Xilanases produzidas por fungo endofítico utilizando sistema micelar reverso

Abstract

Enzymes are proteins that catalyze with great efficiency several biochemical reactions, being crucial for the viability of industrial and biotechnological processes due to their high specificity. Xylanases (EC 3.2.1.8) are hydrolytic enzymes produced by several microorganisms, responsible for the decomposition of plant cell walls, with great applicability in industrial processes, such as in the production of feed, paper and food. These enzymes have been extensively studied, since despite their varied applications, their high cost, their instability, and also their difficult purification from the culture medium, limit its commercialization. The search for efficient methods of extracting and recovering enzymes with reduced loss of activity, therefore, is of great relevance. In this context, the objective of this work was to evaluate the production of xylanases from the cultivation in liquid medium of an Amazonian endophytic fungi, and to investigate the use of the liquid-liquid extraction method by Reverse Micellar System (RMS) in the recovery of the enzymatic activity. The influence of different concentrations of the cationic surfactant hexadecyl trimetrylammonium bromide (CTAB) and the ionic strength in the medium used for the extraction were studied, throught a RCCD experimental design. The Aspergillus sp. F36 was cultured in liquid medium containing 1% xylan for 48 h. After separating the mycelium, the enzymatic extract was used for liquid-liquid extraction by RMS. Protein concentration and enzymatic activity were determined in the samples before and after extraction. The production of xylanases was detected in the fermented broth (6.87 U/mL) and after liquid liquid extraction by RMS under all conditions studied. The assay with 0.564 M of CTAB and 12.0 mS/cm of ionic strength promoted the greatest recovery of the enzymatic activity (202.7%). It was demonstrated that the xylanases recovery through RMS extraction presented critical values at the minimum point of 0.47 M for the surfactant and 13.830 mS/cm of ionic strength, according to the equation obtained in the mathematical model. The value of total protein yield (np) showed that the method has a high selectivity in extracting the enzyme under the conditions studied, in addition to favoring the maintenance of its integrity, which was also observed by an increase in the purification factor. The experimental conditions tested did not inhibit the enzymatic activity of xylanases after extraction by the RMS, indicating the method for recovering these enzymes from the fungal culture medium. With this study, it is concluded that the RMS method can be used for the isolation of the enzyme of interest and that the cationic surfactant CTAB in its highest concentration tested promotes a higher yield of the enzyme in the extraction by RMS.