Modulating the production of xylanase by Bacillus pumilus BS131 through optimization using waste fiber sludge / Modulando a produção de xilanase por Bacillus pumilus BS131 por meio da otimização usando lodo de fibra residual

Abstract In recent days, cheapest alternative carbon source for fermentation purpose is desirable to minimize production cost. Xylanases have become attractive enzymes as their potential in bio-bleaching of pulp and paper industry. The objective of the present study was to identify the potential ability on the xylanase production by locally isolated Bacillus pumilus BS131 by using waste fiber sludge and wheat bran media under submerged fermentation. Culture growth conditions were optimized to obtain significant amount of xylanase. Maximum xylanase production was recorded after 72 hours of incubation at 30 °C and 7 pH with 4.0% substrate concentration. In the nutshell, the production of xylanase using inexpensive waste fiber sludge and wheat-bran as an alternative in place of expensive xylan substrate was more cost effective and environment friendly.

Resumo Nos últimos dias, a fonte alternativa de carbono mais barata para fins de fermentação é desejável para minimizar o custo de produção. As xilanases têm se tornado enzimas atraentes como seu potencial no biobranqueamento da indústria de papel e celulose. O objetivo do presente estudo foi identificar a capacidade potencial na produção de xilanase por Bacillus pumilus BS131 isolado localmente usando lodo de fibra residual e farelo de trigo em meio de fermentação submersa. As condições de crescimento da cultura foram otimizadas para obter uma quantidade significativa de xilanase. A produção máxima de xilanase foi registrada após 72 horas de incubação a 30 °C e pH 7 com concentração de substrato de 4,0%. Resumindo, a produção de xilanase usando lodo de fibra residual de baixo custo e farelo de trigo como uma alternativa no lugar do substrato de xilano caro foi mais econômica e ecológica.

Immobilization of mutated xylanase from Neocallimastix patriciarum in E. coli and application for kraft pulp biobleaching / Imobilização de xilanase mutada de Neocallimastix patriciarum em E. coli e aplicação para polpa kraft

Abstract As an important enzyme, xylanase is widely used in the food, pulp, and textile industry. Different applications of xylanase warrant specific conditions including temperature and pH. This study aimed to carry out sodium alginate beads as carrier to immobilize previous reported mutated xylanase from Neocallimastix patriciarum which expressed in E. coli, the activity of immobilization of mutated xylanase was elevated about 4% at pH 6 and 13% at 62 °C. Moreover, the immobilized mutated xylanase retained a greater proportion of its activity than the wide type in thermostability. These properties suggested that the immobilization of mutated xylanase has potential to apply in biobleaching industry.

Resumo Como importante enzima, a xilanase é amplamente utilizada na indústria alimentícia, de celulose e têxtil. Diferentes aplicações de xilanase garantem condições específicas, incluindo temperatura e pH. Este estudo teve como objetivo realizar grânulos de alginato de sódio como carreador para imobilizar xilanase mutada relatada anteriormente de Neocallimastix patriciarum que expressa em E. coli, a atividade de imobilização da xilanase mutada foi elevada em cerca de 4% em pH 6 e 13% a 62 °C. Além disso, a xilanase mutada imobilizada reteve uma proporção maior de sua atividade do que o tipo amplo em termoestabilidade. Essas propriedades sugerem que a imobilização da xilanase mutada tem potencial para aplicação na indústria de biobranqueamento.

Immobilization of mutated xylanase from Neocallimastix patriciarum in E coli and application for kraft pulp biobleaching / Imobilização de xilanase mutada de Neocallimastix patriciarum em E coli e aplicação para polpa kraft

As an important enzyme, xylanase is widely used in the food, pulp, and textile industry. Different applications of xylanase warrant specific conditions including temperature and pH. This study aimed to carry out sodium alginate beads as carrier to immobilize previous reported mutated xylanase from Neocallimastix patriciarum which expressed in E. coli, the activity of immobilization of mutated xylanase was elevated about 4% at pH 6 and 13% at 62 °C. Moreover, the immobilized mutated xylanase retained a greater proportion of its activity than the wide type in thermostability. These properties suggested that the immobilization of mutated xylanase has potential to apply in biobleaching industry.

Como importante enzima, a xilanase é amplamente utilizada na indústria alimentícia, de celulose e têxtil. Diferentes aplicações de xilanase garantem condições específicas, incluindo temperatura e pH. Este estudo teve como objetivo realizar grânulos de alginato de sódio como carreador para imobilizar xilanase mutada relatada anteriormente de Neocallimastix patriciarum que expressa em E. coli, a atividade de imobilização da xilanase mutada foi elevada em cerca de 4% em pH 6 e 13% a 62 °C. Além disso, a xilanase mutada imobilizada reteve uma proporção maior de sua atividade do que o tipo amplo em termoestabilidade. Essas propriedades sugerem que a imobilização da xilanase mutada tem potencial para aplicação na indústria de biobranqueamento.

