Sharma Sushma, Kong Ziqing, Jia Shaodong, Tran Phong, Nilsson Anna Karin, Chabes Andrei
Department of Medical Biochemistry and Biophysics, Umeå University, Umeå, Sweden.
Methods Mol Biol. 2023;2615:267-280. doi: 10.1007/978-1-0716-2922-2_19.
Defects in deoxyribonucleoside triphosphate (dNTP) metabolism are associated with a number of mitochondrial DNA (mtDNA) depletion syndromes (MDS). These disorders affect the muscles, liver, and brain, and the concentrations of dNTPs in these tissues are already normally low and are, therefore, difficult to measure. Thus, information about the concentrations of dNTPs in tissues of healthy animals and animals with MDS are important for mechanistic studies of mtDNA replication, analysis of disease progression, and the development of therapeutic interventions. Here, we present a sensitive method for the simultaneous analysis of all four dNTPs as well as all four ribonucleoside triphosphates (NTPs) in mouse muscles using hydrophilic interaction liquid chromatography coupled with triple quadrupole mass spectrometry. The simultaneous detection of NTPs allows them to be used as internal standards for the normalization of dNTP concentrations. The method can be applied for measuring dNTP and NTP pools in other tissues and organisms.
脱氧核糖核苷三磷酸（dNTP）代谢缺陷与多种线粒体DNA（mtDNA）耗竭综合征（MDS）相关。这些疾病会影响肌肉、肝脏和大脑，而这些组织中dNTP的浓度通常已经很低，因此难以测量。因此，关于健康动物和患有MDS的动物组织中dNTP浓度的信息，对于mtDNA复制的机制研究、疾病进展分析以及治疗干预的开发都很重要。在此，我们提出了一种灵敏的方法，利用亲水相互作用液相色谱与三重四极杆质谱联用，同时分析小鼠肌肉中的所有四种dNTP以及所有四种核糖核苷三磷酸（NTP）。同时检测NTP可使其用作dNTP浓度标准化的内标。该方法可用于测量其他组织和生物体中的dNTP和NTP库。
