Production, Pre-Purification and Encapsulation of Proteolytic Enzyme Secreted by Filamentous Fungus

Abstract

Encapsulation of enzymes allows the preservation of their biological activities in various environmental conditions, and is important for industrial application. Thus, the objective of this work was to produce, partially purify and encapsulate the microbial protease enzyme obtained by semisolid fermentation. The enzyme extract was produced from the fermentation of substrate with different physicochemical compositions by the fermentation agent the filamentous fungus Aspergillus oryzae. An experimental mixture design was used to map out the experimental region and determine the optimal ratios between substrates. The enzyme extract obtained from the optimized formulation was subjected to partial purification in ammonium sulfate, dialysis, and then lyophilized. With the freeze-dried enzyme the enzymatic determination, milk coagulation, thermogravimetric analysis, and differential thermal analysis were performed. The encapsulation process was carried out using the ionic gelation technique in the presence of the polymer sodium alginate and calcium chloride dihydrate. X-ray diffractometry (XRD) identified that the studied babassu mesocarp, wheat bran and babassu coconut fiber samples showed amorphous structural features, presence of cellulose and lignin. The protease obtained in the formulation of the optimized medium, precipitated and freeze-dried, presented milk coagulation capacity, proteolytic activity of 261.5 U/g and through thermal analysis it was verified that the material has thermal stability around 40 °C. It was possible to encapsulate the enzyme extract and preserve the enzymatic activity and the coagulation capacity of the milk. The results showed the optimization of the medium formulation, as well as the potential for industrial application of the purified and encapsulated enzyme, preserving the activity and enabling the application and reuse of the enzyme.