Modulating the production of xylanase by Bacillus pumilus BS131 through optimization using waste fiber sludge / Modulando a produção de xilanase por Bacillus pumilus BS131 por meio da otimização usando lodo de fibra residual

In recent days, cheapest alternative carbon source for fermentation purpose is desirable to minimize production cost. Xylanases have become attractive enzymes as their potential in bio-bleaching of pulp and paper industry. The objective of the present study was to identify the potential ability on the xylanase production by locally isolated Bacillus pumilus BS131 by using waste fiber sludge and wheat bran media under submerged fermentation. Culture growth conditions were optimized to obtain significant amount of xylanase. Maximum xylanase production was recorded after 72 hours of incubation at 30 °C and 7 pH with 4.0% substrate concentration. In the nutshell, the production of xylanase using inexpensive waste fiber sludge and wheat-bran as an alternative in place of expensive xylan substrate was more cost effective and environment friendly.

Nos últimos dias, a fonte alternativa de carbono mais barata para fins de fermentação é desejável para minimizar o custo de produção. As xilanases têm se tornado enzimas atraentes como seu potencial no biobranqueamento da indústria de papel e celulose. O objetivo do presente estudo foi identificar a capacidade potencial na produção de xilanase por Bacillus pumilus BS131 isolado localmente usando lodo de fibra residual e farelo de trigo em meio de fermentação submersa. As condições de crescimento da cultura foram otimizadas para obter uma quantidade significativa de xilanase. A produção máxima de xilanase foi registrada após 72 horas de incubação a 30 °C e pH 7 com concentração de substrato de 4,0%. Resumindo, a produção de xilanase usando lodo de fibra residual de baixo custo e farelo de trigo como uma alternativa no lugar do substrato de xilano caro foi mais econômica e ecológica.

Immobilization of mutated xylanase from Neocallimastix patriciarum in E. coli and application for kraft pulp biobleaching

Abstract As an important enzyme, xylanase is widely used in the food, pulp, and textile industry. Different applications of xylanase warrant specific conditions including temperature and pH. This study aimed to carry out sodium alginate beads as carrier to immobilize previous reported mutated xylanase from Neocallimastix patriciarum which expressed in E. coli, the activity of immobilization of mutated xylanase was elevated about 4% at pH 6 and 13% at 62 °C. Moreover, the immobilized mutated xylanase retained a greater proportion of its activity than the wide type in thermostability. These properties suggested that the immobilization of mutated xylanase has potential to apply in biobleaching industry.

Resumo Como importante enzima, a xilanase é amplamente utilizada na indústria alimentícia, de celulose e têxtil. Diferentes aplicações de xilanase garantem condições específicas, incluindo temperatura e pH. Este estudo teve como objetivo realizar grânulos de alginato de sódio como carreador para imobilizar xilanase mutada relatada anteriormente de Neocallimastix patriciarum que expressa em E. coli, a atividade de imobilização da xilanase mutada foi elevada em cerca de 4% em pH 6 e 13% a 62 °C. Além disso, a xilanase mutada imobilizada reteve uma proporção maior de sua atividade do que o tipo amplo em termoestabilidade. Essas propriedades sugerem que a imobilização da xilanase mutada tem potencial para aplicação na indústria de biobranqueamento.

Modulating the production of xylanase by Bacillus pumilus BS131 through optimization using waste fiber sludge

Abstract In recent days, cheapest alternative carbon source for fermentation purpose is desirable to minimize production cost. Xylanases have become attractive enzymes as their potential in bio-bleaching of pulp and paper industry. The objective of the present study was to identify the potential ability on the xylanase production by locally isolated Bacillus pumilus BS131 by using waste fiber sludge and wheat bran media under submerged fermentation. Culture growth conditions were optimized to obtain significant amount of xylanase. Maximum xylanase production was recorded after 72 hours of incubation at 30 °C and 7 pH with 4.0% substrate concentration. In the nutshell, the production of xylanase using inexpensive waste fiber sludge and wheat-bran as an alternative in place of expensive xylan substrate was more cost effective and environment friendly.

Resumo Nos últimos dias, a fonte alternativa de carbono mais barata para fins de fermentação é desejável para minimizar o custo de produção. As xilanases têm se tornado enzimas atraentes como seu potencial no biobranqueamento da indústria de papel e celulose. O objetivo do presente estudo foi identificar a capacidade potencial na produção de xilanase por Bacillus pumilus BS131 isolado localmente usando lodo de fibra residual e farelo de trigo em meio de fermentação submersa. As condições de crescimento da cultura foram otimizadas para obter uma quantidade significativa de xilanase. A produção máxima de xilanase foi registrada após 72 horas de incubação a 30 °C e pH 7 com concentração de substrato de 4,0%. Resumindo, a produção de xilanase usando lodo de fibra residual de baixo custo e farelo de trigo como uma alternativa no lugar do substrato de xilano caro foi mais econômica e ecológica.

