Structures of aliphatic amino acid proton-bound dimers by infrared multiple photon dissociation spectroscopy in the 700-2,000 cm(-1) region

Structural aspects of proton-bound dimers composed of amino acids with aliphatic side chains are investigated using infrared multiple photon dissociation (IRMPD) spectroscopy and electronic structure calculations. Features in the IRMPD spectra in the 700-2,000 cm-1 range are due primarily to C=O stretching, NH2 bending, and COH bending. It was possible to distinguish between isomeric structures by comparing the experimental IRMPD spectra and those predicted using B3LYP/6-31+G(d,p). It was possible, based on the calculations and IRMPD spectra, to assign the experimental spectrum of the glycine proton-bound dimer to a structure which was slightly different from that assigned by previous spectroscopic investigations and in agreement with recent thermochemical studies. Since all proton-bound dimers studied here, composed of the different amino acids, have very similar spectra, it is expected that they also have very similar lowest-energy structures including the mixed alanine/glycine proton-bound dimer. In fact, the spectra are so similar that it would be very challenging to distinguish, for example, the glycine proton-bound dimer from the alanine or valine proton-bound dimers in the 700-2,000 cm-1 range. According to the calculated IR spectra it is shown that in the approximately 2,000-3,200 cm-1 range differentiating between different structures as well as different proton-bound dimers may be possible. This is due mainly to differences in the asymmetric stretch of the binding proton which is predicted to occur in this region.     

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.

Infrared spectra of homogeneous and heterogeneous proton-bound dimers in the gas phase

Infrared multiphoton dissociation spectra of three homogeneous and two heterogeneous proton-bound dimers were recorded in the gas phase. Comparison of the experimental infrared spectra recorded in the fingerprint region of the proton-bound dimers with spectra predicted by electronic structure calculations shows that all modes which are observed contain motion of the proton oscillating between the two monomers. The O-H-O asymmetric stretch for the homogeneous dimers is shown to occur at around 800 cm-1. As expected, the O-H-O asymmetric stretching modes for the heterogeneous proton-bound dimers are observed to shift to significantly higher energy with respect to those for the homogeneous proton-bound dimers due to the asymmetry of the O-H-O moeity. This shift is shown to be predictable from the difference in proton affinities between the two monomers. Density functional predictions of the infrared spectra based on the harmonic oscillator model are demonstrated to predict the observed spectra of the homogeneous proton-bound dimers with reasonable accuracy. Calculations of the structure and infrared spectrum of protonated diglyme at the B3LYP/6-31+G** level and basis also agree well with an infrared spectrum recorded previously. For both heterogeneous proton-bound dimers, however, the predicted spectra are blue-shifted with respect to experiment.     

Vibrational Spectroscopy of Homo- and Heterochiral Amino Acid Dimers: Conformational Landscapes

The homo- and heterochiral protonated dimers of asparagine with serine and with valine were investigated using infrared multiple-photon dissociation (IRMPD) spectroscopy. Extensive quantum-chemical calculations were used in a three-tiered strategy to screen the conformational spaces of all four dimer species. The resulting binary structures were further grouped into five different types based on their intermolecular binding topologies and subunit configurations. For each dimer species, there are eight to fourteen final conformational geometries within a 10 kJ mol⁻¹ window of the global minimum structure for each species. The comparison between the experimental IRMPD spectra and the simulated harmonic IR features allowed us to clearly identify the types of structures responsible for the observation. The monomeric subunits of the observed homo- and heterochiral dimers are compared to the corresponding protonated/neutral amino acid monomers observed experimentally in previous IRMDP/rotational spectroscopic studies. Possible chirality and kinetic influences on the experimental IRMPD spectra are discussed.     

Deamination of protonated amines to yield protonated imines

Primary and secondary amines, when examined in atmospheric pressure chemical ionization, electrospray ionization, or chemical ionization, display protonated imines in their mass spectra. These products arise formally by nucleophilic substitution at the α-carbon with loss of both ammonia and molecular hydrogen. Collision-induced dissociation (CID) is used to characterize the product ions by comparison with authentic protonated imines. Gas-phase ion/molecule reactions of protonated amines with neutral amines also yield products that correspond to protonated imines (deamination and dehydrogenation), as well as providing simple deamination products. The reaction mechanism was investigated further by reacting the deamination product, the alkyl cation, with a neutral amine. The observed dehydrogenation of the nascent protonated secondary amine indicates that the reaction sequence is loss of ammonia followed by dehydrogenation even though the isolated protonated secondary amines did not undergo dehydrogenation upon CID. Formation of the deamination products in the protonated amine/amine reaction is competitive with proton-bound dimer formation. The proton-bound dimers do not yield deamination products under CID conditions in the ion trap or in experiments performed using a pentaquadrupole instrument. This demonstrates that the geometry of the proton-bound dimer, in which the α-carbons of the alkylamines are well separated [C _a -N-H-N-C _a ], is an unsuitable entry point on the potential energy hypersurface for formation of the imine [C _a -N-C _a ]. Isolation of the proton-bound dimers in the quadrupole ion trap is achieved with low efficiency and this characteristic can be used to distinguish them from their covalently bound isomers.     

