Cross-sectional analysis of the polysaccharide composition in cellulosic fiber materials by enzymatic peeling/high-performance capillary zone electrophoresis

A combined enzymatic, chemical, and analytical approach was used to determine the cross-sectional carbohydrate composition in cellulosic fibers. The outer surface of cellulosic fibers was enzymatically removed layer-by-layer with precise quantitative control, and the monosaccharides in the peelings were subsequently analyzed by high-performance capillary electrophoresis (HPCE) after precolumn derivatization with a UV label. This method was applied to dissolving pulps and regenerated cellulose fibers, with special emphasis on the cross-sectional distribution of hemicelluloses. Commercially available enzyme solutions were used, resulting in a reproducible peeling. Significant differences were found in the hemicellulose distribution across the fiber of different dissolving pulps, dependent on both natural source (beech or spruce) and preparation process (acidic sulfite cook or prehydrolysis kraft cook). Among the dissolving pulps, beech prehydrolysis kraft pulp showed the highest enrichment of surface xylan. Similar, albeit smaller, differences were noticed between various regenerated fibers (viscose, viscose Modal, and Lyocell): a thin hemicellulose-rich outermost layer was found in all the regenerated fibers studied.     

Pulp properties and their influence on enzymatic degradability

Endoglucanase treatment of pulp for the adjustment of viscosity and the increase in pulp reactivity is a promising step in the concept for the beneficial production of dissolving pulps from paper grade pulps. To promote the commercial applicability of these enzymes, the influence of pulp properties such as carbohydrate composition, pulp type and cellulose morphology on the enzymatic degradability of a pulp was examined. High contents of hemicelluloses and lignin were shown to impair the accessibility of the cellulose to the enzymes. Due to the elevated swelling capacity of cellulose II, conversion of the cellulose morphology from I to II upon alkaline treatments showed a large increasing effect on the cellulose accessibility, and enzymatic degradability. Reactivity measurements of softwood sulfite pulps after enzymatic degradation and acid-catalyzed hydrolysis, respectively, revealed elevated reactivity for the pulp after acid treatment. This is in contrast to effects of enzyme treatments reported for CCE treated kraft pulps.     

Nanofibrillation of wood pulp using a high-speed blender

We report the successful use of a high-speed blender in nanofibrillating never-dried pulp to cellulose nanofibers (CNFs) with a uniform diameter of 15-20 nm. Pulp treated for 30 min in a blender showed the same degree of fibrillation with less damage to the CNF compared with that treated in a grinder. Observing the process of nanofibrillation clarified that the straw-like pulp was fibrillated in a very characteristic way, by forming many "balloon-like structures". As the balloons extended to the edges, the fibrils were rapidly individualized. However, the pulp fragments with ripped cell walls were split into finer fragments and gradually disintegrated into nanofibers. Changing the agitation speed and pulp concentration during the treatment revealed that the pulp concentration of 0.7 wt % at 37,000 rpm was the optimum fibrillation condition in this blender method. Through treatments in various NaCl solutions, the effect of the surface charge of CNF on the fibrillation was studied from the viewpoint of colloidal surface physics. It was found that repulsion due to the electric double layer of the CNF surface may play a critical role in the occurrence of fibrillation.     

Predicting the most appropriate wood biomass for selected industrial applications: comparison of wood,pulping, and enzymatic treatments using fluorescent-tagged carbohydrate-binding modules

Background

Lignocellulosic biomass will progressively become the main source of carbon for a number of products as the Earth’s oil reservoirs disappear. Technology for conversion of wood fiber into bioproducts (wood biorefining) continues to flourish, and access to reliable methods for monitoring modification of such fibers is becoming an important issue. Recently, we developed a simple, rapid approach for detecting four different types of polymer on the surface of wood fibers. Named fluorescent-tagged carbohydrate-binding module (FTCM), this method is based on the fluorescence signal from carbohydrate-binding modules-based probes designed to recognize specific polymers such as crystalline cellulose, amorphous cellulose, xylan, and mannan.