Screening,identification,enzymatic characteristics and optimization of fermentation condition of a high alkaline xylanase-producing strain / 中国生物制品学杂志

@#ObjectiveTo screen a high alkaline xylanase-producing strain,subject to molecular identification and characterization of enzymatic property,and optimize its fermentation condition.MethodsThe farmland soil samples from Shanxi,Henan and Zhejiang Provinces were collected aseptically,from which the high xylanase-producing strains were screened by enrichment culture,isolation and identification of 16S r DNA,and determined for enzymatic properties.The carbon source(xylan,lactose,soluble starch,sucrose,bran and glucose),nitrogen source[ammonium oxalate,peptone,yeast powder,(NH_4)_2SO_4,NH_4Cl,Na NO_3,urea,beef extract,bean powder,KNO_3,(NH_4)_2HPO_4or random mixture of two of peptone,yeast powder,beef extract,(NH_4)_2HPO_4and bean powder],metal ion[CuCl_2,MgCl_2,ZnCl_2,Al_2(SO_4)_3,CoCl_2,MnCl_2,AgNO_3,NaCl,CaCl_2,FeCl_2,BaCl_2 and FeCl_3],pH value(3.0～10.0)in medium and the fermentation temperature(25,28,30,33,35,37 and 40℃)were optimized by single factor test,while the contents of components[xylan,(NH_4)_2HPO_4and bean powder]by response surface method.The high alkaline xylanase-producing strain was fermented by using the optimized medium under the optimized condition,and determined for xylanase activity,and the result was compared with that predicated in model.ResultsA high alkaline xylanase-producing strain named as SX6-18 was screened and identified as Paemibacillus based on 16s rDNA sequence analysis.The xylanase produced by the strain maintained more than 70%of relative activity at temperatures of 25～55℃and pH 6.0～10.0.Mg(2+)promoted while Fe(2+)promoted while Fe(3+),Mn(3+),Mn(2+)and Cu(2+)and Cu(2+)inhibited the activity of xylanase.The optimal carbon source,nitrogen source and metal ion of the medium were xylan,bean powder+(NH_4)_2HPO_4 and Fe(2+)inhibited the activity of xylanase.The optimal carbon source,nitrogen source and metal ion of the medium were xylan,bean powder+(NH_4)_2HPO_4 and Fe(3+),while the optimal p H value and temperature for fermentation were 8.0 and 30℃，respectively.However,the optimal component contents were 15.00 g/L xylan,3.03 g/L(NH_4)_2HPO_4and 3.28 g/L bean powder.The activity of xylanase cultured in optimized medium under opti-mized condition for fermentation reached 701.08 U/m L,which was closed to the expected value(693.96 U/m L).ConclusionA high alkaline xylanase-producing strain SX6-18 was successfully screened,which maintained relatively high enzyme activity in alkaline condition.This study laid a foundation of application of xylanase in papermaking and washing fields.

Effects of site-directed mutagenesis of A79Y and T81Q at Loop structure on thermal stability of xylanase XynASP / 中国生物制品学杂志

@#ObjectiveTo identify the sites of thermal stability at Loop structure of xylanase Xyn ASP in Aspergillus saccharolyticus JOP 1030-1 and improve the thermal stability.MethodsThe amino acid sites related to thermal stability of xylanase were predicted,and beneficial mutation sites at Loop structure were screened by Fireprot online server.Singlepoint mutants Xyn(A79Y),Xyn(A79Y),Xyn(T81Q)and double-point mutants Xyn(T81Q)and double-point mutants Xyn(LQ)(A79Y/T81Q)were constructed by site-directed mutagenesis.The recombinant mutant plasmid was transformed to E.coli BL21(DE3)and induced by IPTG.The recombinant xylanase was purified by Ni-NTA protein purification kit,and determined for the optimum temperature,thermal stability,optimum p H and p H stability.ResultsBeneficial mutation sites A79Y and T81Q were screened at Loop structure,and the purified protein samples showed high purity.Compared with that of wild type Xyn ASP,the optimum temperature of mutants Xyn(LQ)(A79Y/T81Q)were constructed by site-directed mutagenesis.The recombinant mutant plasmid was transformed to E.coli BL21(DE3)and induced by IPTG.The recombinant xylanase was purified by Ni-NTA protein purification kit,and determined for the optimum temperature,thermal stability,optimum p H and p H stability.ResultsBeneficial mutation sites A79Y and T81Q were screened at Loop structure,and the purified protein samples showed high purity.Compared with that of wild type Xyn ASP,the optimum temperature of mutants Xyn(A79Y),Xyn(A79Y),Xyn(T81Q)and Xyn(T81Q)and Xyn(LQ)increased by 10℃，5℃and 5℃，respectively.After heat treatment at 40℃for 30 min,wild type Xyn ASP retained only 30.2%residual relative activity,while the mutant Xyn(LQ)increased by 10℃，5℃and 5℃，respectively.After heat treatment at 40℃for 30 min,wild type Xyn ASP retained only 30.2%residual relative activity,while the mutant Xyn(T81Q)retained 37.7%,Xyn(T81Q)retained 37.7%,Xyn(A79Y)and Xyn(A79Y)and Xyn(LQ )still retained more than 65%.After heat treatment at 60℃for 30 min,the enzyme activity of wild type was 7.1%,while that of Xyn(LQ )still retained more than 65%.After heat treatment at 60℃for 30 min,the enzyme activity of wild type was 7.1%,while that of Xyn(LQ)remained 26.4%.The thermal stability of mutants was improved compared with that of wild type Xyn ASP.The optimum p H of Xyn(LQ)remained 26.4%.The thermal stability of mutants was improved compared with that of wild type Xyn ASP.The optimum p H of Xyn(A79Y),Xyn(A79Y),Xyn(T81Q)and Xyn(T81Q)and Xyn(LQ)was 5.0,which was lower than that of wild type Xyn ASP(p H 6.0).The p H stability of Xyn(LQ)was 5.0,which was lower than that of wild type Xyn ASP(p H 6.0).The p H stability of Xyn(A79Y),Xyn(A79Y),Xyn(T81Q)and Xyn(T81Q)and Xyn(LQ)at pH 3.0～8.0 showed no significant change compared with the wild type.ConclusionSite-directed mutagenesis(A79Y and T81Q)was carried out at Loop structure,and xylanase mutants with obviously improved thermal stability were obtained,which laid a foundation of the later research on the structure,function and relationship of the enzyme and its industrial application.

