Dataset Information

Overcoming biomass recalcitrance by synergistic pretreatment of mechanical activation and metal salt for enhancing enzymatic conversion of lignocellulose.

ABSTRACT:

Background

Due to biomass recalcitrance, including complexity of lignocellulosic matrix, crystallinity of cellulose, and inhibition of lignin, the bioconversion of lignocellulosic biomass is difficult and inefficient. The aim of this study is to investigate an effective and green pretreatment method for overcoming biomass recalcitrance of lignocellulose.

Results

An effective mechanical activation (MA)?+?metal salt (MAMS) technology was applied to pretreat sugarcane bagasse (SCB), a typical kind of lignocellulosic biomass, in a stirring ball mill. Chlorides and nitrates of Al and Fe showed better synergistic effect with MA, especially AlCl₃, ascribing to the interaction between metal salt and oxygen-containing groups induced by MA. Comparative studies showed that MAMS pretreatment effectively changed the recalcitrant structural characteristics of lignocellulosic matrix and reduced the inhibitory action of lignin on enzymatic conversion of SCB. The increase in hydroxyl and carboxyl groups of lignin induced by MAMS pretreatment led to the increase of its hydrophilicity, which could weaken the binding force between cellulase and lignin and reduce the nonproductive binding of cellulase enzymes to lignin.

Conclusions

MAMS pretreatment significantly enhanced the enzymatic digestibility of polysaccharides substrate by overcoming biomass recalcitrance without the removal of lignin from enzymatic hydrolysis system.

SUBMITTER: Zhang Y

PROVIDER: S-EPMC6327530 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Publications

Overcoming biomass recalcitrance by synergistic pretreatment of mechanical activation and metal salt for enhancing enzymatic conversion of lignocellulose.

Zhang Yanjuan Y Huang Min M Su Jianmei J Hu Huayu H Yang Mei M Huang Zuqiang Z Chen Dong D Wu Juan J Feng Zhenfei Z

Biotechnology for biofuels 20190110

<h4>Background</h4>Due to biomass recalcitrance, including complexity of lignocellulosic matrix, crystallinity of cellulose, and inhibition of lignin, the bioconversion of lignocellulosic biomass is difficult and inefficient. The aim of this study is to investigate an effective and green pretreatment method for overcoming biomass recalcitrance of lignocellulose.<h4>Results</h4>An effective mechanical activation (MA) + metal salt (MAMS) technology was applied to pretreat sugarcane bagasse (SCB), ...[more]

PMID: 30647772

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Project description:BackgroundLignocellulose biomass is known as a recalcitrant material towards enzymatic hydrolysis, increasing the process cost in biorefinery. In nature, filamentous fungi naturally degrade lignocellulose, using an arsenal of hydrolytic and oxidative enzymes. Assessment of enzyme hydrolysis efficiency generally relies on the yield of glucose for a given biomass. To better understand the markers governing recalcitrance to enzymatic degradation, there is a need to enlarge the set of parameters followed during deconstruction.ResultsIndustrially-pretreated biomass feedstocks from wheat straw, miscanthus and poplar were sequentially hydrolysed following two steps. First, standard secretome from Trichoderma reesei was used to maximize cellulose hydrolysis, producing three recalcitrant lignin-enriched solid substrates. Then fungal secretomes from three basidiomycete saprotrophs (Laetisaria arvalis, Artolenzites elegans and Trametes ljubarskyi) displaying various hydrolytic and oxidative enzymatic profiles were applied to these recalcitrant substrates, and compared to the T. reesei secretome. As a result, most of the glucose was released after the first hydrolysis step. After the second hydrolysis step, half of the remaining glucose amount was released. Overall, glucose yield after the two sequential hydrolyses was more dependent on the biomass source than on the fungal secretomes enzymatic profile. Solid residues obtained after the two hydrolysis steps were characterized using complementary methodologies. Correlation analysis of several physico-chemical parameters showed that released glucose yield was negatively correlated with lignin content and cellulose crystallinity while positively correlated with xylose content and water sorption. Water sorption appears as a pivotal marker of the recalcitrance as it reflects chemical and structural properties of lignocellulosic biomass.ConclusionsFungal secretomes applied to highly recalcitrant biomass samples can further extend the release of the remaining glucose. The glucose yield can be correlated to chemical and physical markers, which appear to be independent from the biomass type and secretome. Overall, correlations between these markers reveal how nano-scale properties (polymer content and organization) influence macro-scale properties (particle size and water sorption). Further systematic assessment of these markers during enzymatic degradation will foster the development of novel cocktails to unlock the degradation of lignocellulose biomass.

