Sayfritz S J, Aasen J A B, Aune T
Department of Food Safety and Infection Biology, Norwegian School of Veterinary Science, P.O. Box 8146 Dep., 0033, Oslo, Norway.
Toxicon. 2008 Aug 1;52(2):330-40. doi: 10.1016/j.toxicon.2008.06.001. Epub 2008 Jun 13.
A novel extraction and clean-up method has been developed for the determination of paralytic shellfish poisoning (PSP) toxins in shellfish samples. Raw shellfish material was extracted with an acidic acetonitrile/water (80:20, v/v) solution, whilst being homogenised. During the homogenisation the sample extraction solution was cooled with ice water. Subsequently, the extract was frozen at -20 degrees C for at least 4h. During freezing, two layers were formed, only the lower predominantly aqueous layer was used for the determination. The final extract solution was cleaned-up using a combination of Oasis HLB and Carbograph activated carbon SPE columns. The developed extraction and clean-up methods combined with gradient elution liquid chromatography (LC)-mass spectrometry/mass spectrometry (MS/MS) has resulted in a method which can determine the analogs GTX 1-5, C1-2, DcGTX 2-3, DcSTX, Neo, STX in a single analysis with an overall detection limit of 313 microg STXdiHCL-eq./kg shellfish meat. The use of the developed extraction method with post-column high performance liquid chromatography (HPLC) with fluorescence detection (FLD) method provided an overall limit of detection of 89 microg STXdiHCL-eq./kg shellfish meat for the same toxins. Both post-column HPLC-FLD and LC-MS/MS was used to investigate the Norwegian PSP toxin profile. It was found that the PSP toxins could be detected in shellfish samples from the Norwegian coastline for 10 months of the year, from March till December. The toxin profile consisted mainly of the carbamate toxins, GTX 1-4, Neo and STX, in terms of both concentrations and contribution to the overall toxicity. In addition, several of the n-sulfo-carbamoyl toxins were either detected in the samples at relatively low concentrations or their presence in the samples were indicated but could not be confirmed by the post-column HPLC-FLD and LC-MS/MS analyses.
已开发出一种新型提取和净化方法，用于测定贝类样品中的麻痹性贝类毒素（PSP）。将生贝类材料用酸性乙腈/水（80:20，v/v）溶液提取，同时进行匀浆。匀浆过程中，样品提取溶液用冰水冷却。随后，提取物在-20℃冷冻至少4小时。冷冻过程中形成两层，仅使用下层主要为水相的层进行测定。最终提取溶液使用Oasis HLB和Carbograph活性炭固相萃取柱组合进行净化。所开发的提取和净化方法与梯度洗脱液相色谱（LC）-质谱/质谱（MS/MS）相结合，形成了一种能够在单次分析中测定GTX 1-5、C1-2、DcGTX 2-3、DcSTX、Neo、STX类似物的方法，总体检测限为313μg STXdiHCL-当量/千克贝肉。使用所开发的提取方法结合柱后高效液相色谱（HPLC）荧光检测（FLD）方法，对相同毒素的总体检测限为89μg STXdiHCL-当量/千克贝肉。柱后HPLC-FLD和LC-MS/MS均用于研究挪威PSP毒素谱。结果发现，一年中有10个月（从3月到12月）在挪威海岸线的贝类样品中可检测到PSP毒素。就浓度和对总体毒性的贡献而言，毒素谱主要由氨基甲酸酯类毒素GTX 1-4、Neo和STX组成。此外，几种n-磺基氨基甲酰毒素要么在样品中以相对较低的浓度被检测到，要么表明它们在样品中的存在，但柱后HPLC-FLD和LC-MS/MS分析无法证实。
