MS Metabolomics Laboratory, Core Facility, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Magyar Tudósok Blvd 2, H-1117 Budapest, Hungary; Department of Inorganic and Analytical Chemistry, Faculty of Chemical Technology and Biotechnology, Budapest University of Technology and Economics, Szent Gellért Sq 4, H-1111 Budapest, Hungary.
School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Centre Médical Universitaire (CMU), Rue Michel-Servet 1, 1206 Geneva, Switzerland.
J Pharm Biomed Anal. 2018 Feb 20;150:258-267. doi: 10.1016/j.jpba.2017.12.014. Epub 2017 Dec 12.
Ultratrace analysis of sample components requires excellent analytical performance in terms of limits of quantitation (LOQ). Micro UHPLC coupled to sensitive tandem mass spectrometry provides state of the art solution for such analytical problems. Using on-line SPE with column switching on a micro UHPLC-MS/MS system allowed to decrease LOQ without any complex sample preparation protocol. The presented method is capable of reaching satisfactory low LOQ values for analysis of thirteen different steroid molecules from human plasma without the most commonly used off-line SPE or compound derivatization. Steroids were determined by using two simple sample preparation methods, based on lower and higher plasma steroid concentrations. In the first method, higher analyte concentrations were directly determined after protein precipitation with methanol. The organic phase obtained from the precipitation was diluted with water and directly injected into the LC-MS system. In the second method, low steroid levels were determined by concentrating the organic phase after steroid extraction. In this case, analytes were extracted with ethyl acetate and reconstituted in 90/10 water/acetonitrile following evaporation to dryness. This step provided much lower LOQs, outperforming previously published values. The method has been validated and subsequently applied to clinical laboratory measurement.
对样品成分进行痕量分析需要在定量限 (LOQ) 方面具有出色的分析性能。微超高效液相色谱与灵敏的串联质谱联用为解决此类分析问题提供了最新的解决方案。通过在微超高效液相色谱-串联质谱系统上进行在线 SPE 和柱切换，可以在不使用最常用的离线 SPE 或化合物衍生化的情况下降低 LOQ。所提出的方法能够达到令人满意的低 LOQ 值，用于分析人血浆中的十三种不同的甾体分子，而无需使用最常用的离线 SPE 或化合物衍生化。甾体通过两种简单的样品制备方法进行测定，这两种方法基于较低和较高的血浆甾体浓度。在第一种方法中，直接用甲醇沉淀蛋白质后，直接测定较高的分析物浓度。从沉淀中获得的有机相用稀释水稀释，然后直接注入 LC-MS 系统。在第二种方法中，通过在甾体提取后浓缩有机相来测定低甾体水平。在这种情况下，用乙酸乙酯提取分析物，然后在蒸发至干燥后用 90/10 水/乙腈重新配制。这一步提供了更低的 LOQ，优于先前发表的值。该方法已经过验证，并随后应用于临床实验室测量。
