Cellulase Hydrolysis Research Articles

Green utilization of sorghum straw by coupled hydrothermal pretreatment and cellulase hydrolysis for bacillomycin D production under fed-batch mode

The present work explored the coupled hydrothermal pretreatment and cellulase hydrolysis (HPCH) for production of bacillomycin D from sorghum straws by Bacillus subtilis NS-174. The results indicated that HPCH treated sorghum straws had weaker absorption, heavier mass loss and higher crystallinity, and displayed loose and destructive fiber structures. Degradation of hemicellulose and lignin contributhed to yielding glucose and 68.94g/L of glucose was obtained in HPCH treated hydrolysate. High level of glucose in pretreated sorghum straw hydrolysate was helpful for improving bacillomycin D production. When performed by 3rd round (feeding at 144h) fed-batch fermentation, bacillomycin D production and production rate were obtained to be 2351.07mg/L and 113.36mg/L.h in B. subtilis NS-174, respectively.

Unlocking full potential of bamboo waster: Efficient co-production of xylooligosaccharides, lignin, and glucose through low-dosage mandelic acid hydrolysis with alkaline processing.

Mandelic acid (MA), a natural and environmentally friendly organic acid, demonstrates high selectivity and efficiency in hydrolyzing hemicellulose, making it an excellent candidate for xylooligosaccharides (XOS) production at low acid dosages. Despite its potential, the application of MA for XOS production has not been evaluated. The study first investigated the effectiveness of MA in hydrolyzing hemicellulose in bamboo into XOS. Under optimized conditions (50 mM MA, 180 °C, 45 min), a high XOS yield of 65.9 % was achieved, with a total xylobiose and xylotriose yield of 43.5 %. Subsequent alkaline pretreatment enabled 92.1 % lignin removal from MA-pretreated bamboo. The recovered lignin exhibited a high purity of 95.2 % and retained fundamental structure and functional groups of native lignin. The resulting residue displayed enhanced crystallinity and accessibility, with reduced hydrophobicity and surface area lignin compared to untreated bamboo. At high substrate concentration of 20 %, cellulase hydrolysis resulted in a glucose conversion efficiency of 83.9 %. Overall, this integrated strategy offered an efficient approach for the co-production of valuable XOS, lignin, and glucose from bamboo. The efficient energy utilization and economic viability further highlight the potential of this method for large-scale industrial applications, making it an attractive option for biomass valorization.

Improvement of okara noodle quality by modifying the soluble/insoluble dietary fibre ratio

This study investigated the effect of okara modified through cellulase hydrolysis and extrusion on noodle quality. Modification increased the soluble dietary fibre/insoluble dietary fibre (SDF/IDF) ratio in okara, improved appearance, cooking, and texture, and reduced starch digestibility of okara noodles. The 4.0 % cellulase enzymolysis-extruded okara noodles exhibited the quality closest to that of wheat noodles, with an estimated glycaemic index (eGI) < 55 (low-GI). As the okara SDF/IDF ratio increased, the water mobility of noodles decreased, indicating that an increase in the SDF/IDF ratio reduced competitive water absorption of okara. In addition, increased SDF/IDF ratio increased β-sheet content and promoted the enhanced hydrogen bond interactions between proteins and polymerisation between gliadin and glutenin. Moreover, the microstructure of noodles with a higher SDF/IDF ratio of okara was more continuous and compact, further confirming the promotional effect of okara with a higher SDF/IDF ratio on the quality of okara noodles.

Enhancement of hydrogen production in dark fermentation of corn stover hydrolysates through composite electric field pretreatment: Improving enzymatic efficiency and regulating microbial metabolic balance.

This study investigates the effects of composite electric field pretreatment (CEP) on hydrogen production from corn stover enzymatic hydrolysates in dark fermentation. The findings reveal that under optimal conditions, the CEP group achieved a cumulative hydrogen yield of 77.3 ± 2.6 mL/g TS, marking a 55.3 % increase compared to the control group without the electric field. CEP significantly enhances the fermentation capacity of enzymatic hydrolysates during the acid-stage of dark fermentation by disrupting the lignin structure and optimizing cellulase hydrolysis efficiency. Additionally, pH self-regulation is facilitated through the interaction between volatile fatty acids (VFAs) and ammonia nitrogen. Microbial community analysis revealed that CEP shifts the metabolic balance between Clostridium_sensu_stricto_1 and Lactococcus, leading to increased hydrogen yield and concentration during acid fermentation. Notably, Terrisporobacter exhibited a superior acid-producing capability compared to Bacteroides during the dark fermentation of enzymatic hydrolysates. This study provides a new perspective for the practical application of corn stover.