Effects of supplemental xylanase on in vitro disappearance of dry matter in feed ingredients for swine / Efectos de la suplementación con xilanasa sobre la desaparición in vitro de materia seca en ingredientes de piensos para cerdos / Efeitos da xilanase suplementar no desaparecimento in vitro de matéria seca em ingredientes de rações para suínos

Abstract Background: Alternative feed ingredients are widely used in swine diets to lower feed costs, but these ingredients contain a large quantity of non-starch polysaccharides. Supplemental xylanase is known to break down non-starch polysaccharides. However, the effects of exogenous xylanase from Bacillus subtilis on various feed ingredients have rarely been compared. Objective: To evaluate the effects of supplemental xylanase on in vitro disappearance of dry matter (DM) in various feed ingredients for pigs. Methods: Nine feed ingredients were used to measure in vitro ileal disappearance and in vitro total tract disappearance of DM. Each ground ingredient was supplemented with either supplemental xylanase (9,000 U/g) or cornstarch at 1.0%. Results: Supplemental xylanase increased in vitro ileal disappearance of DM in wheat, barley, wheat flour, and wheat bran (p<0.05). The in vitro total tract disappearance of DM for barley and wheat bran increased with xylanase addition (p<0.05). Conclusion: Exogenous xylanase could increase in vitro ileal DM disappearance in barley, wheat, wheat flour, and wheat bran, but did not affect in vitro total tract DM disappearance in wheat and wheat flour.

Resumen Antecedentes: Los ingredientes alternativos se utilizan ampliamente en las dietas porcinas para reducir los costos del pienso, pero estos ingredientes contienen una gran cantidad de polisacáridos no-amiláceos. Se sabe que la xilanasa suplementaria descompone los polisacáridos diferentes al almidón. Sin embargo, rara vez se han comparado los efectos de la xilanasa exógena de Bacillus subtilis en algunos ingredientes del alimento. Objetivo: Evaluar los efectos de la xilanasa suplementaria sobre la desaparición in vitro de la materia seca (MS) en varios ingredientes alimentarios para cerdos. Métodos: Se utilizaron nueve ingredientes del alimento para medir la desaparición ileal in vitro y la desaparición del tracto total in vitro de MS. Cada ingrediente molido se complementó con xilanasa suplementaria (9,000 U/g) o almidón de maíz al 1,0%. Resultados: La xilanasa suplementaria aumentó la desaparición ileal in vitro de MS en trigo, cebada, harina de trigo y salvado de trigo (p<0,05). La desaparición de tracto total de MS in vitro para la cebada y el salvado de trigo aumentó con la adición de xilanasa (p<0,05). Conclusión: La xilanasa exógena podría aumentar la desaparición de la MS ileal in vitro en cebada, trigo, harina de trigo y salvado de trigo, pero no afecta la desaparición de la MS en tracto total in vitro del trigo y la harina de trigo.

