Spectral diffusion and 14N quadrupole splittings in absorption detected magnetic resonance hole burning spectra of photosynthetic reaction centers

Zero field absorption detected magnetic resonance hole burning measurements were performed on photosynthetic reaction centers of the bacteria Rhodobacter sphaeroides R26 and Rhodopseudomonas viridis. Extrapolation to zero microwave power yielded pseudohomogeneous linewidths of 2.0 MHz for Rhodopseudomonas viridis, 1.0 and 0.9 MHz for the protonated forms of Rhodobacter sphaeroides R26 with and without monomer bacteriochlorophyll exchanged, and 0.25 MHz as an upper limit for fully deuterated reaction centers of Rhodobacter sphaeroides R26. The measured linewidths were interpreted as being due to unresolved hyperfine interaction between the nuclear spins and the triplet electron spin, the line shape being determined by spectral diffusion among the nuclei. The difference in linewidths between Rhodobacter sphaeroides R26 and Rhodopseudomonas viridis is then explained by triplet delocalization on the special pair in the former, and localization on one dimer half on the latter. In the fully deuterated sample, four quadrupole satellites were observed in the hole spectra arising from the eight 14N nitrogens in the special pair. The quadrupole parameters seem to be very similar for all nitrogens and were determined to =1.25±0.1 MHz and =0.9±0.1 MHz. The Journal of Chemical Physics is copyrighted by The American Institute of Physics

Cryogenic infrared spectroscopy provides mechanistic insight into the fragmentation of phospholipid silver adducts

Tandem mass spectrometry is arguably the most important analytical tool for structure elucidation of lipids and other metabolites. By fragmenting intact lipid ions, valuable structural information such as the lipid class and fatty acyl composition are readily obtainable. The information content of a fragment spectrum can often be increased by the addition of metal cations. In particular, the use of silver ions is deeply rooted in the history of lipidomics due to their propensity to coordinate both electron-rich heteroatoms and C = C bonds in aliphatic chains. Not surprisingly, coordination of silver ions was found to enable the distinction of sn-isomers in glycerolipids by inducing reproducible intensity differences in the fragment spectra, which could, however, not be rationalized. Here, we investigate the fragmentation behaviors of silver-adducted sn- and double bond glycerophospholipid isomers by probing fragment structures using cryogenic gas-phase infrared (IR) spectroscopy. Our results confirm that neutral headgroup loss from silver-adducted glycerophospholipids leads to dioxolane-type fragments generated by intramolecular cyclization. By combining high-resolution IR spectroscopy and computational modelling of silver-adducted fragments, we offer qualitative explanations for different fragmentation behaviors of glycerophospholipid isomers. Overall, the results demonstrate that gas-phase IR spectroscopy of fragment ions can significantly contribute to our understanding of lipid dissociation mechanisms and the influence of coordinating cations

Unveiling Glycerolipid Fragmentation by Cryogenic Infrared Spectroscopy

Mass spectrometry is routinely employed for structure elucidation of molecules. Structural information can be retrieved from intact molecular ions by fragmentation; however, the interpretation of fragment spectra is often hampered by poor understanding of the underlying dissociation mechanisms. For example, neutral headgroup loss from protonated glycerolipids has been postulated to proceed via an intramolecular ring closure but the mechanism and resulting ring size have never been experimentally confirmed. Here we use cryogenic gas-phase infrared (IR) spectroscopy in combination with computational chemistry to unravel the structures of fragment ions and thereby shed light on elusive dissociation mechanisms. Using the example of glycerolipid fragmentation, we study the formation of protonated five-membered dioxolane and six-membered dioxane rings and show that dioxolane rings are predominant throughout different glycerolipid classes and fragmentation channels. For comparison, pure dioxolane and dioxane ions were generated from tailor-made dehydroxyl derivatives inspired by natural 1,2- and 1,3-diacylglycerols and subsequently interrogated using IR spectroscopy. Furthermore, the cyclic structure of an intermediate fragment occurring in the phosphatidylcholine fragmentation pathway was spectroscopically confirmed. Overall, the results contribute substantially to the understanding of glycerolipid fragmentation and showcase the value of vibrational ion spectroscopy to mechanistically elucidate crucial fragmentation pathways in lipidomics

