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Prefoldin (PFD) is a group II chaperonin that is ubiquitously present in the eukaryotic kingdom. Six subunits (PFD1-6) form a jellyfish-like heterohexameric PFD complex and function in protein folding and cytoskeleton organization. However, little is known about its function in plant cell wall-related processes. Here, we report the functional characterization of a PFD gene from Populus deltoides, designated as PdPFD2.2. There are two copies of PFD2 in Populus, and PdPFD2.2 was ubiquitously expressed with high transcript abundance in the cambial region. PdPFD2.2 can physically interact with DELLA protein RGA1_8g, and its subcellular localization is affected by the interaction. In P. deltoides transgenic plants overexpressing PdPFD2.2, the lignin syringyl/guaiacyl ratio was increased, but cellulose content and crystallinity index were unchanged. In addition, the total released sugar (glucose and xylose) amounts were increased by 7.6% and 6.1%, respectively, in two transgenic lines. Transcriptomic and metabolomic analyses revealed that secondary metabolic pathways, including lignin and flavonoid biosynthesis, were affected by overexpressing PdPFD2.2. A total of eight hub transcription factors (TFs) were identified based on TF binding sites of differentially expressed genes in Populus transgenic plants overexpressing PdPFD2.2. In addition, several known cell wall-related TFs, such as MYB3, MYB4, MYB7, TT8 and XND1, were affected by overexpression of PdPFD2.2. These results suggest that overexpression of PdPFD2.2 can reduce biomass recalcitrance and PdPFD2.2 is a promising target for genetic engineering to improve feedstock characteristics to enhance biofuel conversion and reduce the cost of lignocellulosic biofuel production.

BackgroundLignocellulosic biomass is recognized as a promising renewable feedstock for the production of biofuels. However, current methods for converting biomass into fermentable sugars are considered too expensive and inefficient due to the recalcitrance of the secondary cell wall. Biomass composition can be modified to create varieties that are efficiently broken down to release cell wall sugars. This study focused on identifying the key biomass components influencing plant cell wall recalcitrance that can be targeted for selection in sugarcane, an important and abundant source of biomass.ResultsBiomass composition and the amount of glucan converted into glucose after saccharification were measured in leaf and culm tissues from seven sugarcane genotypes varying in fiber composition after no pretreatment and dilute acid, hydrothermal and ionic liquid pretreatments. In extractives-free sugarcane leaf and culm tissue, glucan, xylan, acid-insoluble lignin (AIL) and acid-soluble lignin (ASL) ranged from 20 to 32%, 15% to 21%, 14% to 20% and 2% to 4%, respectively. The ratio of syringyl (S) to guaiacyl (G) content in the lignin ranged from 1.5 to 2.2 in the culm and from 0.65 to 1.1 in the leaf. Hydrothermal and dilute acid pretreatments predominantly reduced xylan content, while the ionic liquid (IL) pretreatment targeted AIL reduction. The amount of glucan converted into glucose after 26 h of pre-saccharification was highest after IL pretreatment (42% in culm and 63.5% in leaf) compared to the other pretreatments. Additionally, glucan conversion in leaf tissues was approximately 1.5-fold of that in culm tissues. Percent glucan conversion varied between genotypes but there was no genotype that was superior to all others across the pretreatment groups. Path analysis revealed that S/G ratio, AIL and xylan had the strongest negative associations with percent glucan conversion, while ASL and glucan content had strong positive influences.ConclusionTo improve saccharification efficiency of lignocellulosic biomass, breeders should focus on reducing S/G ratio, xylan and AIL content and increasing ASL and glucan content. This will be key for the development of sugarcane varieties for bioenergy uses.

Cationic cell-wall-bound peroxidase (CWPO-C) from Populus alba is the only Class III peroxidase that has been shown to be able to oxidize high molecular weight lignin polymers from sinapyl alcohol and previously, has been believed to be a lignin polymerization-specific peroxidase. However, using an Arabidopsis heterologous expression system, we showed recently that CWPO-C contributes to differentiation or early growth and is involved in auxin catabolism. In this study, to clarify the function of CWPO-C in poplar, we analyzed CWPO-C gene expression and phenotypic changes with CWPO-C overexpression and suppression. Real-time PCR and monitoring promoter activity of CWPO-C using β-glucuronidase (GUS) assay revealed that CWPO-C was strongly expressed in immature tissues, such as the upper stem, axillary buds, and young leaves, in addition to expression in developing xylem. In transgenic poplars in which the expression of CWPO-C was upregulated or suppressed, changes in stem growth, gravitropism bending time, lignin content and syringyl/guaiacyl (S/G) composition were observed. Overexpressing CWPO-C enhanced stem growth and gravitropic response (shorter bending time). With suppressed CWPO-C expression, the lignin content was reduced approximately 45% and the S/G ratio decreased by half. These results strongly suggest that CWPO-C plays a role in differentiation and early growth, as well as in lignin polymerization. [ABSTRACT FROM AUTHOR]

Biofuels produced from lignocellulosic biomass provide energy security and reduce greenhouse gas emissions with positive impacts on sustainability. Lignocellulose byproducts are expected to impact the production of cellulosic ethanol. However, lignin is a major factor imposing biomass recalcitrance to saccharification. Although structural phenolics can be removed from lignocellulosic biomass, physical and chemical pretreatments are expensive. In turn, the structure and composition of lignin is very flexible. Up- and downregulation of genes and, thereby, enzymes involved in core and non-core lignin biosynthesis can reduce lignin content and improve biomass saccharification. Alternatively, insertion of heterologous genes to the phenylpropanoid pathway may lead to the incorporation of alternative monomers into lignin, creating desirable properties and preserving the biological roles of lignin with a positive effect on saccharification. Studies reporting the effect of fine regulation on lignin structures are mapping how plants can be transformed to enhance saccharification while preserving productivity. Based on these findings and events, this review updates the state-of-art changes in lignin biosynthesis to improve saccharification efficiency. The main findings reported over the last decade were summarized to permit researchers an overview of this relevant scientific subject. [ABSTRACT FROM AUTHOR]

As the most abundant aromatic renewable polymer in nature, lignin has been widely utilized in various fields such as industry, agriculture, medicine, and environmental protection, making it an important biomass resource. The characterization of lignin's structural units and their distribution in the cell wall is crucial for its isolation. Histochemical staining, a traditional method for detecting lignin, can rapidly and visually analyze important information such as the ratio of lignin structural units and their distribution in the cell wall. This paper provides a review of the color and fluorescent reactions in lignin histochemical staining. The staining mechanisms of each reaction and their applications in lignin detection are elucidated separately. In particular, in-depth discussions are conducted on the synergistic application of staining reactions with instrumental analytical methods. The paper concludes with an overview of the challenges and prospects for future developments in this research field. [Display omitted] • Histochemical staining for the detection of lignin structural units and their distribution. • Mechanism and application of the Wiesner, Mäule, Cross and Bevan Color reactions. • Mechanism and application of lignin autofluorescence. • Mechanism and application of Safranine O fluorescent dye. [ABSTRACT FROM AUTHOR]

Eucalyptus globulus Labill is an important raw material for pulpwood for which low lignin content and high S/G ratios are important parameters. This work shows a highly significant positive and strong correlation between S/G ratio and acid-soluble lignin (ASL) content among E. globulus trees. Although significant, the negative correlation between the S/G ratio and Klason lignin is weak, and there is no correlation with total lignin content. The Klason residue has a lower S/G ratio than that of the original wood. This suggests that the contribution of the S units to the solubilisation of lignin in acidic conditions is not limited to the S units but also to the G units. This effect becomes more pronounced as the percentage of S units increases. [ABSTRACT FROM AUTHOR]

