Application Of Nuclear Magnetic Resonance Spectroscopy Research Articles

NMR and Metabolomics-A Roadmap for the Future.

Metabolomics investigates global metabolic alterations associated with chemical, biological, physiological, or pathological processes. These metabolic changes are measured with various analytical platforms including liquid chromatography-mass spectrometry (LC-MS), gas chromatography-mass spectrometry (GC-MS) and nuclear magnetic resonance spectroscopy (NMR). While LC-MS methods are becoming increasingly popular in the field of metabolomics (accounting for more than 70% of published metabolomics studies to date), there are considerable benefits and advantages to NMR-based methods for metabolomic studies. In fact, according to PubMed, more than 926 papers on NMR-based metabolomics were published in 2021—the most ever published in a given year. This suggests that NMR-based metabolomics continues to grow and has plenty to offer to the scientific community. This perspective outlines the growing applications of NMR in metabolomics, highlights several recent advances in NMR technologies for metabolomics, and provides a roadmap for future advancements.

Conformational and Structural Characterization of Carbohydrates and their Interactions Studied by NMR.

Carbohydrates, either free or as glycans conjugated with other biomolecules, participate in a plethora of essential biological processes. Their apparent simplicity in terms of chemical functionality hides an extraordinary diversity and structural complexity. Deeply deciphering at the atomic level their structures is essential to understand their biological function and activities, but it is still a challenging task in need of complementary approaches and no generalized procedures are available to address the study of such complex, natural glycans. The versatility of Nuclear Magnetic Resonance spectroscopy (NMR) often makes it the preferred choice to study glycans and carbohydrates in solution media. The most basic NMR parameters, namely chemical shifts, coupling constants, and nuclear Overhauser effects, allow defining short or repetitive chain sequences and characterize their structures and local geometries either in the free state or when interacting with other biomolecules, rendering additional information on the molecular recognition processes. The increased accessibility to carbohydrate molecules extensively or selectively labeled with 13C is boosting the resolution and detail which analyzed glycan structures can reach. In turn, structural information derived from NMR complemented with molecular modeling and theoretical calculations can also provide dynamic information on the conformational flexibility of carbohydrate structures. Furthermore, using partially oriented media or paramagnetic perturbations, it has been possible to introduce additional longrange observables rendering structural information on longer and branched glycan chains. In this review, we provide examples of these studies and an overview of the recent and most relevant NMR applications in the glycobiology field.

Intrinsic anisotropy parameters of a series of lanthanoid complexes deliver new insights into the structure-magnetism relationship

Intrinsic anisotropy parameters of a series of lanthanoid complexes deliver new insights into the structure-magnetism relationship

Application of NMR and Chemometrics for the Profiling and Classification of Ale and Lager American Craft Beer.

In this paper, Nuclear Magnetic Resonance spectroscopy (NMR)-based metabolomics were applied for the discrimination of ale and lager craft American beers. A modified pulse sequence that allows the efficient suppression of the water and ethanol peaks was used to achieve high-quality spectra with minimal sample preparation. The initial chemometrics analysis generated models of low predictive power, indicating the high variability in the groups. Due to this variability, we tested the effect of various data pretreatment and chemometrics approaches to improve the model’s performance. Spectral alignment was found to improve the classification significantly, while the type of normalization also played an important role. NMR combined with statistical and machine-learning techniques such as orthogonal projection to latent structures discriminant analysis (OPLS-DA) and random forest was able to discriminate between ale and lager beers, thus providing an important tool for the quality control and analysis of these products.

Versatile modules enable automated multi-column purifications on the ÄKTA pure chromatography system

Protein purification processes in basic research using ÄKTA™ liquid chromatography systems are often limited to single sample injections and simple one-column purifications. Because many target proteins in structural biology require complex purification protocols the work easily becomes laborious. To streamline and accelerate downstream protein production, an ALIAS™ autosampler and a modular sample in-line dilution process coupled to ion-exchange chromatography were incorporated into the workflow to automate two of the most commonly performed purification strategies - ion-exchange to size exclusion and nickel-ion metal affinity to size exclusion. The chromatographic setup enabled purification of a large array of cytosolic and membrane proteins from small-scale expression cultures produced in insect cells necessary to develop and optimize isotope-labeling strategies for nuclear magnetic resonance spectroscopy applications, resulting in a reduction in experiment time of about 20% per run for both cytosolic and membrane protein purification schemes. However, when queuing multiple samples the throughput increased by 66% and 75%, respectively. In addition, a novel system configuration is presented, where two column valves can be operated independently. This allows for the design of purification loops to increase purity of the target protein.

