Poplar Cell Walls Research Articles

Mapping single molecular binding kinetics of carbohydrate-binding module with crystalline cellulose by atomic force microscopy recognition imaging.

We studied the binding kinetics of family 3 carbohydrate-binding module (CBM3a) molecules to crystalline cellulose fibrils extracted from the poplar cell wall by atomic force microscopy (AFM) recognition imaging. The free CBM3a molecules of different concentrations were added to the buffer solution to bind to the crystalline cellulose sample immobilized on the AFM substrate. During in-situ AFM imaging, the CBM molecules were observed to bind to cellulose efficiently and regularly, especially in the first 60-120 min. A 1:1 single-molecule binding model was used to study the kinetics of the CBM3a-cellulose interaction. The saturation time when the concentration of occupied binding sites is 99% of the maximum bound CBM3a concentration at the end of reaction, t(0.99), was determined by fitting different concentrations of CBM3a against reaction time using the high resolution AFM images and the single-molecule kinetics equations. Based on the experimental data and kinetics calculations, the minimal effective initial CBM3a concentration was estimated to be 5.1 × 10(-7) M at 287 min reaction time. This study provides an in-depth understanding of the binding mechanism of CBM with crystalline cellulose at single molecule level.

Using confocal Raman microscopy to real-time monitor poplar cell wall swelling and dissolution during ionic liquid pretreatment.

The ionic liquids (ILs) are recognized as the potential solvents for the pretreatment of lignocellulosic materials before biomass conversion. However, little knowledge of how the cell wall of biomass responds to the IL locally and dynamically during the pretreatment is available. In the current work, the process of IL pretreatment of poplar using 1-ethyl-3-methylimidazolium acetate ([C2 mim][OAc]) was real-time monitored on a cellular level by employing confocal Raman microscopy. The results showed that the biomass dissolution during the IL pretreatment can be clearly divided into two stages: (1) slow penetration of IL, and (2) rapid dissolution of lignin and carbohydrates. In this case, the onset of the dissolution of these compositions occurred only after the cell wall of biomass swelled to a certain extent. Because the first stage was a slow process which determined the process reaction rate, it can be deduced that enhancing the penetration capacity of IL was crucial for improving the pretreatment efficiency. Based on the obtained results, a model was proposed to better understand how the plant cell wall responds to the IL before, during, and after pretreatment.

Impact of Alkali Pretreatment on the Chemical Component Distribution and Ultrastructure of Poplar Cell Walls

Alkali pretreatment is one of the leading pretreatment technologies for biofuel applications. The histochemical and structural characteristics of poplar cell walls were investigated before and after sodium hydroxide pretreatment (121 oC, 2%) to understand the alterations in biomass cellular structure, which were correlated with saccharification yield. Results showed that alkali pretreatment preferentially removed lignin from the S2 of fibers, which was similar to the behaviors of coniferyl alcohol and aldehyde (lignin-CAA), exhibiting a positive correlation between removal of the two structures. Additionally, the cellulose microfibril angle was enlarged as the residence time increased during pretreatment. Scanning electron microscopy (SEM) analysis further suggested that pretreatment caused ultrastructure changes in cell walls with cracks formation on cell wall surface, especially in the areas adjacent to the cell corner middle lamellar (CCML). Accordingly, the cellulose digestibility of residues increased from 32.1% for the raw material to 53.7% for the treated samples obtained in 72 h. It can be concluded that the changes in topochemistry and ultrastructure of poplar cell walls resulting from alkali pretreatment mediated the efficiency of enzymatic hydrolysis of residues.

