A kinetic study of ethanol production by Zymomonas mobilis

The purpose of this study was to determine the effects of temperature, pH, glucose concentration, and ethanol concentration on the fermentation kinetics of the ethanol-producing bacterium Zymomonas mobilis (ATCC 10988). The effects of these parameters on the specific growth rate and the specific ethanol production rate of Z. mobilis were studied in batch and continuous culture. Initial-rate batch experiments were used to investigate the effects of temperature, pH, and glucose concentration. The growth medium was an aqueous solution of glucose (5 to 20%), yeast extract (Difco, 0.5%) and succinate (0.01 M).

The effect of temperature on the specific growth and ethanol production rates was consistent with the Arrhenius Law in the range of 24 to 35°C. The measured "activation energy" was 6 (Kcal/g mole) in both cases. The temperature corresponding to maximum specific growth rate was 35°C, and the maximum specific ethanol production rate occurred at 37°C. Above these temperatures, the rates declined rapidly. At 36°C, maximum cell growth rates were found over the pH range 5.6 to 7.5, and maximum ethanol production rates were found over the pH range 5.2 to 7.5. Specific growth rates were more sensitive to acidic conditions than were ethanol production rates.

At 36°C, reaction rates were shown to increase with increasing glucose concentration up to approximately 10% and then to decline at higher concentrations. Classical kinetic models incorporating substrate inhibition could not be fit to the experimental data, suggesting that simple substrate inhibition may not be the primary mechanism responsible for the declining rates at high glucose concentrations.

Chemostat experiments were conducted at 36°C in a 2% glucose medium spiked with various concentrations of ethanol. At relatively high dilution rates (0.34 hr^-1), a floe population developed in the reactor. At lower dilution rates, a true steady-state condition was not achieved, as evidenced by a gradual increase in the glucose uptake rate over a period of two weeks. It is suggested that an adaptation of Z. mobilis to the experimental conditions might have been occurring during this time. No quantitative conclusions could be drawn from these experiments on the effects of ethanol on the fermentation kinetics.

The results obtained in this study may be useful in the design of an ethanol-producing bioreactor. A temperature of 36°C and pH range of 5.6—7.5 appear to allow approximately maximum rates of cell growth and ethanol production. The pH control may also be used to selectively inhibit growth, if desired, under acidic conditions. Ethanol inhibition of

Z.

mobilis fermentation is poorly characterized and understood at present, and should be investigated further.