Rui Katahira - Academia.edu (original) (raw)
Papers by Rui Katahira
OSTI OAI (U.S. Department of Energy Office of Scientific and Technical Information), May 1, 2018
Chemsuschem, Nov 16, 2022
Lignin is the largest source of bio‐based aromatic compounds in nature, and its valorization is e... more Lignin is the largest source of bio‐based aromatic compounds in nature, and its valorization is essential to the sustainability of lignocellulosic biorefining. Characterizing lignin‐derived compounds remains challenging due to the heterogeneity of this biopolymer. Tandem mass spectrometry is a promising tool for lignin structural analytics, as fragmentation patterns of model compounds can be extrapolated to identify characteristic moieties in complex samples. This work extended previous resonance excitation‐type collision‐induced dissociation (CID) methods that identified lignin oligomers containing β–O–4, β–5, and β–β bonds, to also identify characteristics of 5–5, β–1, and 4–O–5 dimers, enabled by quadrupole time‐of‐flight (QTOF) CID with energy‐resolved mass spectrometry (ERMS). Overall, QTOF‐ERMS offers in‐depth structural information and could ultimately contribute to tools for high‐throughput lignin dimer identification.
Journal of Wood Chemistry and Technology, Jan 4, 2003
ABSTRACT A novel degradation method of arylglycerol-β-aryl (β-O-4) ether substructure in lignins ... more ABSTRACT A novel degradation method of arylglycerol-β-aryl (β-O-4) ether substructure in lignins without any secondary condensation reactions named TIZ method was developed. The TIZ method consists of three reaction steps: (1) selective tosylation (T) of primary hydroxyl group, (2) iodination (I), and (3) zinc-metal treatment (Z) to cleave of β-O-4 linkage. This method was applied to two nonphenolic β-O-4 lignin substructure compounds of guaiacyl and syringyl type: benzylguaiacylglycerol-β-guaiacyl ether (1a) and benzylsyringylglycerol-β-syringyl ether (1b). Respective reaction steps were performed quantitatively concerning both guaiacyl and syringyl models. After zinc treatment, almost 1 mole of olefinic compounds, 3-benzylguaiacyl-3-hydroxy-1-propene (4a) and 3-benzylsyringyl-3-hydroxy-1-propene (4b), were formed from 1 mol of compound 1a and 1b, respectively. This indicates that β-O-4 linkage in compound 1a and 1b was cleaved quantitatively without formation of any by-products. The structures of the degradation products from compound 1a and 1b by TIZ method were determined by H and C NMR spectroscopy and GC–MS, and by comparison with authentic compounds synthesized independently. The degradation products were quantified by GC. The TIZ method, carried out under weak alkaline and neutral reaction conditions, was shown to be useful for the selective cleavage of β-O-4 linkage of compound 1a and 1b without secondary condensation reaction on side chain. Therefore, this method is an efficient analytical method for lignin structure studies.
Metabolic Engineering, May 1, 2021
Biotechnology for Biofuels, Oct 19, 2011
Background: In converting biomass to bioethanol, pretreatment is a key step intended to render ce... more Background: In converting biomass to bioethanol, pretreatment is a key step intended to render cellulose more amenable and accessible to cellulase enzymes and thus increase glucose yields. In this study, four cellulose samples with different degrees of polymerization and crystallinity indexes were subjected to aqueous sodium hydroxide and anhydrous liquid ammonia treatments. The effects of the treatments on cellulose crystalline structure were studied, in addition to the effects on the digestibility of the celluloses by a cellulase complex. Results: From X-ray diffractograms and nuclear magnetic resonance spectra, it was revealed that treatment with liquid ammonia produced the cellulose III I allomorph; however, crystallinity depended on treatment conditions. Treatment at a low temperature (25°C) resulted in a less crystalline product, whereas treatment at elevated temperatures (130°C or 140°C) gave a more crystalline product. Treatment of cellulose I with aqueous sodium hydroxide (16.5 percent by weight) resulted in formation of cellulose II, but also produced a much less crystalline cellulose. The relative digestibilities of the different cellulose allomorphs were tested by exposing the treated and untreated cellulose samples to a commercial enzyme mixture (Genencor-Danisco; GC 220). The digestibility results showed that the starting cellulose I samples were the least digestible (except for corn stover cellulose, which had a high amorphous content). Treatment with sodium hydroxide produced the most digestible cellulose, followed by treatment with liquid ammonia at a low temperature. Factor analysis indicated that initial rates of digestion (up to 24 hours) were most strongly correlated with amorphous content. Correlation of allomorph type with digestibility was weak, but was strongest with cellulose conversion at later times. The cellulose III I samples produced at higher temperatures had comparable crystallinities to the initial cellulose I samples, but achieved higher levels of cellulose conversion, at longer digestion times. Conclusions: Earlier studies have focused on determining which cellulose allomorph is the most digestible. In this study we have found that the chemical treatments to produce different allomorphs also changed the crystallinity of the cellulose, and this had a significant effect on the digestibility of the substrate. When determining the relative digestibilities of different cellulose allomorphs it is essential to also consider the relative crystallinities of the celluloses being tested.
