Toxicarioside N (TN) is a strophanthidol cardenolide isolated from the seeds of Antiaris toxicaria (Pers.) Lesch. (Moraceae) and showed cytotoxic activity against cancer cells. The aim of this study was to investigate the cytotoxic activity and possible molecular mechanism of TN against human gastric cancer BGC-823 cells using cell counting kit-8 (CCK-8), flow cytometry and western blot assays. The results of CCK-8 assay indicated that TN showed cytotoxic activity against BGC-823 cells with a dose-dependent manner (IC50=24.87 μg/ml). The results of flow cytometry assay exhibited that TN induced apoptosis of BGC-823 cells, suggesting that the cytotoxic activity of TN against BGC-823 cells was related to apoptosis. The results of western blot assay revealed that TN-induced apoptosis in BGC-823 cells was characterized by down-regulations of phosphorylated inhibitors of NF-κB-α (p-IκBα), phosphorylated IκB kinase β (p-IKKβ), IKKβ and nuclear NF-κB p65 and up-regulation of IκBα, and also featured by up-regulations of Fas, FasL, Fas-associated protein with death domain, cleaved-caspase-8 (c-caspase-8), c-caspase-10, c-caspase-3. In conclusion, TN showed cytotoxic activity against BGC-823 cells, and the cytotoxic activity was related to apoptosis. The mechanism of action was associated with NF-κBmediated activation of Fas/FasL pathway. Thus, TN may be a promising candidate drug for treatment of gastric cancer. This, however, needs to be further investigated to explore the actual potentials of TN.