Aims: The increased importance of biosurfactant in the recent past is mainly due to their applications in various industries ranging from petroleum to pharmaceuticals. Their biodegradability and environmental compatibility with low toxicity makes it even more interesting. Microbial production of biosurfactant is found to be a viable option as they are diverse, eco-friendly, facilitate large scale production, able to perform under extreme conditions etc. One class of microbes that is endophytes are known to show great potential in producing different varieties of medically and industrially significant biological compounds. The present study focuses on the screening and production of biosurfactant from endophytic bacteria. Methodology and results: Of all the isolates tested, one endophyte identified as Bacillus cereus HM998898 was found to produce maximum biosurfactant. Statistical method Plackett burman was used to optimize the media for the maximum production and the ideal composition was found to be KNO3 (1 g/L), Gingley oil (2 mL), K2HPO4 (2.5 g/L), KH2PO4 (0.75 g/L), MgSO4·5H2O (0.5 g/L), FeSO4.7H2O (0.005 g/L) and NaCl (0.025 g/L). The extracted biosurfactant was characterized and was identified to be glycolipid. This was further tested for biocompatibility against Fibroblast (3T3) cells and was evaluated for their anti tumor activity against Hep2 cells. Conclusion, significance and impact of study The biosurfactant produced was found to induce toxicity to cancer cells at appreciable levels while they remained non-toxic to normal cells supporting the possible applications of biosurfactant in medical field.