Colors for Molecular Masses: Fusion of Spectroscopy and Mass Spectrometry for Identification of Biomolecules

We present an approach that integrates ultraviolet (UV) photofragmentation spectroscopy of cold ions with high-resolution Orbitrap mass spectrometry (MS) and uses mathematical analysis of the recorded 2D data arrays for structural identification of biomolecules. The synergy of the two orthogonal techniques makes these arrays unique fingerprints of molecular ions, enabling their reliable identifications. Using preliminary created libraries of fingerprints, the UV-MS approach was successfully applied for quantitative identification of exact isobaric molecules in their mixtures, which is one of the challenging cases for mass spectrometry. We also demonstrate how the UV and fragmentation mass spectra of unknown chemical components of a mixture can be recovered from its fingerprint even without a use of library

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 <i>T</i> = 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

Identification of Tyrosine-Phosphorylated Peptides Using Cold Ion Spectroscopy

The accurate and unambiguous detection of post-translational modifications in proteins and peptides remains a challenging task. We report here the use of cold ion spectroscopy for the identification of phosphorylated tyrosine residues in peptides. This approach employs the wavelength-specific UV fragmentation of cryogenically cooled protonated peptides in the gas phase. In addition to the appearance of specific photofragments, the phosphorylation of tyrosine induces large spectral shifts of the peptide electronic band origins. Quantum chemical calculations and experiments together suggest a certain generality of the use of such shifts in the spectroscopic identification of phosphotyrosines. The enhanced selectivity offered by the joint application of wavelength-specific fragmentation and mass spectrometry of cold molecules can also be used in the identifications of aromatic residues in protonated peptides and, potentially, of other UV-absorbing groups in a variety of large polyatomic ions

Microhydration Effects on the Encapsulation of Potassium Ion by Dibenzo-18-Crown‑6

We have measured electronic and conformer-specific vibrational spectra of hydrated dibenzo-18-crown-6 (DB18C6) complexes with potassium ion, K⁺•DB18C6•(H₂O)_<i>n</i> (<i>n</i> = 1–5), in a cold, 22-pole ion trap. We also present for comparison spectra of Rb⁺•DB18C6•(H₂O)₃ and Cs⁺•DB18C6•(H₂O)₃ complexes. We determine the number and the structure of conformers by analyzing the spectra with the aid of quantum chemical calculations. The K⁺•DB18C6•(H₂O)₁ complex has only one conformer under the conditions of our experiment. For K⁺•DB18C6•(H₂O)_<i>n</i> with <i>n</i> = 2 and 3, there are at least two conformers even under the cold conditions, whereas Rb⁺•DB18C6•(H₂O)₃ and Cs⁺•DB18C6•(H₂O)₃ each exhibit only one isomer. The difference can be explained by the optimum matching in size between the K⁺ ion and the crown cavity; because the K⁺ ion can be deeply encapsulated by DB18C6 and the interaction between the K⁺ ion and the H₂O molecules becomes weak, different kinds of hydration geometries can occur for the K⁺•DB18C6 complex, giving multiple conformations in the experiment. For K⁺•DB18C6•(H₂O)_<i>n</i> (<i>n</i> = 4 and 5) complexes, only a single isomer is found. This is attributed to a cooperative effect of the H₂O molecules on the hydration of K⁺•DB18C6; the H₂O molecules form a ring, which is bound on top of the K⁺•DB18C6 complex. According to the stable structure determined in this study, the K⁺ ion in the K⁺•DB18C6•(H₂O)_<i>n</i> complexes tends to be pulled largely out from the crown cavity by the H₂O molecules with increasing <i>n</i>. Multiple conformations observed for the K⁺ complexes will have an advantage for the effective capture of the K⁺ ion over the other alkali metal ions by DB18C6 because of entropic effects on the formation of hydrated complexes

A Decapeptide Hydrated by Two Waters: Conformers Determined by Theory and Validated by Cold Ion Spectroscopy

The intrinsic structures of biomolecules in the gas phase may not reflect their native solution geometries. Microsolvation of the molecules bridges the two environments, enabling a tracking of molecular structural changes upon hydration at the atomistic level. We employ density functional calculations to compute a large pool of structures and vibrational spectra for a gas-phase complex, in which a doubly protonated decapeptide, gramicidin S, is solvated by two water molecules. Though most vibrations of this large complex are treated in a harmonic approximation, the water molecules and the vibrations of the host ion coupled to them are locally described by a quantum mechanical vibrational self-consistent field theory with second-order perturbation correction (VSCF-PT2). Guided and validated by the available cold ion spectroscopy data, the computational analysis identifies structures of the three experimentally observed conformers of the complex. They, mainly, differ by the hydration sites, of which the one at the Orn side chain is the most important for reshaping the peptide toward its native structure. The study demonstrates the ability of a quantum chemistry approach that intelligently combines the semiempirical and <i>ab initio</i> computations to disentangle a complex interplay of intra- and intermolecular hydrogen bonds in large molecular systems

Ion Selectivity of Crown Ethers Investigated by UV and IR Spectroscopy in a Cold Ion Trap

Electronic and vibrational spectra of benzo-15-crown-5 (B15C5) and benzo-18-crown-6 (B18C6) complexes with alkali metal ions, M⁺•B15C5 and M⁺•B18C6 (M = Li, Na, K, Rb, and Cs), are measured using UV photodissociation (UVPD) and IR–UV double resonance spectroscopy in a cold, 22-pole ion trap. We determine the structure of conformers with the aid of density functional theory calculations. In the Na⁺•B15C5 and K⁺•B18C6 complexes, the crown ethers open the most and hold the metal ions at the center of the ether ring, demonstrating an optimum matching in size between the cavity of the crown ethers and the metal ions. For smaller ions, the crown ethers deform the ether ring to decrease the distance and increase the interaction between the metal ions and oxygen atoms; the metal ions are completely surrounded by the ether ring. In the case of larger ions, the metal ions are too large to enter the crown cavity and are positioned on it, leaving one of its sides open for further solvation. Thermochemistry data calculated on the basis of the stable conformers of the complexes suggest that the ion selectivity of crown ethers is controlled primarily by the enthalpy change for the complex formation in solution, which depends strongly on the complex structure