A method that uses liquid chromatography with tandem mass spectrometry (LC/MS/MS) has been developed for the highly sensitive and specific determination of amnesic shellfish poisoning toxins, diarrhetic shellfish poisoning toxins, and other lipophilic algal toxins and metabolites in shellfish. The method was subjected to a full single-laboratory validation and a limited interlaboratory study. Tissue homogenates are blended with methanol-water (9 + 1), and the centrifuged extract is cleaned up with a hexane wash. LC/MS/MS (triple quadrupole) is used for quantitative analysis with reversed-phase gradient elution (acidic buffer), electrospray ionization (positive and negative ion switching), and multiple-reaction monitoring. Ester forms of dinophysis toxins are detected as the parent toxins after hydrolysis of the methanolic extract. The method is quantitative for 6 key toxins when reference standards are available: azaspiracid-1 (AZA1), domoic acid (DA), gymnodimine (GYM), okadaic acid (OA), pectenotoxin-2 (PTX2), and yessotoxin (YTX). Relative response factors are used to estimate the concentrations of other toxins: azaspiracid-2 and -3 (AZA2 and AZA3), dinophysis toxin-1 and -2 (DTX1 and DTX2), other pectenotoxins (PTX1, PTX6, and PTX11), pectenotoxin secoacid metabolites (PTX2-SA and PTX11-SA) and their 7-epimers, spirolides, and homoYTX and YTX metabolites (45-OHYTX and carboxyYTX). Validation data have been gathered for Greenshell mussel, Pacific oyster, cockle, and scallop roe via fortification and natural contamination. For the 6 key toxins at fortification levels of 0.05–0.20 mg/kg, recoveries were 71–99% and single laboratory reproducibilities, relative standard deviations (RSDs), were 10–24%. Limits of detection were <0.02 mg/kg. Extractability data were also obtained for several toxins by using successive extractions of naturally contaminated mussel samples. A preliminary interlaboratory study was conducted with a set of toxin standards and 4 mussel extracts. The data sets from 8 laboratories for the 6 key toxins plus DTX1 and DTX2 gave within-laboratories repeatability (RSDr) of 8–12%, except for PTX-2. Between-laboratories reproducibility (RSDR) values were compared with the Horwitz criterion and ranged from good to adequate for 7 key toxins (HorRat values of 0.8–2.0).

An individual non-motile (NM) cell was isolated from a surface sediment sample collected in Guangxi, China and subsequently established as a dinoflagellate strain in culture. The motile cells are 22.5Á32.5 mm long and 20.0Á30.0 mm wide, with a plate formula of Po, X, 4?, 3a, 7ƒ, 6c, 5(?)s, 5ƒ?, 2ƒƒ, fitting the description of Vulcanodinium rugosum. The Chinese strain shares 99.8%, 97.4% and 96.7% similarity (LSU sequence) with those from Australasia, France and Japan. Asexual division of V. rugosum takes place either in the thecate motile stage or within a NM division cell. Motile cells divided by binary fission inside the parent cell and transformed to NM division cells within 24 h. The NM cell underwent one to three consecutive divisions within the parent wall. The divisions were not always synchronous and neither was the release of motile cells from the NM cells. It generally took 4 to 6 days for the NM cells to complete one division. NM cells survived for 1 month at 4 8C in the dark, suggesting that they might play an important role in species dispersal. A novel pinnatoxin was detected at 20 pg cell(1 and no other known pinnatoxins (AÁG) were detected.