Results

Here we used FTCM to characterize pulps made from softwood and hardwood that were prepared using Kraft or chemical-thermo-mechanical pulping. Comparison of chemical analysis (NREL protocol) and FTCM revealed that FTCM results were consistent with chemical analysis of the hemicellulose composition of both hardwood and softwood samples. Kraft pulping increased the difference between softwood and hardwood surface mannans, and increased xylan exposure. This suggests that Kraft pulping leads to exposure of xylan after removal of both lignin and mannan. Impact of enzyme cocktails from Trichoderma reesei (Celluclast 1.5L) and from Aspergillus sp. (Carezyme 1000L) was investigated by analysis of hydrolyzed sugars and by FTCM. Both enzymes preparations released cellobiose and glucose from pulps, with the cocktail from Trichoderma being the most efficient. Enzymatic treatments were not as effective at converting chemical-thermomechanical pulps to simple sugars, regardless of wood type. FTCM revealed that amorphous cellulose was the primary target of either enzyme preparation, which resulted in a higher proportion of crystalline cellulose on the surface after enzymatic treatment. FTCM confirmed that enzymes from Aspergillus had little impact on exposed hemicelluloses, but that enzymes from the more aggressive Trichoderma cocktail reduced hemicelluloses at the surface.

Conclusions

Overall, this study indicates that treatment with enzymes from Trichoderma is appropriate for generating crystalline cellulose at fiber surface. Applications such as nanocellulose or composites requiring chemical resistance would benefit from this enzymatic treatment. The milder enzyme mixture from Aspergillus allowed for removal of amorphous cellulose while preserving hemicelluloses at fiber surface, which makes this treatment appropriate for new paper products where surface chemical responsiveness is required.

     

Differences in residual lignin properties between Betula verrucosa and Eucalyptus urograndis kraft pulps

By comparing the ultrastructural features of two oxygen delignified hardwood kraft pulps (Eucalyptus urograndis and Betula verrucosa), we have demonstrated a marked difference in their residual lignin properties. In this study, properties such as crystallinity and crystal size of cellulose, molecular weights, carboxyl group contents, and carbohydrate compositions of the two kraft pulps were compared. The examined pulps were in our observations relatively similar. A significant difference, however, was observed in the size exclusion chromatography measurements, which indirectly suggested that a significant portion of residual lignin in eucalyptus pulp was associated with cellulose. Birch pulp, in contrast, exhibited a more conventional tendency for hardwood pulps: lignin mainly associated with hemicelluloses.     

Extracting hemicelluloses prior to aspen chemi-thermomechanical pulping: Effects of pre-extraction on pulp properties

Pre-extraction of hemicelluloses prior to pulping and conversion of the extracted hemicelluloses to other by-products will provide additional revenues to traditional pulp and paper industry. The effects of hemicelluloses pre-extraction with sulfuric acid on aspen (Populus tremuloides) chemi-thermomechanical pulp (CTMP) properties were investigated in this study. The sulfuric acid pre-extraction resulted in a release of 11% of hemicelluloses (determined as xylan) and acetic acid, especially at the second press-impregnation stage. Compared with CTMP without acid pre-extraction, the acid pre-extraction significantly reduced refining energy to a given freeness, and the pulps produced from acid pre-extracted chips have lower shives, higher strength properties except zero-span breaking length, lower bulk, but lower brightness at a similar specific refining energy. This study provides useful information for integration of biorefinery in mechanical pulp mills in the future.     

N-(Carboxyacyl)chitosans

Hemicelluloses were isolated from pineapple-leaf fibers under different conditions. Study of the properties of these hemicelluloses gave direct evidence of some ester linkages between the hemicellulose and the lignin in this fiber. An aldobiouronic acid was isolated from this fiber hemicellulose, and characterized as 2-O-(4-O-methyl-α-d-glucopyranosyluronic acid)-d-xylose. This indicates that the general nature of the hemicellulose is similar to those of jute and other fiber hemicelluloses.     

Effect of alkaline and autohydrolysis processes on the purity of obtained hemicelluloses from corn stalks

A study of the potential of autohydrolysis and alkaline extraction processes from corn stalks was performed for high purity hemicellulose extraction. The influence of process parameters on the purity of obtained hemicelluloses was analyzed. An experimental design was developed for the autohydrolysis treatments to determine the optimal conditions to solubilize the hemicelluloses with lowest content in contaminants. On the other hand, alkaline extraction, including raw material pretreatment (dewaxing and delignification step) was carried out analyzing the effectiveness of this processes for maximum pure hemicellulose recovery. The maximum yield (54% of the raw material hemicelluloses) and the best physicochemical properties (highest hemicellulose content free of lignin) were obtained with these pretreatments in alkaline extraction. Moreover, the effect of lignin removal by sulfuric acid from the autohydrolysis liquors before hemicellulose precipitation was studied. This purification step has allowed to obtain lignin-free autohydrolysis hemicellulose but with the presence of sulfur as predominant contaminant.     

