Cellulase Research Articles

Activity of fungal \u03b2-glucosidases on cellulose

BackgroundFungal beta-glucosidases (BGs) from glucoside hydrolase family 3 (GH3) are industrially important enzymes, which convert cellooligosaccharides into glucose; the end product of the cellulolytic process. They are highly active against the β-1,4 glycosidic bond in soluble substrates but typically reported to be inactive against insoluble cellulose.ResultsWe studied the activity of four fungal GH3 BGs on cellulose and found significant activity. At low temperatures (10 ℃), we derived the approximate kinetic parameters kcat = 0.3 ± 0.1 s−1 and KM = 80 ± 30 g/l for a BG from Aspergillus fumigatus (AfBG) acting on Avicel. Interestingly, this maximal turnover is higher than reported values for typical cellobiohydrolases (CBH) at this temperature and comparable to those of endoglucanases (EG). The specificity constant of AfGB on Avicel was only moderately lowered compared to values for EGs and CBHs.ConclusionsOverall these observations suggest a significant promiscuous side activity of the investigated GH3 BGs on insoluble cellulose. This challenges the traditional definition of a BG and supports suggestions that functional classes of cellulolytic enzymes may represent a continuum of overlapping modes of action.

Enhanced Activity and Stability of Acidothermus cellulolyticus Endoglucanase 1 in Ionic Liquids via Engineering Active Site Residues and Non-Native Disulfide Bridges

We report the rational mutagenesis and engineering of endoglucanase 1 (E1) from Acidothermus cellulolyticus, an industrially relevant cellulase, for improved biomass conversion in ionic liquids. Th...

Combined mechanical grinding and enzyme post-treatment leading to increased yield and size uniformity of cellulose nanofibrils

The low yield and size nonuniformity of cellulose nanofibrils (CNFs) prepared by mechanical grinding restrict their extensive application in many fields. In this study, microfibrillated cellulose (MFC) with varying grinding time (0–2.5 h) was post-treated with endoglucanase (EG) to improve the yield and size uniformity of the CNFs. With the same grinding time, the yield of the CNFs post-treated with EG increased approximately one-fold, and the diameter of the microfibrils decreased according to the SEM and TEM images. The yield of the CNFs ground for 0.5 h and post-treated by EG was similar to that of the sample ground for 2.5 h, and approximately 26.1 kWh/kg mechanical grinding energy were saved by the EG post-treatment while obtaining the same CNF yield. There was an abundance of micropores that affected the specific surface area of samples, and the width of the micropores was centered between 1.3 and 2.0 nm. The crystallinity index (CrI) of the MFC decreased gradually from 74.90 to 69.30% with prolonged grinding time, whereas the CrI of the samples post-treated by EG slightly increased between 1.7 and 3.0%. The tensile strength and elongation at break of the films prepared by the EG post-treated samples slightly decreased, while the Young’s modulus and transmittance increased. This study provides a valid and an eco-friendly biological method to improve the yield and size uniformity of CNFs and reduce energy consumption.

Soil labile organic carbon fractions and soil enzyme activities after 10\xa0years of continuous fertilization and wheat residue incorporation

Labile organic carbon (LOC) fractions and related enzyme activities in soils are considered to be early and sensitive indicators of soil quality changes. We investigated the influences of fertilization and residue incorporation on LOC fractions, enzyme activities, and the carbon pool management index (CPMI) in a 10-year field experiment. The experiment was composed of three treatments: (1) no fertilization (control), (2) chemical fertilizer application alone (F), and (3) chemical fertilizer application combined with incorporation of wheat straw residues (F + R). Generally, the F + R treatment led to the highest concentrations of the LOC fractions. Compared to the control treatment, the F + R treatment markedly enhanced potential activities of cellulase (CL), β-glucosidase (BG), lignin peroxidase (LiP), and manganese peroxidase (MnP), but decreased laccase (LA) potential activity. Partial least squares regression analysis suggested that BG and MnP activities had a positive impact on the light-fraction organic carbon (LFOC), permanganate-oxidizable carbon (POXC), and dissolved organic carbon (DOC) fractions, whereas laccase activity had a negative correlation with those fractions. In addition, the F + R treatment significantly increased the CPMI compared to the F and control treatments. These results indicated that combining fertilization with crop residues stimulates production of LOC and could be a useful approach for maintaining sustainable production capacity in lime concretion black soils along the Huai River region of China.