Project description:BackgroundBiomass recalcitrance towards pretreatment and further processing can be related to the compositional and structural features of the biomass. However, the exact role and relative importance to those structural attributes has still to be further evaluated. Herein, ten different types of biomass currently considered to be important raw materials for biorefineries were chosen to be processed by the recently developed, acid-catalyzed OrganoCat pretreatment to produce cellulose-enriched pulp, sugars, and lignin with different amounts and qualities. Using wet chemistry analysis and NMR spectroscopy, the generic factors of lignocellulose recalcitrance towards OrganoCat were determined.ResultsThe different materials were processed applying different conditions (e.g., type of acid catalyst and temperature), and fractions with different qualities were obtained. Raw materials and products were characterized in terms of their compositional and structural features. For the first time, generic correlation coefficients were calculated between the measured chemical and structural features and the different OrganoCat product yields and qualities. Especially lignin-related factors displayed a detrimental role for enzymatic pulp hydrolysis, as well as sugar and lignin yield exhibiting inverse correlation coefficients. Hemicellulose appeared to have less impact, not being as detrimental as lignin factors, but xylan-O-acetylation was inversely correlated with product yield and qualities.ConclusionThese results illustrate the role of generic features of lignocellulosic recalcitrance towards acidic pretreatments and fractionation, exemplified in the OrganoCat strategy. Discriminating between types of lignocellulosic biomass and highlighting important compositional variables, the improved understanding of how these parameters affect OrganoCat products will ameliorate bioeconomic concepts from agricultural production to chemical products. Herein, a methodological approach is proposed.

Project description:BackgroundThe recalcitrance of cellulosic biomass is widely recognized as a key barrier to cost-effective biological processing to fuels and chemicals, but the relative impacts of physical, chemical and genetic interventions to improve biomass processing singly and in combination have yet to be evaluated systematically. Solubilization of plant cell walls can be enhanced by non-biological augmentation including physical cotreatment and thermochemical pretreatment, the choice of biocatalyst, the choice of plant feedstock, genetic engineering of plants, and choosing feedstocks that are less recalcitrant natural variants. A two-tiered combinatoric investigation of lignocellulosic biomass deconstruction was undertaken with three biocatalysts (Clostridium thermocellum, Caldicellulosiruptor bescii, Novozymes Cellic® Ctec2 and Htec2), three transgenic switchgrass plant lines (COMT, MYB4, GAUT4) and their respective nontransgenic controls, two Populus natural variants, and augmentation of biological attack using either mechanical cotreatment or cosolvent-enhanced lignocellulosic fractionation (CELF) pretreatment.ResultsIn the absence of augmentation and under the conditions tested, increased total carbohydrate solubilization (TCS) was observed for 8 of the 9 combinations of switchgrass modifications and biocatalysts tested, and statistically significant for five of the combinations. Our results indicate that recalcitrance is not a trait determined by the feedstock only, but instead is coequally determined by the choice of biocatalyst. TCS with C. thermocellum was significantly higher than with the other two biocatalysts. Both CELF pretreatment and cotreatment via continuous ball milling enabled TCS in excess of 90%.ConclusionBased on our results as well as literature studies, it appears that some form of non-biological augmentation will likely be necessary for the foreseeable future to achieve high TCS for most cellulosic feedstocks. However, our results show that this need not necessarily involve thermochemical processing, and need not necessarily occur prior to biological conversion. Under the conditions tested, the relative magnitude of TCS increase was augmentation?>?biocatalyst choice?>?plant choice?>?plant modification?>?plant natural variants. In the presence of augmentation, plant modification, plant natural variation, and plant choice exhibited a small, statistically non-significant impact on TCS.

