University of Crete, Department of Chemistry, Environmental Chemical Processes Laboratory, Heraklion 71003, Crete, Greece.
Anal Chim Acta. 2012 Jun 20;731:49-59. doi: 10.1016/j.aca.2012.04.016. Epub 2012 Apr 20.
A liquid chromatography-electrospray-tandem mass spectrometry (ES-MS/MS) method was developed for the speciation analysis of four organic selenium species of relevance to human urinary metabolism, namely trimethylselenomium ion (TMSe(+)), selenomethionine (SeMet) and the two selenosugars, methyl 2-acetamido-2-deoxy-1-seleno-β-D-galactos/-glucos-amine (SeGalNAc and SeGluNAc, respectively). Their chromatographic separation was achieved by using a cation exchange pre-column coupled in-series with a reversed-phase high-performance liquid chromatography column, along with an isocratic mobile phase. Online detection was performed using ES-MS/MS in selective reaction monitoring mode. SeGalNAc was detected as the major human urinary metabolite of selenium in the samples analysed, whereas TMSe(+) was detected in the urine of one volunteer before and after receiving a selenium supplement. SeMet was not detected as a urine excretory metabolite in this study. Spiking experiments performed with the urine samples revealed significant signal suppression caused by coeluting matrix constituents. To overcome such interferences, isotopically labelled (13)CD(3)(82)SeGalNAc was used as an internal standard, whereas in the absence of an isotopically labelled internal standard for TMSe(+), the standard addition method was applied. Quality control for the accurate quantitation of TMSe(+) and SeGalNAc was carried out by analysing spiked human urine samples with appropriate selenium standards over a concentration range of 10-50 μg Se L(-1). The method has achieved a limit of detection in the presence of urine matrix comparable to that of HPLC-inductively coupled plasma-mass spectrometry for the four selenium species: 1.0 μg Se L(-1) for TMSe(+), 5.6 μg Se L(-1) for SeMet, and 0.1 μg Se L(-1) for both SeGalNAc and SeGluNAc.
建立了一种液相色谱-电喷雾串联质谱（ES-MS/MS）方法，用于分析与人体尿代谢有关的四种有机硒形态，即三甲基硒离子（TMSe(+））、硒蛋氨酸（SeMet）和两种硒糖，分别为甲基 2-乙酰氨基-2-脱氧-1-硒代-β-D-半乳糖/葡萄糖胺（SeGalNAc 和 SeGluNAc）。采用阳离子交换预柱与反相高效液相色谱柱串联，以等度流动相实现它们的色谱分离。在线检测采用 ES-MS/MS 进行选择反应监测模式。在分析的样品中，SeGalNAc 被检测为硒的主要人体尿代谢物，而 TMSe(+) 则在一名志愿者接受硒补充前后的尿液中被检测到。在本研究中，未检测到 SeMet 作为尿液排泄代谢物。用尿液样品进行的加标实验表明，共洗脱基质成分会导致显著的信号抑制。为了克服这种干扰，使用同位素标记的（13）CD(3)(82)SeGalNAc 作为内标，而对于 TMSe(+) 没有同位素标记的内标，则应用标准加入法。通过用适当的硒标准品分析加标人尿样品，对 TMSe(+) 和 SeGalNAc 的准确定量进行了质量控制，硒标准品的浓度范围为 10-50 μg Se L(-1)。该方法在尿液基质存在下的检测限与 HPLC-电感耦合等离子体质谱法相当，适用于四种硒形态：TMSe(+) 的检测限为 1.0 μg Se L(-1)，SeMet 的检测限为 5.6 μg Se L(-1)，SeGalNAc 和 SeGluNAc 的检测限均为 0.1 μg Se L(-1)。
