In Vitro Metabolism of Azaspiracids 1–3 with a Hepatopancreatic Fraction from Blue Mussels (Mytilus edulis)

Azaspiracids (AZAs) are a group of biotoxins produced by the marine dinoflagellates Azadinium and Amphidoma spp. that can accumulate in shellfish and cause food poisoning in humans. Of the 60 AZAs identified, levels of AZA1, AZA2, and AZA3 are regulated in shellfish as a food safety measure based on occurrence and toxicity. Information about the metabolism of AZAs in shellfish is limited. Therefore, a fraction of blue mussel hepatopancreas was made to study the metabolism of AZA1–3 in vitro. A range of AZA metabolites were detected by liquid chromatography–high-resolution tandem mass spectrometry analysis, most notably the novel 22α-hydroxymethylAZAs AZA65 and AZA66, which were also detected in naturally contaminated mussels. These appear to be the first intermediates in the metabolic conversion of AZA1 and AZA2 to their corresponding 22α-carboxyAZAs (AZA17 and AZA19). α-Hydroxylation at C-23 was also a prominent metabolic pathway, producing AZA8, AZA12, and AZA5 as major metabolites of AZA1–3, respectively, and AZA67 and AZA68 as minor metabolites via double-hydroxylation of AZA1 and AZA2, but only low levels of 3β-hydroxylation were observed in this study. In vitro generation of algal toxin metabolites, such as AZA3, AZA5, AZA6, AZA8, AZA12, AZA17, AZA19, AZA65, and AZA66 that would otherwise have to be laboriously purified from shellfish, has the potential to be used for the production of standards for analytical and toxicological studies.