Potential applications of enzymes on the extraction of vitexin from dried Mas Cotek leaves

Mas Cotek or Ficus deltoidea var. deltoidea leaves extracts contain various beneficial properties such as antidiabetic, antihypoglycemic and antioxidant activities, which can be enzymatically extracted in water. Vitexin is one of the antioxidants and remarkable compound of flavone C-glycosides found in Mas Cotek leaves. Cellulase is one of the essential enzymes applied to hydrolyze cellulose into fermentable sugars and therefore, this enzyme was used to extract vitexin in this study. Single parameter approach was used to determine factors influencing the enzymatic-mediated extraction. These factors were enzyme concentration, sample-to-water ratio, temperature, incubation time and agitation. The maximum yield of vitexin found was 0.547% under the optimum conditions: sample-to-water ratio 1:10 g/mL, enzyme concentration 0.4%, incubation time of 240 min, temperature of 50 ± 0.02°C and stirring speed of 200 rpm. The highest amount of reducing sugar obtained was 4.039 mg/mL with the use of 0.4% cellulose. A correlation was also observed between the yield of vitexin and reducing sugars. Key words: Cellulose, enzymatic assisted extraction, vitexin, reducing sugars, cellulose, Mas Cotek, Ficus deltoidea var. deltoidea, antioxidants.

Engineering Pichia pastoris with surface-display minicellulosomes for carboxymethyl cellulose hydrolysis and ethanol production

BackgroundsEngineering yeast as a consolidated bioprocessing (CBP) microorganism by surface assembly of cellulosomes has been aggressively utilized for cellulosic ethanol production. However, most of the previous studies focused on Saccharomyces cerevisiae, achieving efficient conversion of phosphoric acid-swollen cellulose (PASC) or microcrystalline cellulose (Avicel) but not carboxymethyl cellulose (CMC) to ethanol, with an average titer below 2 g/L.ResultsHarnessing an ultra-high-affinity IM7/CL7 protein pair, here we describe a method to engineer Pichia pastoris with minicellulosomes by in vitro assembly of three recombinant cellulases including an endoglucanase (EG), an exoglucanase (CBH) and a β-glucosidase (BGL), as well as a carbohydrate-binding module (CBM) on the cell surface. For the first time, the engineered yeasts enable efficient and direct conversion of CMC to bioethanol, observing an impressive ethanol titer of 5.1 g/L.ConclusionsThe research promotes the application of P. pastoris as a CBP cell factory in cellulosic ethanol production and provides a promising platform for screening the cellulases from different species to construct surface-assembly celluosome.

Involvement of PaSNF1 in Fungal Development, Sterigmatocystin Biosynthesis, and Lignocellulosic Degradation in the Filamentous Fungus Podospora anserina.

The sucrose non-fermenting 1/AMP-activated protein kinase (SNF1/AMPK) is a central regulator of carbon metabolism and energy production in the eukaryotes. In this study, the functions of the Podospora anserina SNF1 (PaSNF1) ortholog were investigated. The ΔPaSNF1 mutant displays a delayed development of mycelium and fruiting bodies and fails to form ascospores. The expression of the PaSNF1 gene in the strain providing female organs in a cross is sufficient to ensure fertility, indicating a maternal effect. Results of environmental stress showed that ΔPaSNF1 was hypersensitive to stress, such as osmotic pressure and heat shock, and resistant to fluconazole. Interestingly, the knockout of PaSNF1 significantly promoted sterigmatocystin (ST) synthesis but suppressed cellulase [filter paperase (FPA), endoglucanase (EG), and β-glucosidase (BG)] activity. Further, transcriptome analysis indicated that PaSNF1 made positive regulatory effects on the expression of genes encoding cellulolytic enzymes. These results suggested that PaSNF1 may function in balancing the operation of primary and secondary metabolism. This study suggested that SNF1 was a key regulator concerting vegetative growth, sexual development, and stress tolerance. Our study provided the first genetic evidence that SNF1 was involved in the ST biosynthesis and that it may also be a major actor of lignocellulose degradation in P. anserina.