Abstract Background Multiple analytical methods have been developed to determine the ratios of aromatic lignin units, particularly the syringyl/guaiacyl (S/G) ratio, of lignin biopolymers in plant cell walls. Chemical degradation methods such as thioacidolysis produce aromatic lignin units that are released from certain linkages and may induce chemical changes rendering it difficult to distinguish and determine the source of specific aromatic lignin units released, as is the case with nitrobenzene oxidation methodology. NMR methods provide powerful tools used to analyze cell walls for lignin composition and linkage information. Pyrolysis-mass spectrometry methods are also widely used, particularly as high-throughput methodologies. However, the different techniques used to analyze aromatic lignin unit ratios frequently yield different results within and across particular studies, making it difficult to interpret and compare results. This also makes it difficult to obtain meaningful insights relating these measurements to other characteristics of plant cell walls that may impact biomass sustainability and conversion metrics for the production of bio-derived fuels and chemicals. Results The authors compared the S/G lignin unit ratios obtained from thioacidolysis, pyrolysis-molecular beam mass spectrometry (py-MBMS), HSQC liquid-state NMR and solid-state (ss) NMR methodologies of pine, several genotypes of poplar, and corn stover biomass. An underutilized approach to deconvolute ssNMR spectra was implemented to derive S/G ratios. The S/G ratios obtained for the samples did not agree across the different methods, but trends were similar with the most agreement among the py-MBMS, HSQC NMR and deconvoluted ssNMR methods. The relationship between S/G, thioacidolysis yields, and linkage analysis determined by HSQC is also addressed. Conclusions This work demonstrates that different methods using chemical, thermal, and non-destructive NMR techniques to determine native lignin S/G ratios in plant cell walls may yield different results depending on species and linkage abundances. Spectral deconvolution can be applied to many hardwoods with lignin dominated by S and G units, but the results may not be reliable for some woody and grassy species of more diverse lignin composition. HSQC may be a better method for analyzing lignin in those species given the wealth of information provided on additional aromatic moieties and bond linkages. Additionally, trends or correlations in lignin characteristics such as S/G ratios and lignin linkages within the same species such as poplar may not necessarily exhibit the same trends or correlations made across different biomass types. Careful consideration is required when choosing a method to measure S/G ratios and the benefits and shortcomings of each method discussed here are summarized.

Abstract Background Understanding lignin biosynthesis and composition is of central importance for sustainable bioenergy and biomaterials production. Species of the genus Miscanthus have emerged as promising bioenergy crop due to their rapid growth and modest nutrient requirements. However, lignin polymerization in Miscanthus is poorly understood. It was previously shown that plant laccases are phenol oxidases that have multiple functions in plant, one of which is the polymerization of monolignols. Herein, we link a newly discovered Miscanthus laccase, MsLAC1, to cell wall lignification. Characterization of recombinant MsLAC1 and Arabidopsis transgenic plants expressing MsLAC1 were carried out to understand the function of MsLAC1 both in vitro and in vivo. Results Using a comprehensive suite of molecular, biochemical and histochemical analyses, we show that MsLAC1 localizes to cell walls and identify Miscanthus transcription factors capable of regulating MsLAC1 expression. In addition, MsLAC1 complements the Arabidopsis lac4–2 lac17 mutant and recombinant MsLAC1 is able to oxidize monolignol in vitro. Transgenic Arabidopsis plants over-expressing MsLAC1 show higher G-lignin content, although recombinant MsLAC1 seemed to prefer sinapyl alcohol as substrate. Conclusions In summary, our results suggest that MsLAC1 is regulated by secondary cell wall MYB transcription factors and is involved in lignification of xylem fibers. This report identifies MsLAC1 as a promising breeding target in Miscanthus for biofuel and biomaterial applications.

Abstract Background Plant secondary cell wall is a renewable feedstock for biofuels and biomaterials production. Arabidopsis VASCULAR-RELATED NAC DOMAIN (VND) has been demonstrated to be a key transcription factor regulating secondary cell wall biosynthesis. However, less is known about its role in the woody species. Results Here we report the functional characterization of Populus deltoides WOOD-ASSOCIATED NAC DOMAIN protein 3 (PdWND3A), a sequence homolog of Arabidopsis VND4 and VND5 that are members of transcription factor networks regulating secondary cell wall biosynthesis. PdWND3A was expressed at higher level in the xylem than in other tissues. The stem tissues of transgenic P. deltoides overexpressing PdWND3A (OXPdWND3A) contained more vessel cells than that of wild-type plants. Furthermore, lignin content and lignin monomer syringyl and guaiacyl (S/G) ratio were higher in OXPdWND3A transgenic plants than in wild-type plants. Consistent with these observations, the expression of FERULATE 5-HYDROXYLASE1 (F5H1), encoding an enzyme involved in the biosynthesis of sinapyl alcohol (S unit monolignol), was elevated in OXPdWND3A transgenic plants. Saccharification analysis indicated that the rate of sugar release was reduced in the transgenic plants. In addition, OXPdWND3A transgenic plants produced lower amounts of biomass than wild-type plants. Conclusions PdWND3A affects lignin biosynthesis and composition and negatively impacts sugar release and biomass production.

Key message: A transcriptome analysis reveals the transcripts and alleles differentially expressed in sugarcane genotypes with contrasting lignin composition. Sugarcane bagasse is a highly abundant resource that may be used as a feedstock for the production of biofuels and bioproducts in order to meet increasing demands for renewable replacements for fossil carbon. However, lignin imparts rigidity to the cell wall that impedes the efficient breakdown of the biomass into fermentable sugars. Altering the ratio of the lignin units, syringyl (S) and guaiacyl (G), which comprise the native lignin polymer in sugarcane, may facilitate the processing of bagasse. This study aimed to identify genes and markers associated with S/G ratio in order to accelerate the development of sugarcane bioenergy varieties with modified lignin composition. The transcriptome sequences of 12 sugarcane genotypes that contrasted for S/G ratio were compared and there were 2019 transcripts identified as differentially expressed (DE) between the high and low S/G ratio groups. These included transcripts encoding possible monolignol biosynthetic pathway enzymes, transporters, dirigent proteins and transcriptional and post-translational regulators. Furthermore, the frequencies of single nucleotide polymorphisms (SNPs) were compared between the low and high S/G ratio groups to identify specific alleles expressed with the phenotype. There were 2063 SNP loci across 787 unique transcripts that showed group-specific expression. Overall, the DE transcripts and SNP alleles identified in this study may be valuable for breeding sugarcane varieties with altered S/G ratio that may provide desirable bioenergy traits. [ABSTRACT FROM AUTHOR]

Abstract Background Lignin is known to hinder efficient enzymatic conversion of lignocellulose in biorefining processes. In particular, nonproductive adsorption of cellulases onto lignin is considered a key mechanism to explain how lignin retards enzymatic cellulose conversion in extended reactions. Results Lignin-rich residues (LRRs) were prepared via extensive enzymatic cellulose degradation of corn stover (Zea mays subsp. mays L.), Miscanthus × giganteus stalks (MS) and wheat straw (Triticum aestivum L.) (WS) samples that each had been hydrothermally pretreated at three severity factors (log R 0) of 3.65, 3.83 and 3.97. The LRRs had different residual carbohydrate levels—the highest in MS; the lowest in WS. The residual carbohydrate was not traceable at the surface of the LRRs particles by ATR-FTIR analysis. The chemical properties of the lignin in the LRRs varied across the three types of biomass, but monolignols composition was not affected by the severity factor. When pure cellulose was added to a mixture of LRRs and a commercial cellulolytic enzyme preparation, the rate and extent of glucose release were unaffected by the presence of LRRs regardless of biomass type and severity factor, despite adsorption of the enzymes to the LRRs. Since the surface of the LRRs particles were covered by lignin, the data suggest that the retardation of enzymatic cellulose degradation during extended reaction on lignocellulosic substrates is due to physical blockage of the access of enzymes to the cellulose caused by the gradual accumulation of lignin at the surface of the biomass particles rather than by nonproductive enzyme adsorption. Conclusions The study suggests that lignin from hydrothermally pretreated grass biomass retards enzymatic cellulose degradation by acting as a physical barrier blocking the access of enzymes to cellulose rather than by inducing retardation through nonproductive adsorption of enzymes.

