Characterization of elusive rhamnosyl dioxanium ions and their application in complex oligosaccharide synthesis.

Attaining complete anomeric control is still one of the biggest challenges in carbohydrate chemistry. Glycosyl cations such as oxocarbenium and dioxanium ions are key intermediates of glycosylation reactions. Characterizing these highly-reactive intermediates and understanding their glycosylation mechanisms are essential to the stereoselective synthesis of complex carbohydrates. Although C-2 acyl neighbouring-group participation has been well-studied, the reactive intermediates in more remote participation remain elusive and are challenging to study. Herein, we report a workflow that is utilized to characterize rhamnosyl 1,3-bridged dioxanium ions derived from C-3 p-anisoyl esterified donors. First, we use a combination of quantum-chemical calculations and infrared ion spectroscopy to determine the structure of the cationic glycosylation intermediate in the gas-phase. In addition, we establish the structure and exchange kinetics of highly-reactive, low-abundance species in the solution-phase using chemical exchange saturation transfer, exchange spectroscopy, correlation spectroscopy, heteronuclear single-quantum correlation, and heteronuclear multiple-bond correlation nuclear magnetic resonance spectroscopy. Finally, we apply C-3 acyl neighbouring-group participation to the synthesis of complex bacterial oligosaccharides. This combined approach of finding answers to fundamental physical-chemical questions and their application in organic synthesis provides a robust basis for elucidating highly-reactive intermediates in glycosylation reactions.

A spectroscopic test suggests that fragment ion structure annotations in MS/MS libraries are frequently incorrect.

Modern untargeted mass spectrometry (MS) analyses quickly detect and resolve thousands of molecular compounds. Although features are readily annotated with a molecular formula in high-resolution small-molecule MS applications, the large majority of them remains unidentified in terms of their full molecular structure. Collision-induced dissociation tandem mass spectrometry (CID-MS2) provides a diagnostic molecular fingerprint to resolve the molecular structure through a library search. However, for de novo identifications, one must often rely on in silico generated MS2 spectra as reference. The ability of different in silico algorithms to correctly predict MS2 spectra and thus to retrieve correct molecular structures is a topic of lively debate, for instance in the CASMI contest. Underlying the predicted MS2 spectra are the in silico generated product ion structures, which are normally not used in de novo identification, but which can serve to critically assess the fragmentation algorithms. Here we evaluate in silico generated MSn product ion structures by comparison with structures established experimentally by infrared ion spectroscopy (IRIS). For a set of three dozen product ion structures from five precursor molecules, we find that virtually all fragment ion structure annotations in three major in silico MS2 libraries (HMDB, METLIN, mzCloud) are incorrect and caution the reader against their use for structure annotation of MS/MS ions.

Anomeric Triflates versus Dioxanium Ions: Different Product-Forming Intermediates from 3-Acyl Benzylidene Mannosyl and Glucosyl Donors.

Minimal structural differences in the structure of glycosyl donors can have a tremendous impact on their reactivity and the stereochemical outcome of their glycosylation reactions. Here, we used a combination of systematic glycosylation reactions, the characterization of potential reactive intermediates, and in-depth computational studies to study the disparate behavior of glycosylation systems involving benzylidene glucosyl and mannosyl donors. While these systems have been studied extensively, no satisfactory explanations are available for the differences observed between the 3-O-benzyl/benzoyl mannose and glucose donor systems. The potential energy surfaces of the different reaction pathways available for these donors provide an explanation for the contrasting behavior of seemingly very similar systems. Evidence has been provided for the intermediacy of benzylidene mannosyl 1,3-dioxanium ions, while the formation of the analogous 1,3-glucosyl dioxanium ions is thwarted by a prohibitively strong flagpole interaction of the C-2-O-benzyl group with the C-5 proton in moving toward the transition state, in which the glucose ring adopts a B2,5-conformation. This study provides an explanation for the intermediacy of 1,3-dioxanium ions in the mannosyl system and an answer to why these do not form from analogous glucosyl donors.

Identification of organic micro-pollutants in surface water using MS-based infrared ion spectroscopy.

