Thiosulfinates, a substance of Allium tuberosum L., is a known folk medicine that has been extensively used in diet to treat diseases. In the present study, we have evaluated the effect of thiosulfinates from Allium tumberosum L. on proliferation of metastasis (DU145) and primary malignant tumor (RC-58T/h/SA#4)-derived human prostate cancer cells. Thiosulfinates decrease viable cell numbers in a dose- and time-dependent manner and induce apoptosis. The apoptosis induced by thiosulfinates is associated with the activation of initiator caspase-8, and -9, and the effector caspase-3. Thiosulfinates stimulated Bid cleavage, indicating that the apoptotic action of caspase-8-mediated Bid cleavage leads to the activation of caspase-9. Thiosulfinates decreased the expression of the anti-apoptotic protein Bcl-2, and increased the expression of the pro-apoptotic protein Bax. Thiosulfinates also increased the expression of AIF, a caspase-independent mitochondrial apoptosis factor, in RC-58T/h/#4 cells and induced DNA fragmentation and chromatin condensation. These results indicate that thiosulfinates from Allium tuberosum L. inhibit cell proliferation by inducing apoptosis in RC-58T/h/#4 cells which may be mediated via both caspase-dependent and caspase-independent pathways.