UV Photofragmentation Research Articles

High-Throughput UV Photoionization and Fragmentation of Neutral Biomolecules as a Structural Fingerprint.

We present UV photofragmentation studies of the structural isomers paracetamol, 3-Pyridinepropionic acid (3-PPIA) and (R)-(-)-2-Phenylglycine. In particular, we utilized a new laser-based thermal desorption source in combination with femtosecond multiphoton ionization at 343 nm and 257 nm. The continuous nature of our molecule source, combined with the 50 kHz repetition rate of the laser, allowed us to perform these experiments at high throughput. In particular, we present detailed laser intensity dependence studies at both wavelengths, producing 2D mass spectra with highly differential information about the underlying fragmentation processes. We show that UV photofragmentation produces highly isomer-specific mass spectra, and assign all major fragmentation pathways observed. The intensity-dependence measurements, furthermore, allowed us to evaluate the appearance intensities for each fragmentation channel, which helped to distinguish competing from consecutive fragmentation pathways.

Accelerating photofragmentation UV Spectroscopy–Mass spectrometry fingerprinting for quantification of isomeric peptides

Identification of isomeric biomolecules remains a challenging analytical problem. A recently developed spectroscopic method that combines UV photofragmentation and mass spectrometry for fingerprinting of cold ions (2D UV-MS), has already demonstrated its high performance in the library-based identification and quantification of different types of biomolecular isomers. The practical use of the method has been hindered by a slow rate of data acquisition, which makes the fingerprinting incompatible with high-throughput analysis and online liquid chromatography (LC) separation. Herein we demonstrate how the use of a few pre-selected wavelengths can accelerate the method by two orders of magnitude without a significant loss of accuracy. As a proof of principle, 2D UV-MS fingerprinting was coupled to online LC separation and tested for quantification of isomeric peptides containing either Asp or isoAsp residues. The relative concentrations of the peptides mixed in solution have been determined, on average, with better than 4% and 6% accuracy for resolving and non-resolving gradients of LC separation, respectively.

UV Photofragmentation of Cold Cytosine–M+ Complexes (M+: Na+, K+, Ag+)

The UV photofragmentation spectra of cold cytosine-M+ complexes (M+: Na+, K+, Ag+) were recorded and analyzed through comparison with geometry optimizations and frequency calculations of the ground and excited states at the SCS-CC2/Def2-SVPD level of theory. While in all complexes, the ground state minimum geometry is planar (Cs symmetry), the ππ* state minimum geometry has the NH2 group slightly twisted and an out-of-plane metal cation. This was confirmed by comparing the simulated ππ* Franck-Condon spectra with the vibrationally resolved photofragmentation spectra of CytNa+ and CytK+. Vertical excitation transitions were also calculated to evaluate the energies of the CT states involving the transfer of an electron from the Cyt moiety to M+. For both CytK+ and CytNa+ complexes, the first CT state corresponds to an electron transfer from the cytosine aromatic π ring to the antibonding σ* orbital centered on the alkali cation. This πσ* state is predicted to lie much higher in energy (>6 eV) than the band origin of the π-π* electronic transition (around 4.3 eV) unlike what is observed for the CytAg+ complex for which the first excited state has a nOσ* electronic configuration. This is the reason for the absence of the Cyt+ + M charge transfer fragmentation channel for CytK+ and CytNa+ complexes.

UV photofragmentation dynamics of acetaldehyde cations prepared by single-photon VUV ionization.