Applied and Environmental Microbiology, Jun 28, 2023
In many catalytic depolymerization processes of lignin polymers, aryl-ether bonds are selectively... more In many catalytic depolymerization processes of lignin polymers, aryl-ether bonds are selectively cleaved, leaving carbon-carbon bonds between aromatic units intact, including dimers and oligomers with β-1 linkages. Therefore, elucidating the catabolic system of β-1-type lignin-derived compounds will aid in the establishment of biological funneling of heterologous lignin-derived aromatic compounds to value-added products.
Green Chemistry, 2022
Catalytic fast pyrolysis of biomass is a promising technology to generate biofuel blendstocks. Th... more Catalytic fast pyrolysis of biomass is a promising technology to generate biofuel blendstocks. This process generates a carbon-rich wastewater, which represents a loss of carbon that could be converted to coproducts. Here, we explored the biological conversion of methyl phenols (cresols), a major component of biomass pyrolysis wastewater, into 2-methyl and 3-methyl muconic acids for use as polymer building blocks and plasticizers. We engineered Pseudomonas putida KT2440 to convert all three cresol isomers, o-, m-, and p-cresol, into their methyl muconic acid counterparts via the heterologous aromatic hydroxylase DmpKLMNOP from Pseudomonas putida CF600. We optimized conversion of cresols by expressing a heterologous (methyl)catechol dioxygenase ClcA from Rhodococcus opacus 1CP, followed by proof-of-concept fed-batch bioreactor cultivations. Methyl muconic acids and the hydrogenated methyl adipic acids were incorporated into nylons and plasticizers to evaluate potential performance advantages relative to existing materials. Methyl muconic acids in nylon-6,6 analogs substantially reduced melting and glass transition temperatures and enable post-polymerization modifications, and incorporating methyl adipic acid into nylon-6,6 analogs leads to a slightly reduced glass transition temperature and a 12% reduction in water permeability relative to nylon-6,6. When methyl diacids were incorporated into plasticizers for poly(vinyl chloride), they exhibit lower glass transition temperatures at the same mass loadings as phthalic acid and adipic acid-based plasticizers. The methyl diacids were also predicted to exhibit reduced health and environmental risks compared to phthalic acid. Overall, this study encompasses the selection of a target product from an exemplary waste stream to the demonstration of multiple industrially relevant performance advantages relative to petroleum-derived analogs and highlights the potential for biological waste stream valorization.
Lignin is an underutilized value stream in current biomass conversion technologies because there ... more Lignin is an underutilized value stream in current biomass conversion technologies because there exist no economic and technically feasible routes for lignin depolymerization and upgrading. Base-catalyzed deconstruction (BCD) has been applied for lignin depolymerization (e.g., the Kraft process) in the pulp and paper industry for more than a century using aqueous-phase media. However, these efforts require treatment to neutralize the resulting streams, which adds significantly to the cost of lignin deconstruction. To circumvent the need for downstream treatment, here we report recent advances in the synthesis of layered double hydroxide and metal oxide catalysts to be applied to the BCD of lignin. These catalysts may prove more cost-effective than liquid-phase, non-recyclable base, and their use obviates downstream processing steps such as neutralization. Synthetic procedures for various transition-metal containing catalysts, detailed kinetics measurements using lignin model compounds, and results of the application of these catalysts to biomass-derived lignin will be presented.