Determinants of Ligand Subtype-Selectivity at α1A-Adrenoceptor Revealed Using Saturation Transfer Difference (STD) NMR.

α1A- and α1B-adrenoceptors (α1A-AR and α1B-AR) are closely related G protein-coupled receptors (GPCRs) that modulate the cardiovascular and nervous systems in response to binding epinephrine and norepinephrine. The GPCR gene superfamily is made up of numerous subfamilies that, like α1A-AR and α1B-AR, are activated by the same endogenous agonists but may modulate different physiological processes. A major challenge in GPCR research and drug discovery is determining how compounds interact with receptors at the molecular level, especially to assist in the optimization of drug leads. Nuclear magnetic resonance spectroscopy (NMR) can provide great insight into ligand-binding epitopes, modes, and kinetics. Ideally, ligand-based NMR methods require purified, well-behaved protein samples. The instability of GPCRs upon purification in detergents, however, makes the application of NMR to study ligand binding challenging. Here, stabilized α1A-AR and α1B-AR variants were engineered using Cellular High-throughput Encapsulation, Solubilization, and Screening (CHESS), allowing the analysis of ligand binding with Saturation Transfer Difference NMR (STD NMR). STD NMR was used to map the binding epitopes of epinephrine and A-61603 to both receptors, revealing the molecular determinants for the selectivity of A-61603 for α1A-AR over α1B-AR. The use of stabilized GPCRs for ligand-observed NMR experiments will lead to a deeper understanding of binding processes and assist structure-based drug design.

Direct detection of carbon and nitrogen nuclei for high-resolution analysis of intrinsically disordered proteins using NMR spectroscopy.

Nuclear magnetic resonance spectroscopy (NMR) is a powerful technique for characterizing the structural and dynamic properties of intrinsically disordered proteins and protein regions (IDPs & IDRs). However, the application of NMR to IDPs has been limited by poor chemical shift dispersion in two-dimensional (2D) 1H-15N heteronuclear correlation spectra. Among the various detection schemes available for heteronuclear correlation spectroscopy, 13C direct-detection has become a mainstay for investigations of IDPs owing to the favorable chemical shift dispersion in 2D 13C'-15N correlation spectra. Recent advances in cryoprobe technology have enhanced the sensitivity for direct detection of both 13C and 15N resonances at high magnetic field strengths, thus prompting the development of 15N direct-detect experiments to complement established 13C-detection experiments. However, the application of 15N-detection has not been widely explored for IDPs. Here we compare 1H, 13C, and 15N detection schemes for a variety of 2D heteronuclear correlation spectra and evaluate their performance on the basis of resolution, chemical shift dispersion, and sensitivity. We performed experiments with a variety of disordered systems ranging in size and complexity; from a small IDR (99 amino acids), to a large low complexity IDR (185 amino acids), and finally a ∼73 kDa folded homopentameric protein that also contains disordered regions (133 amino acids/monomer). We conclude that, while requiring high sample concentration and long acquisition times, 15N-detection often offers enhanced resolution over other detection schemes in studies of disordered protein regions with low complexity sequences.

Nuclear Magnetic Resonance Spectroscopy Applications In Foods

Nuclear magnetic resonance spectroscopy (NMR) is the most powerful technique for determining the structure of organic compounds. NMR techniques are used successfully in various food systems for quality control and research. NMR spectroscopy is used to determine structure of proteins, aminoacid profile, carotenoids, organic acids, lipid fractions, the mobility of the water in foods. NMR spectroscopy is also used to identify and quantify the metabolites in foods. Also vegetable oils, fish oils, fish and meat, milk, cheese, wheat, fruit juices, coffee, green tea, foods such as wine and beer are among the last NMR applications. In addition, NMR spectroscopy is utilized for foodomics which is a new discipline that brings food science and nutritional research together. NMR techniques used for the food authentication are one- and two-dimensional NMR techniques, high resolution liquid state 1H and 13C NMR techniques, N15 and P-31 NMR techniques, 1H HR/MAS (high resolution magic angle spinning) NMR techniques. At this study, usage purposes of nuclear magnetic resonance spectroscopy for foods were collected.

