Rate Of Cellulose Hydrolysis Research Articles

Effect of cellulose reducing ends and primary hydroxyl groups modifications on cellulose-cellulase interactions and cellulose hydrolysis.

Cellulose reducing ends are believed to play a vital role in the cellulose recalcitrance to enzymatic conversion. However, their role in insoluble cellulose accessibility and hydrolysis is not clear. Thus, in this study, reducing ends of insoluble cellulose derived from various sources were modified by applying reducing and/or oxidizing agents. The effects of cellulose reducing ends modification on cellulose reducing ends, cellulose structure, and cellulose accessibility to cellulase were evaluated along with the impact on cellulose hydrolysis with complete as well purified cellulase components. Sodium borohydride (NaBH4) reduction and sodium chlorite-acetic acid (SC/AA) oxidation were able to modify more than 90% and 60% of the reducing ends, respectively, while the bicinchoninic acid (BCA) reagent applied for various cycles oxidized cellulose reducing ends to various extents. X-ray diffractograms of the treated solids showed that these treatments did not change the cellulose crystalline structure and the change in crystallinity index was insignificant. Surprisingly, it was found that the cellulose reducing ends modification, either through selective NaBH4 reduction or BCA oxidation, had a negligible impact on cellulose accessibility as well on cellulose hydrolysis rates or final conversions with complete cellulase at loadings as low as 0.5 mg protein/g cellulose. In fact, in contrast to what is traditionally believed, modifications of cellulose reducing ends by these two methods had no apparent impact on cellulose conversion with purified cellulase components and their synergy. However, SC/AA oxidation resulted in significant drop in cellulose conversion (10%-50%) with complete as well purified cellulase components. Nonetheless, further research revealed that the cause for drop in cellulose conversion for the SC/AA oxidation case was due to primary hydroxyl groups (PHGs) oxidation and not the oxidation of reducing ends. Furthermore, it was found that the PHGs modification affects cellulose accessibility and slows the cellulase uptake as well resulting in significant drop in cellulose conversions.

Rice husk-based biogenic silica-anchored Sn-Co bimetallic catalysts for cellulose hydrogenolysis: Structural regulation and mechanism study

The production of high value-added chemicals and materials from renewable biomass is significant for the sustainable development of chemical industry. Herein, rice husk-based biogenic silica-anchored Sn-Co catalysts with adjustable activity are successfully prepared for biomass valorization. By regulating the pretreatment method and carbothermal reduction temperature of catalysts, the interaction between Co, Sn and biogenic silica can be regulated, thereby regulating the catalytic activity of the catalyst for cellulose hydrogenolysis. 50.7 % yield of acetol and 63.4 % yield of C3 products can be obtained. A series of characterizations show that the hydrothermal pretreatment in CoCl2 solution promotes the combination of Co2+ and biogenic silica, which inhibits the excessive doping of Sn4+ and promotes the formation of Co3Sn2 alloy. The Sn-O-Si and Co3Sn2 species in 3Sn-5Co@RHC are the key active sites catalyzing glucose isomerization and fructose retro-aldol condensation. The intermediates experiments show that controlling the hydrolysis rate of cellulose to avoid excessive concentration of monosaccharide intermediates in the reaction system is crucial for inhibiting side reactions and improving the yield of acetol. The use of agroforestry waste-based catalytic materials for biomass conversion in this work can provide inspiration for exploiting the intrinsic characteristics of biomass to develop high value-added materials and chemicals.

Spatiotemporal dynamics of cellulose during enzymatic hydrolysis studied by infrared spectromicroscopy

Spatial and temporal resolved mapping of infrared spectra of cellulose during enzymatic hydrolysis towards developing sustainable bioconversion processes to materials, fuels, and chemicals.

