Rhizopus microsporus var. rhizopodiformis: a thermotolerant fungus with potential for production of thermostable amylases

The effect of several nutritional and environmental parameters on growth and amylase production from Rhizopus microsporus var. rhizopodiformis was analysed. This fungus was isolated from soil of the Brazilian “cerrado” and produced high levels of amylolytic activity at 45°C in liquid medium supplemented with starch, sugar cane bagasse, oat meal or cassava flour. Glucose in the culture medium drastically repressed the amylolytic activity. The products of hydrolysis were analysed by thin layer chromatography, and glucose was detected as the main component. The amylolytic activity hydrolysed several substrates, such as amylopectin, amylase, glycogen, pullulan, starch, and maltose. Glucose was always the main end product detected by high-pressure liquid chromatography analysis. These results indicated that the amylolytic activity studied is a glucoamylase, but there were also low levels of α-amylase. As compared to other fungi, R. microsporus var. rhizopodiformis can be considered an efficient producer of thermostable amylases, using raw residues of low cost as substrates. This information is of technological value, considering the importance of amylases for industrial hydrolysis

Production of xylanase and β-xylosidase from autohydrolysis liquor of corncob using two fungal strains

Agroindustrial residues are materials often rich in cellulose and hemicellulose. The use of these substrates for the microbial production of enzymes of industrial interest is mainly due to their high availability associated with their low cost. In this work, corncob (CCs) particles decomposed to soluble compounds (liquor) were incorporated in the microbial growth medium through autohydrolysis, as a strategy to increase and undervalue xylanase and b-xylosidase production by Aspergillus terricola and Aspergillus ochraceus. The CCs autohydrolysis liquor produced at 200 C for 5, 15, 30 or 50 min was used as the sole carbon source or associated with untreated CC. The best condition for enzyme synthesis was observed with CCs submitted to 30 min of autohydrolysis. The enzymatic production with untreated CCs plus CC liquor was higher than with birchwood xylan for both microorganisms. A. terricola produced 750 total U of xylanase (144 h cultivation) and 30 total U of b-xylosidase (96–168 h) with 0.75% untreated CCs and 6% CCs liquor, against 650 total U of xylanase and 2 total U of b-xylosidase in xylan; A. ochraceus produced 605 total U of xylanase and 56 total U of b-xylosidase (168 h cultivation) with 1% untreated CCs and 10% CCs liquor against 400 total U of xylanase and 38 total U of b-xylosidase in xylan. These results indicate that the treatment of agroindustrial wastes through autohydrolysis can be a viable strategy in the production of high levels of xylanolytic enzymes.This work was supported by State of Sao Paulo Research Foundation (FAPESP/Brazil), National Counsel of Technological and Scientific Development (CNPq/Brazil), National System for Research on Biodiversity (SISBIOTA-Brazil, CNPq 563260/2010-6/FAPESP 2010/52322-3), and Portuguese Foundation for Science and Technology (FCT/Portugal). Hector A. Ruiz thanks to Mexican Science and Technology Council (CONACYT, Mexico) for PhD fellowship support (CONACYT grant number: 213592/308679)

Xylanase and β‐xylosidase production: alternatives for the autohydrolysis liquor application

Fundação para a Ciência e a Tecnologia (FCT)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Production of xylanolytic enzymes by Aspergillus terricola in stirred tank and airlift tower loop bioreactors

Fungi producing high xylanase levels have attracted considerable attention because of their potential industrial applications. Batch cultivations of Aspergillus terricola fungus were evaluated in stirred tank and airlift bioreactors, by using wheat bran particles suspended in the cultivation medium as substrate for xylanase and b-xylosidase production. In the stirred tank bioreactor, in physical conditions of 30°C, 300 rpm, and aeration of 1 vvm (1 l/min), with direct inoculation of fungal spores, 7,475 U/l xylanase was obtained after 36 h of operation, remaining constant after 24 h. In the absence of air injection in the stirred tank reactor, limited xylanase production was observed (final concentration 740 U/l). When the fermentation process was realized in the airlift bioreactor, xylanase production was higher than that observed in the stirred tank bioreactor, being 9,265 U/l at 0.07 vvm (0.4 l/min) and 12,845 U/l at 0.17 vvm (1 l/min) aeration rate.Fundação para a Ciência e a Tecnologia (FCT)CNPq (Brasil)FAPESP (Brasil

Production of xylanases by Aspergillus terricola using wheat bran as carbon source : comparative studies between different bioreactors and influence of aeration and inoculum conditions

Fundação de Amparo à Pesquisa do Estado de São Paulo (Biota-FAPESP/Brazil )Fundação para a Ciência e a Tecnologia (FCT

A new strategy for xylanase production using wheat straw autohydrolysis liquor as substrate