Dissociation of proton-bound complexes and proton affinity of benzamides

Proton-bound heterodimers of substituted benzamides 1–15 and N,N-dimethyl benzamides 16–30, respectively, with a series of reference bases were generated under chemical ionization conditions. Their dissociation into the protonated amide AH⁺ and protonated reference base BH⁺ was studied by metastable ion techniques and by collision-induced dissociation (CID) to examine substituent effects on the proton affinity (PA) of the benzarnides and to elucidate some aspects of the dissociation dynamics of proton-bound clusters. The PAs of the substituted benzarnides were determined by bracketing the amide by a pair of reference bases to give rise to more and less abundant signals of the protonated base in the mass-analyzed ion kinetic energy (MIKE) spectra of the proton-bound heterodimers. The substituent effects observed agree with O-protonation in both the primary and the tertiary benzamides. However, the susceptibility of the benzamide to polar substituent effects is remarkably small, which indicates a “resonance saturation”), of the amide group. The relative abundances of AH⁺ and BH⁺ in the MIKE and collisional activation (CA) mass spectra depend strongly on the pressure of the collision gas during CID, and in certain cases a reversal of the relative abundances with increasing pressure that favors the formation of BH+ from a less basic reference base is observed. Although this effect underlines the limited possibilities of the “kinetic method” for PA determination by CID of proton-bound heterodimers, it uncovers important kinetic effects during the dissociation of proton-bound heterodimers and of proton transfer reactions in the gas phase.. In the case of the protonated amide clusters, the observed intensity effects in the CA mass spectra are explained by a double-well potential energy surface caused by solvation of the protonated base by the polar amide in the protonated heterodimer.     

Heterogeneous proton-bound dimers with a high dipole moment monomer: how could we experimentally observe these anomalous ionic hydrogen bonds?

Electronic structure calculations (CBS-QB3 and G3MP2) have been used to predict a suitable method to experimentally observe the anomalous structure which is predicted to exist in a proton-bound dimer with a high dipole moment monomer. The enthalpy associated with forming the proton-bound dimer from its protonated and neutral monomers is shown to be linearly related to the difference in proton affinities which has been observed experimentally. However, unlike previous experimental studies, the linear correlation is not predicted to depend strongly, if at all, on whether the basic sites are C=O, C=N, or O(H) n-donor bases. Thermochemical measurements, then, are probably not the best method to distinguish between the structures of heterogeneous proton-bound dimers. It has been shown that a suitable method to experimentally observe the anomalous structure of proton-bound dimers containing a high dipole moment monomer (or very polar monomer) is by spectroscopic measurement. The O-H+-O asymmetric stretch is probably not the best infrared band to try to correlate with structure. The best band to observe is one which is in a region of the spectrum not masked by other absorptions and is also sensitive to the proximity of the binding proton. For example, it is shown that the methanol-free O-H stretch is very sensitive to the O-H+ bond distance for a series of heterogeneous proton-bound dimers containing methanol. It is predicted that the free O-H stretch of the methanol/acetonitrile proton-bound dimer is more closely related to the O-H stretch in protonated methanol than the O-H stretch in neutral methanol. Observations of these bands should confirm that the proton is closer to methanol in the methanol/acetonitrile proton-bound dimer despite acetonitrile having a higher proton affinity.     

