Exhaustive laccase-catalysed oxidation of a lignin model compound (vanillyl glycol) produces methanol and polymeric quinoid products.
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ABSTRACT:
SUBMITTER: Lundquist K
PROVIDER: S-EPMC1145179 | biostudies-other | 1985 Jul
REPOSITORIES: biostudies-other
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Project description:The selective lignin conversion into bio-based organic mono-aromatics is a major general challenge due to complex structure itself/additional macromolecule modifications, caused by the cleavage of the ether chemical bonds during the lignocellulosic biomass organosolv pulping in acidified aqueous ethanol. Herein, the acido-lysis of connected benzyl phenyl (BPE), being a representative model compound with ?-O-4 linkage, was investigated in methanol, EtOH and 75 vol% EtOH/water mixture solutions, progressing each time with protonating sulfuric acid. The effect of the physical solvent properties, acidity of the reaction process media and temperature on rate was determined. Experiments suggested BPE following SN1 mechanism due to the formation of a stable primary carbocation/polarity. The product species distribution in non-aqueous functional alcohols was strongly affected. The addition of H2O was advantageous, especially for alkoxylation. Yield was reduced by a factor of 3, consequently preserving reactive hydroxyl group. Quantitative experimental results indicated key performance parameters to achieve optimum. Organosolv lignins were further isolated under significantly moderate conditions. Consecutive structural differences observed supported findings, obtained when using BPE. H2O presence was again found to grant a higher measured -OH content. Mechanistic pathway analysis thus represents the first step when continuing to kinetics, structure-activity relationships or bio-refining industrial resources.
| S-EPMC7338503 | biostudies-literature
Project description:We have developed a multienzyme functionalized membrane reactor for bioconversion of lignin model compound involving enzymatic catalysis. Layer-by-layer approach was used to immobilize three different enzymes (glucose oxidase, peroxidase and laccase) into pH-responsive membranes. This novel membrane reactor couples the in situ generation of hydrogen peroxide (by glucose oxidase) to oxidative conversion of a lignin model compound, guaiacylglycerol-B-guaiacylether (GGE). Preliminary investigation of the efficacy of these functional membranes towards GGE degradation is demonstrated under convective flow mode. Over 90% of the initial feed could be degraded with the multienzyme immobilized membranes at a residence time of approximately 22 seconds. GGE conversion product analysis revealed formation of oligomeric oxidation products with peroxidase, which might be potential hazard to membrane bioreactors. These oxidation products could be further degraded by laccase enzymes in the multienzymatic membranes explaining the potential of multienzyme membrane reactors. The multienzyme incorporated membrane reactors were active for about a month time of storage at 4 °C, and retention of activity was demonstrated after repetitive use.
| S-EPMC6358281 | biostudies-literature
Project description:Raising the awareness of carbon dioxide emissions, climate global warming and fossil fuel depletion has renewed the transition towards a circular economy approach, starting by addressing active bio-economic precepts that all portion amounts of wood are valorised as products. This is accomplished by minimizing residues formed (preferably no waste materials), maximizing reaction productivity yields, and optimising catalysed chemical by-products. Within framework structure determination, the present work aims at drawing a parallel between the characterisation of cellulose-lignin mixture (derived system model) liquefaction and real conversion process in the acidified ethylene glycol at moderate process conditions, i.e., 150 °C, ambient atmospheric pressure and potential bio-based solvent, for 4 h. Extended-processing liquid phase is characterized considering catalyst-transformed reactant species being produced, mainly recovered lignin-based polymer, by quantitative 31P, 13C and 1H nuclear magnetic resonance (NMR) spectroscopy, as well as the size exclusion- (SEC) or high performance liquid chromatography (HPLC) separation for higher or lower molecular weight compound compositions, respectively. Such mechanistic pathway analytics help to understand the steps in mild organosolv biopolymer fractionation, which is one of the key industrial barriers preventing a more widespread manufacturing of the biomass-derived (hydroxyl, carbonyl or carboxyl) aromatic monomers or oligomers for polycarbonates, polyesters, polyamides, polyurethanes and (epoxy) resins.
