Precise Identification of Inhibitors for Coagulation Reactions from Complex Extracts through Monitoring of Biological Aggregates Combined with a Targeted Fishing Technique

Biological aggregates play a crucial role in the pathogenesis of thrombotic diseases, especially thrombin-induced biological aggregates. Therefore, the efficient discovery of thrombin inhibitors is of great significance for the prevention and treatment of thrombotic diseases. In this study, the aggregation precursor protein fluorescent probe was successfully prepared for monitoring the production of biological aggregates induced by thrombin. In this program, the aggregation degree of biomolecules can be quickly monitored through a fluorescence sensing technology. To facilitate the modulation of the biological aggregation process, the application of this advanced fluorescence sensing technology was utilized for the screening of thrombin inhibitors, which are pivotal regulatory molecules in biological aggregation. In addition, by combining the target fishing technique, an integrated model for rapid screening of potential inhibitors in complex extracts was further established. This model not only swiftly detects the presence of inhibitory components within complex systems but also precisely captures and identifies active monomers. After positive drug validation of the screening model, three active monomers, namely, ginkgetin, isoginkgetin, and bilobetin, were accurately screened from 30 natural products. These results highlighted the immense potential of the proposed approach for screening active ingredients from a wide range of natural products.