Elucidating the Anti-Atherosclerotic Mechanisms of Sanzi San: An Integrated Approach Using Serum Pharmacochemistry, Network Pharmacology, and Pharmacokinetics

Abstract Background: Atherosclerosis (AS) serves as the primary pathological foundation for numerous cardiovascular diseases, such as thromboembolic conditions, coronary artery disease, and ischemic cardiac-cerebral vascular disorders. Sanzi San, a traditional Chinese medicinal formulation with a long-standing clinical reputation for efficacy against AS, has not yet had its pharmacological mechanisms fully elucidated. Objective: Our objective is to investigate the impact of Sanzi San on AS in ApoE −/− mice models and elucidate the pharmacological mechanism via an integrated approach of serum pharmacochemistry, network pharmacology, and pharmacokinetic analysis. Materials and Methods: The chemical constituents in Sanzi San and serum were identified using high-performance liquid chromatography-quadrupole/Orbitrap high-resolution mass spectrometry (HPLC-Q/Orbitrap HRMS). A network pharmacology approach, including protein–protein interaction (PPI) and gene ontology enrichment analyses, was then used to investigate the connection between these components and their anti-atherosclerotic activity. Finally, ultra-HPLC-tandem mass spectrometry was used to determine the formula’s pharmacokinetic parameters following oral administration. Results: Through a comparison of serum samples from control and treated groups, 41 prototype compounds originating from Sanzi San were identified. These constituents were classified into three major groups – terpenes, iridoids, and phenolic acids – and were regarded as the potential bioactive components due to their absorption into the bloodstream. Network pharmacology analysis indicated that Sanzi San directly influences blood vessels by regulating their diameter to achieve therapeutic effects. Furthermore, Sanzi San demonstrated therapeutic potential by specifically targeting natural killer (NK) cells within plaque tissue and adventitial cells in the aorta. In vivo validation in ApoE −/− mice showed that Sanzi San improved aortic lesions and regulated body weight. Pharmacokinetic studies also revealed that seven major components of Sanzi San were present in plasma, confirming its pharmacological effects on AS. Conclusion: This study provides evidence of Sanzi San’s clinical efficacy in treating AS. The integrated strategy combining serum pharmacochemistry, network pharmacology, and pharmacokinetics offers a powerful approach for uncovering the key pharmacological mechanisms of herbal formulations.