Acetaldehyde cations (CH3CHO+) were prepared using single-photon vacuum ultraviolet ionization of CH3CHO in a molecular beam and the fragmentation dynamics explored over the photolysis wavelength range 390-210 nm using velocity-map ion imaging and photofragment yield (PHOFY) spectroscopy. Four fragmentation channels are characterized: CH3CHO+→ C2H3O+ + H (I), CH3CHO+→ HCO+ + CH3 (II), CH3CHO+→ CH3+ + HCO (III), CH3CHO+→ CH4+ + CO (IV). Channels (I), (II), and (IV) are observed across the full photolysis wavelength range while channel (III) is observed only at λ < 317 nm. Maximum fragment ion yields are obtained at ∼250 nm. Ion images were recorded over the range 316-228 nm, which corresponds to initial excitation to the B[combining tilde]2A' and C[combining tilde]2A' states of CH3CHO+. The speed and angular distributions are distinctly different for each detected ion and show evidence of both statistical and dynamical fragmentation pathways. At longer wavelengths, fragmentation via channel (I) leads to modest translational energies (ET), consistent with dissociation over a small barrier and production of highly internally excited CH3CO+. Additional components with EINT greater than the CH3CO+ secondary dissociation threshold appear at shorter wavelengths and are assigned to fragmentation products of vinyl alcohol cation or oxirane cation formed by isomerization of energized CH3CHO+. The ET distribution observed for channel (III) products peaks at zero but is notably colder than that predicted by phase space theory, particularly at longer photolysis wavelengths. The colder-than-statistical ET distributions are attributed to contributions from secondary fragmentation of energized CH3CO+ formed via channel (I), which are attenuated by CH3CHO+ isomerization at shorter wavelengths. Fragmentation via channels (II) and (IV) results in qualitatively similar outcomes, with evidence of isotropic statistical components at low-ET and anisotropic components due to excited state dynamics at higher ET.

Radical Anions of Oxidized vs. Reduced Oxytocin: Influence of Disulfide Bridges on CID and Vacuum UV Photo-Fragmentation.

The nonapeptide oxytocin (OT) is used as a model sulfur-containing peptide to study the damage induced by vacuum UV (VUV) radiations. In particular, the effect of the presence (or absence in reduced OT) of oxytocin's internal disulfide bridge is evaluated in terms of photo-fragmentation yield and nature of the photo-fragments. Intact, as well as reduced, OT is studied as dianions and radical anions. Radical anions are prepared and photo-fragmented in two-color experiments (UV + VUV) in a linear ion trap. VUV photo-fragmentation patterns are analyzed and compared, and radical-induced mechanisms are proposed. The effect of VUV is principally to ionize but secondary fragmentation is also observed. This secondary fragmentation seems to be considerably enabled by the initial position of the radical on the molecule. In particular, the possibility to form a radical on free cysteines seems to increase the susceptibility to VUV fragmentation. Interestingly, disulfide bridges, which are fundamental for protein structure, could also be responsible for an increased resistance to ionizing radiations. Graphical Abstract.

Photodissociation Spectroscopy of Cold Protonated Synephrine: Surprising Differences between IR-UV Hole-Burning and IR Photodissociation Spectroscopy of the O-H and N-H Modes.

We report the UV and IR photofragmentation spectroscopies of protonated synephrine in a cryogenically cooled Paul trap. Single (UV or IR) and double (UV-UV and IR-UV) resonance spectroscopies have been performed and compared to quantum chemistry calculations, allowing the assignment of the lowest-energy conformer with two rotamers depending on the orientation of the phenol hydroxyl (OH) group. The IR-UV hole burning spectrum exhibits the four expected vibrational modes in the 3 μm region, i.e., the phenol OH, Cβ-OH, and two NH2+ stretches. The striking difference is that, among these modes, only the free phenol OH mode is active through IRPD. The protonated amino group acts as a proton donor in the internal hydrogen bond and displays large frequency shifts upon isomerization expected during the multiphoton absorption process, leading to the so-called IRMPD transparency. More interestingly, while the Cβ-OH is a proton acceptor group with moderate frequency shift for the different conformations, this mode is still inactive through IRPD.

Vibronic spectra of protonated hydroxypyridines: contributions of prefulvenic and planar structures.

Various hydroxypyridine derivatives are endogenous or synthetic photosensitizers which could contribute to solar radiation damage. The study of their excited states could lead to a better understanding of their action mechanisms. We present here the ultraviolet (UV) spectra of the protonated 2-, 3- and 4-hydroxypyridine. These spectra were obtained with an experimental device coupling an electrospray ion source with a cold quadrupole ion trap and a time of flight mass spectrometer. They display well resolved vibrational structures, with a clear influence of the position of the OH group. These results are interpreted with excited states calculations at the coupled cluster CC2 level.

Identification of Isomeric Ephedrines by Cold Ion UV Spectroscopy: Toward Practical Implementation.

