Uso de cisteíno proteases imobilizadas do látex de Calotropis procera para fabricação de queijos

Abstract

With the increasing consumption of cheese and the decline in the production of chymosin of animal origin, new sources of enzymes capable of promoting milk clotting are being studied. Among them, the cysteine proteases originating from the latex of Calotropis procera have promising applications in the biotechnological food industry, including cheese production. The use of free enzymes has some disadvantages, such as low stability at different temperatures and pH, in addition to the loss of enzymes in the process, increasing the cost of production. A possible alternative to these problems is enzymatic immobilization, as it is a technique that helps in the stability of the molecule, prevents contamination of the final product by the enzyme, as well as the enzyme can be used in several cycles of the process, which is extremely important. relevance in the food industry. Thus, the present work explored the immobilization of cysteine proteases obtained from C. procera latex on a glyoxyl-agarose support and evaluated its activity in milk coagulation and cheese production. The enzymes were successfully immobilized on glyoxyl-agarose support (Glyoxyl-CpCPs). When immobilized, they showed activities ranging from 254% to 45%, according to the enzymatic loads (5, 10, 20, 40 and 50 mg/g), when compared to the enzymes, in solution, under similar reaction conditions. The hydrolysis of casein by glyoxyl-CpCPs was similar to its soluble form. Furthermore, glyoxyl-CpCPs exhibited better specific milk clotting activity than their soluble form and chymosin. Immobilized enzymes maintained stable catalytic activity for at least six months at 8 °C. Atomic force microscopy (AFM) and dynamic light scattering (DLS) techniques showed that the aggregation process of casein micelles after treatment with glyoxyl-CpCPs was very similar to that observed with the use of soluble chymosin. Finally, the glyoxyl-CpCPs support, reused in five reaction cycles of cheese manufacturing, maintained the same yield. The study suggests that the immobilization of lacticiferous enzymes potentiated their use as an input, alternative to chymosin, for the production of coalho cheese.