Resumo Antecedentes: Ingredientes alternativos para rações são amplamente usados em dietas para suínos para reduzir os custos da alimentação, mas esses ingredientes contêm uma grande quantidade de polissacarídeos não amiláceos. A xilanase suplementar é conhecida por quebrar polissacarídeos não amiláceos. No entanto, os efeitos da xilanase exógena de Bacillus subtilis em vários ingredientes da ração raramente foram comparados. Objetivo: Avaliar os efeitos da xilanase suplementar no desaparecimento in vitro da matéria seca (MS) em vários ingredientesde rações para suínos. Métodos: Nove ingredientes da ração foram usados para medir o desaparecimento ileal in vitro e o desaparecimento de MS in vitro do trato total. Cada ingrediente moído foi suplementado com xilanase suplementar (9.000 U/g) ou amido de milhoa 1,0%. Resultados: A xilanase suplementar aumentou o desaparecimento ileal in vitro de MS em trigo, cevada, farinha de trigo e farelo de trigo (p<0,05). O desaparecimento in vitro de MS do trato total para cevada e farelo de trigo aumentou com a adição de xilanase (p<0,05). Conclusão: A xilanase exógena pode aumentar o desaparecimento in vitro da MS ileal em cevada, trigo, farinha de trigo e farelo de trigo, mas não afetou o desaparecimento in vitro do trato total da MS no trigo e na farinha de trigo.

Biomass of unconventional plants from Brazilian semiarid as substrate for hydrolytic enzymes production by Aspergillus niger under solid and submerged fermentation

Aspergillus niger KIJH was grown in solid and submerged fermentation using leaves and roots (with and without bark) of plants typically from Brazilian semiarid as substrate to produce a multienzymatic extract, which was characterised for its potential biotechnological applications. Solid-state fermentation (SSF) was applied to select the most promising plants biomass as induction substrates for the production of hydrolytic enzymes by fungus. The best biomasses were used as substrate in submerged fermentation (SmF) assays at two scales. Samples of up scale fermented culture were partially purified by ultrafiltration and activity and pH and temperature stability of CMCase and xylanase were evaluated. A. niger KIJH produced hydrolytic enzymes under SSF containing unconventional plants biomass from Brazilian semiarid. In SmF conditions, maximum CMCase (0.264 U mL-1) and xylanase (1.163 U mL-1) activities were induced by Jacaratia corumbensis. Scaling up the SmF to 500 mL of medium was able to maintain constant the production of CMCase (0.346 U mL-1) and xylanase (1.273 U mL-1) on the fermented culture. Ultrafiltered and concentrated extract presented CMCase activities practically constant in all temperature ranges (30-80°C) and pH (3.0-9.0), while xylanase optimum activity temperature was 50°C and pH in the range of 3.0 to 5.0. CMCase activity remained stable for 24 hours at 50°C

Biomass of unconventional plants from Brazilian semiarid as substrate for hydrolytic enzymes production by Aspergillus niger under solid and submerged fermentation

Aspergillus niger KIJH was grown in solid and submerged fermentation using leaves and roots (with and without bark) of plants typically from Brazilian semiarid as substrate to produce a multienzymatic extract, which was characterised for its potential biotechnological applications. Solid-state fermentation (SSF) was applied to select the most promising plants biomass as induction substrates for the production of hydrolytic enzymes by fungus. The best biomasses were used as substrate in submerged fermentation (SmF) assays at two scales. Samples of up scale fermented culture were partially purified by ultrafiltration and activity and pH and temperature stability of CMCase and xylanase were evaluated. A. niger KIJH produced hydrolytic enzymes under SSF containing unconventional plants biomass from Brazilian semiarid. In SmF conditions, maximum CMCase (0.264 U mL-1) and xylanase (1.163 U mL-1) activities were induced by Jacaratia corumbensis. Scaling up the SmF to 500 mL of medium was able to maintain constant the production of CMCase (0.346 U mL-1) and xylanase (1.273 U mL-1) on the fermented culture. Ultrafiltered and concentrated extract presented CMCase activities practically constant in all temperature ranges (30-80°C) and pH (3.0-9.0), while xylanase optimum activity temperature was 50°C and pH in the range of 3.0 to 5.0. CMCase activity remained stable for 24 hours at 50°C a

Unveiling Xylanolytic Enzymes Production of Talaromyces wortmannii DR49 on Industrial Agro Wastes

Abstract Xylan degradation is an important step in different industries, such as in biorefinery for biomass hydrolysis. Talaromyces wortmannii is a known fungus due to second metabolite production but only few works showed the xylanolytic potential of this fungus. In this way, the aim of this study was to evaluate the production of xylanolytic enzymes from T. wortmannii DR49 on industrial agro wastes. Cultivation in shake flask showed highest xylanase titration (10.3 U/mL; 9.5 U/mL) for wheat bran (WB) and hydrothermal pretreated sugar cane bagasse (HB); in β-xylosidase production WB and xylose were the best carbon sources (0.57 U/mL; 0.34 U/mL) respectively. STR cultivation revealed that 29°C and pH 6.0 were the best conditions for xylanase (14.5 U/mL) and β-xylosidase (1.7 U/mL) production. T. wortmannii DR49 showed to be a potential candidate for xylanolytic enzymes production using agro wastes in bioreactors, which has never been previously reported in this fungus.