Improving inter-fiber bonding by cellulase under a high surface lignin level

Lignin is behaving negatively in cellulase hydrolysis, leading to the high cost of cellulase in many areas. Instead of removing lignin, the fiber surface composition was changed in structure and distribution in this work through a light mechanical refining. 2 h of cellulase adsorption and hydrolysis were performed after that. The cellulase hydrolysis was found to be improved with the glucose production increasing from 0.031 to 0.074 g L−1. Hydrolyzed fibers were then made into handsheet for inter-fiber bonding properties characterization. With most lignin retained and more hydrogen bonding formed, both tensile index and zero-span tensile index were found to be improved after the cellulase modification. It can be concluded that the cellulase hydrolysis of fibers is positively correlated with the inter-fiber bonding properties.

Understanding the role of porous particle characteristics and water state distribution at multi-scale variation on cellulase hydrolysis of lignocellulosic biomass

Porous particle characteristics of lignocellulosic biomass and the water state distribution surrounding particles determine enzyme/solute transfer, while inherently affecting the liquefaction and hydrolysis reactions. For submillimeter biomass particles (<500 μm), deep eutectic solvent pretreatment tended to create additional macropores (∼4%, pore volume of 4.3 mL/g) and micropores (∼3%, pore area of 2.26 m2/g) at the extragranular and cell wall scales, simultaneously with reducing particle size by 15%. The pretreated biomass exhibited preferential rapid liquefaction over sugar production, leading to a ∼70% reduction in particle size only after 3 h hydrolysis. This further increased particle porosity (∼85% after 12 h hydrolysis) with the transition from large surface-pores to interior micropores. The increases in the tissue/cell-scale pore size and intraparticle area had greater significance for improving hydrolysis performance than particle size reduction. Additionally, particle liquefaction released constrained water in capillary pores and enhanced slurry fluidity; capillary water and free water became main transfer carriers during the hydrolysis. Free water activity was positively related to sugar production (r = 0.80–0.85), and integrating it with particle component and porosity allowed for accurate prediction of hydrolysis yields (R2 = 0.98–0.99). Inspired by particle liquefaction, a short-time feeding strategy was proposed to reconstruct high-solid hydrolysis.

Study on interfacial regulation in real food system: Cellulase-induced steady-state reconstruction of flaxseed-based plant milk and its molecular mechanisms

Cellulase is commonly used for the better release of phytonutrients due to its destructive coordinate effect on plant cell walls. In this study, cellulase hydrolysis showed a new feature: it significantly increased the physicochemical stability of flaxseed-based plant milk (FPM) due to interfacial modification of the oil bodies. Multispectroscopy and rheological measurement were performed to evaluate the stability of FPM, while the mechanical and oxidation stability of flaxseed oil bodies (FOBs) were characterized by rheological and induction period analysis, respectively. Additionally, Cryo-SEM, FTIR, and XRD analysis were used to provide structural information of FOBs' interface. Results revealed that cellulase treatment reduced the particle size and TSI of FPM, and significantly increased the adsorption of total polysaccharides at the FOBs’ interface, with an increase of 20.25 ± 3.77% (P < 0.05). Furthermore, FOBs with enzyme treatment (EN-FOBs) possessed higher elastic modulus (G′) and viscous modulus (G″) at 90 °C than untreated ones. Meanwhile, the induction period of the EN-FOBs was prolonged by 10.47 ± 2.10% (P < 0.05). These findings indicated that cellulase promoted steady-state reconstruction of FPM by contributing to formation of complex interfaces with high mechanical strength. These results offer insight into innovation in plant-based milk processing technology towards green, clean labeling, and high-quality.

