Artificial Lignin Research Articles

Biomimetic Dehydrogenation of Non-Conventional Lignin Monomers on Cellulose Ferulate Interfaces.

Cellulose ferulate, synthesized by Mitsunobu reaction, is shaped into thin films and also used as an aqueous dispersion to perform artificial lignin polymerization on anchor groups. This biomimetic approach is carried out in a Quartz crystal microbalance with a dissipation monitoring (QCM-D) device to enable online monitoring of the dehydrogenation, applying H2O2 and adsorbed horseradish peroxidase (HRP). The systematic use of phenylpropanoids with different oxidation states, i.e., ferulic acid, coniferyl aldehyde, coniferyl alcohol, and eugenol allowed to conclude structure-property relationships. Both the deposited material, as well as the surface roughness increased with the hydrophobicity of the monomers. Beyond surface characterizations, py-GC-MS, HSQC NMR spectroscopy and Size exclusion chromatography (SEC) measurements revealed the linkage types β-β, β-5, 5-5, and β-O-4, as well as the oligomeric character of the dehydrogenation products. All samples possessed an antibacterial activity against B. subtilis and can be used in the field of antimicrobialbiomaterials.

Synthesis and structural design of microspheres comprising cellulose nanofibers and artificial lignin polymer by enzyme-mediated Pickering emulsion templating

An enzyme-mediated Pickering emulsion templating strategy to fabricate spherical microparticles comprising cellulose nanofibers and artificial lignin expands the greater use of renewable, carbon dioxide-fixed wood in green cosmetics.

Two-way reaction of versatile peroxidase with artificial lignin enhances low-molecular weight fractions.

In recent years, versatile peroxidase (VP) has emerged as a promising enzyme for biotechnological applications, as it can oxidize lignin without the external mediators. To gain insights into the breakdown process of artificial lignin by VP, reaction between the two was studied. Degradation products were fractionated using ultrafiltration and analyzed by RP- high performance liquid chromatography with mass detection (HPLC-MS) chromatography. Four fractions were obtained based on their molecular sizes: >10, 3-10, 1-3, and <1kDa. Interestingly, while VP did not significantly alter the yields of these fractions, the chromatograms revealed the presence of oligomers with different molecular weights (MWs) resulting from the enzymatic activity. The VP exhibits a dual role in its enzymatic activity: both degrading and synthesizing these oligomers. This was confirmed by principal component analysis (PCA). The positive correlations were found between certain oligomers (D1 and D2, D5 and D6, as well as between D7, D10, T2, and T4), suggesting their simultaneous degradation. On the other hand, a negative correlation was found between the monomer and some oligomers (D7, D10, T2, and T4), indicating the decomposition of these oligomers into monomers. These findings shed light on the intricate interplay between VP and artificial lignin, offering valuable insights for potential applications in lignin valorization.

Growing of Artificial Lignin on Cellulose Ferulate Thin Films.

Thin films of cellulose ferulate were designed to study the formation of dehydrogenation polymers (DHPs) on anchor groups of the surface. Trimethylsilyl (TMS) cellulose ferulate with degree of substitution values of 0.35 (ferulate) and 2.53 (TMS) was synthesized by sophisticated polysaccharide chemistry applying the Mitsunobu reaction. The biopolymer derivative was spin-coated into thin films to yield ferulate moieties on a smooth cellulose surface. Dehydrogenative polymerization of coniferyl alcohol was performed in a Quartz crystal microbalance with a dissipation monitoring device in the presence of H2O2 and adsorbed horseradish peroxidase. The amount of DHP formed on the surface was found to be independent of the base layer thickness from 14 to 75 nm. Pyrolysis-GC-MS measurements of the DHP revealed β-O-4 and β-5 linkages. Mimicking lignification of plant cell walls on highly defined model films enables reproducible investigations of structure-property relationships.

Lignin as a multifunctional photocatalyst for solar-powered biocatalytic oxyfunctionalization of C–H bonds

Each year, the pulp and paper industry produces approximately 50 million metric tons of lignin as waste, 95% of which is combusted or abandoned. Here, we report the use of lignin as a photocatalyst that forms H2O2 by O2 reduction and H2O oxidation under visible light. We investigated the photophysical and electronic properties of two lignin models, lignosulfonate and kraft lignin, by spectroscopic and photoelectrochemical analyses, and demonstrated the photoredox chemistry of lignin using these and other lignin models (for example, native-like cellulolytic enzyme lignin, artificial lignin dehydrogenation polymer and phenolic β-aryl ether-type lignin dimer). Furthermore, the integration of lignin and H2O2-dependent unspecific peroxygenases (UPOs) enabled the highly enantioselective oxyfunctionalization of various C–H bonds. The use of lignin photocatalysts solves a number of the challenges relating to the sustainable activation of UPOs, notably, eliminating the need for artificial electron donors and suppressing the HO·-mediated inactivation of UPOs. Thus, the lignin–UPO hybrid catalyst achieved a total turnover number of UPO of 81,000 for solar-powered biocatalytic oxyfunctionalization in photochemical platforms.

