Commercial Xylanase Research Articles

A xylanase-aided enzymatic pretreatment facilitates cellulose nanofibrillation

Although biological pretreatment of cellulosic fiber based on endoglucanases has shown some promise to facilitate cellulose nanofibrillation, its efficacy is still limited. In this study, a xylanase-aided endoglucanase pretreatment was assessed on the bleached hardwood and softwood Kraft pulps to facilitate the downstream cellulose nanofibrillation. Four commercial xylanase preparations were compared and the changes of major fiber physicochemical characteristics such as cellulose/hemicellulose content, gross fiber properties, fiber morphologies, cellulose accessibility/degree of polymerization (DP)/crystallinity were systematically evaluated before and after enzymatic pretreatment. It showed that the synergistic cooperation between endoglucanase and certain xylanase (Biobrite) could efficiently “open up” the hardwood Kraft pulp with limited carbohydrates degradation (<7%), which greatly facilitated the downstream cellulose nanofibrillation during mild sonication process (90Wh) with more uniform disintegrated nanofibril products (50–150nm, as assessed by scanning electron microscopy and UV–vis spectroscopy).

Direct hydrolysis of palm oil mill effluent by xylanase enzyme to enhance biogas production using two-steps thermophilic fermentation under non-sterile condition

Palm oil mill effluent (POME) is well known as the potential raw material for biogas production. However, the high hemicellulose and cellulose content (10–14% w/w in dry basis) of the palm fibre in POME are less accessible for biodegradable to sugars, therefore, limiting the carbon source for microorganisms for the biogas production. This study attempted to enhance the hydrolysis step of the biomass by enzymatic pretreatment of POME. The optimum temperature and hydrolysis time using the commercial xylanase (5 unit/mL) were at 50 °C for 12 h. Two-steps thermophilic process consisting of POME pretreatment by various concentrations of xylanase (5–20 unit/mL) under the optimum condition followed by biogas production at 60 °C for 45 days was carried out. In all cases, the substrate to inoculum volatile solids (VS) ratio and volume ratio were 1:1 and 4:1, respectively. The maximum biomethane (CH4) yield of 914 mL CH4/g VS was obtained from the fermentation of POME hydrolysed by 15 unit/mL of xylanase. However, it was not significantly different from those of the other three concentrations of xylanase (845, 870 and 851 mL CH4/g VS, respectively). They were about 2.5–3 folds higher than that of the control (POME without enzymatic hydrolysis) (297 mL CH4/g VS). Microbial community analysis revealed the presence of Clostridium sp. and Methanocaldococcus sp. as the dominant strains.

Production of a recombinant swollenin from Trichoderma harzianum in Escherichia coli and its potential synergistic role in biomass degradation

BackgroundFungal swollenins (SWOs) constitute a class of accessory proteins that are homologous to canonical plant expansins. Expansins and expansin-related proteins are well known for acting in the deagglomeration of cellulose structure by loosening macrofibrils. Consequently, SWOs can increase the accessibility and efficiency of the other enzymes involved in the saccharification of cellulosic substrates. Thus, SWOs are promising targets for improving the hydrolysis of plant biomass and for use as an additive to enhance the efficiency of an enzyme cocktail designed for the production of biofuels.ResultsHere, we report the initial characterization of an SWO from Trichoderma harzianum (ThSwo) that was successfully produced using Escherichia coli as a host. Initially, transcriptome and secretome data were used to compare swo gene expression and the amount of secreted ThSwo. The results from structural modeling and phylogenetic analysis of the ThSwo protein showed that ThSwo does preserve some structural features of the plant expansins and family-45 glycosyl hydrolase enzymes, but it evolutionarily diverges from both of these protein classes. Recombinant ThSwo was purified at a high yield and with high purity and showed secondary folding similar to that of a native fungal SWO. Bioactivity assays revealed that the purified recombinant ThSwo created a rough and amorphous surface on Avicel and displayed a high synergistic effect with a commercial xylanase from T. viride, enhancing its hydrolytic performance up to 147 ± 7%.ConclusionsMany aspects of the structure and mechanism of action of fungal SWOs remain unknown. In the present study, we produced a recombinant, active SWO from T. harzianum using a prokaryotic host and confirmed its potential synergistic role in biomass degradation. Our work paves the way for further studies evaluating the structure and function of this protein, especially regarding its use in biotechnology.

