LAUSR

Lactic acid bacteria play an important role in the food industry and can produce many aromatic compounds during the fermentation process, including diacetyl, an important compound with a buttery flavor. However, existing methods for screening strains that produce high levels of diacetyl are time-consuming and inefficient. An integrated strategy for high-throughput screening of diacetyl-producing strains was therefore developed in this study. This strategy uses 96-deep-well plates for microbial strain cultivation and milk fermentation and trichloroacetic acid for protein precipitation. A colorimetric method based on the reaction of diacetyl with O-phenylenediamine was then developed to determine the diacetyl concentrations in the wells. Good agreement was observed between high-throughput cultivation and traditional method, confirming that the deep-well microplates could replace beaker flasks to scale down cell cultivation and fermentation. The diacetyl concentrations measured with a microplate reader were comparable to those determined via gas chromatography (R 2  = 0.9927) and ultraviolet spectrophotometry (R 2  = 0.9943), and a high recovery level (94.88%) was obtained. Overall, this strategy increased the screening efficiency by 5- to 30-fold compared with traditional methods. Next, this strategy was used successfully to screen two strains, 1-16 (37.05 ± 1.32 mg/L) and CN-7 (32.35 ± 0.54 mg/L), and the diacetyl yields were verified in a 5-L reactor. Our findings suggest that this newly developed high-throughput strategy can accelerate screening efficiency at a low cost and with a high level of accuracy. This strategy has great potential for targeted screening of strains that produce high levels of diacetyl in fermented milk.