DFT energy surfaces for aminopurine homodimers and their conjugate acid ions

Dimers of free nucleobases with their conjugate acid ions can be assigned to either of two categories: protonated dimers or proton-bound dimers. In the former, the extra proton attaches to a lone pair of a neutral dimer. In the latter, the extra proton is situated between two lone pairs and participates in a proton bridge. In general, proton-bound dimers are found to be more tightly held together than protonated dimers. While neutral adenine and its isomer 8-aminopurine (C(5)H(5)N(5)) are substantially more stable than their 7H tautomers, their conjugate acid ions and those of their respective 7H tautomers have nearly the same heats of formation. Correspondingly, the most stable (C(5)H(5)N(5))2H+ structures contain 7H tautomers as the neutral partner. Proton transit from one partner to the other within the most stable protonated dimer of 8-aminopurine has a low barrier (6 kJ mol(-1)). The potential energy curve for the NH stretch in that case is better fitted as a double minimum rather than as a harmonic potential. Purine-purine mismatches have been observed in nucleic acids, to which calculated (C(5)H(5)N(5))2H+ dimer geometries appear nearly isosteric.     

Tridentate ionic hydrogen-bonding interactions of the 5-fluorocytosine cationic dimer and other 5-fluorocytosine analogues characterized by IRMPD spectroscopy and electronic structure calculations

Ionic hydrogen-bonding interactions have been found in several clusters formed by 5-fluorocytosine (5-FC). The chloride and trimethylammonium cluster ions, along with the cationic (proton-bound) dimer have been characterized by infrared multiple-photon dissociation (IRMPD) spectroscopy and electronic structure calculations performed at the B2PLYP/aug-cc-pVTZ//B3LYP/6-311+G(d,p) level of theory. IRMPD action spectra, in combination with calculated spectra and relative energetics, indicate that it is most probable that predominantly a single isomer exists in each experiment. For the 5-FC-trimethylammonium cluster specifically, the calculated spectrum of the lowest-energy isomer convincingly matches the experimental spectrum. Interestingly, the cationic dimer of 5-FC was found to have a single energetically relevant isomer (Cationic-IV) involving a tridentate ionic hydrogen-bonding interaction. The three sites of intermolecular ionic hydrogen bonds in this isomer interact very efficiently, leading to a significant calculated binding energy of 180 kJ/mol. The magnitude of the calculated binding energy for this species, in combination with the strong correlation between the simulated and IRMPD spectra, suggests that a tridentate-proton-bound dimer was observed predominantly in the experiments. Comparison of the calculated relative Gibbs free energies (298 K) for this species and several of the other isomers considered also supports the likelihood of the dominant protonated dimer existing as Cationic-IV.     

Ion-neutral complexes and the proton-bound dimer of acetone

High- and low-energy collision-induced dissociation, and multi-photon infrared dissociation spectra of the proton-bound dimer of acetone all show the formation of protonated-acetone and an acetone molecule as the only dissociation products. In the present paper evidence is presented that the proton-bound dimer of acetone can exist as three isomeric ion-neutral complexes, two of which may interconvert. The dissociation channels and energetics of these three complexes are modelled using the MINDO/3 quantum chemical method.     

Infrared multiphoton dissociation spectra as a probe of ion molecule reaction mechanism: the formation of the protonated water dimer via sequential bimolecular reactions with 1,1,3,3-tetrafluorodimethyl ether

The gas-phase ion-molecule reactions of 1,1,3,3-tetrafluorodimethyl ether and water have been examined using Fourier transform ion cyclotron resonance mass spectrometry, infrared multiphoton dissociation (IRMPD) spectroscopy, and ab initio molecular orbital calculations. This reaction sequence leads to the efficient bimolecular production of the proton-bound dimer of water (H5O2+). Evidence for the dominant mechanistic pathway involving the reaction of CF2H-O=CHF+, an ion of m/z 99, with water is presented. The primary channel occurs via nucleophilic attack of water on the ion of m/z 99 (CF2H-O=CHF+), to lose formyl fluoride and yield-protonated difluoromethanol (m/z 69). Association of a second water molecule with protonated difluoromethanol generates a reactive intermediate that decomposes via a 1,4-elimination to release hydrogen fluoride and yield the proton-bound dimer of water and formyl fluoride (m/z 67). Last, the elimination of formyl fluoride occurs by the association of a third water molecule to produce H5O2+ (m/z 37). The most probable isomeric forms of the ions with m/z 99 and 69 were found using IRMPD spectroscopy and electronic structure theory calculations. Thermochemical information for reactant, transition state, and product species was obtained using MP2(full)/6-311+G**//6-31G* level of theory.     