Expanding the feruloyl esterase gene family of Aspergillus niger by characterization of a feruloyl esterase, FaeC

A feruloyl esterase (FAE) from Aspergillus niger N402, FaeC was heterologously produced in Pichia pastoris X-33 in a yield of 10mg/L. FaeC was most active at pH 7.0 and 50°C, and showed broad substrate specificity and catalyzed the hydrolysis of methyl 3,4-dimethoxycinnamate, ethyl ferulate, methyl ferulate, methyl p-coumarate, ethyl coumarate, methyl sinapate, and methyl caffeate. The enzyme released both ferulic acid and p-coumaric acid from wheat arabinoxylan and sugar beet pectin (up to 3mg/g polysaccharide), and acted synergistically with a commercial xylanase increasing the release of ferulic acid up to six-fold. The expression of faeC increased over time in the presence of feruloylated polysaccharides. Cinnamic, syringic, caffeic, vanillic and ferulic acid induced the expression of faeC. Overall expression of faeC was very low in all tested conditions, compared to two other A. niger FAE encoding genes, faeA and faeB. Our data showed that the fae genes responded differently towards the feruloylated polysaccharides and tested monomeric phenolic compounds suggesting that the corresponding FAE isoenzymes may target different substrates in a complementary manner. This may increase the efficiency of the degradation of diverse plant biomass.

Post-hydrolysis of the prehydrolysate from eucalyptus pulping with xylanase

Sugar-rich effluent from prehydrolysis of eucalyptus wood (prehydrolysate) is an abundant byproduct of paper and pulp industries. It can be used as a resource for xylose production and biorefinery. However, it is mostly concentrated and burned for energy production. In this paper, a commercial xylanase was applied to examine the feasibility of enzymatic treatment of the prehydrolysate. The xylanase efficiently hydrolyzed the xylooligosaccharides in the prehydrolysate to xylose. The optimal hydrolysis conditions were determined to be at pH 5.0, a temperature of 50 °C, a concentration of xylooligosaccharides at 23.46 g/L, and xylanase loading of 75 U/g xylooligosaccharides. Using the aforementioned reaction conditions, xylooligosaccharides in the prehydrolysate were completely hydrolyzed within 24 h. The total xylose concentration reached 50.41 g/L. The xylose was recovered by active carbon decolorization, ion exchange chromatography, desalination, and crystallization, which produced crystal xylose with a 97% purity at a 35% recovery rate. The results demonstrated that xylanase is an efficient and ecofriendly tool for eucalyptus pulping prehydrolysate. Moreover, the prehydrolysate from hardwood pulp industry can serve as an abundant and low cost resource for industrial xylose production.

Development of efficient cellulolytic enzyme display on the cell surface of the yeast Saccharomyces cerevisiae

Sugar-rich effluent from prehydrolysis of eucalyptus wood (prehydrolysate) is an abundant byproduct of paper and pulp industries. It can be used as a resource for xylose production and biorefinery. However, it is mostly concentrated and burned for energy production. In this paper, a commercial xylanase was applied to examine the feasibility of enzymatic treatment of the prehydrolysate. The xylanase efficiently hydrolyzed the xylooligosaccharides in the prehydrolysate to xylose. The optimal hydrolysis conditions were determined to be at pH 5.0, a temperature of 50 °C, a concentration of xylooligosaccharides at 23.46 g/L, and xylanase loading of 75 U/g xylooligosaccharides. Using the aforementioned reaction conditions, xylooligosaccharides in the prehydrolysate were completely hydrolyzed within 24 h. The total xylose concentration reached 50.41 g/L. The xylose was recovered by active carbon decolorization, ion exchange chromatography, desalination, and crystallization, which produced crystal xylose with a 97% purity at a 35% recovery rate. The results demonstrated that xylanase is an efficient and ecofriendly tool for eucalyptus pulping prehydrolysate. Moreover, the prehydrolysate from hardwood pulp industry can serve as an abundant and low cost resource for industrial xylose production.