Physicochemical, structural and functional properties of pomelo peel pectin extracted by combination of pulsed electric field and cellulase hydrolysis

In this study, pectin extracted from pomelo peel was investigated using three different combination methods of pulsed electric field (PEF) and cellulase. Three action sequences were performed, including PEF treatment followed by enzymatic hydrolysis, enzymatic hydrolysis followed by PEF treatment, and enzymatic hydrolysis simultaneously treated by PEF. The three corresponding pectins were namely PEP, EPP and SP. The physiochemical, molecular structural and functional properties of the three pectins were determined. The results showed that PEP had excellent physiochemical properties, with the highest yield (12.08 %), total sugar (80.17 %) and total phenol content (38.20 %). The monosaccharide composition and FT-IR analysis indicated that the three pectins were similar. The molecular weights of PEP, EPP and SP were 51.13, 88.51 and 40.00 kDa, respectively. PEP showed the best gel properties, emulsification stability and antioxidant capacity among the three products, due to its high galacturonic acid and total phenol content, appropriate protein and low molecular weight. The mechanism of PEF-assisted cellulase hydrolysis of pomelo peel was also revealed by SEM analysis. These results suggested that PEF pretreatment was the best method, which not only improved the efficiency of enzymatic extraction, but also reduced resource waste and increased financial benefits.

Effect of incubation conditions of cellulase hydrolysis on mechanical pulp fibre morphology

The mechanical pulp industry is diversifying through the manufacture of high-value paper products, such as microfibrillated cellulose. However, the development of fibre quality is still energy-intensive. Enzymatic hydrolysis is hypothesized to promote fibre cutting, greater fibrillation, and reduce refining energy costs. Despite potential benefits, there is little understanding of the mechanisms behind fibre development during enzymatic hydrolysis of mechanical pulp. This work investigates how incubation pH and temperature during enzymatic hydrolysis impact the refining of mechanical pulp short fibres. Incubation with endoglucanase at pH 5 and 60 °C increased fibre cutting by approximately 20 %. Fibrillation was negatively affected at this condition, resulting in increased slim fines formation with refining. Incubation at pH 8 and 80 °C promoted >15 % reduction in fibre length, despite such conditions being associated with low enzyme activity. The pH variation modified the sedimentation height of the fibres and the conductivity of suspensions, indicating a change in fibre surface charge. Fibre morphology changes were induced by enzyme hydrolysis conducted at conditions representative of the full range of pH and temperature observed in mechanical pulp mills.

Taklimakanibacter deserti gen. nov., sp. nov. and Taklimakanibacter lacteus sp. nov., isolated from desert soil.

Two Gram-stain-negative, aerobic, milk-white coloured, non-motile, short rod-shaped bacteria, designated as strains SYSU D60010T and SYSU D60012T, were isolated from sand samples collected from the Taklimakan Desert of Xinjiang Province in China. Both strains were positive for oxidase, catalase and nitrate reduction, but negative for amylase, H2S production, hydrolysis of gelatin and cellulase. Strains SYSU D60010T and SYSU D60012T grew well at 28 °C, at pH 7 and had the same NaCl tolerance range of 0-1 % (w/v). The major fatty acids (>5 %) of strains SYSU D60010T and SYSU D60012T were summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c), iso-C19 : 0 cyclo ω8c, C16 : 0 and iso-C18 : 1 2-OH. Q-10 was the only respiratory ubiquinone. Strains SYSU D60010T and SYSU D60012T showed high 16S rRNA gene sequence similarities to Aestuariivirga litoralis SYSU M10001T (94.2 and 94.1 %), Rhodoligotrophos jinshengii BUT-3T (92.0 and 91.9 %) and Rhodoligotrophos appendicifer 120-1T (91.8 and 91.7 %), and the genomes were 7.4 and 5.8 Mbp in size with DNA G+C contents of 62.8 and 63.0 mol%, respectively. Phylogenetic, phenotypic and chemotaxonomic characteristics indicated that these two strains represent a novel genus and two novel species within the family Aestuariivirgaceae. We propose the name Taklimakanibacter deserti gen. nov., sp. nov. for strain SYSU D60010T, representing the type strain of this species (=KCTC 52783T =NBRC 113344T) and Taklimakanibacter lacteus gen. nov., sp. nov. for strain SYSU D60012T, representing the type strain of this species (=KCTC 52785T=NBRC 113128T).