Secretome analysis of Talaromyces emersonii reveals distinct CAZymes profile and enhanced cellulase production through response surface methodology

This study reports comprehensive insight into the repertoire of proteins present in the secretome of a hypercellulolytic fungus Talaromyces emersonii and designing of new production medium containing combination of cellulose and gram flour for enhanced production of cellulases and hemicellulases. Probe into the secretome of the fungus through Quadrupole Time Of Flight Liquid Chromatography/Mass Spectrometry (Q-TOF LC/MS) revealed a total of 266 proteins that were represented by high abundance of Carbohydrate Active enzymes (CAZymes) comprising of 93 glycosyl hydrolase family, 13 cellulose binding module, 14 carbohydrate esterase, 8 auxiliary activity proteins. The zymography of endoglucanase (EG), cellobiohydrolase (CBHI), β-glucosidase (β-G) and xylanase revealed expression of five EG, two CBHI, four β-G and two xylanases as the major proteins. The optimization of culture conditions using RSM resulted in 113.74, 6.07, 31.23 and 177.02 Units/mL corresponding to 1.62, 1.34, 1.25 and 1.22 fold higher endoglucanase, cellobiohydrolase (CBHI), β-glucosidase and xylanase activity, respectively, when compared to unoptimized control. The secretome produced by this strain was highly active against acid and alkali pre-treated rice straw showing 59.05 and 74.33 % saccharification, respectively, which was further enhanced when cocktail of T. emersonii and Achaetomium strumarium was used.

Effect of acute exposure to ammonia and BFT alterations on Rhynchocypris lagowski: Digestive enzyme, inflammation response, oxidative stress and immunological parameters

Biofloc technology (BFT) is a new green culture technology that is intended not merely to eradicate nitrogenous residues but also enhance immunity and antioxidant activity in aquatic animals. A 56-day feeding trial and a 96 h ammonia challenge test were implemented to evaluate the effect of acute exposure to ammonia and BFT alterations on Rhynchocypris lagowski: digestive enzyme, inflammation response, oxidative stress and immunological parameters in zero water exchange tanks. According to the differences of C/N ratios, the experiment was divided into four groups: C/N 10.8:1 (control group), C/N 15:1, C/N 20: 1 and C/N 25:1. The results provided evidence that weight gain rate (WGR), specific growth rate (SGR) and protein efficiency ratio (PER) were significantly elevated in C/N 20, while food conversion rate (FCR) was considerably decreased (P < 0.05) compared to the control. Significant increases in digestive enzyme amylase (AMS), lipase (LPS), protease (PRS) and cellulase (CES) activity; Immune enzyme complement C3, complement C4, Nitric Oxide Synthase (NOS) and immunoglobulin M (IgM) activity; Serum biochemical lysozyme (LSZ), glutamic pyruvic transaminase (GPT), glutamic oxaloacetic transaminase (GOT) and alkaline phosphatase (AKP) activity; Inflammation TNF-α, IL-1β, IL-2, IL-6 of R. lagowskis were found in the C/N 20 group after a 56-day feeding trial and a challenging trial (P < 0.05). Comparing the antioxidant capacity of R. lagowski in gills, brains and spleen of juveniles from the four experimental groups, the activity of superoxide dismutase (SOD), total antioxidant capacity (T-AOC), catalase (CAT) activity and glutathion reductases (GR) activity of juveniles in the C/N 20 group were significant higher (P < 0.05), and the content of malondialdehyde (MDA) was considerably lower than that in the control. Overall, these findings suggest that BFT not only can enhance R. lagowski growth performance, digestive enzyme activity, and strengthen antioxidant status but also upgrade immune response, improve the ability of resistance to ammonia stress.

Pectin Degradation is an Important Determinant for Alfalfa Silage Fermentation through the Rescheduling of the Bacterial Community.