Effect of pH on the formation of benzyl ester bonds between glucuronic acid and dehydrogenation polymer

The effect of pH on the addition reaction of glucuronic acid to quinone methides generated in the synthesis of dehydrogenation polymer (DHP or artificial lignin) was investigated. The DHP-glucuronic acid complexes were formed during DHP polymerization catalyzed by a mixture of laccase, β-glucosidase, and O2 within the pH range 7 to 4 in the presence of coniferin as a precursor. The structure of the product and the content of benzyl ester bonds were characterized by Fourier transform infrared spectroscopy, solid-state cross-polarization magic angle spinning carbon-13 nuclear magnetic resonance spectroscopy, ion chromatography, high performance liquid chromatography, and elemental analysis. The results showed that the pH of the reaction system had an important role in the formation of the benzyl ester bonds. Acidic conditions favored the reaction of quinone methide intermediates with carboxyl groups of glucuronic acid in the biosynthesis of DHP. However, weakly acidic conditions (pH 6) enhanced the reaction of quinone methide intermediates with glucose. In neutral conditions, the DHP-protein complex can be efficiently synthesized by the addition reaction of quinone methide intermediates with amino acids in protein.

Enzymatic Preparation and Characterization of Spherical Microparticles Composed of Artificial Lignin and TEMPO-Oxidized Cellulose Nanofiber.

A one-pot and one-step enzymatic synthesis of submicron-order spherical microparticles composed of dehydrogenative polymers (DHPs) of coniferyl alcohol as a typical lignin precursor and TEMPO-oxidized cellulose nanofibers (TOCNFs) was investigated. Horseradish peroxidase enzymatically catalyzed the radical coupling of coniferyl alcohol in an aqueous suspension of TOCNFs, resulting in the formation of spherical microparticles with a diameter and sphericity index of approximately 0.8 μm and 0.95, respectively. The ζ-potential of TOCNF-functionalized DHP microspheres was about −40 mV, indicating that the colloidal systems had good stability. Nanofibrous components were clearly observed on the microparticle surface by scanning electron microscopy, while some TOCNFs were confirmed to be inside the microparticles by confocal laser scanning microscopy with Calcofluor white staining. As both cellulose and lignin are natural polymers known to biodegrade, even in the sea, these woody TOCNF−DHP microparticle nanocomposites were expected to be promising alternatives to fossil resource-derived microbeads in cosmetic applications.

Alternative lignopolymer-based composites useful as enhanced functionalized support for enzymes immobilization

Amino-derivatized lignocomposite (ADL-composite) has been designed as an artificial lignin polymer (P1) of monolignol (coniferyl alcohol, CA) enhanced by aniline insertion (P2). The derivatized lignopolymeric (P2) layer covered the surface of a methacrylate particles (SC2/SC6) functionalized previously with amino phenolic crosslinker (p-phenylenediamine (p-Ph-2-NH2) or p-amino-2-hydroxybenzoic acid (p-NH2-SalA)). One-pot biocatalytic approach allowed to combine the preparation of the polymeric materials and also its attachment on the particles surface. Thus, the monolignol (CA) will be oxi-copolymerized together with aniline on the amino pre-functionalized support surface. The oxidation process was performed using H2O2 reagent and catalyzed by peroxidase enzyme (horseradish peroxidase, HRP). The biocatalytic process was evaluated in term of aromatic monomers (CA and aniline) conversion. The chemical structure and properties of the resulted ADL-composites were investigated using specific techniques (e.g. FTIR, TPD, TGA, static contact angle, elemental analysis, and gel permeation chromatography). Attachment of P1/P2 polymer on the particles improved the hydrophobicity and also the basicity of the composite surface. Lipase enzyme was immobilized on the ADL-composites for testing the applicability of ADL-composites for biocatalyst preparation. SEM analysis allowed to notice the modification of ADL-composites surface after enzyme immobilization. Immobilized lipase exhibited better activity compared with free lipase demonstrating the efficiency of ADL-composite as support for enzyme immobilization.

