Lignin-carbohydrate Complexes Research Articles

Chemical and thermal properties of precipitates made from hydrolysate of spruce wood chips

Autohydrolysis is a widely used technology for extracting hemicelluloses from wood chips. In addition to hemicelluloses, hydrolysates produced in this process contain lignin. Hemicelluloses and lignin could be isolated from hydrolysates and used for generating value-added products. In this study, hydrolysates were produced via autohydrolysis of spruce wood chips. Then, acidification and ethanol precipitation were employed to isolate lignocelluloses from hydrolysates. Generally, acidification and ethanol treatment led to the precipitation of lignin–carbohydrate complexes, but their removals depended on the hydrolysis conditions. The heat capacity (Cp) of the dried hydrolysates was in the range of 0.41–0.45 J/g °C, while the precipitates of acidification and ethanol treatment had the Cp of 1.18 J/g °C and 1.38 J/g °C, respectively. This increment could be related to their higher lignin content and thus their molecular mobility as these segments had a higher amount of methoxyl groups and more Hα and Hβ inter-linkages. The results of this study suggested that the lignocellulosic materials obtained after acid or ethanol treatment of hydrolysates could be considered as additives for heat-resistant biopolymers or hemicellulose-based films, respectively.

Mechanochemical deconstruction of lignocellulosic cell wall polymers with ball-milling

In this work, the deconstruction mechanism of corn stover cell wall polymers during ball milling was evaluated. The characterization showed that ball milling not only brought about the dissociation of the cross-linked cellulose-hemicellulose-lignin complex but also led to the depolymerization of the cell-wall polymers especially the carbohydrates. Micromorphology characterization revealed that mechanical treatment disrupted the orderly fibrillar matrices with a porous structure. The breakage of β-1,4 glycosidic bonds in cellulose and the decomposition of arabinoxylans indicated the modification in polysaccharide chains. The degradation of lignin-carbohydrate complex (LCC) linkages and the cleavage of β-O-4′ linkages in lignin approved the partial degradation of lignin. In conclusion, mechanochemistry is an efficient force to make the polymers in plant fibers more digestible.

Pulp improvement of oil palm empty fruit bunches associated to solid-state biopulping and biobleaching with xylanase and lignin peroxidase cocktail produced by Aspergillus sp. LPB-5

Oil palm empty fruit bunches is a lignocellulosic feedstock with biotechnological potential and thousands of tons are generated in the world each year. Filamentous fungi producing xylanases and ligninases in biopulping to obtain cellulose is a pulp improvement alternative. The enzymatic cocktail was produced in solid-state biopulping by Aspergillus sp. LPB-5 with 54.32 U/g xylanase, 13.41 U/g lignin peroxidase and low cellulase activity. Biological, thermal and chemical pretreatments were compared and enzymatic biobleaching was applied to pretreated pulps. Biopulping and biobleaching combination had 36.80% lignin loss, 26.27% hemicellulose reduction, 74.36% pulp yield with 36.56% digestibility. Alkaline and biobleaching combination removed 81.97% hemicellulose and 93.89% lignin with 73.59% digestibility. Enzymatic biobleaching increased the pulp digestibility in all pretreatments. Finally, the development of a bio-pretreatment to remove hemicellulose and alter the lignin-carbohydrate complex interface presented a soft process with great eco-friendly potential, where mild pre-treatments would reduce the use of aggressive agents.

Unveiling TMP Process Water Potential As an Industrial Sourcing of Valuable Lignin–Carbohydrate Complexes toward Zero-Waste Biorefineries

The development of zero-waste biorefineries is a promising yet challenging pathway toward a sustainable bioeconomy. Deep insights on the composition of biorefineries side streams along with the development of green technologies are mandatory to achieve such a goal. In this Article, a sustainable pilot-scale membrane process was used to recover valuable organic matter dissolved in the main side stream of a thermomechanical pulping (TMP) plant. A great number of analytical tools were combined to get an extensive characterization of the generated fraction, including inductively coupled plasma atomic emission spectroscopy (ICP-AES), high-performance ion chromatography coupled with pulse amperometric detection (HPIC-PAD), Fourier transform infrared (FTIR), size-exclusion chromatography (SEC), and C-13 and 2D heteronuclear single quantum correlation (HSQC) NMR. The results revealed the presence of a considerable amount of native lignin-carbohydrate complexes (LCCs) in the fraction representing 40% of the dry weight. The LCCs are composed of partially acetylated galactoglucomannans (GGMs) covalently linked to low condensed and ) beta-O-4-rich lignin through phenyl glycosidic bonds (25.5 bonds/100 aromatic rings (ar)) and characterized by a relatively low molecular weight around 7 kDa. Such complexes present an interesting potential for valorization with unique properties resulting from the combination of their aromatic and sugar moieties. Different applications are discussed in regards to the generated data in this work as an industrial sourcing of LCC is reported.

