核桃饼粕蛋白提取、多肽制备条件优化及其酶解液的抗氧化性研究

本研究以去除多酚的核桃粕为原料,利用均匀试验优化核桃粕碱溶性蛋白提取条件,利用二次通用组合旋转设计优化双酶法制备核桃粕多肽制备条件,结合项目组前期建立的模拟胃肠道消化体系及模拟胃肠道消化体系+混合乳酸菌两种体系,制备核桃粕蛋白酶解液,采用体外实验对比三种体系制备核桃粕蛋白酶解液的抗氧化活性。核桃粕蛋白质最佳提取条件为:提取溶液的pH11、料液比为1∶21(g/mL)、提取温度65℃、提取时间70 min,提取2次,在此条件下提取液蛋白含量为221.6233 mg/g。双酶法制备核桃粕多肽工艺条件分为两个阶段,第一阶段胃蛋白酶酶解阶段,其最佳条件为:底物质量浓度为1 g/100 mL、pH2.4、加酶量为133.53 U/g、温度为40℃、时间139 min,在此条件下酶解,核桃粕多肽酶解液水解度25.90%;第二阶段胰蛋白酶酶解阶段,其最佳条件为:加酶量800 U/g、温度20℃、pH为6、酶解时间240 min,在此条件下酶解,核桃粕多肽酶解液的水解度为35.57%。三种酶解液的总抗氧化能力VC当量值分别为:双酶酶解液0.0516 mg/mL,体外模拟胃肠道消化酶解液0.0634 mg/mL、体外模拟胃肠道消化+混合乳酸菌酶解液0.0411 mg/mL,用双酶、体外模拟胃肠道消化、体外模拟胃肠道消化+混合乳酸菌三种体系制备的核桃粕蛋白酶解液均具有抗氧化性活性,而双酶法制备的核桃粕酶解液具有较强的抗氧化活性。

Study on Protein Extraction,Polypeptide Preparation and Antioxidant Activity of Walnut Pulp Hydrolysate

In this study,walnut pulp after polyphenol removal was used as raw material,the uniform test was used to optimize hydrolysis conditions of alkali soluble protein extraction,the secondary universal combined rotation design was used to optimize preparation conditions of walnut pulp polypeptides by double enzymes. Simulated gastrointestinal digestive system and simulated gastrointestinal digestive system+mixed lactobacillus system were established by project team at the early stage,which were combined to prepare the proteolytic solution of walnut pulp. The antioxidant activity of alkali solution of walnut pulp prepared by three systems was compared in vitro. The optimal extraction conditions of walnut pulp protein were as follows:pH11,solid-liquid ratio 1:21 (g/mL),water bath extraction at 65 ℃ for 70 min,extraction twice,under this condition the protein content of the extract was 221.6233 mg/g. The process conditions for the preparation of walnut pulp polypeptide by double enzyme method were divided into two stages. The first stage was pepsin enzymolysis,and the optimal conditions were:substrate mass concentration 1 g/100 mL,pH2.4,adding pepsin enzyme amount 133.53 U/g,temperature 40 ℃,and enzymolysis time 139 min. Under this condition,the degree of enzymolysis was 25.90%. Trypsin digestion phase,the second stage,the best conditions were:pH6,adding plus enzyme amount 800 U/g,temperature 20 ℃,and enzymolysis time 240 min. Under this condition,the degree of enzymolysis was 35.57%. Three kinds of total antioxidant capacity of enzymolysis liquid V_C equivalent value were:Enzymolysis solution preparated by double enzymes in vitro 0.0516 mg/mL,enzymolysis solution preparated by gastrointestinal digestion in vitro 0.0634 mg/mL,enzymolysis solution preparated by gastrointestinal digestion in vitro+mixed lactic acid bacteria in gut 0.0411 mg/mL. These three enzymolysis solutiones had antioxidant activity. In the meantime,enzymolysis solution preparated by double enzymes had strong antioxidant activity.