Live-cell high resolution magic angle spinning magnetic resonance spectroscopy for in vivo analysis of Pseudomonas aeruginosa metabolomics

Pseudomonas aeruginosa (PA) is a pathogenic gram-negative bacterium that is widespread in nature, inhabiting soil, water, plants and animals. PA is a prevalent cause of deleterious human infections, particularly in patients whose host defense mechanisms have been compromised. Metabolomics is an important tool used to study host-pathogen interactions and to identify novel therapeutic targets and corresponding compounds. The aim of the present study was to report the metabolic profile of live PA bacteria using in vivo high-resolution magic angle spinning (HRMAS) nuclear magnetic resonance spectroscopy (NMR), in combination with 1- and 2-dimensional HRMAS NMR. This methodology provides a new and powerful technique to rapidly interrogate the metabolome of intact bacterial cells and has several advantages over traditional techniques that identify metabolome components from disrupted cells. Furthermore, application of multidimensional HRMAS NMR, in combination with the novel technique total through-Bond correlation Spectroscopy (TOBSY), is a promising approach that may be used to obtain in vivo metabolomics information from intact live bacterial cells and can mediate such analyses in a short period of time. Moreover, HRMAS 1H NMR enables the investigation of the associations between metabolites and cell processes. In the present study, we detected and quantified several informative metabolic molecules in live PA cells, including N-acetyl, betaine, citrulline, alanine and glycine, which are important in peptidoglycan synthesis. The results provided a complete metabolic profile of PA for future studies of PA clinical isolates and mutants. In addition, this in vivo NMR biomedical approach might have clinical utility and should prove useful in gene function validation, the study of pathogenetic mechanisms, the classification of microbial strains into functional/clinical groups, the testing of anti-bacterial agents and the determination of metabolic profiles of bacterial mutants.

Automated Solid Phase Extraction (SPE) LC/NMR Applied to the Structural Analysis of Extractable Compounds from a Pharmaceutical Packaging Material of Construction

The structural analysis (i.e., identification) of organic chemical entities leached into drug product formulations has traditionally been accomplished with techniques involving the combination of chromatography with mass spectrometry. These include gas chromatography/mass spectrometry (GC/MS) for volatile and semi-volatile compounds, and various forms of liquid chromatography/mass spectrometry (LC/MS or HPLC/MS) for semi-volatile and relatively non-volatile compounds. GC/MS and LC/MS techniques are complementary for structural analysis of leachables and potentially leachable organic compounds produced via laboratory extraction of pharmaceutical container closure/delivery system components and corresponding materials of construction. Both hyphenated analytical techniques possess the separating capability, compound specific detection attributes, and sensitivity required to effectively analyze complex mixtures of trace level organic compounds. However, hyphenated techniques based on mass spectrometry are limited by the inability to determine complete bond connectivity, the inability to distinguish between many types of structural isomers, and the inability to unambiguously determine aromatic substitution patterns. Nuclear magnetic resonance spectroscopy (NMR) does not have these limitations; hence it can serve as a complement to mass spectrometry. However, NMR technology is inherently insensitive and its ability to interface with chromatography has been historically challenging. This article describes the application of NMR coupled with liquid chromatography and automated solid phase extraction (SPE-LC/NMR) to the structural analysis of extractable organic compounds from a pharmaceutical packaging material of construction. The SPE-LC/NMR technology combined with micro-cryoprobe technology afforded the sensitivity and sample mass required for full structure elucidation. Optimization of the SPE-LC/NMR analytical method was achieved using a series of model compounds representing the chemical diversity of extractables. This study demonstrates the complementary nature of SPE-LC/NMR with LC/MS for this particular pharmaceutical application. The identification of impurities leached into drugs from the components and materials associated with pharmaceutical containers, packaging components, and materials has historically been done using laboratory techniques based on the combination of chromatography with mass spectrometry. Such analytical techniques are widely recognized as having the selectivity and sensitivity required to separate the complex mixtures of impurities often encountered in such identification studies, including both the identification of leachable impurities as well as potential leachable impurities produced by laboratory extraction of packaging components and materials. However, while mass spectrometry-based analytical techniques have limitations for this application, newer analytical techniques based on the combination of chromatography with nuclear magnetic resonance spectroscopy provide an added dimension of structural definition. This article describes the development, optimization, and application of an analytical technique based on the combination of chromatography and nuclear magnetic resonance spectroscopy to the identification of potential leachable impurities from a pharmaceutical packaging material. The complementary nature of the analytical techniques for this particular pharmaceutical application is demonstrated.

