Production and characterization of glucoamylase from fungus Aspergillus awamori expressed in yeast Saccharomyces cerevisiae using different carbon sources

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Protease production by different thermophilic fungi

A comparative study was carried out to evaluate protease production in solid-state fermentation (SSF) and submerged fermentation (SmF) by nine different thermophilic fungi - Thermoascus aurantiacus Miehe, Thermomyces lanuginosus, T. lanuginosus TO.03, Aspergillus flavus 1.2, Aspergillus sp. 13.33, Aspergillus sp. 13.34, Aspergillus sp. 13.35, Rhizomucor pusillus 13.36 and Rhizomucor sp. 13.37 - using substrates containing proteins to induce enzyme secretion. Soybean extract (soybean milk), soybean flour, milk powder, rice, and wheat bran were tested. The most satisfactory results were obtained when using wheat bran in SSF. The fungi that stood out in SSF were T. lanuginosus, T. lanuginosus TO.03, Aspergillus sp. 13.34, Aspergillus sp. 13.35, and Rhizomucor sp. 13.37, and those in SmF were T. aurantiacus, T. lanuginosus TO.03, and 13.37. In both fermentation systems, A. flavus 1.2 and R. pusillus 13.36 presented the lowest levels of proteolytic activity.

Screening of filamentous fungi for production of enzymes of biotechnological interest

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Purification and characterization of a cyclomaltodextrin glucanotransferase from Paenibacillus campinasensis strain H69-3

A cyclomaltodextrin glucanotransferase (E.C. 2.4.1.19) from a newly isolated alkalophilic and moderately thermophilic Paenibacillus campinasensis strain H69-3 was purified as a homogeneous protein from culture supernatant. Cyclomaltodextrin glucanotransferase was produced during submerged fermentation at 45 degrees C and purified by gel filtration on Sephadex G50 ion exchange using a Q-Sepharose column and ion exchange using a Mono-Q column. The molecular weight of the purified enzyme was 70 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the pI was 5.3. The optimum pH for enzyme activity was 6.5, and it was stable in the pH range 6.0-11.5. The optimum temperature was 65 degrees C at pH 6.5, and it was thermally stable up to 60 degrees C without substrate during 1 h in the presence of 10 mm CaCl2. The enzyme activity increased in the presence of Co2+, Ba2+, and Mn2+. Using maltodextrin as substrate, the K-m and K-cat were 1.65 mg/mL and 347.9 mu mol/mg.min, respectively.

Production, characterization and properties of polysaccharide depolymerizing enzymes from a strain of Curvularia inaequalis

Xylanase, β-glucosidase, β-xylosidase, endoglucanase and polygalacturonase production from Curvularia inaequalis was carried out by means of solid-state and submerged fermentation using different carbon sources. β-Glucosidase, β-xylosidase, polygalacturonase and xylanase produced by the microorganisms were characterized. β-Glucosidase presented optimum activity at pH 5.5 whereas xylanase, polygalacturonase and β-xylosidase activities were optimal at pH 5.0. Maximal activity of β-glucosidase was determined at 60°C, β-xylosidase at 70°C, and polygalacturonase and xylanase at 55°C. These enzymes were stable at acidic to neutral pH and at 40-45°C. The crude enzyme solution was studied for the hydrolysis of agricultural residues.

A specific short dextrin-hydrolyzing extracellular glucosidase from the thermophilic fungus Thermoascus aurantiacus 179-5

The thermophilic fungus Thermoascus aurantiacus 179-5 produced large quantities of a glucosidase which preferentially hydrolyzed maltose over starch. Enzyme production was high in submerged fermentation, with a maximal activity of 30 U/ml after 336 h of fermentation. In solid-state fermentation, the activity of the enzyme was 22 U/ml at 144 h in medium containing wheat bran and 5.8 U/ml at 48 h when cassava pulp was used as the culture medium. The enzyme was specific for maltose, very slowly hydrolyzed starch, dextrins (2-7G) and the synthetic substrate (α-PNPG), and did not hydrolyze sucrose. These properties suggest that the enzyme is a type II α-glucosidase. The optimum temperature of the enzyme was 70°C. In addition, the enzyme was highly thermostable (100% stability for 10 h at 60°C and a half-life of 15 min at 80°C), and stable within a wide pH range. Copyright © 2006, The Microbiological Society of Korea.

Análise dos parâmetros cinéticos para produção de levana por Zymomonas mobilis utilizando fermentação submersa

Levan is an exopolysaccharide synthesized by several microorganisms during fermentation of a culture medium containing sucrose, yeast extract and minerals. This biopolymer has applications in the food segment as stabilizers, thickeners, as carriers for flavor and fragrances, as well as in the pharmaceutical segment as a hypocholesterolemic agent and for exhibiting antitumor activity. This work aimed to analyze the kinetic parameters for levan production. The microorganism used was Zymomonas mobilis CCT 4494, incubated in a synthetic medium containing 200.0 g L-1 of sucrose, in a rotary shaker at 200 rpm and 30°C. Samples were taken every 24h, during a period of 96h, in order to determine variations of pH, biomass, reducing sugar, total reducing sugar and levan formation. It was observed that yields of biomass and synthesized biopolymer in 24h were superior to those obtained in 48, 72 and 96h of fermentation.

