Produção concomitante de proteases e peptídeos bioativos por fermentação em estado sólido

Abstract

Biopeptides are compounds that have beneficial effects on health, such as: lowering blood pressure, antioxidant activity, reducing problems caused by diabetes, anticancer and antimicrobial actions, among others. They can be obtained through the hydrolysis of proteins by enzymes, among them, enzymes produced by microorganisms through solid-state fermentation (SSF). This process mimics the natural habitat of the microorganisms, causing them to act in the environment, producing enzymes, such as proteases, and other compounds, all in the same process. One of its main advantages is the use of residues and agro-industrial by-products as substrates, reducing environmental pollution, and allowing the valuation of these compounds. Thus, the objective of this work was to develop a solid-state fermentation biotechnological process for the simultaneous production of proteases and biopeptides. For this, the experiment was divided into three stages. The first was carried out according to a Mixture Design, with the use of three substrates in different proportions: soy bran, soy husk, and wheat bran, in the solid-state fermentation by A. niger. It was evaluated the antioxidant activity and soluble proteins in an acid medium in each medium and, also, the synergism and antagonism of the substrates. Other parameters such as pH (7), temperature (30°C) and moisture (60%) were fixed, following the literature. The second step corresponded to a Complete Factorial Design 23, with three central points, with the study of the variables: pH (5, 6 and 7), moisture (50, 60 and 70%), and nitrogen sources concentrations (0, 5, 1.25 and 2%). The substrates used in the fermentation by A. niger were soy bran and soy husk (70:30). As response variables, protease activity, soluble proteins in acid medium, and antioxidant potential were evaluated. In the third stage, was made the scale-up, and the pH variation (6, 7, and 8) of the medium was studied. The analytical determinations were again performed and the peptides and proteases were characterized in terms of size by gel electrophoresis. As a result, the simultaneous production of proteases and peptides by SSF occurred. The best condition presented in stage 1 was with the soy bran and soy husk mixture. In stage 2, the highest protease production was obtained with assay 2 (pH 7, 50% moisture, and 0.5% nitrogen) in a 48h fermentation with 677 U g-1 and in 96h with 426 U g-1. The highest antioxidant potential was obtained with a 96h fermentation, 2875.5 Trolox equivalent (mg/mL) with the assay 2, and 2988.67 mg/mL with the assay 6 (pH 7, 50% moisture and 2% nitrogen). In the last stage, stage 3, were not observed significant differences concerning the studied pH, and the scale-up was favorable towards the production of proteases and antioxidant biopeptides, with results of 479 U g-1 of proteases in a 96 hours fermentation and 1678.88 Trolox equivalent (mg/mL) for the antioxidant activity, when in pH 7. The solid state fermentation proved to be an excellent bioprocess for the simultaneous production of proteases and biopeptides by A. niger.