The present research involves cytotoxic evaluation of Chloris barbata leaves by employing various in vitro models. Extraction of active constituents of the leaves of Chloris barbata was done by using soxhlet apparatus with polar and non-polar solvents. Further the extract was subjected to phytochemical screening and the methanolic extract of the leaf reveals the presence of flavonoids, glycosides, alkaloids, saponins and triterpinoids. Invitro cytotoxic efficacy of the Chloris barbata leaf methanolic extract was assessed with Human epidermoid larynx carcinoma (Hep 2), African green monkey kidney normal (Vero), Dalton's Ascitic Lymphoma (DAL) cell lines by using tryphan blue dye exclusion test, MTT, Sulphorodamine B (SRB) and Neutral Red Uptake (NRU)assay. In Tryphan blue dye exclusion assay it was found that there is an increase in percentage of dead vero (92±0.49), DAL (76±0.47) and Hep-2 cells (82±0.36) with chloris barbata leaf extract 200μg/mlwhen compared to control. Standard 5-Fluorouracil (5μg/ml) has shown an increase in percentage of dead vero, DAL and Hep-2 cells to an extent of 100±1.49, 94±1.35 and 99±0.21 when compared to control. In MTT assay the percentage viability of DAL, Hep-2 and vero cells has been significantly reduced by chloris barbata leaf extract at 200μg/ml (DAL 0.941±0.95, Hep-2 0.714±1.62 and vero cells 1.364±1.48) when compared to control (% viability is 100). In SRB assay, the percentage growth inhibition of cell lines by methanolic leaf extract was found to be 56.362 (vero cells), 69.128 (DAL cells) and 61.354 (Hep-2 cells) when compared to control group. NRU assay has shown the IC50 values 24.11±3.82 (vero), 47.16±0.14 (DAL) and 31.26±2.15 (Hep-2 cells) with leaf extract when compared to control. The above results signify that the methanolic extract of Chloris barbata leaves have remarkable cytotoxic potential. Further isolation of active constituents and in vivo cytotoxic studies has to be carried out to establish the exact mechanism involved in anticancer activity of Chloris barbata leaf extract.