ABSTRACT Research conducted at NREL has shown that lignin undergoes a phase transition during the... more ABSTRACT Research conducted at NREL has shown that lignin undergoes a phase transition during thermochemical pretreatments conducted above its glass transition temperature. The lignin coalesces within the plant cell wall and appears as microscopic droplets on cell surfaces. It is clear that pretreatment causes significant changes in lignin distribution in pretreatments at all scales from small laboratory reactors to pilot scale reactors. A method for selectively extracting lignin droplets from the surfaces of pretreated cell walls has allowed us to characterize the chemical nature and molecular weight distribution of this fraction. The effect of lignin redistribution on the digestibility of pretreated solids has also been tested. It is clear that removal of the droplets increases the digestibility of pretreated corn stover. The improved digestibility could be due to decreased non-specific binding of enzymes to lignin in the droplets, or because the droplets no longer block access to cellulose.
Journal of Wood Science, Feb 1, 2002
Residual wood meal after extraction of milled wood lignin (WMEM) of Eucalyptus globulus was extra... more Residual wood meal after extraction of milled wood lignin (WMEM) of Eucalyptus globulus was extracted with alkali and LiC1/N,N-Dimethylacetamide (DMAc). These agents dissolve mainly hemicellulose and cellulose, respectively. The extractability of WMEM in alkali solutions was influenced by the degree of swelling of the cellulose. Under good swelling conditions, considerable amounts of cellulose and lignin were extracted with the hemicellulose. Maximum extractability was about 60% of the WMEM under optimum conditions (3 M or 5 M LiOH or 3M NaOH solution). Some portion of cellulose was extracted with LiC1/DMAc at room temperature. Thus, lignin in E. globulus WMEM was divided into three fractions: hemicellulose-lignin fraction, cellulose-lignin fraction, and insoluble-lignin fraction.
Microbial biotechnology, May 1, 2020
In the originally published article by Salvach ua et al. (2019), the author Anna Furches was acci... more In the originally published article by Salvach ua et al. (2019), the author Anna Furches was accidentally omitted from the byline of manuscript, 'Metabolic engineering of Pseudomonas putida for increased polyhydroxyalkanoate production from lignin' Volume 13, Issue 1, Pages 290-298. The byline should read as given above.
Journal of Agricultural and Food Chemistry, Mar 19, 2013
The lignin content measured after dilute sulfuric acid pretreatment of corn stover indicates more... more The lignin content measured after dilute sulfuric acid pretreatment of corn stover indicates more lignin than could be accounted for on the basis of the untreated corn stover lignin content. This phenomenon was investigated using a combination of 13 C cross-polarization/magic-angle spinning (CP/MAS) solid-state nuclear magnetic resonance (NMR) spectroscopy and lignin removal using acid chlorite bleaching. Only minimal contamination with carbohydrates and proteins was observed in the pretreated corn stover. Incorporating degradation products from sugars was also investigated using 13 C-labeled sugars. The results indicate that sugar degradation products are present in the pretreatment residue and may be intimately associated with the lignin. Studies comparing whole corn stover (CS) to extractives-free corn stover [CS(Ext)] clearly demonstrated that extractives are a key contributor to the high-lignin mass balance closure (MBC). Sugars and other low molecular weight compounds present in plant extractives polymerize and form solids during pretreatment, resulting in apparent Klason lignin measurements that are biased high.
ACS Sustainable Chemistry & Engineering, May 20, 2014
Enzymatic depolymerization of polysaccharides is often a key step in the production of fuels and ... more Enzymatic depolymerization of polysaccharides is often a key step in the production of fuels and chemicals from lignocellulosic biomass. Historically, model cellulose substrates have been utilized to reveal insights into enzymatic saccharification mechanisms. However, translating findings from model substrates to realistic biomass substrates is critical for evaluating enzyme performance. Here, we employ a commercial fungal enzyme cocktail, purified cellulosomes, and combinations of these two enzymatic systems to investigate saccharification mechanisms on corn stover deconstructed either via clean fractionation (CF) or deacetylated dilute sulfuric acid pretreatments. CF is an organosolv pretreatment method utilizing water, MIBK, and either acetone or ethanol with catalytic amounts of sulfuric acid to fractionate biomass components. The insoluble cellulose-enriched fraction (CEF) from CF contains mainly cellulose, with minor amounts of residual hemicellulose and lignin. Enzymatic digestions at both low and high solid loadings demonstrate that CF reduces the amount of enzyme required to depolymerize polysaccharides relative to deacetylated dilute acid-pretreated corn stover. Transmission and scanning electron microscopy of the digested biomass provides evidence for the different mechanisms of enzymatic deconstruction between free and cellulosomal enzyme systems and reveals the basis for the synergistic relationship between the two enzyme paradigms on a process-relevant substrate. These results also demonstrate that the presence of lignin is more detrimental to cellulosome action than to free fungal cellulases. As enzyme costs are a major driver for biorefineries, this study provides key inputs for evaluation of CF as a pretreatment method and synergistic mixed enzyme systems as a saccharification strategy for biomass conversion.