One-Pot Enzymatic Production of Lignin-Composites

A novel and efficient one-pot system for green production of artificial lignin bio-composites has been developed. Monolignols such as sinapyl (SA) and coniferyl (CA) alcohols were linked together with caffeic acid (CafAc) affording a polymeric network similar with natural lignin. The interaction of the dissolved SA/CA with CafAc already bound on a solid support (SC2/SC6-CafAc) allowed the attachment of the polymeric product direct on the support surface (SC2/SC6-CafAc-L1 and SC2/SC6-CafAc-L2, from CA and SA, respectively). Accordingly, this procedure offers the advantage of a simultaneous polymer production and deposition. Chemically, oxi-copolymerization of phenolic derivatives (SA/CA and CAfAc) was performed with H2O2 as oxidation reagent using peroxidase enzyme (2-1B mutant of versatile peroxidase from Pleurotus eryngii) as catalyst. The system performance reached a maximum of conversion for SA and CA of 71.1 and 49.8%, respectively. The conversion is affected by the system polarity as resulted from the addition of a co-solvent (e.g., MeOH, EtOH, or THF). The chemical structure, morphology, and properties of the bio-composites surface were investigated using different techniques, e.g., FTIR, TPD-NH3, TGA, contact angle, and SEM. Thus, it was demonstrated that the SA monolignol favored bio-composites with a dense polymeric surface, high acidity, and low hydrophobicity, while CA allowed the production of thinner polymeric layers with high hydrophobicity.

New materials used for the consolidation of archaeological wood–past attempts, present struggles, and future requirements

Abstract Given the perilous state of the Oseberg find from Norway, the Museum of Cultural History and the Department of Chemistry both at the University of Oslo, are looking into new methods for treating archaeological wood. While numerous polymers have been previously tested, most do not stabilise the wood sufficiently, penetrate far enough, or remain stable without producing toxic fumes. A few of the more common examples are: Alum salt, KAl(SO 4 ) 2 ·12H 2 O, which was used for treatment earlier but does not penetrate well and leaves the wood very acidic. Poly(oxy ethylene) (POE or Polyethylene glycol [PEG]) is widely used as a consolidant today but this material degrades over time and thus cannot support the finds for a very long time. Melamine-formaldehyde (Kauramin) has also been used and while it is fairly stable, it may also fill the wood and turn it into a ‘block’ of plastic. Since new consolidants would be advantageous, it is discussed what the requirements of such consolidants are and how material sciences may help procure them. It is proposed that an important requirement for a future stabilising agent is to leave an airy structure in order to allow retreatment in the future. This might be accomplished by foaming a polymer, or by combining nanoparticles with a polymer ‘spider web’ network to keep them in place. Such particles may help stabilise pH inside the wood by neutralising any acid generated inside treated artefacts. Special attention is given to the field of biomimetics–the discipline of constructing materials inspired by existing natural designs. It may be possible to construct a frame using bio-inspired materials (possibly an ‘artificial lignin’ mixed with other compounds optimise strength and flexibility) or through biomineralisation (an inorganic ‘skeleton’). Tests on biomimetic cellulose and chitosan have begun and the initial evaluation of these materials is given. Chitosan is made from modified chitin (primarily from shrimp and crabs) and may be dissolved in acidic solutions. Crystalline cellulose is interesting in conservation as the individual particles are resistant to acid and not as hygroscopic as the amorphous part of cellulose. The materials and the procedures used in testing are described. It is shown that crystalline cellulose particles usually flocculate when used to treat archaeological wood but that they may be treated with surfactants in order to improve penetration of archaeological finds.

Electropolymerization of coniferyl alcohol

Electropolymerization of coniferyl alcohol was carried out in an aqueous system (0.2 M NaOH) and in an organic solvent system [CH2Cl2/methanol (4:1 v/v) in the presence of 0.2 M LiClO4] to produce a dehydrogenation polymer (DHP) - artificial lignin. In both systems, the polymerization of coniferyl alcohol was visually confi rmed. In the aqueous system, no dimer was detected in the reaction medium after electropolymerization, suggesting that endwise polymerization occurred on the electrode surface. Thioacidolysis degradation revealed that the obtained polymers had numerous 8-O-4′ linkages. The electropolymerization products obtained in the organic solvent system also had numerous 8-O-4′ linkages; in particular, the polymers obtained in the initial polymerization stage. This was probably because of the limited area available for reaction and the orientation of coniferyl alcohol on the electrode surface controlled the polymerization.

