Development of a Liquid Chromatography–Mass Spectrometry-Based In Vitro Assay to Assess Changes in Steroid Hormones Due to Exposure to Per- and Polyfluoroalkyl Substances
Development of a Liquid Chromatography–Mass Spectrometry-Based In Vitro Assay to Assess Changes in Steroid Hormones Due to Exposure to Per- and Polyfluoroalkyl Substances
复制标题
开发基于液相色谱-质谱的体外测定法，以评估由于暴露于全氟烷基和多氟烷基物质而导致的类固醇激素变化
DOI：
10.1021/acs.chemrestox.2c00116
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发表时间：
2022
影响因子：
4.1
通讯作者：
Aga, Diana S.
中科院分区：
文献类型：
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作者：
Running, Logan;Atilla-Gokcumen, G. Ekin;Aga, Diana S.
全氟化物和多氟化物 (PFAS) 是有机污染物，与多种健康影响有关，包括糖尿病、癌症和内分泌系统失调。本研究旨在开发一种液相色谱串联质谱 (LC-MS/MS) 检测方法，用于测量 H295R 细胞系（一种产生类固醇的肾上腺皮质细胞）暴露于 PFAS 后 17 种激素的变化。由于使用电喷雾电离 MS 分析类固醇激素存在挑战，因此采用化学衍生方法在 LC-MS/MS 中实现 0.07–2 μg/L 的检测限。此外，通过固相萃取 (SPE) 实现的 10 倍浓缩因子可实现十亿分之一的一致检测。衍生化条件的优化显示了一些激素分析物（包括孕酮、皮质酮和皮质醇）中的双衍生产物，并且电离效率比单衍生产物信号高出 20 倍。使用 SPE 进行样品净化，使用弱阴离子交换吸附剂分析细胞介质中的激素，17 种目标激素的回收率达到 80-100%。该方法通过将 H295R 细胞暴露于两种已知的内分泌干扰物（毛喉素和咪鲜胺）进行了验证，这显示了预期的激素变化。 H295R 细胞与各种 PFAS 标准品及其混合物（1 μM）的初始接触显示，使用某些 PFAS 处理（包括 PFBS、PFHxA、PFOS、PFDA 和 PFDS）后，孕激素显着增加。此外，在用其他磺化或羧化头基 PFAS 处理的细胞中观察到激素水平的适度变化。这种用于 H295R 细胞激素分析的灵敏 LC-MS/MS 方法将允许在基于细胞的测定和其他体外模型中研究因暴露于各种环境损害而引起的激素产生的变化。
Per- and poly-fluorinated substances (PFASs) are organic pollutants that have been linked to numerous health effects, including diabetes, cancers, and dysregulation of the endocrine system. This study aims to develop a liquid chromatography with tandem mass spectrometry (LC-MS/MS) assay to measure changes in 17 hormones in H295R cell line (a steroid producing adrenocortical cells) upon exposure to PFASs. Due to the challenges in the analysis of steroid hormones using electrospray ionization MS, a chemical derivatization method was employed to achieve 0.07–2 μg/L detection limits in LC-MS/MS. Furthermore, a 10-fold concentration factor through solid-phase extraction (SPE) allows for consistent sub-parts per billion detections. Optimization of the derivatization conditions showed doubly-derivatized products in some hormone analytes, including progesterone, corticosterone, and cortisol, and gave improved ionization efficiency up to 20-fold higher signal than the singly-derivatized product. The use of SPE for sample cleanup to analyze hormones from cellular media using weak anion exchange sorbent yielded 80–100% recovery for the 17 targeted hormones. The method was validated by exposing H295R cells to two known endocrine disruptors, forskolin and prochloraz, which showed expected changes in hormones. An initial exposure of H295R cells with various PFAS standards and their mixtures at 1 μM showed significant increases in progestogens with some PFAS treatments, which include PFBS, PFHxA, PFOS, PFDA, and PFDS. In addition, modest changes in hormone levels were observed in cells treated with other sulfonated or carboxylated headgroup PFASs. This sensitive LC-MS/MS method for hormone analysis in H295R cells will allow for the investigations of the alterations in the hormone production caused by exposure to various environmental insults in cell-based assays and otherin vitromodels.