In-Cell NMR Spectroscopy–In vivo Monitoring of the Structure, Dynamics, Folding, and Interactions of Proteins at Atomic Resolution

Thallapuranam K Suresh Kumar, Ryan Thurman and Srinivas Jayanthi-In-Cell NMR Spectroscopy–<em>In vivo</em> Monitoring of the Structure, Dynamics, Folding, and Interactions of Proteins at Atomic Resolution

Experimental study from linear to chaotic regimes on a terahertz-frequency gyrotron oscillator

Basic wave-particle interaction dynamics from linear to chaotic regimes is experimentally studied on a frequency tunable gyrotron generating THz radiation in continuous mode (200 W) at 263 GHz which will be used for dynamic nuclear polarization nuclear magnetic resonance spectroscopy applications. In the studied system, the nonlinear dynamics associated to the wave-particle interaction is dominated by longitudinal mode competition of a given transverse TEm,p cavity-mode. This study covers a wide range of control parameter from gyro-traveling wave tube (gyro-TWT) to gyro-backward wave oscillator (gyro-BWO) like interactions for which extensive theoretical studies have been performed in the past on a simplified system. Besides the common route to chaos characterized by period doubling, other routes have been identified among which some are characterized by line-width frequency-broadening on the side-bands. The complex nonlinear dynamics is in good agreement with the theory and the experimental results are discussed on the basis of the prediction obtained with the nonlinear time-dependent self-consistent codes TWANG and EURIDICE both based on a slow-time scale formulation of the self-consistent equations governing the wave-particle dynamics.

An overview of NMR-based metabolomics to identify secondary plant compounds involved in host plant resistance.

Secondary metabolites provide a potential source for the generation of host plant resistance and development of biopesticides. This is especially important in view of the rapid and vast spread of agricultural and horticultural pests worldwide. Multiple pests control tactics in the framework of an integrated pest management (IPM) programme are necessary. One important strategy of IPM is the use of chemical host plant resistance. Up to now the study of chemical host plant resistance has, for technical reasons, been restricted to the identification of single compounds applying specific chemical analyses adapted to the compound in question. In biological processes however, usually more than one compound is involved. Metabolomics allows the simultaneous detection of a wide range of compounds, providing an immediate image of the metabolome of a plant. One of the most universally used metabolomic approaches comprises nuclear magnetic resonance spectroscopy (NMR). It has been NMR which has been applied as a proof of principle to show that metabolomics can constitute a major advancement in the study of host plant resistance. Here we give an overview on the application of NMR to identify candidate compounds for host plant resistance. We focus on host plant resistance to western flower thrips (Frankliniella occidentalis) which has been used as a model for different plant species.

Temporal Chemical Shift Correlations in Reactions Studied by Hyperpolarized Nuclear Magnetic Resonance

High-resolution nuclear magnetic resonance spectroscopy (NMR) has the capability of providing often unrivaled detail on molecular structure and dynamics. Through hyperpolarization, a decisive gain in signal strength can be realized, which extends the applicability of NMR to the investigation of rapid processes far from equilibrium. The progress of irreversible chemical and biochemical reactions can be followed by hyperpolarized NMR with relative ease, within an observable window encompassing the subsecond to second time scales. Here, we present a scheme that uses real-time, hyperpolarization enhanced NMR to make temporal correlations accessible in addition to simply monitoring reaction progress. Since nuclear spin states can be preserved even if the spin carrying atoms directly participate in a reaction, it becomes possible to correlate the positions of these atoms between the reactant and the product species, over time. We demonstrate the application of this technique to the Grignard addition of methylmagnesium bromide to 3-methylbenzophenone. The same experiment may be used for the determination of mechanisms and intermediate states in non-equilibrium processes in fields as varied as organic chemistry, enzymology, or protein folding.

How close is the bench to the bedside? Metabolic profiling in cancer research

Metabolic profiling using mass spectrometry (MS) and nuclear magnetic resonance spectroscopy (NMR) is integral to the rapidly expanding field of metabolomics, which is making progress in toxicology, plant science and various diseases, including cancer. In the area of oncology and metabolic phenotyping, researchers have probed the known changes in malignant cellular pathways using new experimental techniques to gain more insights, and others are exploiting these same cellular pathways for therapeutic drug targets and for novel cancer biomarkers, with the ultimate goal of translation to the clinic. Here, we discuss the challenges and opportunities in metabolic phenotyping for discovering novel cancer biomarkers, and we assess the clinical applicability of MS and NMR.

