Department of Pharmacy, University of Eastern Finland, Yliopistonranta 1 C, FI-70211 Kuopio, Finland.
J Steroid Biochem Mol Biol. 2011 Nov;127(3-5):396-404. doi: 10.1016/j.jsbmb.2011.06.006. Epub 2011 Jun 12.
A fast and sensitive LC-MS/MS method for the quantitative analysis of seven steroid hormones in 150 μl of human serum was developed and validated. The following compounds were included: 17α-hydroxypregnenolone, 17α-hydroxyprogesterone, androstenedione, dehydroepiandrosterone, testosterone, pregnenolone, and progesterone. Individual stable isotope-labeled analogues were used as internal standards. Sample preparation was performed by liquid-liquid extraction, followed by oxime derivatization to improve the ionization efficiency of the analytes. In contrast to the common derivatization-based methods, the reaction was incorporated into the sample preparation process and the only additional step due to the derivatization was a short heating of the autosampler vials before the sample injection. Chromatographic separation was achieved on a reversed-phase column using a methanol-water gradient. For the analyte detection, a triple quadrupole instrument with electrospray ionization was used. Total run time was 7.0 min and the lower limits of quantification were in the range of 0.03-0.34 nM (0.01-0.10 ng/ml), depending on the analyte. The method was validated using human serum samples from both sexes and applied for the serum steroid profiling of endometriosis patients.
建立并验证了一种快速灵敏的 LC-MS/MS 法，可用于在 150μl 人血清中同时定量分析 7 种甾体激素。包括以下化合物：17α-羟孕烯醇酮、17α-羟孕酮、雄烯二酮、脱氢表雄酮、睾酮、孕烯醇酮和孕酮。采用单个稳定同位素标记的类似物作为内标。样品制备采用液液萃取，然后进行肟衍生化以提高分析物的离子化效率。与常见的衍生化方法不同，该反应被整合到样品制备过程中，由于衍生化而增加的唯一额外步骤是在进样前短时间加热自动进样瓶。采用甲醇-水梯度在反相柱上进行色谱分离。使用电喷雾电离的三重四极杆仪器进行分析物检测。总运行时间为 7.0 分钟，定量下限范围为 0.03-0.34 nM（0.01-0.10ng/ml），具体取决于分析物。该方法使用来自两性的人血清样本进行了验证，并应用于子宫内膜异位症患者的血清甾体谱分析。
