The worldwide growing environmental concern and existing control regulations led researchers to study alternative ways to process industrial waste and the use of natural surfactants, or biosurfactants, is among one of the most promising methods due the fact that such compounds are metabolic products of fungi, bacteria and certain strains of yeast. Biosurfactants are amphiphilic molecules with low toxicity, high biodegradability and structural diversity that are capable to reduce interfacial tension of mixtures of water and hydrocarbons and thus an excellent alternative to replace synthetic surfactants. Besides, several different microorganisms were proven able to produce biosurfactants by digesting renewable-based culture medium. This paper describes the biosurfactant production by Bacillus subtilis, Gram positive and non-pathogenic bacteria, using as substrate industrial waste containing high levels of glucose. Were tested several percentages of inoculum (5, 7.5 and 10%) and the produced biosurfactant was evaluated according to the emulsification index (using soybean oil or engine oil in proportion 1:1 with the broth), surface tension analysis and glucose content. The tests were carried out in an orbital-stirred shaker and temperature kept at 37°C. The biosurfactant produced using 10% of inoculum showed a surface tension reduction of 36%, and emulsification index of 75% and 83% for soy oil and motor oil, respectively.