Determination of five nucleosides by LC-MS/MS and the application of the method to quantify N6-methyladenosine level in liver messenger ribonucleic acid of an acetaminophen-induced hepatotoxicity mouse model
Determination of five nucleosides by LC-MS/MS and the application of the method to quantify N6-methyladenosine level in liver messenger ribonucleic acid of an acetaminophen-induced hepatotoxicity mouse model
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LC-MS/MS 测定五种核苷及其在对乙酰氨基酚诱导的肝毒性小鼠模型肝脏信使核糖核酸中 N6-甲基腺苷水平的应用
DOI：
10.1002/jssc.201900041
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发表时间：
2019-08-01
影响因子：
3.1
通讯作者：
Tian, Xin
中科院分区：
文献类型：
--
作者：
Liu, Ruijuan;Zhao, Fang;Tian, Xin
核糖核酸n -6-甲基腺苷甲基化在多种生物过程和疾病中起着重要作用。对乙酰氨基酚引起的肝毒性是临床医生面临的主要挑战之一。迄今为止，尚未研究N(6)甲基腺苷与对乙酰氨基酚诱导的肝毒性之间的联系。本研究建立了同时测定信使核糖核酸中5种核苷（腺苷、尿苷、胞苷、鸟苷和n -6-甲基腺苷）的高效液相色谱-串联质谱法。酶解信使核糖核酸后，核苷样品在Acquity UPLC柱上分离，甲醇和0.02%甲酸水梯度洗脱，Qtrap 4500质谱仪电喷雾电离模式检测。腺苷、尿苷、胞苷和鸟苷的浓度范围为4 ~ 800 ng/mL， n -6-甲基腺苷的浓度范围为0.1 ~ 20 ng/mL。该方法成功地应用于对乙酰氨基酚肝毒性小鼠模型和对照组肝脏信使核糖核酸中n- 6-甲基腺苷水平的测定。本研究为表观遗传学研究中核苷含量的测定提供了一种方法，为研究对乙酰氨基酚诱导的肝毒性中核糖核酸甲基化迈出了第一步，有助于阐明其机制。
Ribonucleic acid N-6-methyladenosine methylation plays an important role in a variety of biological processes and diseases. Acetaminophen-induced hepatotoxicity is one of the major challenges faced by clinicians. To date, the link between N(6)methyladenosine and acetaminophen-induced hepatotoxicity has not been studied. In this study, a simple ultra high performance liquid chromatography with tandem mass spectrometry method was developed for the simultaneous determination of five nucleosides (adenosine, uridine, cytidine, guanosine, and N-6-methyladenosine) in messenger ribonucleic acid. After enzymatic digestion of messenger ribonucleic acid, the nucleosides sample was separated on an Acquity UPLC column with gradient elution using methanol and 0.02% formic acid water, and detected by a Qtrap 4500 mass spectrometer with an electrospray ionization mode. The method was validated over the concentration ranges of 4-800 ng/mL for adenosine, uridine, cytidine, and guanosine and 0.1-20 ng/mL for N-6-methyladenosine. It was successfully applied to the determination of N-6-methyladenosine levels in liver messenger ribonucleic acid in an acetaminophen-induced hepatotoxicity mouse model and a control group. This study offers a method for the determination of nucleoside contents in epigenetic studies and constitutes the first step toward the investigation of ribonucleic acid methylation in acetaminophen-induced hepatotoxicity, which will facilitate the elucidation of its mechanism.