Immobilization of thrombin on magnetic layered double hydroxide ZIF-8 composite materials and establishment of a thrombin inhibitor screening system

In this study, magnetic Fe₃O₄@ZnCr-LDH/ZIF-8 composites were fabricated by co-precipitation and in situ growth, combining the merits of LDHs and ZIF-8 to address Fe₃O₄ particle aggregation and limited enzyme loading capacity. Thrombin was efficiently immobilized within this material for rapid thrombin inhibitors screening. The composites were characterized by TEM, FTIR and other techniques, verifying successful synthesis with good paramagnetic property and hierarchical pores. Compared to free enzymes, the immobilized enzyme had better temperature and pH tolerance, retaining good activity after six cycles. Two potential inhibitors, cryptotanshinone and tanshinone I, were isolated from Salvia miltiorrhiza via combining immobilized enzyme with LC-MS. Molecular docking validated that both compounds bound to thrombin through hydrogen bonds and hydrophobic interactions, and cryptotanshinone exhibited a more favorable binding energy (-7.41 kcal mol^-1) than tanshinone I (-6.52 kcal mol^-1). This strategy provides a new pathway to rapidly screen bioactive natural compounds using functional magnetic nanomaterials.