Cyclic imines evaluation in European commercial shellfish samples

Cyclic imines constitute a quite recently discovered group of marine biotoxins that act on neural receptors and that bioaccumulate in seafood. They are grouped together due to the imino group functioning as their common pharmacore, responsible for acute neurotoxicity in mice. Cyclic imines have not been linked yet to human poisoning and are not regulated in Europe, although the EFSA requires more data to perform conclusive risk assessment for consumers. Spirolides (SPXs) are produced by the dinoflagellate Alexandrium ostenfeldii, gymnodimines (GYMs) are also produced by A. ostenfeldii and by Karenia selliformis. The dinoflagellate Vulcanodinium rugosum produces pinnatoxins (PnTXs). In addition, not all cyclic imines are equally potent: SPX-1 showed about 300 fold more activity than GYM-A on equimolar basis in a in vivo study about neuromuscular excitability in mice. Oral toxicity of SPXs is much lower (10-100 times less toxic orally, depending on the toxin and how the toxins are administered). In contrast to spirolides, PnTXs have proven to be almost as toxic via oral dosing as they are by i.p. injection to mice. Levels of toxicity of spirolide C and pinnatoxin E+F in feed were 500 and 60 (LD50, mice, µg/kg), respectively, which is more relevant to protect consumers. Several commercial samples from eight different countries (Italy, Portugal, Slovenia, Spain, Ireland, Norway, Netherlands and Denmark) were obtained over 2 years. Emerging cyclic imine concentrations in all the samples were analysed on a LC-3000QTRAP and LC-HRMS QExactive mass spectrometer. All this data will be used in an European risk evaluation.