超高效液相色谱-高分辨质谱确证分析泥鳅体内五氯酚代谢物

建立了超高效液相色谱-高分辨质谱(UPLC-HRMS)鉴定泥鳅体内五氯酚代谢物五氯酚磺酸酯的方法。将在低浓度五氯酚下暴露的泥鳅样品粉碎,采用含8%(体积分数)三乙胺的70%(体积分数)乙腈水溶液提取,经混合阴离子交换小柱萃取净化,在ACQUITY BEH C18色谱柱(100 mm×2.1 mm,1.7μm)上分离,采用电喷雾负离子(ESI-)一级质谱全扫描加数据依赖的二级质谱扫描(full mass-ddMS2)模式测定,获得代谢物的准分子离子、同位素离子和二级质谱碎片离子的精确质量数。结果表明,五氯酚在泥鳅体内的代谢以磺化为主,没有发现羟基化和葡萄糖醛酸化。代谢物主要为五氯酚磺酸酯,其含量随着暴露时间(t)的延长逐渐增加,当暴露时间为36 h时达到峰值,随后逐渐减小,当t≥120 h时,五氯酚磺酸酯含量基本维持不变。该方法可用于生物体内五氯酚的代谢研究。

Confirmation and analysis of the metabolites of pentachlorophenol in loaches using ultra-performance liquid chromatography-high resolution mass spectrometry

A method for the characterization of the metabolite pentachlorophenol hydrogen sulfate (PCP-SO₃H) of pentachlorophenol (PCP) in loaches was developed based on ultra-performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS). The loach samples exposed in low concentration of PCP solution were firstly crushed, then extracted by acetonitrile-water solution (70:30, v/v) containing 8% (v/v) triethylamine and purified by mixed anion exchange solid-phase extraction (SPE) cartridges. The chromatographic separation was carried out on a Waters ACQUITY BEH C₁₈ column (100 mm×2.1 mm, 1.7 μm). The qualitative analysis of the metabolites of PCP was operated in a negative electrospray ion mode (ESI^-) under full mass-data dependent MS² (full mass-ddMS²) mode, and the data of quasi-molecular ion, isotope ions, and MS² fragmentation ions of metabolites were obtained. The results revealed that the sulfonation was the main metabolic pathway for PCP in loaches, not the hydroxylation or glucuronate pathway. And the metabolite was found to be PCP-SO₃H. Besides, with the increase of exposure time in PCP solution, the concentration of metabolite PCP-SO₃H firstly increased; when the exposure time was up to 36 h, the concentration of metabolite in loach reached a maximum value; and then extending the exposure time, the concentration of PCP-SO₃H gradually decreased. When the exposure time was up to 120 h, the concentration of PCP-SO₃H in loaches reached a lowest value, and no significant change occurred for further time. The developed UPLC-HRMS method in this study could be used for the investigation of the metabolism of PCP in living beings.