Abstract: The conditions for the synthesis of oxidized tea polyphenol (OTP) from green tea polyphenols (TP) were optimized, employing polyphenol oxidase (PPO) sourced from Huangguan pear as the catalyst and utilizing α-glucosidase inhibitory activity as the evaluation criterion. A comprehensive non-targeted metabolomics analysis was carried out using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), aiming to elucidate the distinct metabolic profiles prior to and following the oxidation process. The potential inhibition mechanism of metabolites on α-glucosidase was investigated by molecular docking. The results showed that the optimal reaction conditions were mass concentration of TP 0.3 mg/mL, reaction time 57 min, 30.5 ℃ and pH4.1, the inhibition rate of OTP on α-glucosidase was 91.85%. The non-targeted metabolomics analysis identified 1372 metabolites in the TP and OTP, of which 161 were classified as differential metabolites. Through principal component analysis (PCA) and cluster analysis, it was determined that, in comparison to TP, OTP exhibited an upregulation of 86 differential metabolites and a downregulation of 75 differential metabolites. A correlation analysis between the differential metabolites of tannins and α-glucosidase inhibitory activity revealed that theaflavins and theasinensins were positively correlated with α-glucosidase inhibitory activity. In addition, molecular docking revealed that these differential metabolites may occupy the potential catalytic active sites of α-glucosidase through hydrogen bonds and hydrophobic interactions, thus hindering the formation of complex between α-glucosidase and substrate. Overall, the enzymatic reaction conditions of OTP identified in this study facilitate the accumulation of theaflavins and theasinensins, while significantly enhancing the α-glucosidase inhibitory activity of OTP. This finding provides valuable insights for the development and utilization of OTP.