Magnetic nanoparticles immobilized thrombin ligand fishing to screen thrombin inhibitors in natural products

• Published in: Journal of pharmaceutical and biomedical analysis 2024-06, Vol.243, p.116110-116110, Article 116110
• Main Authors: Guo, Hui, Guo, Lili, Yu, Jianwei, Zhao, Fengju, Yang, Wei, Li, Jiaxin, Chen, Hanqi, Qian, Junqing
• Format: Article
• Language: English
• Subjects: Anticoagulants; Biological Products - pharmacology; Drugs, Chinese Herbal - chemistry; Enzymes, Immobilized - chemistry; Immobilized enzyme; Ligand fishing; Ligands; Magnetic nanoparticles; Magnetite Nanoparticles; Molecular docking; Molecular Docking Simulation; Thrombin; Thrombin inhibitors
• ISSN: 0731-7085; 1873-264X
• DOI: 10.1016/j.jpba.2024.116110

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. [Display omitted] •Preparation and characterization of thrombin immobilized on Fe3O4@SiO2-NH2.•Detection of enzymatic properties of immobilized thrombin.•Six thrombin inhibitors fished out with Traditional Chinese medicine extracts.•The thrombin inhibitory activity was verified by molecular docking technique.