Department of Chemistry, University of Louisville, 2210 South Brook Street, Louisville, KY, 40208, USA.
Alcohol Research Center, University of Louisville, Louisville, KY, 40208, USA.
J Am Soc Mass Spectrom. 2019 Jun;30(6):987-1000. doi: 10.1007/s13361-019-02140-7. Epub 2019 Mar 7.
We report a reverse phase chromatography mass spectrometry (LC-MS) method for simultaneous quantification of nucleosides and nucleotides from biological samples, where compound identification was achieved by a tier-wise approach and compound quantification was achieved via external calibration. A total of 65 authentic standards of nucleosides and nucleotides were used for the platform development. The limit of detection (LOD) of those compounds ranged from 0.05 nmol/L to 1.25 μmol/L, and their limit of quantification (LOQ) ranged from 0.10 nmol/L to 2.50 μmol/L. Using the developed method, nucleosides and nucleotides from human plasma, human urine, and rat liver were quantified. Seventy-nine nucleosides and nucleotides were identified from human urine and 28 of them were quantified with concentrations of 13.0 nmol/L-151 μmol/L. Fifty-five nucleosides and nucleotides were identified from human plasma and 22 of them were quantified with concentrations of 1.21 nmol/L-8.54 μmol/L. Fifty-one nucleosides and nucleotides were identified from rat liver and 23 were quantified with concentrations of 1.03 nmol/L-31.7 μmol/L. These results demonstrate that the developed method can be used to investigate the concentration change of nucleosides and nucleotides in biological samples for the purposes of biomarker discovery or elucidation of disease mechanisms.
我们报道了一种反相色谱-质谱（LC-MS）方法，用于同时定量生物样品中的核苷和核苷酸，其中通过分级方法实现化合物鉴定，通过外部校准实现化合物定量。总共使用了 65 种核苷和核苷酸的真实标准品用于平台开发。这些化合物的检测限（LOD）范围为 0.05 nmol/L 至 1.25 μmol/L，定量限（LOQ）范围为 0.10 nmol/L 至 2.50 μmol/L。使用开发的方法，定量了人血浆、人尿和大鼠肝中的核苷和核苷酸。从人尿中鉴定出 79 种核苷和核苷酸，其中 28 种用 13.0 nmol/L-151 μmol/L 的浓度定量。从人血浆中鉴定出 55 种核苷和核苷酸，其中 22 种用 1.21 nmol/L-8.54 μmol/L 的浓度定量。从大鼠肝中鉴定出 51 种核苷和核苷酸，其中 23 种用 1.03 nmol/L-31.7 μmol/L 的浓度定量。这些结果表明，该方法可用于研究生物样品中核苷和核苷酸浓度的变化，以用于生物标志物的发现或疾病机制的阐明。
