Effect of xylanase supplementation of cellulase on digestion of corn stover solids prepared by leading pretreatment technologies
Introduction
Highly efficient conversion of carbohydrates in biomass to fermentable sugars is essential to commercially competitive biological processes for making cellulosic ethanol (Wyman, 2007, Yang and Wyman, 2008), and although enzymes realize the high yields required, the corresponding high doses are very expensive (Merino Sandra and Cherry, 2007, Tu et al., 2007, Wingren et al., 2005). Furthermore, current commercial enzymes are mainly intended for pulp and paper and food industries and still lack the proportions of different enzymes and their components required for effective biological production of fermentable sugars from cellulosic biomass (Hespell et al., 1997, Merino Sandra and Cherry, 2007). Typically cellulase, β-glucosidase, xylanase, β-xylosidase, and some accessory activities are required to hydrolyze sugar polymers effectively (Bisaria and Ghose, 1981, Kuhad et al., 1997, Saha and Bothast, 1997), with the proportions of each depending on the type of biomass and pretreatment used. Among leading thermochemical pretreatments, those at low pH hydrolyze xylan to xylose and xylooligomers (Allen et al., 2001, Kabel et al., 2007a, Vazquez et al., 2002) but can also degrade both of these (Kumar and Wyman, 2008d, Lloyd and Wyman, 2005, Saeman, 1945). Furthermore, because the resulting degradation products are strong inhibitors to cellulase and fermenting microorganisms (Kumar and Wyman, 2008e, Larsson et al., 1999, Palmqvist et al., 1996), the pretreatment liquor must be detoxified prior to fermentation (Lynd et al., 2008, Palmqvist and Hahn-Hagerdal, 2000, Weil et al., 2002). On the other hand, high pH alkaline pretreatments leave most of the xylan in the solids, with the result that xylanase as well as possibly other accessory enzymes are needed in addition to cellulase to realize high sugar yields (Chandra et al., 2007, Kumar and Wyman, in press, Mosier et al., 2005, Yang and Wyman, 2008). It has been reported that thermochemical or enzymatic removal of xylan enhances cellulose digestion by reducing the xylan coating and linkages to cellulose (Allen et al., 2001, Ishizawa et al., 2007), but the mechanism of why xylan impacts cellulose digestion is still not entirely clear. Furthermore, data has not been developed to compare the effect of supplementing cellulase with xylanase on sugar release from solids prepared by different promising pretreatments.
In this study, baseline sugar release data was developed for a cellulase plus β-glucosidase mass loading of 29.0 mg/g glucan in unpretreated2 corn stover for solids produced by the leading pretreatment technologies of ammonia fiber expansion (AFEX), ammonia recycled percolation (ARP), dilute sulfuric acid, lime, controlled pH, and sulfur dioxide (SO2). Then, xylanase and β-xylosidase supplementation were employed to determine whether these two activities enhanced performance at the same cellulase loading plus a lower cellulase loading of 14.5 mg/g glucan in unpretreated corn stover. In addition, the influence of acetyl content on cellulase–xylanase interactions was studied, and factors and possible mechanisms for enhancement of glucan digestion by xylanase supplementation were also determined.
Section snippets
Materials
Pure cellulose, Avicel PH-101, was purchased from FMC Corporation, Philadelphia, PA (Cat 11365, Lot 1094627). Birchwood xylan was purchased from Sigma Chemicals, St. Louis, MO. Corn stover was generously provided by the National Renewable Energy Laboratory (NREL, Golden, CO) from a lot that they had collected at the Kramer farm in nearby Wray, CO. Solids prepared by corn stover pretreatment were generously given to us by our CAFI partners from Auburn University, Michigan State University, NREL,
Effect of xylanase supplementation on glucose and xylose release
Pretreated corn stover solids were enzymatically hydrolyzed at fixed CTB mass loadings of 14.5 and 29.0 mg/g glucan in unpretreated corn stover with xylanase supplementation to various degrees for a total of 72 h to establish trends in enzyme effectiveness. However, longer hydrolysis times (e.g., 7 days) would likely be employed commercially to capitalize on the additional sugar release expected.
Conclusions
Glucan and xylan digestion data for cellulase plus β-glucosidase mass loadings of 14.5 and 29.0 mg/g glucan in raw corn stover with xylanase mass supplementation ratios of 1 and 5 are summarized in Table 6. Xylanase supplementation substantially increased xylose release from solids resulting from all pretreatments but had less effect on glucose release. Overall, xylanase supplementation enhanced pretreatment performance at a given enzyme loading in the following order of increasing impact with
Acknowledgements
Support from the US Department of Energy Office of the Biomass Program (contract DE-FG36-04GO14017) and the National Institute of Standards and Technology (award 60NANB1D0064) made this research possible. We are also grateful to the Center for Environmental Research and Technology of the Bourns College of Engineering at the University of California, Riverside and the Thayer School of Engineering at Dartmouth College for providing key equipment and facilities. Auburn, Michigan State, Purdue, and
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