Preparative Isolation and Purification of Calycosin and Formononetin from Astragalus membranaceus Bge. var. mongholicus (Bge.) Hsiao by High-Speed Counter-Current Chromatography

Preparative high-speed counter-current chromatography (HSCCC) was used for the separation of calycosin and formononetin from the ethyl acetate extract of Astragalus membranaceus Bge. var. mongholicus (Bge.) Hsiao. Calycosin was obtained by HSCCC with a binary phase solvent system composed of n-hexane-chloroform-methanol-water (1.5: 3 : 3 : 2, v/v). Then, formononetin was obtained with a system composed of n-hexane-chloroform-methanol water (4: 4:5: 4, v/v). The mobile phase was the lower phase and operated at a flow-rate of 2.0 mL/min, while the apparatus rotated at 800 r/min. High performance liquid chromatographic analysis of calycosin and formononetin revealed that their purities were over 95%. HSCCC is a useful method for the separation of natural products.