Alcohol fermentation of milk by multiple fermentation

Saccharomyces cerevisiae is a representative yeast used for alcohol production. Production of alcohol from milk using S. cerevisiae is not easy, as this species is unable to hydrolyse lactose. Lactobacillus delbrueckii subsp. bulgaricus (LDB) is able to hydrolyse lactose into galactose and glucose, and is widely used in yogurt production. In this study, production of alcohol from milk was attempted by multiple fermentation using S. cerevisiae and LDB. Firstly, strain LDB48A-12, released from catabolite repression, was obtained from LDB48P by N-methyl-N ’-nitro-N-nitrosoguanidine mutagenesis. LDB48A-12 showed high ability of lactose degradation. Surprisingly, growth speed and lactate production of LDB48A-12 were found to be lower than those of LDB48P. Fermentation of 10% skim milk solution using LDB48A-12 and S. cerevisiae SBC3207 for 4 days at 30℃ or 37℃ produced approximately 1~1.5% of alcohol, which was 1.5 times that produced using LDB48P. In addition, decrease in pH was inhibited. These results were thought to demonstrate novel alcoholic milk fermentation. Comparison of genome sequences revealed mutations in RpoA and DnaA in LDB48A-12. These genes are related to catabolite repression and replication of genome DNA, respectively, suggesting that combined mutation of these genes leads to the preferable phenotype for alcoholic milk fermentation.