2,4-Dihydroxy and O2 Protonated Tautomers of dThd and Thd Coexist in the Gas Phase: Methylation Alters Protonation Preferences versus dUrd and Urd

The gas-phase structures of protonated thymidine, [dThd + H]⁺, and its modified form, protonated 5-methyluridine, [Thd + H]⁺, are examined by infrared multiple photon dissociation (IRMPD) action spectroscopy combined with electronic structure calculations. IRMPD action spectra are measured over the ranges extending from ~600 to 1900 cm^–1 and ~2800 to 3800 cm^–1 using the FELIX free electron laser and an optical parametric oscillator/amplifier (OPO/OPA) laser system, respectively. Comparisons between the B3LYP/6-311+G(d,p) linear IR spectra calculated for the stable low-energy conformers and the measured IRMPD spectra are used to determine the most favorable tautomeric conformations of [dThd + H]⁺ and [Thd + H]⁺ and to identify those populated in the experiments. Both B3LYP and MP2 levels of theory predict a minor 2,4-dihydroxy tautomer as the ground-state conformer of [dThd + H]⁺ and [Thd + H]⁺ indicating that the 2'-hydroxyl substituent of Thd does not exert a significant impact on the structural features. [dThd + H]⁺ and [Thd + H]⁺ share parallel IRMPD spectral profiles and yields in both the FELIX and OPO regions. Comparisons between the measured IRMPD and calculated IR spectra suggest that minor 2,4-dihydroxy tautomers and O2 protonated conformers of [dThd + H]⁺ and [Thd + H]⁺ are populated in the experiments. Comparison of this work to our previous IRMPD spectroscopy study of protonated 2'-deoxyuridine and uridine suggests that the 5-methyl substituent alters the preferences of O2 versus O4 protonation.

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Infrared Multiple Photon Dissociation Action Spectroscopy and Theoretical Studies of Triethyl Phosphate Complexes: Effects of Protonation and Sodium Cationization on Structure

The gas-phase structures of protonated and sodium cationized complexes of triethyl phosphate, [TEP + H]⁺ and [TEP + Na]⁺, are examined via infrared multiple photon dissociation (IRMPD) action spectroscopy using tunable IR radiation generated by a free electron laser, a Fourier transform ion cyclotron resonance mass spectrometer with an electrospray ionization source, and theoretical electronic structure calculations. Measured IRMPD action spectra are compared to linear IR spectra calculated at the B3LYP/6-31 G(d,p) level of theory to identify the structures accessed in the experimental studies. For comparison, theoretical studies of neutral TEP are also performed. Sodium cationization and protonation produce changes in the central phosphate geometry, including an increase in the alkoxy ∠OPO bond angle and shortening of the alkoxy P–O bond. Changes associated with protonation are more pronounced than those produced by sodium cationization.     

Infrared Multiple Photon Dissociation Action Spectroscopy and Theoretical Studies of Diethyl Phosphate Complexes: Effects of Protonation and Sodium Cationization on Structure

The gas-phase structures of deprotonated, protonated, and sodium-cationized complexes of diethyl phosphate (DEP) including [DEP − H]⁻, [DEP + H]⁺, [DEP + Na]⁺, and [DEP − H + 2Na]⁺ are examined via infrared multiple photon dissociation (IRMPD) action spectroscopy using tunable IR radiation generated by a free electron laser, a Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS) with an electrospray ionization (ESI) source, and theoretical electronic structure calculations. Measured IRMPD spectra are compared to linear IR spectra calculated at the B3LYP/6-31G(d,p) level of theory to identify the structures accessed in the experimental studies. For comparison, theoretical studies of neutral complexes are also performed. These experiments and calculations suggest that specific geometric changes occur upon the binding of protons and/or sodium cations, including changes correlating to nucleic acid backbone geometry, specifically P–O bond lengths and ∠OPO bond angles. Information from these observations may be used to gain insight into the structures of more complex systems, such as nucleotides and solvated nucleic acids.     

