Abstract: In order to explore the potential antioxidative properties of poultry blood, several proteases were screened for use in the production of antioxidative peptide from chicken red blood cells and the enzymatic hydrolysis process was optimized using response surface methodology. The hydrolysate was purified sequentially by ultrafiltration, cation exchange chromatography, gel filtration chromatography, and high-performance liquid chromatography. The antioxidative peptide obtained was identified by mass spectrometry. The results showed that the hydrolysate obtained using acidic protease had the highest scavenging activity against 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical (> 80%) among 6 proteases used. The optimal hydrolysis conditions were obtained as follows: enzyme/substrate (E/S) ratio 2%, temperature 45 ℃ and time 3.0 h. The DPPH radical scavenging activity, superoxide ion-scavenging activity and reducing power of the hydrolysate obtained using the optimized conditions were (98.31 ± 0.66)%, (28.89 ± 0.31)% and (1.94 ± 0.03), respectively. After purification, the fraction with the highest antioxidative activity was obtained, whose DPPH radical-scavenging activity and reducing power were (87.16 ± 1.59)% and (0.21 ± 0.01) at 0.1 mg/mL, respectively. Finally, the peptide was identified by liquid chromatography-mass spectrometry (LC-MS/MS) as MGQKDSYVGDEAQSKRGILT with a molecular mass of 2 182.1 Da.

Key words: antioxidative peptide, chicken blood corpuscle, response surface methodology, isolation and purification, structural?identification