Development and validation of a green UHPLC-MS/MS method for simultaneous determination of pyrrolizidine and tropane alkaloids in honey and pollen

Pyrrolizidine (PA) and tropane alkaloids (TA) are widely distributed phytotoxins considered by the European Food Safety Authority as hepatotoxic and neurotoxic agents respectively. The main source of exposure are plant-derived products such as teas, cereals, species, honey, and pollen. This study presents a rapid, simple, and green analytical method for the determination of 28 PAs and 2 TAs in honey and pollen matrices by ultra-high performance liquid chromatography coupled to tandem mass spectrometry. The methodology was validated in both matrices, demonstrating accuracy values ranging from 70 % to 130 %, relative standard deviations (RSDs) between 3 % and 23 %, and LOD and LOQ ranging from 0.6 to 2.7 µg Kg −1 and from 2.0 to 9.1 µg Kg −1 , respectively, in compliance with the criteria established by the European Union. The method has been applied to the analysis of 6 honey and 7 pollen samples. Among the honey samples, four of them showed a total concentration ranging from 6.3 to 12.3 µg Kg −1 . Pollen samples showed higher concentrations, ranging from 11.2 to 1654.1 µg Kg −1 , with one sample exceeding the maximum levels set by the European Union. Furthermore, the greenness, applicability and analytical performances of the developed method were assessed using evaluation metric tools. The results confirmed that the method is not only analytically robust and broadly applicable but also aligns with green chemistry principles. • A method for simultaneous quantification of 28 PAs and 2 TAs has been validated. • This method is appliable indistinctly to both matrices, honey and pollen. • Proposed method proven greener using GAPI, RAPI and AGREE metrics. • Pyrrolizidine alkaloid levels in one pollen sample found to exceed EU limits.