The objective of this study was to identify anti-inflammatory peptides from simulated gastrointestinal digest (pepsin-pancreatin hydrolysate) of velvet antler protein. The hydrolysate was purified using ultrafiltration and consecutive chromatographic methods. The anti-inflammatory activity of the purified fraction was evaluated by the inhibition of NO production in lipopolysaccharide-induced RAW 264.7 macrophages. Four anti-inflammatory peptides, VH (Val-His), LAN (Leu-Ala-Asn), AL (Ala-Leu), and IA (Ile-Ala), were identified by liquid chromatography/tandem mass spectrometry. Each of these peptides demonstrated a U-shaped dose-effect relationship. VH, LAN, AL and IA showed the strongest anti-inflammatory activities at 200 μg/mL, that is, 15.5%, 13.0%, 16.0% and 11.2% inhibition of lipopolysaccharide-induced NO production, respectively. Additionally, the enhanced NO inhibitory activity was observed for the peptides mixture, indicating the possible synergistic effects. These results suggested that the peptides derived from velvet antler protein could potentially be used as a promising ingredient in functional foods or nutraceuticals against inflammatory diseases.
