A Facile and Sensitive Method for Quantification of Cyclic Nucleotide Monophosphates in Mammalian Organs: Basal Levels of Eight cNMPs and Identification of 2',3'-cIMP

• Type of Material: Article
• Authors: Xin Jia, Emory University; Benjamin M. Fontaine, Emory University; Fred Strobel, Emory University; Emily Weinert, Emory University
• Language: English
• Date: 2014-12-12
• Publisher: MDPI
• Publication Version: Final Published Version
• Copyright Statement: © 2014 by the authors; licensee MDPI, Basel, Switzerland.
• License: Creative Commons Attribution 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.3390/biom4041070
• Title of Journal or Parent Work: Biomolecules
• ISSN: 2218-273X
• Volume: 4
• Issue: 4
• Start Page: 1070
• End Page: 1092
• Keywords: mammalian cNMP basal levels; liquid chromatography; 3',5'-cyclic nucleotides monophosphate; 2',3'-cyclic nucleotides monophosphate; tandem mass spectrometry; inosine 2',3'-cyclic monophosphate; LC-MS/MS
• Research Categories: Chemistry, General

Abstract

A sensitive, versatile and economical method to extract and quantify cyclic nucleotide monophosphates (cNMPs) using LC-MS/MS, including both 3',5'-cNMPs and 2',3'-cNMPs, in mammalian tissues and cellular systems has been developed. Problems, such as matrix effects from complex biological samples, are addressed and have been optimized. This protocol allows for comparison of multiple cNMPs in the same system and was used to examine the relationship between tissue levels of cNMPs in a panel of rat organs. In addition, the study reports the first identification and quantification of 2',3'-cIMP. The developed method will allow for quantification of cNMPs levels in cells and tissues with varying disease states, which will provide insight into the role(s) and interplay of cNMP signalling pathways.