Project description:BackgroundBiomass recalcitrance to enzymatic hydrolysis has been assigned to several structural and chemical factors. However, their relative importance remains challenging to evaluate. Three representative biomass species (wheat straw, poplar and miscanthus) were submitted to four standard pretreatments (dilute acid, hot water, ionic liquid and sodium chlorite) in order to generate a set of contrasted samples. A large array of techniques, including wet chemistry analysis, porosity measurements using NMR spectroscopy, electron and fluorescence microscopy, were used in order to determine possible generic factors of biomass recalcitrance.ResultsThe pretreatment conditions selected allowed obtaining samples displaying different susceptibility to enzymatic hydrolysis (from 3 up to 98% of the initial glucose content released after 96 h of saccharification). Generic correlation coefficients were calculated between the measured chemical and structural features and the final saccharification rates. Increases in porosity displayed overall strong positive correlations with saccharification efficiency, but different porosity ranges were concerned depending on the considered biomass. Lignin-related factors displayed highly negative coefficients for all biomasses. Lignin content, which is likely involved in the correlations observed for porosity, was less detrimental to enzymatic hydrolysis than lignin composition. Lignin influence was highlighted by the strong negative correlation with fluorescence intensity which mainly originates from monolignols in mature tissues.ConclusionsOur results provide a better understanding of the factors responsible for biomass recalcitrance that can reasonably be considered as generic. The correlations with specific porosity ranges are biomass species-dependent, meaning that enzymes cocktails with fitted enzyme size are likely to be needed to optimise saccharification depending on the biomass origin. Lignin composition, which probably influences its structure, is the most important parameter to overcome to enhance enzymes access to the polysaccharides. Accordingly, fluorescence intensity was found to be a rapid and simple method to assess recalcitrance after pretreatment.

Project description:BackgroundLow-temperature swelling of cotton linter cellulose and subsequent gelatinization in trifluoroacetic acid (TFA) greatly enhance rates of enzymatic digestion or maleic acid-AlCl3 catalyzed conversion to hydroxymethylfurfural (HMF) and levulinic acid (LA). However, lignin inhibits low-temperature swelling of TFA-treated intact wood particles from hybrid poplar (Populus tremula?×?P. alba) and results in greatly reduced yields of glucose or catalytic conversion compared to lignin-free cellulose. Previous studies have established that wood particles from transgenic lines of hybrid poplar with high syringyl (S) lignin content give greater glucose yields following enzymatic digestion.ResultsLow-temperature (-?20 °C) treatment of S-lignin-rich poplar wood particles in TFA slightly increased yields of glucose from enzymatic digestions and HMF and LA from maleic acid-AlCl3 catalysis. Subsequent gelatinization at 55 °C resulted in over 80% digestion of cellulose in only 3 to 6 h with high-S-lignin wood, compared to 20-60% digestion in the wild-type poplar hybrid and transgenic lines high in guaiacyl lignin or 5-hydroxy-G lignin. Disassembly of lignin in woody particles by Ni/C catalytic systems improved yields of glucose by enzymatic digestion or catalytic conversion to HMF and LA. Although lignin was completely removed by Ni/C-catalyzed delignification (CDL) treatment, recalcitrance to enzymatic digestion of cellulose from the high-S lines was reduced compared to other lignin variants. However, cellulose still exhibited considerable recalcitrance to complete enzymatic digestion or catalytic conversion after complete delignification. Low-temperature swelling of the CDL-treated wood particles in TFA resulted in nearly complete enzymatic hydrolysis, regardless of original lignin composition.ConclusionsGenetic modification of lignin composition can enhance the portfolio of aromatic products obtained from lignocellulosic biomass while promoting disassembly into biofuel and bioproduct substrates. CDL enhances rates of enzymatic digestion and chemical conversion, but cellulose remains intrinsically recalcitrant. Cold TFA is sufficient to overcome this recalcitrance after CDL treatment. Our results inform a 'no carbon left behind' strategy to convert total woody biomass into lignin, cellulose, and hemicellulose value streams for the future biorefinery.

