In vitro identification and in vivo metabolic profiling of chemical constituents in Moringa oleifera seeds

<i>Moringa oleifera</i> seeds (MOS), recognized as a nutritionally valuable food, are rich in diverse bioactive compounds and widely utilized for disease prevention and adjunctive therapy. However, their in vitro chemical constituents and in vivo blood-absorbed/metabolized components remain underexplored. This study pioneered the application of UPLC-Q-Exactive Orbitrap-MS to characterize MOS-derived compounds in vitro and in vivo, concurrently elucidating major constituents' fragmentation pathways via mass spectrometry. In vitro, 81 chemical components of MOS were preliminarily characterized. 38 blood-borne components, comprising 11 prototypes and 27 metabolites, were identified in serum samples from MOS-treated rats. A validated LC-MS/MS method was developed to quantify the pharmacokinetics of key bioactive compounds (6-gingerol, vanillin, and eugenol) in rat plasma. These findings establish a theoretical foundation for clarifying MOS composition, evaluating its dietary applications, and guiding further exploration of its pharmacological potential.