Mass spectrometric monitoring of oxidation of aliphatic C6–C8 hydrocarbons and ethanol in low pressure oxygen and air plasmas

Experimental and theoretical studies on the oxidation of saturated hydrocarbons (n‐hexane, cyclohexane, n‐heptane, n‐octane and isooctane) and ethanol in 28 Torr O₂ or air plasma generated by a hollow cathode discharge ion source were made. Ions corresponding to [M + 15]⁺ and [M + 13]⁺ in addition to [M ? H]⁺ and [M ? 3H]⁺ were detected as major ions where M is the sample molecule. The ions [M + 15]⁺ and [M + 13]⁺ were assigned as oxidation products, [M ? H + O]⁺ and [M ? 3H + O]⁺, respectively. By the tandem mass spectrometry analysis of [M ? H + O]⁺ and [M ? 3H + O]⁺, H₂O, olefins (and/or cycloalkanes) and oxygen‐containing compounds were eliminated from these ions. Ozone as one of the terminal products in the O₂ plasma was postulated as the oxidizing reagent. As an example, the reactions of C₆H₁₄^+? with O₂ and of C₆H₁₃⁺ (CH₃CH₂CH⁺CH₂CH₂CH₃) with ozone were examined by density functional theory calculations. Nucleophilic interaction of ozone with C₆H₁₃⁺ leads to the formation of protonated ketone, CH₃CH₂C(=OH⁺)CH₂CH₂CH₃. In air plasma, [M ? H + O]⁺ became predominant over carbocations, [M ? H]⁺ and [M ? 3H]⁺. For ethanol, the protonated acetic acid CH₃C(OH)₂⁺ (m/z 61.03) was formed as the oxidation product. The peaks at m/z 75.04 and 75.08 are assigned as protonated ethyl formate and protonated diethyl ether, respectively, and that at m/z 89.06 as protonated ethyl acetate. Copyright © 2016 John Wiley & Sons, Ltd.     

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.     

Influence of Transition Metal Cationization versus Sodium Cationization and Protonation on the Gas-Phase Tautomeric Conformations and Stability of Uracil: Application to [Ura+Cu]<Superscript>+</Superscript> and [Ura+Ag]<Superscript>+</Superscript>

The gas-phase conformations of transition metal cation-uracil complexes, [Ura+Cu]⁺ and [Ura+Ag]⁺, were examined via infrared multiple photon dissociation (IRMPD) action spectroscopy and theoretical calculations. IRMPD action spectra were measured over the IR fingerprint and hydrogen-stretching regions. Structures and linear IR spectra of the stable tautomeric conformations of these complexes were initially determined at the B3LYP/6-31G(d) level. The four most stable structures computed were also examined at the B3LYP/def2-TZVPPD level to improve the accuracy of the predicted IR spectra. Two very favorable modes of binding are found for [Ura+Cu]⁺ and [Ura+Ag]⁺ that involve O2N3 bidentate binding to the 2-keto-4-hydroxy minor tautomer and O4 monodentate binding to the canonical 2,4-diketo tautomer of Ura. Comparisons between the measured IRMPD and calculated IR spectra enable elucidation of the conformers present in the experiments. These comparisons indicate that both favorable binding modes are represented in the experimental tautomeric conformations of [Ura+Cu]⁺ and [Ura+Ag]⁺. B3LYP suggests that Cu⁺ exhibits a slight preference for O4 binding, whereas Ag⁺ exhibits a slight preference for O2N3 binding. In contrast, MP2 suggests that both Cu⁺ and Ag⁺ exhibit a more significant preference for O2N3 binding. The relative band intensities suggest that O4 binding conformers comprise a larger portion of the population for [Ura+Ag]⁺ than [Ura+Cu]⁺. The dissociation behavior and relative stabilities of the [Ura+M]⁺ complexes, M⁺ = Cu⁺, Ag⁺, H⁺, and Na⁺) are examined via energy-resolved collision-induced dissociation experiments. The IRMPD spectra, dissociation behaviors, and binding preferences of Cu⁺ and Ag⁺ are compared with previous and present results for those of H⁺ and Na⁺.

