Development and evaluation of a hydrophilic interaction liquid chromatography-MS/MS method to quantify 19 nucleobases and nucleosides in rat plasma
Development and evaluation of a hydrophilic interaction liquid chromatography-MS/MS method to quantify 19 nucleobases and nucleosides in rat plasma
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DOI：
10.1002/bmc.3860
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发表时间：
2017-04-01
影响因子：
1.8
通讯作者：
Tang, Dao-quan
中科院分区：
文献类型：
--
作者：
Du, Yan;Li, Yin-jie;Tang, Dao-quan
作为生物体内必需的内源性化合物，核碱基和核苷在生物体内发挥着多种功能。本研究提出了一种新的亲水性相互作用液相色谱串联质谱法，用于同时定量大鼠血浆中19种核碱基和核苷。在样品制备中，根据分析物的色谱谱和离子响应对15种蛋白质沉淀剂进行了评价。分别采用反相液相色谱和亲水性相互作用液相色谱模式进行色谱分离优化；每种分离方式包括两个固定相不同的测试柱。采用色谱剖面和半宽（W-1/2）、容量因子(K)、尾尾因子（f(t)）等参数评价分离效率。此外，还研究了两种流动相体系的组成和最佳缓冲体系中添加剂的浓度。所建立的方法得到了充分的验证，并成功地定量测定了正常和糖尿病肾病（DN）大鼠血浆中的19种核碱基和核苷。正常大鼠与DN大鼠血浆中胞嘧啶、黄嘌呤、胸腺嘧啶、腺苷、鸟苷、肌苷和8-羟基-2-脱氧鸟苷水平差异有统计学意义。这些信息可能为发现DN的潜在生物标志物提供有用的参考。
As essential endogenous compounds, nucleobases and nucleosides fulfill various functions in living organisms. This study presents the development and validation of a new hydrophilic interaction liquid chromatography tandem mass spectrometry method for simultaneous quantification of 19 nucleobases and nucleosides in rat plasma. For the sample preparation, 15 kinds of protein precipitants were evaluated according to the chromatographic profile and ion response of analytes. The optimization of chromatographic separation was respectively performed using reversed-phase liquid chromatography and hydrophilic interaction liquid chromatography mode; each separation mode included two test columns with different stationary phases. The chromatographic profile and parameters such as half-width (W-1/2), capacity factor (K) and tailing factor (f(t)) were used to evaluate the separation efficiencies. Furthermore, the adopted composition of two mobile phase systems and the concentrations of the additives in the optimum buffer system were also investigated. The developed method was fully validated and successfully applied quantitatively to determine 19 nucleobases and nucleosides in plasma from normal and diabetic nephropathy (DN) rats. Significant differences between normal and DN rats were found in plasma levels of cytosine, xanthine, thymidine, adenosine, guanosine, inosine and 8-hydroxy-2-deoxyguanosine. This information may provide a useful reference for the discovery of potential biomarkers of DN.