Biosynthesis of artificial starch and microbial protein from agricultural residue

Growing populations and climate change pose great challenges to food security. Humankind is confronting a serious question: how will we feed the world in the near future? This study presents an out-of-the-box solution involving the highly efficient biosynthesis of artificial starch and microbial proteins from available and abundant agricultural residue as new feed and food sources. A one-pot biotransformation using an in vitro coenzyme-free synthetic enzymatic pathway and baker’s yeast can simultaneously convert dilute sulfuric acid-pretreated corn stover to artificial starch and microbial protein under aerobic conditions. The β-glucosidase-free commercial cellulase mixture plus an ex vivo two-enzyme complex containing cellobiose phosphorylase and potato α-glucan phosphorylase displayed on the surface of Saccharomyces cerevisiae, showed better cellulose hydrolysis rates than a commercial β-glucosidase-rich cellulase mixture. This is because the channeling of the hydrolytic product from the solid cellulosic feedstock to the yeast mitigated the inhibition of the cellulase cocktail. Animal tests have shown that the digestion of artificial amylose results in slow and relatively small changes in blood sugar levels, suggesting that it could be a new health food component that prevents obesity and diabetes. A combination of the utilization of available agricultural residue and the biosynthesis of starch and microbial protein from non-food biomass could address the looming food crisis in the food–energy–water nexus.

Model of Processive Catalysis with Site Clustering and Blocking and Its Application to Cellulose Hydrolysis.

Interactions between particles moving on a linear track and their possible blocking by obstacles can lead to crowding, impeding the particles' transport kinetics. When the particles are enzymes processively catalyzing a reaction along a linear polymeric substrate, these crowding and blocking effects may substantially reduce the overall catalytic rate. Cellulose hydrolysis by exocellulases processively moving along cellulose chains assembled into insoluble cellulose particles is an example of such a catalytic transport process. The details of the kinetics of cellulose hydrolysis and the causes of the often observed reduction of hydrolysis rate over time are not yet fully understood. Crowding and blocking of enzyme particles are thought to be one of the important factors affecting the cellulose hydrolysis, but its exact role and mechanism are not clear. Here, we introduce a simple model based on an elementary transport process that incorporates the crowding and blocking effects in a straightforward way. This is achieved by making a distinction between binding and non-binding sites on the chain. The model reproduces a range of experimental results, mainly related to the early phase of cellulose hydrolysis. Our results indicate that the combined effects of clustering of binding sites together with the occupancy pattern of these sites by the enzyme molecules play a decisive role in the overall kinetics of cellulose hydrolysis. It is suggested that periodic desorption and rebinding of enzyme molecules could be a basis of a strategy to partially counter the clustering of and blocking by the binding sites and so enhance the rate of cellulose hydrolysis. The general nature of the model means that it could be applicable also to other transport processes that make a distinction between binding and non-binding sites, where crowding and blocking are expected to be relevant.

Effect of additives on the enzymatic hydrolysis of pre-treated wheat straw

Enzymatic hydrolysis of pre-treated wheat straw has been studied. Several additives have been checked as potential improvers of glucose yield in the enzymatic hydrolysis, trying to avoid the cellulases inhibition by the lignin as a cause of the decrease of the saccharification rate of lignocellulosic biomass. Non-ionic surfactants, such as Tween 80 (poly(oxyethylene)20-sorbitan-monooleate) and polyethylene glycol-6000, and proteins, as bovine serum albumin, casein and skimmed milk powder, have been used to improve the sugars yield in the enzymatic hydrolysis. Two pre-treated wheat straw materials, one with high delignification and another with low delignification, have been tested. Three concentrations of each additive have been considered. The results obtained point out that the lignin content of the pre-treated lignocellulosic biomass has a decisive influence in reaching a high yield in the enzymatic saccharification. The additives checked have a moderate effect on the cellulose hydrolysis rate. The more favorable option combines high delignification with the use of a cheap protein such as casein, reaching a glucose yield of 88.6% when the best conditions are employed.

Surface-Related Kinetic Models for Anaerobic Digestion of Mi-crocrystalline Cellulose: The Role of Particle Size.

