Infrared multiple photon dissociation spectroscopy of trapped ions

This tutorial review presents the technique of infrared multiple-photon dissociation (IRMPD) spectroscopy of mass-selected trapped ions. This requires coupling of a tunable infrared laser with mass spectrometry instrumentation. IRMPD spectroscopy has recently blossomed due to the emergence of widely tunable free electron lasers, as well as on-going developments of benchtop lasers. The merits of different trapping approaches in mass spectrometry are discussed in the light of photodissociation experiments. This tutorial discusses current capabilities, as well as limitations of the technique.     

Structure of electron-capture dissociation fragments from charge-tagged peptides probed by tunable infrared multiple photon dissociation

In this study, we propose the first spectroscopic structural characterization of c-type ions produced by ECD of a peptide. The structure of c-type ions formed by electron capture dissociation and the overall mechanism leading to their formation are still a question of debate. Depending on the mechanism, c-type ions have been proposed to have either an enol-imine structure (-C(OH)NH) or an amide one (-C(O)-NH2). Since these ions are isomeric, mass spectrometry only cannot discriminate between them, but infrared spectroscopy can bring experimental evidence and help determine which scheme is operative. Using the coupling between a tunable free electron laser and a FT-ICR mass spectrometer, we show that c-type ions have an amide structure, characterized by an IR signature of the C=O stretch at 1731 cm(-1). This result is particularly interesting from the perspective of the elucidation of the ECD mechanism.     

Some aspects of the infrared multiple photon dissociation of ammonia

The infrared multiple photon dissociation of NH₃ by a CO₂ laser is investigated through laser-induced fluorescence and chemiluminescence techniques. A linear dependence of the fractional dissociation yield on pulse energy is found, the dependence exemplifying two distinct thresholds. Significant collisional enhancement of the fractional yield is observed. The results are discussed in terms of the existence of bottlenecks in the laser up-pumping process.     

Pi-complex structure of gaseous benzene-NO cations assayed by IR multiple photon dissociation spectroscopy

[C(6)H(6)NO](+) ions, in two isomeric forms involved as key intermediates in the aromatic nitrosation reaction, have been produced in the gas phase and analyzed by IR multiple photon dissociation (IRMPD) spectroscopy in the 800-2200 cm(-)(1) fingerprint wavenumber range, exploiting the high fluence and wide tunability of a free electron laser (FEL) source. The IRMPD spectra were compared with the IR absorption spectra calculated for the optimized structures of potential isomers, thus allowing structural information on the absorbing species. [C(6)H(6)NO](+) ions were obtained by two routes, taking advantage of the FEL coupling to two different ion traps. In the first one, an FT-ICR mass spectrometer, a sequence of ion-molecule reactions was allowed to occur, ultimately leading to an NO(+) transfer process to benzene. The so-formed ions displayed IRMPD features characteristic of a [benzene,NO](+) pi-complex structure, including a prominent band at 1963 cm(-)(1), within the range for the N-O bond stretching vibration of NO (1876 cm(-)(1)) and NO(+) (2344 cm(-)(1)). A quite distinct species is formed by electrospray ionization (ESI) of a methanol solution of nitrosobenzene. The ions transferred and stored in a Paul ion trap showed the IRMPD features of substituent protonated nitrosobenzene, the most stable among conceivable [C(6)H(6)NO](+) isomers according to computations. It is noteworthy that IRMPD is successful in allowing a discrimination between isomeric [C(6)H(6)NO](+) species, whereas high-energy collision-induced dissociation fails in this task. The [benzene,NO](+) pi-complex is characterized by IRMPD spectroscopy as an exemplary noncovalent ionic adduct between two important biomolecular moieties.     

Structural characterization by infrared multiple photon dissociation spectroscopy of protonated gas-phase ions obtained by electrospray ionization of cysteine and dopamine

