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Inter- and intrachain hydrogen bonding pattern on {100} cellulose surface. Dashed lines denote possible hydrogen bonds, with intrachain bonds in blue and interchain bonds in green. In this work the hydroxyl groups of O2, O3, and O6 oxygens are referred to as O2H2, O3H3, and O6H6 respectively. The hydrogen bond decomposition consideration is limited to hydrogen bonding of the three hydroxyl groups within the cellulose surface layer
Source publication
Enzymatic cleavage of glycocidic bonds is an important, green and biocompatible means to refine lignocellulosic biomass. Here, the effect of the resulting oxidation point defects on the structural and water interactions of crystalline cellulose {100} surface are explored using classical molecular dynamics simulations. We show that even single oxida...
Citations
... In a study by Sharifzadeh et al. [36], a regenerated cellulose/Hal nanocomposite was prepared in EMIMCL, with no noticeable aggregation observed at low Hal contents. Additionally, oxidation on cellulose can improve cellulose solubility as glycosidic bonds are damaged, allowing the broken bonds to interact with the surrounding water, thereby enhancing water solubility [37]. ...
... These peaks indicate water ordering caused by the presence of the cellulose surface. Immobilization and ordering of water molecules on the CNC surface have been shown before both by experimental and simulations studies (Lindh et al. 2017;Malm et al. 2010;Yurtsever et al. 2022;Heiner et al. 1998;Matthews et al. 2006;Biermann et al. 2001;Maurer et al. 2013;Mudedla et al. 2021). ...
Crystalline nanocellulose is widely used for example, in the paper-making and food industries, as support matrix material or reinforcement of polymer materials, but also in drug carrier and nanomedicine applications. Interestingly, aqueous solutions of cellulose are extremely sensitive to small amounts of added salt yet mere considerations of charge screening leave open questions regarding the mechanisms, especially for unmodified cellulose in aqueous solutions. Here, we map NaCl ion distributions and the effect of added NaCl salt on the hydration of Iβ cellulose nanocrystal (CNC) surfaces by atomistic detail molecular dynamics simulations with explicit water solvent. The simulations reveal the dependency of the hydration layers of the six surfaces of CNCs on the ions, as well as NaCl ion binding sites, and preferences in terms of binding free energy for the ions near CNC surfaces at different NaCl concentrations. We discuss the modelling results against our prior rheology characterization of cellulose solutions. Together, the results indicate that the high sensitivity of cellulose aqueous solutions to added salt rises from the ions near the surface changing locally the ordering and structure of the hydration layers of the CNC surfaces. The revealed mechanism of salt-induced viscosity changes in cellulose aqueous solutions allows advanced design of gelling CNC systems for various end uses and may also guide tuning cellulose interactions by different solvent environments.
... Instead, it was suggested that LPMO promotes time-dependent decrystallization of the substrate that improves accessibility for the classical hydrolytic enzymes [40]. Indeed, several studies support the notion that LPMO action promotes decrystallization of cellulose [41][42][43][44]. Recently, Cannella et al. [45] showed that oxidation of filter paper with an LPMO, or chemically, using TEMPO [(2,2,6,6-tetramethylpiperidin-1-yl)oxyl] increases the amount of water retained by the fibers, due to the increased negative surface charge. ...
... It is generally believed that LPMOs help break down these recalcitrant structures by introducing oxidative modifications, creating new sites of accessibility that enable cellulases and other enzymes to continue degrading the substrate. Importantly, recent studies indicate that the LPMO-cellulase synergism may be more complex than creating access points [40][41][42][43]45]. The cleavage of a glycosidic bond and concomitant oxidation of the cleavage point allows surrounding water molecules to access the highly ordered fibril structure, leading to decrystallization and amorphization over time [45]. ...
... The extent of these larger, and potentially slower, effects will likely vary between C1-and C4-oxidizing LPMOs. Generation of aldonic acids (C1 oxidation) is thought to have the largest effect due to the open ring structure allowing more water to penetrate the crystalline structure [41][42][43]. On the other hand, recent work by Angeltveit has shown that, with time, the increase in overall accessibility of the substrate for the traditional hydrolytic enzymes will be governed by a timedependent non-enzymatic decrystallization phase that follows the oxidative action of LPMOs and that does not clearly depend on the oxidative regiospecificity of the enzymes [40]. ...
Background
The polysaccharides in lignocellulosic biomass hold potential for production of biofuels and biochemicals. However, achieving efficient conversion of this resource into fermentable sugars faces challenges, especially when operating at industrially relevant high solid loadings. While it is clear that combining classical hydrolytic enzymes and lytic polysaccharide monooxygenases (LPMOs) is necessary to achieve high saccharification yields, exactly how these enzymes synergize at high solid loadings remains unclear.
