Metabolomics combined with extraction optimization and biochemical assays revealed flavonoids and in vitro and in vivo antioxidant activity of Ponkan Chenpi.

Ponkan Chenpi (PC), derived from the dried and aged peel of ripe Ponkan citrus, is a traditional product with both medicinal and edible value. In this study, an untargeted metabolomics approach utilizing UPLC-QTOF-MS/MS was employed to characterize the metabolite profiles of PCs aged for 1, 3, 5, and 8 years (PC1, PC3, PC5, and PC8). A total of 1455 metabolites were characterized, with flavonoids exhibiting relatively high quantity and abundance. Clear differences in both metabolite profiles and metabolic pathways were observed among PCs of different aging periods. Furthermore, flavonoid extracts were prepared from the PCs, with the content of flavonoid compounds determined by HPLC as the response value. The optimal extraction parameters, determined by single-factor analysis and response surface methodology, were 65 % ethanol concentration, 50 min ultrasonic time, 65 °C ultrasonic temperature, and a 1:65 solid-liquid ratio. Subsequent free radical scavenging assays demonstrated that the PC5 extract exhibited superior antioxidant activity. Correlation analysis indicated that flavonoid monomers-hesperidin, nobiletin, tangeretin, and 5-demethylnobiletin were significantly correlated with antioxidant activity. Network pharmacology and molecular docking analyses revealed that proteins such as PPARγ, COX-2, and ALB were the core targets through which these flavonoids exert antioxidant effects. Finally, the in vivo efficacy of PC5was verified in a D-gal-induced oxidative damage mouse model, which demonstrated that PC5 extract significantly improve the hepatic oxidative stress state and alleviate tissue damage in mice. This work provides a theoretical foundation for the quality evaluation and high-value utilization of PC.