Clinical Chemistry, Laboratory Medicine of Skåne, Sweden.
Scand J Clin Lab Invest. 2012 Oct;72(6):441-6. doi: 10.3109/00365513.2012.679963. Epub 2012 Aug 30.
Methylmalonic acid (MMA), a sensitive biomarker of functional vitamin B12 deficiency, is commonly determined by gas chromatography-mass spectrometry (GC-MS) or liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods using manual extraction and derivatization of MMA to reduce polarity prior to separation.
In the present study we introduce a semi-automated extraction on a strong anion exchanger, HPLC separation on a BEH-amide column to separate serum MMA from its abundant isoform, succinic acid, followed by MS/MS detection and quantification.
The extraction of MMA plus internal standard provides full recovery and the method is linear between 0.03 μmol/L and 20.0 μmol/L (r(2) = 1.0) with intra-and inter-assay imprecision of 2.2%. Agreement with other laboratories has been demonstrated in external proficiency testing. Compared to both conventional GC-MS and LC-MS/MS methods, the correlation is r(2) > 0.99.
The use of robotic pipetting, elimination of derivatization and improved separation by the BEH-amide column combined with HILIC chromatographic conditions significantly improve sample throughput compared to conventional methods. Using a single pipetting robot and LC-MS/MS instrument, this method is currently performing 180 analyses per day from 10 regional hospitals and several additional distant sites.
甲基丙二酸（MMA）是功能性维生素 B12 缺乏的敏感生物标志物，通常通过气相色谱-质谱联用（GC-MS）或液相色谱-串联质谱（LC-MS/MS）方法测定，这些方法在分离前采用手动提取和 MMA 衍生化来降低其极性。
本研究中，我们采用强阴离子交换剂半自动提取，在 BEH-酰胺柱上进行 HPLC 分离，以分离血清 MMA 与其丰富的同型物琥珀酸，然后进行 MS/MS 检测和定量。
MMA 加内标物的提取可实现完全回收，该方法在 0.03 μmol/L 至 20.0 μmol/L 之间呈线性（r(2) = 1.0），批内和批间精密度为 2.2%。与其他实验室的外部能力验证结果具有一致性。与传统 GC-MS 和 LC-MS/MS 方法相比，相关性 r(2) > 0.99。
与传统方法相比，使用机器人移液、消除衍生化以及 BEH-酰胺柱的改进分离，显著提高了样品通量。使用单一移液机器人和 LC-MS/MS 仪器，该方法目前每天可从 10 家区域医院和其他几个远程站点进行 180 次分析。
