Anouck Habrant

Concomitant changes in viscoelastic properties and amorphous polymers during the hydrothermal treatment of hardwood and softwood.

The aim of this study was to understand how the molecular structures of amorphous polymers influence wood viscoelastic properties. Wood from oak and spruce was subjected to hydrothermal treatments at 110 or 135 degrees C. Wood rigidity, reflected by the wood storage modulus, showed different modification patterns according to the wood species or the temperature level. Because viscoelasticity is dependent on wood amorphous polymers, modifications of lignins and noncellulosic polysaccharides were examined. Hemicellulose degradation occurred only at 135 degrees C. In contrast, lignins displayed major structural alterations even at 110 degrees C. In oak lignins, the beta-O-4 bonds were extensively degraded and wood rigidity decreased dramatically during the first hours of treatment. Spruce lignins have a lower beta-O-4 content and, relative to oak, the wood rigidity decrease due to treatment was less pronounced. Wood rigidity was restored to its initial value by prolonged treatment, probably due to the formation of condensed bonds in cell wall polymers.

Changes in Phenolics Distribution After Chemical Pretreatment and Enzymatic Conversion of Miscanthus × giganteus Internode

In addition to lignin, grass cell walls are characterized by the presence of hydroxycinnamic acids that play a significant role in cross-linking polymers into a cohesive network, and pretreatments are required to overcome the recalcitrance of lignocelluloses prior to enzymatic bioconversion of polysaccharides. The effects of dilute acid and ammonium hydroxide pretreatments were studied on the chemical composition and enzymatic saccharification of Miscanthus internodes fragments. The hydroxycinnamic acid content was reduced after both pretreatments, while lignin got enriched in condensed linked structures. In addition, dilute acid pretreatment was effective in decreasing xylan content of Miscanthus, while ammonia treatment induced a marked swelling effect on the cell walls of parenchyma, vascular sclerenchyma, and epidermal sclerenchyma. The phenol distribution at the cell level was estimated using UV transmission microspectrophotometry. Internode cell walls displayed different UV spectra according to the cell type. However, the secondary cell walls had similar UV spectra after pretreatment, whereas spectra recorded at the cell corner region displayed variations according to cell type and pretreatment. Acid pretreatment was more efficient than ammonia to improve the conversion of polysaccharides by a Trichoderma cellulolytic cocktail. Although pretreatments achieved moderate saccharification yields, the secondary cell walls were altered at some pit regions of the vascular sclerenchyma whereas parenchyma appeared recalcitrant. Variations in the UV spectra of enzyme-digested cell walls suggest pretreatment-dependent heterogeneity of the phenolic distribution in the more recalcitrant cell walls.

Changes in hemp secondary fiber production related to technical fiber variability revealed by light microscopy and attenuated total reflectance Fourier transform infrared spectroscopy

Interest in hemp (Cannabis sativa L.) is increasing due to the development of a new range of industrial applications based on bast fibers. However the variability of bast fiber yield and quality represents an important barrier to further exploitation. Primary and secondary fiber content was examined in two commercial hemp varieties (Fedora 17, Santhica 27) grown under contrasted sowing density and irrigation conditions. Both growing conditions and hemp varieties impact stem tissue architecture with a large effect on the proportion of secondary fibers but not primary fibers. Attenuated total reflectance infrared spectroscopy allowed the discrimination of manually-isolated native primary fibers and secondary fibers but did not reveal any clustering according to growing conditions and variety. Infrared data were confirmed by wet chemistry analyses that revealed slight but significant differences between primary and secondary fiber cell wall composition. Infrared spectroscopy of technical fibers obtained after mechanical defibering revealed differences with native primary, but not secondary fibers and also discriminated samples obtained from plants grown under different conditions. Altogether the results suggested that the observed variability of hemp technical fibers could be partially explained by i) differences in secondary fiber production and ii) differential behavior during mechanical defibering resulting in unequal separation of primary and secondary fibers.

