Bussy Ugo, Chung-Davidson Yu-Wen, Buchinger Tyler, Li Ke, Smith Scott A, Daniel Jones A, Li Weiming
Department of Fisheries and Wildlife, Michigan State University, 480 Wilson Rd., East Lansing, MI, 48824, USA.
RTSF Mass Spectrometry and Metabolomics Core, Michigan State University, Biochemistry Building, 603 Wilson Rd., East Lansing, MI, 48824, USA.
Anal Bioanal Chem. 2018 Feb;410(6):1763-1774. doi: 10.1007/s00216-017-0831-7. Epub 2017 Dec 26.
The sea lamprey (Petromyzon marinus) is a destructive invasive species in the Great Lakes. Since the 1960s, tons of the lampricide 3-trifluoromethyl-4-nitrophenol (TFM) has been applied to selected tributaries each year to eliminate or reduce sea lamprey larval populations. Therefore, the environmental impact of TFM needs to be evaluated. However, the metabolism of TFM and its mechanism of selective toxicity in sea lamprey is not yet fully understood. Based upon our previous report on the identification, synthesis, and characterization of TFM metabolites observed in liver incubates from sea lamprey and non-target fishes, we now provide a robust assay for quantifying TFM and its metabolites in fish liver tissue. This method is important for assessing bioaccumulation of TFM in the ecosystems. The compounds purified in our previous report were used to develop and validate a quantitative ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) assay for TFM and TFM metabolites formed in vivo. Several sample preparation techniques were compared, and a protein precipitation method was selected. The unavailability of stable isotopic internal standards was overcome by using a matrix matching method. After a thorough validation, this method was applied to determine the concentrations of TFM and its metabolites in fish liver tissues from animals exposed to TFM, and in the comparison between dead animals and survivors. Seven of eight expected metabolites were observed, some for the first time in vivo. Our results indicate that in vivo nitroreduction, glucuronidation, sulfation, and glutathione conjugation are involved in TFM metabolism in sea lamprey.
海七鳃鳗（Petromyzon marinus）是五大湖地区具有破坏性的入侵物种。自20世纪60年代以来，每年都有大量的杀七鳃鳗剂3-三氟甲基-4-硝基苯酚（TFM）被应用于特定支流，以消灭或减少海七鳃鳗幼体数量。因此，需要评估TFM对环境的影响。然而，TFM在海七鳃鳗体内的代谢及其选择性毒性机制尚未完全明确。基于我们之前关于在海七鳃鳗和非目标鱼类肝脏孵育物中观察到的TFM代谢物的鉴定、合成和表征的报告，我们现在提供一种可靠的方法来定量鱼肝脏组织中的TFM及其代谢物。该方法对于评估TFM在生态系统中的生物累积至关重要。我们之前报告中纯化的化合物被用于开发和验证一种定量超高效液相色谱-串联质谱（UHPLC-MS/MS）方法，用于测定体内形成的TFM和TFM代谢物。比较了几种样品制备技术，并选择了蛋白质沉淀法。通过使用基质匹配法克服了稳定同位素内标物不可用的问题。经过全面验证后，该方法被应用于测定暴露于TFM的动物肝脏组织中TFM及其代谢物的浓度，并用于比较死亡动物和存活动物。观察到了八种预期代谢物中的七种，其中一些是首次在体内观察到。我们的结果表明，体内硝基还原、葡萄糖醛酸化、硫酸化和谷胱甘肽结合参与了海七鳃鳗体内TFM的代谢。