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Rational Control of Conformational Distributions and Mixed‐Valence Characteristics in Diruthenium Complexes

The electronic characteristics of mixed‐valence complexes are often inferred from the shape of the inter‐valence charge transfer (IVCT) band, which usually falls in the near infrared (NIR) region, and relationships derived from Marcus‐Hush theory. These analyses typically assume one single, dominant molecular conformation. The NIR spectra of the prototypical delocalised (Class III Robin–Day mixed‐valence) complexes [{Ru(pp)Cp’}₂(μ‐C≡C?C≡C)]⁺ ([ 1 ]⁺: Cp’=Cp, pp=(PPh₃)₂; [ 2 ]⁺: Cp’=Cp, pp=dppe; [ 3 ]⁺: Cp’=Cp*, pp=dppe) feature a ‘two‐band’ pattern, which complicates band‐shape analysis using these traditional methods. In the past, the appearance of sub‐bands within or near the IVCT transition has been attributed to vibronic effects or localised d‐d transitions. Quantum‐chemical modelling of a series of rotational conformers of [ 1 ]⁺–[ 3 ]⁺ reveals the two components that contribute to the NIR absorption band envelope to be a π‐π* transition and an MLCT transition. The MLCT components only gain appreciable intensity when the orientation of the half‐sandwich ruthenium ligand spheres deviates from idealised cis (Ω P?Ru?Ru?P=0°) or trans (Ω P?Ru?Ru?P=180°) conformations. The increased steric demand of the supporting ligands, together with some underlying inter‐phosphine ligand T‐shaped CH???π stacking interactions across the series [ 1 ]⁺ to [ 2 ]⁺ to [ 3 ]⁺ results in local minima biased towards such non‐idealised conformations of the metal‐ligand fragments (Ω P?Ru?Ru?P=33–153°). Experimentally, this is indicated by appearance of multiple bands within the IR (C≡C) band envelopes and increasing intensity of the higher‐energy MLCT transition(s) relative to the π‐π* transition across the series, and the appearance of a pronounced ‘two‐band’ pattern in the experimental NIR absorption envelopes. These conformational effects and the methods of analysis presented here, which combine analysis of IR and NIR spectra with quantum‐chemical calculations on a range of energetically similar conformational minima, are expected to be quite general for mixed‐valence systems.     

Self‐Assembled Multimetallic/Peptide Complexes: Structures and Unimolecular Reactions of [Mn(GlyGly−H)2n−1]+ and Mn+1(GlyGly−H2n]2+ Clusters in the Gas Phase

The unimolecular chemistry and structures of self‐assembled complexes containing multiple alkaline‐earth‐metal dications and deprotonated GlyGly ligands are investigated. Singly and doubly charged ions [M_n(GlyGly?H)_n‐1]⁺ (n=2–4), [M_n+1(GlyGly?H)_2n]²⁺ (n=2,4,6), and [M(GlyGly?H)GlyGly]⁺ were observed. The losses of 132 Da (GlyGly) and 57 Da (determined to be aminoketene) were the major dissociation pathways for singly charged ions. Doubly charged Mg²⁺ clusters mainly lost GlyGly, whereas those containing Ca²⁺ or Sr²⁺ also underwent charge separation. Except for charge separation, no loss of metal cations was observed. Infrared multiple photon dissociation spectra were the most consistent with the computed IR spectra for the lowest energy structures, in which deprotonation occurs at the carboxyl acid groups and all amide and carboxylate oxygen atoms are complexed to the metal cations. The N?H stretch band, observed at 3350 cm^?1, is indicative of hydrogen bonding between the amine nitrogen atoms and the amide hydrogen atom. This study represents the first into large self‐assembled multimetallic complexes bound by peptide ligands.     

On the Interaction between Deprotonated Cytosine [C(−H)]− and Ba2+: Infrared Multiphoton Spectroscopy and Dynamics

Gas-phase interactions between Ba²⁺ and deprotonated cytosine (C_(−H)) were studied in [C_(−H)Ba]⁺ and [C_(−H)BaC]⁺ complexes by IRMPD spectroscopy coupled to tandem mass-spectrometry in combination with DFT calculations. For the [C_(−H)BaC]⁺ complex only one [C_(−H)KAN1O−Ba-C_anti]⁺ isomer was found, although the presence of another structure cannot be excluded. This isomer features a central tetracoordinated Ba²⁺ that simultaneously interacts with keto-amino [C_(−H)]⁻ deprotonated on N1 and neutral keto-amino C. Both moieties are in different planes as a consequence of an additional NH…O=C hydrogen bond between C and [C_(−H)] ⁻ . A sequential IRMPD dynamics is observed in this complex. For the [C_(−H)Ba]⁺ complex produced by electrospray ionization two isomers ([C_(−H)KAN1OBa]⁺ and [C_(−H)KAN3OBa]⁺) were identified, in which Ba²⁺ interacts simultaneously with the C=O group and the N1 or N3 atom of the keto-amino [C_(−H)]⁻, respectively. A comparison with the related [C_(−H)Pb]⁺ complex (J. Y. Salpin et al., Chem. Phys. Chem. 2014 , 15, 2959–2971) is also presented.     

