Production of cellulolytic enzymes by Trichoderma reesei CCT-2768 through solid-state fermentation using green coconut (Cocos nucifera) fiber pretreated by the steam explosion as substrate.
semi-solid fermentation, green coconut fiber, pretreatments, cellulases.
The residue generated by the green coconut production chain has caused a series of environmental and public health problems as it is a bulky organic material with slow decomposition. An alternative for its use would be in the application in production processes of lignocellulolytic enzymes via semi-solid fermentation (FSS), but a pre-treatment step is necessary to reduce the lignocellulose recalcitrance and the lignin and hemicellulose content in the material, because of that these compounds limit the accessibility of microorganisms. Studies indicate that, for a better induction of cellulase production by FSS processes, pre-treatment or their combination more efficiently exposes cellulose to filamentous fungi. In addition, the use of a suitable solvent during enzyme extraction after FSS also has a strong impact on the performance of cellulase production. Therefore, this study aimed to evaluate the production of cellulases using coconut fiber pretreated by steam explosion combined with alkaline pre-treatment (PV+A) by FSS using filamentous fungus Trichoderma reesei CCT-2768, evaluating the culture conditions (water activity and amount of substrate) through central rotational composite design (CCR), with Carboxymethylcellulase (CMCase) and Filter Paper enzyme activity (FPase) as responses. The combination of pre-treatments was more favorable for the removal of hemicellulose from green coconut fiber, reducing the content of this polymer from 10.15%, about the untreated biomass, to 5.38%, while the content of lignin increased from 21.32% to 34.82%, which may be associated with the lignin rearrangement effect during the biomass cooling process in the pre-treatment. Regarding the SSF of the pre-treated PV+A coconut fiber, it was observed that the solids load significantly influenced the behavior of CMCase in the cultivation, and water activity was shown to be a significant factor in both analyzed enzymes. It could be noticed that greater production of cellulolytic enzymes occurred in the semi-solid culture conditions in which the water activity is 0.995 and 5.25 g of substrate, obtaining an enzymatic activity of CMCases close to 3.0 IU/g, and for FPases, a value of 0.31 IU/g. When the best solvent for cellulase extraction was evaluated, the saline solution of 154 mM NaCl was more efficient in the recovery of the enzymes, producing an extract with the activity of 5.19 IU/g for endoglucanase and 1.19 IU/g for the FPase. Therefore, the best conditions of water activity and solids loading for the production of cellulases were identified, when using the fungus Trichoderma reesei CCT-2768 and coconut fiber pretreated by a steam explosion followed by alkaline pre-treatment, achieving good values of CMCase and FPase activities are found.