Produção e caracterização de coagulante por fungo filamentoso: uma fonte alternativa e renovável de enzimas proteolíticas

Abstract

The growing demand for enzymes with milk-clotting activity in bovine milk has driven research in the field, leading to an increased search for sources of these enzymes to meet market demands. Proteases are enzymes of significant economic relevance and can be synthesized by various species of filamentous fungi, including those belonging to the genus Aspergillus. This research aimed to evaluate the production and characterization of milk-clotting proteases synthesized by the species Aspergillus oryzae DPUA1624. The culture matrix was developed using CYA [Czapek 0,5% (w/v) + yeast extract 0,5% (w/v)] and maintained for seven days at 25°C. After this period, a spore suspension was inoculated into a liquid medium containing Czapek Yeast Extract (CYA), Sabouraud Extract (SAB), Glucose, Yeast Extract, and Peptone (GYP), and fermentation took place on an orbital shaker at 150 rpm and 25 ºC. After 72 hours, the crude extract was recovered and vacuum-filtered. The activity of the proteases was assessed using 1% (w/v) azocasein as a substrate in Tris-HCl buffer solution, pH 7,2. The production of bovine milk-clotting proteases was also determined regarding the optimal pH and temperature, pH stability, and temperature stability. The results indicated that A. oryzae excreted proteases with significant values in the CYA medium, showing proteolytic activity (17,33 U/mL). The optimal activity of these enzymes was determined at pH 6,0 and 50°C. Coagulating enzymes were detected only in CYA (30,89 U), with optimal activity at pH 5,0 and a temperature of 50°C. The results suggest that A. oryzae proteases can be applied in various industrial sectors, such as the food, textile, and baking industries, expanding their potential use as important biotechnological tools.