Infrared multiple photon dissociation spectra of proton- and sodium ion-bound glycine dimers in the N-H and O-H stretching region

The proton- and the sodium ion-bound glycine homodimers are studied by a combination of infrared multiple photon dissociation (IRMPD) spectroscopy in the N-H and O-H stretching region and electronic structure calculations. For the proton-bound glycine dimer, in the region above 3100 cm (-1), the present spectrum agrees well with one recorded previously. The present work also reveals a weak, broad absorption spanning the region from 2650 to 3300 cm (-1). This feature is assigned to the strongly hydrogen-bonded and anharmonic N-H and O-H stretching modes. As well, the shared proton stretch is observed at 2440 cm (-1). The IRMPD spectra for the proton-bound glycine dimer confirms that the lowest energy structure is an ion-dipole complex between N-protonated glycine and the carboxyl group of the second glycine. This spectrum also helps to eliminate the existence of any of the higher-energy structures considered. The IRMPD spectrum for the sodium ion-bound dimer is a much simpler spectrum consisting of three bands assigned to the O-H stretch and the asymmetric and symmetric NH 2 stretching modes. The positions of these bands are very similar to those observed for the proton-bound glycine dimer. Numerous structures were considered and the experimental spectrum agrees with the B3LYP/6-31+G(d,p) predicted spectrum for the lowest energy structure, two bidentate glycine molecules bound to Na (+). Though some of the structures cannot be completely ruled out by comparing the experimental and theoretical spectra, they are energetically disfavored by at least 20 kJ mol (-1).     

Structural Investigation of Protonated Azidothymidine and Protonated Dimer

Infrared multiple photon dissociation (IRMPD) spectroscopy experiments and quantum chemical calculations have been used to explore the possible structures of protonated azidothymidine and the corresponding protonated dimer. Many interesting differences between the protonated and neutral forms of azidothymidine were found, particularly associated with keto-enol tautomerization. Comparison of computational vibrational and the experimental IMRPD spectra show good agreement and give confidence that the dominant protonated species has been identified. The protonated dimer of azidothymidine exhibits three intramolecular hydrogen bonds. The IRMPD spectrum of the protonated dimer is consistent with the spectrum of the most stable computational structure. This work brings to light interesting keto-enol tautomerization and exocyclic hydrogen bonding involving azidothymidine and its protonated dimer. The fact that one dominant protonated species is observed in the gas phase, despite both the keto and enol structures being similar in energy, is proposed to be the direct result of the electrospray ionization process in which the dominant protonated dimer structure dissociates in the most energetically favorable way.

IRMPD spectra of Gly.NH4 + and proton-bound betaine dimer: evidence for the smallest gas phase zwitterionic structures

Zwitterionic structures exist extensively in biological systems and the electric field resulting from zwitterion formation is the driving force for determination of the properties, function and activity of biological molecules, such as amino acids, peptides and proteins. It is of considerable interest and import to investigate the stabilization of zwitterionic structures in the gas phase. Infrared multiple photon dissociation (IRMPD) spectroscopy is a very powerful and sensitive technique, which may elucidate clearly the structures of both ions and ionic clusters in the gas phase, since it provides IR vibrational fingerprint information. The structures of the clusters of glycine and ammonium ion and of the betaine proton-bound homodimer have been investigated using IRMPD spectroscopy, in combination with electronic structure calculations. The experimental and calculated results indicate that zwitterionic structure of glycine may be effectively stabilized by an ammonium ion. This is the smallest zwitterionic structure of an amino acid to be demonstrated in the gas phase. On the basis of the experimental IRMPD and calculated results, it is very clear that a zwitterionic structure exists in the proton-bound betaine dimer. The proton is bound to one of the carboxylate oxygens of betaine, rather than being equally shared. Investigations of zwitterionic structures in the isolated state are essential for an understanding of the intrinsic characteristics of zwitterions and salt bridge interactions in biological systems.     

Self-Assembling of cytosine nucleoside into triply-bound dimers in acid Media. A comprehensive evaluation of proton-bound pyrimidine nucleosides by electrospray tandem mass spectrometry,X-rays diffractometry,and theoretical calculations

Electrospray tandem mass spectrometry (ESI-MS/MS) is used to evaluate the assembling of cytosine and thymine nucleosides in the gas phase, through the formation of hydrogen bonded supermolecules. Mixtures of cytidine analogues and homologues deliver in the gas phase proton-bound heterodimers stabilized by multiple interactions, as proven by the kinetics of their dissociation into the corresponding protonated monomers. Theoretical calculations, performed on initial structures of methylcytosine homodimers available in the literature, converged to a minimized structure whereby the two pyrimidine rings interact through the formation of three hydrogen bonds of similar energy. The crystallographic data here reported show the equivalency of the two interacting pyrimidines which is attributable to the presence of an inversion center. Thymine and uracil pyrimidyl nucleosides form, by ESI, gaseous proton-bound dimers. The kinetic of their dissociation into the related protonated monomers shows that the nucleobases are weekly interacting through a single hydrogen bond. The minimized structure of the protonated heterodimer formed by thymine and N-1-methylthymine confirmed the existence of mainly one hydrogen bond which links the two nucleobases through the O4 oxygens. No crystallographic data exists on thymine proton-bound species, nor have we been able to obtain these aggregates in the solid phase. The gaseous phase, under high vacuum conditions, seems therefore a suitable environment where vanishing structures produced by ESI can be studied with a good degree of approximation.     