This study aimed to evaluate the effects of the four kinds of additives on the silage quality and the relevant bacterial community diversity by Illumina HiSeq 16S rRNA sequencing. The four kinds of additives were Lactobacillus plantarum (LP), organic acids including gallic acid (GA) and phenyllactic acid (PA), pectin (PEC), and enzymes including pectinase (PEE) and cellulase (CE). After 30 d of fermentation, the pH value was shown to have the lowest value in the PEE and PEC groups, followed by the PA group, and then in CE and GA groups; the highest value of pH was found in both LP and control groups. The ammonia nitrogen concentration was lower in the PEE group compared to the other groups except for the PA group. In the comparisons among the seven groups, Lactobacillus was higher in the LP group, Paracoccus was higher in the GA group, Weissella was higher in the PA group, Leuconostoc was higher in the PEC group, Bacillus, Aeromonas, and Curvibacter were higher in the PEE group, and Coriobacteriaceae_UCG_002 was higher in the CE group compared to the other groups. This study proposed that the addition of PEC and PEE improved the fermentation quality of alfalfa silage compared to other additives by improving the bacterial community of Leuconostoc, and Bacillus and Aeromonas, respectively. Moreover, the enhanced fermentation quality of alfalfa silage by the supplementation of PEC and PEE might be attributed to other unclassified genera. This study provides an implication that pectin degradation is an important determinant for alfalfa silage fermentation through the rescheduling of bacterial community diversity.

Screening of Cellulolytic Actinomycetes for Decomposition of Agricultural Waste

Each year, Viet Nam produces around 37 Mt of rice, 17-18 Mt of sugarcane and 4.5 Mt of maize and the total waste created by agriculture is estimated more than 50 Mt. Approximately nearly half of the straw is burned on the field, which is a common practice in intensive rice cultivation systems in this region and significantly impacts greenhouse gas emissions in the country. It has been a challenge for the government to reduce the burning of straw. Using microbials is a potential solution for the reason that fertilized straw, which contains essential nutrients as nitrogen, phosphorous and potassium, can be used as compost to enrich the soil. This study aims to screen for Actinomycetes with high cellulase activities for the decomposition of agricultural wastes. Among 21 isolated actinomycetes, 7 isolates including G1, G3, G11, D4, D5, DE2, X1 were selected with high enzymatic activities. The incubation of selected strains with rice straw and cane bagasse indicated that the ability of all these strains to degrade agricultural wastes. Rice straw showed better degradation results in comparison to sugarcane bagasse. The strains G3 and D4 showed the best degradation performance after 12 d of incubation. The remaining substrate including rice straw and sugarcane bagasse after incubation in turn with D4 and G3 were both 10.5%.

Effect of xylanase, cellulase and natural maguey extract on the chemical composition of corn silage and in vitro rumen gas production

This study considered the application of two exogenous enzymes, xylanase (XYL) and cellulase (CELL), and maguey extract (ME), (applying 1 ml per kg as fresh matter) and a control (without additive) in four maize silage varieties (San Diego, Cacahuacintle, P-1832 and Victoria), to investigate their effect on the chemical composition (CC), gas production (GP) and in vitro ruminal fermentation. The GP was measured at 0, 3, 6, 9, 12, 24, 36, 48, 72 and 96 hours of incubation. Dry matter disappearance (DMD), organic matter disappearance (OMD), metabolizable energy (ME), and short chain fatty acids (SCFAs) were determined after 96 h of incubation. Data were analyzed using a completely randomized design with a 4×4 factorial arrangement with three replications. The CC showed a significant effect (P<0.05) for varieties with the exception of organic matter (OM), and the inclusion of additives increased the dry matter (DM) and crude protein (CP). A significant effect (P<0.01) was observed for the varieties in the GP parameters and ruminal fermentation. The addition of XYL, CEL and ME promoted dry matter degradation and increased energy availability, with increased in vitro gas production.

