In vitro antioxidant activity of a peptide isolated from Nile tilapia (Oreochromis niloticus) scale gelatin in free radical-mediated oxidative systems

In the present study, a peptide possessing antioxidant properties was isolated from Nile tilapia (Oreochromis niloticus) scale gelatin. Gelatin protein was hydrolyzed using alcalase, pronase E, trypsin and pepsin. Antioxidant efficacy of respective hydrolysates were evaluated using 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, hydroxyl radical and superoxide radical anion scavenging activities. Moreover, protective effect on DNA damage caused by hydroxyl radicals generated was determined. Further, the level of reactive oxygen species (ROS) was determined using a fluorescence probe, 2',7'-dichlorofluorescin diacetate (DCFH-DA), which could be converted to highly fluorescent dichlorofluorescein (DCF) with the presence of intracellular ROS on mouse macrophages, RAW 264.7 cells. Among hydrolysates, alcalase-derived hydrolysate exhibited the highest antioxidant activity compared to other enzymatic hydrolysates. Therefore, it was further analyzed and the sequence of an active peptide present in it was identified as Asp-Pro-Ala-Leu-Ala-Thr-Glu-Pro-Asp-Pro-Met-Pro-Phe (1382.57Da). This peptide showed no cytotoxic effect on mouse macrophages (RAW 264.7) and human lung fibroblasts (MRC-5). In addition, it scavenged hydroxyl, DPPH and superoxide radicals at the IC₅₀ values of 7.56, 8.82 and 17.83μM, respectively. These results suggest that the peptide derived from Nile tilapia (O. niloticus) scale gelatin acts as a candidate against oxidative stress and could be used as a potential functional food ingredient.