Background: Multiple analytical methods have been developed to determine the ratios of aromatic lignin units, particularly the syringyl/guaiacyl (S/G) ratio, of lignin biopolymers in plant cell walls. Chemical degradation methods such as thioacidolysis produce aromatic lignin units that are released from certain linkages and may induce chemical changes rendering it difficult to distinguish and determine the source of specific aromatic lignin units released, as is the case with nitrobenzene oxidation methodology. NMR methods provide powerful tools used to analyze cell walls for lignin composition and linkage information. Pyrolysis-mass spectrometry methods are also widely used, particularly as high-throughput methodologies. However, the different techniques used to analyze aromatic lignin unit ratios frequently yield different results within and across particular studies, making it difficult to interpret and compare results. This also makes it difficult to obtain meaningful insights relating these measurements to other characteristics of plant cell walls that may impact biomass sustainability and conversion metrics for the production of bio-derived fuels and chemicals. Results: The authors compared the S/G lignin unit ratios obtained from thioacidolysis, pyrolysis-molecular beam mass spectrometry (py-MBMS), HSQC liquid-state NMR and solid-state (ss) NMR methodologies of pine, several genotypes of poplar, and corn stover biomass. An underutilized approach to deconvolute ssNMR spectra was implemented to derive S/G ratios. The S/G ratios obtained for the samples did not agree across the different methods, but trends were similar with the most agreement among the py-MBMS, HSQC NMR and deconvoluted ssNMR methods. The relationship between S/G, thioacidolysis yields, and linkage analysis determined by HSQC is also addressed. Conclusions: This work demonstrates that different methods using chemical, thermal, and non-destructive NMR techniques to determine native lignin S/G ratios in plant cell walls may yield different results depending on species and linkage abundances. Spectral deconvolution can be applied to many hardwoods with lignin dominated by S and G units, but the results may not be reliable for some woody and grassy species of more diverse lignin composition. HSQC may be a better method for analyzing lignin in those species given the wealth of information provided on additional aromatic moieties and bond linkages. Additionally, trends or correlations in lignin characteristics such as S/G ratios and lignin linkages within the same species such as poplar may not necessarily exhibit the same trends or correlations made across different biomass types. Careful consideration is required when choosing a method to measure S/G ratios and the benefits and shortcomings of each method discussed here are summarized. [ABSTRACT FROM AUTHOR]

Key message: The HbCAld5H1 gene cloned from Hevea brasiliensis regulates the cambial activity, xylem differentiation, syringyl–guaiacyl ratio, secondary wall structure, lignification pattern and xylan distribution in xylem fibres of transgenic tobacco plants. Molecular characterization of lignin biosynthesis gene coniferaldehyde-5-hydroxylase (CAld5H) from Hevea brasiliensis and its functional validation was performed. Both sense and antisense constructs of HbCAld5H1 gene were introduced into tobacco through Agrobacterium-mediated genetic transformation for over expression and down-regulation of this key enzyme to understand its role affecting structural and cell wall chemistry. The anatomical studies of transgenic tobacco plants revealed the increase of cambial activity leading to xylogenesis in sense lines and considerable reduction in antisense lines. The ultra-structural studies showed that the thickness of secondary wall (S2 layer) of fibre had been decreased with non-homogenous lignin distribution in antisense lines, while sense lines showed an increase in S2 layer thickness. Maule color reaction revealed that syringyl lignin distribution in the xylem elements was increased in sense and decreased in antisense lines. The immunoelectron microscopy revealed a reduction in LM 10 and LM 11 labelling in the secondary wall of antisense tobacco lines. Biochemical studies showed a radical increase in syringyl lignin in sense lines without any significant change in total lignin content, while S/G ratio decreased considerably in antisense lines. Our results suggest that CAld5H gene plays an important role in xylogenesis stages such as cambial cell division, secondary wall thickness, xylan and syringyl lignin distribution in tobacco. Therefore, CAld5H gene could be considered as a promising target for lignin modification essential for timber quality improvement in rubber. [ABSTRACT FROM AUTHOR]

Sustainability of cork oak (Quercus suber) forests is threatened by biotic and abiotic factors and characterization of potentially differing genetic resources has therefore gained importance. This work addresses the chemical variation of the three tissues of cork oak stems – cork, phloem and wood – in relation to tree and provenance, looking for genetic chemical diversity and for physiological derived differences. The three tissues differ with cork clearly differentiating regarding summative composition, component ratios and monomeric composition. Cork is the only tissue where suberin is present (42.3% o.d. mass) as the main cell wall component, and it has a high content of extractives (11.7%) with significant proportion of lipophilic compounds. Phloem is more lignified than wood (38.0% vs. 23.4%) and has less polysaccharides (49.1% vs. 64.6%) with glucose-to-other sugars relation of 1:1.3 in phloem and 1:0.7 in wood. Analytical pyrolysis showed that lignification is a heterogeneous process and the lignin monomeric composition depends on tissue and cell type: cork lignin has a H:G:S ratio of 1:2.5:0.3 and S/G ratio of 0.12, while phloem and wood lignins have mainly G and S units with a S/G ratio of respectively 1.1 and 2.3. No significant differences were found between the three provenances, but some chemical variation occurred between the trees within a provenance. NIR spectroscopy and principal component analysis differentiated cork, phloem and wood, while the dispersion within each group highlighted the significant tree variability, while provenances were a non-significant factor of chemical variation.

Background: Understanding lignin biosynthesis and composition is of central importance for sustainable bioenergy and biomaterials production. Species of the genus Miscanthus have emerged as promising bioenergy crop due to their rapid growth and modest nutrient requirements. However, lignin polymerization in Miscanthus is poorly understood. It was previously shown that plant laccases are phenol oxidases that have multiple functions in plant, one of which is the polymerization of monolignols. Herein, we link a newly discovered Miscanthus laccase, MsLAC1, to cell wall lignification. Characterization of recombinant MsLAC1 and Arabidopsis transgenic plants expressing MsLAC1 were carried out to understand the function of MsLAC1 both in vitro and in vivo. Results: Using a comprehensive suite of molecular, biochemical and histochemical analyses, we show that MsLAC1 localizes to cell walls and identify Miscanthus transcription factors capable of regulating MsLAC1 expression. In addition, MsLAC1 complements the Arabidopsis lac4–2 lac17 mutant and recombinant MsLAC1 is able to oxidize monolignol in vitro. Transgenic Arabidopsis plants over-expressing MsLAC1 show higher G-lignin content, although recombinant MsLAC1 seemed to prefer sinapyl alcohol as substrate. Conclusions: In summary, our results suggest that MsLAC1 is regulated by secondary cell wall MYB transcription factors and is involved in lignification of xylem fibers. This report identifies MsLAC1 as a promising breeding target in Miscanthus for biofuel and biomaterial applications. [ABSTRACT FROM AUTHOR]

