Bajad Sunil, Shulaev Vladimir
Sutro Biopharma Inc., South San Francisco, CA, USA.
Methods Mol Biol. 2011;708:213-28. doi: 10.1007/978-1-61737-985-7_13.
Metabolomics involves qualitative and/or quantitative analysis of hundreds of metabolites in a complex sample. As most of the metabolites are polar in nature (for example, amino acids, nucleotides, carboxylic acids), their chromatographic separation and analysis present a difficult challenge. Recently, hydrophilic interaction chromatography (HILIC) has become a useful tool for analysis of such polar molecules. In this chapter, we present a simple HILIC LC-MS/MS method for targeted multiple reaction monitoring (MRM)-based relative quantitation of hundreds of polar metabolites in a complex sample. The method uses an aminopropyl column with acetonitrile as weak solvent and 20 mM ammonium acetate buffer (pH 9.4) as strong solvent. The method does not use any ion pairing reagent making it suitable for analysis in both positive and negative ionization mode using a simple LC-MS/MS instrument setup. The method has been thoroughly tested, validated, and applied to a variety of samples such as bacteria, yeast, plants, human body fluids, and cell cultures.
代谢组学涉及对复杂样品中数百种代谢物进行定性和/或定量分析。由于大多数代谢物本质上是极性的（例如氨基酸、核苷酸、羧酸），它们的色谱分离和分析面临着艰巨的挑战。最近，亲水相互作用色谱法（HILIC）已成为分析此类极性分子的有用工具。在本章中，我们介绍了一种简单的HILIC液相色谱-串联质谱法，用于基于靶向多反应监测（MRM）对复杂样品中数百种极性代谢物进行相对定量。该方法使用氨基丙基柱，以乙腈作为弱溶剂，20 mM乙酸铵缓冲液（pH 9.4）作为强溶剂。该方法不使用任何离子对试剂，使其适用于使用简单的液相色谱-串联质谱仪器设置在正离子和负离子模式下进行分析。该方法已经过全面测试、验证，并应用于多种样品，如细菌、酵母、植物、人体体液和细胞培养物。
