Purification and characterization of antioxidant peptide from hoki (Johnius belengerii) frame protein by gastrointestinal digestion.
To extract antioxidant peptide from hoki frame protein hydrolysate (APHPH), we employed six proteases (pepsin, trypsin, papain, alpha-chymotrypsin, Alcalase and Neutrase) for enzymatic hydrolysis, and the antioxidant activities of their hydrolysates were investigated using both lipid peroxidation inhibition assay and free radical scavenging assay by electron spin resonance spin-trapping technique. Among hydrolysates, peptic hydrolysate, having the highest antioxidant activity, further separated into four groups using ultrafiltration membranes and purified consecutive chromatographic methods. Finally, the purified peptide had a molecular mass of 1801 Da, and amino acid sequence was identified as Glu-Ser-Thr-Val-Pro-Glu-Arg-Thr-His-Pro-Ala-Cys-Pro-Asp-Phe-Asn. APHPH inhibited lipid peroxidation higher than that of alpha-tocopherol as positive control and efficiently quenched different sources of free radical: 1,1-diphenyl-2-pycryl-hydrazyl (IC(50)=41.37 microM), hydroxyl (IC(50)=17.77 microM), peroxyl (IC(50)=18.99 microM) and superoxide radicals (IC(50)=172.10 microM). Furthermore, APHPH decreased t-butylhydroperoxide-induced cytotoxicity on human embryonic lung fibroblasts and efficiently protected free-radical-induced DNA damage.
Kim
Soo-Yong
SY
Department of Chemistry, Pukyong National University, Busan 608-737, Korea.
Je
Jae-Young
JY
Kim
Se-Kwon
SK
D000595:Amino Acid Sequence; D000818:Animals; D000975:Antioxidants / Q000302:isolation & purification* / Q000494:pharmacology; D016923:Cell Death / Q000187:drug effects; D004249:DNA Damage / Q000187:drug effects; D004578:Electron Spin Resonance Spectroscopy; D004622:Embryo, Mammalian; D004625:Embryo, Nonmammalian; D005347:Fibroblasts / Q000187:drug effects; D005396:Fish Products / Q000032:analysis*; D029941:Fish Proteins / Q000737:chemistry* / Q000378:metabolism; D041981:Gastrointestinal Tract / Q000201:enzymology*; D006801:Humans; D017665:Hydroxyl Radical / Q000494:pharmacology; D008969:Molecular Sequence Data; D010447:Peptide Hydrolases / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D010473:Perciformes; D011492:Protein Hydrolysates / Q000737:chemistry; D020122:tert-Butylhydroperoxide / Q000494:pharmacology
D000975:Antioxidants; D029941:Fish Proteins; D010455:Peptides; D011492:Protein Hydrolysates; D017665:Hydroxyl Radical; D020122:tert-Butylhydroperoxide; D010447:Peptide Hydrolases