Structure and Conformation Determine Gas-Phase Infrared Spectra of Detergents

Native mass spectrometry of membrane proteins relies on non-ionic detergents which protect the protein during transfer from solution into the gas phase. Once in the gas phase, the detergent micelle must be efficiently removed, which is usually achieved by collision-induced dissociation (CID). Recently, infrared multiple photon dissociation (IRMPD) has emerged as an alternative activation method for the analysis of membrane proteins, which has led to a growing interest in detergents that efficiently absorb infrared light. Here we investigate whether the absorption properties of synthetic detergents can be tailored by merging structural motifs of existing detergents into new hybrid detergents. We combine gas-phase infrared ion spectroscopy with density functional theory to investigate and rationalize the absorption properties of three established detergents and two hybrid detergents with fused headgroups. We show that, although the basic intramolecular interactions in the parent and hybrid detergents are similar, the three-dimensional structures differ significantly and so do the infrared spectra. Our results outline a roadmap for guiding the synthesis of tailored detergents with computational chemistry for future mass spectrometry applications

Characterization and Fate of a Septanosyl Ferrier Cation in the Gas and Solution Phases

Ferrier reactions follow a mechanistic pathway whereby Lewis acid activation of a cyclic enol ether facilitates departure of an allylic leaving group to form a glycosyl Ferrier cation. Attack on the Ferrier cation provides a new acetal linkage concurrent with the transposition of the alkene moiety. The idiosyncratic outcomes of Ferrier reactions of seven-membered ring carbohydrate-based oxepines prompted an investigation of its corresponding septanosyl Ferrier cation. Experiments that characterized the ion, including gas-phase cryogenic IR spectroscopy matched with density functional theory-calculated spectra of candidate cation structures, as well as product analysis from solution-phase Ferrier reactions, are reported here. Results from both approaches revealed an inclination of the seven-membered ring cation to contract to five-membered ring structures. Gas-phase IR spectra matched best to calculated spectra of structures in which five-membered dioxolenium formation opened the oxepine ring. In the solution phase, an attack on the ion by water led to an acyclic enal that cyclized to a C-methylene-aldehydo arabinofuranoside species. Attack by allyl trimethylsilane, on the other hand, was diastereoselective and yielded a C-allyl septanoside

Cryogenic infrared spectroscopy reveals remarkably short NH⁺⋯F hydrogen bonds in fluorinated phenylalanines

In past decades, hydrogen bonds involving organic fluorine have been a highly disputed topic. Obtaining clear evidence for the presence of fluorine-specific interactions is generally difficult because of their weak nature. Today, the existence of hydrogen bonds with organic fluorine is widely accepted and supported by numerous studies. However, strong bonds with short H⋯F distances remain scarce and are primarily found in designed model compounds. Using a combination of cryogenic gas-phase infrared spectroscopy and density functional theory, we here analyze a series of conformationally unrestrained fluorinated phenylalanine compounds as protonated species. The results suggest proximal NH+⋯F hydrogen bonds with an exceptionally close H⋯F distance (1.79 Å) in protonated ortho-fluorophenylalanine

Studying the Key Intermediate of RNA Autohydrolysis by Cryogenic Gas-Phase Infrared Spectroscopy