Action of lytic polysaccharide monooxygenase on plant tissue is governed by cellular type

Lignocellulosic biomass bioconversion is hampered by the structural and chemical complexity of the network created by cellulose, hemicellulose and lignin. Biological conversion of lignocellulose involves synergistic action of a large array of enzymes including the recently discovered lytic polysaccharide monooxygenases (LPMOs) that perform oxidative cleavage of cellulose. Using in situ imaging by synchrotron UV fluorescence, we have shown that the addition of AA9 LPMO (from Podospora anserina) to cellulases cocktail improves the progression of enzymes in delignified Miscanthus x giganteus as observed at tissular levels. In situ chemical monitoring of cell wall modifications performed by synchrotron infrared spectroscopy during enzymatic hydrolysis demonstrated that the boosting effect of the AA9 LPMO was dependent on the cellular type indicating contrasted recalcitrance levels in plant tissues. Our study provides a useful strategy for investigating enzyme dynamics and activity in plant cell wall to improve enzymatic cocktails aimed at expanding lignocelluloses biorefinery.

Evaluation of Lignocellulosic Biomass Degradation by Combining Mid- and Near-Infrared Spectra by the Outer Product and Selecting Discriminant Wavenumbers Using a Genetic Algorithm.

Mid-infrared (MIR) and near-infrared (NIR) spectroscopy provide useful information on the molecular composition of biological systems. Because they are sensitive to organic and mineral components, there is a growing interest in these techniques for the development of biomarkers that reflect intrinsic characteristics of plants and their mode of degradation. Due to their complexity and complementary nature, an important challenge is the combining of MIR and NIR information to identify discriminating wavenumbers in each wavenumber region, with the ultimate goal of assessing the biodegradation process of a lignocellulosic biomass at different time scales. This work investigates the potential of using the outer product to combine MIR and NIR spectra to highlight the connections between fundamental molecular vibrations and their combinations and bonds. Because this operation yields high-dimensional spectra, we propose to use a genetic algorithm to select the most discriminant wavenumbers within the degradation process. The results from two lignocellulosic biomasses with different biodegradation kinetics, miscanthus aerial parts and maize roots, confirm that the outer product combination of MIR and NIR spectral information allows a better discrimination of the biodegradation kinetic compared with the simple concatenation of MIR and NIR spectra or with the use of MIR or MIR spectral information separately. We show that the genetic algorithm selects wavenumbers that correspond to principal vibrations of chemical functional groups of compounds that undergo degradation/conversion during the biodegradation of the lignocellulosic biomass.

Fluorescent Nano-Probes to Image Plant Cell Walls by Super-Resolution STED Microscopy

Lignocellulosic biomass is a complex network of polymers making up the cell walls of plants. It represents a feedstock of sustainable resources to be converted into fuels, chemicals, and materials. Because of its complex architecture, lignocellulose is a recalcitrant material that requires some pretreatments and several types of catalysts to be transformed efficiently. Gaining more knowledge in the architecture of plant cell walls is therefore important to understand and optimize transformation processes. For the first time, super-resolution imaging of poplar wood samples has been performed using the Stimulated Emission Depletion (STED) technique. In comparison to standard confocal images, STED reveals new details in cell wall structure, allowing the identification of secondary walls and middle lamella with fine details, while keeping open the possibility to perform topochemistry by the use of relevant fluorescent nano-probes. In particular, the deconvolution of STED images increases the signal-to-noise ratio so that images become very well defined. The obtained results show that the STED super-resolution technique can be easily implemented by using cheap commercial fluorescent rhodamine-PEG nano-probes which outline the architecture of plant cell walls due to their interaction with lignin. Moreover, the sample preparation only requires easily-prepared plant sections of a few tens of micrometers, in addition to an easily-implemented post-treatment of images. Overall, the STED super-resolution technique in combination with a variety of nano-probes can provide a new vision of plant cell wall imaging by filling in the gap between classical photon microscopy and electron microscopy.