Application of ESI FT-ICR MS to Study Kraft Lignin Modification by the Exoenzymes of the White Rot Basidiomycete Fungus TrametesHirsutaLE-BIN 072

Trameteshirsuta is a wood rotting fungus that possesses a vast array of lignin degrading enzymes, including7 laccases, 7 ligninolyticmanganese peroxidases, 9 lignin peroxidases and 2 versatile peroxidases. In this study,electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI FT-ICR MS)was used to examine kraft lignin modification by the enzymatic system of this fungus.The observed pattern of lignin modification suggested that before the 6th day of cultivation,the fungal enzymatic system tended to degrade more oxidized moleculesand, hence, less recalcitrant molecules, with the production of hard-to-modify reduced molecular species. At some point after the 6th day of cultivation,the fungal enzymatic system tended to degrade more oxidized moleculesand, hence, less recalcitrant molecules, with the production of hard-to-modify reduced molecular species. At some point after the 6th day of cultivation,the fungus started to degrade less oxidized, more recalcitrant, compounds, converting them into the more oxidized forms. The altered pattern of lignin modification enabled changes in the fungal enzymatic system. These changes were further attributed to the appearance of the particular ligninolyticmanganese peroxides enzyme(MnP7), which was added by the fungus to the mixture of enzymes that had already been secreted (VP2 and MnP5). Keywords: wood rotting fungi, kraft lignin, mass spectrometry, peroxidases

The Inducible Accumulation of Cell Wall-Bound p-Hydroxybenzoates Is Involved in the Regulation of Gravitropic Response of Poplar

Lignin in Populus species is acylated with p-hydroxybenzoate. Monolignol p-hydroxybenzoyltransferase 1 (PHBMT1) mediates p-hydroxybenzoylation of sinapyl alcohol, eventually leading to the modification of syringyl lignin subunits. Angiosperm trees upon gravistimulation undergo the re-orientation of their growth along with the production of specialized secondary xylem, i.e., tension wood (TW), that generates tensile force to pull the inclined stem or leaning branch upward. Sporadic evidence suggests that angiosperm TW contains relatively a high percentage of syringyl lignin and lignin-bound p-hydroxybenzoate. However, whether such lignin modification plays a role in gravitropic response remains unclear. By imposing mechanical bending and/or gravitropic stimuli to the hybrid aspens in the wild type (WT), lignin p-hydroxybenzoate deficient, and p-hydroxybenzoate overproduction plants, we examined the responses of plants to gravitropic/mechanical stress and their cell wall composition changes. We revealed that mechanical bending or gravitropic stimulation not only induced the overproduction of crystalline cellulose fibers and increased the relative abundance of syringyl lignin, but also significantly induced the expression of PHBMT1 and the increased accumulation of p-hydroxybenzoates in TW. Furthermore, we found that although disturbing lignin-bound p-hydroxybenzoate accumulation in the PHBMT1 knockout and overexpression (OE) poplars did not affect the major chemical composition shifts of the cell walls in their TW as occurred in the WT plants, depletion of p-hydroxybenzoates intensified the gravitropic curving of the plantlets in response to gravistimulation, evident with the enhanced stem secant bending angle. By contrast, hyperaccumulation of p-hydroxybenzoates mitigated gravitropic response. These data suggest that PHBMT1-mediated lignin modification is involved in the regulation of poplar gravitropic response and, likely by compromising gravitropism and/or enhancing autotropism, negatively coordinates the action of TW cellulose fibers to control the poplar wood deformation and plant growth.

A Comparison among Lignin Modification Methods on the Properties of Lignin–Phenol–Formaldehyde Resin as Wood Adhesive

The research aim of this work is to determine the influence of lignin modification methods on lignin–phenol–formaldehyde (LPF) adhesive properties. Thus, glyoxal (G), phenol (P), ionic liquid (IL), and maleic anhydride (MA) were used to modify lignin. The modified lignins were used for phenol substitution (50 wt%) in phenol–formaldehyde adhesives. The prepared resins were then used for the preparation of wood particleboard. These LPF resins were characterized physicochemically, namely by using standard methods to determine gel time, solids content, density, and viscosity, thus the physicochemical properties of the LPF resins synthesized. The panels dimensional stability, formaldehyde emission, bending modulus, bending strength, and internal bond (IB) strength were also measured. MA-modified lignin showed by differential scanning calorimetry (DSC) the lowest temperature of curing than the resins with non-modified lignin and modified with IL, phenolared lignin, and glyoxal. LPF resins with lignin treated with maleic anhydride presented a shorter gel time, higher viscosity, and solids content than the resins with other lignin modifications. Equally, the particleboard panels prepared with LPF resins with maleic anhydride or with ionic liquid had the lowest formaldehyde emission and the highest mechanical strength among all the synthesized resins. The dimensional stability of all panels bonded with modified lignin LPF resins presented no difference of any significance.

