The indiscriminate use of pesticides and herbicides to increase crop productivity has aroused a great concern among the environmental and health scientists due to their adverse effects in both target as well as non-target species. Although substantial information is available regarding their environmental and ecological impact, not much is known in regard to its toxicity in the mammalian system. Therefore a study was conducted for the assessment of cytotoxic and genotoxic effects of cypermethrin (Type II pyrethroid) dichlorvos (organophosphate) and pendimethalin (dinitroaniline herbicide) in Chinese hamster ovary (CHO) cells. CHO cells were exposed to 1 microM, 10 microM, 100 microM, 1000 microM, and 10,000 microM, cypermethrin, pendimethalin and dichlorvos for 3h and cytotoxicity was assessed by MTT assay. Their genotoxic potential was also evaluated by Comet assay. The results demonstrate that dichlorvos and pendimethalin exhibited higher extent of cytotoxicity as compared to cypermethrin. A significant (p<0.05) concentration dependent increase in DNA damage was observed with dichlorvos (0.01 microM and above) and pendimethalin (0.1 microM and above) as evident by Comet assay parameters viz., Olive tail moment (arbitrary units), tail DNA (%) and tail length (muM). Cypermethrin induced a significant (p<0.05) DNA damage only at higher concentrations (1000 and 5000 microM). Our data indicates that these chemicals produce cytotoxicity and DNA damage in mammalian cells and should be used with caution.