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目的分离纯化血红素肽-铁,并分析其清除自由基的能力。方法以聚丙烯酰胺为载体,对利用真空及低压高频脉冲胰蛋白酶降解猪血红蛋白的产物进行金属螯合亲和层析分离纯化,利用基质辅助激光解析串联飞行时间质谱(MALDI-TOFMS)及生物信息学网站分析血红素肽-铁的分子结构;以维生素C(Vc)为对照,分析血红素肽-铁的抗氧化能力;以邻苯三酚法分析血红素肽-铁清除超氧阴离子的能力;以脱氧核糖法分析血红素肽-铁清除羟基自由基(·OH)的能力;以三氯甲烷-甲醇法分析血红素肽-铁对脂质过氧化的抑制作用。结果分离纯化的血红素肽-铁由139个氨基酸组成,相对分子质量为16065.08,铁离子含量为11013.5μg/L,理论pI为8.10,消光系数为6990,脂肪系数为92.73,不稳定系数为1.18,总平均疏水性为-0.023;在体外具有较强的清除超氧阴离子和抗氧化自由基能力。结论分离纯化的血红素肽-铁具有清除自由基的能力。
Objective To isolate and purify heme peptide - iron and analyze its ability to scavenge free radicals. Methods Polyacrylamide was used as carrier to separate and purify porcine hemoglobin by using vacuum and low-pressure high-frequency pulsed trypsin. Metal chelate affinity chromatography was used to separate and purify porcine hemoglobin. MALDI-TOF MS and bioassay The molecular structure of heme peptide-iron was analyzed by informatics website. The anti-oxidation ability of heme peptide-iron was analyzed by vitamin C (Vc). The pyrogallol method was used to analyze the heme peptide-iron scavenging superoxide anion Ability of clearing heme hydroxyl radical (· OH) by deoxyribose method; and the inhibitory effect of heme peptide-iron on lipid peroxidation by chloroform-methanol method. Results The purified heme peptide-iron consisted of 139 amino acids with a relative molecular mass of 16065.08, an iron content of 11013.5 μg / L, a theoretical pI of 8.10, an extinction coefficient of 6990, a fat coefficient of 92.73 and an instability coefficient of 1.18 , The total average hydrophobicity of -0.023; in vitro has a strong ability to scavenge superoxide anion and antioxidant free radicals. Conclusion The isolated and purified heme peptide-iron has the ability to scavenge free radicals.