Feruloyl esterase production by Aspergillus terreus CECT 2808 and subsequent application to enzymatic hydrolysis.

Ferulic acid esterases (FAE) were produced by Aspergillus terreus CECT 2808 from vine trimming shoots (VTS) and corn cob. Later, the fungal extracts thus obtained were used to enzymatically release ferulic acid (FA) from both substrates. Our findings showed a higher FAE activity in the enzymatic extracts produced on corn cob (0.070±0.004U/mL). Nevertheless, the enzymatic extracts produced on VTS demonstrated a better performance for FA release from both corn cob (2.05±0.01mg/g) and VTS (0.19±0.003mg/g). This result was probably because of the higher xylanase/FAE ratio determined in VTS extract. Therefore, an additional assay was carried out by supplementing corn cob extract with a commercial xylanase to test the influence of FAE/xylanase ratio in FA release. The results revealed the relevance of the FAE/xylanase ratio for an optimal FA release.

A xylanase from Streptomyces sp. FA1: heterologous expression, characterization, and its application in Chinese steamed bread.

Xylanases (EC 3.2.1.8) are hydrolytic enzymes that have found widespread application in the food, feed, and paper-pulp industries. Streptomyces sp. FA1 xynA was expressed as a secreted protein in Pichia pastoris, and the xylanase was applied to the production of Chinese steamed bread for the first time. The optimal pH and the optimal temperature of XynA were 5.5 and 60 °C, respectively. Using beechwood as substrate, the K m and V max were 2.408 mg mL(-1) and 299.3 µmol min(-1) mg(-1), respectively. Under optimal conditions, a 3.6-L bioreactor produced 1374 U mL(-1) of XynA activity at a protein concentration of 6.3 g L(-1) after 132 h of fermentation. Use of recombinant XynA led to a greater increase in the specific volume of the CSB than could be achieved using commercial xylanase under optimal conditions. This study provides the basis for the application of the enzyme in the baking industry.

Feruloyl esterase from Aspergillus clavatus improves xylan hydrolysis of sugarcane bagasse

Feruloyl esterase is a subclass of carboxylic acid esterases with the capacity to release ferulic acid and other cinnamic acids from plant cell walls and synthetic substrates. Feruloyl esterases act synergistically with xylanases removing ferulic acid residues esterified to arabinoxylans. Feruloyl esterase type D from Aspergillus clavatus (AcFAE) was expressed in Escherichia coli, purified, and applied with a commercial xylanase consortium (Novozymes) for hydrolysis of sugarcane bagasse. Feruloyl esterase plus xylanase increased 5.13-fold the releasing of ferulic acid from sugarcane bagasse. Removal of only 7.7% of ferulic acid content by AcFAE increased 97.3% the sugarcane bagasse hydrolysis by xylanase. These data support the use of AcFAE as an interesting adjuvant enzyme to improve lignocellulose digestion and biotechnological tool for biorefineries.

Stability of Commercial Xylanases stored singly or Combined with Vitamins and/or Minerals Premix at different Temperatures

Stability of Commercial Xylanases stored singly or Combined with Vitamins and/or Minerals Premix at different Temperatures

Amenability of Acacia and Eucalyptus Hardwood Pulps to Elemental Chlorine-Free Bleaching: Application and Efficacy of Microbial Xylanase

This study outlines the results of a biobleaching study of acacia (A. mangium) and eucalyptus (E. globulus) hardwood kraft pulps with commercial xylanase (Optimase CX 72 L). The comparative study was carried out using an elemental chlorine-free (ECF) bleaching sequence (D0EPD1D2) after the enzyme (X) stage. The enzyme treatment resulted in improved optical properties with a reduction in bleach chemical consumption. At an equivalent bleach chemical consumption, a brightness gain of 2.1 and 1.7 units and a whiteness gain of 2.7 and 2.3 units were observed with xylanase treatment in acacia and eucalyptus pulps, respectively. In ECF bleaching using the D0EPD1D2 sequence, a final brightness was achieved to the extent of 90% ISO and 89% ISO for acacia and eucalyptus, respectively, at an equivalent charge of bleach chemicals. The post-color (PC) number was also reduced by up to 45% for both hardwood pulps compared with the control. The bleachability of acacia was observed to be significantly higher than that of eucalyptus. In addition, a 17.0% and 23.0% reduction in chlorine dioxide and sodium hydroxide, respectively, were obtained for both hardwood pulps after xylanase pre-bleaching, thus indicating an environmentally friendly approach to the process.