Production and optimization of enzyme xylanase by Aspergillus flavus using agricultural waste residues

This study aims to isolate xylanase-producing fungi, i.e., Aspergillus flavus, isolated from the litter of theOrchha forest, and efforts are made to culture fungi on cheaper agricultural substrates, i.e., wheat bran and corncobs. After isolation, the xylanolytic activity was tested on the malt extract agar culture medium. Optimizationof growth parameters were also carried out with wheat bran and corn cobs. It was found that maximum xylanasewas produced on the sixthand eighth days of incubation, with pH of 6.0 and 7.0, and substrates amount for corncobs and wheat bran of 16 and 18 mg/ml, respectively. Temperature of 30°C, peptone concentration of 0.5 mg/ml, and yeast extract concentration of 0.75 mg/ml were the same for both substrates. The enzyme producedthrough optimized conditions was assayed for its maximum activity. It was found that the enzyme showedmaximum activity at 15 and 30 minutes of incubation, and at 60° and 55°C temperature for corn cobs andwheat bran, respectively. The pH and substrate (oat spelt xylan) concentrations were optimized at 5.5 and 15mg/ml, respectively. In. this study, the production of xylanase with the use of cheaper agricultural waste, suchas wheat bran and corn cobs, will not only reduce the cost of production but will also help in their eradicationfrom the environment.

Albizia lebbeck fruit pods as a substrate for the production of lignocellulolytic enzymes by Aspergillus niger

Aspergillus niger isolated from the soil was investigated for its capability to produce various lignocellulolyticenzymes, such as LiP, endoglucanase, FPase, xylanase by solid-state fermentation, using Albizia lebbeck fruitpods as a substrate. The chemical composition of the fruit pods was studied, and the production pattern of theenzymes was examined by growing the fungi for 25 days. The LiP activity was low, whereas a good productionof endoglucanase, FPase, and xylanase enzymes was noted. A dye decolorization capacity of the A. niger wasalso studied with Congo red. Therefore, A. lebbeck fruit pods which are considered as waste and burnt off canbe utilized for the production of holocellulolytic enzymes using A. niger.

Isolation of thermotolerant Bacillus subtilis DCH4 from Chancos hot spring (Carhuaz, Peru) with potential to degrade lignocellulosic agriculture wastes / Aislamiento de Bacillus subtilis DCH4 termotolerante de la fuente termal Chancos (Carhuaz, Perú) con potencial para degradar residuos lignocelulósicos agrícolas

Abstract It was isolated bacteria strains from three different types of samples: fresh water, in situ baits and ex situ enrichment. Serial dilutions were prepared and culture was carried at 50 °C using a Basal-Saline medium. Isolated strains were screened for endoglucanase and xylanase activities with qualitative (Congo Red) and quantitative (DNS) methods. Molecular 16S rDNA sequencing analysis was performed for taxonomic identification. It was isolated 31 strains of which 14 showed hydrolytic activities and belonged to Bacillus subtilis and Bacillus licheniformis species. Moreover, the strain B. subtilis DCH4 showed the highest endoglucanase activity at 45°C and pH 5, and xylanase activity at 55°C and pH 6. Then, DCH4 was cultivated by submerged fermentation with two different media supplemented with sugar cane bagasse, wheat straw, or quinoa stalk to evaluate its saccharification capability. Likewise, it was screening its xylanase and cellulase genes employing specific primers; the amplicons obtained were sequenced, and analyzed. It was found that, enzymatic extracts of DCH4 prepared with cane bagasse or quinoa stalk media achieved the highest endoglucanase and xylanase activities. According to molecular analysis of genes involved in the hydrolytic process, the endoglucanase and xylanase activities exhibited by DCH4 could be attributed to a bifunctional cellulase conformed by endo-beta-1,4-glucanase (GH5) joined to cellulose binding domain 3 (CBM3), and an endo-1,4-beta-xylanase (GH11), respectively. Further transcriptomic experiments would be considered to accomplish optimization strategies for biofuel production from lignocellulosic biomass.

Resumen Se aislaron cepas de bacterias provenientes de tres tipos de muestras: agua fresca, cebos enriquecidos in situ y ex situ. Se prepararon diluciones seriadas y el cultivo fue a 50 °C usando un medio Salino-Basal. Las cepas aisladas fueron tamizadas para las actividades endoglucanasa y xilanasa con métodos cualitativos (Rojo Congo) y cuantitativos (DNS). Se usó el análisis molecular 16S rDNA para la identificación taxonómica. Se aislaron 31 cepas, de las cuales 14 mostraron actividades hidrolíticas y pertenecían a Bacillus subtilis y Bacillus licheniformis. Además, B. subtilis DCH4 mostró la mayor actividad endoglucanasa a 45 °C y pH 5, y xilanasa a 55 °C y pH 6. Entonces, DCH4 se cultivó por fermentación sumergida con dos medios diferentes suplementado con bagazo de caña de azúcar, paja de trigo o tallo de quinua para evaluar su capacidad de sacarificación. También, se exploraron los genes de xilanasa y celulasa mediante cebadores específicos; los amplicones obtenidos fueron secuenciados y analizados. Se encontró que los extractos enzimáticos de DCH4 preparados con bagazo de caña o tallos de quinua mostraron las actividades endoglucanasa y xilanasa más elevadas. De acuerdo a los análisis moleculares de los genes involucrados en el proceso hidrolítico, las actividades de endoglunacasa y xilanasa exhibidas por DCH4 podrían atribuirse a una celulasa bifuncional conformada por una endo-beta-1,4-glucanasa (GH5) unida al dominio celulosa 3 (CBM3), y una endo-1,4-beta-xilanasa (GH11), respectivamente. Posteriores experimentos transcriptómicos podrían ser considerados para lograr estrategias de optimización para la producción de biocombustibles a partir de biomasa lignocelulósica.

