Preparation of Polypeptides from Hemp by Two-step Enzymatic Hydrolysis with Complex Enzymes and Its Antioxidant Properties
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摘要: 汉麻籽由于其营养丰富，且不含有胰蛋白酶抑制剂等抗营养因子，近年来成为食品、医药等领域的研究热点。本研究以汉麻分离蛋白为原料，采用碱性蛋白酶和木瓜蛋白酶复合酶分步酶解的方式对其进行酶解，通过响应面优化酶解参数，制取汉麻多肽。并采用分光光度计测定其DPPH自由基清除率与铁还原能力，对汉麻多肽抗氧化能力进行评价。结果表明，采用先碱性蛋白酶后木瓜蛋白酶的酶解方式，制得的汉麻多肽含量最高。酶解最优条件为：第一步碱性蛋白酶水解：底物浓度为5%，酶解时间2 h，pH为8.5，酶解温度为54℃，加酶量为10100 U/g；第二步木瓜蛋白酶水解：pH为6.5，温度为50℃，加酶量为5000 U/g，酶解时间1.5 h，最终获得汉麻多肽混合物的水解度为24.48%，肽含量为8.48 mg/mL；汉麻多肽的DPPH自由基清除率为82.32%，与汉麻分离蛋白相比具有较强的铁还原能力，表明汉麻多肽具备较强的抗氧化能力。本研究为汉麻多肽的开发利用提供理论基础。Abstract: Since hemp seeds because of rich in nutrients and do not contain anti-nutritional factors such as trypsin inhibitors have become a research hotspot in the fields of food and medicine in recent years. In this study,hemp protein isolate was enzymolysis step by step with alkaline protease and papain complex enzyme,and the hemp polypeptide was prepared by response surface. The DPPH free radical scavenging rate and iron reducing capacity were measured by spectrophotometer,and the antioxidant capacity of hemp polypeptide was evaluated. The results showed that the contents of hemp polypeptide was highest obtained by enzymatic hydrolysis of alkaline protease followed by papain. The optimal conditions of enzymatic hydrolysis were as follows:The first step was alkaline protease hydrolysis:The substrate concentration was 5%,the time of enzymatic hydrolysis was 2 h,the pH was 8.5,the temperature of enzymatic hydrolysis was 54 ℃,the amount of enzyme added was 10100 U/g.The second step was papain hydrolysis. The pH was 6.5,the temperature was 50 ℃,the amount of enzyme added was 5000 U/g,the enzymolysis time was 1.5 h,the degree of hydrolysis of hemp peptide mixture was 24.48%,and the contents of peptide was 8.48 mg/mL.The DPPH free radical scavenging rate of hemp polypeptide was 82.32%,the hemp polypeptide had stronger iron reducing ability than hemp protein isolate,which indicated that hemp poly peptide had strong antioxidant ability. This study would provide a theoretical basis for the development and utilization of hemp protein poly peptide.
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Keywords:
- hemp protein isolate /
- alkaline protease /
- papain /
- step enzymolysis /
- polypeptide /
- antioxidant ability
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