Comprehensive monitoring of organic micro-pollutants (OMPs) in drinking water sources relies on non-target screening (NTS) using liquid-chromatography and high-resolution mass spectrometry (LC-HRMS). Identification of OMPs is typically based on accurate mass and tandem mass spectrometry (MS/MS) data by matching against entries in compound databases and MS/MS spectral libraries. MS/MS spectra are, however, not always diagnostic for the full molecular structure and, moreover, emerging OMPs or OMP transformation products may not be present in libraries. Here we demonstrate how infrared ion spectroscopy (IRIS), an emerging MS-based method for structural elucidation, can aid in the identification of OMPs. IRIS measures the IR spectrum of an m/z-isolated ion in a mass spectrometer, providing an orthogonal diagnostic for molecular identification. Here, we demonstrate the workflow for identification of OMPs in river water and show how quantum-chemically predicted IR spectra can be used to screen potential candidates and suggest structural assignments. A crucial step herein is to define a set of candidate structures, presumably including the actual OMP, for which we present several strategies based on domain knowledge, the IR spectrum and MS/MS spectrum.

Distinguishing Oligosaccharide Isomers Using Far-Infrared Ion Spectroscopy: Identification of Biomarkers for Inborn Errors of Metabolism.

Distinguishing isomeric saccharides poses a major challenge for analytical workflows based on (liquid chromatography) mass spectrometry (LC-MS). In recent years, many studies have proposed infrared ion spectroscopy as a possible solution as the orthogonal, spectroscopic characterization of mass-selected ions can often distinguish isomeric species that remain unresolved using conventional MS. However, the high conformational flexibility and extensive hydrogen bonding in saccharides cause their room-temperature fingerprint infrared spectra to have broad features that often lack diagnostic value. Here, we show that room-temperature infrared spectra of ion-complexed saccharides recorded in the previously unexplored far-infrared wavelength range (300-1000 cm-1) provide well-resolved and highly diagnostic features. We show that this enables distinction of isomeric saccharides that differ either by their composition of monosaccharide units and/or the orientation of their glycosidic linkages. We demonstrate the utility of this approach from single monosaccharides up to isomeric tetrasaccharides differing only by the configuration of a single glycosidic linkage. Furthermore, through hyphenation with hydrophilic interaction liquid chromatography, we identify oligosaccharide biomarkers in patient body fluid samples, demonstrating a generalized and highly sensitive MS-based method for the identification of saccharides found in complex sample matrices.

An In Silico Infrared Spectral Library of Molecular Ions for Metabolite Identification.

Infrared ion spectroscopy (IRIS) continues to see increasing use as an analytical tool for small-molecule identification in conjunction with mass spectrometry (MS). The IR spectrum of an m/z selected population of ions constitutes a unique fingerprint that is specific to the molecular structure. However, direct translation of an IR spectrum to a molecular structure remains challenging, as reference libraries of IR spectra of molecular ions largely do not exist. Quantum-chemically computed spectra can reliably be used as reference, but the challenge of selecting the candidate structures remains. Here, we introduce an in silico library of vibrational spectra of common MS adducts of over 4500 compounds found in the human metabolome database. In total, the library currently contains more than 75,000 spectra computed at the DFT level that can be queried with an experimental IR spectrum. Moreover, we introduce a database of 189 experimental IRIS spectra, which is employed to validate the automated spectral matching routines. This demonstrates that 75% of the metabolites in the experimental data set are correctly identified, based solely on their exact m/z and IRIS spectrum. Additionally, we demonstrate an approach for specifically identifying substructures by performing a search without m/z constraints to find structural analogues. Such an unsupervised search paves the way toward the de novo identification of unknowns that are absent in spectral libraries. We apply the in silico spectral library to identify an unknown in a plasma sample as 3-hydroxyhexanoic acid, highlighting the potential of the method.

Secondary prevention in diabetic and nondiabetic coronary heart disease patients: Insights from the German subset of the hospital arm of the EUROASPIRE IV and V surveys.

