采用碱性蛋白酶水解酪蛋白,制备水解度为13.5%、IC50为45.23μg/mL的酪蛋白水解物,然后利用中性蛋白酶对水解物进行类蛋白反应修饰,并研究酶添加量、底物浓度、反应温度和时间对修饰反应的影响。结果表明,修饰反应体系中水解反应占优势,表现为游离氨基含量增加;酶添加量、底物浓度、反应时间对修饰反应的影响显著,而反应温度的影响不大;在低酶添加量、高底物浓度和短反应时间下,修饰反应体系的游离氨基的增加幅度减少,水解反应相对降低。制备6个不同反应程度的修饰产物,ACE抑制活性分析结果显示,修饰产物的IC50降至15.56~19.98μg/mL,表明中性蛋白酶催化的类蛋白反应修饰可以提高酪蛋白水解物的ACE抑制活性。

Casein hydrolysates that have the degree of hydrolysis of 13.5 % and IC50 value of 45.23μg/mL were prepared from casein with a protease Alcalase,and then modified by plastein reaction with another protease Neutrase.The effects of the addition level of Neutrase,the concentration of casein hydrolysates,temperature and time on the plastein reaction of casein hydrolysates were studied.The results indicated that hydrolysis reaction was the dominant one in the Plastein reaction system as the content of free amino groups in reaction system increased in all conditions.The addition level of Neutrase,the concentration of casein hydrolysates and reaction time all had significant impact on the plastein reaction of casein hydrolysates,while reaction temperature gave some influence.Under lower addition level of Neutrase,higher concentration of casein hydrolysates and shorter reaction time,the extent of increase in free amino groups in Plastein reaction system tended to decrease which implied that hydrolysis reaction were weakened relatively.Six modified casein hydrolysates with different reaction extents were prepared and their ACE inhibitory activities were determined.The results showed that the values of IC50 of the modified products were in range of 15.56 to 19.98 μg/mL,indicating that the ACE inhibitory activity of casein hydrolysates could be improved significantly by plastein reaction catalyzed by Neutrase.