In spite of tremendous developments in the field of Allopathy during the 20th century, plants still remain one of the major sources of drugs in the modern as well as traditional systems of medicine throughout the world, as alternative medicine. Many of the Indian medicines, after prolonged use in folk medical practices, are screened and used for treating and preventing various chronic disorders such as cardiovascular diseases and cancer. The anticancer property of one of the Siddha formulations, Rasagenthi Mezhugu (RM) was tested by standard MTT cell proliferation assay method. In MTT assay, the cytotoxicity and GI50 values were observed for three cell lines obtained from ATCC. The cell lines used are COLO205 (ATCC® CCL222) ™), (adherent, epithelial, colon derived from the lymph nodes of metastatic site), MDA-MB-231 (ATCC® HTB-26™), (adherent, epithelial, mammary gland (breast) derived from pleural effusion sites of metastasis) and HCT 116 (ATCC® CCL247™), (adherent, epithelial colon derived from colorectal carcinoma). The percentage of cytotoxicity, Growth Inhibition (GI50) and viability percentage were calculated and inferred accordingly. Results showed that the percentage cytotoxicity of RM in MDA-MB-231 cell line was found to be Maximum (93.9%) followed by COLO205 Cell line (90%) and HCT-116 Cell line (59.2% ) at 2000 μg/ml (GI50) respectively. The results of cytotoxicity assay demonstrated that among all the three tested human cell lines, growth inhibition was found to be directly proportional to the concentration used. The GI50 data represent that among tested human cancer cell lines, the MDAMB-231Cell line is more receptive towards Rasagenthi Mezhugu. One of the prerequisites for an anti-cancer drug, be it an NCE or herbal alternative or a biological compound, is to be cytotoxic in order to kill the cancer cells and many anticancer products in the market are basically cytotoxic. Therefore, the results of the present study on the herbal product, RM strongly support the usefulness of Rasagenthi Mezhugu as one of the key Complementary Alternative Medicines in cancer therapy.