Determination of lipophilic marine toxins in mussels. Quantificationand confirmation criteria using high resolution mass spectrometry

Abstract

tA multitoxin method has been developed for quantification and confirmation of lipophilic marine biotox-ins in mussels by liquid chromatography coupled to high resolution mass spectrometry (HRMS), using anOrbitrap-Exactive HCD mass spectrometer. Okadaic acid (OA), yessotoxin, azaspiracid-1, gymnodimine,13-desmethyl spirolide C, pectenotoxin-2 and Brevetoxin B were analyzed as representative compoundsof each lipophilic toxin group. HRMS identification and confirmation criteria were established. Fragmentand isotope ions and ion ratios were studied and evaluated for confirmation purpose. In depth character-ization of full scan and fragmentation spectrum of the main toxins were carried out. Accuracy (truenessand precision), linearity, calibration curve check, limit of quantification (LOQ) and specificity were theparameters established for the method validation. The validation was performed at 0.5 times the currentEuropean Union permitted levels. The method performed very well for the parameters investigated. Thetrueness, expressed as recovery, ranged from 80% to 94%, the precision, expressed as intralaboratoryreproducibility, ranged from 5% to 22% and the LOQs range from 0.9 to 4.8 pg on column. Uncertaintyof the method was also estimated for OA, using a certified reference material. A top-down approachconsidering two main contributions: those arising from the trueness studies and those coming from theprecision's determination, was used. An overall expanded uncertainty of 38% was obtained.