Optimization of Enzymolysis Modification Conditions of Dietary Fiber from Bayberry Pomace and Its Structural Characteristics and Physicochemical and Functional Properties.

Bayberry pomace, a nutrient-rich material abundant in dietary fiber (DF), has historically been underutilized due to a lack of thorough research. This study aimed to investigate the physicochemical and functional properties of the DF. Ultrasonic enzymatic treatment was performed to extract the total DF, which was then optimized to produce modified soluble dietary fiber (MSDF) and insoluble dietary fiber (MIDF). The optimized conditions yielded 15.14% of MSDF with a water-holding capacity (WHC) of 54.13 g/g. The DFs were evaluated for their structural, physicochemical, and functional properties. The MSDF showed a higher (p < 0.05) WHC, oil-holding capacity (OHC), swelling capacity (SC), cation exchange capacity (CEC), and glucose adsorption capacity (GAC) (about 14.15, 0.88, 1.23, 1.22, and 0.34 times) compared to the DF. Additionally, the MSDF showed strong, superior radical scavenging and blood sugar-lowering capabilities, with a more porous surface morphology. A Fourier-transform infrared (FT-IR) spectroscopy analysis indicated that enzymatic modification degraded the cellulose and hemicellulose, reducing the DF crystallinity. Overall, the results demonstrated that cellulase hydrolysis could effectively improve the physicochemical and functional properties of DF, thereby paving the way for its development into functional food products.

Unlocking the potential of oat straw: Efficient pretreatment methods for enhanced glucose production

The excellent physical and chemical properties of deep eutectic solvents, especially the ability of some of them to dissolve biomass, make them broadly applicable in biomass pretreatment. In this study, oat straw was pretreated with deep eutectic solvents composed of choline chloride and an acid (formic, lactic, or oxalic acid). The highest reducing sugar yield was obtained for the formic acid/choline chloride mixture. Using a pretreatment temperature of 110 °C, a reaction time of 2 h, and a solid-liquid ratio of 1:20, the reducing sugar yield obtained by cellulase hydrolysis was 23.5%, the degradation rate of cellulose reached 76.9%, and hemicellulose was completely degraded. The pretreated oat straw was then analyzed by scanning electron microscopy, Fourier transform infrared spectroscopy, and X-ray diffraction. The changes in its crystallinity and lignin content compared to the untreated specimen were determined. A preliminary mechanism for the pretreatment of oat straw with formic acid/choline chloride was revealed. The study could provide an opportunity to expand the application in biochemicals or biological feed processes.

Valorization of lignocellulosic biomass: Progress in the production of second-generation bioethanol

Cellulosic ethanol, a biofuel product derived from microbial fermentation of cellulose-rich agricultural waste biomass, holds the potential to significantly contribute to sustainable development, energy security, and environmental friendliness as a green and renewable alternative energy source. Cellulosic ethanol offers remarkable energy benefits and carbon emission reductions. However, the biorefining process of cellulosic ethanol faces challenges and complexities. This article revolves around five aspects: feedstock, composition, pretreatment, cellulase hydrolysis, and ethanol fermentation processes in the biorefining of cellulosic ethanol. It provides an overview of the process flow and characteristics of cellulosic ethanol biorefining, emphasizing its significance. The main technological bottlenecks in cellulosic ethanol production are analyzed. Recent research advancements in cellulosic ethanol biorefining are summarized, followed by an outlook on the research focus and future prospects in the field of cellulosic ethanol.

Effects of Physicochemical and Biological Treatment on Structure, Functional and Prebiotic Properties of Dietary Fiber from Corn Straw.

Corn straw is one kind of agricultural by-product containing 70-80% insoluble dietary fiber (IDF). In order to develop corn straw dietary fiber, this study was conducted to increase soluble dietary fiber (SDF) yield and improve the structure, functional and prebiotic properties of IDF and SDF from corn straw treated by alkali oxidation treatment, enzymatic hydrolysis, microbial fermentation and the combination of these methods. The results demonstrated that the yield of SDF was significantly increased from 2.64% to 17.15% after corn straw was treated by alkali oxidation treatment + Aspergillus niger fermentation + cellulase hydrolysis, compared with untreated corn straw. The SDF extracted from corn straw treated by alkali oxidation treatment + Aspergillus niger fermentation + cellulase hydrolysis (F-SDF) exhibited a honeycomb structure, low crystallinity (11.97%), good antioxidant capacity and high capacities of water holding, water solubility and cholesterol absorption and promoted short-chain fatty acids production by chicken cecal microbial fermentation in vitro. F-SDF enhanced the antibacterial activity against Escherichia coli and Staphylococcus aureus proliferations of Lactobacillus plantarum when it was used as a substrate for Lactobacillus plantarum fermentation. It could be concluded that the combined treatments could increase SDF yield from corn straw and improve its functional and prebiotic properties.