Expression and characterization of two glucuronoyl esterases from Thielavia terrestris and their application in enzymatic hydrolysis of corn bran.

The thermophilic fungus Thielavia terrestris when cultured on cellulose produces a cocktail of thermal hydrolases with potential application in saccharification of lignocellulosic biomass and other biotechnological areas. Glucuronoyl esterases are considered to play a unique role as accessory enzymes in lignocellulosic material biodegradation by cleaving the covalent ester linkage between 4-O-methyl-D-glucuronic acid (MeGlcA) and lignin in lignin-carbohydrate complexes (LCCs). Two glucuronoyl esterases from T. terrestris named TtGE1 and TtGE2 were expressed in Pichia pastoris. Both esterases displayed features of thermophilic enzymes, with the optimal temperature at 45°C and 55°C. TtGE1 and TtGE2 exhibited activity towards methyl (4-nitrophenyl β-D-glucopyranosid) uronate (Me-GlcA-pNP) but no catalytic activity to benzyl-D-glucuronate (BnzGlcA), indicating the difference in substrate specificity from previously studied fungal GEs. A substantial increase in the release of monomeric sugars and glucuronic acid from autohydrolysis of corn bran was observed by the supplementing TtGEs into commercial xylanase; the results clearly demonstrated that the TtGEs played a significant role in this degradation process. This research on TtGEs enriches our knowledge of this novel class of fungal GEs. These newly characterized TtGEs could be used as promising accessory enzymes to improve the hydrolysis efficiency of commercial enzymes in saccharification of lignocellulosic materials due to their thermophilic characteristics.

Facile Isolation of LCC-Fraction from Organosolv Lignin by Simple Soxhlet Extraction.

A new fractionation protocol for wheat straw organosolv lignin was developed on the basis of the dominating H-bonding orientations of its components. Acetone as H-bond accepting aprotic polar solvent and methanol as H-bond donating and accepting protic polar solvent were used in sequence. Obtained fractions were structurally and thermally analysed. The protocol allowed for the generation of purified lignin fractions and the isolation of a novel, yet unobserved lignin carbohydrate complex (LCC) fraction. This LCC fraction was found to contain exclusively phenyl glycosides and γ-esters as LCC motifs.

The influence of lignin content and structure on hemicellulose alkaline extraction for non-wood and hardwood lignocellulosic biomass

The extractability of hemicellulose from different lignocellulosics depends on the source of biomass. Differences in hemicellulose extractability are believed to be due to plant-specific hemicellulose arrangement alongside lignin within the cell wall. In this research, six biomasses were used to probe hemicellulose alkaline extractability as a function of the native lignin within the biomasses. Quantitative 2D-HSQC and 13C NMR analysis were performed to determine the S/G (S: syringyl, G: guaiacyl) and lignin-carbohydrate complex (LCC) linkages of milled wood lignin isolated from these biomasses. A strong negative correlation was observed between total lignin content and hemicellulose extractability, demonstrating that a greater presence of lignin in the original material results in lower xylan solubilization. In addition, a correlation between S/G of lignin and xylan dissolution was found within a group of hardwoods and within a group of non-woods. This suggests that monomeric constituency also influences xylan’s propensity for dissolution in 10% NaOH. Although there is some uncertainty in the quantification of LCC linkages, both non-woods and hardwoods exhibited negative correlations between alkaline-stable LCC linkages content and xylan extractability. This suggests that alkaline-stable LCC structures are associated with a decrease in the alkaline extractability of hemicellulose.

Search for Drugs Used in Hospitals to Treat Stomatitis.

