Produção de xilanases por uma cepa selvagem de Aspergillus nidulans (original) (raw)
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Xylanase production by a wild strain of Aspergillus nidulans
A wild strain of Aspergillus nidulans isolated from soil produce cellulase-free xylanase activity when developed on submerged cultures using corn cob powder as the main substrate. Maximum xylanase production (220 U/mL) was obtained when the strain was developed in mineral medium supplemented with 3% (w/v) corn cob for 6 days. SDS-PAGE revealed the occurrence of four isoxylanases with molecular weights of 50, 43, 20 and 18 kDa. Crude xylanase was resistant to acetone precipitation with 80% of recovery. The enzyme had optimal activity at pH values between 5.0 and 6.0 and temperature between 50-55 o C. The enzyme exhibited high stability under alkaline conditions and temperature up to 55 o C. It retained 50% and 23% of its activity when heated for 1 h at 60 and 65 o C, respectively.
Aspergillus terreus, UL 4209 strain, isolated from the soil in South Africa was used to produce an extracellular cellulase-free xylanase in shake flask cultures containing oat spelt and/or birch wood xylans. Maximum xylanase activity (35 U/ml) was observed after 96 h at 35ºC and pH 6 in 1% oat spelt xylan. The xylanase was purified to homogeneity by gel filtration on Sephacryl S-200. This enzyme was found to be a single subunit protein of 22 kDa showing optimal activity at 35ºC and pH 6. The enzyme retained 95% activity at 35 - 40ºC after 4 h incubation at pH 6 and at 50ºC the half-life was 5.8 h. The apparent Km and Vmax values were 3.57 mg/ml and 55.5 mol/min per mg protein, respectively. MALDITOF and LC mass spectroscopy gave 8 peptide ions whose sequence alignments showed that the xylanase produced by this strain has homology with those of other Aspergillus strains such as A. terreus and A. versicolor. These observations showed that our strain produced a low molecular weight, acidophilic, and thermostable xylanase that may be considered for processes operated at moderate temperatures and pH such as preparation of baked cereal food, clarification of fruit juices and saccharification of agro-residues.
Agro-industrial residues, namely wheat bran (WB), red sawdust (RSD), white sawdust (WSD), corn cobs (CC), rice husk (RH) and sugarcane bagasse (SB) were used as alternatives of pure xylan for production of xylanase, carboxy methyl cellulase (CMCase) and avicellase in solid state (SSF) and submerged (SmF) fermentation using Aspergillus flavus AUMC 10331. In SSF, WB was the most utilizable yielding the highest concentration of xylanase (792.4 IU.gds-1) and CMCase (101.8 IU.gds-1) and RH was the best for avicellase (152.4 IU.gds-1). In SmF, A. flavus regularly utilized all substrates to produce the three enzymes. From the six plant residues, CC was the superior source for xylanase production (9660 IU.gds-1) and CMCase production (307 IU.gds-1), while RH registered the highest avicellase (5554 IU.gds-1). Xylanase from CC was purified using ion exchange (IR-120 EP) and Sephadex G-75 column chromatography. The purified xylanase showed activity of 32.77 IU.ml-1 .min-1 and a specific activity of 273.1 IU.mg-1 proteins. The enzyme was active over a pH range of 4.5-8.0, and its highest activity was detected at pH 7.0 and 65 ºC. Xylanase activity was stimulated by FeSO4 up to 121.97 %. Km and Vmax were 12.18 mg.ml-1 and 204.1 IU.min-1 , respectively.
World Journal of Microbiology and Biotechnology, 2005
A locally isolated strain of Aspergillus foetidus MTCC 4898 was studied for xylanase (EC 3.2.1.8) production using lignocellulosic substrates under solid state fermentation. Corncobs were found as the best substrates for high yield of xylanases with poor cellulase production. The influence of various parameters such as temperature, pH, moistening agents, moisture level, nitrogen sources and pretreatment of substrates were evaluated with respect to xylanase yield, specific activity and cellulase production. Influence of nitrogen sources on protease secretion was also examined. Maximum xylanase production (3065 U/g) was obtained on untreated corncobs moistened with modified Mandels and Strenberg medium, pH 5.0 at 1:5 moisture levels at 30°C in 4 days of cultivation. Submerged fermentation under the same conditions gave higher yield (3300 U/g) in 5 days of cultivation, but productivity was less. Ammonium sulphate fractionation yielded 3.56-fold purified xylanase with 76% recovery. Optimum pH and temperature for xylanase activity were found to be 5.3 and 50°C respectively. Kinetic parameters like K m and V max were found to be 3.58 mg/ml and 570 lmol/mg/min. Activity of the enzyme was found to be enhanced by cystiene hydrochloride, CoCl 2 , xylose and Tween 80, while significantly inhibited by Hg ++ , Cu ++ and glucose. The enzyme was found to be stable at 40°C. The half life at 50°C was 57.53 min. However thermostability was enhanced by glycerol, trehalose and Ca ++. The crude enzyme was stable during lyophilization and could be stored at less than 0°C.
2015
Fungi are well known for their ability to produce xylanase into the environment. The aim of this work was to optimize xylanase production by fungi isolated from soil diverse habitats such as manures, dead and decaying wood and soil samples. The fungus Aspergillus flavus ARC-12 was selected for the production of cellulase-free xylanase. Various agro-residues were used as carbon sources and pearl millet stover produced maximum xylanase production (1345.44 IU/gds) under solid-state fermentation. Various culture conditions were optimized by using one factor at the time and the maximum xylanase production (1699.50 IU/gds) was obtained after 48 h of incubation at a temperature of 30 °C, an initial pH of 6.0 and initial moisture content of 77.5%.