Background: Plant secondary cell wall is a renewable feedstock for biofuels and biomaterials production. Arabidopsis VASCULAR-RELATED NAC DOMAIN (VND) has been demonstrated to be a key transcription factor regulating secondary cell wall biosynthesis. However, less is known about its role in the woody species. Results: Here we report the functional characterization of Populus deltoides WOOD-ASSOCIATED NAC DOMAIN protein 3 (PdWND3A), a sequence homolog of Arabidopsis VND4 and VND5 that are members of transcription factor networks regulating secondary cell wall biosynthesis. PdWND3A was expressed at higher level in the xylem than in other tissues. The stem tissues of transgenic P. deltoides overexpressing PdWND3A (OXPdWND3A) contained more vessel cells than that of wild-type plants. Furthermore, lignin content and lignin monomer syringyl and guaiacyl (S/G) ratio were higher in OXPdWND3A transgenic plants than in wild-type plants. Consistent with these observations, the expression of FERULATE 5-HYDROXYLASE1 (F5H1), encoding an enzyme involved in the biosynthesis of sinapyl alcohol (S unit monolignol), was elevated in OXPdWND3A transgenic plants. Saccharification analysis indicated that the rate of sugar release was reduced in the transgenic plants. In addition, OXPdWND3A transgenic plants produced lower amounts of biomass than wild-type plants. Conclusions: PdWND3A affects lignin biosynthesis and composition and negatively impacts sugar release and biomass production. [ABSTRACT FROM AUTHOR]

Fourier transform infrared spectroscopy (FTIR) is a simple nondestructive technique that allows the user to obtain quick and accurate information about the structure of the constituents of wood. Spectra deconvolution is a computational technique, complementary to FTIR analysis, which improves the resolution of overlapped or unobserved bands in the raw spectra. High performance liquid chromatography (HPLC) is an analytical technique useful to determine the ratio of the lignin monomers obtained by the alkaline nitrobenzene oxidation method. Furthermore, lignin content has been commonly determined by wet chemical methods; Klason lignin determination is a quick and accessible method. Here, we detail the procedures for chemical analysis of the wood lignin using these techniques. Additionally, the deconvolution process of FTIR spectra for the determination of the S/G ratio, in lignin isolated by this or other methods, is explained in detail., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Summary: The aromatic composition of lignin is an important trait that greatly affects the usability of lignocellulosic biomass. We previously identified a rice (Oryza sativa) gene encoding coniferaldehyde 5‐hydroxylase (OsCAld5H1), which was effective in modulating syringyl (S)/guaiacyl (G) lignin composition ratio in rice, a model grass species. Previously characterized OsCAld5H1‐knockdown rice lines, which were produced via an RNA‐interference approach, showed augmented G lignin units yet contained considerable amounts of residual S lignin units. In this study, to further investigate the effect of suppression of OsCAld5H1 on rice lignin structure, we generated loss‐of‐function mutants of OsCAld5H1 using the CRISPR/Cas9‐mediated genome editing system. Homozygous OsCAld5H1‐knockout lines harboring anticipated frame‐shift mutations in OsCAld5H1 were successfully obtained. A series of wet‐chemical and two‐dimensional NMR analyses on cell walls demonstrated that although lignins in the mutant were predictably enriched in G units all the tested mutant lines produced considerable numbers of S units. Intriguingly, lignin γ‐p‐coumaroylation analysis by the derivatization followed by reductive cleavage method revealed that enrichment of G units in lignins of the mutants was limited to the non‐γ‐p‐coumaroylated units, whereas grass‐specific γ‐p‐coumaroylated lignin units were almost unaffected. Gene expression analysis indicated that no homologous genes of OsCAld5H1 were overexpressed in the mutants. These data suggested that CAld5H is mainly involved in the production of non‐γ‐p‐coumaroylated S lignin units, common in both eudicots and grasses, but not in the production of grass‐specific γ‐p‐coumaroylated S units in rice. Significance Statement: CRISPR/Cas9‐mediated gene knockout of rice coniferaldehyde 5‐hydroxylase (OsCAld5H1) resulted in lignins enriched in guaiacyl (G) units. However, the mutants still produced considerable numbers of syringyl (S) units. The enrichment of G units was limited to the non‐γ‐p‐coumaroylated lignin units, and the grass‐specific γ‐p‐coumaroylated lignin units were apparently unchanged, suggesting that OsCAld5H1 is mainly involved in the biosynthesis of non‐γ‐p‐coumaroylated monolignols common in both eudicots and grasses, but not in that of grass‐specific γ‐p‐coumaroylated monolignols. [ABSTRACT FROM AUTHOR]

In spite of the increasing number of myxomycete studies in the tropical Southeast Asia over the last decades, many forest patches are still left unexplored for the region, in particular in the Philippine archipelago. Thus, an assessment of myxomycete diversity, occurrence, and composition was carried out in forest fragments from two municipalities of the province of Laguna, Philippines. From the 12 established 5 m x 5 m plots, a total of 240 moist chamber cultures were prepared for this study and yielded 42 myxomycete species belonging to 14 genera. Rarefaction curves and several heterogeneity indices revealed higher species diversity in Los Baños than in Calauan. Moreover, the taxonomic diversity index also showed that the taxonomic diversity of myxomycete in Los Baños is more intuitively diverse than in Calauan. In terms of community analysis between the two municipalities, it showed that myxomycete communities clustered between different microhabitats. High values of coeffi- cient of community and percentage similarity indices possibly implicates that spore dispersal in forest fragments in closer proximities may cause high number of shared species (60%). With an addition of Cribraria lepida as a new record for the country, this study now updates the myxomycete profile for Laguna, Philippines. [ABSTRACT FROM AUTHOR]

Abstract Plants are occasionally exposed to environmental perturbations that limit their growth. One of these perturbations is the exposure to and interaction with various nanoparticles (NPs) that are discarded continuously into the environment. Hitherto, no study has been carried out evaluating the effects of iron oxide (γ-Fe 2 O 3) NPs on soybean growth and lignin formation, as proposed herein. For comparative purposes, we also submitted soybean plants to non-nanoparticulate iron (FeCl 3). Exposure of the plants to γ-Fe 2 O 3 NPs increased cell wall-bound peroxidase (POD) activity but decreased phenylalanine ammonia lyase (PAL) activity due, probably, to the negative feedback of accumulated phenolic compounds. In contrast, FeCl 3 decreased cell wall-bound POD activity. Both γ-Fe 2 O 3 NPs and FeCl 3 increased the lignin content of roots and stems. However, significant lignin-induced growth inhibition was noted only in stems after exposure to NPs, possibly due to changes in lignin monomer composition. In this case, γ-Fe 2 O 3 NPs decreased the guaiacyl monomer content of roots but increased that of stems. The high levels of monomer guaiacyl in stems resulting from the action of γ-Fe 2 O 3 NPs decreased syringyl/guaiacyl ratios, generating more highly cross-linked lignin followed by the stiffening of the cell wall and growth inhibition. In contrast, FeCl 3 increased the contents of monomers p- hydroxyphenyl and syringyl in roots. The observed increase in the syringyl/guaiacyl ratio in plant roots submitted to FeCl 3 agrees with the lack of effect on growth, due to the formation of a less condensed lignin. In brief, we here describe that γ-Fe 2 O 3 NPs and FeCl 3 act differently in soybean plants. Graphical abstract Image 1 Highlights • γ-Fe 2 O 3 NPs and ionic iron (FeCl 3) stimulated lignin formation in roots and stems. • Lignin-induced growth inhibition was noted only in stems of plants exposed to NPs. • Inhibition of stem growth by NPs is due to changes in lignin monomer composition. • FeCl 3 and γ-Fe 2 O 3 NPs act distinctly on soybean lignin metabolism. [ABSTRACT FROM AUTHOR]