In this work, for modelling the anaerobic digestion of microcrystalline cellulose, two surface-related models based on cylindrical and spherical particles were developed and compared with the first-order kinetics model. A unique dataset consisting of particles with different sizes, the same crystallinity and polymerisation degree was used to validate the models. Both newly developed models outperformed the first-order kinetics model. Analysis of the kinetic constant data revealed that particle size is a key factor determining the anaerobic digestion kinetics of crystalline cellulose. Hence, crystalline cellulose particle size should be considered in the development and optimization of lignocellulose pre-treatment methods. Further research is necessary for the assessment of impact of the crystalline cellulose particle size and surface properties on the microbial cellulose hydrolysis rate.

Correction: High enhancement of the hydrolysis rate of cellulose after pretreatment with inorganic salt hydrates

Correction for ‘High enhancement of the hydrolysis rate of cellulose after pretreatment with inorganic salt hydrates’ by Marta Lara-Serrano et al., Green Chem., 2020, 22, 3860–3866, DOI: 10.1039/D0GC01066A.

Is Steam Explosion a Promising Pretreatment for Acid Hydrolysis of Lignocellulosic Biomass?

For the production of sugars and biobased platform chemicals from lignocellulosic biomass, the hydrolysis of cellulose and hemicelluloses to water-soluble sugars is a crucial step. As the complex structure of lignocellulosic biomass hinders an efficient hydrolysis via acid hydrolysis, a suitable pretreatment strategy is of special importance. The pretreatment steam explosion was intended to increase the accessibility of the cellulose fibers so that the subsequent acid hydrolysis of the cellulose to glucose would take place in a shorter time. Steam explosion pretreatment was performed with beech wood chips at varying severities with different reaction times (25–34 min) and maximum temperatures (186–223 °C). However, the subsequent acid hydrolysis step of steam-exploded residue was performed at constant settings at 180 °C with diluted sulfuric acid. The concentration profiles of the main water-soluble hydrolysis products were recorded. We showed in this study that the defibration of the macrofibrils in the lignocellulose structure during steam explosion does not lead to an increased rate of cellulose hydrolysis. So, steam explosion is not a suitable pretreatment for acid hydrolysis of hardwood lignocellulosic biomass.

Synergy between endo/exo-glucanases and expansin enhances enzyme adsorption and cellulose conversion

Effective binding between cellulases and cellulose is essential for enzymatic hydrolysis of lignocellulose. Expansin can loosen the cellulose structure and can enhance the efficiency of cellulase. However, possible synergy between cellulases and expansin is not clear. In this work, the real-time adsorption of exoglucanases (Cel7A) or endoglucanases (Cel7B) with Bacillus subtilis expansin (BsEXLX1) and the enzymatic hydrolysis of cellulose were followed using quartz crystal microbalance with dissipation (QCM-D). Initial adsorption rate, adsorption capacity, and pseudo-steady-state rate of cellulose hydrolysis by Cel7A/Cel7B increased in the presence of BsEXLX1. When injecting Cel7A or Cel7B together with BsEXLX1 at a mass ratio of 1:1, the hydrolysis rate was almost 5 times the rate for Cel7A or Cel7B alone at 25 °C. These results increase our understanding of the real-time synergism between cellulases and expansin on cellulose, as well as the impact of their synergy on the enzymatic hydrolysis of cellulose.

Synergistic effect of ionic liquid and surfactant for enzymatic hydrolysis of lignocellulose by Paenibacillus sp. LLZ1 cellulase

