Produção de celulases por fermentação submersa utilizando micro-organismos prospectados em coleções de culturas nacionais

Abstract

The research about alternative sources of fuels has been mainly focused on biomass composed of cellulose, hemicellulose and lignin. The structural carbohydrates contained in the cellulosic matrix represent the substrate that can be used for biofuel production by fermentative processes. The choice of the biomass enzymatic hydrolysis of plants using cellulases, can generate an option less environmental impact compared to other processes of hydrolysis. Studies about microbial new sources, and more accurate analysis of the steps that make up the production of cellulases are essential as a strategy to reduce the costs generated by the use of cellulases in the process of obtaining fermentable sugars. This study aims at the selection of filamentous fungi producers of cellulolytic enzymes, as well as investigating the parameters of cellulase production by submerged fermentation process. In the initial stage were selected micro-organisms by methodology in plates. Was investigated the potential cellulolytic enzyme through the index in a specific medium and were selected strains that reached a diameter of colony growth of 150 mm in Petri dishes in a shorter time of incubation (48 hours). Were selected micro-organisms with the greatest potential enzymatic tests that were then tested in submerged fermentation. The first test consisted in selecting the best microorganisms as producers of FPase in non-baffled flask containing 100 mL of culture medium specific. In the subsequent test were examined the three best strains from the previous step, which were then subjected to submerged fermentation in baffled flasks. The next step was designed to investigate the enzyme production in fermentation with four types of soluble sugars: glucose, lactose, sucrose and xylose. The fungal strain was selected in the previous steps used in the tests in a bioreactor, which analyzed three different strategies of inoculation. The strains Fusarium sp. SAP 09, Lasiodiplodia theobromae CNPAT 040, Trichoderma sp. LCB 79, Trichoderma sp. INPA 666, Trichoderma sp. INPA 1014 and Trichoderma sp. INPA 1218 were selected from the first stage. In the selection stage by submerged fermentation in non-baffled flasks, the best FPase activity was achieved by the strain Trichoderma sp. INPA 666 (48.0 FPU /L) and CMCase by the fungus Lasiodiplodia theobromae CNPAT 040 (350.0 U /L). In comparison with baffled flasks, there was a greater production both FPase and CMCase in Erlenmeyer without baffles, indicating that the shear forces applied to the fungal cultures were potentially harmful for enzyme production. The use of sucrose proved to be the best option among soluble sugars tested, with higher rates of FPase activity (49.9 FPU / L) and CMCase (119.7 U / L). The best strategy for the inoculation was a spore suspension obtained from a solid state fermentation of wheat bran, in the time of 72 hours of fermentation.