Cellulase and xylanase production by Aspergillus species
Annals of Microbiology, 2011
This study aimed to produce cellulases and xylanase from Aspergillus species in submerged cultivation using sugarcane bagasse as substrate. Nine strains of Aspergillus were obtained from the Coleção de Culturas Micoteca [University Recife Mycology (URM)] and were cultivated in medium containing raw and processed sugarcane bagasse (RSB and PSC, respectively) for 5 days at 30°C and 90 rpm. The production of enzymes was evaluated by determining the activity of total cellulase, endoglucanase, exoglucanase, cellobiase, and xylanase. RSB was the best inducer for total cellulase and xylanase whereas PSB was best for exoglucanase. The production of endoglucanase and cellobiase at higher levels was independent of the type of sugarcane bagasse used. The strains that produced cellulases and xylanase at the highest levels were A. phoenicis URM 4924 and A. aculeatus URM 4953 using RSB and A. carbonarius URM 1546 using PSB medium.
A cellulase-free xylanase from alkali-tolerant Aspergillus fischeri Fxn1
Biotechnology Letters, 1995
An alkali-tolerant fungusAsperqillus fischeri Fxn1 isolated from xylan enrichment grew in the pH range 5–10 and secreted an extracellular cellulase-free xylanase. Arabinose, lactose, maltose, cellobiose and glucose induced low levels of xylanase (1.8–9.0 IU/ml), whereas xylose, xylan and wheat bran induced higher level (34–45 IU/ml).CMcellulose and FPcellulose did not support growth. The optimum pH of xylanase was 6.0–6.5 and it was stable in a wide range of pH 5–9.5. The optimum temperature was 60°C and it was stable upto 55°C. The half-lives at 50 and 55 °C were 240 and 40 min. respectively. This enzyme released reducing sugars from pulp at pH 9.0 and 40°C.
Xylanolytic and cellulolytic potential of a soil isolate, Aspergillus fumigatus (MS16) was studied by growing it on a variety of lignocellulosics, purified cellulose and xylan supplemented media. It was noted that carboxymethyl cellulose, salicin and xylan induce the production of endoglucanase, β-glucosidase and xylanase, respectively. The study revealed that Aspergillus fumigatus (MS16) co-secretes xylanase and cellulase in the presence of xylan; the ratio of the two enzymes was influenced by the initial pH of the medium. The maximum titers of xylanase and cellulase were noted at initial pH of 5.0. Relatively higher titers of both the enzymes were obtained when the fungus was cultivated at 35 o C. Whereas, cellulase production was not detected when the fungus was cultivated at 40 o C. The volumetric productivity (Q p) of xylanase was much higher than cellulases. The organism produced 2-3 folds higher titers of xylanase when grown on lignocellulosic materials in submerged cultivation than under solid-state cultivation, suggesting a different pattern of enzyme production in presence and in absence of free water. The partial characterization of enzymes showed that xylanase from this organism has higher melting temperature than endoglucanase and β-glucosidase. The optimum temperature for activity was higher for xylanases than cellulases, whereas the optimum pH differed slightly i.e. in the range of 4.0-5.0. Enzyme preparation from this organism was loaded on some crude substrates and it showed that the enzyme preparation can be used to hydrolyze a variety of vegetable and agricultural waste materials.
African Journal of Biotechnology, 2008
Aspergillus terreus, UL 4209 strain, isolated from the soil in South Africa was used to produce an extracellular cellulase-free xylanase in shake flask cultures containing oat spelt and/or birchwood xylans. Maximum xylanase activity (35 U/ml) was observed after 96 h at 35ºC and pH 6 in 1% oat spelt xylan. The xylanase was purified to homogeneity by gel filtration on Sephacryl S-200. This enzyme was found to be a single subunit protein of 22 kDa showing optimal activity at 35ºC and pH 6. The enzyme retained 95% activity at 35-40ºC after 4 h incubation at pH 6 and at 50ºC the half-life was 5.8 h. The apparent K m and V max values were 3.57 mg/ml and 55.5 µmol/min per mg protein, respectively. MALDI-TOF and LC mass spectroscopy gave 8 peptide ions whose sequence alignments showed that the xylanase produced by this strain has homology with those of other Aspergillus strains such as A. terreus and A. versicolor. These observations showed that our strain produced a low molecular weight, acidophilic, and thermostable xylanase that may be considered for processes operated at moderate temperatures and pH such as preparation of baked cereal food, clarification of fruit juices and saccharification of agro-residues.
Xylan is a principal structural component in plant cell walls and could interfere in many industries and biological activities. Bio-synthesis of the xylan-degrading enzyme xylanase by fungal isolates was investigated, using agricultural wastes as substrates for production. Aspergillus fumigatus was shown to be the superior isolate to produce xylanase, and wheat bran proved to be the most appropriate substrate for enzyme production and activity. Media composition and growth conditions were optimized for the production of fungal xylanase using Plackett-Burman multifactorial mixture design and Box-Behnken experimental design. Characterization of the purified xylanase revealed that the most favorable conditions for the maximum enzyme activity (57 IU/mL) were temperature of 55-60°C with thermal stability for 30 min and pH of 9 with enzyme stability for 20 min. Application of xylanase for biodegradation of sugarcane pulp fibers was examined using scanning electron micrographs, which exhibited a vigorous alteration in the topography of fiber surface and structure with a prolonged enzymatic treatment period.