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United States Department of Agriculture

Agricultural Research Service

Research Project: ADVANCED CONVERSION TECHNOLOGIES FOR SUGARS AND BIOFUELS: SUPERIOR FEEDSTOCKS, PRETREATMENTS, INHIBITOR REMOVAL, AND ENZYMES

Location: Bioenergy Research

2011 Annual Report


1a. Objectives (from AD-416)
The long-term goal of this project is to develop lignocellulosic materials as a feedstock for producing sugars and biofuels. The research will focus more specifically on the following objectives: Objective 1: Develop commercially-viable analytical tools that producers and biorefiners of lignocellulosic feedstocks can use to evaluate the quality of harvested biomass for enzymatic and fermentative conversion to ethanol and butanol and plant breeders can use to select superior cultivars for biorefining. Objective 2: Develop new, commercially-viable enzyme and/or protein systems that increase the efficiency of lignocellulosic saccharification. Objective 3: Identify components in lignocellulosic hydrolysates which reduce saccharification or fermentation efficiencies and develop commercially-viable mitigation strategies. Objective 4: Develop new technologies that enable commercially-viable pretreatment processes for lignocellulosic biomass, inhibitor abatement strategies, and enzyme preparations which are optimized for saccharifying particular lignocellulosic feedstocks.


1b. Approach (from AD-416)
Renewable biofuels have the potential to reduce United States dependence on imported oil, lower greenhouse gas emissions, and to further develop the rural economy. Renewable fuels produced from lignocelluloses should be able to replace up to 30% of the United States oil consumption. While technologically proven, commercializing lignocellulosic biofuels is stymied by prohibitively high processing and capital costs. There are opportunities to reduce expenses by developing higher-quality feedstocks, more active enzymes, better managing side-products, and using faster/higher yielding biocatalysts. Our research targets improvements in each of these areas. Crop quality for bioprocessing will be improved by collaborating with plant scientists to breed bioenergy cultivars, including reduced lignin mutants that are more amenable to processing, without sacrificing production yield. The enzymes used are too expensive because their specific activities are too low. Activities can be improved by creating balanced mixtures from blending individual enzymes selected for superior kinetics and by discovery of auxiliary proteins that aid cellulase binding to cellulose. Processing of biomass prior to fermentation releases a wide range of biologically active byproducts that can retard or stall fermentation and frustrate water recycling schemes. Biological abatement has the promise to selectively remove complex organic compounds without generating further waste. It is expected that when combined with an advanced pretreatment and engineered microbes, the final result will be a significantly improved process for converting lignocelluloses into sugars and renewable fuels.


3. Progress Report
The overall goals of this project are to develop technologies for lowering the cost and increasing the efficiency for converting biomass to biofuels and chemicals. The specific areas targeted are developing higher-quality feedstocks, more active enzymes, and better managing undesirable side-products present in processing streams. Scientists working in the Agricultural Research Service (ARS) Bioenergy Research Unit at the National Center for Agricultural Utilization Research (NCAUR), Peoria, IL, have made substantial progress during 2011 in each of these areas. In particular, a novel process has been developed for conversion of reed canarygrass into ethanol that relies on dilute ammonia, which can be recycled and that requires minimum preparation for fermentation. The pretreated biomass was successfully converted to ethanol using a xylose-fermenting yeast strain, developed by NCAUR scientists, at very good yields (e.g., 81-84%). We have also applied our novel technology for conditioning pretreated biomass for fermentation to rice hulls; a significant agricultural residue. Rice hulls are particularly difficult to ferment following pretreatment because the fermentation microbes die when introduced into the hydrolysates. We have successfully treated rice hulls with dilute-acid and fermented the sugars into ethanol, again using our xylose-fermenting yeast. The strategy relied on using our patented biological abatement approach to remove microbial inhibitors following pretreatment and prior to fermentation. Finally, we have made important strides in our enzyme work. Xylan represents 30-40% of the plant cell wall carbohydrates, and enzymes will be needed to convert xylan to simple sugars so that they can be fermented by yeasts. Enzymatic hydrolysis of this xylan fraction in biomass, even following extensive pretreatment, has given us poor yields (e.g. 28-40%). We have embarked on a rational approach to overcome this barrier. We have collected residual xylan following a yeast fermentation of pretreated switchgrass, which included commercial enzymes for conversion of the carbohydrates into fermentable sugars. There is currently no appropriate method for determining the structure of the recalcitrant xylan. We have developed a novel method for chemically modifying the side-branches of the xylan and thereby allowing determination of specific structure information. This information in turn will be used to improve yields by either altering pretreatment conditions to avoid these xylan structures or supplementing with enzymes capable of specifically hydrolyzing these groups.


