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首页> 外文期刊>Environmental toxicology and chemistry >Quantitative measurement of fathead minnow vitellogenin by liquid chromatography combined with tandem mass spectrometry using a signature peptide of vitellogenin.
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Quantitative measurement of fathead minnow vitellogenin by liquid chromatography combined with tandem mass spectrometry using a signature peptide of vitellogenin.

机译：通过液相色谱结合串联质谱法定量测定黑头min鱼卵黄蛋白原，采用卵黄蛋白原的特征肽。

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摘要

Vitellogenin (VTG) has been proposed as a sensitive biomarker of exposure to environmental estrogenic contaminants that induce VTG production in oviparous species. Enzyme-linked immunosorbent assay (ELISA) methods are currently widely used to measure the VTG levels. In this paper, a new liquid chromatography combined with tandem mass spectrometry (LC/ESI-MS/MS) method for the quantitative analysis of VTG in the plasma of fathead minnows exposed to 17alpha-ethinylestradiol (EE2) has been developed. This method includes, first, the selection of the signature peptide, which involves sodium dodecyl sulfate-polyarylamide gel electrophoresis separation, in-gel digestion, LC/ESI-MS/MS analysis with an ion trap mass spectrometer, and sequence determination with the TurboSEQUEST MS/MS database application; second, optimization of the selected signature peptide in multireaction monitor (MRM) mode with a triple quadrupole mass spectrometer; and third, trypsin digestion of plasma and VTG quantitation via MRM-mode LC/ESI-MS/MS. A series of plasma samples from fathead minnows following exposure to EE2 was assayed. A good correlation was found when EE2-induced plasma samples from fathead minnows were analyzed with ELISA and the described new method. Although used here with fathead minnow, the new LC/ESI-MS/MS method could be easily applied to the analysis of VTG expressed in any other fish species. Quantitation of VTG by this method was found to be highly specific and linear. The absence of potential artifactual measurements of VTG at low exposure levels could also be critical in future studies that evaluate weakly estrogenic compounds in aquatic species.
机译：卵黄蛋白原（VTG）已被提议作为暴露于环境雌激素污染物的敏感生物标志物，这些雌激素污染物可在卵子物种中诱导VTG的产生。酶联免疫吸附测定（ELISA）方法目前广泛用于测量VTG水平。本文开发了一种新的液相色谱-串联质谱法（LC / ESI-MS / MS），用于定量分析暴露于17α-乙炔雌二醇（EE2）的黑头the血浆中的VTG。该方法首先包括选择标记肽，包括十二烷基硫酸钠-聚芳基酰胺凝胶电泳分离，凝胶内消化，离子阱质谱仪进行LC / ESI-MS / MS分析以及使用TurboSEQUEST进行序列确定MS / MS数据库应用程序；其次，使用三重四极杆质谱仪在多反应监测器（MRM）模式下优化选定的特征肽。第三，通过MRM模式LC / ESI-MS / MS对血浆进行胰蛋白酶消化和VTG定量。分析了暴露于EE2后的黑头fat的一系列血浆样品。当用ELISA和描述的新方法分析EE2诱导的黑头min鱼血浆样品时，存在良好的相关性。尽管在这里与黑头min鱼一起使用，但是新的LC / ESI-MS / MS方法可以轻松地用于分析在任何其他鱼类中表达的VTG。发现通过这种方法对VTG的定量是高度特异性和线性的。在未来的评估水生生物中弱雌激素化合物的研究中，缺乏低暴露水平的VTG的潜在人为测量也可能至关重要。

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