Structural characterization of protonated gas-phase ions of cysteine and dopamine by infrared multiple photon dissociation (IRMPD) spectroscopy using a free electron laser in combination with theory based on DFT calculations reveals the presence of two types of protonated dimer ions in the electrospray mass spectra of the metabolites. In addition to the proton-bound dimer of each species, the covalently bound dimer of cysteine (bound by a disulfide linkage) has been identified. The dimer ion of m/z 241 observed in the electrospray mass spectra of cysteine has been identified as protonated cystine by comparison of the experimental IRMPD spectrum to the IR absorption spectra predicted by theory and the IRMPD spectrum of a standard. Formation of the protonated covalently bound disulfide-linked dimer ions (i.e. protonated cystine) from electrospray of cysteine solution is consistent with the redox properties of cysteine. Both the IRMPD spectra and theory indicate that in protonated cystine the covalent disulfide bond is retained and the proton is involved in intramolecular hydrogen bonding between the amine groups of the two cysteine amino acid units. For cysteine, the protonated covalently bound dimer (m/z 241) dominated the mass spectrum relative to the proton-bound dimer (m/z 243), but this was not the case for dopamine, where the protonated monomer and the proton-bound dimer were both observed as major ions. An extended conformation of the ethylammonium side chain of gas-phase protonated dopamine monomer was verified from the correlation between the predicted IR absorption spectra and the experimental IRMPD spectrum. Dopamine has the same extended ethylamine side chain conformation in the proton-bound dopamine dimer identified in the mass spectra of electrosprayed dopamine. The structure of the proton-bound dimer of dopamine is confirmed by calculations and the presence of an IR band due to the shared proton. The presence of the shared proton in the protonated cystine ion can be inferred from the IRMPD spectrum.

Multiple infrared photon dissociation of vinyl chloride

Dehydrohalogenation of vinyl chloride, to form acetylene and HCL, is produced by irradiation with infrared laser pulses in the 9–11 μm region. The acetylene itself is broken down to yield diacetylene and elemental carbon and hydrogen when irradiated by focussed laser pulses. Initial excitation into vibrational modes of different symmetry appears to be about equally effective in converting absorbed energy to chemical reaction. The results are analyzed in terms of several models for multiple infrared photon dissociation; threshold energies of 20· 50 J/cm² are deduced.     

Dissociation of heme from gaseous myoglobin ions studied by infrared multiphoton dissociation spectroscopy and Fourier-transform ion cyclotron resonance mass spectrometry

Detachment of heme prosthetic groups from gaseous myoglobin ions has been studied by collision-induced dissociation and infrared multiphoton dissociation in combination with Fourier-transform ion cyclotron resonance mass spectrometry. Multiply charged holomyoglobin ions (hMbn+) were generated by electrospray ionization and transferred to an ion cyclotron resonance cell, where the ions of interest were isolated and fragmented by either collision with Ar atoms or irradiation with 3 mum photons, producing apomyoglobin ions (aMbn+). Both charged heme loss (with [Fe(III)-heme]+ and aMb(n-1)+ as the products) and neutral heme loss (with [Fe(II)-heme] and aMbn+ as the products) were detected concurrently for hMbn+ produced from a myoglobin solution pretreated with reducing reagents. By reference to Ea = 0.9 eV determined by blackbody infrared radiative dissociation for charged heme loss of ferric hMbn+, an activation energy of 1.1 eV was deduced for neutral heme loss of ferrous hMbn+ with n = 9 and 10.     

Vibrational spectra of small silicon monoxide cluster cations measured by infrared multiple photon dissociation spectroscopy

The first gas-phase infrared spectra of silicon monoxide cations (SiO)(n)(+), n = 3-5, using multiple photon dissociation in the 550-1250 cm(-1) frequency range, are reported. All clusters studied here fragment via loss of a neutral SiO unit. The experimental spectra are compared to simulated linear absorption spectra from calculated low energy isomers for each cluster. This analysis indicates that a "ring" isomer is the primary contributor to the (SiO)(3)(+) spectrum, that the (SiO)(4)(+) spectrum results from two close-lying bicyclic ring isomers, and that the (SiO)(5)(+) spectrum is from a bicyclic ring with a central, fourfold-coordinated Si atom. Experiment and theory indicate that the energies and energetic orderings of (SiO)(n)(+) isomers differ from those for neutral (SiO)(n) clusters.     

Infrared multiple photon dissociation spectroscopy of potassiated proline

The structure of proline in [proline + K]+ has been investigated in the gas phase using high level DFT and MP2 calculations and infrared photo dissociation spectroscopy with a free electron laser (FELIX). The respective FELIX spectrum of [proline + K]+ matches convincingly the calculated spectra of two structurally closely related and nearly iso-energetic zwitterionic salt bridge (SB) structures. An additional unresolved band at approximately 1725 cm(-1) matching with the characteristic CO stretching mode of charge solvation (CS) structures points toward the presence of a minor population of these conformers of proline in [proline + K]+. However, theory predicts a significant energy gap of 18.9 kJ mol(-1) (B3LYP/6-311++G(2d,2p)) or 15.6 kJ mol(-1) (MP2) between the lowest CS conformer of proline and the clearly favored SB structure.     

