Development and validation of a single analytical method for the determination of tryptophan, and its kynurenine metabolites in rat plasma
Development and validation of a single analytical method for the determination of tryptophan, and its kynurenine metabolites in rat plasma
复制标题
DOI：
10.1016/j.jchromb.2012.04.030
复制
发表时间：
2012-06-01
影响因子：
3
通讯作者：
Harvey, Brian H.
中科院分区：
文献类型：
--
作者：
Moeller, Marisa;Du Preez, Jan L.;Harvey, Brian H.
在一个单一的实验方案中，以高精度和高可靠性监测大量样品中犬尿氨酸途径的活性是非常有益的。建立了一种快速测定大鼠血浆中色氨酸、犬尿氨酸、犬尿酸（KYNA）、3-羟基苯甲酸（30haa）、邻氨基苯酸和喹啉酸（QA）的固相萃取-液相色谱-电喷雾串联质谱分析方法。我们还在该方法中评估了吡啶酸（PA），但它存在不可接受的验证参数。该方法采用固相萃取法进行预纯化，然后采用C18反相色谱法进行色谱分离。质谱检测采用质谱仪进行正、负电喷雾电离；流速为0.2 mL/min，进样量为10 μ l，包括样品清理在内的总运行时间为12 min。结果表明，该方法线性良好（R-2 > 0.95），验证参数均在可接受范围内。该技术还表明，在断奶后社会隔离饲养的大鼠中，血浆色氨酸、犬尿氨酸、邻氨基苯酸和QA显著升高，KYNA显著降低，这是一种被认为与抑郁症和精神分裂症相关的动物模型。因此，该方法可用于LC-MS/MS准确准确地测量血浆中犬尿氨酸途径的代谢物，从而有助于实现药理学和诊断研究的新机遇。(C) 2012 Elsevier B.V.版权所有
It is highly beneficial to monitor the activity of the kynurenine pathway in a large series of samples with high accuracy and reliability in a single experimental protocol. We have developed a rapid specific solid-phase extraction (SPE)-liquid chromatography-electrospray ionization tandem mass spectrometry method for assaying tryptophan, kynurenine, kynurenic acid (KYNA),3-hydroxyanthranilic acid (3OHAA), anthranilic acid and quinolinic acid (QA) in rat plasma. We also evaluated picolinic acid (PA) in this method, but it presented with unacceptable validation parameters. The assay involves pre-purification by SPE followed by chromatographic separation by C18 reversed phase chromatography. Mass spectrometric detection was performed using a mass spectrometer in positive and negative electrospray ionization; with a flow rate of 0.2 mL/min and an injection volume of 10 mu L. Total run time including sample cleanup was 12 min. The assay method was found to be linear (R-2 > 0.95) and all the validation parameters were within acceptance range. The developed technique also demonstrated a significant elevation in plasma tryptophan, kynurenine, anthranilic acid and QA, and a significant decrease in KYNA, in rats subjected to post-weaning social isolation rearing, a putative animal model of relevance for depression and schizophrenia. This method can therefore be applied to measure metabolites of the kynurenine pathway in plasma accurately and precisely by LC-MS/MS, thereby helping to realize new opportunities in pharmacological and diagnostic research. (C) 2012 Elsevier B.V. All rights reserved.