Electrospray mass spectrometry (ESIMS) is now used extensively to establish the structural integrity of synthetic oligonucleotides which play an important role in molecular biology. There have been fewer studies, however, of electrospray tandem mass spectrometry (ESI-MS/MS) for the characterisation of oligonucleotides.
ESI-MS/MS spectra of a range of dinucleotides and oligonucleotides containing up to 8 bases have been examined on a triple quadrupole mass spectrometer. These spectra showed abundant peaks due to free bases and phosphate, however, there were generally limited backbone cleavages. A number of strategies for generating more extensive fragmentation were investigated. The use of higher collision energies and/or skimmer potential had no significant effect on the types of fragments observed, though there were some differences in relative intensities of the fragment ions. For dinucleotides MS/MS of different types of parent ions were compared, i.e., [M-H-]-, [M+H+]+, [M+Li+]+, [M+Na+]+. The most extensive fragmentation was observed from MS/MS of [M+Li+]+ similar to that shown by Ramsey et al. [1]. Many of the additional fragments involved cleavage of the sugar residue, which is consistent with previous reports for sugars [2]. In MS/MS spectra of multiply charged peaks (i.e., 2- and 3-), of larger oligonucleotides (3mers to 8mers) obtained on the triple quadrupole, ions due to individual bases and phosphate groups again dominate the spectra. This contrasts to previous studies by McLuckey et al. who observed some backbone fragmentation in MS/MS spectra of multiply charged ions, using an ion trap [3].
Finally, preliminary high resolution MS/MS spectra of oligonucleotides obtained on a magnetic-sector orthogonal acceleration time-of-flight tandem mass spectrometer will be presented.