Studies on sporulation and butanol production by Clostridium acetobutylicum

The low n-butanol concentration (2% w/v) produced from the fermentation of carbohydrates by Clostridium acetobutylicum limits the commercial feasibility of the acetone-butanol-ethanol (ABE) fermentation. Although this low product concentration has been attributed to both butanol toxicity and sporulation of solvent-producing cells, only butanol toxicity has been thus far studied in detail.; The present study was undertaken to determine if butanol production could be enhanced at the expense of sporulation by manipulating certain environmental and nutritional conditions. The effects on these two processes of inoculum age and size, the type of carbohydrate source, the type of nitrogen source, phosphate concentration, pressurization of fermentors by sealing, cell-immobilization, and butanol concentration were examined.; Sealing of fermentation vessels, or cell-immobilization had negative or inconclusive effects on both butanol production and sporulation in batch culture, and thus could not be used to enhance butanol production.; Inoculum age and size, the type of carbohydrate substrate, and phosphate concentration all had effects on butanol production and sporulation, but could not be effectively used to enhance butanol production at the expense of sporulation.; Varying the type of nitrogen source, or continuous removal of butanol by extractive fermentation, was used successfully to enhance butanol production. Cultures which contained aspartic acid as the nitrogen source produced a higher butanol concentration (10.4 g/L) than that produced by control cultures (8.1 g/L) which contained the inorganic nitrogen source di-ammonium hydrogen o-phosphate (DAHP). There was a 75% decrease in sporulation in association with this increased butanol production. The inhibition of sporulation was specific, and could be maintained even in the presence of small amounts of aspartic acid. However, any reduction in aspartic acid concentration was detrimental to butanol production.; Continuous extraction of butanol into an organic phase (consisting of oleyl alcohol) gave the highest recorded butanol production (14.1 g/L) from fermentation of glucose, and a higher level of sporulation (40%) than the nonextractive control fermentation. These results showed that butanol toxicity was the main factor limiting both processes. It was concluded that butanol toxicity had a considerably greater effect than sporulation on the production of butanol.