IRMPD Action Spectroscopy,ER-CID Experiments,and Theoretical Studies of Sodium Cationized Thymidine and 5-Methyluridine: Kinetic Trapping During the ESI Desolvation Process Preserves the Solution Structure of [Thd+Na]<Superscript>+</Superscript>

Thymidine (dThd) is a fundamental building block of DNA nucleic acids, whereas 5-methyluridine (Thd) is a common modified nucleoside found in tRNA. In order to determine the conformations of the sodium cationized thymine nucleosides [dThd+Na]⁺ and [Thd+Na]⁺ produced by electrospray ionization, their infrared multiple photon dissociation (IRMPD) action spectra are measured. Complementary electronic structure calculations are performed to determine the stable low-energy conformations of these complexes. Geometry optimizations and frequency analyses are performed at the B3LYP/6-311+G(d,p) level of theory, whereas energies are calculated at the B3LYP/6-311+G(2d,2p) level of theory. As protonation preferentially stabilizes minor tautomers of dThd and Thd, tautomerization facilitated by Na⁺ binding is also considered. Comparisons of the measured IRMPD and computed IR spectra find that [dThd+Na]⁺ prefers tridentate (O2,O4',O5') coordination to the canonical 2,4-diketo form of dThd with thymine in a syn orientation. In contrast, [Thd+Na]⁺ prefers bidentate (O2,O2') coordination to the canonical 2,4-diketo tautomer of Thd with thymine in an anti orientation. Although 2,4-dihydroxy tautomers and O2 protonated thymine nucleosides coexist in the gas phase, no evidence for minor tautomers is observed for the sodium cationized species. Consistent with experimental observations, the computational results confirm that the sodium cationized thymine nucleosides exhibit a strong preference for the canonical form of the thymine nucleobase. Survival yield analyses based on energy-resolved collision-induced dissociation (ER-CID) experiments suggest that the relative stabilities of protonated and sodium cationized dThd and Thd follow the order [dThd+H]⁺ < [Thd+H]⁺ < [dThd+Na]⁺ < [Thd+Na]⁺.

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Mass spectra of sulfinamide derivatives and identification of their thermal decomposition products

The mass spectra of 30 sulfinamide derivatives (RSONHR', R' alkyl or p-XC₆H₄) are reported. Most of the spectra had peaks attributable to thermal decomposition products. For some compounds these were identified by pyrolysis under similar conditions to be: RSO₂NHR', RSO₂SR, RSSR and NH₂R' (in all kinds of sulfinyl amides); RSNHR' (in the case of arylsulfinyl arylamides); RSO₂C₆H₄NH₂, RSOC₆H₄NH₂ and RSC₆H₄NH₂ (in the case of arylsulfinyl arylamides of the type of X = H) The mass spectra of the three thermally stable compounds showed that there are several kinds of common fragment ions. The mass spectra of the thermally labile compounds had two groups of ions; (i) characteristic fragment ions of the intact molecules and (ii) the molecular ions of the thermal decomposition products. It was concluded that the sulfinamides give the following ions after electron impact: [M]⁺, [M ? R]⁺, [M ? R + H]⁺, [M ? SO]⁺, [RS]⁺, [NHR']⁺, [NHR' + H]⁺, [RSO]⁺, [RSO + H]⁺, [R]⁺, [R + H]⁺, [R']⁺ and [M ? OH]⁺, and that the thermal decomposition products give the following ions: [RSO₂SR]⁺, [RSSR]⁺, [M ? O]⁺, [M + O]⁺ and [RSOC₆H₄NH₂]⁺.     