Investigation of lignin-carbohydrate complexes in kraft pulps by selective enzymatic treatments

The occurrence of covalent bonds between residual lignin and polysaccharides in birch and pine kraft pulps was investigated by specific enzymatic treatments. Pure enzymes degrading cellulose, xylan and mannan were used both separately and in combination. Comparison of the molar masses of polysaccharides and lignin in the orginal pulps and in the residual pulps after enzymatic treatments showed that residual lignin in birch kraft pulp is linked at least to xylan. A minor portion may also be linked to cellulose. In pine kraft pulp some of the residual lignin appears to be linked to cellulose, glucomannan and xylan. The linkages between lignin and cellulose and hemicelluloses may be either native or formed during pulp processing. The results also provided new information on the synergistic action of cellulose- and hemicellulose-degrading enzymes on pulp fibres. The synergism appears to be mainly due to the structure of the pulp fibres, with different layers of cellulose sheets, hemicelluloses and lignin. On the other hand the results also provided information about fibre structure. The degradation of xylan clearly enhanced the action of enzymes on cellulose, suggesting that xylan partially covers the cellulose. A similar phenomenon was not observed in the simultaneous hydrolysis of glucomannan and cellulose. However, the results suggest that glucomannan does interact with cellulose, possibly by non-covalent linkages. Received: 8 July 1998 / Received revision: 7 October 1998 / Accepted: 11 October 1998     

Routes to Potential Bioproducts from Lignocellulosic Biomass Lignin and Hemicelluloses

An essential feature of proposed fermentation-based lignocellulose to biofuel conversion processes will be the co-production of higher value chemicals from lignin and hemicellulose components. Over the years, many routes for chemical conversion of lignin and hemicelluloses have been developed by the pulp and paper industry and we propose that some of these can be applied for bioproducts manufacturing. For lignin products, thermochemical, chemical pulping, and bleaching methods for production of polymeric and monomeric chemicals are reviewed. We conclude that peroxyacid chemistry for phenol and ring-opened products looks most interesting. For hemicellulose products, preextraction of hemicelluloses from woody biomass is important and influences the mixture of solubilized material obtained. Furfural, xylitol, acetic acid, and lactic acid are possible targets for commercialization, and the latter can be further converted to acrylic acid. Pre-extraction of hemicelluloses can be integrated into most biomass-to-biofuel conversion processes.     

Nerve growth factor receptor-like immunoreactivity in primary and permanent canine tooth pulps of the cat

Summary The distribution of nerve growth factor receptor (NGF receptor)-like immunoreactivity in pulps of developing primary and mature permanent cat canine teeth was examined, by use of a monoclonal antibody against NGF receptor detected by fluorescence immunohistochemistry and pre-embedding immunocytochemical light- and electron microscopy. Both primary and permanent pulps contained a vast number of NGF receptor-like immunoreactive nerves. Immunolabelling appeared to be localized both to axons and Schwann cells. In addition, many blood vessel walls in immature primary tooth pulps showed NGF receptor-like immunoreactivity, in contrast to permanent pulps where blood vessels rarely were NGF receptor-immunoreactive. Double-labelling immunofluorescence experiments revealed that in the permanent pulp a majority of the NGF receptor-positive nerves also showed calcitonin gene-related peptide (CGRP)-like immunoreactivity, and many showed substance P-like immunoreactivity. However, nerve fibers with neuropeptide Y-like immunoreactivity lacked NGF receptor-like immunoreactivity. In developing primary tooth pulps fewer NGF receptor-positive nerves were CGRP-like immunoreactive or substance P-like immunoreactive, as compared to the permanent pulp. Neuropeptide Y-like immunoreactive nerve fibers were not detected in the primary tooth pulp. The results suggest a role for nerve growth factor in both developing and mature sensory nerves of the tooth pulp.     

Preliminary studies on TCF bleaching of Pinus pinaster acetosolv pulps.