Revealing the Changes in Topochemical Characteristics of Poplar Cell Wall During Hydrothermal Pretreatment

Understanding of ultrastructural organization related to the biomass recalcitrance is critical for optimizing the deconstruction of cell wall to release fermentable sugars. In this study, the changes in topochemical characteristics of poplar cell wall pretreated with liquid hot water at 170 °C for 5, 10, 20, 30, and 40 min were investigated. The hydrothermal pretreatment (HTP) caused the alterations of surface morphological and chemical features in different cell wall sublayers in a time-dependent manner. Scanning electron microscopy observations revealed a reduction in re-deposited lignin droplets in the middle layer of secondary wall (S2 layer) after 20 min of pretreatment. Confocal Raman microscopy images exhibited that the loss of lignin concentration in the S2 layer was more than that in the compound middle lamella (CML) after 30 min of pretreatment, whereas the drop of hemicellulose concentration exhibited an opposite trend. Meanwhile, the decline of lignin and hemicellulose concentrations was accompanied by the increase in relative concentration of cellulose, indicating the exposure of embedded cellulose. The higher yield of enzymatic hydrolysis obtained from the poplars after 30 min of pretreatment were 38 %, indicating that the removal of lignin and hemicelluloses from the S2 layer and CML is likely to be important factors in increasing enzymatic hydrolysis of poplar. This study is expected to potentially promote understanding of ultrastructural features hindering cell wall deconstruction, making it effective to convert lignocellulosic biomass into sugars.

Multi-Scale Visualization of Dynamic Changes in Poplar Cell Walls During Alkali Pretreatment

Alkali pretreatment is a promising pretreatment technology that can effectively deconstruct plant cell walls to enhance sugar release performance. In this study, multi-scale visualization of dynamic changes in poplar cell walls during sodium hydroxide pretreatment (2% w/v, 121°C) was carried out by light microscopy (LM), confocal Raman microscopy (CRM) and atomic force microscopy (AFM). LM observations indicated that swelling occurred primarily in the secondary wall (S) but alkali had little effect on the cell corner middle lamella (CCML). Correspondingly, there was a preferential delignification in the S at the beginning of pretreatment, while the level of delignification in CCML (~88%) was higher than that in the S (~83%) for the whole process revealed by Raman spectra. It also suggested that prolonging residence time to 180 min would not remove lignin completely but cause rapid loss of carbohydrates, which was further visualized by Raman spectroscopy images. Furthermore, AFM measurements illustrated that pretreatment with alkali exposed the embedded microfibrils from noncellulosic polymers clearly, enlarged the diameter of microfibrils, and decreased the surface porosity. These results suggested that there was a synergistic mechanism of lignocellulose deconstruction regarding cell wall swelling, main components dissolution, and microfibril morphological changes that occurred during alkali pretreatment.

Wood chemistry analysis and expression profiling of a poplar clone expressing a tyrosine-rich peptide

Our study has identified pathways and gene candidates that may be associated with the greater flexibility and digestibility of the poplar cell walls. With the goal of facilitating lignin removal during the utilization of woody biomass as a biofuel feedstock, we previously transformed a hybrid poplar clone with a partial cDNA sequence encoding a tyrosine- and hydroxyproline-rich glycoprotein from parsley. A number of the transgenic lines released more polysaccharides following protease digestion and were more flexible than wild-type plants, but otherwise normal in phenotype. Here, we report that overexpression of the tyrosine-rich peptide encoding sequence in these transgenic poplar plants did not significantly alter total lignin quantity or quality (S/G lignin ratio), five- and six-carbon sugar contents, growth rate, or susceptibility to a major poplar fungal pathogen, Septoria musiva. Whole-genome microarray analysis revealed a total of 411 differentially expressed transcripts in transgenic lines, all with decreased transcript abundance relative to wild-type plants. Their corresponding genes were overrepresented in functional categories such as secondary metabolism, amino acid metabolism, and energy metabolism. Transcript abundance was decreased primarily for five types of genes encoding proteins involved in cell-wall organization and in lignin biosynthesis. The expression of a subset of 19 of the differentially regulated genes by qRT-PCR validated the microarray results. Our study has identified pathways and gene candidates that may be the underlying cause for the enhanced flexibility and digestibility of the stems of poplar plants expressing the TYR transgene.

