Purificação, caracterização e aplicação de uma exopoligalacturonase de Aspergillus terreus PA3A5T na clarificação de sucos de frutas

Abstract

With advances in the biotech sector and the growing demand to optimize and reduce production costs, the use of enzymes, mainly of microbial origin, has increased. Among these enzymes, pectinases have been used in fruit juice clarification processes, reducing their viscosity and turbidity, thus allowing for clear and stable juices with high added value. This study aimed to purify, characterize and evaluate the application of an exopolygalacturonase from Aspergillus terreus PA3A5T in the clarification of fruit juices. From the optimization of the cultivation conditions by the methodology of the Central Composite Rotational Design (CCRD), an increase of 1.45 times in the enzymatic activity of the pectinase was obtained, adopting as the best condition the Vogel medium supplemented with 3% of the source of carbon (passion fruit husk), at a temperature of 28°C, for 120 hours (test 6). For pectinase present in the crude extract, the highest enzymatic activity was observed at 65°C, in pH 6.0, being stable at 40°C in acidic pH. Purified pectinase presented a relative molecular mass of approximately 95.2 kDa, maximum activity at 70°C in pH 4.0, remaining stable in acidic pH and at 60°C. FeCl2, SnCl2, CoCl2, ZnSO4, NaCl, Al2(SO4)3, MgCl2, MnCl2 e CuSO4 ions induced the enzymatic activity, while PbCl2 completely inhibited it. The values of Km and Vmax were 1.83 mg mL-1 and 335 U mg- 1, respectively. Pectinase showed high catalytic activity on polygalacturonic acid and no activity on beech xylan and CMC, thus it was classified as a polygalacturonase. The analysis of the hydrolysis products showed that the enzyme was able to release only monogalacturonate as a degradation product of polygalacturonate, being characterized as an exopolygalacturonase (EC 3.2.1.67). The exo-PG from Aspergillus terreus PA3A5T proved to be very effective in the clarification process of the juices studied, with an increase of 44 and 22% for the transmittance, with a color reduction of 48 and 52% for orange and mango juice, respectively.