Consumption of ciguatoxin-contaminated seafood can lead to ciguatera poisoning (CP). The diagnosis of CP in humans is based on the clinical symptoms after eating the fish from tropical or subtropical areas because no confirmatory clinical tests are available. One of the challenges for ciguatoxin analysis is their extremely low but toxicologically relevant concentration in biological samples. We previously reported a method using acetonitrile to precipitate proteins and extract the ciguatoxins simultaneously in whole blood samples from animals for toxin quantification by N2A cell-based assay. However, a test method for unambiguous confirmation of exposure of marine animals or humans to ciguatoxins is still needed. In the present study, we adopted the acetonitrile extraction method and added sample clean-up in the sample preparation for the determination of Pacific ciguatoxins CTX1B (aka P-CTX-1), 52-epi-54-deoxyCTX1B (aka P-CTX-2), and CTX3C (aka P-CTX-3C) in blood plasma by LC-MS/MS. We investigated sample clean-up, LC mobile phases, LC solvent programming, and settings of the two mass spectrometers (4000 Q TRAP and AB SCIEX Triple Quad 5500) in order to improve the ability to detect the Pacific ciguatoxins at ppt level. Rat blood plasma was used for the method development. Average recoveries of the three toxins in the rat plasma samples ranged from 90% to 116% with relative standard deviations of less than 15%. The method detection limits were still not low enough for the determination of the Pacific ciguatoxins in individual blood samples from Hawaiian monk seals with the two LC-MS systems. The methods were applied to a pooled sample of blood plasma collected from Hawaiian monk seals for confirmation of toxin exposure. This study will benefit monitoring of Pacific ciguatoxins in marine mammals and potentially humans by LC-MS/MS.

中文翻译：

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样品制备和液相色谱-串联质谱法用于分析血液样品中选定的太平洋雪茄毒素。

食用被雪茄毒素污染的海鲜会导致雪茄中毒（CP）。对人类CP的诊断是基于从热带或亚热带地区食用鱼后的临床症状，因为尚无确证的临床测试。雪茄毒素分析的挑战之一是它们在生物样品中的浓度极低，但在毒理学上相关。我们先前报道了一种使用乙腈沉淀蛋白质并同时从动物的全血样品中提取雪茄毒素的方法，用于通过基于N2A细胞的测定进行毒素定量。但是，仍然需要一种明确确认海洋动物或人类接触雪茄毒素的测试方法。在目前的研究中，我们采用了乙腈提取方法，并在样品制备中添加了样品净化功能，用于测定太平洋雪茄毒素CTX1B（aka P-CTX-1），52-epi-54-deoxyCTX1B（aka P-CTX-2）和CTX3C LC-MS / MS检测血浆中的P-CTX-3C（aka P-CTX-3C）。我们研究了样品净化，液相色谱流动相，液相色谱溶剂编程以及两个质谱仪（4000 Q TRAP和AB SCIEX Triple Quad 5500）的设置，以提高在ppt水平检测太平洋雪茄毒素的能力。大鼠血浆用于方法开发。大鼠血浆样品中三种毒素的平均回收率从90％到116％不等，相对标准偏差小于15％。该方法的检出限仍不足以通过两个LC-MS系统从夏威夷僧海豹的单个血液样品中测定太平洋西瓜毒素。将该方法应用于从夏威夷僧海豹收集的血浆集合样品中，以确认毒素暴露。这项研究将有利于通过LC-MS / MS监测海洋哺乳动物和潜在人类中的太平洋雪茄毒素。