Isolation and identification of antioxidative peptides from rice endosperm protein enzymatic hydrolysate by consecutive chromatography and MALDI-TOF/TOF MS/MS

The defatted rice endosperm protein (REP) was, respectively, digested by five different protease treatments (Alcalase, Chymotrypsin, Neutrase, Papain and Flavorase), and Neutrase appears to be the most desirable for producing high quality antioxidant peptides from REP. Specially, the DPPH and hydroxyl radical scavenging activities of NHREP were higher than its superoxide radical scavenging activity, and the percentage inhibition of autooxidation of NHREP (80.09%) was similar to that of α-tocopherol (86.59%) on day 5. Furthermore, NHREP was purified consecutively, and the antioxidant peptides were identified to be Phe-Arg-Asp-Glu-His-Lys-Lys (FRDEHKK, 959.5 Da) and Lys-His-Asp-Arg-Gly-Asp-Glu-Phe (1002.5 Da) by MALDI-TOF/TOF MS/MS. Lastly, FRDEHKK was chemically synthesised. It significantly inhibited lipid peroxidation in an linoleic acid emulsion system more effectively than α-tocopherol, and enhanced the viability of t-BHP induced cytotoxicity up to 74.38% (for MRC-5) and 78.39% (for RAW264.7) at 80 μg/ml. Conclusively, it was feasible to produce natural antioxidants from REP.