Proceedings of the National Academy of Sciences
Lignin valorization is being intensely pursued via tandem catalytic depolymerization and biologic... more Lignin valorization is being intensely pursued via tandem catalytic depolymerization and biological funneling to produce single products. In many lignin depolymerization processes, aromatic dimers and oligomers linked by carbon–carbon bonds remain intact, necessitating the development of enzymes capable of cleaving these compounds to monomers. Recently, the catabolism of erythro -1,2-diguaiacylpropane-1,3-diol ( erythro -DGPD), a ring-opened lignin-derived β-1 dimer, was reported in Novosphingobium aromaticivorans . The first enzyme in this pathway, LdpA (formerly LsdE), is a member of the nuclear transport factor 2 (NTF-2)-like structural superfamily that converts erythro -DGPD to lignostilbene through a heretofore unknown mechanism. In this study, we performed biochemical, structural, and mechanistic characterization of the N. aromaticivorans LdpA and another homolog identified in Sphingobium sp. SYK-6, for which activity was confirmed in vivo. For both enzymes, we first demonstra...
Sphingobiumsp. SYK-6 is an efficient aromatic catabolic bacterium that can consume all four stere... more Sphingobiumsp. SYK-6 is an efficient aromatic catabolic bacterium that can consume all four stereoisomers of 1,2-diguaiacylpropane-1,3-diol (DGPD), which is a ring-opened β-1-type dimer. Recently, LdpA-mediated catabolism oferythro-DGPD was reported in SYK-6, but the catabolic pathway forthreo-DGPD was heretofore unknown. Here we elucidated the catabolism ofthreo-DGPD, which proceeds through conversion toerythro-DGPD. Whenthreo-DGPD was incubated with SYK-6, the Cα alcohol groups ofthreo-DGPD (DGPD I and II) were initially oxidized to produce the Cα carbonyl form (DGPD-keto I and II). This initial oxidation step is catalyzed by Cα-dehydrogenases, which belong to the short-chain dehydrogenase/reductase (SDR) family and are involved in the catabolism of β-O-4-type dimers. Analysis of seven candidate genes revealed that NAD+-dependent LigD and LigL are mainly involved in the conversion of DGPD I and II, respectively. Next, we found that DGPD-keto I and II were reduced toerythro-DGPD (D...
Green Chemistry, 2022
Lignin is an abundant renewable resource that is a promising substrate for upgrading to fuels and... more Lignin is an abundant renewable resource that is a promising substrate for upgrading to fuels and chemicals.
Lignin is an abundant and recalcitrant component of plant cell walls. While lignin degradation in... more Lignin is an abundant and recalcitrant component of plant cell walls. While lignin degradation in nature is typically attributed to fungi, growing evidence suggests that bacteria also catabolize this complex biopolymer. However, the spatiotemporal mechanisms for lignin catabolism remain unclear. Improved understanding of this biological process would aid in our collective knowledge of both carbon cycling and microbial strategies to valorize lignin to value-added compounds. Here, we examine lignin modifications and the exoproteome of three aromatic catabolic bacteria: Pseudomonas putida KT2440, Rhodoccocus jostii RHA1, and Amycolatopsis sp. ATCC 39116. P. putida cultivation on lignin-rich media is characterized by an abundant exoproteome that is dynamically and selectively packaged into outer membrane vesicles (OMVs). Interestingly, many enzymes known to exhibit activity towards lignin-derived aromatic compounds are enriched in OMVs from early to late stationary phase, corresponding ...