Stomatitis is an inflammatory disease of the oral mucosa, often accompanied by pain. Usually it is represented by aphthous stomatitis, for which treatment steroid ointment is commonly used. However, in the cases of refractory or recurrent stomatitis, traditional herbal medicines have been used with favorable therapeutic effects. Chemotherapy, especially in the head and neck region, induces stomatitis at higher frequency, which directly affects the patient’s quality of life and treatment schedule. However, effective treatment for stomatitis has yet to be established. This article presents the clinical report of Kampo medicines on the stomatitis patients in the Nihon university, and then reviews the literature of traditional medicines for the treatment of stomatitis. Among eighteen Kampo medicines, Hangeshashinto has been the most popular for the treatment of stomatitis, due to its prominent anti-inflammatory activity. It was unexpected that clinical data of Hangeshashinto on stomatitis from Chinese hospital are not available. Kampo medicines have been most exclusively administered to elder person, as compared to pediatric population. Supplementation of alkaline plant extracts rich in lignin-carbohydrate complex may further extend the applicability of Kampo medicines to viral diseases.

Downregulation of pectin biosynthesis gene GAUT4 leads to reduced ferulate and lignin-carbohydrate cross-linking in switchgrass

Knockdown (KD) expression of GAlactUronosylTransferase 4 (GAUT4) in switchgrass improves sugar yield and ethanol production from the biomass. The reduced recalcitrance of GAUT4-KD transgenic biomass is associated with reduced cell wall pectic homogalacturonan and rhamnogalacturonan II content and cross-linking, and the associated increases in accessibility of cellulose to enzymatic deconstruction. To further probe the molecular basis for the reduced recalcitrance of GAUT4-KD biomass, potential recalcitrance-related factors including the physicochemical properties of lignin and hemicellulose are investigated. We show that the transgenic switchgrass have a lower abundance of ferulate and lignin-carbohydrate complex cross-linkages, reduced amounts of residual arabinan and xylan in lignin-enriched fractions after enzymatic hydrolysis, and greater coalescence and migration of lignin after hydrothermal pretreatment in comparison to the wild-type switchgrass control. The results reveal the roles of both decreased lignin-polymer and pectin cross-links in the reduction of recalcitrance in PvGAUT4-KD switchgrass.

Structural characterization of lignin and its carbohydrate complexes isolated from bamboo (Dendrocalamus sinicus)

Isolation of earth abundant biopolymer, Lignin, from Dendrocalamus sinicus and their structural properties were investigated to achieve its large-scale practical applications in value-added products. Two lignin fractions (MWL, DSL) were isolated with successive treatments of dioxane and dimethylsulfoxide (DMSO) from dewaxed and ball milled bamboo (D. sinicus) sample. The two-step treatments yielded 52.1% lignin based on the total lignin content in the dewaxed bamboo sample. Spectroscopy analyses indicated that the isolated bamboo lignin was a typical grass lignin, consisting of p-hydroxyphenyl, guaiacyl, and syringyl units. The major interunit linkages presented in the obtained bamboo lignin were β-O-4′ aryl ether linkages, together with lower amounts of β-β', β-5′, and β-1′ linkages. The tricin was detected to be linked to lignin polymer through the β-O-4′ linkage in the bamboo. In addition, phenyl glycoside and benzyl ether lignin-carbohydrate complexes (LCC) linkages were clearly detected in bamboo (D. sinicus), whereas the γ-ester LCC linkages were ambiguous due to the overlapping NMR signals with other substructures. The detailed structural properties of the obtained lignin fraction together with the light-weight will benefit efficient utilization of natural polymers as a possibly large-scale bio-based precursor for making polymeric materials, biochemicals, functional carbon and biofuels, and multifunctional polymer nanocomposites.

Recent Progress of Basic Studies of Natural Products and Their Dental Application

The present article reviews the research progress of three major polyphenols (tannins, flavonoids and lignin carbohydrate complexes), chromone (backbone structure of flavonoids) and herbal extracts. Chemical modified chromone derivatives showed highly specific toxicity against human oral squamous cell carcinoma cell lines, with much lower toxicity against human oral keratinocytes, as compared with various anticancer drugs. QSAR analysis suggests the possible correlation between their tumor-specificity and three-dimensional molecular shape. Condensed tannins in the tea extracts inactivated the glucosyltransferase enzymes, involved in the biofilm formation. Lignin-carbohydrate complexes (prepared by alkaline extraction and acid-precipitation) and crude alkaline extract of the leaves of Sasa species (SE, available as an over-the-counter drug) showed much higher anti-HIV activity, than tannins, flavonoids and Japanese traditional medicine (Kampo). Long-term treatment with SE and several Kampo medicines showed an anti-inflammatory and anti-oxidant effects in small size of clinical trials. Although the anti-periodontitis activity of synthetic angiotensin II blockers has been suggested in many papers, natural angiotensin II blockers has not yet been tested for their possible anti-periodontitis activity. There should be still many unknown substances that are useful for treating the oral diseases in the natural kingdom.