Characterization of five [C4H7O]+ Ion structures. Fragmentation of the 2-pentanone molecular ion

The [C₄H₇0]⁺ ions [CH₂?CH? C(?OH)CH₃]⁺ (1), [CH₃CH?CH? C(?OH)H]⁺ (2), [CH₂?C(CH₃)C(?OH)H]⁺ (3), [Ch₃CH₂CH₂C?O]⁺ (4) and [(CH₃)₂CHC?O]⁺ (5) have been characterized by their collision-induced dissociation (CID) mass spectra and charge stripping mass spectra. The ions 1–3 were prepared by gas phase protonation of the relevant carbonyl compounds while 4 and 5 were prepared by dissociative electron impact ionization of the appropriate carbonyl compounds. Only 2 and 3 give similar spectra and are difficult to distinguish from each other; the remaining ions can be readily characterized by either their CID mass spectra or their charge stripping mass spectra. The 2-pentanone molecular ion fragments by loss of the C(1) methyl and the C(5) methyl in the ratio 60:40 for metastable ions; at higher internal energies loss of the C(1) methyl becomes more favoured. Metastable ion characteristics, CID mass spectra and charge stripping mass spectra all show that loss of the C(1) methyl leads to formation of the acyl ion 4, while loss of the C(5) methyl leads to formation of protonated vinyl methyl ketone (1). These results are in agreement with the previously proposed potential energy diagram for the [C₅H₁₀O]⁺˙ system.     

Fragmentation of protonated O,O-diethyl O-aryl phosphorothionates in tandem mass spectral analysis

The gas-phase ion chemistry of protonated O,O-diethyl O-aryl phosphorothionates was studied with tandem mass spectrometric and ab initio theoretical methods. Collision-activated dissociation (CAD) experiments were performed for the [M+H]⁺ ions on a triple quadrupole mass spectrometer. Various amounts of internal energy were deposited into the ions upon CAD by variation of the collision energy and collision gas pressure. In addition to isobutane, deuterated isobutane C₄D₁₀ also was used as reagent gas in chemical ionization. The daughter ions [M+H?C₂H₄]⁺ and [M+H?2C₂H₄]⁺ dominate the CAD spectra. These fragments arise via various pathways, each of which involves γ-proton migration. Formation of the terminal ions [M+H?2C₂H₄?H₂O]⁺, [M+H?2C₂H₄?H₂S]⁺, [ZPhOH₂]⁺, [ZPhSH₂]⁺, and [ZPhS]⁺ [Z = substituent(s) on the benzene ring] suggests that (1) the fragmenting [M+H]⁺ ions of O,O-diethyl O-aryl phosphorothionates have protons attached on the oxygen of an ethoxy group and on the oxygen of the phenoxy group; (2) thiono-thiolo rearrangement by aryl migration to sulfur occurs; (3) the fragmenting rear-ranged [M+H]⁺ ions have protons attached on the oxygen of an ethoxy group and on the sulfur of the thiophenoxy group. To get additional support for our interpretation of the mass spectrometric results, some characteristics of three protomers of O,O-diethyl O-phenyl phosphorothionate were investigated by carrying out ab initio molecular orbital calculations at the RHF/3–21G* level of theory.     