| S-EPMC8234995 | biostudies-literature
Project description:Laccase efficiently catalyses polymerization of phenolic compounds. However, knowledge on applications of polymers synthesized in this manner remains scarce. Here, the potential of laccase-catalysed polymerization of natural phenols to form products useful in hair dyeing was investigated. All 15 tested phenols yielded coloured products after laccase treatment and colour diversity was attained by using mixtures of two phenolic monomers. After exploring colour differentiation pattern of 120 different reactions with statistical regression analysis, three monomer combinations, namely gallic acid and syringic acid, catechin and catechol, and ferulic acid and syringic acid, giving rise to brown, black, and red materials, respectively, were further characterized because such colours are commercially important for grey hair dyeing. Selected polymers could strongly absorb visible light and their hydrodynamic sizes ranged from 100 to 400 nm. Analyses of enzyme kinetic constants, liquid chromatography and electrospray ionization-mass spectrometry (ESI-MS) coupled with collision-induced dissociation MS/MS indicate that both monomers in reactions involving catechin and catechol, and ferulic acid and syringic acid, are coloured by heteropolymer synthesis, but the gallic acid/syringic acid combination is based on homopolymer mixture formation. Comparison of colour parameters from these three reactions with those of corresponding artificial homopolymer mixtures also supported the idea that laccase may catalyse either hetero- or homo-polymer synthesis. We finally used selected materials to dye grey hair. Each material coloured hair appropriately and the dyeing showed excellent resistance to conventional shampooing. Our study indicates that laccase-catalysed polymerization of natural phenols is applicable to the development of new cosmetic pigments.
| S-EPMC3815374 | biostudies-literature
Project description:Laccase-mediator systems (LMS) have been widely studied for their capacity to oxidize the nonphenolic subunits of lignin (70-90% of the polymer). The phenolic subunits (10-30% of the polymer), which can also be oxidized without mediators, have received considerably less attention. Consequently, it remains unclear to what extent the presence of a mediator influences the reactions of the phenolic subunits of lignin. To get more insight in this, UHPLC-MS was used to study the reactions of a phenolic lignin dimer (GBG), initiated by a laccase from Trametes versicolor, alone or in combination with the mediators HBT and ABTS. The role of HBT was negligible, as its oxidation by laccase occurred slowly in comparison to that of GBG. Laccase and laccase/HBT oxidized GBG at a comparable rate, resulting in extensive polymerization of GBG. In contrast, laccase/ABTS converted GBG at a higher rate, as GBG was oxidized both directly by laccase but also by ABTS radical cations, which were rapidly formed by laccase. The laccase/ABTS system resulted in Cα oxidation of GBG and coupling of ABTS to GBG, rather than polymerization of GBG. Based on these results, we propose reaction pathways of phenolic lignin model compounds with laccase/HBT and laccase/ABTS.
| S-EPMC5805406 | biostudies-literature
Project description:The Trametes versicolor laccase (TvL)-catalysed oligomerisation of the aniline dimer p-aminodiphenylamine (PADPA) was investigated in an aqueous medium of pH = 3.5, containing 80-100 nm-sized anionic vesicles formed from AOT, the sodium salt of bis(2-ethylhexyl)sulfosuccinic acid. If run under optimal conditions, the reaction yields oligomeric products which resemble the emeraldine salt form of polyaniline (PANI-ES) in its polaron state, known to be the only oxidation state of linear PANI which is electrically conductive. The vesicles serve as "templates" for obtaining products with the desired PANI-ES-like features. For this complex, heterogeneous, vesicle-assisted, and enzyme-mediated reaction, in which dissolved dioxygen also takes part as a re-oxidant for TvL, small changes in the composition of the reaction mixture can have significant effects. Initial conditions may not only affect the kinetics of the reaction, but also the outcome, i.e., the product distribution once the reaction reaches its equilibrium state. While a change in the reaction temperature from T ≈ 25 to 5 °C mainly influenced the rate of reaction, increase in enzyme concentration and the presence of millimolar concentrations of chloride ions were found to have significant undesired effects on the outcome of the reaction. Chloride ions, which may originate from the preparation of the pH = 3.5 solution, inhibit TvL, such that higher TvL concentrations are required than without chloride to yield the same product distribution for the same reaction runtime as in the absence of chloride. With TvL concentrations much higher than the elaborated value, the products obtained clearly were different and over-oxidised. Thus, a change in the activity of the enzyme was found to have influence not only on kinetics but also led to a change in the final product distribution, molecular structure and electrical properties, which was a surprising find. The complementary analytical methods which we used in this work were in situ UV/vis/NIR, EPR, and Raman spectroscopy measurements, in combination with a detailed ex situ HPLC analysis and molecular dynamics simulations. With the results obtained, we would like to recall the often neglected or ignored fact that it is important to describe and pay attention to the experimental details, since this matters for being able to perform experiments in a reproducible way.