Agaricus blazei as a Substrate for the Production of β-1,3-Glucanase by Trichoderma harzianum Rifai

Extracellular β-1,3-glucanase was produced by Trichoderma harzianum Rifai cultivated in the Agaricus blazei (Agaricus brasiliensis) extract as a substrate in submerged fermentation. A 22-central composite factorial design was developed using the time of culture (x1/day) and Agaricus blazei extract concentration (x2/(g/L)) as variables, and the results were analyzed using response surface methodology (RSM). The results showed that the Agaricus blazei extract concentration was the most important variable in the production of β-1,3-glucanase, and the maximum β-1,3-glucanase activity (0.77 U/mL) was obtained in one day of cultivation. The β-glucan present in the cell wall of Agaricus blazei mushroom proved to be a good substrate for inducing the production of specific β-1,3-glucanase by Trichoderma harzianum Rifai.

Wine aroma improvement using a β-glucosidase preparation from aureobasidium pullulans

Microbial β-glucosidases have been used for the enhancement of wine aroma. Nevertheless, few enzymes are active in the conditions of winemaking. In this work, the production of a β-glucosidase by an Aureobasidium pullulans strain (Ap-β-gl) isolated from grape ecosystems was evaluated. The maximum enzymatic synthesis using submerged fermentation was after 96 h of growth in complex media containing 20 g/L of cellobiose as the sole carbon source. The crude enzyme (Ap-β-gl) showed optimal pH at 5.5 and two peaks of optimum temperature (at 45 and 70 C). It showed a wide range of pH stability, stability at low temperatures, and tolerance to ethanol, showing suitable characteristics for winemaking conditions. The hydrolysis of glycosidic terpenes by Ap-β-gl was studied, and its ability to efficiently release free terpenols was demonstrated by gas chromatography/mass spectrometry. The enzymatic treatment notably increased the amount of monoterpenes, showing good prospects for its potential application for the development of aroma in wines. © 2012 Springer Science+Business Media New York.

Avaliação de dois diferentes meios de cultura para o desenvolvimento de biopesticidas à base de Bacillus thuringiensis e sua aplicação em larvas de Aedes aegypti

The bacteria Bacillus thuringiensis var. israelensis (Bti) generates certain toxins with pesticide action, which can be used on the control of transmissible diseases by culicides, specially Aedes aegypti, the dengue vector. This biopesticide has been produced by submerged fermentation and, in Brazil, this production has been made by very little research centers and, more recently, by a unique small enterprise. For the implementation of a viable vectors control program through biopesticides, some studies about culture media are essential in order to join efficiency and low costs. In this way, agroindustrial wastes or by-products have been used as a nutrient source for the culture media production. In this study, corn steep liquor, a corn industrial processing by-product and tryptose, both with/without sugar addition, were compared as culture media. Cellular growth was evaluated by optical density at 620 nm, spore production by total viable cell count and LC50 by bioassays against 4th instar larvae. Among the four examined substrates, the medium composed by glucose plus corn steep liquor presented the best spore production and bioassay results.

Lipase production by Botryosphaeria ribis EC-01 on soybean and castorbean meals: Optimization, immobilization, and application for biodiesel production

The effects of soybean and castorbean meals were evaluated separately, and in combinations at different ratios, as substrates for lipase production by Botryosphaeria ribis EC-01 in submerged fermentation using only distilled water. The addition of glycerol analytical grade (AG) and glycerol crude (CG) to soybean and castorbean meals separately and in combination, were also examined for lipase production. Glycerol-AG increased enzyme production, whereas glycerol-CG decreased it. A 24 factorial design was developed to determine the best concentrations of soybean meal, castorbean meal, glycerol-AG, and KH2PO4 to optimize lipase production by B. ribis EC-01. Soybean meal and glycerol-AG had a significant effect on lipase production, whereas castorbean meal did not. A second treatment (22 factorial design central composite) was developed, and optimal lipase production (4,820 U/g of dry solids content (ds)) was obtained when B. ribis EC-01 was grown on 0.5 % (w/v) soybean meal and 5.2 % (v/v) glycerol in distilled water, which was in agreement with the predicted value (4,892 U/g ds) calculated by the model. The unitary cost of lipase production determined under the optimized conditions developed ranged from US$0.42 to 0.44 based on nutrient costs. The fungal lipase was immobilized onto Celite and showed high thermal stability and was used for transesterification of soybean oil in methanol (1:3) resulting in 36 % of fatty acyl alkyl ester content. The apparent K m and V max were determined and were 1.86 mM and 14.29 μmol min -1 mg-1, respectively. © 2013 Springer Science+Business Media New York.

Purificação e caracterização bioquímica da invertase extracelular produzida pelo fungo filamentoso Aspergillus terreus

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Purificação e caracterização bioquímica de tanases produzidas pelo fungo filamentoso Aspergillus phoenicis

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Estudo das atividades xilanásicas produzidas pelo fungo Aspergillus caespitosus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Isolamento de linhagens microbianas termofílicas amilolíticas, produção, caracterização e aplicação das amilases na hidrólise do amido de mandioca

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)