Development of antioxidant rich peptides from milk protein by microbial proteases and analysis of their effects on lipid peroxidation in cooked beef

Three microbial proteases, validase (Val) from Aspergillus oryze, alkaline protease (AP) from Bacillus licheniformis, and neutral protease (NP) from Bacillus subtilis, were investigated for producing antioxidant hydrolysates/peptides from milk protein isolate. The hydrolysates were fractionated by sequential ultra-filtration and their antioxidant properties examined. The oxygen radical absorbance capacity (ORAC) of the twelve hydrolysate fractions varied significantly ranging from 62.9 to 246.6 μmol Trolox equivalents (TE) per gram with Val-F3 fraction possessing the highest value. The 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) scavenging activities of the hydrolysate fractions also varied significantly ranging from 3.1 to 35.4 μmol TE/g. The results revealed that most hydrolysate fractions exerted weak chelating activities of ranging from 0.1 to 0.9 mg ethylenediaminetetraacetic acid (EDTA) equivalents/g. However, the NP-F2 fraction exerted an exceptionally higher activity of 4.7 mg EDTA equivalents/g. Val-F3, Val-F2, and NP-F2 were incorporated into ground beef to determine their impact on lipid peroxidation. Only the Val-F3 fraction (200 μg/g in the meat) significantly reduced meat lipid peroxidation. Increasing the dosage of VAL-F3 (800 μg/g) did not enhance the inhibition on meat lipid oxidation.