Comparison and Optimization of Thermostable Xylanase Production by Bacillus Pumilus and Bacillus Cereus using Corn Husk

One of the most studied and widely used commercial Xylanase is endo-1, 4-s-xylanases.It is useful because the specific activity of Xylanase is higher than the other hydrolytic xylanases. Therefore, xylanases are commonly used in processes involved in degradation of plant materials, paper and pulp bleaching. The present study investigate the production and optimization of thermostable Xylanase by the bacteria Bacillus pumilus (MTCC 10209) and Bacillus cereus (MTCC 10202) under solid state fermentation using corn husk as carbon source since corn husk contain Xylan (28%) and Xylose (17.2%). Maximum production of Xylanase was observed to be 2.2019 (U/ml) and 2.5010 (U/ml) for B.cereus and B.pumilus respectively. The Response Surface Methodology revealed that the optimum temperature, pH and inoculums size for the enzyme activity were 65⁰C, pH 7.0 and 1 (%v/w) for B.cereus, whereas for B.pumilus it was 65⁰C, pH 7.0 and 1.50 (%v/w) respectively. The increasing concentration of metal ion (FeSo₄) ranging from 1mM- 10mM increases the enzyme activity of B.pumilus. Meanwhile, the enzyme activity of B.cereus was strongly inhibited by increasing metal ion concentration.

Effect of enzymatic and technological treatments on solubilisation of arabinoxylans from brewer's spent grain

Brewer's spent grain (BSG), the most abundant brewing by-product, has hidden and underexploited nutritional potential. In order to valorize BSG, the effects of three commercial xylanases and a peptidase on water unextractable arabinoxylans (WUAX) were studied. Comparing all treatments, higher addition of xylanase resulted in an increase in water extractable arabinoxylans (WEAX). In the most efficient treatment, xylanase alone was able to solubilise 23.7% of WUAX, while the peptidase showed no effect. However, when added together with xylanase, peptidase increased the solubilisation of WUAX up to 1.6 folds. A positive correlation between particle size reduction and solubilisation of WUAX was also proved through milling BSG. These results suggest that access to xylan backbone increases with proteolytic activities, proving a synergistic effect of these specific enzymes. Therefore, if properly treated before being added as ingredient, BSG could add health functionalities to foodstuff while reducing the environmental impact of brewing industries.

Growth performance and endogenous losses of broilers fed wheat-based diets with and without essential oils and xylanase supplementation

An experiment was conducted to compare the effect of a supplementary mixture of essential oils, with and without exogenous xylanase, on performance, carcass composition, dietary nitrogen (N)–corrected apparent metabolizable energy (AMEn), dry matter retention (DMR), N retention (NR), fat digestibility (FD) coefficients, and endogenous mucin losses (measured as sialic acid, SA) when fed to broiler chickens.Three hundred male Ross 308 broilers in total were reared in floor pens from 0 to 21 d of age. Birds were fed 1 of 3 wheat-based diets: basal diet (215 g/kg CP, 12.12 MJ/kg AME) with either no additive (control diet; C) or 100 g/tonne of a standardized combination of 5% carvacrol, 3% cinnamaldehyde, and 2% capsicum oleoresin (diet XT); or a combination of XT and commercial xylanase enzyme at a rate of 100 g of XT and 2,000 units (U) of xylanase/kg (diet XYL), respectively. Each diet was randomly allocated to 10 pens with 10 birds.Feeding XT and XYL diets improved birds’ growth performance (P < 0.05). Birds fed XT and XYL diets had an improved caloric conversion ratio (P < 0.05) and consumed 1.3 MJ less AMEn per kilogram of growth compared to birds fed the control diet only. Feeding XT improved only the dietary FD coefficient (P < 0.05) compared to control-fed birds, but the dietary FD coefficient did not differ for XYL diet (P > 0.05). Birds fed XYL diet excreted 35% less endogenous mucin compared to control-fed birds (P < 0.05). Birds fed XT alone gained more carcass protein than the control-fed birds (P < 0.05) but did not differ from the birds fed XYL diet (P > 0.05). There was no indication of a negative interaction between dietary essential oils and xylanase.

