• Energy Research

Developing feedstock-independent biomass pretreatment would be vital to second generation biorefineries that would fully utilize diverse non-food lignocellulosic biomass resources, decrease transportation costs of low energy density feedstock, and conserve natural biodiversity. Cellulose solvent- and organic solvent-based lignocellulose fractionation (COSLIF) was applied to a variety of feedstocks, including Miscanthus, poplar, their mixture, bagasse, wheat straw, and rice straw. Although non-pretreated biomass samples exhibited a large variation in enzymatic digestibility, the COSLIF-pretreated biomass samples exhibited similar high enzymatic glucan digestibilities and fast hydrolysis rates. Glucan digestibilities of most pretreated feedstocks were ∼93% at five filter paper units per gram of glucan. The overall glucose and xylose yields for the Miscanthus:poplar mixture at a weight ratio of 1:2 were 93% and 85%, respectively. These results suggested that COSLIF could be regarded as a feedstock-independent pretreatment suitable for processing diverse feedstocks by adjusting pretreatment residence time only.

Fractionated corn bran was processed to maximize ethanol production from starch, cellulose, and xylan. After various bench-scale experiments, an optimized process with dilute acid pretreatment (1.5 % w/w H2SO4) at 90 °C for 60 min was utilized followed by enzymatic hydrolysis using cellulase and hemicellulase for 48 hr. After simultaneous saccharification (regarding starch) and fermentation at 150 L using an engineered yeast, which consumes both glucose and xylose to make ethanol, the 86 % total sugar conversion yield was achieved, including conversions of 95 % for starch, 77 % for cellulose and 77 % for xylan. Also, an accurate mass balance was formulated for ethanol-producing carbohydrates including starch, cellulose, and xylan from feedstock to final ethanol. A highly efficient process of converting corn fiber to ethanol was successfully scaled up to 150 L.

AbstractWhile many pretreatments attempt to improve the enzymatic digestibility of biomass by removing lignin, this study shows that improving the surface area accessible to cellulase is a more important factor for achieving a high sugar yield. Here we compared the pretreatment of switchgrass by two methods, cellulose solvent‐ and organic solvent‐based lignocellulose fractionation (COSLIF) and soaking in aqueous ammonia (SAA). Following pretreatment, enzymatic hydrolysis was conducted at two cellulase loadings, 15 filter paper units (FPU)/g glucan and 3 FPU/g glucan, with and without BSA blocking of lignin absorption sites. The hydrolysis results showed that the lignin remaining after SAA had a significant negative effect on cellulase performance, despite the high level of delignification achieved with this pretreatment. No negative effect due to lignin was detected for COSLIF‐treated substrate. SEM micrographs, XRD crystallinity measurements, and cellulose accessibility to cellulase (CAC) determinations confirmed that COSLIF fully disrupted the cell wall structure, resulting in a 16‐fold increase in CAC, while SAA caused a 1.4‐fold CAC increase. A surface plot relating the lignin removal, CAC, and digestibility of numerous samples (both pure cellulosic substrates and lignocellulosic materials pretreated by several methods) was also developed to better understand the relative impacts of delignification and CAC on glucan digestibility. Biotechnol. Bioeng. 2011; 108:22–30. © 2010 Wiley Periodicals, Inc.

Biomass was heated (200-240°C) in the presence of glycerol, for 4-12 min, under shear to disrupt the native cell wall architecture. The impact of this method, named glycerol thermal processing (GTP), on saccharification efficiency of the hardwood Liquidambar styraciflua, and a control cellulose sample was studied as a function of treatment severity. Furthermore, the enzymatic conversion of samples with varying compositions was studied after extraction of the structural polymers. Interestingly, the sweet gum processed materials crystallinity index increased by 10% of the initial value. The experiments revealed that the residual lignin was not a barrier to limiting the digestibility of cellulose after pretreatment yielding up to 70% glucose based on the starting wood material. Further xylan removal greatly improved the cellulose hydrolysis rate, converting nearly 70% of the cellulose into glucose within 24h, and reaching 78% of ultimate glucan digestibility after 72 h.

Decreasing lignin content of plant biomass by genetic engineering is believed to mitigate biomass recalcitrance and improve saccharification efficiency of plant biomass. In this study, we compared two different pretreatment methods (i.e., dilute acid and cellulose solvent) on transgenic plant biomass samples having different lignin contents and investigated biomass saccharification efficiency. Without pretreatment, no correlation was observed between lignin contents of plant biomass and saccharification efficiency. After dilute acid pretreatment, a strong negative correlation between lignin content of plant samples and overall glucose release was observed, wherein the highest overall enzymatic glucan digestibility was 70% for the low-lignin sample. After cellulose solvent- and organic solvent-based lignocellulose fractionation pretreatment, there was no strong correlation between lignin contents and high saccharification efficiencies obtained (i.e., 80-90%). These results suggest that the importance of decreasing lignin content in plant biomass to saccharification was largely dependent on pretreatment choice and conditions.

