衍生化后通过 LC-MS/MS 对人体血浆中的甲基丙二酸进行定量分析

IF 2.8 3区医学 Q2 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography B Pub Date : 2024-11-01 DOI:10.1016/j.jchromb.2024.124359

Danhong Shen , Wei Chen , Jindi Liu , Yanhua Liu , Hongjun Shi

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引用次数: 0

摘要

甲基丙二酸（MMA）是维生素 B12 的一种反向生物标记物，在临床实践中的应用越来越广泛。然而，甲基丙二酸的灵敏度低且易受异构体的强烈干扰，这给色谱分析带来了挑战。我们开发了一种室温下快速衍生血浆 MMA 的方法，利用 N-（3-二甲基氨基丙基）-N'-乙基碳二亚胺盐酸盐（EDC）和 2,2,2-三氟乙胺盐酸盐（TFEA）将其转化为相应的 2,2,2-三氟乙酰胺衍生物。酰胺化过程在 10 分钟内完成，然后用三氯乙酸对酰胺进行蛋白质沉淀萃取，再进行液相色谱-串联质谱（LC-MS/MS）分析。该技术显著提高了 MMA 在色谱分析中的信噪比。衍生化后的 MMA 在 42.4-2711.9 nmol/L 浓度范围内呈良好的线性关系，相关系数 (R2) 为 0.9990。重复测量的日内精密度和日间精密度分别为 2.4 % 至 4.4 % 和 2.6 % 至 2.8 %，回收率为 97.9 % 至 100.1 %。

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Quantitative analysis of methylmalonic acid in human plasma by LC-MS/MS after derivatization

Methylmalonic acid (MMA) is a reverse biomarker of vitamin B12 that is increasingly utilized in clinical practice. However, its low sensitivity and susceptibility to strong interference from isomer present chromatographic challenges. We have developed a rapid derivatization method for plasma MMA at room temperature, converting it to the corresponding 2,2,2-trifluoroethylamide derivative using N-(3-dimethylaminopropyl)-N’-ethylcarbodiimide hydrochloride (EDC) and 2,2,2-trifluoroethylamine hydrochloride (TFEA). Amidization was completed within 10 min, followed by protein precipitation extraction of the amides with trichloroacetic acid for Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. This technique notably enhanced the signal-to-noise ratio of MMA in chromatography. The derivatized MMA exhibited excellent linearity within a concentration range of 42.4–2711.9 nmol/L, with a correlation coefficient (R²) of 0.9990. The intraday and interday precision of replicate measurements ranged from 2.4 % to 4.4 % and 2.6 % to 2.8 %, respectively, while the recovery fell between 97.9 % and 100.1 %.

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来源期刊

Journal of Chromatography B 医学-分析化学

CiteScore

5.60

自引率

3.30%

发文量

306

审稿时长

44 days

期刊介绍： The Journal of Chromatography B publishes papers on developments in separation science relevant to biology and biomedical research including both fundamental advances and applications. Analytical techniques which may be considered include the various facets of chromatography, electrophoresis and related methods, affinity and immunoaffinity-based methodologies, hyphenated and other multi-dimensional techniques, and microanalytical approaches. The journal also considers articles reporting developments in sample preparation, detection techniques including mass spectrometry, and data handling and analysis. Developments related to preparative separations for the isolation and purification of components of biological systems may be published, including chromatographic and electrophoretic methods, affinity separations, field flow fractionation and other preparative approaches. Applications to the analysis of biological systems and samples will be considered when the analytical science contains a significant element of novelty, e.g. a new approach to the separation of a compound, novel combination of analytical techniques, or significantly improved analytical performance.