Quality attributes of fresh common bean during storage as postharvest treatment with 1-MCP

ABSTRACT Fresh common beans were treated with 1-MCP at 0.5, 1.0, and 1.5 μL L−1 for 20 h at 25°C in an airtight chamber along with a control sample to evaluate the effect of 1-MCP concentration on the quality of fresh common bean during storage. The result showed that postharvest treatment with 0.5 μL L−11-MCP significantly suppressed the increase of peroxidase (POD), cinnamyl-alcohol dehydrogenase (CAD), phenylalnine ammonialyase (PAL), and cellulase (CEL) activity in fresh common bean, inhibited the transport of reducing sugar and increase in cellulose, delayed the increase in the length of pod tendon and electrolyte leakage, and retarded the decrease in the relative thickness of pods and firmness, and maintained the quality of fresh common beans. Our finding that postharvest 0.5 μL L−11-MCP could be used to maintain the quality of the fresh common bean.

Enhancement and bioavailability of phenolic content in Kappaphycus alvarezii through solid substrate fermentation

Known as carrageenan, Kappaphycus alvarezii is a popular species of red edible seaweed which is mainly cultivated for its hydrocolloids. K. alvarezii is a good source of phenolics which are considered to be natural antioxidants that promote myriads of health benefits. Extraction of phenolic compounds using conventional solvent extraction may result in hazards and toxicological effect and low yield of bound phenolics. Therefore, this study was conducted to increase the release of bound phenolics through solid substrate fermentation. In the present study, three different varieties of K. alvarezii were fermented with Aspergillus oryzae through solid substrate fermentation (SSF) at 30 ℃ for 2 to 6 days. The changes of total phenolic content and antioxidant activity due to the effects of SSF were investigated. Results obtained revealed that the highest total phenolic content for green (10.022 mg GAE/g) and purple (4.037 mg GAE/g) varieties of K.alvarezii were on the fourth day of fermentation while for yellow (4.479 mg GAE/g) variety was on the sixth day of fermentation. A remarkable enhancement of antioxidant activity was discovered through DPPH radical scavenging activity on the fourth day of fermentation for all varieties of K. alvarezii tested. Total phenolic content also showed a significant correlation with antioxidant activity of fermented seaweed. Cellulase, xylanase, and β-glucosidase enzymes produced by A.oryzae during SSF were also investigated to see the relationship with the release of phenolic compounds after fermentation. The highest cellulase activity was observed on the fourth day of fermentation. The similar observation was also found for xylanase enzyme in yellow and purple varieties of K. alvarezii , except for green variety where the highest xylanase activity was on the sixth day of fermentation. The maximum β-glucosidase activity in yellow and green varieties was observed on the fourth day of fermentation while purple variety exhibited the maximum β-glucosidase activity on the sixth day. There are positive correlation between the enzymes studied and the polyphenols content in all varieties of K. alvarezii . This study demonstrated that fermented K. alvarezii contained more phenolic content compared to non-fermented seaweed.

Silage fermentation and ruminal degradation of cassava foliage prepared with microbial additive

To effectively utilize the tropical cassava (Manihot esculenta Crantz) foliage (CF) resources, the CF silages were prepared with microbial additives, including Chikuso-1 (CH1, Lactobacillus plantarum), Snow Lact L (SN, L. rhamnosus), Acremonium cellulase (CE), SN + CE and CH1 + CE. Silage fermentation, chemical composition and ruminal degradation were studied in Hainan, China. CF silages prepared with lactic acid bacteria (LAB) and CE were well preserved, with a higher (P < 0.05) lactic acid, a lower (P < 0.05) pH value, butyric acid content and NH3-N ⁄ total-N compared with the controls. The additive-treated silages showed increased crude protein (CP) content, but decreased (P < 0.05) NDF and ADF contents. Meanwhile, the additive treatment improved relative feed value and ruminal degradability of dry matter (DM), CP, neutral detergent fiber and acid detergent fiber. In addition, the combination of LAB and CE resulted in better fermentation quality and ruminal degradability compared with LAB or CE single treatment. The results demonstrated that the CF could be prepared as ruminant feed, and the combination of LAB and CE might exert beneficial synergistic effect on silage fermentation.

