In this study, thrombin was immobilized with magnetic particles modified by glutaraldehyde. The changes in secondary structures of immobilized enzyme revealed an increment in conformational rigidity and stability, which can be reflected in temperature and pH stability as well as the tolerance of organic reagents. The optimal reutilization times of magnetic particle immobilized thrombin were 7 times, and the half-life of enzyme activity preserved at room temperature was 5 days, which was 2.5 times higher than that of free enzyme. Ligusticum chuanxiong and Anemarrhenae Rhizoma with high enzyme inhibitory activity were selected for primary screening, and six potential inhibitors of thrombin were identified by HPLC/MS. The results showed that three compounds in Anemarrhenae Rhizoma had better predictive thrombin inhibitory activity. Through the in vitro thrombin activity inhibition experiment, it was also verified that mangiferin and neo-mangiferin had an ideal thrombin activity inhibition effect, which was consistent with the results of molecular docking.

中文翻译：

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磁性纳米颗粒固定凝血酶配体捕获筛选天然产物中的凝血酶抑制剂

在本研究中，凝血酶被戊二醛修饰的磁性颗粒固定化。固定化酶二级结构的变化揭示了构象刚性和稳定性的增加，这可以反映在温度和pH稳定性以及有机试剂的耐受性上。磁粒固定化凝血酶的最佳重复利用次数为7次，室温保存酶活性半衰期为5 d，比游离酶提高了2.5倍。选取酶抑制活性较高的川芎和知母进行初步筛选，通过HPLC/MS鉴定出6种潜在的凝血酶抑制剂。结果表明，知母中的三种化合物具有较好的预测凝血酶抑制活性。通过体外凝血酶活性抑制实验，也验证了芒果苷和新芒果苷具有理想的凝血酶活性抑制效果，这与分子对接的结果一致。