Newly Isolated Penicillium sp. for Cellulolytic Enzyme Production in Soybean Hull Residue

Abstract (1) Background: The aim of this study was to evaluate the production and partial characterization of xylanase and avicelase by a newly isolated Penicillium sp. in solid-state fermentation, using soybean hulls as substrate. (2) Methods: Temperature, time, number of spores, and substrate moisture on xylanase and avicelase bioproduction were evaluated, maximizing activity with 30°C, 1x106 spores/g substrate, 14 and 7 days of fermentation with 70 and 76% substrate moisture contents, for xylanase and avicelase, respectively. (3) Results: Different solvents, temperatures, and agitation in the enzymatic extraction were evaluated, obtaining higher activities, 430.77 and 26.77 U/g for xylanase and avicelase using 30 min extraction and 0.05 M citrate buffer solution (pH 4.5 ), respectively at 60°C and 175 rpm and 50°C and 125 rpm. The optimum pH and temperature for enzymatic activity determination were 5.3 and 50°C. Enzyme extract stability was evaluated, obtaining higher stability with pH between 4.5 and 5.5, higher temperature of up to 40°C. The kinetic thermal denaturation (Kd), half-life time, D-value, and Z-value were similar for both enzymes. The xylanase Ed value (89.1 kJ/mol) was slightly lower than the avicelase one (96.7 kJ/mol), indicating higher thermostability for avicelase. (4) Conclusion: In this way, the production of cellulases using alternative substrates is a way to reduce production costs, since they represent about 10% of the world demand of enzymes, with application in animal feed processing, food production and breweries, textile processing, detergent and laundry production, pulp manufacturing and the production of biofuels.

Enhancing thermostability of xylanase from rumen microbiota by molecular cyclization / 生物工程学报

The capacity for thermal tolerance is critical for industrial enzyme. In the past decade, great efforts have been made to endow wild-type enzymes with higher catalytic activity or thermostability using gene engineering and protein engineering strategies. In this study, a recently developed SpyTag/SpyCatcher system, mediated by isopeptide bond-ligation, was used to modify a rumen microbiota-derived xylanase XYN11-6 as cyclized and stable enzyme C-XYN11-6. After incubation at 60, 70 or 80 ℃ for 10 min, the residual activities of C-XYN11-6 were 81.53%, 73.98% or 64.41%, which were 1.48, 2.92 or 3.98-fold of linear enzyme L-XYN11-6, respectively. After exposure to 60-90°C for 10 min, the C-XYN11-6 remained as soluble in suspension, while L-XYN11-6 showed severely aggregation. Intrinsic and 8-anilino-1-naphthalenesulfonic acid (ANS)-binding fluorescence analysis revealed that C-XYN11-6 was more capable of maintaining its conformation during heat challenge, compared with L-XYN11-6. Interestingly, molecular cyclization also conferred C-XYN11-6 with improved resilience to 0.1-50 mmol/L Ca²⁺ or 0.1 mmol/L Cu²⁺ treatment. In summary, we generated a thermal- and ion-stable cyclized enzyme using SpyTag/SpyCatcher system, which will be of particular interest in engineering of enzymes for industrial application.

Progress in improving the properties of dissolving pulp by enzymes / 生物工程学报

Dissolving pulp consists of high purity cellulose and is widely used to as raw materials for the production of regenerated cellulose fiber, cellulose ester and cellulose ether. The characteristic of dissolving pulp affects greatly the production and processing performance of subsequent products. The α-cellulose content, hemicellulose content, pulp viscosity, ash, transition metal ion content, fiber morphology, molecular weight distribution of cellulose and the reactivity are the important properties. Because of its green, mild and high efficiency, the application of enzymes in improving the properties of dissolving pulp has a promising application prospect and has been researched significantly. In this review, the main properties of dissolving pulp are presented first, followed by a recommendation of the enzymes to improve these properties. The application and current research of cellulase and xylanase in improving the properties of dissolving pulp are emphasized. The main problems and the future research areas in improving the properties of dissolving pulp by enzymes are revealed. Finally, the technology prospects in this field are proposed.

