Abstract: Traditional medicines or herbal formulations can serve as the source of potential new chemotherapeutic drugs. Nowadays novel plant derived natural products have been designed to determine biological structure-activity relationships. A new family of betulinic acid derivatives were synthesized and the structure-activity relationship of the compounds was initially evaluated by comparing their in vitro cytotoxicity against 4 human cancer cell lines U937, HepG2, HT29, MCF-7 and non cancerous human PBMC. Betulinic Acid derivative, BC-SM-05 was the most potent inducer of apoptosis and the most effective inhibitor of cell growth and proliferation with IC50 value 19.6 µM for colon carcinoma cell line HT-29. In addition, BC-SM-05 at IC50 concentration was also found to generate increased ROS production and caspases activation. Moreover, HT-29 cells treated with BC-SM-05 and stained with Hoechst 33258 showed formation of apoptotic bodies and DNA degradation whereas untreated cells had intact nuclei. All together our data indicates that BC-SM-05 induces apoptosis in HT-29 cells via mitochondrial dependent pathway may prove itself to be a potential therapeutic agent for colon cancer, providing a basis for the development of the compound as a novel anticancer agent.