Quantitative profiling of biomarkers related to B-vitamin status, tryptophan metabolism and inflammation in human plasma by liquid chromatography/tandem mass spectrometry
Quantitative profiling of biomarkers related to B-vitamin status, tryptophan metabolism and inflammation in human plasma by liquid chromatography/tandem mass spectrometry
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DOI：
10.1002/rcm.4013
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发表时间：
2009-05-01
影响因子：
2
通讯作者：
Ueland, Per M.
中科院分区：
文献类型：
--
作者：
Midttun, Oivind;Hustad, Steinar;Ueland, Per M.
维生素B-2和B-6是参与色氨酸和同型半胱氨酸代谢的酶反应的辅助因子。这些维生素和氨基酸的血浆浓度与吸烟和炎症有关，也与免疫激活的其他标志相关。由于缺乏合适的分析方法，对这些关系的大规模研究一直受到阻碍。测定方法包括核黄素、5‘-磷酸吡哆醛、4-吡哆酸、吡哆醇和吡哆胺等5种形式的维生素B-6、色氨酸和6种色氨酸代谢物(犬尿氨酸、犬尿酸、邻氨基苯甲酸、3-羟基犬尿氨酸、黄原酸和3-羟基邻氨基苯甲酸)、半胱硫氨酸、新喋呤和可替宁。将含有13个同位素标记内标的三氯乙酸加入60亩L血浆中，混合物离心，所得上清液用于分析。在稳定键C8色谱柱上，以乙腈、七氟丁酸和高浓度(650mmo1/L)冰醋酸为流动相，5min内分离，电喷雾电离串联质谱仪(ESI-MS/MS)检测。流动相确保了所有分析物的充分分离和高电离效率。回收率为75-123%，日内和日间变异系数分别为2.5-9.5%和5.4-16.9%。检出限为0.05~7nmol/L。该方法可定量测定16种与维生素B状态和炎症有关的内源性血浆浓度，可能在大规模流行病学研究中有用。版权所有(C)2009 John Wiley&Sons，Ltd.
Vitamins B-2 and B-6 serve as cofactors in enzymatic reactions involved in tryptophan and homocysteine metabolism. Plasma concentrations of these vitamins and amino acids are related to smoking and inflammation, and correlate with other markers of immune activation. Large-scale studies of these relations have been hampered by lack of suitable analytical methods. The assay described includes riboflavin, five vitamin B-6 forms (pyridoxal 5'-phosphate, pyridoxal, 4-pyridoxic acid, pyridoxine and pyridoxamine), tryptophan and six tryptophan metabolites (kynurenine, kynurenic acid, anthranilic acid, 3-hydroxykynurenine, xanthurenic acid and 3-hydroxyanthranilic acid), cystathionine, neopterin and cotinine. Trichloroacetic acid containing 13 isotope-labelled internal standards was added to 60 mu L of plasma, the mixture was centrifuged, and the resulting supernatant used for analysis. The analytes were separated within 5 min on a stable-bond C8 column by a gradient-type mobile phase containing acetonitrile, heptafluorobutyric acid and high concentration (650 mmol/L) of acetic acid, and detected using electrospray ionization tandem mass spectrometry (ESI-MS/MS). The mobile phase ensured sufficient separation and high ionization efficiency of all analytes. Recoveries were 75-123% and within-day and between-day coefficients of variance (CVs) were 2.5-9.5% and 5.4-16.9%, respectively. Limits of detection ranged from 0.05 to 7 nmol/L. The method enables quantification of endogenous plasma concentrations of 16 analytes related to B-vitamin status and inflammation, and may prove useful in large-scale epidemiological studies. Copyright (C) 2009 John Wiley & Sons, Ltd.