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In order to achieve high butanol production by Clostridium saccharoperbutylacetonicum N1-4, the effect of lactic acid on acetone-butanol-ethanol fermentation and several fed-batch cultures in which lactic acid is fed have been investigated. When a medium containing 20 g/l glucose was supplemented with 5 g/l of closely racemic lactic acid, both the concentration and yield of butanol increased; however, supplementation with more than 10 g/l lactic acid did not increase the butanol concentration. It was found that when fed a mixture of lactic acid and glucose, the final concentration of butanol produced by a fed-batch culture was greater than that produced by a batch culture. In addition, a pH-controlled fed-batch culture resulted in not only acceleration of lactic acid consumption but also a further increase in butanol production. Finally, we obtained 15.5 g/l butanol at a production rate of 1.76 g/l/h using a fed-batch culture with a pH-stat continuous lactic acid and glucose feeding method. To confirm whether lactic acid was converted to butanol by the N1-4 strain, we performed gas chromatography-mass spectroscopy (GC-MS) analysis of butanol produced by a batch culture during fermentation in a medium containing [1,2,3-(13)C(3)] lactic acid as the initial substrate. The results of the GC-MS analysis confirmed the bioconversion of lactic acid to butanol.

30Sourdough, a traditional fermented dough, is made via natural fermentation by lactic 31 acid bacteria (LAB). Its pH changes from near neutral to acid during the subculture 32 process. However, the product quality of subcultured sourdough depends on the 33 unpredictable succession of LAB communities, the influential factors of which are still 34 unclear. To elucidate one end of the LAB community succession mechanism, we 35 evaluated the effect of pH by designing four subculture experiments using a model 36 medium adjusted to pH 6.7, 5.5, and 4.5, as well as a natural sourdough subculture. All 37 experiments began by inoculating a sourdough LAB mixture, and both bacterial 38 successions and fermentative properties were monitored until ten subculture steps. In 39 media subcultures, lactic acid production was higher in higher pH media. Three LAB 40 genera, Weissella, Pediococcus, and Lactobacillus, each represented by one operational 41 taxonomic unit (OTU), were successively detected in all subcultures. In later steps with 42 lower pH media, an OTU closely related to Lactobacillus brevis dominated, replacing 43 an OTU closely related to the Weissella cibaria-confusa group that was more dominant 44 than the L. brevis OTU in the near-neutral pH medium. In the sourdough subculture, the 45 three genera were also detected, while Lactobacillus was dominant in earlier steps due 46 4 to the emergence of an OTU closely related to Lactobacillus sanfranciscensis. These 157

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