Pia Dima Anna, De Santis Lucia, Verlengia Cristina, Lombardo Francesco, Lenzi Andrea, Mazzarino Monica, Botrè Francesco, Paoli Donatella
Laboratory of Seminology - Sperm Bank "Loredana Gandini", Department of Experimental Medicine, "Sapienza" University of Rome, Italy.
IVF Unit, San Raffaele Scientific Institute, Vita-Salute University, Milan, Italy.
Clin Mass Spectrom. 2020 Oct 28;18:54-65. doi: 10.1016/j.clinms.2020.10.002. eCollection 2020 Nov.
Phthalates and bisphenol A interfere with the synthesis, secretion, transport, binding, metabolism, and excretion of endogenous hormones and, for this reason, are classified as endocrine disruptors. We are here presenting an analytical method for the simultaneous detection of six phthalates metabolites and bisphenol A in different biological fluids (urine, serum and follifular fluid) by liquid chromatography coupled to tandem mass spectrometry. The quantification was carried out in negative electrospray ionization mode using selected reaction monitoring as acquisition mode. Different extraction protocols, using either solid phase or liquid/liquid extraction, were comparatively evaluated to optimize the sample preparation procedure. Solid-phase extraction was chosen as it ensured the best recovery and overall sensitivity. The method was successfully validated: recovery varying in the range 71 ± 2%-107 ± 6% depending on the target analyte and the matrix considered, and ≤ 12% for follicular fluid, ≤11% for serum and ≤ 10% for urine and accuracy ≤ 115% for follicular fluid, ≤113% for serum ≤ 115% for urine , linearity with R > 0.99, with the exception of MEP (recovery 64 ± 8%, ≤ 20%, precision ≤ 16% for follicular fluid). The actual applicability of the method developed and validated in this study was assessed by the analysis of real samples, including 10 specimens of follicular fluid, serum and urine samples, that showed the presence of phthalates metabolites and Bisphenol A, and confirming that the newly developed method can be applied in the routine clinical laboratory for the identification and quantitation of these endocrine-disrupting chemicals.
邻苯二甲酸盐和双酚A会干扰内源性激素的合成、分泌、运输、结合、代谢及排泄，因此被归类为内分泌干扰物。本文介绍了一种采用液相色谱-串联质谱法同时检测不同生物体液（尿液、血清和卵泡液）中六种邻苯二甲酸盐代谢物和双酚A的分析方法。定量分析在负电喷雾电离模式下进行，采用选择反应监测作为采集模式。对使用固相萃取或液-液萃取的不同提取方案进行了比较评估，以优化样品制备程序。选择固相萃取是因为它能确保最佳回收率和整体灵敏度。该方法成功得到验证：回收率根据目标分析物和所考虑的基质不同，在71±2%-107±6%范围内变化，卵泡液中≤12%，血清中≤11%，尿液中≤10%；准确度方面，卵泡液中≤115%，血清中≤113%，尿液中≤115%；线性关系R>0.99，但2-甲基-1,3-丙二醇单酯（MEP，回收率64±8%，卵泡液中≤20%，精密度≤16%）除外。通过对实际样品（包括10份卵泡液、血清和尿液样本）的分析，评估了本研究中开发和验证的方法的实际适用性，这些样品显示存在邻苯二甲酸盐代谢物和双酚A，证实新开发的方法可应用于常规临床实验室，用于鉴定和定量这些内分泌干扰化学物质。
