Selective liquid chromatographic determination of trace domoic acid in seawater and phytoplankton: improvement using the ophthaldialdehyde/9-fluorenylmethylchloroformate derivatization
Limnol. Oceanogr. Methods 11:327-336 (2013) | DOI: 10.4319/lom.2013.11.327
ABSTRACT: Domoic acid (DA), a toxin produced worldwide by some species of the genus Pseudo-nitzschia, is responsible for contamination of marine molluscs, mammals, birds, and for human intoxication, and when detected in high levels results in closures of shellfish farms, thus causing severe economic losses to aquaculture. Studies on algal production of DA in cultures and field samples require sensitive methods capable of measuring trace concentrations of domoic acid. Measuring domoic acid concentrations at trace levels is still a significant challenge. A sensitive and reliable double derivatization using o-phthaldialdehyde-mercaptoethanol/9-fluorenylmethylchloroformate (OPA-MeSH/FMOC-Cl) followed by high-performance liquid chromatography with fluorescence detection has been adapted for the determination of trace levels of dissolved (dDA) and particulate domoic acid (pDA). The selectivity toward domoic acid is greatly improved by eliminating the reaction between FMOC-Cl and primary amines. Compared with the original Pocklington method, the sensitivity obtained by the double derivatization is largely enhanced (×6) and the reproducibility is significantly increased (<4% relative standard deviation [RSD] for nanomolar domoic acid concentrations). A better precision and an improved interoperator reproducibility were obtained with this two-step derivatization optimized protocol. This methodology will be useful for monitoring domoic acid production both in Pseudo-nitzschia cultures and in natural environmental marine samples.