Evaluation of Reaction Efficiency of Thioacidolysis for Cleavage of β-O-4 Interunitary Linkages by Using β-O-4 Type Artificial Lignin Polymer

Dimeric lignin model compounds with non-phenolic and phenolic moieties and a β -O-4 type artificial lignin polymer were subjected to thioacidolysis to evaluate the reaction efficiency of thioacidolysis for cleavage of β -O-4 interunitary linkage. The obtained yields of thioethylated monomeric products from the dimeric lignin model compounds reached nearly 100% under the conventional condition, whereas that from the artificial lignin polymer was as low as 74%. Neither prolonged reaction time nor increased concentration of ethanethiol (the thioacidolysis reagent) enhanced the resulting monomer yields from the polymer (69–79%). Thioacidolysis of the lignin model compounds followed by HPSEC analysis also showed the dimeric model compounds were degraded almost quantitatively, but that the artificial lignin polymer was not. Thioacidolysis followed by desulfurization gave at least one dimeric product resulting from incomplete β -O-4 cleavage at significant yield. These results suggested the conventional thioacidolysis could not achieve quantitative cleavage of β -O-4 linkages in lignin macromolecules.

Synthesis of Bromoacetophenone Derivatives as Starting Monomers for β-O-4 Type Artificial Lignin Polymers

Guaiacyl, syringyl, and p-hydroxyphenyl-type bromoacetophenone derivatives were synthesized as the starting materials for β-O-4 type artificial lignin polymers. They were prepared from 4-hydroxyacetophenone instead of 4-hydroxybenzaldehyde, which was used in the previous investigations. The new route required 4 reaction steps. Overall yields of the monomers were 60, 79, and 68%, respectively.

Synthesis of β-O-4-type artificial lignin polymers and their analysis by NMR spectroscopy

We describe the synthesis and NMR spectroscopic analysis of three artificial lignin polymers containing only the beta-O-4 substructure: syringyl-type homopolymer, p-hydroxyphenyl-type homopolymer and guaiacyl/syringyl-type heteropolymer. Using gel permeation chromatography, the weight-average degree of polymerization (DP(w)) of the three polymers was determined as 19.2, 38.6, and 13.9, respectively. The polymers were prepared based on the synthetic methodology of guaiacyl-type homopolymer, and were fully characterized using (1)H-, (13)C-, and (1)H-(13)C NMR spectroscopy of the acetylated and non-acetylated forms. The spectra of guaiacyl/syringyl-type heteropolymers were in good agreement with those of the beta-O-4 substructure of milled wood lignin obtained from the hardwood of Japanese white birch.

Chemical synthesis of β-O-4 type artificial lignin

An artificial lignin polymer containing only the beta-O-4 substructure was synthesized. The procedure consists of two key steps: 1) polycondensation of a brominated monomer by aromatic Williamson reaction; and 2) subsequent reduction of the carbonyl polymer. 13C-NMR and HMQC spectra of the polymer were consistent with beta-O-4 substructures in milled wood lignin isolated from Japanese fir wood. The weight average degree of polymerization (DP(w)) ranged from 19.5 to 30.6, which is comparable to enzymatically synthesized artificial lignin from p-hydroxycinnamyl alcohols (dehydrogenation polymer, DHP) and some isolated lignins. Using this new lignin model polymer, it will now be possible to reinvestigate the properties and reactivity of the main lignin structure in terms of its polymeric character.

A Comparative Study of Enzymatically and Photochemically Polymerized Artificial Lignin Supramolecular Structures Using Environmental Scanning Electron Microscopy

Environmental scanning electron microscopy images of the self-assembled structures of enzymatically (DHP) and photochemically polymerized (PCP) artificial lignin are herein presented. Differences in the structural organization between DHP and PCP polymer at the supramolecular level were reported. Based on topological information, we proposed a hypothesis about possible new physiological roles of lignin in live plant cells and the ecological significance of possible in-vivo photochemical lignin polymerization.

Incorporation of [2-13C]ferulic acid, a lignin precursor, into Leucaena leucocephala and its analysis by solid state 13C n.m.r. spectroscopy

Solid state 13C n.m.r. analysis of lignin in situ, in the root tissue of the woody angiosperm, Leucaena leucocephala, and specifically labelled at a carbon site thought to be involved in the majority of inter-unit linkages, revealed significant bonding differences when compared to an artificial lignin preparation generally viewed more or less to represent native lignin structure.

Weitere Versuche mit deuteriertem Coniferylalkohol

Abstract6‐ und 2‐Deutero‐coniferylalkohol wurden enzymatisch zu künstlichem Lignin dehydriert. Dabei verliert die C9‐Einheit des 6‐Deutero‐coniferylalkohols 8% ihres Deuteriums und die des 2‐Deutero‐coniferylalkohols 4%. Daß sich die C‐Atome 6 und 2 an der Kondensation beteiligen, geht auch aus dem Vorkommen der methoxylierten O‐Phthalsäuren unter den Abbausäuren des Lignins hervor.