Basal Stem rot (BSR) disease of oil palm caused by Ganoderma boninense is one of the major obstacle in oil palm production. The root contact with unattended infected debris left in the plantations during replanting is known to be the primary source of infection spread. Abiding with the law, potentially effective technique of managing Ganoderma infected oil palm debris is deemed necessary because of the zero-burning policy in Malaysian oil palm plantations. The wood degradation abilities of four white-rot hymenomycetes were assessed using oil palm blocks obtained from healthy and diseased portions of oil palm trunks infected with G. boninens e. In general, out of four white-rot hymenomycetes, Pycnoporus sanguineus FBR secreted a significantly ( P ≤ 0.05) greater quantity of oxidative enzymes than that secreted by the Grammothele fuligo strains. The ability of G. fuligo ST2 and P. sanguineus FBR to secrete amylase enzymes has been well documented and the enzyme profiles were about 4.5-fold ( G. fuligo ST2) and 2.8-fold ( P. sanguineus FBR) higher in samples collected at day 120 than at day 30 of biodegradation, supported with Fourier transform infrared (FTIR) spectra. The crystallinity ratio was found to increase in the middle of decay process, owing to the preferential degradation of non-crystalline lignins and hemicelluloses by both strains. Syringyl (S)-lignin units were favorably degraded than guaiacyl (G)-lignin units by both strains, resulting in a decreased S/G ratio in the diseased portions, at the end of decay process. Scanning electron microscopy (SEM) revealed that penetration of fungal hyphae within the wood vessels and the parenchymal tissues with boreholes, resulting in the loosening of the parenchymal tissue and wood rays. Based on the lignocellulolytic enzymes production and the level of degradation, P. sanguineus FBR and G. fuligo ST2 regarded as the best bio-degraders of G. boninense colonized wood blocks. This study provides the novel insight on understanding the biochemical and anatomical changes induced by white-rot hymenomycetes during biodegradation of oil palm wood blocks with an ultimate aim of reducing the G anoderma inoculum under heavy BSR infection pressure in ecofriendly manner. [ABSTRACT FROM AUTHOR]

Main conclusion : Regulation of a gene encoding coniferaldehyde 5-hydroxylase leads to substantial alterations in lignin structure in rice cell walls, identifying a promising genetic engineering target for improving grass biomass utilization. The aromatic composition of lignin greatly affects utilization characteristics of lignocellulosic biomass and, therefore, has been one of the primary targets of cell wall engineering studies. Limited information is, however, available regarding lignin modifications in monocotyledonous grasses, despite the fact that grass lignocelluloses have a great potential for feedstocks of biofuel production and various biorefinery applications. Here, we report that manipulation of a gene encoding coniferaldehyde 5-hydroxylase (CAld5H, or ferulate 5-hydroxylase, F5H) leads to substantial alterations in syringyl (S)/guaiacyl (G) lignin aromatic composition in rice ( Oryza sativa), a major model grass and commercially important crop. Among three CAld5H genes identified in rice, OsCAld5H1 ( CYP84A5) appeared to be predominantly expressed in lignin-producing rice vegetative tissues. Down-regulation of OsCAld5H1 produced altered lignins largely enriched in G units, whereas up-regulation of OsCAld5H1 resulted in lignins enriched in S units, as revealed by a series of wet-chemical and NMR structural analyses. Our data collectively demonstrate that OsCAld5H1 expression is a major factor controlling S/G lignin composition in rice cell walls. Given that S/G lignin composition affects various biomass properties, we contemplate that manipulation of CAld5H gene expression represents a promising strategy to upgrade grass biomass for biorefinery applications. [ABSTRACT FROM AUTHOR]

Selected and tested poplar clones are very suitable biomass resources for various applications such as biofuels, the pulp and paper industry as well as chemicals production. In this study, we determined the content of lignin, cellulose, holocellulose, and extractives, syringyl to guaiacyl (S/G) ratio in lignin, and also calculated higher heating values (HHV) among eight examined clones of Populus grown on three different experimental sites. The highest lignin content for all the examined sites was determined in ‘I-214’ and ‘Baka 5’ clones, whereas the highest content of extractives was found in ‘Villafranca’ and ‘Baka 5’ clones. The highest S/G ratio for all the examined sites was determined in ‘Villafranca’ and ‘Agathe F’ clones. The chemical profiles of main wood components, extractives, and the S/G ratio in lignin were also influenced by both the experimental site and the clone × site interaction. Higher heating values, derived from calculations based on the contents of lignin and extractives (or lignin only), were in close agreement with the previously published data. The highest heating values were found for ‘Baka 5’ and ‘I-214’ clones. The optimal method of poplar biomass utilization can be chosen on basis of the lignocellulosics chemical composition and the S/G ratio in lignin.

Pyrolysis-gas chromatography/mass spectrometry (Py-GC-MS) was applied to measure the lignin syringyl/guaiacyl (S/G) ratio in E. dunni, E. grandis, E. nitens, E. urograndis, and E. urophylla woods. A total of 41 compounds were identified, of which 11 were derived from carbo-hydrates and 30 from lignins. The S/G ratio was calculated on the basis of the areas of peaks recorded in the pyrograms and compared with the results obtained by alkaline nitrobenzene oxidation. The values of S/G found by pyrolysis were similar for all the species using the compounds guaiacol, 4-methylguaiacol, 4-vinylguaiacol, vanillin, 4-ethylsyringol, 4-vinylsyringol, homosyringaldehyde, acetosyringone, and syringylacetone, as lignin markers. The selected markers were efficient for the deter-mination of S/G ratio in eucalyptus wood by Py-GC-MS. The Py-GC-MS technique produced results that are comparable to the nitrobenzene oxidation method, with the advantage of requiring small wood samples and a short analysis time.

The use of analytical pyrolysis combined with gas chromatography/mass spectrometry (Py-GC/MS) to determine the syringyl/guaiacyl ratio (S/G) in lignins from Eucalyptus spp woods was investigated. Sample of E. grandis and "E. urograndis" wood, with and without extractives, were subjected to pyrolysis from 300 ºC to 600 ºC. The products that results from pyrolysis were identified by mass spectrometry and the S/G ratio was determined based on the areas of the peaks corresponding to the guaiacyl and syringyl derivatives. The best S/G estimation is achieved when pyrolysis is carried out at 550 ºC. Extractives and carbohydrate present in the woods do not interfere with the results.

