Abstract: OBJECTIVE To establish an efficient method for extracting and isolating the alkaloids from strychni semen. METHODS With the content of brucine and strychnine as index, the ultrasonic assisted extraction process was optimized by uniform design method, and high-speed counter-current chromatography(HSCCC) was employed to separate the alkaloids. RESULTS The optimum condition of extraction was as follows: ratio of solid to liquid was 1∶10, ethanol concentration was 50%(pH=5), ultrasonic power was 600 W, and the extraction time was 60 min; the two-phase solvent system of HSCCC composed of n-hexane-ethyl acetate-methanol-water(3∶5∶3∶5) was selected, brucine with the purity of 94.1% and strychnine with the purity of 92.5% were obtained. CONCLUSION The optimum extraction method is economical and feasible. HSCCC is suitable for the isolation of alkaloids from strychni semen.