An alkaline lipase from Trichosporon asahii MSR 54 was used to develop presoak formulation for removing oil stains at ambient temperature. The lipase was produced in a reactor followed by concentration by ultrafiltration and then it was dried with starch. The biochemical characteristics of enzyme showed that it was an alkaline lipase having pH activity in the range of pH 8.0-10.0 and temperature in the range of 25-50 degrees C. The present lipase was active >80% at 25 degrees C. The lipase was cystein activated with fourfold enhancement in presence of 5 mM cystein and likewise the activity was also stimulated in presence of papain hydrolysate which served as source of cystein. The presoak formulation consisted of two components A and B, component A was enzyme additive and B was a mixture of carbonate/bicarbonate source of alkali and papain hydrolysate as source of cystein. The results indicated that the presoaking in enzyme formulation followed by detergent washing was a better strategy for stain removal than direct washing with detergent in presence of lipase. Further, it was observed that 0.25% presoak component B in presence of 100 U enzyme component A (0.1 g) was the best formulation in removing maximum stain from mustard oil/triolein soiled clothes as indicated by increase in reflectance which was found equal to that of control cloth. The lipase action in presoaked formulation was clearly indicated by quantitated fatty acid release and also the TLC results of wash water, where oil hydrolytic products were visible only in presence of enzyme in the treatment. The wash performance carried at 25 degrees C indicated that washing at 25 degrees C was at par with that at 40 degrees C as indicated by similar reflectance of the washed cloth piece though qualitative fatty acid release was higher at 40 degrees C.