羊血红蛋白水解物中新型抗氧化肽的纯化、鉴定和对H_(2)O_(2)损伤Caco-2细胞保护作用  

Purification and Identification of Novel Antioxidant Peptides from Sheep Hemoglobin Hydrolysate and Their Protective Effect on H2O2-Induced Oxidative Injury in Caco-2 Cells

作　　者：马泽浩 耿玉坤 王静云[1] 卢士玲[1] 曹逗逗 刘星语 MA Zehao;GENG Yukun;WANG Jingyun;LU Shiling;CAO Doudou;LIU Xingyu(School of Food Science and Technology,Shihezi University,Shihezi 832000,China)

机构地区：[1]石河子大学食品学院,新疆石河子832000

出　　处：《食品科学》2024年第11期41-51,共11页Food Science

基　　金：石河子大学青年创新培育人才项目(CXPY202108);石河子大学高层次人才科研启动项目(RCZK202021);兵团指导性科技计划项目(2022ZD064);五师科技计划项目(2023XM02)。

摘　　要：以新鲜羊血为原料制备羊血红蛋白抗氧化肽,采用葡萄糖凝胶G-25色谱、DEAE葡萄糖凝胶A-50阴离子交换色谱、反相高效液相色谱和液相色谱-串联质谱对羊血红蛋白水解物进行分离纯化和鉴定,通过研究自由基清除能力、模拟胃肠消化后的稳定性及对H2O2诱导的Caco-2细胞保护作用评价羊血红蛋白肽的抗氧化活性。结果表明,从羊血红蛋白粗肽中纯化并鉴定出Ala-Tyr-Glu-Val-Asp(AYEVD)、Phe-His-Thr-Met-Glu(FHTME)、Ser-PheMet-Tyr-Glu-Lys(SFMYEK)3种新型抗氧化活性肽,分子质量分别为595.60、663.74、803.92 Da;其中AYEVD的1,1-二苯基-2-三硝基苯肼自由基清除能力最强;AYEVD在模拟胃肠道消化后显示出更强的抗氧化活性。此外,AYEVD还能抑制H2O2诱导Caco-2细胞的氧化损伤,显著减少活性氧积累、抑制早期细胞凋亡和丙二醛的形成,并提高细胞内抗氧化酶活性。该研究可为羊血红蛋白肽作为新型抗氧化剂应用于功能食品提供理论依据。In this study,antioxidant peptides derived from sheep hemoglobin were prepared from fresh sheep blood.Sephadex G-25 chromatography,DEAE Sephadex A-50 chromatography,reverse phase high performance liquid chromatography(RP-HPLC)and liquid chromatography-tandem mass spectrometry(LC-MS/MS)were used to separate,purify,and identify peptides from the hydrolysate of sheep hemoglobin.The antioxidant activity of the obtained peptides was evaluated by their free radical scavenging capacity,stability against simulated gastrointestinal digestion and protective effect on H2O2-induced oxidative injury in Caco-2 cells.The results indicated that three novel antioxidant peptides,Ala-Tyr-Glu-Val-Asp(AYEVD),Phe-His-Thr-Met-Glu(FHTME)and Ser-Phe-Met-Tyr-Glu-Lys(SFMYEK),were purified and identified from the hydrolysate,with molecular masses of 595.60,663.74 and 803.92 Da,respectively.Among them,AYEVD had the strongest scavenging capacity to scavenge 1,1-diphenyl-2-picrylhydrazyl(DPPH)radical,and AYEVD showed stronger antioxidant activity after simulated gastrointestinal digestion.In addition,AYEVD inhibited H2O2-induced oxidative injury in Caco-2 cells,significantly reduced the accumulation of reactive oxygen species(ROS),suppressed early cell apoptosis and the formation of malondialdehyde(MDA),and increased the activity of intracellular antioxidant enzymes.Therefore,this study could provide a theoretical basis for the application of novel antioxidant peptides from sheep hemoglobin in functional foods.

关 键 词：羊血红蛋白 抗氧化肽 纯化 鉴定 Caco-2细胞 细胞保护 

分 类 号：TS251.93[轻工技术与工程—农产品加工及贮藏工程]