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In this study; a spectrum–effect relationship analysis combined with a high-performance liquid chromatography–mass spectrometry (LC–MS) analysis was established to screen and identify active components that can inhibit thrombin and factor Xa (THR and FXa) in Salviae Miltiorrhizae Radix et Rhizoma–Chuanxiong Rhizoma (Danshen–Chuanxiong) herbal pair. Ten potential active compounds were predicted through a canonical correlation analysis (CCA), and eight of them were tentatively identified through an LC–MS analysis. Furthermore; the enzyme inhibitory activity of six available compounds; chlorogenic acid; Z-ligustilide; caffeic acid; ferulic acid; tanshinone I and tanshinone IIA; were tested to verify the feasibility of the method. Among them; chlorogenic acid was validated to possess a good THR inhibitory activity with IC50 of 185.08 µM. Tanshinone I and tanshinone IIA are potential FXa inhibitors with IC50 of 112.59 µM and 138.19 µM; respectively. Meanwhile; molecular docking results show that tanshinone I and tanshinone IIA; which both have binding energies of less than −7.0 kcal·mol−1; can interact with FXa by forming H-bonds with residues of SER214; GLY219 and GLN192. In short; the THR and FXa inhibitors in the Danshen–Chuanxiong herbal pair have been successfully characterized through a spectrum–effect relationship analysis and an LC–MS analysis.

Screening of Potential Thrombin and Factor Xa Inhibitors from the Danshen–Chuanxiong Herbal Pair through a Spectrum–Effect Relationship Analysis