Oxymatrine, one of the main active components of extracts from the dry roots of Sophora flavescens, has been reported to possess anticancer activities in vitro and in vivo However, the precise mechanism involved remains largely unknown. The present study is conducted to investigate the anticancer activity and the underlying mechanisms of oxymatrine in human hepatoma cells (Hep-G2 and SMMC-7721) in vitro and in vivo Hep-G2 and SMMC-7721 cells were treated by oxymatrine and subjected to methyl thiazolyl tetrazolium analysis, Hoechst 33342 staining, annexin V/propidium iodide double staining, reverse transcription polymerase chain reaction, and Western blot analysis. In addition, SMMC-7721 xenograft tumors were established in male nude BALB/c mice, and oxymatrine was intravenously administered to evaluate the anticancer capacity in vivo Our results showed that oxymatrine inhibited the proliferation and induced apoptosis of Hep-G2 and SMMC-7721 cells in a dose-dependent manner in vitro Furthermore, the RNA and protein expression of Bax and caspase 3 levels were significantly upregulated, whereas the expression of Bcl-2 was downregulated. These protein interactions may play a pivotal role in the regulation of proliferation and apoptosis. More importantly, our in vivo studies showed that administration of oxymatrine decreased tumor growth in a dose-dependent manner. Immunohistochemistry analysis demonstrated an increase of Bax and caspase 3 and a decrease of Bcl-2 in tumor tissues following oxymatrine treatment which are consistent with the in vitro results. Taken together, our findings indicated that oxymatrine can inhibit cell proliferation and induce apoptosis of human hepatoma Hep-G2 and SMMC-7721 cells and might offer a therapeutic potential advantage for human hepatoma chemoprevention or chemotherapy.