Isolation and characterization of bioethanol producing wild yeasts from bio-wastes and co-products of sugar factories

Purpose: Yeasts are widely used for the production of bioethanol from biomasses rich in sugar. The present study was aimed at isolating, screening, and characterizing fermentative wild yeast recovered from bio-waste and co-products of Ethiopian sugar factories for bioethanol production using sugarcane molasses as a substrate. Method: The wild yeasts were identified according to their cellular morphology and D1/D2 and ITS1-5.8S-ITS2 rDNA sequencing. Analysis of ethanol and by-product concentration was done by HPLC equipped with a UV detector. Higher alcohols, acetaldehyde, and methanol were analyzed using GC-MS equipped with a flame ionization detector (FID). Result: Seven strains (Meyerozyma caribbica MJTm3, Meyerozyma caribbica MJTPm4, Meyerozyma caribbica SHJF, Saccharomyces cerevisiae TA2, Wickerhamomyces anomalus MJTPm2, Wickerhamomyces anomalus 4m10, and Wickerhamomyces anomalus HCJ2F) were found tolerant to 18% (v/v) ethanol, whereas one strain Meyerozyma caribbica MJTm3 tolerated 20%. These strains also showed tolerance to 45°C, 50% of sugar, and pH 2–10. Meyerozyma caribbica MJTm3 produced 12.7% (v/v) of alcohol with an actual ethanol concentration of 26 g L⁻¹, an ethanol yield of 47%, 78% of theoretical yield, and a productivity of 0.54 g L⁻¹ h⁻¹ from 30 °Brix of molasses at 48 h incubation under laboratory scale. Based on the one variable at a time optimization (OVAT), the optimal parameters for maximum bioethanol production were at initial pH 5.5, 35 °Brix, 30°C, 15% inoculum size, 150 rpm, 4 g L⁻¹ di-ammonium phosphate supplement, and 48 h incubation. Under these optimum conditions, 14% (v/v) alcohol, 42 g L⁻¹ actual ethanol concentration, 69% ethanol yield, 89% of theoretical yield, and productivity of 0.88 g L⁻¹ h⁻¹ were obtained. Conclusion: These results indicated that M. caribbica MJTm3 should further be evaluated, optimized, and improved for industrial bioethanol production due to its fermentation potential.