Louzao M C, Vieytes M R, Yasumoto T, Botana L M
Departamento de Farmacologia and Departamento de Fisiologia Animal, Facultad de Veterinaria de Lugo Universidad de Santiago de Compostela, 27002 Lugo, Spain.
Chem Res Toxicol. 2004 Apr;17(4):572-8. doi: 10.1021/tx0342262.
Marine toxins such as brevetoxins and ciguatoxins are produced by dinoflagellates and can accumulate in seafood. These toxins affect humans through seafood consumption. Intoxication is mainly characterized by gastrointestinal and neurological disorders and, in most severe cases, by cardiovascular problems. To prevent the consumption of food contaminated with these toxins, shellfish have been tested by mouse bioassay. However, this method is expensive, time-consuming, and ethically questionable. The objective of this study was to use a recently developed fluorimetric microplate assay to rapidly detect brevetoxins and ciguatoxins. The method is based on the pharmacological effect of brevetoxins and ciguatoxins known to activate sodium channels and involves (i). the incubation of excitable cells in 96 well microtiter plates with the fluorescent dye bis-oxonol, whose distribution across the membrane is potential-dependent, and (ii). dose-dependent cell depolarization by the toxins. Our findings demonstrate that measuring changes in membrane potential induced by brevetoxins and ciguatoxins allowed their quantitation. Active toxins could be reliably detected at concentrations in the nanomolar range. The simplicity, sensitivity, and possibility of being automated provide the basis for development of a practical alternative to conventional testing for brevetoxins and ciguatoxins.
诸如短裸甲藻毒素和雪卡毒素之类的海洋毒素由双鞭毛藻产生，并可在海产品中蓄积。这些毒素通过食用海产品影响人类。中毒主要表现为胃肠道和神经系统紊乱，在最严重的情况下，还会出现心血管问题。为防止食用受这些毒素污染的食物，已通过小鼠生物测定法对贝类进行检测。然而，这种方法成本高、耗时且在伦理上存在问题。本研究的目的是使用最近开发的荧光微孔板测定法快速检测短裸甲藻毒素和雪卡毒素。该方法基于已知能激活钠通道的短裸甲藻毒素和雪卡毒素的药理作用，包括：（i）将可兴奋细胞与荧光染料双羟萘酚在96孔微量滴定板中孵育，该染料在膜上的分布取决于电位；（ii）毒素引起剂量依赖性细胞去极化。我们的研究结果表明，测量短裸甲藻毒素和雪卡毒素引起的膜电位变化可对其进行定量。在纳摩尔浓度范围内能够可靠地检测到活性毒素。其简单性、敏感性以及自动化的可能性为开发一种替代传统短裸甲藻毒素和雪卡毒素检测方法的实用方法提供了基础。