To explore the potential values of lignin from agricultural residues (apple tree pruning, olive tree pruning and almond shell) and industrial residues (kraft black liquor) were used as raw materials for lignin extraction by different fractionation processes (kraft, organosolv, acetosolv and acetosolv and formosolv processes). The isolated lignins were characterized by Py-GC/MS, FTIR and GPC. The fractionation process influenced greatly the average molecular weight (Mw) that was measured by GPC. The harsh conditions during the acetosolv and acetosolv-formosolv processes promoted the repolymerization phenomena giving high Mw lignins. The EKL presented lower Mw due to higher retention times. All lignins exhibit higher relative abundances of S-type of phenols except for the almond shell lignin where G-type of phenols were the most abundant ones. The S/G ratios for hardwood type of lignin were analogous which ranged from 2.30 to 2.71, whereas almond shells lignin revealed an S/G ratio of 0.94. Kraft liquid fraction showed the highest lignin content (6.39%) and consequently the highest S/G ratio (2.71). [ABSTRACT FROM AUTHOR]

Background: Higher ratios of syringyl-to-guaiacyl (S/G) lignin components of Populus were shown to improve sugar release by enzymatic hydrolysis using commercial blends. Cellulolytic microbes are often robust biomass hydrolyzers and may offer cost advantages; however, it is unknown whether their activity can also be significantly influenced by the ratio of different monolignol types in Populus biomass. Hydrolysis and fermentation of autoclaved, but otherwise not pretreated Populus trichocarpa by Clostridium thermocellum ATCC 27405 was compared using feedstocks that had similar carbohydrate and total lignin contents but differed in S/G ratios. Results: Populus with an S/G ratio of 2.1 was converted more rapidly and to a greater extent compared to similar biomass that had a ratio of 1.2. For either microbes or commercial enzymes, an approximate 50 % relative difference in total solids solubilization was measured for both biomasses, which suggests that the differences and limitations in the microbial breakdown of lignocellulose may be largely from the enzymatic hydrolytic process. Surprisingly, the reduction in glucan content per gram solid in the residual microbially processed biomass was similar (17-18 %) irrespective of S/G ratio, pointing to a similar mechanism of solubilization that proceeded at different rates. Fermentation metabolome testing did not reveal the release of known biomass-derived alcohol and aldehyde inhibitors that could explain observed differences in microbial hydrolytic activity. Biomass-derived p-hydroxybenzoic acid was up to ninefold higher in low S/G ratio biomass fermentations, but was not found to be inhibitory in subsequent test fermentations. Cellulose crystallinity and degree of polymerization did not vary between Populus lines and had minor changes after fermentation. However, lignin molecular weights and cellulose accessibility determined by Simons' staining were positively correlated to the S/G content. Conclusions: Higher S/G ratios in Populus biomass lead to longer and more linear lignin chains and greater access to surface cellulosic content by microbe-bound enzymatic complexes. Substrate access limitation is suggested as a primary bottleneck in solubilization of minimally processed Populus, which has important implications for microbial deconstruction of lignocellulose biomass. Our findings will allow others to examine different Populus lines and to test if similar observations are possible for other plant species. [ABSTRACT FROM AUTHOR]

Milled wood lignins from alkaline pretreated wood with very low sugar content and a wide range of syringyl-to-guaiacyl (S/G) ratio between 1.2 and 3.0 were isolated from 12 industrially valuable hardwood (HW) species. The lignin preparations were investigated by means of a comprehensive 13C nuclear magnetic resonance (NMR) methodology to address the possibilities and limitations of this approach for HW native lignins and to estimate the structural variations within HW lignins. Good correlations were found for different independent methods for the quantification of major lignin moieties. The results were reliable at the C6 level and not only for relative comparison. The correlation was good between methoxyl group determinations by wet chemistry and those by 13C NMR spectroscopy. The limitations of the 13C NMR method were also pointed out. The differences in the S/G ratios can be large, but other structural deviations are less significant. Strong correlations between the S/G ratios and the amounts of other structural peculiarities could not be found by the 13C NMR approach. However, with increasing S/G ratios, the β-O-4 content showed increasing tendencies and the degree of condensation showed decreasing tendencies. [ABSTRACT FROM AUTHOR]

The lignin from Cynara cardunculus stalks was isolated by the classical Björkman method and characterized by pyrolysis coupled with gas chromatography and mass spectrometry (Py-GC/MS), two-dimensional nuclear magnetic resonance spectroscopy (2D-NMR), and derivatization followed by reductive cleavage (DFRC). The milled Cynara lignin (MCyL) was constituted mainly by guaiacyl (G) and syringyl-units(S) (S/G molar ratio of 0.7), with the complete absence of p-hydroxyphenyl (H) units. The 2D-NMR analysis indicated a predominance of alkyl-aryl ether linkages (70 % of all inter unit linkages are β- O-4′) and significant amounts of condensed structures such as phenylcoumarans (β-5′, 14 %), resinols (β-β′, 7 %), spirodienones (β-1′, 5 %), and dibenzodioxocins (5-5′, 4 %). Furthermore, the analyses indicated that the lignin is partially acylated at the γ-OH (12 % acylation) by acetate groups and that acetylation occurs preferentially on syringyl-units. As in other plants, acetylation occurs at the monomer stage, and sinapyl acetate behaves as a real lignin monomer participating in lignification in cardoon stalks. The detailed structural characterization of cardoon lignin reported here will foster the industrial use of this biomass for the production of biofuels and other bio-based chemicals under the lignocellulosic biorefinery. [ABSTRACT FROM AUTHOR]

A simple extraction protocol and an ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS) method for the determination of the syringyl/guaiacyl (S/G) ratio in lignin is reported herein. The method was entirely developed using stems of three Eucalyptus species, which were hydrolyzed with NaOH and partitioned with ethyl ether; vanillin (from the G monomer) and syringaldehyde (from S monomer) were quantified. The S/G ratios obtained were comparable to those usually reported for eucalyptus. The data for one of the eucalyptus species were compared with those obtained with a widely accepted method using thioacidolysis and gas chromatography-mass spectrometry (GC-MS). The method was also applied to sugarcane and showed to be reliable. The yield of the NaOH hydrolysis of the monolignols ranged from 89.94 to 95.69 %, with more than 77.12 % of recuperation in the liquid-liquid extraction. The whole analytical procedure was validated, achieving results with less than 4.38 % of variation. The lowest LOD and LOQ were 0.01 and 0.05 μg/mL, respectively. In addition, the method combines reliability and a fast and direct quantification. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

Eucalypt feedstock was pretreated with Myceliophthora thermophila laccase and methyl syringate, as mediator, in a multistage sequence consisting of successive enzymatic and alkaline peroxide stages, directly on the ground wood. Lignin modification was studied by two-dimensional (2D) nuclear magnetic resonance (NMR) of wood (at the gel state) after each stage of the sequence. Decrease of guaiacyl and syringyl lignin units (with preferential removal of the former) and aliphatic (mainly β- O-4′-linked) side chains was observed along the sequence, without a substantial change in polysaccharide cross-signals along the sequence, was observed. However, the most noticeable modification was the formation of C-oxidized syringyl units in the enzymatic stages, which were partially removed in the alkaline extractions. Further insight was attained by 2D NMR of the cellulolytic-enzyme lignins, isolated from the pretreated samples, and the filtrates from the different stages. Additionally, a comparison of eucalypt delignification with the high redox potential laccase from Pycnoporus cinnabarinus in the presence of the synthetic mediator 1-hydroxybenzotriazole was carried out. Wood delignification already occurred at the first cycle of the pretreatment with P. cinnabarinus laccase and 1-hydroxybenzotriazole, which directly correlated with increases in glucose (~10 %) yields after enzymatic hydrolysis. In contrast, the first cycle of pretreatment with M. thermophila laccase and methyl syringate did not produce any delignification or saccharification improvement. However, at the end of the sequence, similar delignification (~50 %) of eucalypt feedstock and increases (~30 %) in glucose yields were attained in both laccase-mediator pretreatments. Finally, the influence of lignin content and the ratio of its syringyl and guaiacyl units on recalcitrance of eucalypt feedstock to enzymatic saccharification is discussed. [ABSTRACT FROM AUTHOR]

