双酶法制备薏米多肽工艺及其体外抗氧化活性研究
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摘要
以薏米蛋白液为原料,采用双酶协同酶解的方法制备薏米多肽。以水解度为评价指标,在单因素试验的基础上,运用正交试验设计优化薏米多肽的制备工艺;利用DPPH自由基、ABTS自由基、羟基自由基(OH·)清除率和铁氰化钾还原法评价了薏米多肽的抗氧化性。结果表明:薏米多肽的最佳制备条件为:选用胰蛋白酶与碱性蛋白酶双酶协同酶解(酶活比6:4),酶解时间3 h、酶解温度50℃、酶解pH9.0、底物浓度3%、加酶量1000 U/g,在此条件下,薏米蛋白液的水解度为18.05%。薏米多肽具有较强的抗氧化活性,随着质量浓度的增大,其对DPPH自由基、ABTS自由基、羟基自由基的清除能力和还原能力均显著增加,呈现出明显的剂量依赖效应。薏米多肽对3种自由基清除活性的半数抑制浓度(IC_50)分别为8.39、0.22 mg/mL和3.33 mg/mL。
In order to prepare coix seed peptides, the coix seed protein was hydrolyzed with double enzyme at the same time in aqueous solution. To screen the optimal conditions of coix seed peptides preparation, an orthogonal design combined with single factor experiments was taken, and the indicators were based on degree of hydrolysis. Four different methods including DPPH radical scavenging, ABTS radical scavenging, hydroxyl radical scavenging and reducing power were used for evaluating the antioxidant activities of coix seed peptides. The optimal hydrolysis conditions were achieved by using pancreatin and alcalase simultaneously(enzymatic activity ratio=6∶4) to hydrolyze coix seed protein for 3 hours at substrate concentration 3%, pH 9.0, 50 ℃ and enzyme dosage 1000 U/g. Under these conditions, the degree of hydrolysis was up to 18.05%. The antioxidant activity assays showed that coix seed peptides presented strong antioxidant activity. Moreover, with the increase of the concentration of coix seed peptides, the scavenging rate of DPPH, ABTS, hydroxyl radical and reducing power increased significantly in a dose-dependent manner. The half inhibitory concentration(IC_50) for the three radical scavenging activity were 8.39, 0.22 mg/mL and 3.33 mg/mL, respectively.
In order to prepare coix seed peptides, the coix seed protein was hydrolyzed with double enzyme at the same time in aqueous solution. To screen the optimal conditions of coix seed peptides preparation, an orthogonal design combined with single factor experiments was taken, and the indicators were based on degree of hydrolysis. Four different methods including DPPH radical scavenging, ABTS radical scavenging, hydroxyl radical scavenging and reducing power were used for evaluating the antioxidant activities of coix seed peptides. The optimal hydrolysis conditions were achieved by using pancreatin and alcalase simultaneously(enzymatic activity ratio=6∶4) to hydrolyze coix seed protein for 3 hours at substrate concentration 3%, pH 9.0, 50 ℃ and enzyme dosage 1000 U/g. Under these conditions, the degree of hydrolysis was up to 18.05%. The antioxidant activity assays showed that coix seed peptides presented strong antioxidant activity. Moreover, with the increase of the concentration of coix seed peptides, the scavenging rate of DPPH, ABTS, hydroxyl radical and reducing power increased significantly in a dose-dependent manner. The half inhibitory concentration(IC_50) for the three radical scavenging activity were 8.39, 0.22 mg/mL and 3.33 mg/mL, respectively.
引文
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[3]职爱民,冯定远.生物活性肽概述及其在动物营养上的应用[J].饲料与畜牧,2008,(7):10-13.
[4]Liu M,Wang Y,Liu Y,et al.Bioactive peptides derived from traditional Chinese medicine and traditional Chinese food:A review[J].Food Research International,2016,89:63-73.
[5]Lorenzo JM,Munekata PES,Gómez B,et al.Bioactive peptides as natural antioxidants in food products-Areview[J].Trends in Food Science&Technology,2018,79:136-147.
[6]邹远东.酶法多肽:人类健康卫士[M].北京:知识产权出版社,2015:110-111.
[7]Zhu F.Coix:Chemical composition and health effects[J].Trends in Food Science&Technology,2017,61:160-175.
[8]韩丽丽.薏米蛋白酶法制备及产品开发[D].南宁:广西大学,2017.
[9]杨文博,张英华.蛋白质水解度的测定方法研究[J].中国调味品,2014,(3):88-90.
[10]董红艳,张守文.薏米蛋白的酶水解工艺优化[J].农产品加工(学刊),2013,(5):46-50.
[11]蔡俊,陈季旺,王茹,等.多肽体外抗氧化活性测定方法的比较[J].食品科学,2016,37(11):52-57.
[12]蒲月华,邓旗,杨萍,等.珍珠贝多肽体外抗氧化活性的研究[J].食品科技,2016,(11):124-128.
[13]荣建华,李小定,谢笔钧.大豆肽体外抗氧化效果的研究[J].食品科学,2002,23(11):118-120.
[14]王霞,苏秀榕,丁进锋,等.响应面法优化双酶水解黄鳍金枪鱼胰脏的工艺研究[J].现代食品科技,2010,26(11):1229-1233.
[15]熊柳,赵云霞,孙庆杰.双酶法制备花生抗氧化肽[J].食品科学,2013,34(3):207-211.
[16]张根生,丁琬莹,王月,等.响应面法优化双酶酶解酪蛋白工艺[J].食品科学,2013,34(21):237-241.
[17]丁树慧,王立梅,齐斌,等.双酶分步水解低值海洋鱼粉制备低聚肽的研究[J].食品研究与开发,2017,38(24):42-47.
[18]吴悠,曹冲,杨玲,等.洋葱蛋白及多肽体外抗氧化活性研究[J].食品科技,2017,(10):235-238.
[19]李丽,阮金兰,钱伟亮,等.多指标评价核桃蛋白及多肽的抗氧化活性[J].食品研究与开发,2017,38(3):1-4.