| S-EPMC9086443 | biostudies-literature
Project description:Laccase-mediator-oxidized lignin offers replacement for conventional chemical binders to produce fiberboards. Compared to the previously reported laccase-mediator system (LMS), a lignin-laccase-mediator-system (LLMS) has an advantage in that it requires much shorter fiber-enzyme incubation time due to significantly increased redox reactions. However, the cost of regularly applying laccase on an industrial scale is currently too high. We have employed CcLcc5 from cultures of the basidiomycete Coprinopsis cinerea as a novel basi-laccase (a CAZy subfamily AA1_1 laccase) in medium-density fiberboard (MDF) production, in comparison to the commercial formulation Novozym 51003 with recombinantly produced asco-laccase MtL (a CAZy subfamily AA1_3 laccase-like multicopper oxidase from the ascomycete Myceliophthora thermophila). With the best-performing natural mediator 2,6-dimethoxyphenol (DMP), unpurified CcLcc5 was almost as good as formulated Novozym 51003 in increasing the molecular weight (MW) of the technical lignins tested, the hydrophilic high-MW Ca-lignosulfonate and the hydrophobic low-MW kraft lignin (Indulin AT). Oxygen consumption rates of the two distantly related, poorly conserved enzymes (31% sequence identity) with different mediators and lignosulfonate were also comparable, but Indulin AT significantly reduced the oxidative activity of Novozym 51003 unlike CcLcc5, regardless of the mediator used, either DMP or guaiacol. Oxygen uptake by both laccases was much faster with both technical lignins with DMP than with guaiacol. In case of lignosulfonate and DMP, 20-30 min of incubation was sufficient for full oxygen consumption, which fits in well in time with the usual binder application steps in industrial MDF production processes. LLMS-bonded MDF was thus produced on a pilot-plant scale with either crude CcLcc5 or Novozym 51003 at reduced enzyme levels of 5 kU/kg absolutely dry wood fiber with lignosulfonate and mediator DMP. Boards produced with CcLcc5 were comparably good as those made with Novozym 51003. Boards reached nearly standard specifications in internal bond strength (IB) and modulus of rupture (MOR), while thickness swelling (TS) was less good based on the hydrophilic character of lignosulfonate. LLMS-bonded MDF with Indulin AT and DMP performed better in TS but showed reduced IB and MOR values.