Effect of pH and pH-shifting on Lignin, Lignin-protein Properties, and Lignin-protein Interactions

Innovation of high-value added lignin derivatives has become a topic of interest, but lignin modification and utilization remain a challenge due to lignin’s complicated structure. In our previous study, we successfully improved water resistance and adhesion performance of soy protein adhesives using unmodified lignin. This study focuses on lignin modification as well as lignin-protein properties and interactions. Lignin modification was achieved by pH changes and pH-shifting. Cleavage of β-O-4 linkage was observed at pH 8.5 and pH 12, resulting in smaller particle sizes and changes in thermal properties. Partial repolymerization was found after pH-shifting treatments. Lignin increased the strength of protein films under high temperature and significantly enhanced the water resistance of soy protein at pH 12. Cross-linking of protein and lignin took place via carbonyl, amino, and hydroxyl groups. Multiple-point interactions between lignin and protein resulted in a stronger lignin-protein network. Additional lignin-protein complexes with high molecular weight were detected with an elevated lignin concentration at pH 8.5. The binding interaction between lignin and protein, although of non-specific nature, was also observed by isothermal titration calorimetry (ITC). Findings of this study contribute to the further development of green lignin-protein products.

Lignin functionalization strategies and the potential applications of its derivatives – A Review

Lignin is one of the most important and widespread carbon sources on Earth. Significant amounts of lignin are delivered to the market by pulp mills and biorefineries, and there have been many efforts to develop routes for its valorization. Over the years, lignin has been used to produce biobased chemicals, materials, and advanced biofuels on the basis of its variable functionalities and physicochemical properties. Today, lignin’s applications are still limited by its heterogeneity, variability, and low reactivity. Thus, modification technologies have been developed to allow lignin to be suitable for a wider range of attractive industrial applications. The most common modifications used for this purpose include amination, methylation, demethylation, phenolation, sulfomethylation, oxyalkylation, acylation or esterification, epoxidation, phosphorylation, nitration, and sulfonation. This article reviews the chemistry involved in these lignin modification technologies, discussing their effect on the finished product while presenting some market perspectives and future outlook to utilize lignin in sustainable biorefineries.

How Do Shipworms Eat Wood? Screening Shipworm Gill Symbiont Genomes for Lignin-Modifying Enzymes

Shipworms are ecologically and economically important mollusks that feed on woody plant material (lignocellulosic biomass) in marine environments. Digestion occurs in a specialized cecum, reported to be virtually sterile and lacking resident gut microbiota. Wood-degrading CAZymes are produced both endogenously and by gill endosymbiotic bacteria, with extracellular enzymes from the latter being transported to the gut. Previous research has predominantly focused on how these animals process the cellulose component of woody plant material, neglecting the breakdown of lignin – a tough, aromatic polymer which blocks access to the holocellulose components of wood. Enzymatic or non-enzymatic modification and depolymerization of lignin has been shown to be required in other wood-degrading biological systems as a precursor to cellulose deconstruction. We investigated the genomes of five shipworm gill bacterial symbionts obtained from the Joint Genome Institute Integrated Microbial Genomes and Microbiomes Expert Review for the production of lignin-modifying enzymes, or ligninases. The genomes were searched for putative ligninases using the Joint Genome Institute’s Function Profile tool and blastp analyses. The resulting proteins were then modeled using SWISS-MODEL. Although each bacterial genome possessed at least four predicted ligninases, the percent identities and protein models were of low quality and were unreliable. Prior research demonstrates limited endogenous ability of shipworms to modify lignin at the chemical/molecular level. Similarly, our results reveal that shipworm bacterial gill-symbiont enzymes are unlikely to play a role in lignin modification during lignocellulose digestion in the shipworm gut. This suggests that our understanding of how these keystone organisms digest and process lignocellulose is incomplete, and further research into non-enzymatic and/or other unknown mechanisms for lignin modification is required.