ABSTRACT Microbial communities that deconstruct plant biomass have broad relevance in biofuel production and global carbon cycling. Biomass pretreatments reduce plant biomass recalcitrance for increased efficiency of enzymatic hydrolysis. We exploited these chemical pretreatments to study how thermophilic bacterial consortia adapt to deconstruct switchgrass (SG) biomass of various compositions. Microbial communities were adapted to untreated, ammonium fiber expansion (AFEX)-pretreated, and ionic-liquid (IL)-pretreated SG under aerobic, thermophilic conditions using green waste compost as the inoculum to study biomass deconstruction by microbial consortia. After microbial cultivation, gravimetric analysis of the residual biomass demonstrated that both AFEX and IL pretreatment enhanced the deconstruction of the SG biomass approximately 2-fold. Two-dimensional nuclear magnetic resonance (2D-NMR) experiments and acetyl bromide-reactive-lignin analysis indicated that polysaccharide hydrolysis was the dominant process occurring during microbial biomass deconstruction, and lignin remaining in the residual biomass was largely unmodified. Small-subunit (SSU) rRNA gene amplicon libraries revealed that although the dominant taxa across these chemical pretreatments were consistently represented by members of the Firmicutes , the Bacteroidetes , and Deinococcus-Thermus , the abundance of selected operational taxonomic units (OTUs) varied, suggesting adaptations to the different substrates. Combining the observations of differences in the community structure and the chemical and physical structure of the biomass, we hypothesize specific roles for individual community members in biomass deconstruction.

Alcohols of increasing alkyl chain length were investigated as precipitants in an ionic liquid (IL) pretreatment system. Switchgrass samples pretreated by 1-ethyl-3-methylimidazolium acetate were characterized after the use of different alkyl chain lengths of alcohols as antisolvents. The resulting IL-pretreated switchgrass (PSG) samples were characterized by enzymatic hydrolysis, cross polarization/magic angle spinning (CP/MAS) 13C nuclear magnetic resonance (NMR), Fourier transform infrared spectroscopy (FTIR), and 2D NMR spectroscopy. Glucan digestibilities of PSG samples were ∼80 % after 72 h at 5 mg protein g−1 glucan regardless of the antisolvent used. The use of 1-octanol as an antisolvent, with 10 % water to allow for use of wet biomass, enabled a partial lignin fractionation and multiphase separation for the IL recycle without compromising the chemical structure of the carbohydrates and lignin from the PSG. Lignin fragments were observed in the IL after pretreatment by gel permeation chromatography (GPC). After separation, both the IL and the octanol antisolvent were reused for switchgrass pretreatment and precipitation for an additional 3 cycles. The PSG samples derived from recycled IL were rapidly hydrolyzed, and a high glucan digestibility of 80 % was obtained even at a low enzyme loading of 5 mg protein g−1 glucan. 2D NMR analysis of residual solids of PSG post-enzymatic hydrolysis revealed that lignin in these residual solids was depolymerized. This strategy enables an ease in separation of pretreated lignocellulosic solids, reduced water use, and recycle of both IL and the antisolvent.

Effective hydrolysis of pretreated lignocellulose mediated by cellulase requires an in-depth understanding of cellulase adsorption and desorption. Here we developed a simple method for determining the adsorbed cellulase on cellulosic materials or pretreated lignocellulose, which involves (i) hydrolysis of adsorbed cellulase in the presence of 10 M of NaOH at 121 degrees C for 20 min, and (ii) the ninhydrin assay for the amino acids released from the hydrolyzed cellulase. The major lignocellulosic components (i.e., cellulose, hemicellulose, and lignin) did not interfere with the ninhydrin assay. A number of cellulase desorption methods were investigated: pH change as well as the use of detergents, high salt solution, and polyhydric alcohols. The pH adjustment to 13.0 and the elution by 72% ethylene glycol at neutral pH were among the most efficient approaches for desorbing the adsorbed cellulase. For the recycling of active cellulase, a modest pH adjustment to 10.0 may be a low-cost viable method to desorb active cellulase. It was found that more than 90% of cellulase for hydrolysis of the pretreated corn stover could be recycled by washing at pH 10.0.