OBJECTIVE： To analyze and identify the metabolites of tuberostemonine in rats plasma，urine and feces，and to clarify metabolic pathway of tuberostemonine. METHODS： Rats were randomly divided into blank group （0.3％ carboxymethyl cellulose） and medication group （tuberostemonine 50 mg/kg，i.g.），with 6 rats in each group. The plasma of rats was collected 0.25，0.5，1，2，4，6，12 h after intragastric administration to prepare samples. Urine and feces were collected 0-12 h and 12-24 h after administration to prepare samples. UPLC-Q-TOF/MS and software of Triple TOFTM high resolution mass spectrometry were used to analyze and identifiy chemical structure of metabolites in samples. RESULTS： A total of 4 compounds were detected in rat plasma，including one prototype compound and 3 metabolites. 4 metabolites were detected in urine and 2 metabolites in feces. Phase Ⅰ metabolites of tuberostemonine mainly included hydration and hydroxylation. Phase Ⅱ metabolites were not found. CONCLUSIONS： Hydration and hydroxylation are the major metabolic transformation forms of tuberostemonine in rats in vivo.