Center for Metabolomics and Bioanalysis (CEMBIO), Facultad de Farmacia, Campus Monteprincipe, Universidad CEU San Pablo, 28668 Boadilla del Monte, Madrid, Spain.
J Chromatogr A. 2013 Aug 23;1304:52-60. doi: 10.1016/j.chroma.2013.07.029. Epub 2013 Jul 11.
The aim of metabolomics studies is the comprehensive and quantitative analysis of all metabolites in a cell, tissue or organism. This approach requires sample preparation methods to be fast, reproducible and able to extract a wide range of analytes with different polarities, as well as analytical platforms able to detect the extracted metabolites. Recently, we have developed a one-step extraction method consisting of a lipophilic and hydrophilic layer within a single vial insert, in-vial dual extraction (IVDE). In order to check possible application of this method to real biological case, analysis of plasma samples obtained from three streptozotocin-induced diabetic and three control rats was performed. Analytical validity of the method was proved by the calculation (in quality control samples) of relative standard deviation (RSD) for detected metabolites. The percentage of metabolites with RSD<30% was 93% for Fatty acyls, 80% for Glycerolipids, 93% for Glycerophospholipids, 68% for Sterol lipids, and 91% for Sphingolipids. IVDE allowed for selection of more than 600 different features discriminating two studied groups. For around 40% of these masses putative identification was possible. Adequate, with several considerations described within this paper, application of IVDE method enables wide metabolite coverage from a single 20μL plasma aliquot. Within the features putatively identified, glycerolipids and glycerophospholipids arose as the most important groups of compounds discriminating diabetic rats from controls. All discriminating metabolites give an idea of the large metabolic differences that can be present in non-controlled type 1 diabetes.
代谢组学研究的目的是全面、定量地分析细胞、组织或生物体中的所有代谢物。这种方法需要快速、可重复的样品制备方法，并且能够提取具有不同极性的广泛分析物，以及能够检测提取代谢物的分析平台。最近，我们开发了一种一步提取方法，即在单个小瓶插入物内包含亲脂层和亲水层，即小瓶内双重提取（IVDE）。为了检查这种方法在实际生物案例中的可能应用，对来自 3 只链脲佐菌素诱导的糖尿病大鼠和 3 只对照大鼠的血浆样本进行了分析。通过计算（在质控样品中）检测到的代谢物的相对标准偏差（RSD），证明了该方法的分析有效性。RSD<30%的代谢物百分比为：脂肪酸 93%、甘油酯 80%、甘油磷酯 93%、固醇脂 68%、鞘脂 91%。IVDE 允许选择超过 600 个不同的特征来区分两个研究组。其中约 40%的物质可以进行假定鉴定。在本文中描述了一些考虑因素的情况下，适当应用 IVDE 方法可以从单个 20μL 血浆等分试样中实现广泛的代谢物覆盖。在假定鉴定的特征中，甘油酯和甘油磷酯是区分糖尿病大鼠和对照大鼠的最重要化合物组。所有区分代谢物都说明了在未控制的 1 型糖尿病中可能存在的大量代谢差异。
