Search Results (933)

This study investigates the effectiveness of hydrothermal and alkaline pretreatment methods in enhancing the concentration of fermentable sugars derived from rice husk waste. After the pretreatments, enzymatic hydrolysis and fermentation processes were executed to evaluate the ethanol production from each pretreatment. Rice husk powder measuring ≤250 µm was used. For the alkaline pretreatment, sodium hydroxide (NaOH) was used at concentrations ranging from 0.5, 1 to 1.5% w/v. The efficacy of the hydrothermal pretreatment method was evaluated after 15, 30 and 45 min at 120 °C. The enzymatic hydrolysis process was performed over 144 h at 50 °C, pH 4.8 with an enzyme loading of 30 FPU (filter paper units). Fermentation was carried out at 37 °C using a strain of Saccharomyces cerevisiae Hansen 1883 (NCYC 366). Results indicated that the optimal conditions for alkaline pretreatment were observed at a 1.5% NaOH, while the best hydrothermal procedure was achieved at 120 °C and 45 min. The impact of these pretreatments was assessed based on the efficiency of enzymatic hydrolysis. The alkaline pretreatment resulted in 81.70% conversion of cellulose to glucose and 96.30% conversion of hemicellulose to xylose. In contrast, the hydrothermal pretreatment achieved 93% cellulose-to-glucose conversion and 83.35% hemicellulose-to-xylose conversion. The ethanol production registered ranged from 13 to 13.23 g·L−1, corresponding to a conversion factor of 0.43 for ethanol from fermentable sugars. Full article

Xylanase is commonly thought to effectively cooperate with cellulase to promote the bioconversion of lignocellulose. In this study, a novel xylanase, SipoEnXyn10A (Xyn10A), previously identified from Streptomyces ipomoeae, was employed to investigate its synergetic effects on sugarcane bagasse (SCB) transformation. It was shown that the relative increase in reducing sugars reached up to 65%, with enhanced yields of glucose and xylose by 78% and 50%, respectively, in the case of the replacement of cellulase with an equivalent amount of Xyn10A at an enzyme loading of 12.5%. The highest degrees of synergy (DS) for glucose and xylose could reach 2.57 and 1.84. Moreover, the hydrolysis rate increased evidently, and the reaction time to reach the same yield of glucose and xylose was shortened by 72 h and 96 h, respectively. This study on synergistic mechanisms demonstrated that the addition of Xyn10A could cause the destruction of substrates’ morphology and the dissolution of lignin components but could not change the accessibility and crystallinity of substrate cellulose. The joint effect of cellulase and xylanase during the hydrolysis process was thought to result in a synergistic mechanism. Full article

The improper treatment of crop straw not only leads to resource wastage but also adversely impacts the ecological environment. However, the application of microorganisms can accelerate the decomposition of crop straw and improve its utilization. In this study, cellulose-degrading microbial strains were isolated from naturally decayed corn straw and screened using Congo red staining, along with assessing variations in carboxymethyl cellulase (CMCase) activity, filter paper enzyme (FPase) activity and β-glucosidase (β-Gase) activity, as well as the degradation rate. The eight strains, namely Neurospora intermedia isolate 29 (A1), Streptomyces isolate FFJC33 (A2), Gibberella moniliformis isolate FKCB-009 (A3), Fusarium fujikuroi isolate EFS3(2) (A4), Fusarium Fujikuroi isolate FZ04 (A5), Lysine bacillus macroides strain LNHL43 (B1), Bacillus subtilis strain MPF30 (B2) and Paenibacilli lautus strain ALEB-P1 (C), were identified and selected for microbial strain consortium design based on their high activities of CMCase, FPase and β-Gase. The fungi, bacteria and actinomycete strains were combined without antagonistic effects on corn straw decomposition. The results showed the A2B2 combination had a significantly higher FPase at 55.44 U/mL and β-Gase at 25.73 U/mL than the other two strain combinations (p < 0.05). Additionally, the degradation rate of this combination was 40.33%, which was considerably higher than that of the other strains/consortia. The strain combination A4B2C also had superior enzyme activity, including CMCase with a value of 35.03 U/mL, FPase with a value of 63.59 U/mL and β-Gase with a value of 26.15 U/mL, which were significantly different to those of the other three strain combinations (p < 0.05). Furthermore, seven single microbial strains with high cellulase activities were selected to construct various microbial consortiums for in situ composting in order to evaluate their potential. Taken as a whole, the results of composting, including temperature, moisture content, pH, E4/E6 value and seed germination index, indicated that the microbial strain consortium consisting of Neurospora intermediate isolate 29, Fusarium fujikuroi isolate EFS3(2), Fusarium fujikuroi isolate FZ04, Lysinibacillus macrolides, Lysinibacillus sphaericus, Bacillus subtilis and Paenibacillus lautus was advantageous for corn straw decomposition and yielded high-quality compost. The screened flora was able to effectively degrade corn straw. This study provides a novel solution for the construction of a microbial consortium for the composting of corn straw. Full article

