Brevetoxin metabolism and elimination in the Eastern oyster (Crassostrea virginica) after controlled exposures to Karenia brevis

Abstract

The metabolism and elimination of brevetoxins were examined in the Eastern oyster (Crassostrea virginica) following controlled exposures to Karenia brevis cultures in the laboratory. After a 2-day exposure period ( approximately 62 million cells/oyster), elimination of brevetoxins and their metabolites was monitored by using liquid chromatography/mass spectrometry (LC/MS). Composite toxin in oyster extracts was measured by in vitro assay (i.e. cytotoxicity, receptor binding, and ELISA). Of the parent algal toxins, PbTx-1 and PbTx-2 were not detectable by LC/MS in K. brevis-exposed oysters. PbTx-3 and PbTx-9, which are accumulated directly from K. brevis and through metabolic reduction of PbTx-2 in the oyster, were at levels initially (after exposure) of 0.74 and 0.49 microg equiv./g, respectively, and were eliminated largely within 2 weeks after dosing. PbTx-7 and PbTx-10, the reduced forms of PbTx-1, were non-detectable. Conjugative brevetoxin metabolites identified previously in field-exposed oysters were confirmed in the laboratory-exposed oysters. Cysteine conjugates of PbTx-1 and PbTx-2, and their sulfoxides, were in the highest abundance, as apparent in LC/MS ion traces, and were detectable for up to 6 months after dosing. Composite toxin measurements by in vitro assay also reflected persistence (up to 6 months) of brevetoxin residues in the oyster. Levels of cysteine conjugates, as determined by LC/MS, were well correlated with those of composite toxin, as measured by ELISA, throughout depuration. Composite toxin levels by cytotoxicity assay were well correlated with those by receptor binding assay. Cysteine-PbTx conjugates are useful LC/MS determinants of brevetoxin exposure and potential markers for composite toxin in the Eastern oyster.