Cellulolytic, amylolytic and xylanolytic potential of thermophilic isolates of Surajkund hot spring

A total of 41 isolates were obtained from various samples (soil, mud, and water) of Surajkund hot spring, Jharkhand, atthree different isolation temperatures of 50C, 60C, and 70C. However, our interest was in the thermophilic strains thatwere isolated at 60C and 70C. Four isolates at 70C (BITSNS038, BITSNS039, BITSNS040, BITSNS041) are theproducers of thermozymes, namely amylase, xylanase, and cellulase, respectively. The highlights of the present study alsoshowed that three out of four isolates demonstrated all three enzymatic activities, i.e. amylolytic, xylanolytic and cellulolytic on agar plate assay conditions at 70C. One of the isolates, BITSNS038, was further chosen for phenotypiccharacterization as well as 16S rRNA gene sequencing and was affiliated to Geobacillus icigianus. The presence ofGeobacillus icigianus was reported first time from hot spring, Surajkund, which showed amylolytic index of 1.58,xylanolytic index of 1.5 and cellulolytic index of 2.3 based on plate assay, and amylase activity of 0.81 U/mL, xylanaseactivity of 0.72 U/mL and very less cellulase activity of 0.15 U/mL after 24 h of growth in submerged conditions. Oneisolate at 60C BITSNS024 was found to exhibit maximum amylase activity with an enzymatic index value of 3.5 and wasidentified as Anoxybacillus gonensis.

Complexos enzimáticos sobre a energia metabolizável e digestibilidade dos nutrientes do milheto para frangos de corte / Enzyme complexes on metabolizable energy and nutrient digestibility of millet for broiler chickens

Objetivou-se avaliar o efeito de complexos enzimáticos sobre a energia metabolizável e o coeficiente de digestibilidade de nutrientes do milheto para frangos de corte. Quinhentos e setenta e seis frangos machos foram distribuídos em 36 gaiolas, com três tratamentos: T1 - composição de milheto sem complexo enzimático; T2 - composição de milheto com complexo enzimático (CES) e T3 - composição de milheto com complexo enzimático (CEV). Os tratamentos foram definidos com base em seis dietas (três dietas referências e três dietas testes). As dietas testes foram obtidas pela substituição de 40% da dieta referência por milheto inteiro, e a adição de enzimas consistiu de dois complexos enzimáticos: CES, constituído pelas enzimas fitase, protease, xilanase, ß-glucanase, celulase, amilase e pectinase; e CEV. constituído pelas enzimas protease, celulase e amilase. Na fase de 11 a 20 dias, a suplementação com o CEV reduziu os valores de EMA, EMAn e CDPB. A suplementação com CES melhorou o CDPB, e não houve efeito significativo para CDMS e CDEB. Na fase de 21 a 30 dias, houve menor aproveitamento da energia e dos nutrientes com as suplementações CES e CEV. Na fase de 31 a 40 dias, as suplementações reduziram os valores de EMA, EMAn, e o complexo CEV foi efetivo em aumentar o valor de CDPB. A inclusão dos complexos enzimáticos CES (fitase, protease, xilanase, ß-glucanase, celulase, amilase e pectinase) e CEV (protease, celulase e amilase) não favoreceu a utilização da energia do milheto, no entanto melhorou o coeficiente de digestibilidade da proteína do milheto nos períodos de 11 a 20 e de 31 a 40 dias de idade.(AU)

The objective of this study was to evaluate the effect of enzymatic complexes on metabolizable energy and nutrient digestibility coefficient of millet for broilers chickens. 576 male chickens, were distributed in 36 cages with three treatments: T1 - millet composition without enzymatic complex; T2 - millet composition with enzymatic complex (ECS); and T3 - millet composition with enzymatic complex (ECV). The treatments were defined from six diets (3 reference diets and 3 test diets). The test diets were obtained from the substitution of 40% for reference diet by whole millet, and the enzyme addition consisted of two enzymatic complex, ECS constituted by phytase, protease, xylanase, ß-glucanase, cellulase, amylase and pectinase enzymes, and ECV constituted by protease, cellulase and amylase enzymes. In the 11 to 20 days phase, a supplementation with the ECV reduced the AME, AMEn and CDPB values, a ECS supplementation improved the CDPB, and there was no significant effect for CDMS and CDEB. In the 21 to 30 days phase, there were less profit of the energy and nutrients with ECS and ECV supplements. In the 31 to 40 days phase as supplements reduced the values of AME, AMEn, and the ECV complex was effective in increasing the value of CDPB. The inclusion of ECS enzymatic complexes, (phytase, protease, xylanase, ß-glucanase, cellulase, amylase and pectinase) and ECV (protease, cellulase and amylase), did not favor millet's energy utilization, however, favored the millet's protein digestibility coefficient on 11 to 20 and 31 to 40 periods.(AU)