During the screening of a traditional Chinese folk herb library against Hep G2 and Hep3 B cell lines, the Et OAc extract from the Tibetan medicine, Caragana tibetica（CT-Et OAc） exhibited potential anti-hepatocellular carcinoma（anti-HCC） activity. HPLC-based activity profiling was performed for targeted identification of anti-HCC activity from CT-Et OAc by MS-directed purification method. CT-Et OAc was separated by time-based fractionation for further anti-HCC bioassay by a semipreparative HPLC column（150 mm × 10 mm i.d., 5 μm） with a single injection of 5 mg. Bioassay-guided and ESIMS-directed large scale purification was performed with a single injection of 400 mg of CT-Et OAc by peak-based fractionation. A 1.4-mm heavy wall micro NMR tube with z-gradient was used to measure one and two dimensional NMR spectra for the minor or trace amounts of components of the extract. Two active compounds could be elucidated as naringenin chalcone（CT-1） and 3-hydroxy-8, 9-dimethoxypterocarpan（CT-2） relevant to anti-HCC effects for the Et OAc extract of C. tibetica rapidly and unambiguously by this protocol.