The Impact of Xylanase and Hot Acid Pretreatment on HexAs in Eucalyptus Kraft Pulp Bleaching

The influence of the presence of hexenuronic acid (HexA) in eucalyptus kraft pulp on the efficiency of D0EpD1 was investigated by separately using both commercial cellulase-free xylanase and hot acid pretreatments. In comparison to the xylanase pretreatment, the hot acid pretreatment was more than four-fold effective in removing the HexAs (61%) and concomitantly reduced the kappa number, but resulted in lower viscosity. After subsequent D0EpD1 bleaching, the final brightness and its stability were improved remarkably, especially under xylanase pretreatment. At the same brightness of 85% ISO, the xylanase pretreatment reduced the consumption of active chlorine (35.6%) more than the acid pretreatment did (27.5%). The total relative active chlorine savings for the xylanase pretreatment over the acid pretreatment was therefore nearly 23%. The HexA component can be removed considerably in both D0 and D1 stages during the D0EpD1 bleaching sequence, while only slightly in the Ep stage. The use of xylanase and acid pretreatment both resulted in insignificant physical strength losses for the bleached pulps.

Effect of exogenous xylanase on rumen in vitro gas production and degradability of wheat straw.

The objective of this study was to determine effects of xylanase on in vitro gas production (GP) and in sacco degradability of wheat straw. Rumen fluid was obtained from three Mongolian native goats fitted with permanent rumen cannulas. The trial consisted of five doses (0, 0.5, 1.0, 1.5, 2.0 μL/g of substrate) of a commercial xylanase (Dyadic® xylanase PLUS, Dyadic International, Inc., Jupiter, FL, USA). For the in sacco degradability, different levels of xylanase enzyme were added directly onto 2 g of wheat straw in nylon bags and incubated in the rumen for 3, 6, 12, 24 and 48 h to estimate degradability of wheat straw. Total GP increased (P < 0.001) at all times of incubation at intermediate levels of xylanase. Methane production had a similar pattern at 3 and 12 h of incubation; increased linearly at 24 h of incubation, and was unaffected at 6 and 48 h of incubation. Rumen NH3 -N concentration increased linearly at 3 h and the highest values were observed with intermediate enzyme levels. All ruminal volatile fatty acids increased linearly with intermediate levels of the fibrolytic enzyme. The in sacco rate of dry matter degradation decreased linearly (P = 0.020) with increasing enzymes. Intermediate levels of xylanase improved rumen kinetic fermentation and degradability. The outcome of this research indicated that the application of xylanase enzyme could improve in vitro GP fermentation of wheat straw.

Enzymatic Production of a Soluble Fiber Hydrolyzate from Chinese Cabbage Waste and Its Health-Related Properties

A feasibility study on the potential use of Chinese cabbage waste as a raw material for the production of soluble dietary fiber was performed. The alkali-soluble fraction obtained by extracting Chinese cabbage waste was hydrolyzed by two commercial xylanase preparations to produce soluble fiber hydrolyzates. The freeze-dried soluble fiber was treated with 85% ethanol and separated into the alcohol-soluble dietary fiber (ASF) and alcohol-insoluble dietary fiber (AIF) fractions. ASF had a significantly greater promoting effect on the growth of lactic acid bacteria. AIF effectively hindered the diffusion of glucose and bile acid from dialysis membranes and displayed a significantly greater bile acid retarding effect than carboxymethylcellulose and pectin. Thus, the water-soluble dietary fiber obtained from Chinese cabbage waste by enzymatic hydrolysis has potential use as a fiber source with prebiotic, hypoglycemic and hypolipidemic effects. Practical Applications Dietary fiber, which has many health-related benefits, is added to various food products as a functional ingredient or acceptability factor. The aim of this study was to produce soluble dietary fiber from Chinese cabbage waste by enzymatic hydrolysis. Our results indicated that soluble fiber hydrolyzate from Chinese cabbage is a practical new resource for preparing functional drinks and nutraceutical products for use in the food industry. The use of agro-industrial waste to produce bioactive ingredients could improve the economic value of such waste.