Evaluation of the effects of compression combined with heat treatment by nanoindentation (NI) of poplar cell walls

Abstract A combination of compression and heat treatment is a modification method that has great potential for improving the mechanical properties and dimensional stability of wood materials in industrial application. The objective of this project was to track changes in the microstructure, chemical composition, cellulose crystallinity, and mechanical properties of the treated poplar cell wall to investigate the mechanism of modification. Poplar boards were compressed at 100°C and subsequently treated in the hot press at 200°C. The results indicated that the treatment contributed to a reduction in porosity without obvious mechanical compression and damage to the cell wall. Hemicellulose degraded, however, and the relative lignin content and cellulose crystallinity increased during the process. The observed increase in relative lignin content and crystallinity may contribute to the improvement of mechanical properties. The longitudinal elastic modulus and hardness of poplar cell walls increased significantly from 12.5 and 0.39 GPa for the control to a maximum of 15.7 and 0.51 GPa for compressed wood with HT, respectively.

EFFECT OF ENZYME TREATMENT ON THE MECHANICAL PROPERTIES OF WOOD CELL WALLS BY NANOINDENTATION

The objective of this research was to study the changes in hardness and elastic modulus of wood cell walls treated with enzymes. Such changes greatly influence the properties of paper and wood composites. Two enzymes, hemicellulase and lipase, were selected for the treatment. Poplar samples (Populus euramevicana) were treated with hemicellulase, while samples of southern yellow pine (Pinus spp.) and Mongolia scotch pine (Pinus sylvestris L. var. mongolica Litv.) were treated with lipase. Mechanical properties of both treated samples were investigated by nanoindentation. The results showed some changes in the hardness and elastic modulus of the poplar cell wall treated by hemicellulase. Hardness and elastic modulus values of southern yellow pine and Mongolia scotch pine cell walls treated by lipase decreased with increasing amounts of the enzyme.

Reversible swelling of the cell wall of poplar biomass by ionic liquid at room temperature

Time-resolved autofluorescence, Raman microspectroscopy, and scanning microprobe X-ray diffraction were combined in order to characterize lignocellulosic biomass from poplar trees and how it changes during treatment with the ionic liquid 1- n-ethyl-3-methylimidazolium acetate (EMIMAC) at room temperature. The EMIMAC penetrates the cell wall from the lumen, swelling the cell wall by about a factor of two towards the empty lumen. However, the middle lamella remains unchanged, preventing the cell wall from swelling outwards. During this swelling, most of the cellulose microfibrils are solubilized but chain migration is restricted and a small percentage of microfibrils persist. When the EMIMAC is expelled, the cellulose recrystallizes as microfibrils of cellulose I. There is little change in the relative chemical composition of the cell wall after treatment. The action of EMIMAC on the poplar cell wall at room temperature would therefore appear to be a reversible swelling and a reversible decrystallization of the cell wall.

Compositional characterization and imaging of “wall-bound” acylesters of Populus trichocarpa reveal differential accumulation of acyl molecules in normal and reactive woods

Acylesterification is one of the common modifications of cell wall non-cellulosic polysaccharides and/or lignin primarily in monocot plants. We analyzed the cell-wall acylesters of black cottonwood (Populus trichocarpa Torr. & Gray) with liquid chromatography-mass spectrometry (LC-MS), Fourier transform-infrared (FT-IR) microspectroscopy, and synchrotron infrared (IR) imaging facility. The results revealed that the cell wall of dicotyledonous poplar, as the walls of many monocot grasses, contains a considerable amount of acylesters, primarily acetyl and p-hydroxycinnamoyl molecules. The "wall-bound" acetate and phenolics display a distinct tissue specific-, bending stress responsible- and developmental-accumulation pattern. The "wall-bound" p-coumarate predominantly accumulated in young leaves and decreased in mature leaves, whereas acetate and ferulate mostly amassed in the cell wall of stems. Along the development of stem, the level of the "wall-bound" ferulate gradually increased, while the basal level of p-coumarate further decreased. Induction of tension wood decreased the accumulation of the "wall-bound" phenolics while the level of acetate remained constant. Synchrotron IR-mediated chemical compositional imaging revealed a close spatial distribution of acylesters with cell wall polysaccharides in poplar stem. These results indicate that different "wall-bound" acylesters play distinct roles in poplar cell wall structural construction and/or metabolism of cell wall matrix components.