BACKGROUND: Patients with coronary heart disease (CHD) with and without diabetes mellitus have an increased risk of recurrent events requiring multifactorial secondary prevention of cardiovascular risk factors. We compared prevalences of cardiovascular risk factors and its determinants including lifestyle, pharmacotherapy and diabetes mellitus among patients with chronic CHD examined within the fourth and fifth EUROASPIRE surveys (EA-IV, 2012-13; and EA-V, 2016-17) in Germany. METHODS: The EA initiative iteratively conducts European-wide multicenter surveys investigating the quality of secondary prevention in chronic CHD patients aged 18 to 79 years. The data collection in Germany was performed during a comprehensive baseline visit at study centers in Würzburg (EA-IV, EA-V), Halle (EA-V), and Tübingen (EA-V). RESULTS: 384 EA-V participants (median age 69.0 years, 81.3% male) and 536 EA-IV participants (median age 68.7 years, 82.3% male) were examined. Comparing EA-IV and EA-V, no relevant differences in risk factor prevalence and lifestyle changes were observed with the exception of lower LDL cholesterol levels in EA-V. Prevalence of unrecognized diabetes was significantly lower in EA-V as compared to EA-IV (11.8% vs. 19.6%) while the proportion of prediabetes was similarly high in the remaining population (62.1% vs. 61.0%). CONCLUSION: Between 2012 and 2017, a modest decrease in LDL cholesterol levels was observed, while no differences in blood pressure control and body weight were apparent in chronic CHD patients in Germany. Although the prevalence of unrecognized diabetes decreased in the later study period, the proportion of normoglycemic patients was low. As pharmacotherapy appeared fairly well implemented, stronger efforts towards lifestyle interventions, mental health programs and cardiac rehabilitation might help to improve risk factor profiles in chronic CHD patients.

Environmentally weathered polystyrene particles induce phenotypical and functional maturation of human monocyte-derived dendritic cells.

Micro- and nanoplastics (MNP) are ubiquitously present in the environment due to their high persistence and bioaccumulative properties. Humans get exposed to MNP via various routes and consequently, they will encounter dendritic cells (DC) which are antigen-presenting cells involved in regulating immune responses. The consequences of DC exposure to MNP are an important, yet understudied, cause of concern. Therefore, this study aimed to assess the uptake and effect of MNP in vitro by exposing human monocyte-derived dendritic cells (MoDC) to virgin and environmentally weathered polystyrene (PS) particles of different sizes (0.2, 1, and 10 µm), at different concentrations ranging from 1 to 100 µg/ml. The effects of these particles were examined by measuring co-stimulatory surface marker (i.e. CD83 and CD86) expression. In addition, T-cell proliferation was measured via a mixed-leukocyte reaction (MLR) assay. The results showed that MoDC were capable of absorbing PS particles, and this was facilitated by pre-incubation in heat-inactivated (HI) plasma. Furthermore, depending on their size, weathered PS particles in particular caused increased expression of CD83 and CD86 on MoDC. Lastly, weathered 0.2 µm PS particles were able to functionally activate MoDC, leading to an increase in T-cell activation. These in vitro data suggest that, depending on their size, weathered PS particles might act as an immunostimulating adjuvant, possibly leading to T-cell sensitization.

Competing C-4 and C-5-Acyl Stabilization of Uronic Acid Glycosyl Cations.

Uronic acids are carbohydrates carrying a terminal carboxylic acid and have a unique reactivity in stereoselective glycosylation reactions. Herein, the competing intramolecular stabilization of uronic acid cations by the C-5 carboxylic acid or the C-4 acetyl group was studied with infrared ion spectroscopy (IRIS). IRIS reveals that a mixture of bridged ions is formed, in which the mixture is driven towards the C-1,C-5 dioxolanium ion when the C-5,C-2-relationship is cis, and towards the formation of the C-1,C-4 dioxepanium ion when this relation is trans. Isomer-population analysis and interconversion barrier computations show that the two bridged structures are not in dynamic equilibrium and that their ratio parallels the density functional theory computed stability of the structures. These studies reveal how the intrinsic interplay of the different functional groups influences the formation of the different regioisomeric products.