Effects of ferulic acid esterase-producing Lactobacillus fermentum and cellulase additives on the fermentation quality and microbial community of alfalfa silage.

BackgroundAlfalfa (Medicago sativa) is an important forage material widely used for animal feed production. Ensiling is an effective method for preserving alfalfa, but it has shown some limitations in the production of high-quality alfalfa silage due to its low water soluble carbohydrates (WSC) content and high buffering capacity. Lactic acid bacteria (LAB) and cellulase are often used as silage additives to promote the ensiling process and enhance fermentation quality.MethodsExperiments were conducted to investigate the effects of ferulic acid esterase (FAE)-producing Lactobacillus fermentum 17SD-2 (LF) and cellulase (CE) on the fermentation quality and microbial community of alfalfa silage. After 60 days of ensiling, analysis of fermentation quality and bacterial diversity in alfalfa silages were conducted using high-performance liquid chromatography and high-throughput sequencing methods.ResultsAlfalfa was ensiled with additives (LF, CE, and LF+CE) or without additives for 60 days. All additives increased lactic acid and decreased pH values and ammonia-N contents compared to control. Among all treatments, the combined addition of LF and CE showed lowest pH (4.66) and ammonia-N (NH3-N, 0.57% DM) content, highest contents of lactic acid (LA, 10.51% DM), dry matter (DM, 22.54%) and crude protein (CP, 24.60% DM). Combined addition of LF and CE performed better in reducing neutral detergent fiber (NDF, 29.76% DM) and acid detergent fiber (ADF, 22.86% DM) contents than the addition of LF (33.71, 27.39% DM) or CE (32.07, 25.45% DM) alone. Moreover, the microbial analysis indicated that LF+CE treatments increased the abundance of desirable Lactobacillus and inhibited the growth of detrimental Enterobacter and Clostridia in alfalfa silage.DiscussionCombined addition of FAE-producing LF and CE is more effective than treatments of LF or CE alone in improving fermentation quality and nutrition values of alfalfa silage. This is likely due to a synergistic effect of CE and FAE produced by LF on plant cell wall degradation, indicating that these additives promote each other to improve fiber degradation and silage fermentation. In conclusion, combined addition of FAE-producing LF and CE could be a feasible way to improve alfalfa silage quality.

Stability/activity features of the main enzyme components of rohapect 10L.

Rohapect 10L is an enzyme cocktail commercialized for juice clarification. Here, we characterized the activity and stability of five enzymatic activities present in this cocktail: total pectinase (PE), polygalacturonase (PG), pectin lyase (PL), pectin methyl esterase (PME), and total cellulase (CE) activities. All these enzyme activities have the maximum activity and stability at pH 4, conditions near those found in most fruit juices. However, if the enzymes need to be handled under different conditions (e.g., to immobilize them), their stability becomes extremely low in some cases, just at pH values slightly higher than the optimal one. For example, at pH 10 only CE was reasonably stable at 25°C, while many other enzyme activities were rapidly almost inactivated, even at 4°C. For these cases, different additives were evaluated, and we found that polyethylene glycol was positive or very positive for all enzyme stabilities, allowing keeping reasonable activities after several hours at pH 10 and 25°C. Another additive, that is, dextran, has a small positive effect for PE, PG, and CE, and a very positive effect for PL, albeit significantly destabilizing PME. Thus, the handling and use of this extract requires some care when is performed out of optimal conditions.

Optimization of fermentation factors to enhance rice straw degradation ability using a microbial consortium LZF-12

