Abstract: Oldenlandia diffusa, a Traditional Chinese Medicine (TCM) commonly used for curing hepatoma and hepatitis, showed promising anti-tumor effects on a number of tumor cell lines. In this study, Stearic Acid (C18H36O2, SA) was isolated and identified from O. diffusa as one of the active components using a bioassay-guided method. SA showed significant inhibitory effects on the growth of human hepatoma cell lines, HepG2 and Hep3B, but not on human normal liver cell line, WRL-68. Both HepG2 and Hep3B cells were more sensitive towards SA when comparing with other fatty acids. Mechanistic study showed that Pa induced apoptosis on both HepG2 and Hep3B cells with induction of sub-G1 population, DNA fragmentation, changing of mitochondrial transmembrane potential and activation of caspase precursors observed. This study firstly reports that SA is an active component isolated from O. diffusa with potential for hepatoma treatment.