Experimental and Theoretical Investigation of the Proton-Bound Dimer of Lysine

The structure of the proton-bound lysine dimer has been investigated by infrared multiple photon dissociation (IRMPD) spectroscopy and electronic structure calculations. The structures of different possible isomers of the proton-bound lysine dimer have been optimized at the B3LYP/6-31 + G(d) level of theory and IR spectra calculated using the same computational method. Based on relative Gibbs free energies (298 K) calculated at the MP2/aug-cc-pVTZ//B3LYP/6-31 + G(d) level of theory, LL-CS01, and followed closely (1.1 kJ mol^–1) by LL-CS02 are the most stable non-zwitterionic isomers. At the MP2/aug-cc-pVTZ//6-31 + G(d) and MP2/aug-cc-pVTZ//6-31 + (d,p) levels of theory, isomer LL-CS02 is favored by 3.0 and 2.3 kJ mol^–1, respectively. The relative Gibbs free energies calculated by the aforementioned levels of theory for LL-CS01 and LL-CS02 are very close and strongly suggest that diagnostic vibrational signatures found in the IRMPD spectrum of the proton-bound dimer of lysine can be attributed to the existence of both isomers. LL-ZW01 is the most stable zwitterionic isomer, in which the zwitterionic structure of the neutral lysine is well stabilized by the protonated lysine moiety via a very strong intermolecular hydrogen bond. At the MP2/aug-cc-pVTZ//B3LYP/6-31 + G(d), MP2/aug-cc-pVTZ//6-31 + G(d) and MP2/aug-cc-pVTZ//6-31 + G(d,p) levels of theory, the most stable zwitterionic isomer (LL-ZW01) is less favored than LL-CS01 by 7.3, 4.1 and 2.3 kJ mol^–1, respectively. The experimental IRMPD spectrum also confirms that the proton-bound dimer of lysine largely exists as charge-solvated isomers. Investigation of zwitterionic and charge-solvated species of amino acids in the gas phase will aid in a further understanding of structure, property, and function of biological molecules.     

Photodissociation pathways and lifetimes of protonated peptides and their dimers

Photodissociation lifetimes and fragment channels of gas-phase, protonated YA(n) (n = 1,2) peptides and their dimers were measured with 266 nm photons. The protonated monomers were found to have a fast dissociation channel with an exponential lifetime of ~200 ns while the protonated dimers show an additional slow dissociation component with a lifetime of ~2 μs. Laser power dependence measurements enabled us to ascribe the fast channel in the monomer and the slow channel in the dimer to a one-photon process, whereas the fast dimer channel is from a two-photon process. The slow (1 photon) dissociation channel in the dimer was found to result in cleavage of the H-bonds after energy transfer through these H-bonds. In general, the dissociation of these protonated peptides is non-prompt and the decay time was found to increase with the size of the peptides. Quantum RRKM calculations of the microcanonical rate constants also confirmed a statistical nature of the photodissociation processes in the dipeptide monomers and dimers. The classical RRKM expression gives a rate constant as an analytical function of the number of active vibrational modes in the system, estimated separately on the basis of the equipartition theorem. It demonstrates encouraging results in predicting fragmentation lifetimes of protonated peptides. Finally, we present the first experimental evidence for a photo-induced conversion of tyrosine-containing peptides into monocyclic aromatic hydrocarbon along with a formamide molecule both found in space.     