Ab Initio Multiplet-Plus-Cumulant Approach for Correlation Effects in X-Ray Photoelectron Spectroscopy.

The treatment of electronic correlations in open-shell systems is among the most challenging problems of condensed matter theory. Current approximations are only partly successful. Ligand-field multiplet theory has been widely successful in describing intra-atomic correlation effects in x-ray spectra, but typically ignores itinerant states. The cumulant expansion for the one-electron Green's function has been successful in describing shake-up effects but ignores atomic multiplets. More complete methods, such as dynamic mean-field theory can be computationally demanding. Here, we show that separating the dynamic Coulomb interactions into local and longer-range parts with ab initio parameters yields a combined multiplet-plus-cumulant approach that accounts for both local atomic multiplets and satellite excitations. The approach is illustrated in transition metal oxides and explains the multiplet peaks, charge-transfer satellites, and distributed background features observed in XPS experiment.

Stabilization of Glucosyl Dioxolenium Ions by "Dual Participation" of the 2,2-Dimethyl-2-(ortho-nitrophenyl)acetyl (DMNPA) Protection Group for 1,2-cis-Glucosylation.

The stereoselective introduction of glycosidic bonds is of paramount importance to oligosaccharide synthesis. Among the various chemical strategies to steer stereoselectivity, participation by either neighboring or distal acyl groups is used particularly often. Recently, the use of the 2,2-dimethyl-2-(ortho-nitrophenyl)acetyl (DMNPA) protection group was shown to offer enhanced stereoselective steering compared to other acyl groups. Here, we investigate the origin of the stereoselectivity induced by the DMNPA group through systematic glycosylation reactions and infrared ion spectroscopy (IRIS) combined with techniques such as isotopic labeling of the anomeric center and isomer population analysis. Our study indicates that the origin of the DMNPA stereoselectivity does not lie in the direct participation of the nitro moiety but in the formation of a dioxolenium ion that is strongly stabilized by the nitro group.

Characterization of Elusive Reaction Intermediates Using Infrared Ion Spectroscopy: Application to the Experimental Characterization of Glycosyl Cations.

A detailed understanding of the reaction mechanism(s) leading to stereoselective product formation is crucial to understanding and predicting product formation and driving the development of new synthetic methodology. One way to improve our understanding of reaction mechanisms is to characterize the reaction intermediates involved in product formation. Because these intermediates are reactive, they are often unstable and therefore difficult to characterize using experimental techniques. For example, glycosylation reactions are critical steps in the chemical synthesis of oligosaccharides and need to be stereoselective to provide the desired α- or ß-diastereomer. It remains challenging to predict and control the stereochemical outcome of glycosylation reactions, and their reaction mechanisms remain a hotly debated topic. In most cases, glycosylation reactions take place via reaction mechanisms in the continuum between SN1- and SN2-like pathways. SN2-like pathways proceeding via the displacement of a contact ion pair are relatively well understood because the reaction intermediates involved can be characterized by low-temperature NMR spectroscopy. In contrast, the SN1-like pathways proceeding via the solvent-separated ion pair, also known as the glycosyl cation, are poorly understood. SN1-like pathways are more challenging to investigate because the glycosyl cation intermediates involved are highly reactive. The highly reactive nature of glycosyl cations complicates their characterization because they have a short lifetime and rapidly equilibrate with the corresponding contact ion pair. To overcome this hurdle and enable the study of glycosyl cation stability and structure, they can be generated in a mass spectrometer in the absence of a solvent and counterion in the gas phase. The ease of formation, stability, and fragmentation of glycosyl cations have been studied using mass spectrometry (MS). However, MS alone provides little information about the structure of glycosyl cations. By combining mass spectrometry (MS) with infrared ion spectroscopy (IRIS), the determination of the gas-phase structures of glycosyl cations has been achieved. IRIS enables the recording of gas-phase infrared spectra of glycosyl cations, which can be assigned by matching to reference spectra predicted from quantum chemically calculated vibrational spectra. Here, we review the experimental setups that enable IRIS of glycosyl cations and discuss the various glycosyl cations that have been characterized to date. The structure of glycosyl cations depends on the relative configuration and structure of the monosaccharide substituents, which can influence the structure through both steric and electronic effects. The scope and relevance of gas-phase glycosyl cation structures in relation to their corresponding condensed-phase structures are also discussed. We expect that the workflow reviewed here to study glycosyl cation structure and reactivity can be extended to many other reaction types involving difficult-to-characterize ionic intermediates.

