The strong UV chromophores thymine (Thy) and uracil (Ura) have identical heteroaromatic rings that only differ by one methyl substituent. While their photophysics has been elucidated in detail, the effect on the excited states of base protonation and single water molecules is less explored. Here we report gas-phase absorption spectra of ThyH(+) and UraH(+) and monohydrated ions and demonstrate that the substituent is not only responsible for spectral shifts but also influences the tautomer distribution, being different for bare and monohydrated ions. Spectra interpretation is aided by calculations of geometrical structures and transition energies. The lowest free-energy tautomer (denoted 178, enol-enol form) accounts for 230-280 nm (ThyH(+)) and 225-270 nm (UraH(+)) bands. ThyH(+) hardly absorbs above 300 nm, whereas a discernible band is measured for UraH(+) (275-320 nm), ascribed to the second lowest free-energy tautomer (138, enol-keto form) comprising a few percent of the UraH(+) population at room temperature. Band widths are similar to those measured of cold ions in support of very short excited-state lifetimes. Attachment of a single water increases the abundance of 138 relative to 178, 138 now clearly present for ThyH(+). 138 resembles more the tautomer present in aqueous solution than 178 does, and 138 may indeed be a relevant transition structure. The band of ThyH(+)(178) is unchanged, that of UraH(+)(178) is nearly unchanged, and that of UraH(+)(138) blue-shifts by about 10 nm. In stark contrast to protonated adenine, more than one solvating water molecule is required to re-establish the absorption of ThyH(+) and UraH(+) in aqueous solution.

Microsolvation of chromophore ions commonly has large effects on their electronic structure and as a result on their optical absorption spectra. Here spectroscopy of protonated adenine (AdeH(+)) and its complex with one water molecule isolated in vacuo was done using a home-built mass spectrometer in combination with a tuneable pulsed laser system. Experiments also included the protonated adenosine 5'-monophosphate nucleotide (AMPH(+)). In the case of bare AdeH(+) ions, one-photon absorption leads to four dominant fragment ions corresponding to ammonium and ions formed after loss of either NH3, HCN, or NH2CN. The yields of these were measured as a function of the wavelength of the light from 210 nm to 300 nm, and they were combined to obtain the total photoinduced dissociation at each wavelength (i.e., action spectrum). A broad band between 230 nm and 290 nm and the tail of a band with maximum below 210 nm (high-energy band) are seen. In the case of AdeH(+)(H2O), the dominant dissociation channel after photoexcitation in the low-energy band was simply loss of H2O while photodissociation of protonated AMP revealed two dominant dissociation channels associated with the formation of either AdeH(+) or loss of H3PO4. The action spectra of AdeH(+), AdeH(+)(H2O), and AMPH(+) are almost identical in the 230-290 nm region, and they resemble the absorption spectrum of protonated adenine in aqueous solution recorded at low pH. Hence from our work it is firmly established that the lowest-energy transitions are independent of the surroundings.

Light absorption in the visible region by isolated Ru(bipy)3(+) (bipy = 2,2'-bipyridine) monocations, prepared in vacuo by reduction of dications in collisional electron transfer from cesium atoms, was recorded using photodissociation mass spectroscopy and found to be broad and similar to that of acetonitrile-solvated ions (maximum at 520 nm).

Vulnerability of biomolecules to ultraviolet radiation is intimately linked to deexcitation pathways: photostability requires fast internal conversion to the electronic ground state, but also intramolecular vibrational redistribution and cooling on a time scale faster than dissociation. Here we present a protocol to disentangle slow and non-hazardous statistical dissociation from prompt cleavage of peptide bonds by 210 nm light based on experiments on protonated peptides isolated in vacuo and tagged by 18-crown-6 ether (CE). The weakest link in the system is between the charged site and CE, which is remote from the initial site of excitation. Hence loss of CE serves as direct proof that energy has reached the charge-site end, leaving the backbone intact. Our work demonstrates that excitation of tertiary amide moieties (proline linkages) results in both prompt dissociation and statistical dissociation after energy randomisation over all vibrational degrees of freedom.

We report on the dissociation channels after UV photoexcitation of peptide cations tagged with 18-crown-6 ether (CE). The model peptides chosen for study were singly protonated (Ala)-Pro (n = 1, 2, 3) and Pro-Pro (Ala = alanine, Pro = proline) that all contain at least one tertiary amide group with high absorption cross section at 210 nm (5.90 eV). Statistical dissociation was identified from the loss of CE, a process occuring remotely from the initial site of excitation, and therefore requiring flow of energy to the ammonium group where the CE is bound. However, homolytic breakage of the peptide backbone at the site of excitation is competitive, resulting in so-called a radical cations. Density functional theory calculations of dissociation energies were done on the simplest system [Ala-Pro + H](CE) and found to be 1.87 eV for CE loss and 3.29 eV for the formation of a(CE) and x. These numbers were used to calculate statistical branching ratios for the dissociation processes based on detailed balance. After the absorption of two 210 nm photons (according to power-dependence measurements), the branching ratio between the two channels is calculated to be less than 10, far below the observed ratio of 0.65. Hence both statistical and non-statistical dissociation contribute to dissociation of these photoexcited peptides.