Two prenylated biflavonoids, podoverines B-C, were isolated from the dried roots and rhizomes of Sinopodophyllum emodi using a Sephadex LH-20 column (SLHC) and high-speed counter-current chromatography (HSCCC). The 95% ethanol extract was partitioned with ethyl acetate in water. Target compounds from the ethyl acetate fraction were further enriched and purified by the combined application of SLHC and HSCCC. n-Hexane-ethyl acetate-methanol-water (3.5:5:3.5:5, v/v) was chosen as the two phase solvent system. The flow rate of mobile phase was optimized at 2.0 mL·min(-1). Finally, under optimized conditions, 13.8 mg of podoverine B and 16.2 mg of podoverine C were obtained from 200 mg of the enriched sample. The purities of podoverines B and C were 98.62% and 99.05%, respectively, as determined by HPLC. For the first time, podoverins B and C were found in the genus Sinopodophyllum. Their structures were determined by spectroscopic methods (HR-ESI-MS, ¹H-NMR, (13)C-NMR, HSQC, HMBC). Their absolute configurations were elucidated by comparison of their experimental and calculated ECD spectra. The cytotoxic activities were evaluated against MCF-7 and HepG2 cell lines. The separation procedures proved to be practical and economical, especially for trace prenylated biflavonoids from traditional Chinese medicine.
Keywords: Sephadex LH-20 column chromatography; Sinopodophyllum emodi; cytotoxic activity; high-speed counter-current chromatography; prenylated biflavonoid.