Unconventional Yeasts as Whole-Cell Biocatalysts for Carboligase-Mediated Synthesis of Optically Active Acyloins

Background/Objectives: α-Hydroxy ketones (acyloins) are valuable chiral building blocks widely used in the synthesis of pharmaceuticals and fine chemicals. Their stereoselective synthesis remains challenging, driving interest in sustainable biocatalytic approaches for asymmetric C–C bond formation. Methods: Here, non-conventional yeasts (NCYs) from six genera were evaluated as lyophilized whole-cell biocatalysts for acyloin production through carboligase-mediated synthesis. Results: Four strains—Pichia carsonii WY-1, Lodderomyces elongisporus WY-2, and Candida guilliermondii WY-7 and WY-8—exhibited high carboligase activity when pyruvate was used as co-substrate, efficiently converting benzaldehyde and pyruvate to (R)-PAC. Factorial design screening of reaction conditions (pH, co-substrate concentration, and temperature) minimized byproduct formation, yielding (R)-PAC in 79–96% yield and 97.6–98.3% ee. Evaluating a substrate scope of 30 functionalized aromatic aldehydes revealed broad native tolerance, particularly toward meta- and para-substituted derivatives, affording corresponding α-hydroxy ketones in 10–90% yield and generally >90% ee. Conclusions: These results demonstrate the potential of lyophilized NCYs as whole-cell biocatalysts for stereoselective acyloin synthesis. Furthermore, these strains serve as promising sources of new enzymes with carboligase activity for future biochemical exploration and biocatalyst development.