Results
An LPMO-poor cellulase cocktail, Celluclast 1.5 L, was spiked with one or both of two fungal LPMOs from Thermothielavioides terrestris and Thermoascus aurantiacus, TtAA9E and TaAA9A, respectively, to assess their impact on cellulose saccharification efficiency at high dry matter loading, using Avicel and steam-exploded wheat straw as substrates. The results demonstrate that LPMOs can mitigate the reduction in saccharification efficiency associated with high dry matter contents. The positive effect of LPMO inclusion depends on the type of feedstock and the type of LPMO and increases with the increasing dry matter content and reaction time. Furthermore, our results show that chelating free copper, which may leak out of the active site of inactivated LPMOs during saccharification, with EDTA prevents side reactions with in situ generated H2O2 and the reductant (ascorbic acid).
Conclusions
This study shows that sustaining LPMO activity is vital for efficient cellulose solubilization at high substrate loadings. LPMO cleavage of cellulose at high dry matter loadings results in new chain ends and thus increased water accessibility leading to decrystallization of the substrate, all factors making the substrate more accessible to cellulase action. Additionally, this work highlights the importance of preventing LPMO inactivation and its potential detrimental impact on all enzymes in the reaction.
... Its chemical composition consists of D-glucose units that are linked with β-(1-4)-glycosidic linkages [83,84]. In plants, cellulose is surrounded by lignin and hemicellulose and contains β-(1-4)-glucose chains in both crystalline order and amorphous regions [85]. Cellulose is insoluble in water mainly due to two reasons: (1) it is tightly packed due to inter-chain hydrogen bonding and hydrophobic interactions [86], and (2) the crystalline regions of cellulose fibers make it thermodynamically stable [87]. ...
Decellularization of plant tissues is an emerging route to fabricate scaffolds for tissue engineering and regenerative medicine. Although significant progress has been made in the field of plant tissue decellularization, functionalization of plant scaffolds is still an emerging field, and loading them with L-ascorbic acid to promote skin regeneration has not yet been reported. L-ascorbic acid is an antioxidant that plays a key role in collagen synthesis as a cofactor of lysyl hydroxylase and prolyl hydroxylase. It has been shown to have significant importance in physiological wound healing by stimulating fibroblasts to produce collagen at both the molecular and the genetic levels. In this work, we aimed to fabricate an ascorbic acid-releasing bioactive scaffold by introducing a stable form of ascorbic acid, L-ascorbic acid 2-phosphate (AA2P), into decellularized baby spinach leaves and investigated its biological activity in vitro. Our results demonstrated that AA2P could be easily introduced into decellularized baby spinach leaf scaffolds and subsequently released within the effective dose range. AA2P-releasing baby spinach leaves were found to increase metabolic activity and enhance collagen synthesis in L929 fibroblasts after 21 days. In conclusion, this study demonstrated the fabrication of a novel functionalized skin tissue engineering scaffold and made a significant contribution to the fields of plant decellularization and skin tissue engineering.
Graphical abstract
... This allows cellulose to be modified easier with other materials. Aldehyde groups can form bonds with polymers bearing amine groups (Mudedla et al. 2021;Zhang et al. 2018;Hou et al. 2018). ...
Bacterial cellulose (BC) is a biomaterial extensively studied in tissue engineering due to its favorable properties. Porosity, biocompatibility, biodegradability and mechanical durability are essential material properties for scaffold use in tissue engineering. This study aims to fabricate porous scaffolds using a moldable and degradable BC-HAp composite for bone tissue engineering. BC was produced by Komagataeibacter sucrofermentans under static culture conditions. The harvested BC membranes were purified and then mechanically shredded. BC oxidation was performed using different sodium periodate concentrations (0.05–0.5 M) and treatment times (0.5–12 h). Oxidized BCs (oxBC) were modified with hydroxyapatite (HAp), then were moulded, lyophilized, and characterized. The degradability of the scaffolds was determined for 45 days. Cytotoxic analysis of oxBC scaffolds was carried out for 7 days using the L929 fibroblast cell line. The oxidation degrees of the shredded BC samples were between 6.75 and 81%, which increased in line with the increasing concentration and application time of periodate. The scaffolds prepared using oxidized cellulose for 30 and 60 min (oxBC30 and oxBC60) preserved their integrity, These scaffolds showed a weight loss of 9% and 14% in 45 days, respectively. The pore distribution was between 50 and 450 µm and concentrated in the 50–150 µm range. The compression moduli were 88.72 kPa and 138.88 kPa for oxBC30-HAp and oxBC60-HAp, respectively. It was determined that oxBC did not show a significant difference in cell viability compared to the control groups and was not cytotoxic. In conclusion, degradable and more porous bone scaffolds were fabricated using mouldable oxBC.