Over the course of the COVID-19 pandemic, mRNA-basedvaccineshave gained tremendous importance. The development and analysis of modified RNA moleculesbenefit from advanced mass spectrometry and require sufficient understanding of fragmentation processes.Analogous tothe degradation of RNA in solution by autohydrolysis,backbone cleavage of RNA strands wasequally observedin the gas phase; however, the fragmentation mechanism remained elusive.In this work,autohydrolysis-like intermediates weregenerated from isolated RNA dinucleotidesin the gas phaseand investigatedusing cryogenic infrared spectroscopy in helium nanodroplets.Data from both experiment and density functional theory provide evidence forthe formation of a five-membered cyclic phosphateintermediateand rule outlinear orsix-membered structures. Furthermore, the experiments show that another prominent condensed-phase reactionof RNA nucleotides can be induced in the gas phase: the tautomerization of cytosine.Both observed reactions aretherefore highlyuniversal and intrinsic properties of the investigated molecules

Studying the Intrinsic Reactivity of Chromanes by Gas-Phase Infrared Spectroscopy

Tandem mass spectrometry is routinely used for the structural analysis of organic molecules, but many fragmentation reactions are not well understood. Because several potential structures can correspond to a measured mass, the assignment of product ions is ambiguous using mass spectrometry alone. Here, we combine mass spectrometry with high-resolution gas-phase infrared spectroscopy and computational chemistry tools to identify product ion structures and derive collision-induced fragmentation mechanisms of the chromane derivatives Trolox and Methyltrolox. We find that protonated Trolox and Methyltrolox fragment identically via dehydration and decarbonylation, while deprotonated ions display substantially diverging reactivities. For deprotonated Methyltrolox, we observe unusual radical fragmentation reactions and suggest a [1,2]-Wittig rearrangement involving aryl migration in the gas phase. Overall, the combined experimental and theoretical approach presented here revealed complex proton dynamics and intramolecular rearrangement reactions, which expand our understanding on structure–reactivity relationships of isolated molecules in different protonation states

Untersuchung des reaktiven Intermediats der RNA Autohydrolyse mittels kryogener Infrarotspektroskopie in der Gasphase

Im Laufe der COVID-19 Pandemie haben mRNA-basierte Impfstoffe an immenser Bedeutung gewonnen. Massenspektrometrie ist für die Entwicklung und Analyse von modifizierten RNA Molekülen unerlässlich, setzt jedoch ein grundlegendes Verständnis über Fragmentierungsprozesse voraus. Analog zu der Zersetzung von RNA in Lösung durch Autohydrolyse, kann die Spaltung des RNA Rückgrats ebenso in der Gasphase stattfinden. Bislang sind die Fragmentierungsmechanismen jedoch unzureichend untersucht. In dieser Arbeit wurden Intermediate aus isolierten RNA Dinukleotiden in der Gasphase generiert und mittels kryogener Infrarotspektroskopie in Helium-Nanotröpfchen untersucht. Die experimentellen Daten, unterstützt durch Dichtefunktionaltheorie, liefern Hinweise dafür, dass die Bildung eines fünfgliedrigen zyklischen Phosphat-Intermediats begünstigt ist, während lineare oder sechsgliedrige Strukturen ausgeschlossen werden können. Weiterhin zeigen die Experimente, dass eine zusätzliche, bekannte Reaktion von RNA Nukleotiden in Lösung auch in der Gasphase induziert werden kann: die Tautomerisierung von Cytosin. Die beiden beobachteten Reaktionen spiegeln daher universelle und intrinsische Eigenschaften der untersuchten Moleküle wider

Direct Experimental Characterization of a Sialyl Cation

Sialic acids are monosaccharide residues involved in several biological processes. Controlling the stereoselectivity of sialylation reactions is challenging and mechanistic studies on the structure of its intermediate, the sialyl cation, are scarce. Here it is shown that a sialyl cation can be generated and isolated from an ionized sialic acid precursor. This short-lived species is structurally characterized for the first time using cryogenic infrared spectroscopy. In combination with quantum chemical calculations, the results reveal that the positive charge at the anomeric carbon of the sialyl cation is stabilized by remote participation of the C5-NHAc group leading to the formation of a bridged structure. In this structure, the β-side is shielded from nucleophilic attack, potentially explaining the α-selectivity of this building block in SN1-type sialylation reactions. Other modes of participation are energetically unfavored and cannot be observed experimentally