Optimization of Biosurfactant Production by Bacillus subtilis NRRL B14322 Using Agro-Industrial Residues: A Plackett-Burman Approach

This study investigates the production of biosurfactants by Bacillus subtilis NRRL B14322 using formulated media derived from industrial residues. The media included two hydrophobic carbon sources (acid- and enzyme-derived soapstock), two hydrophilic carbon sources (sugarcane and soy molasses), and four nitrogen sources (sugarcane yeast extract, brewer’s yeast extract, crude yeast extract powder, and corn steep liquor). These substrates were selected for their nutritional content and large-scale availability, with soapstock production exceeding 500,000 tons annually, sugarcane molasses reaching tens of millions of tons, corn steep liquor derived from over 1 billion tons of corn processed globally, and brewer’s yeast byproducts contributing up to 558,000 tons per year. Micronutrients, pH, and temperature were also varied. A Plackett–Burman design was applied to identify key factors influencing biosurfactant production, measured by the emulsification index (E24) using hexane. The best performance (E24 = 0.475) was obtained using a medium containing 40 g/L enzyme-derived soapstock, 3 g/L acid-derived soapstock, 30 g/L sugarcane molasses, 20 g/L soy molasses, 20 g/L corn steep liquor, 20 g/L sugarcane yeast extract, 80 g/L brewer’s yeast extract, and 6 g/L crude yeast extract powder, along with KH₂PO₄, K₂HPO₄, MgSO₄·7H₂O, CaCl₂·2H₂O, FeSO₄·7H₂O, and MnSO₄·H₂O. Pareto analysis identified CaCl₂·2H₂O as the most significant factor. These findings support the strategic use of agro-industrial residues in sustainable biosurfactant production with potential for scale-up and environmental applications.