... Wetting is governed by the properties of both the solid substrate and the liquid. While the CHARMM carbohydrate forcefield employed in this work has been shown previously to satisfactorily predict properties of both isolated glucan chains (Guvench et al. 2009) and cellulose crystal structures (Oehme et al. 2018;Matthews et al. 2012;Beckham et al. 2011;Payne et al. 2011;Mudedla et al. 2021), the liquid models were subject to validation by comparing their Fig. 3 The evolution of contact angles for water (SPC/E force field) with time on native and acetylated cellulose surfaces. Blue lines show the instantaneous values and black lines are ten-nanosecond running averages. ...
Wetting of cellulose by different liquids is interesting from the point of view of the processing of cellulose-based nanomaterials. Here, the contact angles formed by water and several organic liquids on both native and acetylated cellulose were calculated from molecular dynamics simulations. It was found that liquid surface tension was crucial for their wetting behavior. Acetylation decreases the work of adhesion to most liquids investigated, even non-polar ones, while others are not affected. Water has the highest affinity to cellulose, both native and acetylated. The results have implications for liquid infiltration of nanocellulose networks and the interaction of cellulose with different liquids in general.
... Some studies suggest that oxidative treatments (such as TEMPO-mediated oxidation) under certain experimental conditions can alter the amorphous cellulose regions while maintaining the crystalline domains and lead to increases in CrI [54,55]. However, other authors point out that oxidative action may cause scission of glycosidic bonds, inducing structural changes in the crystalline domains of cellulose, and decreasing crystallinity [14,56]. Therefore, further studies are needed to investigate any potential changes in cellulose crystallinity and crystal size that may occur as a result of oxidative events within the HC reactor, and how these changes could impact the structure of cellulose nanocrystals. ...
Enzymatic hydrolysis to produce cellulose nanocrystals (CNCs) offers economical and sustainability advantages. However, since this is still an emerging technology, further improvements are needed to make the process feasible for scale-up. In our study, we propose a novel approach using hydrodynamic cavitation (HC) as a pretreatment technology to enhance the efficiency and yield of CNC production. HC treatments conducted under various scenarios induced structural modifications of the cellulose fibers that facilitated their defibrillation and subsequent enzymatic hydrolysis. Isolation through hydrolysis involved the synergistic action of specific enzymes, including xylanase and endoglucanase. Xylanase acted on the hemicellulosic fraction, breaking down the xylan structure and exposing the cellulose fibers. Subsequently, endoglucanase targeted the amorphous regions of cellulose, releasing CNCs. By integrating the HC pretreatment with enzymatic hydrolysis, we achieved remarkable results. High-yield CNCs, reaching approximately 60%, were obtained with a crystallinity index ranging from 81% to 85%. Moreover, this process demonstrated a significant reduction in total energy consumption of approximately 56%, contributing to its economic viability. Furthermore, CNCs with diverse morphologies, aspect ratios, and viscosity profiles were produced, enabling the tailoring of their properties for specific and new applications.
... Studies on fiber structural integrity upon LPMO oxidation (Villares et al. 2017) and mathematical modeling (Vermaas et al. 2015;Trentin and Skaf 2019;Mudedla et al. 2021) have speculated that water could be strongly retained or bound in the proximity of the cellulose oxidation sites. The otherwise structured layer of water could also be disrupted around the structural "defects" introduced by polysaccharide chain breaking, affecting in some extent the binding of water. ...
... The oxidation introduced by LPMOs or TEMPO although differing in the carbon position might cause structural reassessments of the rod-shaped fibers, thus likely influencing the water:cellulose interaction. Similar conclusions were indicated by mathematical modeling studies when comparing C1-oxidized cellulose surfaces against pristine cellulose (Mudedla et al. 2021). ...
The cellulose-water interface is a dynamic environment mostly dominated by interactions between water molecules and hydroxyl groups protruding from the top layer of the polysaccharide chains. This interface has attracted increasing interest within the context of hydrolysis with glycosyl hydrolases, and studies on the role of tightly bound and free water has emerged. At the molecular level, cellulose-bound water has been considered important to allow enzymatic hydrolysis at industrial relevant conditions, i.e. at high dry matter (HDM) contents. In the presence of lytic polysaccharide monooxygenase enzymes, the hydrolysis can with effective yields be run at well beyond the dry matter limit previously set by the 1st generation of enzyme preparations lacking LPMOs. The oxidative cleavage of the cellulose chain performed by LPMOs allow a higher level of synergism with GH in terms of accessibility of the cellulose surface. In this work, we studied how cellulose oxidation by LPMO increases the cellulose-water interaction and the impact of this on cellulose saccharification. Low-field NMR, water constraint and enzyme kinetics at high dry matter contents were used to characterize the cellulose-water interaction and its implications in enzymatic cellulose hydrolysis.
