In this study, a new formulation of low-cost, biodegradable, and non-toxic biosurfactant by Candida sphaerica UCP 0995 was investigated. The study was conducted in a bioreactor on an industrial waste-based medium, and a central composite rotatable design was used for optimization. The best results, namely a 25.22 mN/m reduction in surface tension, a biosurfactant yield of 10.0 g/L, and a critical micelle concentration of 0.2 g/L, were achieved in 132 h at an agitation speed of 175 rpm and an aeration rate of 1.5 vvm. Compositional and spectroscopic analyses of the purified biosurfactant by chemical methods, Fourier transform infrared spectroscopy, and nuclear magnetic resonance suggested that it is a glycolipid-type biosurfactant, and it showed no cytotoxicity in the MTT assay. The biosurfactant, submitted to different formulation methods as a commercial additive, remained stable for 120 days at room temperature. Tensioactive properties and stability were evaluated at different pH values, temperatures, and salt concentrations. The biosurfactant obtained with all formulation methods demonstrated good stability, with tolerance to wide ranges of pH, temperature and salinity, enabling application under extreme environmental conditions. Bioremediation tests were performed to check the efficacy of the isolated biosurfactant and the selected microbial species in removing oil from soil. The results demonstrated that the biosurfactant produced has promising properties as an agent for the bioremediation of contaminated soil.