The presence of diverse carbohydrate-active enzymes (CAZymes) is crucial for the direct bioconversion of lignocellulose. In this study, various anaerobic microbial consortia were employed for the degradation of 10 g/L of minimally pretreated corncob. The involvement of lactic acid bacteria (LAB) and a CAZyme-rich bacterium (Bacteroides cellulosilyticus or Paenibacillus lautus) significantly enhanced the lactic acid production by Ruminiclostridium cellulolyticum from 0.74 to 2.67 g/L (p < 0.01), with a polysaccharide conversion of 67.6%. The supplement of a commercial cellulase cocktail, CTec 2, into the microbial consortia continuously promoted the lactic acid production to up to 3.35 g/L, with a polysaccharide conversion of 80.6%. Enzymatic assays, scanning electron microscopy, and Fourier transform infrared spectroscopy revealed the substantial functions of these CAZyme-rich consortia in partially increasing enzyme activities, altering the surface structure of biomass, and facilitating substrate decomposition. These results suggested that CAZyme-intensified consortia could significantly improve the levels of bioconversion of lignocellulose. Our work might shed new light on the construction of intensified microbial consortia for direct conversion of lignocellulose. Full article

Anthracnose is one of the destructive diseases of pitaya that seriously affects the plant growth and fruit quality and causes significant yield and economic losses worldwide. However, information regarding the species of pathogens that cause anthracnose in pitaya (Hylocereus undatus) fruits in Gansu Province, China, and its pathogenic mechanism is unknown. Thus, the purposes of our present study were to identify the species of pathogens causing H. undatus fruits anthracnose based on the morphological and molecular characteristics and determine its pathogenic mechanism by physiological and biochemical methods. In our present study, forty-six isolates were isolated from the collected samples of diseased H. undatus fruits and classified as three types (named as H-1, H-2, and H-3), according to the colony and conidium morphological characteristics. The isolation frequencies of H-1, H-2, and H-3 types were 63.04%, 21.74%, and 15.22%, respectively. The representative single-spore isolate of HLGTJ-1 in H-1 type has significant pathogenicity, and finally we identified Colletotrichum truncatum as the pathogen based on the morphological characteristics as well as multi-locus sequence analysis. Moreover, the H. undatus fruits inoculated with C. truncatum had a significantly increased activity of cell wall-degrading enzymes (CWDEs) cellulase (Cx), β-glucosidase (β-Glu), polygalacturonase (PG), and pectin methylgalacturonase (PMG), while having a decreased level of cell wall components of original pectin and cellulose in comparison to control. The average increased activities of Cx, β-Glu, PG, and PMG were 30.73%, 40.40%, 51.55%, and 32.23% from day 0 to 6 after inoculation, respectively. In contrast, the average decreased contents of original pectin and cellulose were 1.82% and 16.47%, respectively, whereas the average increased soluble pectin content was 38.31% in comparison to control. Our results indicate that C. truncatum infection increased the activities of CWDEs in H. undatus fruits to disassemble their cell wall components, finally leading to the fruits’ decay and deterioration. Thus, our findings will provide significant evidence in the controlling of pitaya anthracnose in the future. Full article

Cellulose is one of the most abundant renewable resources in nature. However, its recalcitrant crystalline structure hinders efficient enzymatic depolymerization. Unlike cellulases, lytic polysaccharide monooxygenases (LPMOs) can oxidatively cleave glycosidic bonds in the crystalline regions of cellulose, playing a crucial role in its enzymatic depolymerization. An AA9 LPMO from Myceliophthora thermophila was previously identified and shown to exhibit a highly efficient catalytic performance. To further enhance its catalytic efficiency, consensus mutagenesis was applied. Compared with the wild-type enzyme, the oxidative activities of mutants A165S and P167N increased by 1.8-fold and 1.4-fold, respectively, and their catalytic efficiencies (k_cat/K_m) improved by 1.6-fold and 1.2-fold, respectively. The mutants also showed significantly enhanced activity in the synergistic degradation of cellulose with cellobiohydrolase. Additionally, the P167N mutant exhibited better H₂O₂ tolerance. A molecular dynamics analysis revealed that the increased activity of mutants A165S and P167N was due to the closer proximity of the active center to the substrate post-mutation. This study demonstrates that selecting appropriate mutation sites via a semi-rational design can significantly improve LPMO activity, providing valuable insights for the protein engineering of similar enzymes. Full article