Metabolic Engineering, 2021
OSTI OAI (U.S. Department of Energy Office of Scientific and Technical Information), May 1, 2018
Chemsuschem, Nov 16, 2022
Lignin is the largest source of bio‐based aromatic compounds in nature, and its valorization is e... more Lignin is the largest source of bio‐based aromatic compounds in nature, and its valorization is essential to the sustainability of lignocellulosic biorefining. Characterizing lignin‐derived compounds remains challenging due to the heterogeneity of this biopolymer. Tandem mass spectrometry is a promising tool for lignin structural analytics, as fragmentation patterns of model compounds can be extrapolated to identify characteristic moieties in complex samples. This work extended previous resonance excitation‐type collision‐induced dissociation (CID) methods that identified lignin oligomers containing β–O–4, β–5, and β–β bonds, to also identify characteristics of 5–5, β–1, and 4–O–5 dimers, enabled by quadrupole time‐of‐flight (QTOF) CID with energy‐resolved mass spectrometry (ERMS). Overall, QTOF‐ERMS offers in‐depth structural information and could ultimately contribute to tools for high‐throughput lignin dimer identification.
Journal of Wood Chemistry and Technology, Jan 4, 2003
ABSTRACT A novel degradation method of arylglycerol-β-aryl (β-O-4) ether substructure in lignins ... more ABSTRACT A novel degradation method of arylglycerol-β-aryl (β-O-4) ether substructure in lignins without any secondary condensation reactions named TIZ method was developed. The TIZ method consists of three reaction steps: (1) selective tosylation (T) of primary hydroxyl group, (2) iodination (I), and (3) zinc-metal treatment (Z) to cleave of β-O-4 linkage. This method was applied to two nonphenolic β-O-4 lignin substructure compounds of guaiacyl and syringyl type: benzylguaiacylglycerol-β-guaiacyl ether (1a) and benzylsyringylglycerol-β-syringyl ether (1b). Respective reaction steps were performed quantitatively concerning both guaiacyl and syringyl models. After zinc treatment, almost 1 mole of olefinic compounds, 3-benzylguaiacyl-3-hydroxy-1-propene (4a) and 3-benzylsyringyl-3-hydroxy-1-propene (4b), were formed from 1 mol of compound 1a and 1b, respectively. This indicates that β-O-4 linkage in compound 1a and 1b was cleaved quantitatively without formation of any by-products. The structures of the degradation products from compound 1a and 1b by TIZ method were determined by H and C NMR spectroscopy and GC–MS, and by comparison with authentic compounds synthesized independently. The degradation products were quantified by GC. The TIZ method, carried out under weak alkaline and neutral reaction conditions, was shown to be useful for the selective cleavage of β-O-4 linkage of compound 1a and 1b without secondary condensation reaction on side chain. Therefore, this method is an efficient analytical method for lignin structure studies.
Metabolic Engineering, May 1, 2021
Biotechnology for Biofuels, Oct 19, 2011
Background: In converting biomass to bioethanol, pretreatment is a key step intended to render ce... more Background: In converting biomass to bioethanol, pretreatment is a key step intended to render cellulose more amenable and accessible to cellulase enzymes and thus increase glucose yields. In this study, four cellulose samples with different degrees of polymerization and crystallinity indexes were subjected to aqueous sodium hydroxide and anhydrous liquid ammonia treatments. The effects of the treatments on cellulose crystalline structure were studied, in addition to the effects on the digestibility of the celluloses by a cellulase complex. Results: From X-ray diffractograms and nuclear magnetic resonance spectra, it was revealed that treatment with liquid ammonia produced the cellulose III I allomorph; however, crystallinity depended on treatment conditions. Treatment at a low temperature (25°C) resulted in a less crystalline product, whereas treatment at elevated temperatures (130°C or 140°C) gave a more crystalline product. Treatment of cellulose I with aqueous sodium hydroxide (16.5 percent by weight) resulted in formation of cellulose II, but also produced a much less crystalline cellulose. The relative digestibilities of the different cellulose allomorphs were tested by exposing the treated and untreated cellulose samples to a commercial enzyme mixture (Genencor-Danisco; GC 220). The digestibility results showed that the starting cellulose I samples were the least digestible (except for corn stover cellulose, which had a high amorphous content). Treatment with sodium hydroxide produced the most digestible cellulose, followed by treatment with liquid ammonia at a low temperature. Factor analysis indicated that initial rates of digestion (up to 24 hours) were most strongly correlated with amorphous content. Correlation of allomorph type with digestibility was weak, but was strongest with cellulose conversion at later times. The cellulose III I samples produced at higher temperatures had comparable crystallinities to the initial cellulose I samples, but achieved higher levels of cellulose conversion, at longer digestion times. Conclusions: Earlier studies have focused on determining which cellulose allomorph is the most digestible. In this study we have found that the chemical treatments to produce different allomorphs also changed the crystallinity of the cellulose, and this had a significant effect on the digestibility of the substrate. When determining the relative digestibilities of different cellulose allomorphs it is essential to also consider the relative crystallinities of the celluloses being tested.