Real-Time Equation-of-Motion CCSD Cumulant Green's Function.

Many-body excitations in X-ray photoemission spectra have been difficult to simulate from first principles. We have recently developed a cumulant-based one-electron Green's function method using the real-time coupled-cluster-singles equation-of-motion approach (RT-EOM-CCS) that provides a general framework for treating these problems. Here we extend this approach to include double excitations in the ground-state energy and in the coupled cluster amplitudes, which have been implemented using subroutines generated by the Tensor Contraction Engine (TCE). As in the case of the singles approximation, RT-EOM-CCSD yields a nonperturbative cumulant form of the Green's function in terms of the time-dependent cluster amplitudes, adding nonlinear corrections to the traditional cumulant forms. The extended approach is applied to the core-hole spectral function for small molecular systems. We find that, when core-optimized basis sets are used, the doubles contributions reduce the mean absolute errors in the core binding energies of the 10e systems from 0.8 to 0.3 eV. They also significantly improve the quasiparticle-satellite gap by reducing its overestimation from about 3-5 to about 0-1 eV in CH4, NH3, and H2O, and also improving the overall shape of the satellite features. Finally, we demonstrate the application of the new implementation to the larger, classical XPS ESCA series of molecules and show that the singles approximation can be paired with a modest basis set to study carbon speciation.

Characterization of Cyclic N-Acyliminium Ions by Infrared Ion Spectroscopy.

N-Acyliminium ions are highly reactive intermediates that are important for creating CC-bonds adjacent to nitrogen atoms. Here we report the characterization of cyclic N-acyliminium ions in the gas phase, generated by collision induced dissociation tandem mass spectrometry followed by infrared ion spectroscopy using the FELIX infrared free electron laser. Comparison of DFT calculated spectra with the experimentally observed IR spectra provided valuable insights in the conformations of the N-acyliminium ions.

Advanced calculations of X-ray spectroscopies with FEFF10 and Corvus.

The real-space Green's function code FEFF has been extensively developed and used for calculations of X-ray and related spectra, including X-ray absorption (XAS), X-ray emission (XES), inelastic X-ray scattering, and electron energy-loss spectra. The code is particularly useful for the analysis and interpretation of the XAS fine-structure (EXAFS) and the near-edge structure (XANES) in materials throughout the periodic table. Nevertheless, many applications, such as non-equilibrium systems, and the analysis of ultra-fast pump-probe experiments, require extensions of the code including finite-temperature and auxiliary calculations of structure and vibrational properties. To enable these extensions, we have developed in tandem a new version FEFF10 and new FEFF-based workflows for the Corvus workflow manager, which allow users to easily augment the capabilities of FEFF10 via auxiliary codes. This coupling facilitates simplified input and automated calculations of spectra based on advanced theoretical techniques. The approach is illustrated with examples of high-temperature behavior, vibrational properties, many-body excitations in XAS, super-heavy materials, and fits of calculated spectra to experiment.

Equation-of-Motion Coupled-Cluster Cumulant Green's Function for Excited States and X-Ray Spectra.

