UPLC-QE-Orbitrap-MS profiling under pathological conditions integrated with symptom-oriented network pharmacology to elucidate the therapeutic basis of Dihuang Yinzi in ischemic stroke

Objective Dihuang Yinzi (DHYZ) is a classic traditional Chinese medicine formula widely used in the treatment of stroke-related disorders. However, the relationship between its active constituents under pathological conditions and specific symptoms of ischemic stroke (IS) has not been systematically elucidated. This study aimed to investigate the material basis and mechanism of DHYZ by integrating pathological condition-based plasma and brain exposure profiling with symptom-oriented network pharmacology. Methods Ultra-performance liquid chromatography coupled with Q-Exactive Orbitrap mass spectrometry (UPLC-QE-Orbitrap-MS) was used to identify DHYZ constituents and to profile its absorbed components in both plasma and brain tissue of transient middle cerebral artery occlusion (tMCAO) rats under pathological conditions; symptom-oriented network pharmacology was applied to map brain-penetrating components to stroke-related symptom targets; and molecular docking followed by molecular dynamics simulations was performed to evaluate the binding potential and stability of representative components with core targets. Results A total of 179 compounds were identified in DHYZ. Among them, 64 components were detected in plasma and 28 components in brain tissue of tMCAO rats, with terpenoids (e.g., ursolic acid, catalpol) considered as the main bioactive components. A total of 239 intersection targets, 6 core targets (IL1B, INS, AKT1, IL6, BCL2, TP53), and key pathways (AGE-RAGE, TNF) were screened. Molecular docking suggested that seven representative components (e.g., ursolic acid, catalpol) exhibited favorable binding potential with core targets including IL1B and AKT1 via hydrogen bonds and hydrophobic interactions. Molecular dynamics simulations further supported the stability and activity of the catalpol-target complex. Conclusion This study integrates pathological condition-based plasma and brain exposure profiling with symptom-oriented network pharmacology to investigate the mechanism of DHYZ against ischemic stroke. Among 179 identified constituents, 48 prototypes were detected in plasma and 14 in brain tissue (10 in the infarcted hemisphere). Symptom-target mapping linked IL1B/AKT1/IL6 to motor-speech deficits and INS to amnesia, converging on AGE-RAGE and TNF pathways. This integrated approach-combining pathological-state exposure analysis, symptom-anchored target mapping, and computational validation-provides a more specific mechanistic framework for understanding DHYZ's neuroprotective effects and offers a methodological reference for future TCM formula research from a pathology-anchored, symptom-centered perspective.