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Abstract Background Pretreatment of lignocellulosic biomass (LCB) is a key step for its efficient bioconversion into ethanol. Determining the best pretreatment and its parameters requires monitoring its impacts on the biomass material. Here, we used fluorescent protein-tagged carbohydrate-binding modules method (FTCM)-depletion assay to study the relationship between surface-exposed polysaccharides and enzymatic hydrolysis of LCB. Results Our results indicated that alkali extrusion pretreatment led to the highest hydrolysis rates for alfalfa stover, cattail stems and flax shives, despite its lower lignin removal efficiency compared to alkali pretreatment. Corn crop residues were more sensitive to alkali pretreatments, leading to higher hydrolysis rates. A clear relationship was consistently observed between total surface-exposed cellulose detected by the FTCM-depletion assay and biomass enzymatic hydrolysis. Comparison of bioconversion yield and total composition analysis (by NREL/TP-510-42618) of LCB prior to or after pretreatments did not show any close relationship. Lignin removal efficiency and total cellulose content (by NREL/TP-510-42618) led to an unreliable prediction of enzymatic polysaccharide hydrolysis. Conclusions Fluorescent protein-tagged carbohydrate-binding modules method (FTCM)-depletion assay provided direct evidence that cellulose exposure is the key determinant of hydrolysis yield. The clear and robust relationships that were observed between the cellulose accessibility by FTCM probes and enzymatic hydrolysis rates change could be evolved into a powerful prediction tool that might help develop optimal biomass pretreatment strategies for biofuel production.

Abstract Background Lignocellulosic biomass will progressively become the main source of carbon for a number of products as the Earth’s oil reservoirs disappear. Technology for conversion of wood fiber into bioproducts (wood biorefining) continues to flourish, and access to reliable methods for monitoring modification of such fibers is becoming an important issue. Recently, we developed a simple, rapid approach for detecting four different types of polymer on the surface of wood fibers. Named fluorescent-tagged carbohydrate-binding module (FTCM), this method is based on the fluorescence signal from carbohydrate-binding modules-based probes designed to recognize specific polymers such as crystalline cellulose, amorphous cellulose, xylan, and mannan. Results Here we used FTCM to characterize pulps made from softwood and hardwood that were prepared using Kraft or chemical-thermo-mechanical pulping. Comparison of chemical analysis (NREL protocol) and FTCM revealed that FTCM results were consistent with chemical analysis of the hemicellulose composition of both hardwood and softwood samples. Kraft pulping increased the difference between softwood and hardwood surface mannans, and increased xylan exposure. This suggests that Kraft pulping leads to exposure of xylan after removal of both lignin and mannan. Impact of enzyme cocktails from Trichoderma reesei (Celluclast 1.5L) and from Aspergillus sp. (Carezyme 1000L) was investigated by analysis of hydrolyzed sugars and by FTCM. Both enzymes preparations released cellobiose and glucose from pulps, with the cocktail from Trichoderma being the most efficient. Enzymatic treatments were not as effective at converting chemical-thermomechanical pulps to simple sugars, regardless of wood type. FTCM revealed that amorphous cellulose was the primary target of either enzyme preparation, which resulted in a higher proportion of crystalline cellulose on the surface after enzymatic treatment. FTCM confirmed that enzymes from Aspergillus had little impact on exposed hemicelluloses, but that enzymes from the more aggressive Trichoderma cocktail reduced hemicelluloses at the surface. Conclusions Overall, this study indicates that treatment with enzymes from Trichoderma is appropriate for generating crystalline cellulose at fiber surface. Applications such as nanocellulose or composites requiring chemical resistance would benefit from this enzymatic treatment. The milder enzyme mixture from Aspergillus allowed for removal of amorphous cellulose while preserving hemicelluloses at fiber surface, which makes this treatment appropriate for new paper products where surface chemical responsiveness is required.