Platycodon grandiflorum saponins: Ionic liquid-ultrasound-assisted extraction, antioxidant, whitening, and antiaging activity

This study explored the use of ionic liquid-ultrasound (ILU)-assisted extraction to enhance the extraction rate of Platycodon grandiflorum saponins (PGSs), and the content, extraction mechanism, antioxidant activity, whitening, and antiaging activity of PGSs prepared using ILU, ultrasound-water, thermal reflux-ethanol, and cellulase hydrolysis were compared. The ILU method particularly disrupted the cell wall, improved PGS extraction efficiency, and yielded a high total saponin content of 1.45 ± 0.02 mg/g. Five monomeric saponins were identified, with platycodin D being the most abundant at 1.357 mg/g. PGSs displayed excellent in vitro antioxidant activity and exhibited inhibitory effects on tyrosinase, elastase, and hyaluronidase. The results suggest that PGSs may have broad antioxidant, skin-whitening, and antiaging potential to a large extent. Overall, this study provided valuable insights into the extraction, identification, and bioactivities of PGSs, which could serve as a reference for future development and application of these compounds in the functional foods industry.

Effects of combined drying techniques and cellulase hydrolysis on the nutritional value and sensory properties of shiitake mushrooms (Lentinus edodes)

Dried shiitake mushrooms offer rich nutritional value and unique sensory properties, prompting further investigation. The effects of different drying techniques (hot air drying (HAD), infrared hot air drying (IRHAD), pulsed vacuum drying (PVD), vacuum freeze drying (VFD), and natural drying (ND)) combined with enzymatic hydrolysis on the release of flavor compounds and nutrients from shiitake mushrooms were explored. The combination of HAD with cellulase hydrolysis yielded notably high levels of umami amino acids (5.4723 ± 0.1501 mg/g) and 5′-nucleotides (4.0536 ± 0.0062 mg/g), and superior volatile flavors. Combined with cellulase hydrolysis, IRHAD achieved the highest level of total sugars (6.57 ± 0.34 mg/mL), VFD resulted in the greatest soluble protein content (153.21 ± 0.23 μg/mL), PVD yielded the highest total phenolics content (93.20 ± 0.41 μg GAE/mL), and ND produced the maximum reducing sugar content (5.79 ± 0.13 mg/mL). This study addresses crucial gap in the post-drying processing of shiitake mushrooms, offering valuable insights for further product development of shiitake mushrooms.

Effective adsorption of norfloxacin from water on magnetic biochar composite derived from cellulase hydrolysis apple branch: Synthesis optimization, performance assessment and mechanism insight

Based on the optimization of feedstock and temperature condition, we synthesized a novel biochar composite (CHMAB) by pyrolyzing apple branch after cellulase hydrolysis and magnetization as well as coating them with humic acid (HA). Compared with the simultaneously prepared magnetic biochar (MAB) and HA-coated magnetic biochar (HMAB), batch adsorption test results showed that CHMAB had the best norfloxacin (NOR) removal performance in a wide pH range (2−12). The endothermic and spontaneous adsorption behavior of NOR on CHMAB could be better suited to the pseudo-second-order model and Langmuir model with maximum absorption capacity (81.08 mg g−1) at 35℃. Pore filling, π-π EDA interactions, hydrogen bonding, surface complexation, electrostatic interactions and hydrophobic interactions might all be implicated in the NOR removal process. Additionally, the regenerated CHMAB still maintained 99.23% removal efficiency for 0.1 mol L−1 NOR and performed well after seven cycles. These findings highlighted the possibility of enhancing the adsorption capacity of biochar for antibiotics by modification through cellulase hydrolysis of precursor and HA coating procedure. It is noteworthy that the use of low-cost agricultural and forestry wastes to produce adsorbents for emerging contaminants such as antibiotics could have greater potential for future practical applications under the ongoing dual carbon policy.