Oxygen pre-treatment of Pinus pinaster acetosolv pulps has been studied as a first step towards TCF bleaching. Using a 2(3) factorial design, the influence of temperature (80-120 degrees C), time (1-2 h) and NaOH concentration (1.5-3%) on pulp yield in the oxygen stage, chemical composition and physical properties of the pulps obtained was studied. Pulps pre-bleached with oxygen in the conditions selected as optimal (80 degrees C, 1 h with 2.25% NaOH) have been bleached with TCF sequences which included stages with hydrogen peroxide or hydrogen peroxide-oxygen under pressure. Even if high degrees of delignification were reached, with a reduction in Kappa number up to 95% and without important loss of viscosity, the carbohydrates degradation, especially hemicelluloses in the acetic acid delignification, reduces the strength potential of the pulps.     

Dissolution behavior of different celluloses

Celluloses from different origins were dissolved stepwise in N,N-dimethylacetamide/lithium chloride (9% v/w; DMAc/LiCl) with the aim to study the time course of the dissolution process, completeness of dissolution in the dissolved fractions, possible discrimination effects, and differences between the celluloses. Cellulosic pulps from both annual plants and different wood species were analyzed. The obtained fractions were subject to gel permeation chromatography (GPC) with multiple detection to monitor the development of molecular mass distribution (MMD), molecular mass, and recovered mass. The dissolution behavior of accompanying xylans was followed by quantitative analysis of the uronic acids by fluorescence labeling--GPC. The morphological changes at the remaining fibers in the stepwise dissolution were addressed by SEM. The time needed to dissolve completely the cellulosic pulp differed from species to species, mainly between pulps from annual plants and pulps from wood. Annual plants generally needed much longer to dissolve completely. In the beginning of the dissolution, the dissolved fractions of annual plants showed a distinct discrimination effect because they were enriched in hemicellulose. By contrast, wood pulps dissolve fast and without distinct changes in the MMD of the dissolved fractions over time. Bagasse pulp is an exception to the observation for annual plants and rather resembled the behavior of wood celluloses. Prolonged dissolution times, as often practiced in cellulose GPC, do not lead to any improvements regarding the determination of molecular mass, MMD, and recovered mass of injected sample, so that the dissolution times required for reliable GPC analysis can be significantly shortened, which will be important for biorefinery analytics with high numbers of samples.     

Selective solubilization of xylan in pulp using a purified xylanase fromTrichoderma harzianum

Summary The specificity of action of a cellulase-free xylanase preparation on pulp fibers was revealed by the composition of the solubilized products after enzyme treatment. The neutral carbohydrates released by the treatment of two hardwood kraft pulps were composed exclusively of xylooligomers. A similar treatment of Solka Floc showed no detrimental effect on the degree of polymerization of the cellulose fibers, as determined by size exclusion chromatographic analysis.     

New approach for upgrading pulp & paper quality: Mild potassium permanganate treatment of already bleached pulps

The present work introduces mild – room temperature – potassium permanganate treatment of cellulosic materials, namely already bleached pulps. Such treatment represents a new approach for upgrading pulp and paper quality, which is lacking in the literature. Potassium permanganate was investigated as a purifying and mild oxidizing agent for commercial already bleached softwood and bagasse pulps. It was found that treatment of the bleached beaten pulps, with 0.25–2% KMnO₄ (based on pulp weight), led to significant improvement in paper properties. The strength (breaking length) increased greatly and the brightness increased significantly due to treatment. The improvements were related to the degree of polymerization, and to the alphacellulose content of pulps.Moreover, potassium permanganate serves as a disinfectant and deodorizer. Thus treatment of bleached pulps with KMnO₄ is a promising remedy for the side effects which pulps suffer, during transportation and storage, before papermaking.     

Delignification of Wood Chips and Pulps by Using Natural and Synthetic Porphyrins: Models of Fungal Decay

Kraft pulps, prepared from softwoods, and small chips of birch wood were treated with heme and tert-butyl hydroperoxide in aqueous solutions at reflux temperature. Analyses of treated pulps showed decreases in kappa number (a measure of lignin content) from about 36 to less than 2, with concomitant increases in brightness (80% increase in the better samples). Analyses of treated wood chips revealed selective delignification and removal of hemicelluloses. After 48 h of treatment, lignin losses from the wood chips approached 40%, and xylose/mannose (hemicellulose) losses approached 70%, while glucose (cellulose) losses were less than 10%. Examination of delignified chips by transmission electron microscopy showed that the removal of lignin occurred in a manner virtually indistinguishable from that seen after decay by white rot fungi. Various metalloporphyrins, which act as biomimetic catalysts, were compared to horseradish peroxidase and fungal manganese peroxidase in their abilities to oxidize syringaldazine in an organic solvent, dioxane. The metalloporphyrins and peroxidases behaved similarly, and it appeared that the activities of the peroxidases resulted from the extraction of heme into the organic phase, rather than from the activities of the enzymes themselves. We concluded that heme-tert-butyl hydroperoxide systems in the absence of a protein carrier mimic the decay of lignified tissues by white rot fungi.     

