Influência das fontes de carbono na indução de celulases e hemicelulases em Chrysoporthe cubensis cultivado em meio líquido

• Autor(a) principal: Dutra, Thiago Rodrigues
• Data de Publicação: 2013
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: LOCUS Repositório Institucional da UFV
• Texto Completo: http://locus.ufv.br/handle/123456789/2463
• topic: Carbono; Hemicelulases; Celulases; Chrysoporthe cubensis; Carbon; Hemicellulases; Cellulases; Chrysoporthe cubensis; CNPQ::CIENCIAS BIOLOGICAS::BIOQUIMICA

Results obtained recently in our research group (Production and Characterization of biotechnological interest Enzyme) have demonstrated that Chrysoporthe cubensis, fungal pathogen of eucalyptus, has showed potential for producing enzymes involved in saccharification of lignocellulosic biomass. In this work C. cubensis was cultivated in several carbon sources with the view to observe the production of cellulases and hemicellulases. Complex carbon sources such as wheat bran, carboxymethilcellulose CMC, Birchwood xylan XB, Locust bean gum LBG, citrus peel CP and also simpler carbon sources such as glucose, cellobiose, lactose, galactose, mannose, xylose and arabinose were tested as enzymatic activity inducers. C. cubensis produced greater levels of celullolytic and hemicellulolytic activities when added with wheat bran and CMC. When cultivated in wheat bran C. cubensis reached values for β-glucosidase, endoglucanase and xylanase activities of 0,36 U/mL, 3,6 U/mL, 14,2 U/mL, respectively. When cultivated in CMC, C. cubensis produced β-glucosidase, endoglucanase and xylanase activities of 0,48 U/mL, 2,8 U/mL, 10,7 U/mL, respectively. When cultivated in birchwood xylan extracted produced by C. cubensis presented xylanase activity of 9,9 U/mL, however it was not efficient for inducing cellulases. Simpler carbon sources generally presented low inducing capacity in C. cubensis. Zimograms were made with the aim to analize qualitatively the enzymes present in extracts produced by fungus C. cubensis. Zimogram containning fluoorescent substrate of 4-Methylumbelliferyl β-D- glucopyranoside, has demonstrated that C. cubensis was able to produce β-glicosidase activity, when cultivated in all sources of carbon, except in mannose and galactose. Extracts produced by C. cubensis, cultivated in complex sources, presented a β-glucosidase of 55,6 kDa. Presences of possible complexes of high molecular weight, containning β-glucosidase activity were also detected. Among extracts produced in the presence of simpler sources, lactose induced the same group of β-glucosidases as complex sources. Xylose and arabinose also showed activity bands with high molecular weights. Xylose also induced a β-glucosidase of 74 kDa and the arabinose was able to induce the production of a β-glucosidase of 48 kDa and other of 55,6 kDa. Glucose induced a β-glucosidase of 106,8 kDa and other of 48 kDa. Zimogram containing birchwood xylan as substrate, has demonstrated that C. cubensis was able of producing a xylanase constitutive of 19 kDa when added in all sources tested. XB-SDS-PAGE has shown that C. cubensis was able to produce a constitutive xylanase of 19 kDa, when grown in any sources tested. Extracts produced by wheat bran, CMC and birchwood xylan, presented a well defined region activity of xylanase between 97 and 39 kDa. Locust bean gum and citrus peel were able to induce the production of a xylanase of 37,3 kDa. Xylose has induced the production of two xylanases, one of 53,2 kDa and other of 47,9 kDa. The extract produced by the fungus added in arabinose, presented a band standard similar to what is found in the extract from grown in xylose. Glucose, lactose, galactose and mannose showed only the basal activity of 19 kDa xylanase. Zimogram containing CMC as substrate has demonstrated that only extract produced by C. cubensis added in mannose and xylose, it was not possible to observe the activity of endoglucanase. Wheat bran, CMC and Birchwood xylan induced the production of two endoglucanases, one of 38 kDa and other of 55,6 kDa. When C. cubensis grew in glucose, LBG, CP, lactose, galactose, arabinose and cellobiose it was possible to view the production of one endoglucanase of 38 kDa. Analizyng results of enzymatic activities together with zimogram analysis, it is clear that complex sources as wheat bran, CMC and Birchwood xylan have high inducing potential in relation to the production of enzymes involved with biomass scarification. Simpler sources, on the other hand, although they do not present meaningful values of enzymatic activities, may trigger production of cellulases as much as hemicellulases, albeit in discrete level.