Molecular mobility of poplar cell wall polymers studied by dielectric techniques

In this work, thermally stimulated currents (TSC) analyzes combined with dynamic dielectric spectroscopy (DDS) have been applied to the investigation of polymers’ molecular mobility involves in poplar cell wall. The molecular origin of the various dielectric relaxations has been determined. Cellulose and lignin effects in wood dielectric response were distinguished. The correlation between results obtained by both dielectric methods allows us to follow molecular mobility involved in delocalized movement as primary relaxation mode. For these two major components of wood, the evolution of relaxation times involve in α-relaxation mode is explained using the strong/fragile pattern. We compared the cellulose and lignin in situ and ex situ responses to interpret wood compound behavior. The importance of structural wood interactions which modified the molecular mobility of polymer components will be underlined.

Lignin dehydrogenative polymerization mechanism: a poplar cell wall peroxidase directly oxidizes polymer lignin and produces in vitro dehydrogenative polymer rich in β-O-4 linkage

An investigation was performed to determine whether lignin dehydrogenative polymerization proceeds via radical mediation or direct oxidation by peroxidases. It was found that coniferyl alcohol radical transferred quickly to sinapyl alcohol. The transfer to syringaresinol was slower, however, the transfer to polymeric lignols occurred very slightly. This result suggests that the radical mediator theory does not sufficiently explain the mechanism for dehydrogenative polymerization of lignin. A cationic cell wall peroxidase (CWPO-C) from poplar (Populus alba L.) callus showed a strong substrate preference for sinapyl alcohol and the sinapyl alcohol dimer, syringaresinol. Moreover, CWPO-C was capable of oxidizing high-molecular-weight sinapyl alcohol polymers and ferrocytochrome c. Therefore, the CWPO-C characteristics are important to produce polymer lignin. The results suggest that CWPO-C may be a peroxidase isoenzyme responsible for the lignification of plant cell walls.

Ultrastructural Changes in Poplar Cell Wall during Steam Explosion Treatment

Ultrastructural Changes in Poplar Cell Wall during Steam Explosion Treatment

Effect of steam explosion treatment on the physico-chemical characteristics and enzymic hydrolysis of poplar cell wall components

The purpose of this study was to determine the effect of steam explosion pretreatment on pore size distribution and enzymic hydrolysis of poplar wood chips under various conditions (duration and temperature of treatment, pH impregnation). The measurements showed an increase in the available surface area with the severity of treatment and a good correlation with the initial rate of enzymic hydrolysis. This increase in enzymic hydrolysis can be explained by the removal of the hemicelluloses, but also by the melting and agglomeration of the depolymerized lignin, as shown on electron micrographs of ultrathin cross-sections.

Fine structure of cellulose microfibrils in poplar gelatinous layer and Valonia

Fine structure of cellulose microfibrils in poplar (Populus euramericana) gelatious layer and Valonia cell walls was observed in the electron microscope by using disintegration and ultrathin section techniques with various electron stains. Staining of the gelatinous layer in poplar showed that the microfibrils had a paracrystalline region surrounding a crystalline core, but such a region was negligible in Valonia. From the facts that kinks and shortened microfibrils were observed after mechanical and hydrolysis treatments, respectively, and that the microfibrils were unaffected by negative staining, it was concluded that the microfibril core is uniformly crystalline in the longitudinal direction although there may be some crystalline dislocations.