Bioconversion Technology Development

Gasification-Fermentation

This thermochemical technology involves the gasification of a variety of feedstocks, producing a synthesis gas that is then cleaned, cooled and placed into storage. The syngas is then bubbled through a bioreactor which contains a unique microorganism that converts carbon monoxide, carbon dioxide, and hydrogen into ethanol and other valuable products. Even though this process has been successfully demonstrated, a number of critical factors must be addressed to improve conversion efficiency prior to moving this technology to commercialization. These factors involve syngas production, syngas fermentation, and microbe development.

Syngas Production

The undesirable tars generated during gasification are problematic because some of the compounds may be toxic to the microorganisms within the bioreactor. Tar elimination reactions are known to be kinetically limited, so reaction rates can be increased through the use of a catalyst. Several different catalysts will be used in our fluidized-bed gasifier to determine their efficiency in catalytic conversion of some of the tars.

Syngas Fermentation

Recent work has shown that acetic acid is primarily produced during the growth phase of one candidate microorganisms (P7) and that ethanol production is much smaller than desired (less than 10 times the acetic acid production). Work will continue, utilizing tools to understand the switch between acetic acid and ethanol production and to develop a strategy to enhance ethanol production. Avenues we will explore include adjustment of the redox potential during fermentation, shock treatment with acetic acid, medium manipulation (changing nutrients, vitamins, and minerals at onset of stationary phase) and/or pH adjustment.

Work will continue in assessing the benefit of dual reactors, one for growth while the other for ethanol production. Additional opportunities include growing the cells on glucose to obtain a high cell mass and then switching the feed to producer gas. The results of this study will be applied to our continually developing bioreactor model. Fixed biofilms for increased cell and product yields will also be investigated.

A large-scale (70-liter working volume) bioreactor is scheduled to come on-line within the next three months to perform pilot-scale studies on ethanol production from biomass-generated synthesis gas. Initially, the studies will be scaled from the 3-liter reactor studies. Analysis of inlet/outlet gas compositions, pH, product concentrations, and associated parameters (such as yields) will be measured. The bioreactor will initially be run in liquid batch mode with subsequent runs using continuous liquid feed and product removal. Mass transfer studies will be performed to assess mass transfer limitations.

Microbe Development

A defined medium has been developed for strain P7. The fermentation of syngas by P7 has been altered by medium composition. Examples include showing the dependence of ethanol production on the presence of iron and shifting end products of fermentation from butanol/butyrate to ethanol/acetate by metabolic inhibitors. A defined medium has not been developed for another candidate strain (P11) and medium alterations, which affect syngas fermentation, do not have similar effects on P11. This illustrates that medium development is an empirical process and also that it is a part of process development that would continue even upon commercialization.

The hydrogenases expressed by P7 during fermentation of carbon monoxide have been attributed to being biochemically novel. The expression of hydrogenases based on substrate provided will be determined using activity stains on anaerobic protein gels. The process of improving fermentation parameters for strain P11 will begin upon completed of its characterization.

Simultaneous Saccharification and Fermentation (SSF)

Activities will continue in identifying potential improvements in process efficiencies. SSF combines enzymatic hydrolysis of cell wall polysaccharides into monomer sugars and microbial fermentation of these sugars to ethanol. One such improvement could be realized through the use of thermotolerant yeasts. SSF reduces product inhibition of enzymes and saves on capital costs by placing two processes into one tank. Unfortunately, ideal temperatures for conventional ethanolgens such as Saccharomyces cerevisiae and Zymomonas mobilis (32 to 37°C) used in SSF processes cannot survive at temperatures at which maximum cellulase enzyme activity occurs (45 to 50°C), so lower temperatures (less than 37 °C) must be used. Research will be conducted on a group of five Kluyveromyces marxianus yeast strains referred to as the IBM strains. These strains have been observed to ferment glucose to ethanol at temperatures up to 50°C, with little if any reduction in fermentation capacity up to 45°C. They are also one of the few types of yeast that have been reported to ferment both cellobiose, a disaccharide of b-glucose that is the product of cellulose hydrolysis by cellulase, and xylose, a major constituent of perennial grasses, to ethanol at 45°C. In addition, these strains have shown ethanol tolerance from 75 to 94 g/l, which is more than many comparable thermotolerant ethanolgens. Using these stains could decrease in enzyme costs of over 40% which translates to lowering enzyme costs to less than $0.10 per gallon of ethanol. A number of perennial grasses identified under “Feedstock Development” will be subjected to IBM strains to determine the conversion efficiency. The most effective pretreatment conditions for the greatest amount of cellulose conversion to ethanol will be determined for each grass and IMB strain tested. Using these pretreatment conditions, SSF parameters of enzyme loading and temperature will be optimized for each IMB strain tested. Optimized parameters can then be used for future scale up and commercialization, as well as strain development and economic studies. Given successful initial results, additional feedstocks will be assessed including a transgenic low-lignin switchgrass recently developed the Noble Foundation, Ardmore, OK.

On-Farm Ethanol Production from Sweet Sorghum

Sweet sorghum has the potential to be used as a renewable energy crop, and has become a viable candidate for ethanol production. Previous research has concentrated on transporting biomass to a central processing facility which extracts and ferments the juice. As such, the greatest barrier to commercialization of this process has been the high capital costs involved in a central processing plant that may operate only seasonally. Research is underway to determine the feasibility of in-farm juice collection and fermentation. The process includes a newly designed field harvester capable of pressing and collecting the juice, large storage bladders for fermentation, and a mobile distillation unit for ethanol concentration. Studies will be conducted to determine the juice production potential of sweet sorghum across Oklahoma, machinery performance, process performance (yeast type, pH adjustment, and nutrient needs), and environmental factors effecting fermentation.