... It compares the intensity of the highest peak (I200) and the local minimum Imin in the diffraction pattern 2θ range of 17−19°. According to recent literature data reported by [83], and Redlinger-Pohn (2022) [81], the objective values of crystallinity index (CrI) from XRD can be measured using modern calculation methods such as a deconvolution approach (fitted peaks). The latter takes into account the amorphous model and compares the areas of the peaks [81]. ...
The aims of this study are to investigate the structure of four historical Moroccan cedar softwood samples of different aging time duration (16th, 17th, 19th, 21st centuries) and compare among these four samples, using two analytical methods, FTIR and XRD, in order to confirm some structural changes and determine the degree of deterioration. The pronounced hemicellulose deterioration was highlighted by a breakdown of IR acetyl groups at 1738 cm−1 from the 19th century sample until aged ones. The cellulose XRD crystallinity index showed an important decrease from recent to oldest samples (51.8 to 20.2%) justifying the damages mainly in the two oldest samples (17th and 16th centuries), also confirmed by FTIR. The alteration of lignin was manifested in the case of the two ancient samples (16th and 17th centuries), proven by the decrease in IR bands related to aromatic nuclei (1595, 1500, 1230 cm−1) evolving towards a new diconjugate C=O formers at 1647 cm−1 (quinone, Ar-CO-Ar, Ar-CO-C=C). For accurate elucidation, the data of two combined techniques were compared and correlated. The obtained results depended on the part of the wood exposed to weathering effects (internal or external) and were influenced by both extended time of aging and effects of natural deterioration agents. The effects of natural aging were investigated in four historical Moroccan cedar softwood samples (16th, 17th, 19th, 21st centuries) using two analytical tools: FTIR and XRD. The pronounced hemicellulose deterioration was highlighted by a breakdown of IR acetyl groups at 1738 cm−1 and declines in the absorption signal at 1268 cm−1 from the 19th century sample until aged ones. The cellulose XRD crystallinity index (CrI) estimation showed an important decrease from recent to oldest samples (51.8 to 20.2%) justifying the damages mainly in the two oldest samples (17th and 16th centuries). These data were also confirmed by FTIR showing a significant reduction in both area profiles of C-O-C (1150–1000 cm−1) and C-H crystalline cellulosic bands (1375, 1318, and 1268 cm−1), respectively. The lignin alteration in both old samples (16th and 17th centuries) was proven by the decrease in IR aromatic skeleton (1595, 1500, and 1230 cm−1) evolving towards a new diconjugate C=O formers at 1647 cm−1 (quinone, Ar-CO-Ar, Ar-CO-C=C). To determine the structural difference and the degree of deterioration, the IR area of C=O band intensities ranging from 1550 to 1800 cm−1 was exploited. For accurate elucidation, the data of two combined techniques were compared and correlated. The obtained results depended on the part of the wood (internal or external) exposed to weathering effects and were influenced by both extended time of aging and effects of natural deterioration agents.
Keywords: cedar softwood (Cedrus atlantica); natural aging; wood degradation; IR spectroscopy; XRD; cellulose crystallinity index (CrI); lignocellulose
... Our simulations showed that, in the absence of NaCl, the ordering of water is affected by the cellulose surface to approximately 1 nm distance from the surface (Fig. 6a), which is consistent with previous works (Heiner and Teleman 1997;Heiner et al. 1998;Hakalahti et al. 2017;Mudedla et al. 2021). The addition of NaCl does not significantly influence this ordering of water, as seen in the comparison of water density curves in Fig. 6a, which is consistent with the low number of ions bound. ...
Hydrogels formed by cellulose nanofibers (CNFs) find use in a variety of applications. CNF hydrogels generally stiffen and ultimately flocculate with increasing salt concentrations. While charge repulsion explains the behavior of nanocellulose variants that have been stabilized by charged groups, it has been a puzzle why ions have such a pronounced effect also on CNFs with unmodified surfaces. We studied the effect of ionic solutes on native CNF hydrogels, and found that already at very low concentrations of around 1 mM, ions cause crowding of the hydrogels. The ionic solutes used were NaCl, Na2SO4, NaI, NaSCN, and sodium acetate. For the hydrogels, we used low densities of CNFs which lead to relatively weak gels that were highly sensitive to salts. Screening of the electrical double layer could not explain the results at such low ion concentrations. To understand cellulose-ion interactions, we used computational molecular dynamics simulations. The results provide an explanation by the effect of ions on the structure of the hydration layers of the cellulose. Understanding how and why ions affect the properties of native CNF hydrogels can help in for example manufacture of CNFs and when using CNFs as material components, substrates for enzymes, or as rheology modifiers. Ion-effects on the hydration layer of cellulose may also be important for more fundamental understanding of interfacial interactions of cellulose with water under different conditions.
Graphical abstract
