| Summary: | The present work is focused on the development of an analytical platform to elucidate the metabolic pathway of PTSO from onion, an organosulfur compound well-known for its functional and technological properties and its potential application in animal and human nutrition. This analytical platform consisted of the use of gas chromatography–mass spectrometry (GC-MS) and ultra-high performance liquid chromatography quadrupole with time-of-flight MS (UHPLC-Q-TOF-MS) in order to monitor volatile and non-volatile compounds derived from the PTSO. For the extraction of the compounds of interest, two different sample treatments were developed: liquid–liquid extraction (LLE) and salting-out assisted liquid–liquid extraction (SALLE) for GC–MS and UHPLC-Q-TOF-MS analysis, respectively. Once the analytical platform was optimised and validated, an in vivo study was planned to elucidate PTSO metabolisation, revealing the presence of dipropyl disulfide (DPDS) in liver samples with concentrations between 0.11 and 0.61 µg g<sup>−1</sup>. The DPDS maximum concentration in the liver was observed at 0.5 h after the intake. DPDS was also present in all plasma samples with concentrations between 2.1 and 2.4 µg mL<sup>−1</sup>. In regard to PTSO, it was only found in plasma at times above 5 h (0.18 µg mL<sup>−1</sup>). Both PTSO and DPDS were excreted via urine 24 h after ingestion.
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