Infrared multiphoton and collision-induced dissociation studies of some gaseous alkylamine ions

Collision-induced dissociation and infrared multiphoton dissociation of ions formed in di- and tri-ethylamine, di- and tri-n-propylamine, and di-isopropylamine were investigated by Fourier-transform ion-cyclotron resonance mass spectrometry. Molecular ions of all amines except di-n-propylamine produced similar fragment ions when subjected to either dissociation technique. The initial fragmentation involved C_αC_β bond cleavage, loss of an alkyl radical, and formation of an immonium ions. Subsequent fragmentations of the immonium ions produced by both dissociation mechanisms involved McLafferty-type rearrangements and loss of alkenes. The molecular ion of di-n-propylamine fragmented by a different mechanism when subjected to infrared irradiation. Protonated molecules of di- and tri-n-propylamine yielded C₃H₆ and an ammonium ion upon infrared multiphoton dissociation, while protonated molecules of the other amines did not dissociate when this technique was applied. In contrast, collision-induced dissociation produced fragmentation for all protonated molecules. Explanation of the different fragmentations observed for the two dissociation techniques is given in terms of a mechanism involving a tight transition state for protonated di- and tri-n-propylamine dissociation.     

Reactivity and infrared spectroscopy of gaseous hydrated trivalent metal ions

Hydrated trivalent rare earth metal ions containing yttrium and all naturally abundant lanthanide metals are formed using electrospray ionization, and the structures and reactivities of these ions containing 17-21 water molecules are probed using blackbody infrared radiative dissociation (BIRD) and infrared action spectroscopy. With the low-energy activation conditions of BIRD, there is an abrupt transition in the dissociation pathway from the exclusive loss of a single neutral water molecule to the exclusive loss of a small protonated water cluster via a charge-separation process. This transition occurs over a narrow range of cluster sizes that differs by only a few water molecules for each metal ion. The effective turnover size at which these two dissociation rates become equal depends on metal ion identity and is poorly correlated with the third ionization energies of the isolated metals but is well correlated with the hydrolysis constants of the trivalent metal ions in bulk aqueous solution. Infrared action spectra of these ions at cluster sizes near the turnover size are largely independent of the specific identity of the trivalent metal ion, suggesting that any differences in the structures of the ions present in our experiment are subtle.     

Infrared spectrum of potassium‐cationized triethylphosphate generated using tandem mass spectrometry and infrared multiple photon dissociation

Tandem mass spectrometry and wavelength‐selective infrared photodissociation were used to generate an infrared spectrum of gas‐phase triethylphosphate cationized by attachment of K⁺. Prominent absorptions were observed in the region of 900 to 1300 cm^?1 that are characteristic of phosphate P?O and P? O? R stretches. The relative positions and intensities of the IR absorptions were reproduced well by density functional theory (DFT) calculations performed using the B3LYP functional and the 6‐31+G(d), 6‐311+G(d,p) and 6‐311++G(3df,2pd) basis sets. Because of good correspondence between experiment and theory for the cation, DFT was then used to generate a theoretical spectrum for neutral triethylphosphate, which in turn accurately reproduces the IR spectrum of the neat liquid when solvent effects are included in the calculations. Copyright © 2009 John Wiley & Sons, Ltd.     

A new optothermal detector of infrared multiple photon absorption

A new optothermal detector has been developed to detect the multiphoton absorption spectra of NH₃, C₂H₄, and SF₆ in a Strong CO₂ laser field. The results coincide with those for optoacoustic detection.     

