Mendiara Isabel, Bentayeb Karim, Nerín Cristina, Domeño Celia
Department of Analytical Chemistry, EINA-University of Zaragoza, María de Luna St. 3, E-50018 Zaragoza, Spain.
Talanta. 2015 Jan;132:690-7. doi: 10.1016/j.talanta.2014.10.029. Epub 2014 Oct 23.
An automated method for analyzing free non-cholesterol sterols in human serum using online solid phase extraction-liquid chromatography-mass spectrometry is proposed herein. The method allows the determination of three phytosterols (sitosterol, stigmasterol and campesterol) and two cholesterol precursors (desmosterol and lanosterol). The analysis of sterols in human serum is critical in the study of cholesterol-related disorders, such as inherited familial hypercholesterolemias. Special effort was made to isolate the analytes from the serum lipoproteins, their natural conveyance through the bloodstream. The sample treatment consisted of a Bligh-Dyer extraction followed by dilution of the extract. This treatment allowed the sample to be injected into the online system and ensured the correct detection of the analytes, while avoiding the matrix effects commonly related to serum samples. The analytical performance showed linear ranges that covered two orders of magnitude, with correlation coefficients above 0.99. Limits of detection and quantification ranged from 0.2 ng/mL to 13 ng/mL and from 1.0 ng/mL to 43 ng/mL, respectively. Recovery when spiking serum with a half or a tenth of the average concentration reported in human serum ranged from 99% to 111% and from 102% to 120%, respectively. Intra-day precision and inter-day precision were below 20%.
本文提出了一种使用在线固相萃取-液相色谱-质谱联用技术分析人血清中游离非胆固醇甾醇的自动化方法。该方法可测定三种植物甾醇（谷甾醇、豆甾醇和菜油甾醇）以及两种胆固醇前体（羊毛甾醇和7-脱氢胆固醇）。人血清中甾醇的分析对于研究与胆固醇相关的疾病，如遗传性家族性高胆固醇血症至关重要。我们特别致力于从血清脂蛋白（它们在血液中的天然载体）中分离分析物。样品处理包括采用布利-戴尔萃取法，然后对提取物进行稀释。这种处理方式使得样品能够注入在线系统，并确保对分析物进行正确检测，同时避免了通常与血清样品相关的基质效应。分析性能显示线性范围覆盖两个数量级，相关系数高于0.99。检测限和定量限分别为0.2 ng/mL至13 ng/mL和1.0 ng/mL至43 ng/mL。当向血清中加入人血清中报告平均浓度的一半或十分之一时，回收率分别为99%至111%和102%至120%。日内精密度和日间精密度均低于20%。
