A lipopeptide biosurfactant from Bacillus natto TK-1 was purified by thin-layer chromatography (TLC) and the molecular weight of the lipopeptide was determined by ESI-MS.The critical micelle concentration (CMC) of lipopeptide was estimated to be around 105 mg/L and the minimum surface tension was 30.1 mN/m.Lipopeptide was found to be an anti-adhesive agent against several bacterial strains.The antimicrobial activity of lipopeptide was evaluated against ten microorganisms using disc diffusion method and the lipopeptide showed a broad spectrum of antimicrobial activity.