Detection of Human Urinary 5-Hydroxymethylcytosine by Stable Isotope Dilution HPLC-MS/MS Analysis
文献类型：期刊论文
| 作者 | Yin, Ruichuan; Mo, Jiezhen; Lu, Meiling; Wang, Hailin |
| 刊名 | ANALYTICAL CHEMISTRY
 |
| 出版日期 | 2015-02-03 |
| 卷号 | 87期号:3页码:1846-1852 |
| 英文摘要 | The sixth DNA base 5-hydroxymethylcytosine (5hmC) is the major oxidation product of the epigenetic modification 5-methylcytosine (5mC), mediating DNA demethylation in mammals. Reduced 5hmC levels are found to be linked with various tumors and neurological diseases; therefore, 5hmC is an emerging biomarker for disease diagnosis, treatment, and prognosis. Due to its advantages of being sterile, easily accessible in large volumes, and noninvasive to patients, urine is a favored diagnostic biofluid for 5hmC analysis. Here we developed an accurate, sensitive, and specific assay for quantification of 5mC, 5hmC, and other DNA demethylation intermediates in human urine. The urinary samples were desalted and enriched using off-line solid-phase extraction, followed by stable isotope dilution HPLC-MS/MS analysis for 5hmC and 5mC. By the use of ammonium bicarbonate (NH4HCO3) as an additive to the mobile phase, we improved the online-coupled MS/MS detection of 5mC, 5hmC, and 5-formylcytosine (5fC) by 1.8-14.3 times. The recovery of the method is approximately 100% for 5hmC, and 70-90% for 5mC. The relative standard deviation (RSD) of the interday precision is about 2.9-10.6%, and that of the intraday precision is about 1.4-7.7%. By the analysis of 13 volunteers using the developed method, we for the first time demonstrate the presence of 5hmC in human urine. Unexpectedly, we observed that the level of 5hmC (22.6 +/- 13.7 nmol/L) is comparable to that of its precursor 5mC (52.4 +/- 50.2 nmol/L) in human urine. Since the abundance of 5hmC (as a rare DNA base) is 1 or 2 orders of magnitude lower than 5mC in genomic DNA, our finding probably implicates a much higher turnover of 5hmC than 5mC in mammalian genomic DNA and underscores the importance of DNA demethylation in daily life. |
| 研究领域[WOS] | Chemistry, Analytical |
| WOS记录号 | WOS:000349059000059 |
| 公开日期 | 2016-03-15 |
| 源URL | [http://ir.rcees.ac.cn/handle/311016/32548]  |
| 专题 | 生态环境研究中心_环境化学与生态毒理学国家重点实验室 |
| 推荐引用方式 GB/T 7714 | Yin, Ruichuan,Mo, Jiezhen,Lu, Meiling,et al. Detection of Human Urinary 5-Hydroxymethylcytosine by Stable Isotope Dilution HPLC-MS/MS Analysis[J]. ANALYTICAL CHEMISTRY,2015,87(3):1846-1852. |
| APA | Yin, Ruichuan,Mo, Jiezhen,Lu, Meiling,&Wang, Hailin.(2015).Detection of Human Urinary 5-Hydroxymethylcytosine by Stable Isotope Dilution HPLC-MS/MS Analysis.ANALYTICAL CHEMISTRY,87(3),1846-1852. |
| MLA | Yin, Ruichuan,et al."Detection of Human Urinary 5-Hydroxymethylcytosine by Stable Isotope Dilution HPLC-MS/MS Analysis".ANALYTICAL CHEMISTRY 87.3(2015):1846-1852. |
入库方式： OAI收割
来源：生态环境研究中心
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