Project description:Background:During the chemical and biochemical decomposition of lignocellulosic biomasses, lignin is highly recalcitrant. Genetic transformation of plants to qualitatively and/or quantitatively modify lignin may reduce these recalcitrant properties. Efficient discovery of genes to achieve lignin manipulation is thus required. Results:To screen for new genes to reduce lignin recalcitrance, we heterologously expressed 50 enzymatic genes under the control of a cinnamate 4-hydroxylase (C4H) gene promoter, derived from a hybrid aspen, which is preferentially active in tissues with lignified cell walls in Arabidopsis plants. These genes encode enzymes that act on metabolites in shikimate, general phenylpropanoid, flavonoid, or monolignol biosynthetic pathways. Among these genes, 30, 18, and 2 originated from plants, bacteria, and fungi, respectively. In our first screening step, 296 independent transgenic plants (T1 generation) harboring single or multiple transgenes were generated from pools of seven Agrobacterium strains used for conventional floral-dip transformation. Wiesner and Mäule staining patterns in the stems of the resultant plants revealed seven and nine plants with apparent abnormalities in the two respective staining analyses. According to genomic PCR and subsequent direct sequencing, each of these 16 plants possessed a gene encoding either coniferaldehyde dehydrogenase (calB), feruloyl-CoA 6'-hydroxylase (F6H1), hydroxycinnamoyl-CoA hydratase/lyase (couA), or ferulate 5-hydroxylase (F5H), with one transgenic plant carrying both calB and F6H1. The effects of these genes on lignin manipulation were confirmed in individually re-created T1 transgenic Arabidopsis plants. While no difference in lignin content was detected in the transgenic lines compared with the wild type, lignin monomeric composition was changed in the transgenic lines. The observed compositional change in the transgenic plants carrying calB, couA, and F5H led to improved sugar release from cell walls after alkaline pretreatment. Conclusions:Simple colorimetric characterization of stem lignin is useful for simultaneous screening of many genes with the potential to reduce lignin recalcitrance. In addition to F5H, the positive control, we identified three enzyme-coding genes that can function as genetic tools for lignin manipulation. Two of these genes (calB and couA) accelerate sugar release from transgenic lignocelluloses.

Project description:BackgroundBiological conversion of lignocellulosic biomass is significantly hindered by feedstock recalcitrance, which is typically assessed through an enzymatic digestion assay, often preceded by a thermal and/or chemical pretreatment. Here, we assay 17 lines of unpretreated transgenic black cottonwood (Populus trichocarpa) utilizing a lignocellulose-degrading, metabolically engineered bacterium, Caldicellulosiruptor bescii. The poplar lines were assessed by incubation with an engineered C. bescii strain that solubilized and converted the hexose and pentose carbohydrates to ethanol and acetate. The resulting fermentation titer and biomass solubilization were then utilized as a measure of biomass recalcitrance and compared to data previously reported on the transgenic poplar samples.ResultsOf the 17 transgenic poplar lines examined with C. bescii, a wide variation in solubilization and fermentation titer was observed. While the wild type poplar control demonstrated relatively high recalcitrance with a total solubilization of only 20% and a fermentation titer of 7.3 mM, the transgenic lines resulted in solubilization ranging from 15 to 79% and fermentation titers from 6.8 to 29.6 mM. Additionally, a strong inverse correlation (R 2 = 0.8) between conversion efficiency and lignin content was observed with lower lignin samples more easily converted and solubilized by C. bescii.ConclusionsFeedstock recalcitrance can be significantly reduced with transgenic plants, but finding the correct modification may require a large sample set to identify the most advantageous genetic modifications for the feedstock. Utilizing C. bescii as a screening assay for recalcitrance, poplar lines with down-regulation of coumarate 3-hydroxylase 3 (C3H3) resulted in the highest degrees of solubilization and conversion by C. bescii. One such line, with a growth phenotype similar to the wild-type, generated more than three times the fermentation products of the wild-type poplar control, suggesting that excellent digestibility can be achieved without compromising fitness of the tree.

Dataset Information

Overcoming biomass recalcitrance by synergistic pretreatment of mechanical activation and metal salt for enhancing enzymatic conversion of lignocellulose.

Background

Results

Conclusions

Publications

Overcoming biomass recalcitrance by synergistic pretreatment of mechanical activation and metal salt for enhancing enzymatic conversion of lignocellulose.

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