Ephedrine and pseudoephedrine are stimulant drugs whose use is prohibited in athletic competition by the World Anti-Doping Agency (WADA) at very different threshold doping violation concentrations. We use a recently developed universal approach that integrates UV photofragmentation spectroscopy of cold ions with Orbitrap mass spectrometry (MS) for highly selective and highly sensitive identification of these diastereomers. Both species can be selectively detected at a solution concentration of a few tens of ng/mL, which is almost 3 orders of magnitude lower than the threshold concentration required by WADA. Relative concentrations of the isomers in solutions have been determined with the standard deviation of 3.1%, when the ions were cooled in an ion trap maintained at T = 6 K. Considering practical implementation of the method, we evaluated its performance for a simplified instrumentation. At an affordable elevated temperature of ∼70 K and with a low-maintenance midbandwidth optical parametric oscillator, a few second measurement should yield nearly the same selectivity and only ten times lower sensitivity than with the current research grade instrument.

Twisted Intramolecular Charge Transfer in Protonated Amino Pyridine.

The excited state properties of protonated ortho (2-), meta (3-), and para (4-) aminopyridine molecules have been investigated through UV photofragmentation spectroscopy and excited state coupled-cluster CC2 calculations. Cryogenic ion spectroscopy allows recording well-resolved vibronic spectroscopy that can be reproduced through Franck-Condon simulations of the ππ* local minimum of the excited state potential energy surface. The excited state lifetimes have also been measured through a pump-probe excitation scheme and compared to the estimated radiative lifetimes. Although protonated aminopyridines are rather simple aromatic molecules, their deactivation mechanisms are indeed quite complex with unexpected results. In protonated 3- and 4-aminopyridine, the fragmentation yield is negligible around the band origin, which implies the absence of internal conversion to the ground state. Besides, a twisted intramolecular charge transfer reaction is evidenced in protonated 4-aminopyridine around the band origin, while excited state proton transfer from the pyridinic nitrogen to the adjacent carbon atom opens with roughly 500 cm(-1) of excess energy.

UV photofragmentation and IR spectroscopy of cold, G-type β-O-4 and β-β dilignol-alkali metal complexes: structure and linkage-dependent photofragmentation.

Ultraviolet photofragmentation spectroscopy and infrared spectroscopy were performed on two prototypical guaiacyl (G)-type dilignols containing β-O-4 and β-β linkages, complexed with either lithium or sodium cations. The complexes were generated by nanoelectrospray ionization, introduced into a multistage mass spectrometer, and subsequently cooled in a 22-pole cold ion trap to T ≈ 10 K. A combination of UV photofragment spectroscopy and IR-UV double resonance spectroscopy was used to characterize the preferred mode of binding of the alkali metal cations and the structural changes so induced. Based on a combination of spectral evidence provided by the UV and IR spectra, the Li(+) and Na(+) cations are deduced to preferably bind to both dilignols via their linkages, which constitute unique, oxygen-rich binding pockets for the cations. The UV spectra reflect this binding motif in their extensive Franck-Condon activity involving low-frequency puckering motions of the linkages in response to electronic excitation. In the pinoresinol•Li(+)/Na(+) complexes involving the β-β linkage, the spectra also showed an inherent spectral broadening. The photofragment mass spectra are unique for each dilignol•Li(+)/Na(+) complex, many of which are also complementary to those produced by collision-induced dissociation (CID), indicating the presence of unique excited state processes that direct the fragmentation. These results suggest the potential for site-selective fragmentation and for uncovering fragmentation pathways only accessed by resonant UV excitation of cold lignin ions.

Mapping the UV Photophysics of Platinum Metal Complexes Bound to Nucleobases: Laser Spectroscopy of Isolated Uracil·Pt(CN)4(2-) and Uracil·Pt(CN)6(2-) Complexes.

We report the first UV laser spectroscopic study of isolated gas-phase complexes of platinum metal complex anions bound to a nucleobase as model systems for exploring at the molecular level the key photophysical processes involved in photodynamic therapy. Spectra of the Pt(IV)(CN)6(2-)·Ur and Pt(II)(CN)4(2-)·Ur complexes were acquired across the 220-320 nm range using mass-selective photodepletion and photofragment action spectroscopy. The spectra of both complexes reveal prominent UV absorption bands (λmax = 4.90 and 4.70 eV) that we assign primarily to excitation of the Ur π-π* localized chromophore. Distinctive UV photofragmentation products are observed for the complexes, with Pt(IV)(CN)6(2-)·Ur photoexcitation resulting in complex fission, while Pt(II)(CN)4(2-)·Ur photoexcitation initiates a nucleobase proton-transfer reaction across 4.4-5.2 eV and electron detachment above 5.2 eV. The observed photofragments are consistent with ultrafast decay of a Ur localized excited state back to the electronic ground state followed by intramolecular vibrational relaxation and ergodic complex fragmentation.