Stable cellulases are critical for in situ hydrolysis of ionic liquid-pretreated lignocellulose, and surfactants were reported to improve the stability of cellulase in practical circumstances. In this study, synergistic effect of Tween-80 and [Emim]OAc was first identified to enhance the hydrolysis of lignocellulose by Paenibacillus sp. LLZ1 cellulase, proving the effectiveness of surfactants in in situ hydrolysis. Protein fluorescence and near-UV circular dichroism spectroscopy analyses illustrated that ionic liquids might move local residues and alter global tertiary structure of cellulase, leading to higher instability. The addition of Tween-80 might turn the microenvironment of cellulase residues more hydrophilic and reduce the contact of protein with deactivation factors. The synergistic system was further applied in the in situ hydrolysis of microcrystalline cellulose and bagasse cellulose by Paenibacillus cellulase. The enzymatic hydrolysis rate of microcrystalline cellulose was enhanced by 2.5-fold and the final conversion was improved by 45%. Scanning electron microscopy demonstrated the substrate solubilizing effect during reaction by monitoring the morphological changes of lignocellulosic substrates. This study highlighted the potential of applying synergistic reaction system in in situ enzymatic hydrolysis of lignocellulose, and revealed the inherent mechanism initially.

Dilute alkali pretreatment and subsequent enzymatic hydrolysis of amur silvergrass for ethanol production

A dilute alkali pretreatment (NaOH) was used to remove lignin and some hemicelluloses, as well as to efficiently increase the accessibility of enzymes to the cellulose in Amur silvergrass. A single factor experiment was designed with 4 factors (1 to 5% w/w NaOH, 1/6 to 1/14 solid to liquid ratio, 15 to 90 min residence time, and 80 to 125 °C digestion temperature) with 3 duplicates of 5 levels for each factor. On the basis of the single factor test, an L8 (24)-orthogonal experiment was conducted to identify the main influencing factor and the optimal factor combinations verified by an enzymatic hydrolysis and fermentation experiment. The main factors influencing ethanol production were NaOH concentration and digestion temperature, while residence time and solid to liquid ratio had a lesser effect. The enzymatic hydrolysis rate of cellulose reached 82.6%, and the highest conversion rate of ethanol was 78.3% with 4.0% (w/w) NaOH and a 1:6 solid to liquid ratio at 100 °C for 15 min. Scanning electron microscope (SEM) images of the lignocellulosic surface structure of non-pretreated and optimum pretreated Amur silvergrass displayed obvious differences. The lignin was the key recalcitrance-causing factor for ethanol production, which can be effectively removed by the NaOH.

High enhancement of the hydrolysis rate of cellulose after pretreatment with inorganic salt hydrates

Cellulose treatment dissolution/precipitation in inorganic salt hydrates produces a dramatic morphology change that yields a reactivity improvement respect the untreated for transformations.

Modification of microcrystalline cellulose structural properties by ball-milling and ionic liquid treatments and their correlation to enzymatic hydrolysis rate and yield

This study investigated the characteristics of cellulose Avicel PH-101 (NPA) and Avicel samples treated by either ball milling (BMA) or ionic liquid 1-ethyl-3-methylimidazolium acetate (ILA), which alter cellulose structural properties specifically and, consequently, affect the enzymatic hydrolysis of these materials differently. All materials were submitted to 48 h hydrolysis using the commercial enzymatic preparations Celluclast 1.5 L and Novozymes N188. For ILA hydrolysis, a high yield of 96.8% was observed, which was related to its high specific surface area, low cellulose crystallinity, and high cellulose II content rather than to the particle size. The hydrolysis yields of BMA and NPA were 74.4% and 41.6%, respectively. BMA crystallinity was higher in comparison to ILA and lower in comparison to NPA, whereas the BMA surface area was lower in comparison to ILA and NPA. X-ray diffraction (XRD) analysis suggested the formation of cellulose II by both pretreatments on Avicel, which was reinforced by the samples’ diffractograms and an additional characterization by XRD/crystallinity index of the partial hydrolyzed BMA insoluble residue. In the presence of amorphous cellulose, cellulose II allomorph presented higher hydrolysis recalcitrance. The set of results showed that crystallinity and surface area provided a straightforward correlation with the hydrolysis rate and yield of Avicel cellulose. In addition, the understanding of the effects that can be caused by pretreatments of cellulosic materials was improved and these findings contribute with the interpretation of the kinetics and yields of enzymatic hydrolysis.

Catalytic hydrolysis of cellulose by phosphotungstic acid-supported functionalized metal-organic frameworks with different electronegative groups.