Pyrolysis coupled with gas chromatography and mass spectrometry (Py-GC/MS) was used for evaluating the syringyl/guaiacyl (S/G) ratios of residual lignins in oxygen pre-bleached kraft pulps from 24 Eucalyptus wood clones. The optimization of the method showed that the best conditions were achieved when pyrolysis is carried out at 550 °C, using a 60 m length SPB-1701 capillary column and the mass spectrometer operating in SIM mode. Pulp pyrograms were analyzed and the compounds produced by pyrolysis of residual lignins identified by mass spectrometry. A total of 18 compounds were identified, being nine derived from guaiacyl units and nine derived from syringyl lignin. The S/G ratios were calculated using the relative area of S-lignin (syringol, 4-methylsyringol, 4-vinylsyringol and trans -4-(prop-2-enyl) syringol) and G-lignin (guaiacol, 4-methylguaiacol, 4-vinylguaiacol and trans -isoeugenol) as lignin markers. These studies showed that Py-GC/MS-SIM is an efficient tool to characterizing the chemical composition of residual lignin in oxygen pre-bleached Eucalyptus pulps. [ABSTRACT FROM AUTHOR]

Detailed structural features of lignin from hydroxycinnamoyl transferase (HCT) down-regulated poplar has not been thoroughly illustrated. In the present study, MWL and CEL samples were successively extracted with higher yield and purity. NMR spectra demonstrated that the MWL and CEL presented similar structural features (β-O-4', β-β', β-5', and β-1'), although the subsequent CEL had a high β-O-4' linkages content and S/G ratios, which are also related to their molecular weights. There were no significant differences in S/G ratio and the distribution of the inter-unit linkages between the control and HCT down-regulated poplar (HCT poplar). However, a minor amount of H units were detected in lignin of the HCT poplar. In addition, p-hydroxybenzoate (PB) content was observably increased in the lignin from the HCT poplar. These findings are of very importance in assessing of the digestibility of HCT down-regulated poplar in the fuel production. [ABSTRACT FROM AUTHOR]

Background: Non-productive cellulase adsorption onto lignin has always been deemed to negatively affect the enzymatic hydrolysis of lignocellulosic feedstocks. Therefore, understanding enzyme-lignin interactions is essential for the development of enzyme mixtures, the processes of lignocellulose hydrolysis, and the genetic modification of lignocellulosic biomass and enzymes. In this work, we examined the properties of six lignins from diverse types of lignocellulosic biomass (aspen, pine, corn stover, kenaf, and two Arabidopsis lines, wild-type and SALK mutant of fah1) to determine the mechanism of differences in their adsorption of enzymes. Results: We found that lignin sources affected enzyme adsorption using structural features, such as functional groups and lignin composition. Guaiacyl (G) lignin had a higher adsorption capacity on enzymes than syringyl (S) lignin. The low S/G ratio and high uniform lignin fragment size had good correlations with high adsorption capacity. A higher content of phenolic hydroxyl groups and a lower content of carboxylic acid groups resulted in stronger adsorption affinity for corn stover lignin (CL) than for kenaf lignin (KL) and aspen lignin (AL). The lower amount of aliphatic hydroxyls that reduced hydrophobic interactions could explain the higher adsorption capacity of pine lignin (PL) than CL. Enzyme activity assays, as well as the hydrolysis of Avicel, phosphoric acid-swollen cellulose (PASC), and holocellulose, were performed to study the behaviors of mono-component enzymes that resulted in adsorption. We found that cellobiohydrolase (CBH) and xylanase were adsorbed the most by all lignins, endoglucanase (EG) showed less inhibition, and β-glucosidase (BG) was the least affected by lignins, indicating the important role of carbohydrate-binding module (CBM) in protein adsorption. Conclusion: Lignin sources affect enzyme adsorption using structural features and lignin composition, such as S/G ratio, carboxylic acid, aliphatic hydroxyl, and phenolic hydroxyl. For mono-component enzymes, the adsorption capacity decreased in the order CBH, xylanase > EG > BG. These investigations revealed the difference in lignin properties between diverse biomass and adsorption capacity of enzymes to lignins, and the possible underlying mechanism. The results can also serve as a reference for the genetic modification of lignocellulosic biomass and enzymes. [ABSTRACT FROM AUTHOR]

Genetic analysis of wood chemical composition is often limited by the cost and throughput of direct analytical methods. The speed and low cost of Fourier transform near infrared (FT-NIR) overcomes many of these limitations, but it is an indirect method relying on calibration models that are typically developed and validated with small sample sets. In this study, we used >1500 young greenhouse grown trees from a clonally propagated single Populus family, grown at low and high nitrogen, and compared FT-NIR calibration sample sizes of 150, 250, 500 and 750 on calibration and prediction model statistics, and heritability estimates developed with pyrolysis molecular beam mass spectrometry (pyMBMS) wood chemical composition. As calibration sample size increased from 150 to 750, predictive model statistics improved slightly. Overall, stronger calibration and prediction statistics were obtained with lignin, S-lignin, S/G ratio, and m/z 144 (an ion from cellulose), than with C5 and C6 carbohydrates, and m/z 114 (an ion from xylan). Although small differences in model statistics were observed between the 250 and 500 sample calibration sets, when predicted values were used for calculating genetic control, the 500 sample set gave substantially more similar results to those obtained with the pyMBMS data. With the 500 sample calibration models, genetic correlations obtained with FT-NIR and pyMBMS methods were similar. Quantitative trait loci (QTL) analysis with pyMBMS and FT-NIR predictions identified only three common loci for lignin traits. FT-NIR identified four QTLs that were not found with pyMBMS data, and these QTLs were for the less well predicted carbohydrate traits. [ABSTRACT FROM AUTHOR]

Abstract: The DFRC (Derivatization Followed by Reductive Cleavage) analysis of lignin gives phenylpropanoid (C6–C3 type) degradation products from arylether linkages and can be used to detect lignin specifically from plant tissues, excluding other (poly) phenolic substances. DFRC analysis was employed to investigate the lignin composition in broad-leaves of typical Japanese angiosperms. Many leaves gave both guaiacyl- (G, 3-methoxyphenyl-) and syringyl- (S, 3,5-dimethoxyphenyl-) type degradation products, which indicates the existence of lignin in angiospermous leaves, and the leaf lignin has common structural elements with xylem lignin of the same tree. However, the results sometimes differed among species. Persimmon and cherry leaves revealed only trace amounts of degradation products. These results show the heterogeneous distribution of leaf lignin in angiospermous tree species. The syringyl/guaiacyl ratio of leaf lignin DFRC products was apparently lower than that of xylem lignin, which suggests the role of leaf lignin as a material to reinforce leaf vascular tissue for water conduction, like the low S/G ratio in woody vessel element cells. [Copyright &y& Elsevier]

Abstract: The ratio among syringyl (S) and guaiacyl (G) lignin substructures is increasingly used as criteria in the selection of Eucalyptus species to pulping processes. The S/G distribution of several Brazilian Eucalyptus species and their crossings was determined by analytical pyrolysis-GC/MS (Py-GC/MS) and alkaline nitrobenzene oxidation directly from wood fibers. The ratio varied among the Eucalyptus hybrids. In Py-GC/MS, the S/G ratio is calculated by summing up the proportions of S and G type lignin pyrolysis products with various 1–3 carbon containing side chain structures. Py-GC/MS revealed that the distribution of the lignin side chain structures varied among the species. Similar trends were seen both in the G and in the S series. The result was verified with principal component analysis (PCA), which also revealed that similar side chains structures in the G and S units are correlated with each other. That PCA result confirmed that all G and S type degradation products should be included in the calculation of S/G ratios instead of selected products. S/G ratios determined by Py-GC/MS and alkaline nitrobenzene oxidation were close to each other, but not equal. Lower values were obtained by Py-GC/MS. The differences between the two methods are probably related the degradation mechanism of lignin with Py-GC/MS and nitrobenzene oxidation methods. Based on these results, it is not possible to say which of the methods is more reliable. However, both methods can well be used to compare the S/G ratios of lignin among samples. [Copyright &y& Elsevier]

