Extraction recoveries and stability of diarrhetic shellfish poisoning (DSP) toxins in naturally contaminated samples

Instituto Nacional dos Recursos Biológicos-IPIMAR, Lisbon, Portugal.
Food Additives and Contaminants - Part A Chemistry, Analysis, Control, Exposure and Risk Assessment (Impact Factor: 1.8). 03/2009; 26(2):229-35. DOI: 10.1080/02652030802290530
Source: PubMed


During the last few years the occurrence of a high percentage of esters of diarrhetic shellfish poisoning (DSP) toxins has been observed in shellfish from the Portuguese coast. Most of the commercial bivalves contain DSP toxins in ester forms, either acyl derivatives of okadaic acid (OA) or of dinophysistoxin-2 (DTX-2). The stability of these toxins in shellfish tissues and in raw methanol extracts was investigated in two different naturally contaminated species, mussel and carpet shell, over a 4-week period. The results for both species revealed that DSP toxins were more stable in tissue than in raw methanol extracts. Losses of DSP toxins were seen in the first 2 weeks and were more than 30%, but after that a period of stabilization was observed. The decrease was due probably from losses of esters of OA and DTX-2, the free toxins were stable over the period studied. The extraction most commonly used for chemical and biochemical assays relied on methanolic extraction with aqueous 80% methanol. In this work we have tested the extraction solvent on the extractability of DSP toxins from several naturally contaminated species. A single dispersive extraction with methanol, with solvent ratios of 70%, 80%, 90% and 100%, were tested. After alkaline hydrolysis of esterified toxins and clean-up with hexane and dichloromethane, the samples were analysed by liquid chromatography-mass spectrometry (LC-MS). The recovery of DSP toxins increased with increasing percentages of methanol up to 90%. A decrease in recovery with 100% methanol was observed probably due to problems during the liquid-liquid partitioning.

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    • "The combined dichloromethane layers were dried under reduced pressure and re-suspended in 500 μl of 100% aqueous methanol. Aliquots of 500 μl were transferred to a vial for LC-MS/ MS analysis (Rodrigues et al. 2009;EURLMB 2011). The total sample toxicity was expressed as μg kg −1 of wet weight. "
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    • "The combined dichloromethane layers were dried under reduced pressure and re-suspended in 500 mL of 80% aqueous methanol. Aliquots of 500 mL were transferred to a vial for LC-MS/MS analysis (Rodriguez et al. 2009). "
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