Quantitative determination of marine toxins associated with diarrhetic shellfish poisoning by liquid chromatography coupled with mass spectrometry

Quantitative determination by liquid chromatography (LC) coupled with mass spectrometry (MS) was achieved for the following 10 toxins found in association with diarrhetic shellfish poisoning: okadaic acid (OA), dinophysistoxin-1 (DTX1), 7-O-palmitoylokadaic acid (palOA), 7-O-palmitoyldinophysistoxin-1 (palDTX1), pectenotoxin-1 (PTX1), pectenotoxin-2 (PTX2), pectenotoxin-2 seco acid (PTX2SA), pectenotoxin-6 (PTX6), yessotoxin (YTX), and 45-hydroxyyessotoxin (YTXOH). Toxins in 2 g of the adductor muscle or the digestive glands of scallops, Patinopecten yessoensis, were extracted with 18 ml of methanol–water (9:1, v/v), freed of polar contaminants by partition between chloroform and water, and treated by solid-phase extraction on a silica cartridge column. Samples containing YTXOH were purified separately on a buffered reversed-phase column. Chromatographic separation was achieved by the following combinations of columns and mobile phases: a Symmetry C18 column with acetonitrile–0.05% acetic acid (7:3, v/v) for OA, DTX1, PTX6 and PTX2SA; a Develosil ODS column with the same mobile phase for PTX1 and PTX2; a Capcellpak column with methanol–2.5% acetic acid (98:2, v/v) for palOA and palDTX1; and an Inertsil ODS column with methanol–0.2 M ammonium acetate (8:2, v/v) for YTX and YTXOH. Carboxylic acid toxins were selectively monitored on [M−H]− ions, sulfated toxins on [M−Na]− ions, and neutral toxins on [M+NH4]+ ions. Average recoveries of the toxins spiked to tissue homogenates ranged from 70 to 134%. Detection limits in the muscle ranged from 5 to 40 ng/g and those in the digestive glands from 10 to 80 ng/g.