Preparative isolation and purification of bergapten and imperatorin from the medicinal plant Cnidium monnieri using high-speed counter-current chromatography by stepwise increasing the flow-rate of the mobile phase

A high-speed counter-current chromatography (HSCCC) method was developed for the preparative separation and purification of bergapten and imperatorin from the Chinese medicinal plant Cnidium monnieri (L.) Cusson. The crude extract was obtained by extraction with ethanol from the dried fruits of Cnidium monnieri (L.) Cusson under sonication. Preparative HSCCC with a two-phase solvent system composed of n-hexane-ethyl acetate-ethanol-water (5:5:5:5, v/v/v/v) was successfully performed by increasing the flow-rate of the mobile phase stepwise from 1.0 to 2.0 ml min(-1) after 180 min. The components purified and collected were analyzed by high-performance liquid chromatography. The method yielded 45.8 mg of bergapten at 96.5% purity and 118.3 mg of imperatorin at 98.2% purity from 500 mg of the crude extract in a single run. The recoveries of bergapten and imperatorin were 92.1 and 93.7%, respectively.