Cervical cancer is fourth most common fatal cancer in women worldwide. Lupeol is a dietary triterpenoid and has shown itsanticancer efficacy against various cancer types with selectivity in targeting cancer cells. In the present study, anticancerefficacy and mechanism of action of a phytochemical, lupeol, in human cervical carcinoma (HeLa) cells has been examined.The anticancer efficacy of lupeol was assessed by trypan blue cell counting, annexin Vassay, cell cycle analysis, expressionof apoptotic proteins by RT-PCR and Western blotting and assessment of mitochondrial ROS generation by mitosox andmitotracker assays. Our results demonstrated that lupeol decreased cell proliferation and viability of HeLa cells significantly(p\0.001). Lupeol induced S-phase cell cycle arrest and also decreased the expression of S-phase Cyclins and CDKs andincreased the expression of cyclin-dependent kinase inhibitors, p21 at transcriptional and translational level. Further, lupeolinduced apoptosis and increased the expression of apoptosis markers such as cleaved PARP and Bax:Bcl-2 ratio. Furthermore,mitosox and mitotracker dye incubation followed by FACS analysis showed an increase in mitochondrialsuperoxide generation and reduction in healthy mitochondrial mass. These results suggest that lupeol could be an effectivechemotherapeutic agent against cervical carcinoma due to its growth inhibitory activity through induction of S-phase cellcycle arrest and apoptosis.