On-line trapping/capillary hydrophilic-interaction liquid chromatography/mass spectrometry for sensitive determination of RNA modifications from human blood

• Published in: Chinese chemical letters Vol. 30; no. 3; pp. 553 - 557
• Main Authors: Qi, Chubo, Jiang, Hanpeng, Xiong, Jun, Yuan, Bifeng, Feng, Yuqi
• Format: Journal Article
• Language: English
• Published: Elsevier B.V 01.03.2019; Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), Department of Chemistry, Wuhan University, Wuhan 430072, China; Hubei Cancer Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430079, China%Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), Department of Chemistry, Wuhan University, Wuhan 430072, China
• Subjects: 5-Methylcytosine; Capillary monolithic column; Hydrophilic-interaction liquid chromatography; Mass spectrometry; N6-Methyladenosine; Capillary monolithic column; N6-Methyladenosine; Mass spectrometry; 5-Methylcytosine; Hydrophilic-interaction liquid chromatography
• ISSN: 1001-8417; 1878-5964
• DOI: 10.1016/j.cclet.2018.11.029

With the developed on-line trapping/cHILIC/MS analytical platform, the detection limits of RNA modifications of m6A and 5-mC can reach to 0.06 fmol and 0.10 fmol. We then investigated the contents of m6A and 5-mC in human blood RNA from healthy persons at the age of 6–14 and 60–68 years. Our results showed that both m6A and 5-mC contents were significantly decreased in elder persons, suggesting the RNA modifications of m6A and 5-mC are correlated to aging. [Display omitted] RNA modification has recently been proposed to play important roles in biological regulation. The detection and quantification of RNA modifications generally are challenging tasks since most of the modifications exist in low abundance in vivo. Here we developed an on-line trapping/capillary hydrophilic-interaction liquid chromatography/electrospray ionization-mass spectrometry (on-line trapping/cHILIC/MS) method for sensitive and simultaneous quantification of RNA modifications of N6-methyladenosine (m6A) and 5-methylcytosine (5-mC) from human blood. The hydrophilic organic-silica hybrid monolith was prepared using sol-gel combined with “thiol-ene” click reaction for the separation of nucleosides. A poly(MAA-co-EGDMA) monolithic capillary was used as the on-line trapping column. With the developed on-line trapping/cHILIC/MS analytical platform, the detection limits of m6A and 5-mC can reach to 0.06 fmol and 0.10 fmol. We then investigated the contents of m6A and 5-mC in human blood RNA from healthy persons at the age of 6–14 and 60–68 years. Our results showed that both m6A and 5-mC contents were significantly decreased in elder persons, suggesting the RNA modifications of m6A and 5-mC are correlated to aging.