Yuye decoction (YYD) has been widely used as a folk Chinese herbal formula in clinical treatment of type 2 diabetes mellitus(T2DM) for many years. However, its mechanism is still unclear.The aim of this study was to explore the potential mechanism of YYD against T2DM initially by UHPLC-MS/MS combining with network pharmacology, molecular docking techniques and experimental validation.The main ingredients in the water extract of YYD were initially identified using UHPLC-MS/MS analysis. Combined with network pharmacology and molecular docking techniques, the YYD key compounds-core targets-key signaling pathways network was constructed and the binding activity of key components to core targets was validated. The T2DM rat model was induced by Streptozotocin combined with high glucose and high fat diets. The apoptosis cell model of mouse islet β-cell of Min6 was induced by high-glucose and palmitic acid. Histopathological and immunofluorescence satining were used to evaluate pancreatic islet β-cell function and apoptosis in rats. Min6 cell viability and apoptosis ratio were evaluated by CCK-8 and TUNEL staining. The predicted targets and pathways were validated by experiments in vitro and in vivo.The 56 compounds from YYD were identified by UHPLC-MS/MS. The potential targets of the above compounds were predicted by online compound target database, among of which 362 targets were associated with T2DM. Protein-protein interaction analysis identified the main targets such as SRC, MAPK1, PIK3R1, AKT1, HRAS and HSP90AA1, which were considered as the therapeutic targets of YYD on against T2DM. Functional enrichment analysis revealed that PI3K/AKT, FoxO and apoptosis signaling pathways were significantly enriched. Molecular docking results showed that compounds of monolinolein, neomangiferin, mangiferin, pelargonidin-3-O-glucoside and acacetin from YYD had high binding activities to PIK3R1, AKT1, Sirt1 and FoxO1. Therefore, PI3K/AKT1, Sirt1/FoxO1 and apoptotic signaling pathways were considered as predicted targets for experimental validation study. Animal experiments showed that YYD reduced blood glucose levels, improved pancreatic dysfunction and pancreatic islet β-cells apoptosis in T2DM rats which contributed to the activation of AKT1 and FoxO1 and their related signaling molecules. These results were confirmed in Min6 cell model induced by high-glucose and palmitic acid.In summary, this study systematically visualized the possible therapeutic effects and mechanisms of YYD on T2DM through the network pharmacology approach and experimental study. The results indicated that YYD could prevent pancreatic islet dysfunction and reverse islet of β-cells apoptosis possibly via PI3K/AKT1, Sirt1/FoxO1 signaling pathways.Copyright © 2023. Published by Elsevier B.V.