Quantification of lipophilic toxins associated with diarrhetic shellfish poisoning in Japanese bivalves by liquid chromatography-mass spectrometry and comparison with mouse bioassay

Abstract

Quantification of lipophilic toxins in bivalves associated with diarrhetic shellfish poisoning was investigated by liquid chromatography-mass spectrometry (LC-MS). Using a C8-silica reversed phase column and a mobile phase of aqueous acetonitrile containing 2 mM ammonium formate and 50 mM formic acid, okadaic acid, dinophysistoxin-1, 7-O-palmitoyldinophysistoxin-1, pectenotoxin-1, pectenotoxin-2, pectenotoxin-6, pectenotoxin-2 seco-acid, yessotoxin, and 45-hydroxyyessotoxin in bivalves were quantified by LC-MS in the negative mode. When the crude 90% methanol extracts were analyzed by LC-MS, there were no significant effects from bivalve matrices on the quantification of toxins. More than 200 bivalve samples collected from various production areas in Japan were analyzed by LC-MS. Pectenotoxin-6 and dinophysistoxin-1 were the dominant toxins in scallops and mussels, respectively. Yessotoxin and 45-hydroxyyessotoxin were also detected in both species. Comparison of the quantitative results obtained for these bivalve samples between LC-MS and mouse bioassay indicates that LC-MS is suitable for routine monitoring of lipophilic toxins in Japanese bivalves.

LC-MS/MS Analysis of Diarrhetic Shellfish Poisoning (DSP) Toxins, Okadaic Acid and Dinophysistoxin Analogues, and Other Lipophilic Toxins

“…Table 5 shows the average toxin profiles obtained from the 61 samples that gave the discrepancies between the toxicities quantified by MBA (<0.05 MU/g) and LC-MS (>0.05 MU/g). 63 The percentage of YTXs was more than 30%. The discrepancies are probably caused by the YTXs that are poorly detected by MBA.…”

“…Recoveries were greater than 90% for each toxin from bivalve extracts fortified at 0.05 μg/mL (0.5 μg/g hepatopancreas (HP)) even when methanol:water (9:1, v/v) extracts of bivalve were directly analyzed by the LC-MS with a column switching valve. 63 This LC-MS method enabled us to elucidate the toxin profile of bivalves in various production areas in Japan. Dominant toxins in bivalves and toxic dinoflagellates were detected using SIM for -of yessotoxin (m/z 1141.5).…”

“…Figure 19 shows comparison of the mouse toxicities quantified by MBA and LC-MS for the 196 scallop and mussel samples in DSP testing. 63 Toxin profiles of these bivalves are listed in Table 4 Ninety-six samples were quantified by both LC-MS and MBA as being below the quarantine level. Thirty-three samples were above the quarantine level by both LC-MS and MBA.…”

“…Table 4 lists the toxin profiles obtained from scallops and mussels collected from various production areas in Japan. 63 PTX2sa was excluded from the toxin profiles because it does not respond in the mouse bioassay, 13 even though it was the dominant PTX analogue reported in Mytilus galloprovincialis contaminated by Dinophysis spp. 56 The standard deviation (SD) in each toxin profile was high because samples in each monitoring site were collected in different seasons through a year.…”

“…Simultaneous LC-MS analysis of OA/DTX analogues, PTXs and YTXs is possible by using C8 or C18 columns. [61][62][63]65 The LC-MS chromatogram of the simultaneous analysis of DSP and lipophilic toxins is illustrated in Fig. 18.…”