A magnetic nanoparticles-based ligand fishing method coupled with parallel artificial membrane permeability assay for rapid discovery and analysis of GSNO reductase inhibitors with good membrane permeability from Salvia miltiorrhiza Bunge.

Salvia miltiorrhiza Bunge (Danshen) is a traditional Chinese medicinal herb that has long been used for its cardiovascular and antioxidant benefits and has a potential role in the treatment of hypertensive disorders associated with pregnancy, such as pre-eclampsia. S-Nitrosoglutathione reductase (GSNOR) has emerged as a promising therapeutic target due to its role in regulating nitric oxide bioavailability and vascular function. Rapid screening and characterization of GSNOR inhibitors with favorable membrane permeability from Danshen by developing a novel magnetic nanoparticle-based ligand fishing method combined with a parallel artificial membrane permeability assay. Functionalized magnetic nanoparticles were synthesized and characterized using transmission electron microscopy, Fourier transform infrared spectroscopy, X-ray diffraction, and thermogravimetric analysis. GSNOR was immobilized on these nanoparticles, and the immobilization yield and enzyme activity were evaluated. The method was used to screen 17 compounds from Danshen extract, with 9 compounds showing significant binding affinity to GSNOR. Molecular docking and in vitro inhibitory activity assays were performed to validate the binding mechanisms and inhibitory effects of the identified compounds. Molecular docking revealed that these compounds form multiple hydrophobic interactions and hydrogen bonds with GSNOR, effectively inhibiting its activity. Tanshinone I and Tanshinone IIA exhibited the highest enrichment (19.7- and 20.0-fold, respectively) and potent inhibitory activity on GSNOR, with IC50 values of 19.47 μM and 17.99 μM. This study not only identified GSNOR inhibitors from Danshen but also established an efficient strategy for screening bioactive compounds from herbal medicines.