Bioethanol production by wild yeasts utilizing sugarcane molasses

Bioethanol production from sugar-rich biomass provides a renewable and sustainable alternative to fossil fuels. However, achieving efficient yields requires robust yeast strains that can withstand industrial stresses and optimized fermentation processes. This dissertation investigated wild and co-cultured yeast strains for their potential to convert sugarcane molasses into bioethanol, focusing on strain identification, stress tolerance, process optimization, and co-fermentation strategies. In the first phase, wild yeasts were isolated from by-products of Ethiopian sugar factories and screened for their fermentative capacity. Seven isolates demonstrated tolerance to high ethanol concentrations, elevated sugar levels, temperature, and pH variation. Among them, Meyerozyma caribbica MJTm3 showed exceptional performance, producing up to 14% alcohol and achieving 89% of the theoretical ethanol yield under laboratory-scale optimized conditions. To confirm the taxonomic identity of MJTm3, whole genome sequencing (WGS) was employed in the second phase. Comparative genomic and phylogenomic analyses validated its classification as M. caribbica, while also highlighting the genetic complexities in differentiating it from its close relative, M. guilliermondii. The third phase optimized fermentation conditions for MJTm3 using response surface methodology. This approach identified key parameters such as pH, inoculum size, molasses concentration, and mixing rate, enabling yields of 86% and ethanol concentrations of 56 g·L⁻¹, confirming the strain’s industrial promise. Finally, co-fermentation experiments with Saccharomyces cerevisiae TA2 and Wickerhamomyces anomalus HCJ2F-19 demonstrated that mixed cultures significantly enhanced ethanol output compared to single strains, with optimized conditions yielding up to 35.5 g·L⁻¹ ethanol.

Keywords: Sugarcane molasses, fermentation, process optimization, bioethanol yield, stress tolerance, and whole genome sequencing.