Electron capture dissociation of gaseous multiply charged ions by Fourier-transform ion cyclotron resonance

Fourier-transform ion cyclotron resonance instrumentation is uniquely applicable to an unusual new ion chemistry, electron capture dissociation (ECD). This causes nonergodic dissociation of far larger molecules (42 kDa) than previously observed (<1 kDa), with the resulting unimolecular ion chemistry also unique because it involves radical site reactions for similarly larger ions. ECD is highly complementary to the well known energetic methods for multiply charged ion dissociation, providing much more extensive protein sequence information, including the direct identification of N- versus C-terminal fragment ions. Because ECD only excites the molecule near the cleavage site, accompanying rearrangements are minimized. Counterintuitively, cleavage of backbone covalent bonds of protein ions is favored over that of noncovalent bonds; larger (>10 kDa) ions give far more extensive ECD if they are first thermally activated. This high specificity for covalent bond cleavage also makes ECD promising for studying the secondary and tertiary structure of gaseous protein ions caused by noncovalent bonding.     

Detailed unfolding and folding of gaseous ubiquitin ions characterized by electron capture dissociation

The unfolding enthalpy of the native state of ubiquitin in solution is 5 to 8 times that of its gaseous ions, as determined by electron capture dissociation (ECD) mass spectrometry. Although two-state folding occurs in solution, the three-state gaseous process proposed for this by Clemmer and co-workers based on ion mobility data is supported in general by ECD mass spectra, including relative product yields, distinct Delta H(unfolding) values between states, site-specific melting temperatures, and folding kinetics indicating a cooperative process. ECD also confirms that the 13+ ions represent separate conformers, possibly with side-chain solvated alpha-helical structures. However, the ECD data on the noncovalent bonding in the 5+ to 13+ ions, determined overall in 69 of the 75 interresidue sites, shows that thermal unfolding proceeds via a diversity of intermediates whose conformational characteristics also depend on charge site locations. As occurs with increased acidity in solution, adding 6 protons to the 5+ ions completely destroys their tertiary noncovalent bonding. However, solvation of the newly protonated sites to the backbone instead increases the stability of the secondary structure (possibly an alpha-helix) of these gaseous ions, while in solution these new sites aid denaturation by solvation in the aqueous medium. Extensive ion equilibration can lead to even more compact and diverse conformers. The three-state unfolding of gaseous ubiquitin appears to involve ensembles of individual chain conformations in a "folding funnel" of parallel reaction paths. This also provides a further caution for characterizing solution conformers from their gas-phase behavior.     

Infrared multiple photon dissociation spectroscopy of sodium and potassium chlorate anions

The structures of gas‐phase, metal chlorate anions with the formula [M(ClO₃)₂]^?, M = Na and K, were determined using tandem mass spectrometry and infrared multiple photon dissociation (IRMPD) spectroscopy. Structural assignments for both anions are based on comparisons of the experimental vibrational spectra for the two species with those predicted by density functional theory (DFT) and involve conformations that feature either bidentate or tridentate coordination of the cation by chlorate. Our results strongly suggest that a structure in which both chlorate anions are bidentate ligands is preferred for [Na(ClO₃)₂]^?. However, for [K(ClO₃)₂]^? the best agreement between experimental and theoretical spectra is obtained from a composite of predicted spectra for which the chlorate anions are either both bidentate or both tridentate ligands. In general, we find that the overall accuracy of DFT calculations for prediction of IR spectra is dependent on both functional and basis set, with best agreement achieved using frequencies generated at the B3LYP/6‐311+g(3df) level of theory. Copyright © 2009 John Wiley & Sons, Ltd.     

Infrared multiple photon dissociation action spectroscopy and computational studies of mass-selected gas-phase fluorescein and 2',7'-dichlorofluorescein ions

Fluorescein (FL) and its derivative 2',7'-dichlorofluoroescein (DCF) are well-known fluorescent dyes used in many biological and biochemical applications. Although extensive studies have been carried out to investigate their chemical and photophysical properties in different solvent media, little is known about their intrinsic behaviors in the gas phase. Here, infrared multiple photon dissociation (IRMPD) action spectra are reported for the three charged prototropic forms of FL and DCF and compared with computed IR spectra from electronic structure calculations. In each case, the measured spectra show good agreement with the calculated spectra of the lowest energy computed conformer. Moreover, the major bands of the monoanion IRMPD spectra show striking similarities to those of the dianions and are quite different from those of the cations. These experimental results clearly indicate that the gaseous monoanions are predominantly deprotonated on the xanthene chromophore, rather than the benzoate deprotonation site favored in solution. Investigations such as this, which provide a better understanding of intrinsic properties of ionic dyes, forms a baseline from which to elucidate solvent effects and will aid the rational design of dyes possessing desirable fluorescence properties.