Abstract: The cultural conditions for the production of biosurfactant by Serratia marcescens DT-1P were optimised by Response Surface Methodology (RSM). A study of complex interactions among glycerol (1-5% w/v), inoculum (1-10 OD600), pH (4-8), temperature (20-40°C) and (NH4)2SO4 (0.5-2.0%) were carried out using central composite rotatable design of experiments. The optimum conditions, which were obtained i.e., glycerol (3.4% w/v), inoculum (1.0 OD600), pH (5.8), temperature (26.6°C) and (NH4)2SO4 (0.2%) yielded 6.0 mg L-1 biomass and glycerol (5% w/v), inoculum (10.0 OD600), pH (8.0), temperature (20.0°C) and (NH4)2SO4 (0.05%), yielded 24.48 mg % of biosurfactant. The surfactant was identified as arabinolipid.