Combination of microwave with acid deep eutectic solvent pretreatment for reed (Phragmites australis) fractionation

The cost-effective and low-carbon fractionation of lignocellulosic biomass will enhance the economic viability of bio-refining. To achieve efficient reed fractionation under favorable conditions, microwave-assisted choline chloride and p-toluene sulfonic acid (M-ChCl/p-TsOH) pretreatment was employed. The fractionation of reed components demonstrated higher effectiveness with M-ChCl/p-TsOH pretreatment compared to conventional DES pretreatment. Under M-ChCl/p-TsOH conditions, a high hemicellulose removal rate (∼90%) and delignification rate (∼65%) were achieved while achieving a saccharification rate of 88% during cellulase hydrolysis. Microwave assistance not only significantly reduced reaction time to less than 60 s but also enhanced the pretreatment efficacy. Furthermore, the obtained lignin products consisted of low-polymerization lignin (MW < 800 g/mol) and acid-soluble lignin, providing a solid foundation for subsequent high-value utilization of lignin. This study provides a promising strategy for the low carbon fractionation of reed using DES, aiming to simultaneously achieve efficient cellulose hydrolysis and obtain tractable lignin.

Emulsification Characteristics of Insoluble Dietary Fibers from Pomelo Peel: Effects of Acetylation, Enzymatic Hydrolysis, and Wet Ball Milling.

To improve the application potential of pomelo peel insoluble dietary fiber (PIDF) in emulsion systems, acetylation (PIDF-A), cellulase hydrolysis (PIDF-E), and wet ball milling (PIDF-M) were investigated in this paper as methods to change the emulsification properties of PIDF. The impact of the methods on PIDF composition, structure, and physicochemical properties was also assessed. The results demonstrated that both acetylation modification and cellulase hydrolysis could significantly improve the emulsification properties of PIDF. The emulsions stabilized with PIDF-A and PIDF-E could be stably stored at 25 °C for 30 d without phase separation at particle concentrations above 0.8% (w/v) and had higher storage stability: The D4,3 increments of PIDF-A- and PIDF-E-stabilized emulsions were 0.98 μm and 0.49 μm, respectively, at particle concentrations of 1.2% (w/v), while the storage stability of PIDF-M-stabilized emulsion (5.29 μm) significantly decreased compared with that of PIDF (4.00 μm). Moreover, PIDF-A showed the highest water retention capacity (21.84 g/g), water swelling capacity (15.40 mL/g), oil retention capacity (4.67 g/g), and zeta potential absolute (29.0 mV) among the PIDFs. In conclusion, acetylation modification was a promising method to improve the emulsifying properties of insoluble polysaccharides.

Cloning of AA9 Polysaccharide Monooxygenase gene AN3860 into pEX2B for expression in Aspergillus oryzae

Polysaccharide monooxygenases (PMOs) catalyze oxidative degradation of recalcitrant carbohydrate chains in cellulose, starch, and chitin. In biofuel industry, the conversion of rich lignocellulose source to fermentable sugars by hydrolytic cellulases can be synergistically boosted by cellulose-active PMOs (AA9 PMOs) found in a vast number of fungi that grow on biomass. Aspergillus nidulans, a filamentous fungus, possess a dozen of PMO-encoding genes, but only the AN3860 is expressed at a high level when cultured with wheat straw as the sole carbon source. Bioinformatic analysis indicates that AN3860 belongs to type 3 AA9 PMO subfamily that is capable of hydroxylating both C1 and C4 of the glycosidic linkages. Therefore, AN3860 may be a potential enzyme to improve cellulose hydrolysis efficiency, which has not been characterized. In this study, we describe the AN3860 cloning into Aspergillus oryzae AUT1-PlD. To facilitate the purification of AN3860, we added a CBM20 tag to its C-terminal. The recombinant vector was designed and constructed successfully. Simultaneously, we have obtained the clone of A. oryzae carrying the target gene by the ATMT method. Further expression optimization and characterization of AN3860 by both activity assays and spectroscopic techniques are underway.