贻贝粉蛋白质酶解产物的动力学、表征和肽功能研究
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摘要
近十年的研究表明,许多食物蛋白和多肽除了具有较好的营养价值外,还表现出特定的生物活性。在酶法蛋白水解产物和发酵乳制品中含有生物活性多肽,它们也有可能在蛋白的胃肠道消化过程中产生。研究表明,通过胃肠道的消化吸收,贻贝蛋白具有抗氧化活性。
本论文研究了通过贻贝粉的蛋白提取物的水解获得相应蛋白和具有功能特性以及生物活性的多肽。
首先,依据蛋白的不同种类确定蛋白提取的分离方法。提取物没有功能活性,但是通过碱性蛋白酶的部分水解改善了其功能活性,同时还比较了各种蛋白酶对提取物水解过程的不同作用。水解动力学表明,贻贝蛋白提取物具有显著的酶特异性,而且酶和水解活性具有相关性。同时我们还研究了在特定的DH和pH下三种不同的酶系统,而且提取物的成分不受初始酶浓度,底物浓度和温度的影响。
因此,不管设定的参数值是多少,DH的发展变化都能预测控制目标蛋白多肽混合物的产生。为了模仿反应的水解动力学,建立了一个根据米氏函数反应曲线的模型。这样,水解中无法显现的各种现象(酶的抑制与失活,底物的改变)都能够明确。
A growing body of scientific evidence in the past decade has revealed that many food proteins and peptides exhibit specific biological activities in addition to their established nutritional value. Bioactive peptides have been found in enzymatic protein hydrolysates and fermented dairy products, but they can also be released during gastrointestinal digestion of proteins. Many studies have demonstrated that mussel protein have potent antioxidant activity. Isolated from gastrointestinal digest.
This thesis made it possible to study obtaining protein content, functional properties or biologically active peptides with through the development of processes for the preparation and the hydrolysis of protein isolates resulting from mussel powder.
First, a method of preparation of protein isolates being different by the type of proteins was developed. The isolates do not have good functional properties but their partial hydrolysis by Alcalase 2.4L improves some of them. Then, the hydrolytic action of commercial proteases (Alcalase 2.4L, Pronase SG, Neutrase 0.8L, Lypaine 6500) on the isolate of different process were compared. It was shown that the kinetics of hydrolysis is significantly influenced by the specificity of the enzyme and there is a relation enzyme/degree of hydrolysis (DH)/targeted activity. Lastly, we showed for three different enzyme/substrate systems that at given DH and pH, the composition of the hydrolysates is independent of the initial enzyme and substrate concentrations and of the temperature.
Thus, the prediction of the temporal evolution of the DH, whatever the values of precedent parameters, allows to control the generation of a peptide mixture with targeted properties. A model based on the reaction pathway of Michaelis-Menten was then built in order to simulate the hydrolysis kinetics in batch reactor. For that, limiting phenomena implied in the hydrolysis (inhibition or inactivation of the enzyme, modification of the substrate) were highlighted.
本论文研究了通过贻贝粉的蛋白提取物的水解获得相应蛋白和具有功能特性以及生物活性的多肽。
首先,依据蛋白的不同种类确定蛋白提取的分离方法。提取物没有功能活性,但是通过碱性蛋白酶的部分水解改善了其功能活性,同时还比较了各种蛋白酶对提取物水解过程的不同作用。水解动力学表明,贻贝蛋白提取物具有显著的酶特异性,而且酶和水解活性具有相关性。同时我们还研究了在特定的DH和pH下三种不同的酶系统,而且提取物的成分不受初始酶浓度,底物浓度和温度的影响。
因此,不管设定的参数值是多少,DH的发展变化都能预测控制目标蛋白多肽混合物的产生。为了模仿反应的水解动力学,建立了一个根据米氏函数反应曲线的模型。这样,水解中无法显现的各种现象(酶的抑制与失活,底物的改变)都能够明确。
A growing body of scientific evidence in the past decade has revealed that many food proteins and peptides exhibit specific biological activities in addition to their established nutritional value. Bioactive peptides have been found in enzymatic protein hydrolysates and fermented dairy products, but they can also be released during gastrointestinal digestion of proteins. Many studies have demonstrated that mussel protein have potent antioxidant activity. Isolated from gastrointestinal digest.
This thesis made it possible to study obtaining protein content, functional properties or biologically active peptides with through the development of processes for the preparation and the hydrolysis of protein isolates resulting from mussel powder.
First, a method of preparation of protein isolates being different by the type of proteins was developed. The isolates do not have good functional properties but their partial hydrolysis by Alcalase 2.4L improves some of them. Then, the hydrolytic action of commercial proteases (Alcalase 2.4L, Pronase SG, Neutrase 0.8L, Lypaine 6500) on the isolate of different process were compared. It was shown that the kinetics of hydrolysis is significantly influenced by the specificity of the enzyme and there is a relation enzyme/degree of hydrolysis (DH)/targeted activity. Lastly, we showed for three different enzyme/substrate systems that at given DH and pH, the composition of the hydrolysates is independent of the initial enzyme and substrate concentrations and of the temperature.
Thus, the prediction of the temporal evolution of the DH, whatever the values of precedent parameters, allows to control the generation of a peptide mixture with targeted properties. A model based on the reaction pathway of Michaelis-Menten was then built in order to simulate the hydrolysis kinetics in batch reactor. For that, limiting phenomena implied in the hydrolysis (inhibition or inactivation of the enzyme, modification of the substrate) were highlighted.
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