This study aims to evaluate the in silico genomic characteristics of five species of the genus Planotetraspora: P. kaengkrachanensis, P. mira, P. phitsanulokensis, P. silvatica, and P. thailandica, with a view to their application in therapeutic research. The 16S rRNA comparison indicated that these species were phylogenetically distinct. Pairwise comparisons of digital DNA-DNA hybridization (dDDH) and OrthoANI values between these studied type strains indicated that dDDH values were below 62.5%, while OrthoANI values were lower than 95.3%, suggesting that the five species represent distinct genomospecies. These results were consistent with the phylogenomic study based on core genes and the pangenome analysis of these five species within the genus Planotetraspora. However, the genome annotation showed some differences between these species, such as variations in the number of subsystem category distributions across whole genomes (ranging between 1979 and 2024). Additionally, the number of CAZYme (Carbohydrate-Active enZYme) genes ranged between 298 and 325, highlighting the potential of these bacteria for therapeutic research applications. The in silico physico-chemical characteristics of cellulases from Planotetraspora species were analyzed. Their 3D structure was modeled, refined, and validated. A molecular docking analysis of this cellulase protein structural model was conducted with cellobiose, cellotetraose, laminaribiose, carboxymethyl cellulose, glucose, and xylose ligand. Our study revealed significant interaction between the Planotetraspora cellulase and cellotetraose substrate, evidenced by stable binding energies. This suggests that this bacterial enzyme holds great potential for utilizing cellotetraose as a substrate in various applications. This study enriches our understanding of the potential applications of Planotetraspora species in therapeutic research. Full article

Mandarin peel (MP) has gained attention as a feedstock for flavonoid recovery via the extraction process based on the biorefinery concept, but residues remain after the extraction. Toward an integrated biorefinery concept, this study aimed to valorize extracted MP (eMP) by using it in bioethanol production. For efficient fermentable sugar production, the effect of enzymatic hydrolysis conditions on sugar conversion from eMP was investigated, and the results showed that combining cellulase and cellobiase resulted in a higher enzymatic glucose conversion (78.2%) than the use of the individual enzymes (37.5% and 45.6%). Pectinase played an essential role in enhancing enzymatic arabinose conversion, and the optimal conditions were determined to be pH 4 and 90 units of the three enzymes. Under optimal conditions, the sugar yield was 199 g glucose and 47 g arabinose/kg eMP, and the hydrolysate was used in bioethanol fermentation. The results showed that the bioethanol production was 3.78 g/L (73.9% yield), similar to the control medium (3.79 g/L; 74.2% yield), although the cell growth of the yeast was slightly delayed in the eMP hydrolysate medium. This study highlights the potential of eMP as a low-cost feedstock for sugar and bioethanol production. Full article

An amylolytic industrial yeast strain named 1974-GA-temA, developed previously by our research team by coexpressing the α-amylase and glucoamylase genes, combines enzyme production, sweet potato residue (SPR) hydrolysis, and glucose fermentation into ethanol in a one-step process. This consolidated bioprocessing (CBP) method has great application potential in the commercial production of bioethanol from SPR, but important fermentation parameters should be optimized to further increase the ethanol concentration and yield. In this study, the effects of the initial fermentation pH, solid-to-liquid ratio, inoculation volume, addition of exogenous enzyme, and supplementation with metal ions were systemically investigated. Single-factor experiments revealed that the optimal pH was 4.0. In the solid-to-liquid ratio test, an increase in the solid-to-liquid ratio corresponded with a gradual increase in the ethanol concentration, peaking at 1:5. However, the ethanol yield gradually decreased, with the optimal solid-to-liquid ratio identified as 1:5. The ethanol concentration and yield reached 9.73 g/L and 5.84%, respectively. Additionally, an increase in the inoculum size resulted in increased ethanol concentration and yield, with the optimal inoculum level determined to be 10%. An ethanol concentration of 7.87 g/L was attained under these specified conditions, equating to an ethanol yield of 4.72%. Further analysis was conducted to assess the effects of exogenous cellulase, hemicellulase, and pectinase, both individually and in combination, on ethanol concentration and yield. The results indicated that pectinase had a particularly significant effect. The highest ethanol concentration was observed when all three enzymes were administered concurrently, yielding 27.27 g/L ethanol. Then, the role of metal ions in SPR fermentation was evaluated. The metal ions did not significantly affect the process, with the exception of copper ions. The addition of copper ions at a specific concentration of 0.2 g/100 g SPR increased the ethanol concentration. However, concentrations exceeding 0.2 g/100 g SPR inhibited yeast cell growth. Finally, orthogonal optimization was employed to determine the optimal combination of factors: pH, 4.0; solid-to-liquid ratio, 1:6; inoculation volume, 10%; cellulase and pectinase addition; and the absence of Cu²⁺ addition. Under these conditions, strain 1974-GA-temA produced 34.83 ± 0.62 g/L ethanol after 8 days of fermentation, corresponding to a 20.90% ± 0.37% ethanol yield. This value markedly exceeds the outcomes of all the conducted orthogonal experiments. The fermentation optimization experiments in this study are expected to increase ethanol production during the CBP fermentation of SPR. Full article

