Impact of cooking on the fate and bioaccessibility of selenium compounds, including SeNPs, in spinach using stable isotope tracers, in vitro gastrointestinal digestion, and (HPLC)-ID-ICP-MS

This work presents the investigation of the fate of three selenium species, including selenium nanoparticles (SeNPs), in selenium-biofortified spinach during cultivation, cooking, and simulated human gastrointestinal digestion. The use of multiple selenium isotopic tracers, combined with ICP-MS methodologies, enabled the quantitative assessment of changes in both total selenium content and selenium speciation along the entire farm-to-digestion pathway. Spinach ( Spinacia oleracea ) plants were biofortified through foliar application of 76 SeMet, 77 Se(IV), and chitosan-stabilized selenium nanoparticles (Ch-SeNPs). Analysis by isotope dilution ICP-MS (ID-ICP-MS) revealed limited selenium leaching into the cooking water, indicating that most of the accumulated selenium remained in the edible tissue after boiling. HPLC-ID-ICP-MS demonstrated that thermal processing promoted the formation of inorganic selenium species derived from SeNPs. From a nutritional perspective, these changes are undesirable, as inorganic selenium species exhibit higher toxicity. Despite these transformations, boiling significantly increased selenium bioaccessibility in spinach biofortified with both 76 SeMet and 77 Se(IV). Furthermore, gastrointestinal digestion induced more pronounced changes in selenium speciation in raw spinach than in cooked spinach, characterized mainly by a decrease in selenomethionine (SeMet) and an increase in Se -methylselenocysteine (SeMeSeCys) derived from 76 SeMet. Inorganic selenium species generated during cooking were not detected in the bioaccessible fraction. Overall, the results highlight the complex transformations undergone by selenium species during food processing and digestion. The combination of multiple isotopic tracers with ID-ICP-MS and HPLC-ID-ICP-MS proved to be a powerful analytical strategy for simultaneously tracking and quantifying trace-element species transformations throughout the entire food chain.