Identification and Quantification of Cyclic Di-Guanosine Monophosphate and Its Linear Metabolites by Reversed-Phase LC-MS/MS

Cyclic dinucleotides such as bis-(3',5')-cyclic dimeric guanosine monophosphate (3',3'-c-di-GMP) represent an important class of second messengers in bacteria and are involved in numerous (patho)physiological settings. Here, we describe a sensitive and specific quantification method for 3',3'-c-di-GMP by HPLC-coupled tandem mass spectrometry (LC-MS/MS). Additionally, linear 3',3'-c-di-GMP metabolites, i.e., 5'-phosphoguanylyl-3',5'-guanosine (pGpG) and 5'-guanosine monophosphate (5'-GMP), as well as cyclic guanosine monophosphate (3',5'-cGMP) and 3',3' c-di-GMP analogues (2',3'-c-di-GMP and 2',2'-c-di-GMP) can be simultaneously determined by this method.