Effect of Ag(+) on the Excited-State Properties of a Gas-Phase (Cytosine)2Ag(+) Complex: Electronic Transition and Estimated Lifetime.

Recently, DNA molecules have received great attention because of their potential applications in material science. One interesting example is the production of highly fluorescent and tunable DNA-Agn clusters with cytosine (C)-rich DNA strands. Here, we report the UV photofragmentation spectra of gas-phase cytosine···Ag(+)···cytosine (C2Ag(+)) and cytosine···H(+)···cytosine (C2H(+)) complexes together with theoretical calculations. In both cases, the excitation energy does not differ significantly from that of isolated cytosine or protonated cytosine, indicating that the excitation takes place on the DNA base. However, the excited-state lifetime of the C2H(+) (τ = 85 fs), estimated from the bandwidth of the spectrum, is at least 2 orders of magnitude shorter than that of the C2Ag(+) (τ > 5000 fs). The increased excited-state lifetime upon silver complexation is quite unexpected, and it clearly opens the question about what factors are controlling the nonradiative decay in pyrimidine DNA bases. This is an important result for the expanding field of metal-mediated base pairing and may also be important to the photophysical properties of DNA-templated fluorescent silver clusters.

UV Photofragmentation Dynamics of Protonated Cystine: Disulfide Bond Rupture.

Disulfide bonds (S-S) play a central role in stabilizing the native structure of proteins against denaturation. Experimentally, identification of these linkages in peptide and protein structure characterization remains challenging. UV photodissociation (UVPD) can be a valuable tool in identifying disulfide linkages. Here, the S-S bond acts as a UV chromophore and absorption of one UV photon corresponds to a σ-σ* transition. We have investigated the photodissociation dynamics of protonated cystine, which is a dimer of two cysteines linked by a disulfide bridge, at 263 nm (4.7 eV) using a multicoincidence technique in which fragments coming from the same fragmentation event are detected. Two types of bond cleavages are observed corresponding to the disulfide (S-S) and adjacent C-S bond ruptures. We show that the S-S cleavage leads to three different fragment ions via three different fragmentation mechanisms. The UVPD results are compared to collision-induced dissociation (CID) and electron-induced dissociation (EID) studies.

A framework for resolving the origin, nature and evolution of the diffuse interstellar band carriers?

The carriers of the diffuse interstellar bands (DIBs) still remain an unknown commodity. Both dust and molecules have been suggested as carriers but none proposed have yet been able to explain the nature and the diversity of the DIBs. Hence, it is perhaps time to review the problem in terms of the intermediate-sized nano-particles. It is here proposed that the DIB carriers are the nm-sized and sub-nm-sized products of the UV photo-fragmentation of hydrogenated amorphous carbon grains, a-C(:H), and their heteroatom-doped variants, a-C:H:X (where X may be O, N, Mg, Si, Fe, S, Ni, P, …). An interstellar hydrogenated amorphous carbon dust evolutionary framework is described within which a solution to the age-old DIB problem could perhaps be found.

IR and UV Spectroscopy of Vapor-Phase Jet-Cooled Ionic Liquid [emim]+[Tf2N]−: Ion Pair Structure and Photodissociation Dynamics

Small gas-phase clusters (ion pairs) of the ionic liquid [emim](+)[Tf2N](-) have been generated in a supersonic expansion. Clusters are investigated via UV photofragmentation and time-of-flight mass spectrometry. Spectra between 42,000 and 45,000 cm(-1) reveal dynamical branching between direct dissociation of the ion pair to the cation and anion and to radical species. The IR spectrum between 2800 and 3200 cm(-1) was measured by action spectroscopy. Multiple conformations of the ion pair are found to be present in the molecular beam, leading to broad spectral features, further complicated by hydrogen bonding and Fermi resonances. The measured and theoretical spectra compare well, and the jet-cooled ion pair structures present in the molecular beam are strongly hydrogen bonded "stacked" conformers.

