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Oxytropis falcata  Bunge, known as the “king of herbs” in Tibetan medicine, is used for treatment of hyperpyrexia, pain, wounds, inflammation, and anthrax. However, it is difficult to efficiently isolate compounds with high purity from  O. falcata  because of the complexity of traditional Tibetan medicines. In this study, the 80% ethanol elution fraction from extract by AB-8 macroporous resin column chromatography was demonstrated to have anticancer activity on human hepatoma SMMC-7721 cells in vitro. Then, a high-speed counter-current chromatography (HSCCC) method was successfully established for separation of compounds by using hexane–ethyl acetate–methanol–water (10:4:10:10,  v / v / v / v ) as the solvent system. Five flavonoids (7-hydroxyflavonone [ 1 ], 5,7-dihydroxy-4′-methoxy flavonol [ 2 ], 5,7-dihydroxyflavanone [ 3 ], 2′,4′-dihydroxychalcone [ 4 ], and 2′,4′-dihydroxydihydrochalcone [ 5 ]) were obtained in one-step separation with purities of 97.7%, 98.1%, 98.3%, 99.0%, and 98.3%, respectively. Finally, anticancer activities against the growth of SMMC-7721 cells of 5 flavonoids were confirmed. The IC 50  values of the separated compounds were 213.45 μg/mL, 197.74 μg/mL, 375.16 μg/mL, 17.44 μg/mL, and 136.83 μg/mL in 24 h, respectively. The present study provided a basis for further development and utilization of this medicinal herb as a source of a new potential anticancer agent.

Efficient separation of five flavonoids from Oxytropis falcata bunge by high-speed counter-current chromatography and their anticancer activity