It is found that strong electronegative groups can selectively adsorb cellulose by hydrogen bonds. Grafting strong negatively charged groups onto catalysts to achieve the functionalization of the catalyst can give it the ability to selectively adsorb cellulose without affecting its catalysis, which is of great significance for the hydrolysis of cellulose. In this study, PTA@MIL-101-X (X = -Br, -NH2, -Cl, -NO2) materials were synthesized to investigate the effect of grafting different electronegative groups on carriers to the directional hydrolysis of cellulose. The synthesized catalysts used phosphotungstic acid as the catalytic center while treated MIL-101 structure as the carrier. The grafting of different electronegative groups changed the crystal structure of the metal organic framework without affecting its stability during the reaction. The strong negative functional groups can selectively adsorb cellulose by forming hydrogen bonds with cellulose hydroxyl groups and weaken the hydrogen bonds within cellulose molecules. This hydrogen bond can reduce the side reaction of glucose, lighten the difficulty of cellulose hydrolysis, and improve the efficiency of cellulose conversion at the same time. The hydrolysis rate of cellulose increased with the electronegativity enhancement of the grafted functional groups, and the grafted -NO2 catalyst PTA@MIL-101-NO2 obtained the highest glucose yield of 16.2% in the cellulose-directed hydrolysis. The -NH2 can form a chemical linkage with PTA through electrostatic interaction to get the highest immobilization stability and exhibit excellent stability in the recycling of catalysts. Graphical abstract.

Humic acid inhibition of hydrolysis and methanogenesis with different anaerobic inocula

There is increasing evidence that humic acid (HA) is hampering the performance of anaerobic digesters treating animal manures and thermally-hydrolysed waste activated sludge. In the present study, HA inhibition and inhibition resilience was examined for hydrolysis (carbohydrate and protein) and acetotrophic methanogenesis with four distinct full-scale anaerobic inocula. The aim was to further understand HA inhibition and to explore potential relationships between microbial factors and inhibition resilience. For two of the four tested inocula, cellulose degradation showed a start-up delay that lengthened as HA concentration increased from 0 to 2 g L−1. This inhibition was reversible because, after the initial delay, subsequent hydrolysis rates and methane yields were not significantly influenced by HA concentration. Cellulose hydrolysis results at HA concentrations below 2 g L−1 support a threshold inhibition mechanism, i.e. HA complexes with hydrolytic enzymes preventing them from binding with cellulose, but once all the HA had been complexed, enzymes subsequently released are free to bind with cellulose. Inocula with higher cellulose hydrolytic activity were less affected by HA inhibition, suggesting a potential link between HA inhibition resilience and microbial activity. However, above 5 gHA L−1, cellulose hydrolysis rates decreased with increasing HA concentration; indicating that the mechanisms of inhibition may change depending on some threshold HA concentration. Protein hydrolysis and acetotrophic methanogenesis were less susceptible to HA inhibition than cellulose hydrolysis, since signs of inhibition were only observed above 5 gHA L−1. Acetotrophic methanogenesis was partially inhibited at 10 gHA L−1 and completely inhibited at 20 gHA L−1. These results further support that HA inhibition is selective towards particular enzymes.

The impact of kinetic parameters on cellulose hydrolysis rates

Abstract Mechanistic kinetic models are vital tools to elucidate rate-limiting interactions and processes hampering complete hydrolysis of cellulose by cellulases. Properly formulated, the models should ultimately simulate experimental cellulose hydrolysis time courses given that model parameters are known a priori. Here, we review reported values of critical parameters in cellulose hydrolysis (catalytic, adsorption/desorption, complexation/decomplexation rate constants and apparent processivity) and examine how estimation methods may impact the magnitudes and interpretation of the values. The sensitivity of the extents and rates of cellulose hydrolysis to the range of reported values were examined by conducting simulations with a general mechanistic model of TrCel7A hydrolysis of cellulose. When the simulation was conducted at enzyme-limiting conditions, initial bursts in cellulose hydrolysis rates were evident and highly sensitive to specific hydrolysis rates of productively bound enzymes and dissociation rates of non-productively bound enzymes. Under substrate limiting conditions, hydrolysis rates were significantly reduced and no initial burst was observed, with more pronounced sensitivity of hydrolysis rates to the range of kinetic values. Enzyme-substrate interaction rate constants reviewed here only capture short-time hydrolysis rate outcomes and future work to elucidate mechanisms of cellulose hydrolysis must also incorporate how substrate-limitations evolve over the course of hydrolysis.

