Piraud Monique, Ruet Séverine, Boyer Sylvie, Acquaviva Cécile, Clerc-Renaud Pascale, Cheillan David, Vianey-Saban Christine
Laboratoire des Maladies Héréditaires du Métabolisme et Dépistage Néonatal, Hospices Civils de Lyon, Centre de Biologie Est, Bron, France.
Methods Mol Biol. 2011;708:25-53. doi: 10.1007/978-1-61737-985-7_2.
The diagnosis of inherited metabolic disorders of amino acid (AA) metabolism is based on the qualitative and/or the quantitative analysis of AAs, mainly in blood and urine. For years, the most widespread technique in use was ion-exchange chromatography followed by post-column derivatization with ninhydrin, a method which is the basis of numerous automated AA analyzers with a throughput of about eight samples/day. The emergence of tandem mass spectrometry (MS/MS) coupled to liquid chromatography (LC) has made possible the measurement of many metabolites for the diagnosis of inborn errors of metabolism. The LC-MS/MS method described here allows the clinical diagnosis of AA disorders by analysis of underivatized AAs and derivative molecules in various biological samples prepared by methanol precipitation. AAs are separated by ion-pairing reversed-phase LC, using perfluorocarboxylic acid as an ion-pairing agent. Each AA is detected in MS/MS-positive ionization mode by its specific transition. The method allows the analysis of about 40 biological samples/day.
氨基酸（AA）代谢遗传性疾病的诊断主要基于对血液和尿液中氨基酸的定性和/或定量分析。多年来，使用最广泛的技术是离子交换色谱法，随后用茚三酮进行柱后衍生化，这是许多自动化氨基酸分析仪的基础方法，每天通量约为八个样本。串联质谱（MS/MS）与液相色谱（LC）联用的出现，使得测量多种代谢物以诊断先天性代谢缺陷成为可能。本文所述的LC-MS/MS方法通过分析经甲醇沉淀制备的各种生物样品中的未衍生化氨基酸和衍生分子，实现氨基酸疾病的临床诊断。氨基酸通过离子对反相液相色谱进行分离，使用全氟羧酸作为离子对试剂。每种氨基酸在MS/MS正离子化模式下通过其特定的跃迁进行检测。该方法每天可分析约40个生物样品。