Applied and Environmental Microbiology, Jun 28, 2023
In many catalytic depolymerization processes of lignin polymers, aryl-ether bonds are selectively... more In many catalytic depolymerization processes of lignin polymers, aryl-ether bonds are selectively cleaved, leaving carbon-carbon bonds between aromatic units intact, including dimers and oligomers with β-1 linkages. Therefore, elucidating the catabolic system of β-1-type lignin-derived compounds will aid in the establishment of biological funneling of heterologous lignin-derived aromatic compounds to value-added products.
Green Chemistry, 2022
Catalytic fast pyrolysis of biomass is a promising technology to generate biofuel blendstocks. Th... more Catalytic fast pyrolysis of biomass is a promising technology to generate biofuel blendstocks. This process generates a carbon-rich wastewater, which represents a loss of carbon that could be converted to coproducts. Here, we explored the biological conversion of methyl phenols (cresols), a major component of biomass pyrolysis wastewater, into 2-methyl and 3-methyl muconic acids for use as polymer building blocks and plasticizers. We engineered Pseudomonas putida KT2440 to convert all three cresol isomers, o-, m-, and p-cresol, into their methyl muconic acid counterparts via the heterologous aromatic hydroxylase DmpKLMNOP from Pseudomonas putida CF600. We optimized conversion of cresols by expressing a heterologous (methyl)catechol dioxygenase ClcA from Rhodococcus opacus 1CP, followed by proof-of-concept fed-batch bioreactor cultivations. Methyl muconic acids and the hydrogenated methyl adipic acids were incorporated into nylons and plasticizers to evaluate potential performance advantages relative to existing materials. Methyl muconic acids in nylon-6,6 analogs substantially reduced melting and glass transition temperatures and enable post-polymerization modifications, and incorporating methyl adipic acid into nylon-6,6 analogs leads to a slightly reduced glass transition temperature and a 12% reduction in water permeability relative to nylon-6,6. When methyl diacids were incorporated into plasticizers for poly(vinyl chloride), they exhibit lower glass transition temperatures at the same mass loadings as phthalic acid and adipic acid-based plasticizers. The methyl diacids were also predicted to exhibit reduced health and environmental risks compared to phthalic acid. Overall, this study encompasses the selection of a target product from an exemplary waste stream to the demonstration of multiple industrially relevant performance advantages relative to petroleum-derived analogs and highlights the potential for biological waste stream valorization.
Lignin is an underutilized value stream in current biomass conversion technologies because there ... more Lignin is an underutilized value stream in current biomass conversion technologies because there exist no economic and technically feasible routes for lignin depolymerization and upgrading. Base-catalyzed deconstruction (BCD) has been applied for lignin depolymerization (e.g., the Kraft process) in the pulp and paper industry for more than a century using aqueous-phase media. However, these efforts require treatment to neutralize the resulting streams, which adds significantly to the cost of lignin deconstruction. To circumvent the need for downstream treatment, here we report recent advances in the synthesis of layered double hydroxide and metal oxide catalysts to be applied to the BCD of lignin. These catalysts may prove more cost-effective than liquid-phase, non-recyclable base, and their use obviates downstream processing steps such as neutralization. Synthetic procedures for various transition-metal containing catalysts, detailed kinetics measurements using lignin model compounds, and results of the application of these catalysts to biomass-derived lignin will be presented.