Modelling the specific pathway of CH4 and CO2 formation using carbon isotope fractionation: an example for a boreal mesotrophic fen

ABSTRACTWe described mathematically the process of peat methanization in a boreal mesotrophic fen. Gaseous and dissolved CH4 and CO2 as well as their δ13C signatures were considered in the dynamic equations for incubation bottles. In accordance with the model, acetate, H2, and CO2 were produced during cellulose hydrolysis and acidogenesis. 13C/12C in CO2 was a key variable reflecting dynamic changes in the rates of cellulose hydrolysis and acidogenesis, acetoclastic and hydrogenotrophic methanogenesis. As CO2 is the substrate in hydrogenotrophic methanogenesis, δ13C–CO2 increased from the start till the dissolved hydrogen concentration became very low. Thereafter, the rate of acetoclastic methanogenesis with the significant current acetate concentration dominated over the rate of hydrogenotrophic methanogenesis leading to the decreasing δ13C–CO2 and the increasing δ13C–CH4. The model was validated by describing the system’s dynamics under strong and weak inhibition of acetoclastic and hydrogenotrophic methanogenesis by methyl fluoride, respectively. During peat methanization at the lowered temperature of 10 °C, the processes of hydrogenotrophic methanogenesis and homoacetogenesis competing for H2 may occur. However, based on dynamics of the carbon isotope signatures, especially on dynamics of δ13C–CO2, the model showed no significant contribution of homoacetogens in peat methanization.

Can We Reduce the Cellulase Enzyme Loading Required To Achieve Efficient Lignocellulose Deconstruction by Only Using the Initially Absorbed Enzymes?

The cost-effective production of sugars from biomass continues to be challenging, partly due to the relatively high enzyme/protein loading required to achieve effective hydrolysis of the insoluble polysaccharides within the pretreated lignocellulosic substrates. Previous work has suggested that those enzymes that initially, strongly adsorb to the insoluble substrate are crucial for effective cellulose hydrolysis. However, most previous work in this topic area has used either purified enzymes or “older” generations of cellulase preparations acting on “model” cellulosic substrates. The results were, in several cases, contradictory or inconclusive. In the work reported here, the roles and functions of the initially adsorbed enzymes in determining the rate and extent of cellulose hydrolysis were assessed when using several different pretreated biomass substrates and the Novozyme enzyme preparation, Cellic CTec3. It was apparent that the initially adsorbed enzymes (irreversible bound to substrate after centrif...

Highly Efficient Conversion of Xylose Residues to Levulinic Acid over FeCl3 Catalyst in Green Salt Solutions

The economically viable synthesis of levulinic acid (LA), a promising and valuable renewable biomass-derived platform for bioproducts, with high carbon efficiency is a challenge. A direct and highly effective catalytic system for conversion of xylose residues (XRs) into LA under mild conditions by using FeCl3 as catalyst and cheaply available NaCl as promoter has been developed. The NaCl solution exhibits high carbon efficiency in LA (68.0 mol %) when compared with the non-NaCl systems (48.5 mol %) due to the moderate increase of the acidity and the higher viscosity of the NaCl system than water. The experimental results demonstrated that the presence of NaCl caused no distinctive changes on reaction pathways but increased the dissolution rate and the hydrolysis rate of XRs cellulose. Moreover, further integration of our degradation process with a reactive extraction step makes energy-efficient separation of LA. The NaCl solutions easily and efficiently extracted LA into LA-derived solvent 2-methyltetrahy...