Separating and probing tautomers of protonated nucleobases using differential mobility spectrometry

Abstract

The protonated nucleobases (C + H) + , (T + H) + , (U + H) + , (A + H) + , and (G + H) + are investigated in a combined experimental and computational study using differential mobility spectrometry (DMS), mass spectrometry, and electronic structure calculations. DMS is used to isolate individual tautomeric forms for each protonated nucleobase prior to characterization with HDX or CID. The population distributions of each protonated nucleobase formed by electrospray ionization (ESI) are dominated by a single tautomeric form, as is predicted by our calculations. However, all nucleobases present additional tautomers upon ESI, with these minor contributions to the ensemble populations attributed to additional higher energy metastable species. In addition to the tautomerderived species, additional ion signals in the DMS data are attributed to larger nucleobasecontaining clusters, which fragment post-DMS to yield bare ion and fragment ion signals that are consistent with those expected for the bare protonated nucleobases. Contributions from larger clustered species are instead distinguished by monitoring DMS ion signal as declustering potential voltages are ramped.

Selecting and identifying gas-phase protonation isomers of nicotineH+ using combined laser, ion mobility and mass spectrometry techniques

“…Not only are protomers isomers-and therefore unable to be mass-resolved by mass spectrometry-changes in ESI conditions are observed to affect relative protomer populations. [1][2][3][4][5][6][7][8][9][10][11][12][13][14] This can be an analytical problem when protomers dissociate to different product ions under activation, a phenomenon that can confound assignment by comparison to reference spectra. Differences in the photodissociation of protomers have been used to assign protomer populations, [10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25][26] although it is difficult to predict how the electronic spectroscopy and photodissociation will be affected by differences in protonation site.…”

“…FAIMS has proven to be a powerful tool for separating and analysing protomer populations at atmospheric pressure prior to detection by mass spectrometry. 1,4,6,8,46 Relative-energy calculations are typically coupled with FAIMS analysis to rationalise experimental results. However, it has been shown that theoretical results cannot reliably predict the relative protomer populations generated by ESI.…”

“…For example, protomer populations can be shifted by changing ESI conditions [1][2][3][4][5][6][7][8][9][10][11][12][13][14] and less stable isomers can be kinetically trapped by the ESI process. 4,6,9,[47][48] Furthermore, the relative energies of protomers can depend strongly on different levels of theory and basis sets, which can result in the prediction of different lowest energy protomers using different computational approaches. 14, 49 Here, we report the integration of FAIMS, ion-trap mass spectrometry and UVPD action spectroscopy to detect and assign the protonation isomers of protonated nicotine generated from ESI.…”

Probing the application range and selectivity of a differential mobility spectrometry–mass spectrometry platform for metabolomics

“…We had detected multiple peaks for some purine derivatives, for example theobromine (see “nucleobases” in 3.2 ). Peak splitting of protonated nucleobases has been observed before and was attributed to the presence of multiple tautomeric forms of the same molecule [17]. However, the methylxanthines (MeX) investigated in this study produced single peaks and could therefore be regarded as present in only one tautomeric form.…”

“…The resolution between of amino-substituted adenine and its hydroxyl analog hypoxanthine, was determined to be R DMS =1.58. Based on recent literature [17], it is likely that tautomerism contributes to the DMS behavior of nucleobases and their derivatives. Untangling the individual contributions from these factors was outside the scope of this study, even though we observed multiple peaks in some cases (see below).…”