Thermophilic ethanol production investigation of ethanol yield and tolerance in continuous culture

Ethanol yield and ethanol tolerance, the two factors that most constrain the utilization of thermophilic bacteria for ethanol production, were investigated in continuous xylose-grown cultures ofClostridium thermosaccharolyticum. Under xylose-limiting conditions, including varying dilution rates and feed concentrations, the ethanol selectivity (Se, mol/mol) relative to acetic acid, lactic acid, and propane diol remained relatively constant at about 2. Product addition and removal experiments indicate that mass action effects related to the concentrations of organic fermentation products play a relatively minor role in determining the ratios of products made. Of much greater apparent importance were as yet uncharacterized regulatory mechanisms that appear to be correlated with nonlimiting concentrations of the carbon and energy-source. Substrate-plentiful transients were found to accompany Se values > 11. Such transients provide a useful model system for the study of end product control, as well as a cultivation mode with considerable applied potential. No apparent ethanol inhibition was observed, as indicated by no decrease in the maximum rate of growth allowing complete substrate utilization (0.22 h-1) for endogenously-produced ethanol concentrations up to 11.4 g/L, and total endogenously-produced + exogenously-added ethanol concentrations up to 21.3 g/L. Higher concentrations of ethanol are tolerated atµ= 0.11 h-1, although the onset of inhibition was not characterized at this growth rate. Results suggest that the ethanol tolerance of C.thermosaccharolyticum grown in continuous culture may be greater than that typically observed previously for thermophiles grown in batch culture.