The study was undertaken to isolate antimicrobial potent biosurfactant producing bacteria from halophilic environment. Five bacterial colonies codes as MK1-MK5 were isolated from seashore soil sample and screened for biosurfactant activity by blood haemolysis and oil spreading assay. The best biosurfactant producer of Pseudomonas sp. MK5 which produced s haemolysis and good oil displacement activity was identified by studying cultural, morphological and biochemical characteristics. The biosurfactant produced by Pseudomonas sp. MK5 was extracted and studied for their antimicrobial effect on bacterial and fungal pathogens. In this study, the biosurfactant exhibited good antibacterial activity against Gram positive than Gram negative bacteria and also showed good antifungal activity on tested fungus. The functional groups of the biosurfactant was analysed by FTIR study, the data confirms the presence of various functional groups/compounds in the biosurfactant and confirm the presence rhamnolipids. This present study substantiates the antimicrobial potential of biosurfactant produced by Pseudomonas sp. MK5. It could be gain more important in future for various pharmaceutical applications.