Gas‐phase interactions of organotin compounds with glycine

Gas‐phase interactions of organotins with glycine have been studied by combining mass spectrometry experiments and quantum calculations. Positive‐ion electrospray spectra show that the interaction of di‐ and tri‐organotins with glycine results in the formation of [(R)₂Sn(Gly)‐H]⁺and [(R)₃Sn(Gly)]⁺ ions, respectively. Di‐organotin complexes appear much more reactive than those involving tri‐organotins. (MS/MS) spectra of the [(R)₃Sn(Gly)]⁺ ions are indeed simple and only show elimination of intact glycine, generating the [(R)₃Sn]⁺ carbocation. On the other hand, MS/MS spectra of [(R)₂Sn(Gly)‐H]⁺complexes are characterized by numerous fragmentation processes. Six of them, associated with elimination of H₂O, CO, H₂O + CO and formation of [(R)₂SnOH]⁺ (?57 u),[(R)₂SnNH₂]⁺( ?58 u) and [(R)₂SnH]⁺ (?73 u), are systematically observed. Use of labeled glycines notably concludes that the hydrogen atoms eliminated in water and H₂O + CO are labile hydrogens. A similar conclusion can be made for hydrogens of [(R₂)SnOH]⁺and [(R₂)SnNH₂]⁺ions. Interestingly, formation [(R)₂SnH]⁺ ions is characterized by a migration of one the α hydrogen of glycine onto the metallic center. Finally, several dissociation routes are observed and are characteristic of a given organic substituent. Calculations indicated that the interaction between organotins and glycine is mostly electrostatic. For [(R)₂Sn(Gly)‐H]⁺complexes, a preferable bidentate interaction of the type η²‐O,NH2 is observed, similar to that encountered for other metal ions. [(R)₃Sn]⁺ ions strongly stabilize the zwitterionic form of glycine, which is practically degenerate with respect to neutral glycine. In addition, the interconversion between both forms is almost barrierless. Suitable mechanisms are proposed in order to account for the most relevant fragmentation processes. Copyright © 2013 John Wiley & Sons, Ltd.     

Relationships between structure,ionization profile and sensitivity of exogenous anabolic steroids under electrospray ionization and analysis in human urine using liquid chromatography–tandem mass spectrometry

The relationships between the ionization profile, sensitivity, and structures of 64 exogenous anabolic steroids (groups I–IV) was investigated under electrospray ionization (ESI) conditions. The target analytes were ionized as [M + H]⁺ or [M + H–nH₂O]⁺ in the positive mode, and these ions were used as precursor ions for selected reaction monitoring analysis. The collision energy and Q3 ions were optimized based on the sensitivity and selectivity. The limits of detection (LODs) were 0.05–20 ng/mL for the 64 steroids. The LODs for 38 compounds, 14 compounds and 12 compounds were in the range of 0.05–1, 2–5 and 10–20 ng/mL, respectively. Steroids including the conjugated keto‐functional group at C3 showed good proton affinity and stability, and generated the [M + H]⁺ ion as the most abundant precursor ion. In addition, the LODs of steroids using the [M + H]⁺ ion as the precursor ion were mostly distributed at low concentrations. In contrast, steroids containing conjugated/unconjugated hydroxyl functional groups at C3 generated [M + H ? H₂O]⁺ or [M + H ? 2H₂O]⁺ ions, and these steroids showed relatively high LODs owing to poor stability and multiple ion formation. An LC‐MS/MS method based on the present ionization profile was developed and validated for the determination of 78 steroids (groups I–V) in human urine. Copyright © 2015 John Wiley & Sons, Ltd.     

Structure and further fragmentation of significant [a3 + Na − H]+ ions from sodium‐cationized peptides

A good understanding of gas‐phase fragmentation chemistry of peptides is important for accurate protein identification. Additional product ions obtained by sodiated peptides can provide useful sequence information supplementary to protonated peptides and improve protein identification. In this work, we first demonstrate that the sodiated a₃ ions are abundant in the tandem mass spectra of sodium‐cationized peptides although observations of a₃ ions have rarely been reported in protonated peptides. Quantum chemical calculations combined with tandem mass spectrometry are used to investigate this phenomenon by using a model tetrapeptide GGAG. Our results reveal that the most stable [a₃ + Na ? H]⁺ ion is present as a bidentate linear structure in which the sodium cation coordinates to the two backbone carbonyl oxygen atoms. Due to structural inflexibility, further fragmentation of the [a₃ + Na ? H]⁺ ion needs to overcome several relatively high energetic barriers to form [b₂ + Na ? H]⁺ ion with a diketopiperazine structure. As a result, low abundance of [b₂ + Na ? H]⁺ ion is detected at relatively high collision energy. In addition, our computational data also indicate that the common oxazolone pathway to generate [b₂ + Na ? H]⁺ from the [a₃ + Na ? H]⁺ ion is unlikely. The present work provides a mechanistic insight into how a sodium ion affects the fragmentation behaviors of peptides. Copyright © 2015 John Wiley & Sons, Ltd.     

