Optimization of Penicillium aurantiogriseum protease immobilization on magnetic nanoparticles for antioxidant peptides' obtainment.

This work reports an optimization of protease from Penicillium aurantiogriseum immobilization on polyaniline-coated magnetic nanoparticles for antioxidant peptides' obtainment derived from bovine casein. Immobilization process was optimized using a full two-level factorial design (2) followed by a response surface methodology. Using the derivative, casein was hydrolyzed uncovering its peptides that were sequenced and had antioxidant properties tested through (2,2'-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt) (ABTS) radical scavenging and hydrogen peroxide scavenging assays. Optimal conditions for immobilization were 2 hr of immobilization, offered protein amount of 200 µg/mL, immobilization pH of 6.3 and 7.3 hr of activation. Derivative keeps over 74% of its original activity after reused five times. Free and immobilized enzyme casein hydrolysates presented similar peptide mass fingerprints, and prevalent peptides could be sequenced. Hydrolysates presented more than 2.5× higher ROS scavenging activity than nonhydrolyzed casein, which validates the immobilized protease capacity to develop casein-derived natural ingredients with potential for functional foods.