Valorization of Lignin–Carbohydrate Complexes from Hydrolysates of Norway Spruce: Efficient Separation, Structural Characterization, and Antioxidant Activity

Hot-water extraction has shown great potential in a green biorefinery; however, efficient separation of lignin and carbohydrates is still a big challenge, especially to lignin–carbohydrate complexes (LCCs). The aim of the present study was to develop a facile approach for efficiently fractionating LCCs from hot-water extraction liquor of spruce wood to understand their comprehensive structure and thus to achieve the valorization of fractions. Approximately 93% of the hemicelluloses were first recovered by a selective adsorptive resin from hot-water hydrolysates. The lignin-rich fractions obtained from desorption of the adsorbed compounds were further subjected to a gradient dialysis procedure toward functional fractionation. Linear relationship between lignin and uronic acids as well as xylose contents in graded fractions of all hydrolysates was observed. Further characterization of LCCs by two-dimensional nuclear magnetic resonance and pyrolysis-gas chromatography-mass spectrometry showed the large diffe...

Activation of Enzymatic Hydrolysis Lignin by NaOH/Urea Aqueous Solution for Enhancing Its Sulfomethylation Reactivity

Enzymatic hydrolysis lignin (EHL), from straw bioethanol process, was activated using NaOH/urea aqueous solution under low temperature condition to enhance its sulfonation degree and dispersion property. The sulfonation degree of EHL increased by 62.60% after the activation of NaOH/urea aqueous solution at −10 °C for 12 h, which is resulted from the breakage of the hydrogen bonds and ether bonds in lignin–carbohydrate complexes (LCCs) and the removal of residual carbohydrate. The functional group content, 2D-HSQC, and AFM analysis indicated that the increase of phenolic groups and the weakened aggregation degree of EHL after the activation are in favor of the increase of activity sites and decrease of steric hindrance, which is beneficial to the enhancement of sulfomethylation reactivity. Owing to the enhancement of reactivity, the dispersion properties of sulfomethylated EHL on graphite suspension were significantly enhanced.

Decay of Carpinus betulus wood by Trametes versicolor - An anatomical and chemical study

Decay patterns occurring in wood of hornbeam (Carpinus betulus) during natural decay in vivo were investigated and compared to decay in artificially inoculated wood in vitro. The fungal species causing decay was determined by morphological characterization and confirmed as Trametes versicolor using rDNA-ITS PCR and sequencing. T. versicolor caused degradation with properties of both simultaneous and selective white-rot. Chemical and FT-IR analyses suggested that T. versicolor caused severe reduction of lignin-carbohydrate complexes and guaiacyl lignin within the cell walls during natural decay. Under the light microscope, it was clear that decomposition of cell elements in the reaction zones of naturally decayed wood was greater than in inoculated wood. Histological analyses of inoculated wood blocks suggested that T. versicolor was able to switch from a typical simultaneous rot to selective delignification, with degradation of the middle lamella, and of lignin in cell corners. The results of our work demonstrated that test fungus able to attack the hornbeam tree by saprotrophic and parasitic life strategies.

Microwave-assisted recovery of monomeric sugars from an acidic steam treated wood hydrolysate

Fractionation of components from bio-refinery wastes streams is complicated by the presence of both oligomer and lignin fractions. Microwave-assisted acid hydrolysis was used in this study to convert oligomer sugars in an industrial prehydrolysis liquor (PHL) to monomeric sugars. A total of 19.6 g/L monomeric sugars was obtained at a combined severity factor (CSF) of 3.2. Furthermore, it was found that xylan linked to lignin in a lignin-carbohydrate complex (LCC) could be liberated, resulting in lignin with a relatively low dispersity (3.12) and average molecular weight (1718 g/mol) that has high commercial value in the phenol–formaldehyde resin industry. This study presents for the first time a relatively inexpensive method for recovery of 100% of available sugars in the PHL without apparent loss in monomeric sugar as well as 50% removal of lignin as a valuable by-product. Application of this method can significantly improve the economic sustainability of forest-based biorefineries.

Classification of fungal glucuronoyl esterases (FGEs) and characterization of two new FGEs from Ceriporiopsis subvermispora and Pleurotus eryngii.

