Title: Production of Single-Cell Protein using Enzymatically Hydrolyzed Defatted Soybean Meal-Derived Sugars by Candida Utilis and Komagataella Pastoris

Plant-based protein has gained significant prominence as a viable alternative to its animal-based counterparts. Soybeans, being the leading source of plant-based protein, have widespread use in animal feeds and other sectors of the food industry. However, in its primary form, soybean does not serve as nutritious feed for non-ruminant animals and humans due to the presence of anti-nutritional oligosaccharides, trypsin inhibitors, lectins, and saponins. This entails a need for a specialized process that not only reduces antinutritional components but also enhances the protein quality in soybean meals. In this study, defatted soybean meal (dSBM) was utilized as substrate and treated enzymatically to break down constituent carbohydrates into monosaccharides. The monosaccharide-rich media was utilized to synthesize single-cell protein (SCP) from yeasts in an aerobic environment. Komagataella pastoris and Candida utilis were the selected strains to produce SCP. Following heat treatment with Termamyl®, dSBM was hydrolyzed using cocktail of α-amylase, pectinase, glucoamylase, and xylanase enzymes at 50 °C, pH~5.5, and for duration of 12 h. The hydrolytic process was optimized by fine-tuning enzyme concentrate loadings and composition, achieving a total monosaccharide yield of 33.8 ± 1.6%, which includes a glucose yield of 35.7 ± 3.3%. During propagation of yeast cultures, it was observed that both strains reached similar final dry cell densities, 11.7 ± 0.0 g/L for K. pastoris and 12.8 ± 0.0 g/L for C. utilis cells. By tracing sugar concentrations, it was observed that both species strongly favored hexose consumption over pentose sugars. Amino acid analyses on SCPs revealed an overall improvement in the pattern over untreated dSBM, with essential amino acids (apart from methionine) satisfying FAO recommended levels.