Peter L. Rogers

Kinetics of alcohol production by zymomonas mobilis at high sugar concentrations

SummaryStudies on the growth ofZ.mobilis revealed that high concentrations of glucose (10-25%) can be efficiently and rapidly converted to ethanol in batch culture. By comparison withS. carlsbergensis,Z.mobilis had specific glucose uptake rates and specific ethanol productivies several times greater than the yeast.Z.mobilis also had ethanol yields of up to 97% of a theoretical value.

Zymomonas mobilis for fuel ethanol and higher value products.

High oil prices, increasing focus on renewable carbohydrate-based feedstocks for fuels and chemicals, and the recent publication of its genome sequence, have provided continuing stimulus for studies on Zymomonas mobilis. However, despite its apparent advantages of higher yields and faster specific rates when compared to yeasts, no commercial scale fermentations currently exist which use Z. mobilis for the manufacture of fuel ethanol. This may change with the recent announcement of a Dupont/Broin partnership to develop a process for conversion of lignocellulosic residues, such as corn stover, to fuel ethanol using recombinant strains of Z. mobilis. The research leading to the construction of these strains, and their fermentation characteristics, are described in the present review. The review also addresses opportunities offered by Z. mobilis for higher value products through its metabolic engineering and use of specific high activity enzymes.

Ethanol production byZymomonas mobilis in continuous culture at high glucose concentrations

SummaryEthanol production byZ.mobilis has been studied in continuous culture with 10, 15 and 20% glucose media. At 10% glucose, steady state conditions were achieved under glucose-limited conditions. At 15 and 20% glucose, the glucose was not fully metabolized even at low dilution rates and oscillatory behavior was evident. It is proposed that ethanol inhibition of growth is responsible for these phenomena. Comparison of kinetic parameters with those from previously published batch data revealed similar values. The maintenance energy coefficient (m) forZ.mobilis was relatively high and was calculated as 1.6 g/g/h for 10% glucose and 3.1 g/g/h for 15% glucose.

Kinetic studies on a highly productive strain of Zymomonas mobilis

SummaryPrevious studies have demonstrated that Zymomonas mobilis is a very promising organism for ethanol production. In the present study comparative kinetic data from batch and continuous cultures on glucose media are presented which show that a new strain of Z. mobilis has higher specific rates of growth and ethanol production as well as a higher tolerance to ethanol.

Continuous ethanol production by immobilized cells of Zymomonas mobilis

SummaryStudies have been carried out with a highly productive strain of Zymomonas mobilis in an immobilized cell reactor using both Ca alginate and κ-carrageenan as supporting matrices. Productivities above 50 g/l/h have been found at ethanol concentrations in excess of 60 g/l. With immobilized cells of Z. mobilis, there was a decline of approximately 30s% in activity after 800 h operation.

Production of L‐phenylacetylcarbinol (L‐PAC) from benzaldehyde using partially purified pyruvate decarboxylase (PDC)

Biotransformation of benzaldehyde to L‐phenylacetylcarbinol (L‐PAC) as a key intermediate for L‐ephedrine synthesis has been evaluated using pyruvate decarboxylase (PDC) partially purified from Candida utilis. PDC activity was enhanced by controlled fermentative metabolism and pulse feeding of glucose prior to the enzyme purification. With partially purified PDC, several enzymatic reactions occurred simultaneously and gave rise to by‐products (acetaldehyde and acetoin) as well as L‐PAC production. Optimal reaction conditions were determined for temperature, pH, addition of ethanol, PDC activity, benzaldehyde, and pyruvate:benzaldehyde ratio to maximize L‐PAC, and minimize by‐products. The highest L‐PAC concentration of 28.6 g/L (190.6 mM) was achieved at 7 U/mL PDC activity and 200 mM benzaldehyde with 2.0 molar ratio of pyruvate to benzaldehyde in 40 mM potassium phosphate buffer (pH 7.0) containing 2.0 M ethanol at 4°C.

Mathematical modelling of ethanol production from glucose/xylose mixtures by recombinant Zymomonas mobilis

A model has been developed for the fermentation of mixtures of glucose and xylose by recombinant Zymomonas mobilis strain ZM4(pZB5), containing additional genes for xylose assimilation and metabolism. A two-substrate model based on substrate limitation, substrate inhibition, and product (ethanol) inhibition was evaluated, and experimental data was compared with model simulations using a Microsoft EXCEL based program and methods of statistical analysis for error minimization. From the results it was established that the model provides good predictions of experimental batch culture data for 25/25, 50/50, and 65/65 g l−1 glucose/xylose media.

The simultaneous production of sorbitol from fructose and gluconic acid from glucose using an oxidoreductase of Zymomonas mobilis

SummaryThe production of sorbitol and gluconic acid by toluene-treated, permeabilized cells of Zymomonas mobilis has been evaluated. From a 60% total sugar solution (300 g/l glucose and 300 g/l fructose), a sorbitol concentration of 290 g/l and a gluconic acid concentration of 283 g/l were achieved after 15 h in a batch process using free toluene-treated cells. A continuous process with immobilized cells was developed and only a small loss of enzyme activity (less than 5%) was evident after 120 h. With a strongly basic anion exchange resin and an eluent of 0.11 M Na2B4O7/0.11 M H3BO3, good separation of sorbitol and gluconic acid was achieved.

Sorbitol production by Zymomonas mobilis

SummaryHigh resolution 13C Nuclear Magnetic Resonance (NMR) spectroscopy has been employed to determine the chemical composition of the unknown major products in a sucrose or fructose plus glucose fermentation to ethanol by the bacterium Zymmonas mobilis. When grown on these sugars Z.mobilis was found to produce significant amounts of sorbitol, up to 43 g·l-1 for strain ZM31 when grown on 250 g·l-1 sucrose.The production of sorbitol and decrease of glucose, fructose, or sucrose was followed throughout batch fermentations by NMR and HPLC. Sorbitol was shown to be derived only from fructose by [14C]-feeding experiments. Additionally 31P NMR spectroscopy was utilized to determine the concentrations of both glucose 6-phosphate and fructose 6-phosphate relative to their respective concentrations in Z.mobilis cells fermenting glucose or fructose alone.It is suggested that free glucose inside the cell inhibits fructokinase. Free intracellular fructose may then be reduced to sorbitol via a dehydrogenase type enzyme. Attempts to grow Z.mobilis on sorbitol were unsuccessful, as were experiments to induce growth via mutagenesis.

The effect of temperature on the kinetics of ethanol production by strains of Zymomonas mobilis

SummaryTwo strains of Z. mobilis were evaluated for temperature sensitivity between 25°C and 40°C. At higher temperatures the cell viability, biomass yield, ethanol yield and final ethanol concentration decreased, and there was evidence of increased ethanol inhibition. However the kinetic parameters μ, qs and qp were largely unaffected by temperature over this range.