Fungal glucuronoyl esterases (FGEs) catalyze cleavage of the ester bond connecting a lignin alcohol to the xylan-bound 4-O-methyl-D-glucuronic acid of glucuronoxylans. Thus, FGEs are capable of degrading lignin-carbohydrate complexes and have potential for biotechnological applications toward woody biomass utilization. Therefore, identification and characterization of new FGEs are of critical importance. Firstly, in this study, we built a phylogenetic tree from almost 400 putative FGEs obtained on BLAST analysis and defined six main clades. In the phylogenetic tree, all the putative FGEs of ascomycetes cluster in clades I to IV, and most of the putative FGEs of basidiomycetes (B-FGEs) cluster in clades V to VI. Interestingly, several B-FGEs were found to cluster in clade II; most FGEs of clade II were found to have higher theoretical isoelectric points than those in the other five clades. To gain an insight into the putative FGEs in the clades that have not been characterized yet, we chose the FGEs of Ceriporiopsis subvermispora (CsGE) and Pleurotus eryngii (PeGE), which belong to clades V and II, respectively. The catalytic domains of both CsGE and PeGE were successfully expressed using Pichia pastoris, and then purified. Benzyl glucuronic acid was used as a substrate to confirm the activities of the CsGE and PeGE, and the hydrolyzed product, glucuronic acid, was quantified spectrophotometrically. Both CsGE and PeGE clearly exhibited the esterase activity. Additionally, we demonstrated that PeGE exhibits high tolerance toward several denaturing agents, which may make it a potentially more applicable enzyme.

Characterization of lignin-derived products from various lignocellulosics as treated by semi-flow hot-compressed water

To elucidate the decomposition behaviors of lignin from different taxonomic groups, five different lignocellulosics were treated with hot-compressed water (230 °C/10 MPa/15 min) to fractionate lignins into water-soluble portions, precipitates, and insoluble residues. The lignin-derived products in each fraction were characterized and compared. The delignification of monocotyledons [nipa palm (Nypa fruticans) frond, rice (Oryza sativa) straw, and corn (Zea mays) cob] was more extensive than that achieved for Japanese cedar (Cryptomeria japonica, gymnosperm) and Japanese beech (Fagus crenata, dicotyledon angiosperm). The water-soluble portions contained lignin monomers like coniferyl alcohol and phenolic acids, while the precipitates contained higher molecular weight lignin with high content of ether-type linkages. Lignin in the insoluble residues was rich in condensed-type structures. In all five lignocellulosics, ether-type linkages were preferentially cleaved, while condensed-type lignin showed resistance to hot-compressed water. In the monocotyledons, lignin–carbohydrate complexes were cleaved and gave lignins that had higher molecular weights than those eluted from the woods. These differences would facilitate the delignification in monocotyledons. Such information provides useful information for efficient utilization of various lignocellulosics.

Unveiling the Structural Properties of Lignin–Carbohydrate Complexes in Bamboo Residues and Its Functionality as Antioxidants and Immunostimulants

Lignin–carbohydrate complexes (LCCs), a significant component of plant cell walls, have been found to bear biological functionality as antioxidants in food and as immunostimulants for living cell. ...

Effect of autohydrolysis on alkaline delignification of mixed hardwood chips and on lignin structure

AbstractAutohydrolysis (AH) prior to the kraft process is one way to valorize hemicelluloses. The presented results show that AH also has a beneficial effect insofar as kraft cooking can be substituted by soda cooking, and higher brightness levels can be achieved under the same bleaching conditions. The lignin has more free phenolic groups after AH, which is attributable to depolymerization, and which could partly explain the better pulping and bleaching results. Another explanation could be the lower amount of lignin involved in lignin carbohydrate complexes after an AH step.

Mass Spectrometry Exposes Undocumented Lignin-Carbohydrate Complexes in Biorefinery Pretreatment Stream

The presence and effects of lignin–carbohydrate complexes (LCCs) in biorefinery processes are largely unknown. Recently identified in a significant hydrothermal pretreatment process stream (autohydrolyzate), these molecules likely influence not only downstream processing but also product purity and performance. However, without an understanding of their structures, it is impossible to analyze and cope with their effects. To identify and elucidate LCCs in autohydrolyzate, a new method employed chloride doping and tandem mass spectrometry. The results showed complexes ranging in mass from 326–714 Da with evidence of xylose and glucose units. Further analysis revealed lignin-like levels of unsaturation, considering the number of carbons present. These results suggest that, although the complexes contain one or two carbohydrate units, their primary structures are closer to lignin compounds. This method provides a glimpse into the structures of LCCs in a biorefinery process stream, laying the foundation for ea...