Infrared multiphoton dissociation of small-interfering RNA anions and cations

Infrared multiphoton dissociation (IRMPD) on a linear ion trap mass spectrometer is applied for the sequencing of small interfering RNA (siRNA). Both single-strand siRNAs and duplex siRNA were characterized by IRMPD, and the results were compared with that obtained by traditional ion trap-based collision induced dissociation (CID). The single-strand siRNA anions were observed to dissociate via cleavage of the 5′ P—O bonds yielding c- and y-type product ions as well as undergo neutral base loss. Full sequence coverage of the siRNA anions was obtained by both IRMPD and CID. While the CID mass spectra were dominated by base loss ions, accounting for ∼25% to 40% of the product ion current, these ions were eliminated through secondary dissociation by increasing the irradiation time in the IRMPD mass spectra to produce higher abundances of informative sequence ions. With longer irradiation times, however, internal ions corresponding to cleavage of two 5′ P—O bonds began to populate the product ion mass spectra as well as higher abundances of [a − Base] and w-type ions. IRMPD of siRNA cations predominantly produced c- and y-type ions with minimal contributions of [a − Base] and w-type ions to the product ion current; the presence of only two complementary series of product ions in the IRMPD mass spectra simplified spectral interpretation. In addition, IRMPD produced high abundances of protonated nucleobases, [G + H]⁺, [A + H]⁺, and [C + H]⁺, which were not detected in the CID mass spectra due to the low-mass cut-off associated with conventional CID in ion traps. CID and IRMPD using short irradiation times of duplex siRNA resulted in strand separation, similar to the dissociation trends observed for duplex DNA. With longer irradiation times, however, the individual single-strands underwent secondary dissociation to yield informative sequence ions not obtained by CID.     

Probing Protonation Sites of Isolated Flavins Using IR Spectroscopy: From Lumichrome to the Cofactor Flavin Mononucleotide

Infrared spectra of the isolated protonated flavin molecules lumichrome, lumiflavin, riboflavin (vitamin B₂), and the biologically important cofactor flavin mononucleotide are measured in the fingerprint region (600–1850 cm^?1) by means of IR multiple‐photon dissociation (IRMPD) spectroscopy. Using density functional theory calculations, the geometries, relative energies, and linear IR absorption spectra of several low‐energy isomers are calculated. Comparison of the calculated IR spectra with the measured IRMPD spectra reveals that the N10 substituent on the isoalloxazine ring influences the protonation site of the flavin. Lumichrome, with a hydrogen substituent, is only stable as the N1‐protonated tautomer and protonates at N5 of the pyrazine ring. The presence of the ribityl unit in riboflavin leads to protonation at N1 of the pyrimidinedione moiety, and methyl substitution in lumiflavin stabilizes the tautomer that is protonated at O2. In contrast, flavin mononucleotide exists as both the O2‐ and N1‐protonated tautomers. The frequencies and relative intensities of the two C?O stretch vibrations in protonated flavins serve as reliable indicators for their protonation site.     

Differences in structure, energy, and spectrum between neutral, protonated, and deprotonated phenol dimers: comparison of various density functionals with ab initio theory

We have carried out extensive calculations for neutral, cationic protonated, anionic deprotonated phenol dimers. The structures and energetics of this system are determined by the delicate competition between H-bonding, H-π interaction and π-π interaction. Thus, the structures, binding energies and frequencies of the dimers are studied by using a variety of functionals of density functional theory (DFT) and M?ller-Plesset second order perturbation theory (MP2) with medium and extended basis sets. The binding energies are compared with those of highly reliable coupled cluster theory with single, double, and perturbative triple excitations (CCSD(T)) at the complete basis set (CBS) limit. The neutral phenol dimer is unique in the sense that its experimental rotational constants have been measured. The geometry of the neutral phenol dimer is governed by the hydrogen bond formed by two hydroxyl groups and the H-π interaction between two aromatic rings, while the structure of the protonated/deprotonated phenol dimers is additionally governed by the electrostatic and induction effects due to the short strong hydrogen bond (SSHB) and the charges populated in the aromatic rings in the ionic systems. Our salient finding is the substantial differences in structure between neutral, protonated, and deprotonated phenol dimers. This is because the neutral dimer involves in both H(π)···O and H(π)···π interactions, the protonated dimer involves in H(π)···π interactions, and the deprotonated dimer involves in a strong H(π)···O interaction. It is important to compare the reliability of diverse computational approaches employed in quantum chemistry on the basis of the calculational results of this system. MP2 calculations using a small cc-pVDZ basis set give reasonable structures, but those using extended basis sets predict wrong π-stacked structures due to the overestimation of the dispersion energies of the π-π interactions. A few new DFT functionals with the empirical dispersion give reliable results consistent with the CCSD(T)/CBS results. The binding energies of the neutral, cationic protonated, and anionic deprotonated phenol dimers are estimated to be more than 28.5, 118.2, and 118.3 kJ mol(-1), respectively. The energy components of the intermolecular interactions for the neutral, protonated and deprotonated dimers are analyzed.