This paper introduces an enzymatic approach to estimate internal mass-transfer resistances during food digestion studies. Cellulase has been used to degrade starch cell walls (where cellulose is a significant component) and reduce the internal mass-transfer resistance, so that the starch granules are released and hydrolysed by amylase, increasing the starch hydrolysis rates, as a technique for measuring the internal mass-transfer resistance of cell walls. The estimated internal mass-transfer resistances for granular starch hydrolysis in a beaker and stirrer system for simulating the food digestion range from 2.2 × 10⁷ m⁻¹ s at a stirrer speed of 100 rpm to 6.6 × 10⁷ m⁻¹ s at 200 rpm. The reaction rate constants for cellulase-treated starch are about three to eight times as great as those for starch powder. The beaker and stirrer system provides an in vitro model to quantitatively understand external mass-transfer resistance and compare mass-transfer and reaction rate kinetics in starch hydrolysis during food digestion. Particle size analysis indicates that starch cell wall degradation reduces starch granule adhesion (compared with soaked starch samples), though the primary particle sizes are similar, and increases the interfacial surface area, reducing internal mass-transfer resistance and overall mass-transfer resistance. Dimensional analysis (such as the Damköhler numbers, Da, 0.3–0.5) from this in vitro system shows that mass-transfer rates are greater than reaction rates. At the same time, SEM (scanning electron microscopy) images of starch particles indicate significant morphology changes due to the cell wall degradation. Full article

Oyster mushroom spherical virus (OMSV) is a mycovirus that inhibits mycelial growth, induces malformation symptoms, and decreases the yield of fruiting bodies in Pleurotus ostreatus. However, the pathogenic mechanism of OMSV infection in P. ostreatus is poorly understood. In this study, RNA sequencing (RNA-seq) was conducted, identifying 354 differentially expressed genes (DEGs) in the mycelium of P. ostreatus during OMSV infection. Verifying the RNA-seq data through quantitative real-time polymerase chain reaction on 15 DEGs confirmed the consistency of gene expression trends. Both Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses highlighted the pivotal role of primary metabolic pathways in OMSV infection. Additionally, significant changes were noted in the gene expression levels of carbohydrate-active enzymes (CAZymes), which are crucial for providing the carbohydrates needed for fungal growth, development, and reproduction by degrading renewable lignocellulose. The activities of carboxymethyl cellulase, laccase, and amylase decreased, whereas chitinase activity increased, suggesting a potential mechanism by which OMSV influenced mycelial growth through modulating CAZyme activities. Therefore, this study provided insights into the pathogenic mechanisms triggered by OMSV in P. ostreatus. Full article

Applications of millet bran dietary fiber (MBDF) in the food industry are limited by its poor hydration properties. Herein, MBDF was modified by heating, xylanase and cellulase treatment separately combined with carboxymethylation, acetylation, and phosphate crosslinking, and the effects of the modified MBDFs on heat-induced egg white protein gel (H-EWG) were studied. The results showed that three composite modifications, especially heating and dual enzymolysis combined with carboxymethylation, increased the surface area, soluble fiber content, and hydration properties of MBDF (p < 0.05). MBDF and the modified MBDFs all made the microstructure of H-EWG denser and decreased its α-helix content. Three composite modifications, especially heating and dual enzymolysis combined with carboxymethylation, enhanced the improving effect of MBDF on the WRA (from 24.89 to 35.53 g/g), pH, hardness (from 139.93 to 323.20 g), chewiness, and gumminess of H-EWPG, and enhanced the gastric stability at 3–5 g/100 g. MBDFs modified with heating and dual enzymolysis combined with acetylation or crosslinking were more effective in increasing the antioxidant activity of the gastrointestinal hydrolysates of H-EWG than MBDF (p < 0.05). Overall, heating, xylanase and cellulase treatment separately combined with carboxymethylation, acetylation and crosslinking can enhance the hydration properties and the improving effect of millet bran fibers on H-EWG properties. Full article

