Objective: To analyze the effects of Clostridium difficile toxin B (TcdB) on the proliferation and apoptosis of colon cancer cell line SW480 and the possible mechanisms related to cell apoptosis. Methods: SW480 cells were treated with different concentrations of TcdB. Cell proliferation was detected by MTT assay. Cell apoptosis and mitochondrial membrane potential were measured with flow cytometry. Results: TcdB significantly inhibited the proliferation of SW480 cells in a time-concentration dependent manner and the inhibition rate reached 46.36% at 48 h. Flow cytometry results showed that TcdB could induce the apoptosis of SW480 cells in a time-concentration dependent manner and a 20.83% apoptosis rate was induced by 800 ng/ml of TcdB at 48 h. Conclusions TcdB could inhibit the proliferation and induce the apoptosis of colon cancer SW480 cells, and the possible mechanisms might be relate to the initiation of mitochondrial apoptosis pathway.