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Project description:Lignin is a complex natural organic polymer and is one of the primary components of lignocellulose. The efficient utilization of lignocellulose is limited because it is difficult to degrade lignin. In this study, we screened a lacz1 gene fragment encoding laccase from the macrotranscriptome data of a microbial consortium WSC-6, which can efficiently degrade lignocellulose. The reverse transcription-quantitative PCR (RT-qPCR) results demonstrated that the expression level of the lacz1 gene during the peak period of lignocellulose degradation by WSC-6 increased by 30.63 times compared to the initial degradation period. Phylogenetic tree analysis demonstrated that the complete lacz1 gene is derived from a Bacillus sp. and encoded laccase. The corresponding protein, LacZ1, was expressed and purified by Ni-chelating affinity chromatography. The optimum temperature was 75°C, the optimum pH was 4.5, and the highest enzyme activity reached 16.39 U/mg. We found that Cu2+ was an important cofactor needed for LacZ1 to have enzyme activity. The molecular weight distribution of lignin was determined by gel permeation chromatography (GPC), and changes in the lignin structure were determined by 1H nuclear magnetic resonance (1H NMR) spectra. The degradation products of lignin by LacZ1 were determined by gas chromatography and mass spectrometry (GC-MS), and three lignin degradation pathways (the gentian acid pathway, benzoic acid pathway, and protocatechuic acid pathway) were proposed. This study provides insight into the degradation of lignin and new insights into high-temperature bacterial laccase. IMPORTANCE Lignin is a natural aromatic polymer that is not easily degraded, hindering the efficient use of lignocellulose-rich biomass resources, such as straw. Biodegradation is a method of decomposing lignin that has recently received increasing attention. In this study, we screened a gene encoding laccase from the lignocellulose-degrading microbial consortium WSC-6, purified the corresponding protein LacZ1, characterized the enzymatic properties of laccase LacZ1, and speculated that the degradation pathway of LacZ1 degrades lignin. This study identified a new, high-temperature bacterial laccase that can degrade lignin, providing insight into lignin degradation by this laccase.
| S-EPMC8580002 | biostudies-literature
Project description:The fractional pyrolysis of lignin model compound para-coumaryl alcohol (p-CMA) containing a propanoid side chain and a phenolic OH group was studied using the System for Thermal Diagnostic Studies at temperatures from 200 to 900 °C, in order to gain mechanistic insight into the role of large substituents in high-lignin feedstocks pyrolysis. Phenol and its simple derivatives p-cresol, ethyl-, propenyl-, and propyl-phenols were found to be the major products predominantly formed at low pyrolysis temperatures (<500 °C). A cryogenic trapping technique was employed combined with EPR spectroscopy to identify the open-shell intermediates registered at pyrolysis temperatures above 500 °C. These were characterized as radical mixtures primarily consisting of oxygen-linked conjugated radicals. A comprehensive potential energy surface analysis of p-CMA and p-CMA + H atom systems was performed using various DFT protocols to examine the possible role of concerted molecular eliminations and free-radical mechanisms in the formation of major products. Other significant unimolecular concerted reactions along with formation and decomposition of primary radicals are also described and evaluated. The calculations suggest that a set of the chemically activated secondary radical channels is relevant to the low temperature product formation under fractional pyrolysis conditions.
| S-EPMC7769150 | biostudies-literature
Project description:Laccases were proposed to catalyze the oxidative polymerization of monolignols. We identified 49 laccase gene models in Populus trichocarpa, of which 29 were predicted to be targets of ptr-miR397a. We overexpressed Ptr-MIR397a in transgenic P. trichocarpa. In each of all 9 transgenic lines tested, 17 PtrLACs were down-regulated as analyzed by RNA-seq. Transgenic lines with severe reduction in the expression of these laccase genes resulted in an approximately 40% decrease in the total laccase activity. Overexpression of Ptr-MIR397a in these transgenic lines also reduced lignin content, whereas levels of all monolignol biosynthetic gene transcripts remained unchanged. A hierarchical genetic regulatory network (GRN) built by a bottom-up Graphic Gaussian Model algorithm provides additional support for a role of ptr-miR397a as a negative regulator of laccases for lignin biosynthesis. Full transcriptome based differential gene expression in the overexpressed transgenics and protein domain analyses implicate previously unidentified transcription factors and their targets in an extended hierarchical GRN including ptr-miR397a and laccases that coregulate lignin biosynthesis in wood formation. Ptr-miR397a, laccases and other regulatory components of this network may provide additional strategies for genetic manipulation of lignin content. Total twelve trees were used. Those include nine individual transgenic trees for overexpressing Ptr-miR397a, as nine biological replicates, and three wild-type trees.
2014-01-17 | E-GEOD-54153 | biostudies-arrayexpress