Characterization of the crystalline structure of cellulose using static and dynamic FT-IR spectroscopy

The cellulose structure is a factor of major importance for the strength properties of wood pulp fibers. The ability to characterize small differences in the crystalline structures of cellulose from fibers of different origins is thus highly important. In this work, dynamic FT-IR spectroscopy has been further explored as a method sensitive to cellulose structure variations. Using a model system of two different celluloses, the relation between spectral information and the relative cellulose Ialpha content was investigated. This relation was then used to determine the relative cellulose Ialpha content in different pulps. The estimated cellulose I allomorph compositions were found to be reasonable for both unbleached and bleached chemical pulps. In addition, it was found that the dynamic FT-IR spectroscopy technique had the potential to indicate possible correlation field splitting peaks of cellulose Ibeta.     

Effect of Residual Lignin Type and Amount on Bleaching of Kraft Pulp by Trametes versicolor

The white rot fungus Trametes (Coriolus) versicolor can delignify and brighten unbleached hardwood kraft pulp within a few days, but softwood kraft pulps require longer treatment. To determine the contributions of higher residual lignin contents (kappa numbers) and structural differences in lignins to the recalcitrance of softwood kraft pulps to biobleaching, we tested softwood and hardwood pulps cooked to the same kappa numbers, 26 and 12. A low-lignin-content (overcooked) softwood pulp resisted delignification by T. versicolor, but a high-lignin-content (lightly cooked) hardwood pulp was delignified at the same rate as a normal softwood pulp. Thus, the longer time taken by T. versicolor to brighten softwood kraft pulp than hardwood pulp results from the higher residual lignin content of the softwood pulp; possible differences in the structures of the residual lignins are important only when the lignin becomes highly condensed. Under the conditions used in this study, when an improved fungal inoculum was used, six different softwood pulps were all substantially brightened by T. versicolor. Softwood pulps whose lignin contents were decreased by extended modified continuous cooking or oxygen delignification to kappa numbers as low as 15 were delignified by T. versicolor at the same rate as normal softwood pulp. More intensive O₂ delignification, like overcooking, decreased the susceptibility of the residual lignin in the pulps to degradation by T. versicolor.     

Extraction of xylans from hardwood chips prior to kraft cooking

The aim of this work is to define an extraction process of hemicelluloses from hardwood chips, prior to their transformation into paper pulp by the kraft process. Eucalyptus wood chips were submitted to autohydrolysis and acid hydrolysis treatments. Autohydrolysis means that no extra acid is added during the heating of the wood suspension, whereas sulphuric acid is added for the acid hydrolysis treatment. The liquor to wood ratio was 4, and temperature and time were varied so as to optimise the hemicelluloses extraction. Quantities of xylans varying between 12 and 38 g/l of hydrolysate (or between 9 and 15% on wood) could be extracted.Kraft cooking process was applied to the pre-hydrolysed wood chips. It was shown that they were easier to delignify than untreated wood chips, and the resulting pulp was also easier to bleach. The resulting cellulosic fibres were characterised for their papermaking properties: the higher the pentosan removal, the lower the strength properties of the pulps produced.     

The effects of hemicelluloses and lignin removal on water uptake behavior of hemp fibers

This study investigated the individual influences of hemicelluloses and lignin removal on the water uptake behavior of hemp fibers. Hemp fibers with different content of either hemicelluloses or lignin were obtained by chemical treatment with 17.5% sodium hydroxide or 0.7% sodium chlorite. Various tests (capillary rise method, moisture sorption, water retention power) were applied to evaluate the change in water uptake of modified hemp fibers. The obtained results show that when the content of either hemicelluloses or lignin is reduced progressively by chemical treatment, the capillary properties of hemp fibers are improved, i.e. capillary rise height of modified fibers is increased up to 2.7 times in relation to unmodified fibers. Furthermore, hemicelluloses removal increases the moisture sorption and decreases the water retention values of hemp fibers, while lignin removal decreases the moisture sorption and increases the water retention ability of hemp fibers.