NG108-15 cell-based and protein phosphatase inhibition assays as alternative semiquantitative tools for the screening of lipophilic toxins in mussels. Okadaic acid detection.
We report the application of a cell-based assay (CBA) using NG108-15 a hybridoma cell strain and a protein phosphatase inhibition-based assay (PPIA) as alternative toxicological or functional semiquantitative tools, respectively, for the screening of lipophilic toxins in mussels (Mytilusgalloprovincialis). Acetonic extracts were directly tested by CBA and PPIA but severe matrix effects were observed. As a solution, a simple 17-fraction protocol with solid-phase extraction (SPE) cartridges was optimised and applied as a previous step to the CBA or the PPIA. LC-MS/MS analyses were performed in parallel to determine the lipophilic toxins content in mussel extracts. Evaluation of the SPE protocol by LC-MS/MS showed okadaic acid (OA) recovery above 90% and negligible effects of mussel matrix on the SPE performance. The whole methods provided limits of detection of 47 and 45mug OA equivalents/kg for CBA and PPIA, respectively. The combined strategy permitted the identification of OA toxicity in two fractions, and allowed us to clearly distinguish between negative and positive samples, the latter being either OA-spiked or naturally-contaminated samples at levels equal or above the regulatory limit. The combination of fractioning with CBA or PPIA allows the quantification of the toxic and functional effects of samples above these concentrations.
Cañete
Elisabeth
E
IRTA, Ctra. Poble Nou, km 5.5, 43540 Sant Carles de la Ràpita, Tarragona, Spain.
Campàs
Mònica
M
de la Iglesia
Pablo
P
Diogène
Jorge
J
D000818:Animals; D001681:Biological Assay / Q000379:methods*; D049872:Bivalvia / Q000378:metabolism*; D002460:Cell Line; D002853:Chromatography, Liquid; D006822:Hybrid Cells; D008387:Marine Toxins / Q000032:analysis*; D019319:Okadaic Acid / Q000737:chemistry / Q000378:metabolism*; D054648:Protein Phosphatase 2 / Q000037:antagonists & inhibitors*; D053719:Tandem Mass Spectrometry
D008387:Marine Toxins; D019319:Okadaic Acid; D054648:Protein Phosphatase 2