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目的通过酶解法制备北冬虫夏草活性多肽并筛选出最佳酶解条件。方法分别用胃蛋白酶和胰蛋白酶对北冬虫夏草进行单酶酶解,然后先加胃蛋白酶后加胰蛋白酶进行双酶酶解。按时间,加酶量和底物浓度不同条件采用正交试验,通过茚三酮比色法检测水解度优化最佳酶解工艺,采用MTT法检测单酶酶解液和双酶酶解液对PC12细胞活性的影响。结果胃蛋白酶和胰蛋白酶水解度测定吸光度最大值分别为0.899和0.788,对应酶解时间、加酶量和底物浓度都是5h、2.5%和12%;双酶酶解在此条件下的吸光度值为0.875。酶解液对PC12细胞修复率影响最大值为139.59%,对应样品为胃蛋白酶酶解原液稀释10~(-6)倍。结论胃蛋白酶酶解效果高于胰蛋白酶和双酶。
OBJECTIVE To prepare the active polypeptide of Cordyceps militaris by enzymolysis method and select the optimal enzymatic hydrolysis conditions. Methods Pepsin and trypsin were respectively used for single-enzyme digestion of Cordyceps militaris, and then trypsin digestion was carried out by adding pepsin and trypsin first. According to different conditions of time, amount of enzyme and substrate concentration, orthogonal test was used to determine the optimal enzymatic hydrolysis degree by ninhydrin colorimetric method. The single and double enzymatic hydrolysis solutions were detected by MTT assay PC12 cell activity. Results The maximum absorbance values of pepsin and trypsin hydrolysis were 0.899 and 0.788, respectively. The corresponding enzymolysis time, enzyme dosage and substrate concentration were 5h, 2.5% and 12%, respectively. The absorbance The value is 0.875. The maximum effect of hydrolyzate on the recovery rate of PC12 cells was 139.59%. The corresponding sample was diluted 10 ~ (-6) times with pepsin solution. Conclusions Pepsin enzymatic hydrolysis is better than trypsin and double enzyme.