Abstract: To investigate the effects of mitochondrial in apoptosis of HepG2 cells induced by reseveratrol. Mitochondrial proteins were prepared from HepG2 cells treated by resveratrol and control.Differentially expressed proteins were separated by two-dimensional electrophoretic, and analyzed by Time of Flight Mass Spectrometry. A effective approach was established to extract mitochondria. Four significant difference proteins were identified by Time of Flight Mass Spectrometry. Kinesin protein and CENP-E were down-regulated in HepG2 cells when treated with resveratrol, which implicated to be related with the function of cell cycle and cell cytoskeleton. Mitochondrial ribosomal protein L7/L12 was highly expressed in HepG2 cells when treated with resveratrol, which suggested the relationship of mitochondrial dysfunction and resveratrol induced apoptosis in HepG2 cells.