Agro-industrial residues are lignocellulosic materials with a high content of cellulose, hemicellulose and lignin. If such residues can be produced in bioprocesses (e.g. xylanase production) there is an attractive possibility of their integral use in biotechnological processes. In general, xylanase biosynthesis is induced by its substrate – xylan, but the high xylan content of some wastes such as corn cobs and wheat bran makes them an accessible and cheap source of inducers. Another alternative to improve the xylanase production, which is the main goal of this work, is the treatment of lignocellulosic materials in autohydrolysis processes which, under optimized conditions, lead to the solubilization of hemicelluloses (liquid phase, liquor) that may be favorable to xylanase production. The inclusion of these components in the nutrient medium composition can be a strategy to optimize the microbial xylanase biosynthesis. The best conditions for xylanase production were observed when the microorganism was cultivated in birchwood xylan for 6 days; however, satisfactory results were obtained using a combination of 1% wheat bran with 2% or 10% autohydrolysis liquor, for 5 days fermentation, once the xylanase production was around 86-87% of production with xylan. Besides, the obtained production with 100% wheat straw autohydrolysis liquor was also interesting, once after 7 days of cultivation, the xylanase production was higher than the ones obtained with wheat bran or by the combination of wheat bran and liquor.Biota FAPESP (Brazil)Fundação para a Ciência e a Tecnologia (FCT

Application of wheat straw autohydrolysis liquor to xylanase and β-xylosidase large-scale production in a stirred tank bioreactor

Research into microbial xylanases production has increased due to its several applications. In this context, studies that make this practice feasible are important. Wheat bran is an inexpensive byproduct, which contains around 28% hemicellulose; however, the wheat bran particles suspended in the cultivation medium have to be decomposed to soluble compounds to be used by the fungi and the treatment of lignocellulosic materials in autohydrolysis processes makes this easier. The inclusion of these treated materials in the nutrient media can be a strategy to increase and undervalue xylanase production. The best conditions for xylanase and β-xylosidase production were observed when A. ochraceus was cultivated with 1% wheat bran added 10% wheat straw autohydrolysis liquor as carbon source, this substrate was more favorable when compared with xylan, wheat bran and wheat straw autohydrolysis liquor used separately. The application of this substrate in a stirred tank bioreactor shows the need for improvements of the fermentation process

Xylanase and β-xylosidase production by Aspergillus ochraceus: new perspectives for the application of wheat straw autohydrolysis liquor

The xylanase biosynthesis is induced by its substrate—xylan. The high xylan content in some wastes such as wheat residues (wheat bran and wheat straw) makes them accessible and cheap sources of inducers to be mainly applied in great volumes of fermentation, such as those of industrial bioreactors. Thus, in this work, the main proposal was incorporated in the nutrient medium wheat straw particles decomposed to soluble compounds (liquor) through treatment of lignocellulosic materials in autohydrolysis process, as a strategy to increase and undervalue xylanase production by Aspergillus ochraceus. The wheat straw autohydrolysis liquor produced in several conditions was used as a sole carbon source or with wheat bran. The best conditions for xylanase and β-xylosidase production were observed when A. ochraceus was cultivated with 1% wheat bran added of 10% wheat straw liquor (produced after 15 min of hydrothermal treatment) as carbon source. This substrate was more favorable when compared with xylan, wheat bran, and wheat straw autohydrolysis liquor used separately. The application of this substrate mixture in a stirred tank bioreactor indicated the possibility of scaling up the process to commercial production.This work was supported by Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP/Brazil), Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq/Brazil), National System for Research on Biodiversity (SISBIOTA-Brazil, CNPq 563260/2010-6/FAPESP no. 2010/52322-3), and Fundacao para a Ciencia e a Tecnologia (FCT/Portugal)

A novel xylan degrading β-D-xylosidase: purification and biochemical characterization

Aspergillus ochraceus, a thermotolerant fungus isolated in Brazil from decomposing materials, produced an extracellular b-xylosidase that was purified using DEAE-cellulose ion exchange chromatography, Sephadex G-100 and Biogel P-60 gel filtration. b-xylosidase is a glycoprotein (39 % carbohydrate content) and has a molecular mass of 137 kDa by SDS-PAGE, with optimal temperature and pH at 70 C and 3.0–5.5, respectively.b-xylosidase was stable in acidic pH (3.0–6.0) and 70 C for 1 h. The enzyme was activated by 5 mM MnCl2 (28 %)and MgCl2 (20 %) salts. The b-xylosidase produced by A. ochraceus preferentially hydrolyzed p-nitrophenyl-b- D-xylopyranoside, exhibiting apparent Km and Vmax values of 0.66 mM and 39 U (mg protein)-1 respectively, and to a lesser extent p-nitrophenyl-b-D-glucopyranoside. The enzyme was able to hydrolyze xylan from different sources,suggesting a novel b-D-xylosidase that degrades xylan. HPLC analysis revealed xylans of different compositions which allowed explaining the differences in specificity observed by b-xylosidase. TLC confirmed the capacity.This work was supported by the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP), and the Conselho de Desenvolvimento Científico e Tecnológico (CNPq). J. A. J. and M. L. T. M. P are Research Fellows of CNPq. M. M. was a recipient of a FAPESP fellowship and this work is part of her Doctoral Thesis. It is also part of the project SISBIOTA CNPq: 563260/2010-6 and FAPESP: 2010/52322-3