Untreated molasses efficiently Enhances 2,3-Butanediol synthesis using engineered Aureobasidium melanogenum P8AC-4.

2,3-Butanediol, an important platform chemical, possesses extensive applications in chemical engineering, pharmaceuticals, and materials science. To address the challenges of resource depletion and environmental impact associated with conventional petroleum-based synthesis, this study focuses on the sustainable production of 2,3-BDO via microbial fermentation. Aureobasidium melanogenum TSYW-58 was employed as the chassis for metabolic engineering. In previous work, genes responsible for the biosynthesis of pullulan, polymalic acid, melanin, and liamocin were deleted, yielding strain PPLM-8, characterized by high gluconic acid titer and reduced by-product formation. Building upon this platform, genes encoding α-acetolactate synthase and α-acetolactate decarboxylase from Bacillus subtilis were heterologously expressed, along with the endogenous butanediol dehydrogenase, leading to the successful construction of strain P8AC-4. This strain is capable of efficient 2,3-BDO production with minimal by-products. Shake-flask fermentation yielded 43.21 ± 1.23 g/L of 2,3-BDO, with a stereoisomer composition of 77.07 % meso- and 22.93 % (R,R)-isomer. Fermentation kinetic analysis in a 5 L bioreactor revealed that 2,3-BDO biosynthesis is tightly coupled with the logarithmic growth phase of the microorganism. By leveraging this metabolic trait, growth-inhibiting components present in untreated straw hydrolysate and molasses were employed to modulate fungal growth rates. Notably, cultivation with untreated molasses as the carbon source effectively extended the product synthesis phase, achieving a 2,3-BDO concentration of 110.37 ± 4.43 g/L in batch culture, representing a 155.42 % increase compared to that obtained with the glucose-based medium. These findings offer an efficient and cost-effective biomanufacturing strategy for the industrial-scale production of 2,3-BDO, contributing to the advancement of sustainable chemical synthesis.