Immobilization of a 22kDa Xylanase on Eudragit L-100 for Xylo-oligosaccharide Production

A commercial xylanase was immobilized on Eudragit L-100 with the recovery activity of 87.3-126.2%. After immobilization, the optimum pH was not changed while its optimum temperature moved from 70 to 75°C. However, the immobilized enzyme did not show a higher thermal stability than the free xylanase. A slightly decrease in the Km values of free enzyme was observed upon immobilization. Meanwhile, Vmax values of the immobilized enzyme were 38% higher than those of the free xylanase. The result of Fluorescence spectroscopy used to probe the changes in the enzyme structure upon immobilization showed minute change and which may result in increase of immobilized enzyme activity. The application of immobilized enzyme hydrolyzing cottonseed husk and corncob pretreated by alkaline peroxide solution was the basically same as the free enzyme. The reusability of immobilized enzyme in the third cycles produced 163 and 126% reducing sugar hydrolyzing cottonseed husk and corncob.

Screening of Xylanolytic Aspergillus fumigatus for Prebiotic Xylooligosaccharide Production Using Bagasse.

Sugarcane bagasse is an important lignocellulosic material studied for the production of xylooligosaccharides (XOS). Some XOS are considered soluble dietary fibre, with low caloric value and prebiotic effect, but they are expensive and not easily available. In a screening of 138 fungi, only nine were shortlisted, and just Aspergillus fumigatus M51 (35.6 U/mL) and A. fumigatus U2370 (28.5 U/mL) were selected as the most significant producers of xylanases. These fungi had low β-xylosidase activity, which is desirable for the production of XOS. The xylanases from Trichoderma reesei CCT 2768, A. fumigatus M51 and A. fumigatus U2370 gave a significantly higher XOS yield, 11.9, 14.7 and 7.9% respectively, in a 3-hour reaction with hemicellulose from sugarcane bagasse. These enzymes are relatively thermostable at 40-50 °C and can be used in a wide range of pH values. Furthermore, these xylanases produced more prebiotic XOS (xylobiose and xylotriose) when compared with a commercial xylanase. The xylanases from A. fumigatus M51 reached a high level of XOS production (37.6%) in 48-72 h using hemicellulose extracted from sugarcane bagasse. This yield represents 68.8 kg of prebiotic XOS per metric tonne of cane bagasse. In addition, in a biorefinery, after hemicellulose extraction for XOS production, the residual cellulose could be used for the production of second-generation ethanol.

In silico Study of Bacillus brevis Xylanase - Structure Prediction and Comparative Analysis with Other Bacterial and Fungal Xylanase

The most important building block of hemicelluloses is xylan. It is broken down into xylose oligomer residues by Xylanase - an enzyme, produced by most organisms, to utilize xylose as primary source of carbon. The Xylanase produced are classified into families, viz 5, 8, 10, 11 and 43 - of Glycoside Hydrolases (GH). Xylanase from family GH 11 are monospecific, they consist solely of Xylanase activity, exclusively active on D-xylose containing substrates.They are inactive on aryl cellobiosides. The fungal Xylanase are produced in higher concentrations, as compared to bacterial Xylanase, but have limited use in pulp bleaching, as they affect the viscosity and strength of the product. In the present study, we have worked upon the Xylanase of Bacillus brevis, which is fulfilling all the required quality needed to be a commercial Xylanase, and thus is used by many industries. The enzyme, when studied after modelling, provided similar structural configuration with high stability. When compared with other bacterial and fungal Xylanase structures, it provided better potential to ‘activity enhancement’ and ‘in silico handling’.