Determination of choline, betaine, and dimethylglycine in plasma by a high-throughput method based on normal-phase chromatography-tandem mass spectrometry
Determination of choline, betaine, and dimethylglycine in plasma by a high-throughput method based on normal-phase chromatography-tandem mass spectrometry
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DOI：
10.1373/49.2.286
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发表时间：
2003-02-01
影响因子：
9.3
通讯作者：
Lien, EA
中科院分区：
文献类型：
--
作者：
Holm, PI;Ueland, PM;Lien, EA
背景：季铵化合物、胆碱、甜菜碱和二甲基甘氨酸（DMG）存在于与神经递质和膜磷脂合成、同型半胱氨酸再甲基化以及叶酸状态相关的代谢途径中。缺乏一种方便、高通量的方法来测定这些化合物，阻碍了基于人群的研究它们与生活方式、营养和慢性疾病的可能关联。将血清或血浆样品与三体积含有d(9)-胆碱和d(9)-甜菜碱作为内标的乙腈混合去蛋白。我们使用正相硅胶柱分离胆碱（保留时间为2.8 min）、甜菜碱（1.3 min）、DMG （1.15 min）和内标，在多反应监测模式下，利用分子跃迁m/z 104—bbb60（胆碱）、m/z 113—bbb69 （d(9)-胆碱）、m/z 118—bbb59（甜菜碱）、m/z 127—bbb68 （d(9)-甜菜碱）和m/z 104—>58 (DMG)，通过串联质谱检测出正离子。结果：三种代谢物在0.4 ~ 400 μ mol/L范围内呈线性关系，检测下限（信噪比= 5）小于或等于0.3 μ mol/L。日内、日间不精确度（cv）分别为2.1 ~ 7.2%和3.5 ~ 8.8%。分析回收率为87 ~ 105%。60名健康献血者的空腹血浆浓度（中位数，25 -75百分位数）为胆碱8.0 (7.0-9.3)μ mol/L，甜菜碱31.7 (27.0-41.1)μ mol/L, DMG 1.66 (1.30-2.02) μ mol/L。在吃清淡早餐的个体中，所有三种代谢物的血浆浓度显著高于禁食个体（25-30%）。结论：首次建立了人血浆或血清中胆碱、甜菜碱和DMG的联合测定方法。该方法的特点是样品制备简单，无衍生化，通量高，不精确（CV）。
Background: The quaternary ammonium compounds, choline and betaine, and dimethylglycine (DMG) reside along a metabolic pathway linked to the synthesis of neurotransmitters and membrane phospholipids and to homocysteine remethylation and, therefore, folate status. Lack of a convenient, high-throughput method for the determination of these compounds has prevented population-based studies of their possible associations with lifestyle, nutrition, and chronic diseases.Methods. Serum or plasma samples were deproteinized by mixing with three volumes of acetonitrile that contained d(9)-choline and d(9)-betaine as internal standards. We used a normal-phase silica column for the separation of choline (retention time, 2.8 min), betaine (1.3 min), DMG (1.15 min), and internal standards, which were detected as positive ions by tandem mass spectroscopy in the multiple-reaction monitoring mode, using the molecular transitions m/z 104-->60 (choline), m/z 113-->69 (d(9)-choline), m/z 118-->59 (betaine), m/z 127-->68 (d(9)-betaine), and m/z 104-->58 (DMG).Results: For all three metabolites, the assay was linear in the range 0.4-400 mumol/L, and the lower limit of the detection (signal-to-noise ratio = 5) was less than or equal to0.3 mumol/L. The within- and between-day imprecision (CVs) was 2.1-7.2% and 3.5-8.8%, respectively. The analytical recovery was 87-105%. The fasting plasma concentrations (median, 25th-75th percentiles) were 8.0 (7.0-9.3) mumol/L for choline, 31.7 (27.0-41.1) mumol/L for betaine, and 1.66 (1.30-2.02) mumol/L for DMG in 60 healthy blood donors. In individuals who had eaten a light breakfast, plasma concentrations of all three metabolites were significantly (25-30%) higher than in fasting individuals.Conclusion: This is the first method for the combined measurement of choline, betaine, and DMG in human plasma or serum. The assay is characterized by simple sample preparation, no derivatization, high throughput, imprecision (CV)