Thermal desorption effects on fragment ion production from multi-photon ionized uridine and selected analogues

Abstract

Experiments on neutral gas-phase nucleosides are often complicated by thermal lability. Previous mass spectrometry studies of nucleosides have identified enhanced relative production of nucleobase ions (e.g. uracil⁺ from uridine) as a function of desorption temperature to be the critical indicator of thermal decomposition. On this basis, the present multi-photon ionization (MPI) experiments demonstrate that laser-based thermal desorption is effective for producing uridine, 5-methyluridine, and 2'-deoxyuridine targets without thermal decomposition. Our experiments also revealed one notable thermal dependence: the relative production of the sugar ion C₅H₉O₄ ⁺ from intact uridine increased substantially with the desorption laser power and this only occurred at MPI wavelengths below 250 nm (full range studied 222-265 nm). We argue that this effect can only be rationalized plausibly in terms of changing populations of different isomers, tautomers, or conformers in the target as a function of the thermal desorption conditions. Furthermore, the wavelength threshold behavior of this thermally-sensitive MPI channel indicates a critical dependence on neutral excited state dynamics between the absorption of the first and second photons. The experimental results are complemented by density functional theory (DFT) optimizations of the lowest-energy structure of uridine and two further conformers distinguished by different orientations of the hydroxymethyl group on the sugar part of the molecule. The energies of the transitions states between these three conformers are low compared with the energy required for decomposition.

Fig. 1. Chemical structures of the molecules studied in the present work. The diagrams are arranged such that the nucleosides with common sugar parts (ribose minus OH or deoxyribose minus OH) align horizontally, while those with common base parts (dehydrogenated uracil or dehydrogenated thymine) align vertically.

Fig. 2. Scheme of the MPI-TOF experiment in laser-based thermal desorption mode. The desorption source is shown in greater detail.

Fig. 3. Illustration of the reaction pathway concerning hydroxymethyl group rotation in ground-state uridine, calculated at M05/aug-cc-pVDZ level. Conformer 1 corresponds to the lowest-energy structure identified by Peña et al. on the basis of ab initio calculations and rotational spectroscopy experiments. Conformer 2 corresponds to the lowest-energy structure from DFT calculations by Delchev and by So and Alavi. Conformer 3 has not been reported previously. Transition states between each conformer correspond to a maximum in the potential energy along the reaction coordinate. Hydrogen bonds are indicated by dotted lines and atoms are colored as follows: blue for nitrogen, dark gray for carbon, red for oxygen, and light gray for hydrogen. Selected parameters of the optimized conformers and transition states are given in Table 1.

Fig. 4. Ratios of nucleobase⁺ (B⁺, e.g. m/z 112 for uracil⁺) over protonated nucleobase (BH⁺, e.g. m/z 113 for protonated uracil) production by 225 nm MPI of (a) uridine, C₉H₁₂N₂O₆, (b) 5-methyluridine, C₁₀H₁₄N₂O₆, (c) 2′-deoxyuridine, C₉H₁₂N₂O₅, and (d) thymidine, C₁₀H₁₄N₂O₅, as a function of the desorption laser power. The desorption laser power ranges correspond to foil temperature ranges of 133–167 °C for uridine, 72–151 °C for 5-methyluridine, 67–93 °C for 2′-deoxyuridine, and 119–151 °C for thymidine.

Fig. 5. (a) Desorption laser power dependence of S⁺/total ion production by 225 nm MPI of uridine (foil temperature range 119–165 °C). (b) Desorption laser power dependence of S⁺/total ion production and B⁺/BH⁺ production by 250 nm MPI of uridine (foil temperature range 136–218 °C). (c) MPI wavelength dependence of the S⁺ and C₃H₅O⁺ ion signals/total ion production from uridine (desorption laser power 0.41 W, foil temperature 144 °C). (d) 265 nm MPI mass spectrum of uridine (desorption laser power 0.41 W, foil temperature 144 °C).