Bioreactor cultures
Strain GSU5 was grown in bioreactor batch cultures using glucose or xylose as carbon sources in order to analyze solvent production in a larger scale. Several tests were performed to determine the best pH conditions for growth and solvent production, since previous studies had indicated that solvent production was enhanced by low pH in otherT. thermosaccharolyticum strains. Preliminary tests in glucose cultures with initial starting pH of 6.7 or 7.5 did not allow adequate growth, so all cultures were performed with an initial pH of 7, and pH was controlled to prevent values lower than 5. Sugars were added at an initial concentration of 10 g/L, and replenished after 17 h by adding half the initial amount to avoid sugar depletion.
Growth was more vigorous in glucose cultures, achieving 52% more biomass than when using xylose (1.9 vs 1.0 g/L). Solvent production was also much higher in glucose, while xylose cultures produced more acids than solvents (Figure 4). Butyric acid remained the main product in xylose cultures, but glucose cultures produced slightly more ethanol than butyrate. The final butanol concentration obtained was 0.33 ± 0.01 g/L in glucose and 0.26 ± 0.02 g/L in xylose, while the final ethanol concentration was 4.34 ± 0.06 g/L in glucose and 0.78 ± 0.49 g/L in xylose (Table 3).
As expected, fermentor cultures corresponding to both sugars had a much higher biomass and solvent production that 5 ml tube cultures. Although the main products were the same observed in tube cultures, the relative amounts of acids and alcohols varied. A higher relative amount of butanol was observed with both substrates when all major metabolites were considered, showing a slight increase in carbon fluxes towards butanol in bioreactor cultures (Figure 5). When cultures grown using glucose were compared to tube cultures, solvent production was observed to increase more than biomass, with the most important increase observed in butanol production. While an 8 fold change was observed in biomass (1.78 vs 0.22 g/L), final concentrations of ethanol and butanol increased 10 and 14 times respectively, so that the amount of alcohols produced in the fermentor was similar to that of acids (Figure 5). Results obtained with xylose were different, as a 6 fold increase in biomass (1.04 vs 0.20 g/L) was accompanied by a similar increase in butanol concentration, while ethanol concentration only increased 1.5 fold when fermentor cultures were compared with 5 ml cultures (Figure 5).