牦牛血抗氧化肽制备方法对比及分离纯化研究

Preparation Comparison of Yak Blood Antioxidant Peptides and Its Purification

摘要:

为研究牦牛血抗氧化肽最佳制备方法，以牦牛血为原料，分别采用枯草芽孢杆菌发酵法、碱性蛋白酶解法、菌酶联合法制备3种牦牛血抗氧化肽，依次简写为BF、AP、BA。应用羟基自由基（·OH）、超氧阴离子自由基（·O2-）清除能力，脂质过氧化抑制能力、还原力4种体外抗氧化实验，对比3种牦牛血抗氧化肽的抗氧化活性。以超滤、凝胶层析法对制备的较高活性产物进行分离。结果表明：3种抗氧化肽活性大小为BA>BF>AP，BA有最大的·O2-清除能力、脂质过氧化抑制能力及还原力，表明制备牦牛血抗氧化肽的最佳制备方法为菌酶联合法；超滤分离菌酶联合制备产物获得分子量<5 kD的抗氧化肽活性高于分子量>10 kDa、介于5~10 kDa的抗氧化肽，对·OH的IC50值为1.62 mg/mL；该组分经凝胶层析分离后得到4个组分，其最高活性组分Ⅰ对·OH的IC50值达到0.72 mg/mL。

Abstract:

To research the best preparation of Yak blood antioxidant peptides，Yak blood was used to prepare three kinds of antioxidant peptides（abbreviated as BF，AB，BA） which obtained through fermentation with Bacillus subtilis，enzymatic hydrolysis with alkaline protease and combination of bacteria and enzyme respectively. Four kinds of antioxidant activities evaluation methods in vitro including scavenging of hydroxyl radical and superoxide anion，anti-lipid peroxidation，reducing power were presented to evaluate the antioxidant activities of Yak blood antioxidant peptides. Ultrafiltration and Gel chromatography were performed to purify the higher activity product. As results：the order of antioxidant activities of three antioxidant peptides was BA>BF>AP. BA had the most antioxidant activities of scavenging of superoxide anion，reducing power，anti-lipid peroxidation which indicated that the optimum preparation method was the combination of bacteria and enzyme.The antioxidant activity of antioxidant peptides with molecular weight <5 kDa which obtained through ultrafiltration was better compared to other molecular weights like >10 kDa and 5~10 kDa. Its IC50 of scaveng hydroxyl radical was 1.62 mg/mL. Then it was purified into four components by gel chromatography，in which component Ⅰwith the highest scavenging rate on hydroxyl radical. Its IC50 of scaveng hydroxyl radical was 0.52 mg/mL.