Biosurfactants are surface active compounds released by microorganisms. They are biodegradable non-toxic and ecofreindly materials. In this review we have updated the information about different microbial surfactants. The biosurfactant production depends on the fermentation conditions, environmental factors and nutrient availability. The extraction of the biosurfactants from the cell-free supernatant using the solvent extraction procedure and the qualitative and quantitative analysis has been discussed with appropriate equipment details. The application of the biosurfactant includes biomedical, cosmetic and bioremediation. Rhamnolipid biosurfactants produced by Pseudomonas aeruginosa DS10-129 showed significant applications in the bioremediation of hydrocarbons in gasoline-spilled soil and petroleum oily sludge. Rhamnolipid biosurfactant enhanced the bioremediation process by releasing the weathered oil from the soil matrices and enhanced the bioavailability of hydrocarbons for microbial degradation. It is having potential applications in the remediation of hydrocarbon contaminated sites.