mobile phase. HSCCC using the above solvent system afforded the isolation of seven compounds: fustin (1), dihydroquercetagetin (2), epifisetinidol-(4β→8)-(+)-catechin (3), fisetin (4), quercetin (5), butein (6), and the new natural compound C-3/C-3“ dimer of 3′,4′,7-trihydroxyflavanone (7). Their structure was elucidated by 1D (1H NMR, 13C NMR) and 2D (HSQC, HMBC) NMR experiments. The here described HSCCC solvent system is suitable for the separation of medium-polarity phenolic compounds from C. coggygria wood.