Heartwood and sapwood samples from Eucalyptus globulus were char-acterized by Py-GC/MS and GC-FID in respect to composition and content of lignin. The pyrolysis lignin-derived compounds were assembled by gmups: "syringol," "S-aldehydes," "S-ketones," "S-alcohols," and "C11H12O3" (S-units); "guaiacol," "eugenol," "G-aldehydes," "G-ketones," "G-alcohols," and "others" (G-units); "phe-nol" and "H-aldehydes" (H-units). Heartwood and sapwood had similar lignin content in an extractive-free basis (23.7% and 23.0%, respectively) and in lignin composition (S-units, 76.0% vs. 76.3%; G-units, 22.0% vs. 21.0%; H-units, 1.9% vs. 2.7%; S/G ratio 3.5 and 3.6). The wood samples were kraft pulped under isothermal conditions at 130°C, 150°C and 170°C and several cooking times. Heartwood and sapwood behaved similarly. Al 130°C llie delignification was slow willi no significant selectivity in respect to lignin composition. At 150°C and 170°C the S-units were more susceptible to reaction and comparatively more removed, inducing a decrease of S/G ratio to 0.6. The main prod-ucts to be extracted belong to "syringol" and "S-aldehydes," while the residual lignin in pulps was enriched in "guaiacol," "eugenol" (G-units), and "phenol" (H-units). [ABSTRACT FROM AUTHOR]

Selected and tested poplar clones are very suitable biomass resources for various applications such as biofuels, the pulp and paper industry as well as chemicals production. In this study, we determined the content of lignin, cellulose, holocellulose, and extractives, syringyl to guaiacyl (S/G) ratio in lignin, and also calculated higher heating values (HHV) among eight examined clones of Populus grown on three different experimental sites. The highest lignin content for all the examined sites was determined in 'I-214' and 'Baka 5' clones, whereas the highest content of extractives was found in 'Villafranca' and 'Baka 5' clones. The highest S/G ratio for all the examined sites was determined in 'Villafranca' and 'Agathe F' clones. The chemical profiles of main wood components, extractives, and the S/G ratio in lignin were also influenced by both the experimental site and the clone × site interaction. Higher heating values, derived from calculations based on the contents of lignin and extractives (or lignin only), were in close agreement with the previously published data. The highest heating values were found for 'Baka 5' and 'I-214' clones. The optimal method of poplar biomass utilization can be chosen on basis of the lignocellulosics chemical composition and the S/G ratio in lignin. [ABSTRACT FROM AUTHOR]

Abstract: Eucalyptus globulus sapwood and heartwood showed no differences in lignin content (23.0% vs. 23.7%) and composition: syringyl-lignin (17.9% vs. 18.0%) and guaiacyl-lignin (4.8% vs. 5.2%). Delignification kinetics of S- and G-units in heartwood and sapwood was investigated by Py-GC–MS/FID at 130, 150 and 170°C and modeled as double first-order reactions. Reactivity differences between S and G-units were small during the main pulping phase and the higher reactivity of S over G units was better expressed in the later pulping stage. The residual lignin composition in pulps was different from wood or from samples in the initial delignification stages, with more G and H-units. S/G ratio ranged from 3 to 4.5 when pulp residual lignin was higher than 10%, decreasing rapidly to less than 1. The S/H was initially around 20 (until 15% residual lignin), decreasing to 4 when residual lignin was about 3%. [Copyright &y& Elsevier]

This study investigated the influence of various hardwood characteristics on enzymatic hydrolysis. Important hardwood species, including three Eucalyptus species, were comprehensively characterized using quantitative 13 C NMR, image analysis and fiber quality analysis. Hydrolysis efficiency from all the hardwoods was correlated to the wood chemical composition and lignin characteristics. Among the key wood components that control enzymatic hydrolysis efficiency, lignin content, enzyme adsorption on substrate and, the ratio of syringyl/guaiacyl (S/G) of the pretreated feedstock were identified as the most important. No wood morphological feature was found to have a significant influence on enzymatic conversion of the pretreated samples. [ABSTRACT FROM AUTHOR]

Lignin content and composition are considered as mandatory traits of eucalyptus breeding programs, especially for pulp, paper, and bioenergy production. In this article, we used 33 Eucalyptus urophylla full-sib families of an 8 × 8 factorial design to provide estimates of genetic parameters for lignin- and growth-related traits. Secondly, from the sequencing of the 16 unrelated founders, we described the nucleotide and haplotype variability of cinnamoyl- CoA reductase ( CCR), a candidate gene for lignin-related traits encoding the cinnamoyl-CoA reductase. Finally, we tested the association between CCR polymorphisms and trait variation using a mixed linear model. A high value of narrow sense heritability was obtained for lignin content ( h² = 0.85) and S/G ratio ( h² = 0.62) indicating that these traits are under strong genetic control. High levels of nucleotide (θ = 0.0131) and haplotype (Hd = 0.958) diversity were detected for CCR. From an initial set of 152 biallelic single nucleotide polymorphisms (SNPs), a subset of 65 nonredundant loci was selected. Three intronic SNPs were found to be associated to the variation of S/G ratio after multiple testing correction. In the line of what has been obtained in forest trees, these SNPs explained between 2.45% and 2.87% of the genetic variance of the trait. This study demonstrates the interest of the candidate gene approach for quantitative trait nucleotide detection in Eucalyptus and paves the way to gene assisted selection of lignin composition in E. urophylla. [ABSTRACT FROM AUTHOR]

Abstract: The degradation of organic macromolecules, including lignin, in plant-derived soil organic matter, is important to the global carbon cycle. In grasslands, saprotrophic (decomposer) fungi are major decomposers of such organic material. The aim of this study was to characterise lignin degradation, particularly with respect to lignin oxidation typical of white-rot basidiomycete fungi. Lignin breakdown products, analysed by gas chromatography–mass spectrometry (GC–MS) with TMAH thermochemolysis, in initial wheat (Triticum aestivum var. Swatham) straw samples were compared with those in samples which had been buried as a “model” resource for 46 months in a sand-dune grassland at Ainsdale National Nature Reserve, Lancashire, UK. Our results showed that lignin oxidation occurred in the straw over the 46 month period, as there were general increases in the [Ac/Al]S and [Ac/Al]G ratios and a clear decrease in the [S/G] ratio. These data provide tentative support for the theory that white-rot basidiomycete fungi are involved in the degradation of lignin in grasslands. [Copyright &y& Elsevier]

In an attempt to explain variations in delignification behaviors among different hardwood species, the kraft pulping delignification rates of Eucalyptus urograndis, E. nitens, E. globulus, sweet gum, maple, red oak, birch, red alder, cottonwood, and acacia were obtained and correlated with their respective lignin chemical structures. Since H-factor for hardwood is calculated based on the softwood activation energy (Ea) value, a comparison between softwood vs. hardwood activation energy was also performed. Lignin was isolated by a modified isolation protocol, using alkaline pretreatment of the wood prior to isolation. The lignin preparations were analyzed via quantitative 13C NMR spectroscopy. Substantial variations were found among the hardwood species studied. A linear correlation between the kraft delignification rate and the amount of syringyl units was found. Activation energy values obtained for kraft pulping of hardwoods were very similar and almost identical to the value obtained for softwood. Birch was the only species with outlier behavior. [ABSTRACT FROM AUTHOR]