ABSTRACT Research conducted at NREL has shown that lignin undergoes a phase transition during the... more ABSTRACT Research conducted at NREL has shown that lignin undergoes a phase transition during thermochemical pretreatments conducted above its glass transition temperature. The lignin coalesces within the plant cell wall and appears as microscopic droplets on cell surfaces. It is clear that pretreatment causes significant changes in lignin distribution in pretreatments at all scales from small laboratory reactors to pilot scale reactors. A method for selectively extracting lignin droplets from the surfaces of pretreated cell walls has allowed us to characterize the chemical nature and molecular weight distribution of this fraction. The effect of lignin redistribution on the digestibility of pretreated solids has also been tested. It is clear that removal of the droplets increases the digestibility of pretreated corn stover. The improved digestibility could be due to decreased non-specific binding of enzymes to lignin in the droplets, or because the droplets no longer block access to cellulose.
Journal of Wood Science, Feb 1, 2002
Residual wood meal after extraction of milled wood lignin (WMEM) of Eucalyptus globulus was extra... more Residual wood meal after extraction of milled wood lignin (WMEM) of Eucalyptus globulus was extracted with alkali and LiC1/N,N-Dimethylacetamide (DMAc). These agents dissolve mainly hemicellulose and cellulose, respectively. The extractability of WMEM in alkali solutions was influenced by the degree of swelling of the cellulose. Under good swelling conditions, considerable amounts of cellulose and lignin were extracted with the hemicellulose. Maximum extractability was about 60% of the WMEM under optimum conditions (3 M or 5 M LiOH or 3M NaOH solution). Some portion of cellulose was extracted with LiC1/DMAc at room temperature. Thus, lignin in E. globulus WMEM was divided into three fractions: hemicellulose-lignin fraction, cellulose-lignin fraction, and insoluble-lignin fraction.
Microbial biotechnology, May 1, 2020
In the originally published article by Salvach ua et al. (2019), the author Anna Furches was acci... more In the originally published article by Salvach ua et al. (2019), the author Anna Furches was accidentally omitted from the byline of manuscript, 'Metabolic engineering of Pseudomonas putida for increased polyhydroxyalkanoate production from lignin' Volume 13, Issue 1, Pages 290-298. The byline should read as given above.
Journal of Agricultural and Food Chemistry, Mar 19, 2013
The lignin content measured after dilute sulfuric acid pretreatment of corn stover indicates more... more The lignin content measured after dilute sulfuric acid pretreatment of corn stover indicates more lignin than could be accounted for on the basis of the untreated corn stover lignin content. This phenomenon was investigated using a combination of 13 C cross-polarization/magic-angle spinning (CP/MAS) solid-state nuclear magnetic resonance (NMR) spectroscopy and lignin removal using acid chlorite bleaching. Only minimal contamination with carbohydrates and proteins was observed in the pretreated corn stover. Incorporating degradation products from sugars was also investigated using 13 C-labeled sugars. The results indicate that sugar degradation products are present in the pretreatment residue and may be intimately associated with the lignin. Studies comparing whole corn stover (CS) to extractives-free corn stover [CS(Ext)] clearly demonstrated that extractives are a key contributor to the high-lignin mass balance closure (MBC). Sugars and other low molecular weight compounds present in plant extractives polymerize and form solids during pretreatment, resulting in apparent Klason lignin measurements that are biased high.
ACS Sustainable Chemistry & Engineering, May 20, 2014
Enzymatic depolymerization of polysaccharides is often a key step in the production of fuels and ... more Enzymatic depolymerization of polysaccharides is often a key step in the production of fuels and chemicals from lignocellulosic biomass. Historically, model cellulose substrates have been utilized to reveal insights into enzymatic saccharification mechanisms. However, translating findings from model substrates to realistic biomass substrates is critical for evaluating enzyme performance. Here, we employ a commercial fungal enzyme cocktail, purified cellulosomes, and combinations of these two enzymatic systems to investigate saccharification mechanisms on corn stover deconstructed either via clean fractionation (CF) or deacetylated dilute sulfuric acid pretreatments. CF is an organosolv pretreatment method utilizing water, MIBK, and either acetone or ethanol with catalytic amounts of sulfuric acid to fractionate biomass components. The insoluble cellulose-enriched fraction (CEF) from CF contains mainly cellulose, with minor amounts of residual hemicellulose and lignin. Enzymatic digestions at both low and high solid loadings demonstrate that CF reduces the amount of enzyme required to depolymerize polysaccharides relative to deacetylated dilute acid-pretreated corn stover. Transmission and scanning electron microscopy of the digested biomass provides evidence for the different mechanisms of enzymatic deconstruction between free and cellulosomal enzyme systems and reveals the basis for the synergistic relationship between the two enzyme paradigms on a process-relevant substrate. These results also demonstrate that the presence of lignin is more detrimental to cellulosome action than to free fungal cellulases. As enzyme costs are a major driver for biorefineries, this study provides key inputs for evaluation of CF as a pretreatment method and synergistic mixed enzyme systems as a saccharification strategy for biomass conversion.