Atmospheric Pressure Acetylene Detection by UV Photo-Fragmentation and Induced C2 Emission

Detection of C(2)H(2) via UV photo-fragmentation, followed by monitoring the C(2) d(3)Π(g)-a(3)Π(u) fluorescence, is explored at atmospheric pressure and at temperatures of 295 K, 600 K, and 800 K, for excitation wavelengths 210 to 240 nm using a broadband laser source (~3 cm(-1) fwhm). At the lower temperature, C(2) emissions correlate closely with C(2)H(2) Ã ← X absorption bands, and the excitation spectra suggest a higher-transition probability for the v(″)(4) = 2 and 3 states than for the v(″)(4) = 0 and 1 states. As temperature increases, the excitation spectra exhibit a higher nonresonant background.

Gas phase Photo-Formation and Vacuum UV Photofragmentation Spectroscopy of Tryptophan and Tyrosine Radical-Containing Peptides

Tryptophan (Trp(•)) and tyrosyl (Tyr(•)) radical containing peptides were produced by UV laser-induced electron detachment from a suitable precursor. Vacuum ultraviolet (VUV) action spectra of these radical peptides were recorded with synchrotron radiation in the 4.5-16 eV range, from which fragmentation pathways and yields are measured as a function of the VUV photon energy. An enhancement in photofragmentation yields of radical species by 1 order of magnitude with respect to nonradical peptides is demonstrated here for the first time. Photofragmentation spectra are compared with absorption spectra for model chromophores calculated in the frame of the time-dependent density functional theory (TDDFT). A qualitative agreement in the position of bands in the 6-8 eV region is observed between experimental photofragmentation and calculated absorption spectra. Photofragmentation spectra of peptide radicals can be useful to better assess the complex deactivation pathways that occur following the absorption of a VUV photon in biomolecular radical anions.

Ultraviolet Photofragmentation Spectroscopy of Alkaline Earth Dication Complexes with Pyridine and 4-Picoline (4-Methyl pyridine)

A detailed experimental and theoretical study has been undertaken of the UV photofragmentation spectroscopy of the alkaline earth metal dications Mg(2+), Ca(2+), and Sr(2+) complexed with pyridine and 4-methyl pyridine (4-picoline). The ion complexes have been prepared using the pick-up technique and held in an ion trap where their internal temperature has been reduced to <150 K. Exposure of the trapped ions to tunable UV laser radiation leads to the appearance of photofragments with intensities that show significant variation as a function of wavelength. For all three metal dications, the resultant spectra show evidence of resolved features. Time-dependent density functional theory (TDDFT) has been used to identify possible electronic transitions that might be present in the [M(pyridine)(4)](2+) complexes (M = Mg, Ca, and Sr) within the wavelength range studied. These calculations show that the spectra are dominated by strong π* ← π and weaker π* ← n transitions localized on the pyridine ligands. The calculations correctly identify those regions of the experimental spectra where UV transitions begin to occur in the complexes and also the wavelengths at which absorption maxima are reached; however, more subtle features of the spectra are difficult to assign with confidence.

UV-photodissociation of non-cyclic and cyclic mononucleotides

Irradiation of nucleotides in the gas phase with ultraviolet light can lead to their fragmentation. We present a comparison of UV photofragmentation data on ribo-, deoxyribo- and cyclic nucleotides with guanine and adenine as nucleobases. The envelope of the UV photofragment spectra does not depend significantly on the detailed structure of the sugar–phosphate backbone. The fragment channels observed in the photofragmentation of ribonucleotides are very similar to those of deoxyribonucleotides with small differences in the relative abundances of the product ions. In fragmentation of cyclic nucleotides, the deprotonated base anions are the most abundant fragments, in contrast to the non-cyclic nucleotides, where this channel is significantly weaker. We discuss the abundances in the context of possible fragmentation mechanisms and structural differences of the parent molecules.

State-resolved UV photofragmentation spectrum of the metal dication complex [Zn(pyridine)4]2+

A combined theoretical and experimental study of electronic transitions in the complex [Zn(pyridine)(4)](2+) provides the first example of a state-resolved electronic spectrum to be recorded for a dication complex in the gas phase.