Vito Biomedical Research Institute, Hangzhou 310024, Zhejiang, China.
Vito Biomedical Research Institute, Hangzhou 310024, Zhejiang, China.
J Chromatogr B Analyt Technol Biomed Life Sci. 2024 Nov 1;1248:124359. doi: 10.1016/j.jchromb.2024.124359. Epub 2024 Oct 31.
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 (R) 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 %.
甲基丙二酸（MMA）是维生素 B12 的反向生物标志物，在临床实践中越来越多地被使用。然而，其低灵敏度和易受异构体的强烈干扰，对色谱分析提出了挑战。我们开发了一种在室温下快速衍生血浆 MMA 的方法，使用 N-(3-二甲氨基丙基)-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。酰胺化在 10 分钟内完成，然后用三氯乙酸沉淀提取酰胺，用于液相色谱-串联质谱（LC-MS/MS）分析。该技术显著提高了 MMA 在色谱中的信噪比。衍生化的 MMA 在 42.4-2711.9 nmol/L 的浓度范围内表现出优异的线性，相关系数（R）为 0.9990。重复测量的日内和日间精密度分别在 2.4%至 4.4%和 2.6%至 2.8%之间，而回收率在 97.9%至 100.1%之间。
