HPLC electrospray mass spectrometric characterization of trimeric building blocks for oligonucleotide synthesis

Trimeric nucleotide building blocks are valuable in synthesis of randomized oligonucleotides. In this study, we have developed HPLC-MS and HPLC-MS/MS methods for quality control of protected trinucleotides. C18 reversed-phase HPLC was used for purity evaluation, and base sequences were verified using negative ion electrospray ionization mass spectrometry (ESI-MS) and collision induced dissociation (CID) MS/MS. The principal dissociation pathway was formation of w-ions, which represent the 3'-5' direction. The other major fragments were d-ions, which are formed by cleavage of 5' C-O bonds of the sugars. The developed method was suitable for verification of purity and structure of the 29 trinucleotides studied.