Pinus massoniana bark extract (PMBE) contains a variety of flavonoids whose antioxidant properties have been confirmed in vitro. This study was undertaken to evaluate the cytotoxic effects and the mechanism of cell death on the PMBE-treated human cervical cancer cell line, HeLa. PMBE treatment led to cell growth inhibition in a dose- and time-dependent manner, and PMBE-induced apoptosis was confirmed by DAPI staining, TUNEL assays and sub-G1 phase accumulation. Cell cycle was also arrested in G2/M phase. Immunoblotting analysis showed that cytochrome c was released, the protein expression of Bax was increased, the protein expression of Bcl-2 was down-regulated and caspase-9 and -3 were activated in PMBE-treated HeLa cells. Taken together, PMBE inhibited proliferation, induces apoptosis and causes cell cycle arrest in HeLa cells, indicating that PMBE may be a potential therapeutic agent for cancer.