Green's function methods provide a robust, general framework within many-body theory for treating electron correlation in both excited states and x-ray spectra. Conventional methods using the Dyson equation or the cumulant expansion are typically based on the GW self-energy approximation. In order to extend this approximation in molecular systems, a non-perturbative real-time coupled-cluster cumulant Green's function approach has been introduced, where the cumulant is obtained as the solution to a system of coupled first order, non-linear differential equations. This approach naturally includes non-linear corrections to conventional cumulant Green's function techniques where the cumulant is linear in the GW self-energy. The method yields the spectral function for the core Green's function, which is directly related to the x-ray photoemission spectra (XPS) of molecular systems. The approach also yields very good results for binding energies and satellite excitations. The x-ray absorption spectrum (XAS) is then calculated using a convolution of the core spectral function and an effective, one-body XAS. Here this approach is extended to include the full coupled-cluster-singles (CCS) core Green's function by including the complete form of the non-linear contributions to the cumulant as well as all single, double, and triple cluster excitations in the CC amplitude equations. This approach naturally builds in orthogonality and shake-up effects analogous to those in the Mahan-Noizeres-de Dominicis edge singularity corrections that enhance the XAS near the edge. The method is illustrated for the XPS and XAS of NH3.

IRMPD Spectroscopy of Homo- and Heterochiral Asparagine Proton-Bound Dimers in the Gas Phase.

We investigate gas-phase structures of homo- and heterochiral asparagine proton-bound dimers with infrared multiphoton dissociation (IRMPD) spectroscopy and quantum-chemical calculations. Their IRMPD spectra are recorded at room temperature in the range of 500-1875 and 3000-3600 cm-1. Both varieties of asparagine dimers are found to be charge-solvated based on their IRMPD spectra. The location of the principal intramolecular H-bond is discussed in light of harmonic frequency analyses using the B3LYP functional with GD3BJ empirical dispersion. Contrary to theoretical analyses, the two spectra are very similar.

Unraveling intrinsic correlation effects with angle-resolved photoemission spectroscopy.

Interaction effects can change materials properties in intriguing ways, and they have, in general, a huge impact on electronic spectra. In particular, satellites in photoemission spectra are pure many-body effects, and their study is of increasing interest in both experiment and theory. However, the intrinsic spectral function is only a part of a measured spectrum, and it is notoriously difficult to extract this information, even for simple metals. Our joint experimental and theoretical study of the prototypical simple metal aluminum demonstrates how intrinsic satellite spectra can be extracted from measured data using angular resolution in photoemission. A nondispersing satellite is detected and explained by electron-electron interactions and the thermal motion of the atoms. Additional nondispersing intensity comes from the inelastic scattering of the outgoing photoelectron. The ideal intrinsic spectral function, instead, has satellites that disperse both in energy and in shape. Theory and the information extracted from experiment describe these features with very good agreement.

Real-Time Coupled-Cluster Approach for the Cumulant Green's Function.

Green's function methods within many-body perturbation theory provide a general framework for treating electronic correlations in excited states and spectra. Here, we develop the cumulant form of the one-electron Green's function using a real-time coupled-cluster equation-of-motion approach, in an extension of our previous study (Rehr J.; et al. J. Chem. Phys. 2020, 152, 174113). The approach yields a nonperturbative expression for the cumulant in terms of the solution to a set of coupled first-order, nonlinear differential equations. The method thereby adds nonlinear corrections to traditional cumulant methods, which are linear in the self-energy. The approach is applied to the core-hole Green's function and is illustrated for a number of small molecular systems. For these systems, we find that the nonlinear contributions yield significant improvements, both for quasiparticle properties such as core-level binding energies and for inelastic losses that correspond to satellites observed in photoemission spectra.

Characterization of holmium(iii)-acetylacetonate complexes derived from therapeutic microspheres by infrared ion spectroscopy.

Microspheres containing radioactive 166holmium-acetylacetonate are employed in emerging radionuclide therapies for the treatment of malignancies. At the molecular level, details on the coordination geometries of the Ho complexes are however elusive. Infrared ion spectroscopy (IRIS) was used to characterize several 165Ho-acetylacetonate complexes derived from non-radioactive microspheres. The coordination geometry of four distinct ionic complexes were fully assigned by comparison of their measured IR spectra with spectra calculated at the density functional theory (DFT) level. The coordination of each acetylacetonate ligand is dependent on the presence of other ligands, revealing an asymmetric chelation motif in some of the complexes. A fifth, previously unknown constituent of the microspheres was identified as a coordination complex containing an acetic acid ligand. These results pave the way for IRIS-based identification of microsphere constituents upon neutron activation of the metal center.