Use of Commercial Enzymes to Boost On-Site Enzyme Efficiency


Efficient conversion of lignocellulosic polysaccharides to fermentable sugars is the key for commercial production of biofuels (Wyman 2007; Yang and Wyman 2008). Despite of many technological improvements, pretreatment and utilization of commercially available lignocellulolytic enzymes to break the recalcitrance is limiting the viability of producing biofuels from lignocellulosic materials (Nguyen and Saddler 1991; von Sivers and Zacchi 1995; Wingreini et al. 2005; Merino and Cherry 2007; Tu et al. 2007). Efficient and economically viable ethanol production requires that all sugars produced from cellulose and hemicellulose should be converted to ethanol or other bioproducts (Hahn-Hagerdal et al. 2007; Tomas-Pejo et al. 2008). Enzymatic hydrolysis plays a pivotal role in efficient conversion of the cellulose and hemicelluloses fraction of pretreated biomass materials. The efficiency of enzymatic hydrolysis depends on the pretreatment used and right kind and proportion of enzyme cocktail.

Numerous studies to investigate the potential of softwood conversion to ethanol have been done in the last two decades (von Sivers and Zacchi 1995; Schell et al. 1998; Boussaid et al. 1999). Pretreatment conditions affect the requisite enzyme mixture employed for polysaccharide hydrolysis. Previously, acid- and SO2- catalyzed steam explosion was extensively used to pretreat softwood for enhancing the enzymatic hydrolysis (Mackie et al. 1985; Ramos et al. 1992; Wu et al. 1999; Boussaid et al. 2000; Yang et al. 2002; Kumar et al. 2010). Among different pretreatment methods developed to date, wet explosion (WEx) is considered one of the most appropriate and cost-effective methods for deconstruction of softwood and has demonstrated unparalleled performance for fermentable sugar production as discussed by Rana et al. (2012).

In-house enzyme production has recently gain interest for reducing the cost associated with enzymatic hydrolysis, but the efficiency of those crude enzymes is limited compared to commercial enzymes. Still commercial enzymes are expensive © The Author(s) 2017

V. Rana, D. Rana, Renewable Biofuels, SpringerBriefs in Applied Sciences and Technology, DOI 10.1007/978-3-319-47379-6_4

and have been problematic for the exploitation of lignocellulosics for production of biofuels (Hespell et al. 1997; Merino and Cherry 2007). In the present study, we examine the possibility of supplementing in-house produced enzymes with small amounts of commercial enzymes to obtain sufficient hydrolytic effects with reduced dosages of commercial enzymes, thereby lowering the overall cost of enzymatic hydrolysis.

Fungal-derived cellulolytic enzymes were investigated extensively in the last few decades for the hydrolysis of lignocelluloses (Persson et al. 1991; Saddler 1998). Lignocellulolytic enzymes can be produced by a diverse group of fungi including ascomycetes (e.g., T. reesei), basidiomycetes including white-rot fungi (e.g., P. chrysosporium), brown-rot fungi (e.g., Fomitopsis palustris), and some anaerobic species (e.g., Orpinomyces sp.) which degrade cellulose in the gastrointestinal tract of ruminant animals (Ljungdahl 2008). Out of these strains, aerobic fungal strains specifically T. reesei are of interest as it produces large amounts of extracellular cellulolytic enzymes when grown in liquid culture. Lignocellulose degradation requires three enzymatic components/domains: EG, CBH, and BG; however, none of the fungal strains including the best mutants described are able to produce all three required enzyme components at the same time (Dashtban et al. 2009). T. reesei exhibits high cellobiohydrolase (CBH) and endoglucanase (EG) activities but lack sufficient p-glucosidase activities (Stockton et al. 1991; Kumar et al. 2008). Therefore, in order to achieve good cellulose hydrolysis, T. reesei cel- lulases are typically supplemented with p-glucosidase from Aspergillus niger.

Fungal strains displaying cellulolytic activity are capable of degradation cellulose and have great potential to be used in a consolidated biorefinery for cost- effective biofuel production, but their efficiency is generally low compared to commercial enzymes. A substantial amount of research has been done to investigate the potential benefit of accessory enzymes to supplement commercial cellulases such as commercial xylanase, arabinase, mannanase, pectinase, and other auxiliary enzymes which substantially can reduce the amount of enzymes required for efficient biomass hydrolysis of acid or alkaline pretreated biomass. Alvira et al. (2011) observed that endoxylanase and a-L-arabinofuranosidase supplementation during enzymatic hydrolysis of steam-exploded wheat straw increased the enzymatic hydrolysis yield by 10 %. According to Kumar and coworkers (1999) (Kumar and Wyman 2009), an incremental increase in glucose release was observed with xylanase supplementation. Varnai et al. (2011) observed that addition of endo-p- mannanase increased the overall hydrolysis yield by 20-25 % of cellulose. Despite a large research effort within this field, there is still a lack of understanding of the effect of supplementing low-cost in-house enzymes with small amounts of commercial enzymes for hydrolyzing pretreated biomass materials such as softwood.

The overall objective of this study is to evaluate the effect and potential application of supplementing in-house produced enzymes with commercial enzymes for reducing the overall cost of enzymatic hydrolysis without sacrificing the hydrolytic efficiency. The proposed research is innovative as we use hydrolysis of softwood which has proven to be more difficult than degrading pretreated agricultural residues. In this study, sugar yields from hydrolyzing wet-exploded loblolly pine under three different scenarios are compared: scenario 1, in-house produced enzymes; scenario 2, commercial enzymes; and scenario 3, in-house enzymes added with commercial enzymes with the objective of finding the lowest amount of commercial enzymes that is needed to achieve optimal saccharification yields when using inhouse produced enzymes.

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