The ESI CAD fragmentations of protonated 2,4,6‐tris(benzylamino)‐ and tris(benzyloxy)‐1,3,5‐triazines involve benzyl–benzyl interactions: a DFT study

The electrospray ionization collisionally activated dissociation (CAD) mass spectra of protonated 2,4,6‐tris(benzylamino)‐1,3,5‐triazine (1) and 2,4,6‐tris(benzyloxy)‐1,3,5‐triazine (6) show abundant product ion of m/z 181 (C₁₄H₁₃⁺). The likely structure for C₁₄H₁₃⁺ is α‐[2‐methylphenyl]benzyl cation, indicating that one of the benzyl groups must migrate to another prior to dissociation of the protonated molecule. The collision energy is high for the ‘N’ analog (1) but low for the ‘O’ analog (6) indicating that the fragmentation processes of 1 requires high energy. The other major fragmentations are [M + H‐toluene]⁺ and [M + H‐benzene]⁺ for compounds 1 and 6, respectively. The protonated 2,4,6‐tris(4‐methylbenzylamino)‐1,3,5‐triazine (4) exhibits competitive eliminations of p‐xylene and 3,6‐dimethylenecyclohexa‐1,4‐diene. Moreover, protonated 2,4,6‐tris(1‐phenylethylamino)‐1,3,5‐triazine (5) dissociates via three successive losses of styrene. Density functional theory (DFT) calculations indicate that an ion/neutral complex (INC) between benzyl cation and the rest of the molecule is unstable, but the protonated molecules of 1 and 6 rearrange to an intermediate by the migration of a benzyl group to the ring ‘N’. Subsequent shift of a second benzyl group generates an INC for the protonated molecule of 1 and its product ions can be explained from this intermediate. The shift of a second benzyl group to the ring carbon of the first benzyl group followed by an H‐shift from ring carbon to ‘O’ generates the key intermediate for the formation of the ion of m/z 181 from the protonated molecule of 6. The proposed mechanisms are supported by high resolution mass spectrometry data, deuterium‐labeling and CAD experiments combined with DFT calculations. Copyright © 2012 John Wiley & Sons, Ltd.     

The unexpected formation of [M − H]+ species during MALDI and dopant‐free APPI MS analysis of novel antineoplastic curcumin analogues

Unusual ionization behavior was observed with novel antineoplastic curcumin analogues during the positive ion mode of matrix‐assisted laser desorption ionization (MALDI) and dopant‐free atmospheric pressure photoionization (APPI). The tested compounds produced an unusual significant peak designated as [M ? H]⁺ ion along with the expected [M + H]⁺ species. In contrast, electrospray ionization, atmospheric pressure chemical ionization and the dopant‐mediated APPI (dopant‐APPI) showed only the expected [M + H]⁺ peak. The [M ? H]⁺ ion was detected with all evaluated curcumin analogues including phosphoramidates, secondary amines, amides and mixed amines/amides. Our experiments revealed that photon energy triggers the ionization of the curcumin analogues even in the absence of any ionization enhancer such as matrix, solvent or dopant. The possible mechanisms for the formation of both [M ? H]⁺ and [M + H]⁺ ions are discussed in this paper. In particular, three proposed mechanisms for the formation of [M ? H]⁺ were evaluated. The first mechanism involves the loss of H₂ from the protonated [M + H]⁺ species. The other two mechanisms include hydrogen transfer from the analyte radical cation or hydride abstraction from the neutral analyte molecule. Copyright © 2014 John Wiley & Sons, Ltd.     

Some aspects of the mass spectra of triptycene and tri- and diphenylmethanes

Mass spectra of isotope-labeled triptycenes, triphenylmethanes and diphenylmethanes rule out the bulk of postulated decomposition mechanisms and fragment-ion structures. The formation of [M ? H]⁺ and [M ? 2H]²⁺ from triptycene, of [M ? H]⁺, [M ? CH₃]⁺ and [M ? CH₄]⁺ from triphenylmethane, and of [M ? H]²⁺ and [M ? 2H]²⁺-as well as the previously reported [M ? H]⁺ and [M ? CH₃]⁺-from diphenylmethane all seem to be preceded or accompanied by complete loss of position identity of the α and ring hydrogens in the original molecules. A statistical preference for loss of α hydrogens is found in the process leading to [M ? 2H]⁺ and [M ? H]²⁺ from triptycene, as in the formation of [M ? H]²⁺ from toluene.