Ecotoxicology and Wildlife Health Division, Wildlife and Landscape Science Directorate, Environment and Climate Change Canada, National Wildlife Research Centre, 1125 Colonel By Drive, Carleton University, Ottawa, ON, K1A 0H3, Canada.
Ecotoxicology and Wildlife Health Division, Wildlife and Landscape Science Directorate, Environment and Climate Change Canada, National Wildlife Research Centre, 1125 Colonel By Drive, Carleton University, Ottawa, ON, K1A 0H3, Canada.
Chemosphere. 2019 Dec;237:124474. doi: 10.1016/j.chemosphere.2019.124474. Epub 2019 Jul 27.
The present study investigated the metabolism of the flame retardant and plasticizer chemical, triphenyl phosphate (TPHP), in a rat liver microsome-based in vitro assay with glutathione (GSH) in order to elucidate metabolic pathways leading to formation of conjugates. A highly sensitive and efficient method was developed for the detection and characterization of GSH reactive metabolites using LC-Q-TOF-MS/MS both in the negative and positive electrospray ionization modes. Seven GSH conjugates formed as a result of microsomal incubation, which were identified as S-conjugates based on MS/MS spectra, and confirmed by subsequent time-dependent incubation assays. With the exception of hydrolysis reactions leading to formation of a diester metabolite, diphenyl phosphate (DPHP), the results demonstrated that Phase I epoxidation on phenyl ring of TPHP leading to mono- and di-hydroxylated TPHP metabolites, which can further conjugate with GSH. Depending on hydroxylated TPHP formation, an o-hydroquinone intermediate formed in vitro via Phase I metabolism, and the o-benzoquinone form reacted with GSH and also formed GSH conjugates. The present study showed that via hydroxylated TPHP Phase I formation that GSH conjugates are important Phase II metabolites for TPHP metabolism in vitro. Some GSH conjugates may be valuable candidate biomarkers for monitoring TPHP exposure in biota.
本研究采用大鼠肝微粒体体外试验，结合谷胱甘肽（GSH），研究阻燃剂和增塑剂化学物质三苯基磷酸酯（TPHP）的代谢情况，以阐明导致形成轭合物的代谢途径。采用 LC-Q-TOF-MS/MS 建立了一种用于检测和表征 GSH 反应性代谢物的高灵敏度和高效方法，分别在负离子和正离子电喷雾电离模式下进行。通过微粒体孵育形成了七种 GSH 轭合物，根据 MS/MS 图谱将其鉴定为 S-轭合物，并通过随后的时程孵育试验进行了确认。除了导致二酯代谢物二苯基磷酸酯（DPHP）形成的水解反应外，结果表明 TPHP 苯环上的 I 相环氧化导致单羟基化和二羟基化 TPHP 代谢物形成，这些代谢物可以进一步与 GSH 结合。根据羟基化 TPHP 的形成情况，体外 I 相代谢会形成邻苯二酚中间产物，邻苯醌形式会与 GSH 反应，也会形成 GSH 轭合物。本研究表明，通过 I 相形成的羟基化 TPHP，GSH 轭合物是 TPHP 代谢体外的重要 II 相代谢物。一些 GSH 轭合物可能是监测生物体内 TPHP 暴露的有价值的候选生物标志物。