β-glucosidase is a key component of cellulase for its function in hydrolyzing cellobiose to glucose in the final step of cellulose degradation. The high-level expression of β-glucosidase is essential for cellulose conversion. Aspergillus niger ATCC 20611 has the potential for efficient protein expression because of its ability to secret enzymes for the industrial production of fructooligosaccharides, but it lacks robust promoters for high-level protein expression. Here, the development of A. niger 20611 as a powerful protein expression system exploited the conserved constitutive promoter Pgpd1 of the glyceraldehyde-3-phosphate dehydrogenase-encoding gene from Trichoerma reesei to drive the expression of the enhanced green fluorescent protein in A. niger ATCC 20611. The mycelium of the transformant AGE9 exhibited intense fluorescence. Then, the promotor Pgpd1 was used to drive the expression of β-glucosidase and the enzyme activity of transformants AGB1 and AGB33 were 1.02 and 0.51 U/mL, respectively. These results demonstrate that the promotor Pgpd1 from T. reesei was applicable for A. niger ATCC 20611. Furthermore, the T. reesei-specific robust promoter Pcdna1 was used to drive the expression of β-glucosidase. The β-glucosidase exhibited a high-level expression with a yield of 15.2 U/mL, which was over 13.9 times higher than that driven by the promoter Pgpd1. The β-glucosidase was thermally stable and accounted for 85% of the total extracellular proteins. Subsequently, the fermentation broth including β-glucosidase was directly added to the cellulase mixture of T. reesei for saccharification of the acid-treated corncob residues and the delignified corncob residues, which increased the saccharification efficiency by 26.21% and 29.51%, respectively. Thus, β-glucosidase exhibited a high level of expression in A. niger ATCC 20611 and enhanced cellulose degradation by addition in vitro. In addition, the robust promoter Pcdna1 of T. reesei could drive the high-level expression of protein in A. niger ATCC 20611. These results demonstrate that the promoters in filamentous fungi could be employed across species in A. niger ATCC 20611 and further facilitated the efficient expression of β-glucosidase to optimize cellulases for efficient cellulose transformation. Full article

Enzymatic degradation of cellulosic biomass represents the most sustainable and environmentally friendly method for producing liquid biofuel, widely utilized in various commercial processes. While cellulases are predominantly produced by bacteria and fungi, the enzymatic potential of cellulase-producing yeasts remains significantly less explored. In this study, the yeast strain Trichosporon insectorum, isolated from the gut of the coprophagous beetle Gymnopleurus sturmii, was utilized for cellulase production in submerged fermentation. A central composite design was employed to optimize cellulase production, with substrate concentration, temperature, and pH as dependent variables. The highest CMCase activity of 0.71 IU/mL was obtained at 1% substrate concentration, pH 5, and an incubation temperature of 40 °C for 72 h of fermentation using cellulose as a carbon source. For FPase production, the high value was 0.23 IU/mL at 0.5% CMC, pH 6, and an incubation temperature of 40 °C for 72 h. After purification, the enzymes produced by T. insectorum represent 39% of the total proteins. The results of this study offer an alternative strategy for utilizing various carbon sources, both soluble (CMC, carboxymethylcellulose) and insoluble (cellulose), to efficiently produce cellulase for the degradation of lignocellulosic materials. This approach holds promising benefits for sustainable waste management. Full article

A new flour blend (F) composed of selected milling and leaving passages with a high content of non-starch polysaccharides underwent thermal (T), hydrothermal (H) or hybrid processing and was used along with cellulase (C) and cellulase-xylanase complex (CX) to produce bread. This modified flour can be considered a clean label product. In this study, blends of common and treated flours were tested for dough properties and rheology. The modified flours were added at 10 and 20% to the base wheat flour. A pan bread was then prepared to test their suitability for bread baking. Dough and bread properties were subsequently assessed. Accordingly, dough with added thermally, hydrothermally, and hybrid modified flours revealed differences in rheology. Addition of hybrid enzymatic-hydrothermal treated flour increased dough tenacity by 23% and baking strength by 26%, but decreased dough extensibility by 19%, whereas hybrid enzymatic-thermal modification increased water absorption by 6% and bread yield from 146.77% to 150.02% when modified flour was added at 20%. Breads with added modified flours demonstrated a 16% increase in bread volume, 8% lower baking loss, and 14% greater density, with no negative effect on color and texture. Thus, hybrid thermal-enzymatic treatment of the developed flours can be recommended as a suitable method for enhancing the utilization of waste flour fractions and increasing their value by enabling them to be considered as clean label bread improvers. Full article