Department of Analytical Chemistry, Faculty of Sciences, Granada, Spain.
Electrophoresis. 2013 Sep;34(18):2615-22. doi: 10.1002/elps.201300247. Epub 2013 Aug 21.
A new MEKC-ESI-MS method for the analysis of amino acids (AAs) in human urine was developed employing ammonium perfluorooctanoate (APFO) as volatile surfactant. The influence of APFO on the MS signal of AAs was evaluated by infusion experiments, which showed that APFO hardly affects analyte responses and presents significantly less ion suppression than equal concentrations of ammonium acetate. In order to obtain efficient separation of AAs, MEKC parameters such as the pH and APFO concentration of the BGE, were optimized. Optimum AA resolution, including baseline separation of leucine and isoleucine, was obtained using 150 mM APFO (pH 9.0) as BGE, representing a considerable selectivity improvement over CE using 50 mM ammonium acetate (pH 9.0). Optimization of CE-MS parameters, such as sheath liquid composition and flow rate, and ESI and MS settings, led to LODs ranging from 9 to 26 ng/mL for the 20 tested AAs, which is highly favorable for an MEKC-MS method. Good linearity (r(2) > 0.99) and repeatability were obtained for all AAs tested with RSD values of 3.0-6.7% for peak area and <1.5% for migration time. The applicability of the MEKC-MS method was demonstrated by the quantitative determination of AAs in urine employing only a 1:1 dilution with BGE as sample pretreatment. All AAs could selectively be detected and quantified obtaining relevant concentration values for normal human urine.
建立了一种新的 MEKC-ESI-MS 方法，用于分析人尿中的氨基酸（AA），采用全氟辛烷铵（APFO）作为挥发性表面活性剂。通过注入实验评估了 APFO 对 AA 的 MS 信号的影响，结果表明 APFO 几乎不会影响分析物的响应，并且比等浓度的乙酸铵产生的离子抑制作用小得多。为了获得 AA 的有效分离，优化了 MEKC 参数，如 BGE 的 pH 值和 APFO 浓度。使用 150mM APFO（pH9.0）作为 BGE，可获得最佳的 AA 分辨率，包括亮氨酸和异亮氨酸的基线分离，与使用 50mM 乙酸铵（pH9.0）的 CE 相比，选择性有了显著提高。优化 CE-MS 参数，如鞘液组成和流速，以及 ESI 和 MS 设置，可使 20 种测试的 AA 的 LOD 范围从 9 到 26ng/mL，这对 MEKC-MS 方法非常有利。对于所有测试的 AA，均获得了良好的线性（r(2)>0.99）和重复性，峰面积的 RSD 值为 3.0-6.7%，迁移时间的 RSD 值<1.5%。该 MEKC-MS 方法的适用性通过仅用 BGE 进行 1:1 稀释作为样品预处理，对尿液中的 AA 进行定量测定得到了证明。可以选择性地检测和定量所有 AA，获得正常人尿液的相关浓度值。