Biological pretreatment has broad application prospects in agricultural waste treatment because of its economic, environmental protection and energy saving characteristics. In this study, a microbial consortium LZF-12 was applied as a biological pretreatment to degrade rice straw. Batch experiments were performed to hydrolysis conditions by using the method of Box-Behnken factorial design (BBD). The results showed that the model multiple correlation coefficient R2 was 0.9816, and the effects of three factors on the degradation of rice straw of LZF-12 as descending order were the initial rice content, chicken manure content, and initial pH. The interaction between straw concentration, chicken manure concentration, and initial pH had significant effects on the degradation of a microbial consortium. Under the optimum conditions of 0.86% rice straw, 0.5% chicken manure and the initial pH of 7.0, the degradation rate of rice straw reached 72.4%. There is only small difference of 0.55% between the experimental value and predicted value from BBD model. Therefore, it is feasible for the established model due to the consistent results between the prediction and experimental value. The microbial consortium LZF-12 has high cellulase enzyme activities and degradation ability over a wide range of temperature and pH, indicating it has a good development potential and application prospects in waste biodegradation and biomass energy production. Keywords: microbial consortium, biological pretreatment, biodegradation, rice straw, optimization DOI: 10.25165/j.ijabe.20191203.4638 Citation: Zheng G X, Zhou C Y, Yin T, Lu Z X, Ai S. Optimization of fermentation factors to enhance rice straw degradation ability using a microbial consortium LZF-12. Int J Agric & Biol Eng, 2019; 12(3): 168–173.

Using a linear pH-responsive zwitterionic copolymer to recover cellulases in enzymatic hydrolysate and to enhance the enzymatic hydrolysis of lignocellulose

A linear pH-responsive zwitterionic copolymer P(MDB) was synthesized from functional monomers methacrylic acid, 2-(dimethylamino) ethyl methacrylate (DMA) and benzyl methacrylate (BMA) through free radical polymerization to recover cellulases and enhance the enzymatic hydrolysis. The hydrophobicity and molecular weight of the copolymer was controlled by the feed ratio of monomers and the mass ratio of initiator to monomers, respectively. UV-turbidity showed P(MDB) exhibited sensitive pH-responsiveness with dissolution–precipitation ΔpH about 1 pH unit. The precipitation pHφ of P(MDB) was 5.0–5.5 and over 90% of P(MDB) could precipitate in the vicinity of its isoelectric point. Increasing the hydrophobicity or molecular weight of P(MDB) could improve its pH sensitivity and precipitation rate. P(MDB) could enhance the enzymatic hydrolysis of dilute acid pretreated Eucalyptus by 10–25%. P(MDB) could recover the free cellulases in enzymatic hydrolysate by co-precipitation. SDS-PAGE analysis showed that P(MDB) could recover 100% β-glucosidase (β-GL) and the recovery rates of cellobiohydrolase (CBH) and endoglucanase (EG) increased as the hydrophobicity and molecular weight of P(MDB) increased. The mechanism was proposed that hydrophobic affiliation was the main binding force of P(MDB) to cellulases. Increasing the hydrophobicity and molecular weight of copolymer could increase its binding force and the number of binding sites to cellulases thus the overall cellulase recovery performance was improved.

Impact of wilting and additives on fermentation quality and carbohydrate composition of mulberry silage.

ObjectiveThe objective of this study was to investigate the effects of wilting and additives on the fermentation quality, structural and non-structural carbohydrate composition of mulberry silages.MethodsThe selected lactic acid bacteria strains Lactobacillus plantarum ‘LC279063’ (L1), commercial inoculant Gaofuji (GF), and Trichoderma viride cellulase (CE) were used as additives for silage preparation. Silage treatments were designed as control (CK), L1, GF, or CE under three wilting rates, that is wilting for 0, 2, or 4 hours (h). After ensiling for 30 days, the silages were analyzed for the chemical and fermentation characteristics.ResultsThe results showed that wilting had superior effects on increasing the non-structural carbohydrate concentration and degrading the structural carbohydrate. After ensiling for 30 days, L1 generally had a higher fermentation quality than other treatments, indicated by the lower pH value, acetic acid, propionic acid and ammonia nitrogen (NH3-N) content, and the higher lactic acid, water soluble carbohydrate, glucose, galactose, sucrose, and cellobiose concentration (p<0.05) at any wilting rate. Wilting could increase the ratio of lactic acid/acetic acid and decrease the content of NH3-N.ConclusionThe results confirmed that wilting degraded the structural carbohydrate and increased the non-structural carbohydrate; and L1 exhibited better properties in improving fermentation quality and maintaining a high non-structural carbohydrates composition compared with the other treatments.