Phytochemical characterization and pharmacological mechanisms of Huazhuo Sanjie Chubi Decoction in treating gouty arthritis: a multivariant approach

Ethnopharmacological relevance: Huazhuo Sanjie Chubi Decoction (HSCD), a Chinese herbal formula, is traditionally used for the treatment of spleen deficiency with dampness accumulation and is commonly used to treat gouty arthritis (GA). However, the potential active compounds and mechanisms of HSCD remain unclear. Aim of the study: To elucidate the key bioactive compounds and pharmacological mechanisms of HSCD in treating GA. Materials and methods: The chemical compounds in HSCD were qualitatively and quantitatively analyzed using ultraperformance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). Network pharmacology and molecular docking were employed to identify key active compounds and associated molecular pathways. Monosodium urate (MSU)-induced RAW264.7 macrophages and GA rat model were used to explore the potential therapeutic effects and mechanisms of HSCD in treating GA. Results: UPLC-MS/MS identified 184 compounds in HSCD, with 28 key compounds quantified. Network pharmacology revealed that verbenalin, limonin, and quercitrin are strongly associated with the molecular mechanisms of HSCD in treating GA via the PI3K-AKT signaling pathway. These compounds exhibited strong binding affinity to PI3K and AKT proteins. In RAW264.7 cells, HSCD and the three identified compounds dose-dependently reduced inflammation by inhibiting nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS). They also downregulated both the PI3K-AKT and apoptosis signaling pathways. In rats, HSCD exerted therapeutic effects against acute GA by alleviating swelling and pathological damage to the ankle joints. Moreover, the molecular mechanisms in vivo were confirmed to be associated with the PI3K-AKT and apoptosis signaling pathways. Conclusion: This study employed a multivariant approach to demonstrate the main bioactive compounds and molecular mechanisms of HSCD in treating GA, thereby supporting its traditional use.