4. Accomplishments


Review Publications
Rosenbaum, M., Bar, H.Y., Beg, Q., Segre, D., Booth, J., Cotta, M.A., Angenent, L.T. 2011. Shewanella oneidensis in a lactate-fed pure-culture and a glucose-fed co-culture with Lactococcus lactis with an electrode as electron acceptor. Bioresource Technology. 102(3):2623-2628.

Bowman, M.J., Dien, B.S., O Bryan, P.J., Sarath, G., Cotta, M.A. 2011. Selective chemical oxidation and depolymerization of switchgrass (Panicum virgatum L.) xylan with oligosaccharide product analysis by mass spectrometry. Rapid Communications in Mass Spectrometry. 25(8):941-950.

Nichols, N.N., Sutivisedsak, N., Dien, B.S., Biswas, A., Lesch, W.C., Cotta, M.A. 2011. Conversion of starch from dry common beans (Phaseolus vulgaris L.) to ethanol. Industrial Crops and Products. 33(3):644-647.

Dien, B.S., Miller, D.J., Hector, R.E., Dixon, R.A., Chen, F., McCaslin, M., Risen, P., Sarath, G., Cotta, M.A. 2011. Enhancing alfalfa conversion efficiencies for sugar recovery and ethanol production by altering lignin composition. Bioresource Technology. 102(11):6479-6486.

Mertens, J.A., Bowman, M.J. 2011. Expression and characterization of fifteen Rhizopus oryzae 99-880 polygalacturonase enzymes in Pichia pastoris. Current Microbiology. 62(4):1173-1178.

Ximenes, E., Kim, Y., Mosier, N., Dien, B.S., Ladisch, M. 2011. Deactivation of cellulases by phenols. Enzyme and Microbial Technology. 48(1):54-60.

Fan, Z., Yuan, L., Jordan, D.B., Wagschal, K.C., Heng, C., Braker, J.D. 2010. Engineering lower inhibitor affinities in beta-D-xylosidase. Applied Microbiology and Biotechnology. 86(4):1099-1113.

Jordan, D.B., Wagschal, K.C. 2010. Properties and applications of microbial beta-D-xylosidases. Applied Microbiology and Biotechnology. 86(6):1647-1658

Vogel, K.P., Dien, B.S., Jung, H.G., Casler, M.D., Masterson, S.D., Mitchell, R. 2011. Quantifying actual and theoretical ethanol yields for switchgrass strains using NIRS analyses. BioEnergy Research. 4(2):96-110. DOI: 10.1007/s12155-010-9104-4.

Kim, Y., Hendrickson, R., Mosier, N.S., Ladisch, M.R., Bals, B., Balan, V., Dale, B.E., Dien, B.S., Cotta, M.A. 2010. Effect of compositional variability of Distillers' Grains on cellulosic ethanol production. Bioresource Technology. 101(14):5385-5393.

Saathoff, A.J., Sarath, G., Chow, E.K., Dien, B.S., Tobias, C.M. 2011. Downregulation of cinnamyl-alcohol dehydrogenase in switchgrass by RNA silencing results in enhanced glucose release after cellulase treatment. PLoS One. 6(1):e16416. DOI: 10.1371/journal.pone.0016416.

Arora, A., Seth, A., Dien, B.S., Belyea, R.L., Singh, V., Tumbleson, M.E., Rausch, K.D. 2011. Microfiltration of thin stillage: Process simulation and economic analyses. Biomass and Bioenergy. 35(1):113-120.

Last Modified: 10/18/2017
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