Proceedings of the National Academy of Sciences
Lignin valorization is being intensely pursued via tandem catalytic depolymerization and biologic... more Lignin valorization is being intensely pursued via tandem catalytic depolymerization and biological funneling to produce single products. In many lignin depolymerization processes, aromatic dimers and oligomers linked by carbon–carbon bonds remain intact, necessitating the development of enzymes capable of cleaving these compounds to monomers. Recently, the catabolism of erythro -1,2-diguaiacylpropane-1,3-diol ( erythro -DGPD), a ring-opened lignin-derived β-1 dimer, was reported in Novosphingobium aromaticivorans . The first enzyme in this pathway, LdpA (formerly LsdE), is a member of the nuclear transport factor 2 (NTF-2)-like structural superfamily that converts erythro -DGPD to lignostilbene through a heretofore unknown mechanism. In this study, we performed biochemical, structural, and mechanistic characterization of the N. aromaticivorans LdpA and another homolog identified in Sphingobium sp. SYK-6, for which activity was confirmed in vivo. For both enzymes, we first demonstra...
Sphingobiumsp. SYK-6 is an efficient aromatic catabolic bacterium that can consume all four stere... more Sphingobiumsp. SYK-6 is an efficient aromatic catabolic bacterium that can consume all four stereoisomers of 1,2-diguaiacylpropane-1,3-diol (DGPD), which is a ring-opened β-1-type dimer. Recently, LdpA-mediated catabolism oferythro-DGPD was reported in SYK-6, but the catabolic pathway forthreo-DGPD was heretofore unknown. Here we elucidated the catabolism ofthreo-DGPD, which proceeds through conversion toerythro-DGPD. Whenthreo-DGPD was incubated with SYK-6, the Cα alcohol groups ofthreo-DGPD (DGPD I and II) were initially oxidized to produce the Cα carbonyl form (DGPD-keto I and II). This initial oxidation step is catalyzed by Cα-dehydrogenases, which belong to the short-chain dehydrogenase/reductase (SDR) family and are involved in the catabolism of β-O-4-type dimers. Analysis of seven candidate genes revealed that NAD+-dependent LigD and LigL are mainly involved in the conversion of DGPD I and II, respectively. Next, we found that DGPD-keto I and II were reduced toerythro-DGPD (D...
Green Chemistry, 2022
Lignin is an abundant renewable resource that is a promising substrate for upgrading to fuels and... more Lignin is an abundant renewable resource that is a promising substrate for upgrading to fuels and chemicals.
Lignin is an abundant and recalcitrant component of plant cell walls. While lignin degradation in... more Lignin is an abundant and recalcitrant component of plant cell walls. While lignin degradation in nature is typically attributed to fungi, growing evidence suggests that bacteria also catabolize this complex biopolymer. However, the spatiotemporal mechanisms for lignin catabolism remain unclear. Improved understanding of this biological process would aid in our collective knowledge of both carbon cycling and microbial strategies to valorize lignin to value-added compounds. Here, we examine lignin modifications and the exoproteome of three aromatic catabolic bacteria: Pseudomonas putida KT2440, Rhodoccocus jostii RHA1, and Amycolatopsis sp. ATCC 39116. P. putida cultivation on lignin-rich media is characterized by an abundant exoproteome that is dynamically and selectively packaged into outer membrane vesicles (OMVs). Interestingly, many enzymes known to exhibit activity towards lignin-derived aromatic compounds are enriched in OMVs from early to late stationary phase, corresponding ...
Metabolic Engineering, 2021