Interactions of chemical and natural stresses in the abalone ( Haliotis rufescens) as measured by surface-Probe localized 31P NMR

Application of nuclear magnetic resonance spectroscopy (NMR) for elucidating the sublethal actions of toxicants in intact aquatic organisms, and the influence of natural stress factors, has recently been demonstrated. Using a flow-through chamber, red abalones ( Haliotis rufescens) were exposed (at 14 °C) to ambient (35%.), hyposaline (25%.), or hyper saline (4 5%.) seawater, with and without a sublethal concentration (1.2 mg l −1) of pentachlorophenol (PCP). They were then maintained in clean seawater (at the same salinity) to check both rate and degree of recovery; phosphagen changes in foot muscle were monitored by surface-probe localized 31P NMR. Whereas different water salinities alone did not produce measurable effects, upon PCP exposure phosphoarginine levels declined, inorganic monophosphate levels increased, and intracellular pH decreased. Whereas during recovery all biochemical endpoints returned to normal, both onset and intensity of effects were significantly influenced in an unexpected manner by changes in seawater salinity. Application of 31P NMR represents a powerful new tool for elucidating the influence of natural stress factors on pollutant actions in vivo, providing a more accurate assessment of environmental effects.

Nuclear Magnetic Resonance Spectroscopy of Pullulan and Isomaltose: Complete Assignment of Chemical Shifts

AbstractOver the past decade, advances in nuclear magnetic resonance spectroscopy (NMR) and increased availability of NMR spectrometers have led to the application of NMR to a wide range of problems in many fields, including environmental chemistry. Complete assignment of carbon and proton resonances is a prerequisite for using NMR spectroscopy to monitor changes in chemical structures during the course of biochemical and biological experiments. Several corrections and additions to previously published 13C chemical shifts of pullulan and isomaltose are reported here. In addition, the 1H assignments of isomaltose are presented.

Amorphous Hydrogenated Semiconductors

AbstractSemiconducting thin films are produced in plasma and CVD reactors and are commercially useful materials because of a ubiquitous presence of hydrogen, a low density of defects, and electro-optical properties which vary systematically with composition.However, little is known about how variations in fundamental growth parameters produce concomitant changes in film properties, particularly hydrogen incorporation.The reason for this gap in knowledge derives partly from the lack of adequate structural probes of disordered materials.In recent years nuclear magnetic resonance spectroscopy (NMR) has emerged as a powerful probe of the microstructure of amorphous semiconductors.I will discuss recent developments in the application of NMR to studies of amorphous hydrogenated semiconductors, and interpret these results in terms of the chemistry and physics of the growth process.

Peptide sequence analysis by nuclear magnetic resonance spectroscopy.

Nuclear magnetic resonance spectroscopy (NMR) has found wide application to biochemical problems. This is reflected in the number of books (61, 130, 212, 243, 473) and review articles (95, 96, 163, 169, 294, 300, 325, 375) that describe this area of research. In recent years, considerable progress has been made in studies of the structure in solution of peptides, proteins, nucleotides and other biomolecules. Major technical developments were responsible for this success. These include: (1) the manufacture of superconducting magnets, (2) the general availability of Fourier transform NMR spectrometers, and (3) the discovery of paramagnetic chemical shift and relaxation enhancement (broadening) agents. In tandem, these advancements have extended the sensitivity and scope of NMR spectroscopy considerably. As a consequence, carbon-13 resonance studies no longer require an isotopic enrichment step (257, 420), and the available resolution has been improved to such a level that even the details of multiplet structure in protein 1H-spectra may be discerned (77–79, 130). The paramagnetic chemical shift and broadening probes may be used to simplify complex NMR spectra, and are useful in making peak assignments. Roberts and Jardetzky (375), in 1970, concluded that sequence determinations by NMR would likely be of limited value because of the inability to resolve adquately the resonances of 1H nuclei of the various amino acid residues. In this chapter, we will show that advances in NMR technology have generated renewed interest in peptide sequence analysis, and that this application of NMR has considerable merit and promise.