Nuclear Magnetic Resonance Probe Research Articles

Noncontact In Situ Multidiagnostic NMR/Dielectric Spectroscopy.

Introduction of a dielectric material in a nuclear magnetic resonance (NMR) probe head modifies the frequency response of the probe circuit, a phenomenon revealed by detuning of the probe. For NMR spectroscopy, this detuning is corrected for by tuning and matching the probe head prior to the NMR measurement. The magnitude of the probe detuning, "the dielectric shift", provides direct access to the dielectric properties of the sample, enabling NMR spectrometers to simultaneously perform both dielectric and NMR spectroscopy. By measuring sample dielectric permittivity as a function of frequency, dielectric permittivity spectroscopy can be performed using the new methodology. As a proof of concept, this was evaluated on methanol, ethanol, 1-propanol, 1-pentanol, and 1-octanol using a commercial cross-polarization magic angle spinning (CPMAS) NMR probe head. The results accurately match the literature data collected by standard dielectric spectroscopy techniques. Subsequently, the method was also applied to investigate the solvent-surface interactions of water confined in the micropores of an MFI-type, hydrophilic zeolite with a Si/Al ratio of 11.5. In the micropores, water adsorbs to Bro̷nsted acid sites and defect sites, resulting in a drastically decreased dielectric permittivity of the nanoconfined water. Theoretical background for the new methodology is provided using an effective electric circuit model of a CPMAS probe head with a solenoid coil, describing the detuning resulting from the insertion of dielectric samples in the probe head.

Large dynamics of a phase separating arginine-glycine-rich domain revealed via nuclear and electron spins

Liquid-liquid phase separation is the key process underlying formation of membrane-less compartments in cells. A highly dynamic cellular body with rapid component exchange is Cajal body (CB), which supports the extensive compositional dynamics of the RNA splicing machinery, spliceosome. Here, we select an arginine-glycine (RG)-rich segment of coilin, the major component of CB, establish its RNA-induced phase separation, and through combined use of nuclear magnetic resonance (NMR) and electron paramagnetic resonance (EPR) probes, interrogate its dynamics within the crowded interior of formed droplets. Taking advantage of glycine-based singlet-states, we show that glycines retain a large level of sub-nanoseconds dynamics inside the coilin droplets. Furthermore, the continuous-wave (CW) and electron-electron dipolar (PELDOR) and electron-nucleus hyperfine coupling EPR data (HYSCORE) support the RNA-induced formation of dynamic coilin droplets with high coilin peptide concentrations. The combined NMR and EPR data reveal the high dynamics of the RG-rich coilin within droplets and suggest its potential role in the large dynamics of CBs.

Automated test apparatus for bench-testing the magnetic field homogeneity of NMR transceiver coils

We describe an automated hands-off bench testing method for measuring the magnetic field profile of transceiver coils for nuclear magnetic resonance (NMR). The scattering parameter (S-parameter) data is measured using a portable network analyzer, and the results are automatically exported to a computer for plotting and viewing. This assay dramatically reduces the time needed to measure the magnetic field (B1) homogeneity profile of a transceiver coil while also improving accuracy relative to manual operation. Here, we demonstrate the method on a saddle coil of a solution-state NMR probe in comparison to profiles obtained using NMR spectroscopy measurements. We also measure the axial homogeneity of a variable-pitch solenoid.

Identifying placental metabolic biomarkers of preterm birth using nuclear magnetic resonance of intact tissue samples

IntroductionOur understanding of the etiology of preterm birth (PTB) is incomplete; however, recent evidence has found a strong association between placental dysfunction and PTB. Altered placental metabolism may precede placental dysfunction and therefore the study of placental metabolic profiles could identify early biomarkers of PTB. In this study, we evaluated the placental metabolome in PTB in intact tissue samples using nuclear magnetic resonance (NMR) and spectral editing. MethodsPlacental tissue samples were collected from nine term pregnancies and nine preterm pregnancies (<37 weeks’ gestation). 1H NMR experiments on unprocessed tissue samples were performed using a high field magnet (500 MHz spectrometer) and a comprehensive multiphase NMR probe. The relative concentrations of 23 metabolites were corrected for gestational age and compared between groups. ResultsThe relative concentration of valine, glutamate and creatine were significantly decreased while alanine, choline and glucose were elevated in placentas from PTB pregnancies compared to controls (p < 0.05). Multivariate analysis using principal component analysis showed the PTB and control groups were significantly separated (p < 0.0001) and pathway analysis identified perturbations in the glycine, serine and threonine metabolism, aminoacyl-tRNA biosynthesis and valine, leucine and isoleucine biosynthesis pathways. ConclusionPTB is associated with significant alterations in placental metabolism. This study helps improve our understanding of the etiology of PTB. It also highlights the potential for small molecule metabolites to serve as placental metabolic biomarkers to aid in the prediction and diagnosis of PTB. The results can be translated to clinical use via in utero magnetic resonance spectroscopy.

Improving the Sensitivity of Enantioanalysis with Densely Fluorinated NMR Probes.

Nuclear magnetic resonance (NMR) spectroscopy has long been utilized as a classic method for chiral discrimination of enantiomers. However, its sensitivity limitations have hindered the detection of analytes at low concentrations. In this study, we present our efforts to overcome this challenge by employing chiral NMR probes that are labeled with a significant number of chemically equivalent 19F atoms. Specifically, we have designed and synthesized three chiral palladium pincer complexes, all of which are labeled with nonafluoro-tert-butoxy groups to enhance detectability. The recognition of enantiomers with the probe induces distinct changes in microenvironments, resulting in differential perturbations on the chemical shift of the 19F atoms in proximity. This method is applicable to the enantiodifferentiation of various amines, amino alcohols, and amino acid esters. The abundance of 19F atoms enables the detection of chiral analytes at low concentrations, which is otherwise challenging to achieve through traditional 1H NMR-based analysis. Two of the probes are constructed with asymmetric pincer ligands with structurally varied sidearms, allowing for facile manipulation of the chiral binding pocket. The C2 symmetrical probe possesses 36 equivalent 19F atoms, enabling the determination of enantiocomposition of samples with concentrations in the low micromolar range.

Development of High-Temperature Superconducting Coil for Solid-State NMR Experiments

Abstract A high-temperature superconducting (HTS) coil, in which YBa2Cu3O7-δ is vapor-deposited on the surface of a sapphire plane, has been developed to increase signal-to-noise ratios (SNR) in solid-state nuclear magnetic resonance (NMR) measurements. The Q-factor of the NMR probe is found to be 16,125 at the resonant frequency of 38.525 MHz. A single sharp signal, full width at half maximum of 20 ppm, can be detected by the HTS coil in a stationary solid-state 79Br NMR spectrum of a polycrystalline potassium bromide at the temperature of 9 K. Compared with the results for an isomorphic copper coil, an improvement of a factor of 6.6 is obtained for SNR, indicating that accumulation time of NMR experiments can be reduced to 1/44.

An Optimized Two-Dimensional Quantitative Nuclear Magnetic Resonance Strategy for the Rapid Quantitation of Diester-Type C19-Diterpenoid Alkaloids from Aconitum carmichaelii.

With the development of nuclear magnetic resonance (NMR) spectrometers and probes, two-dimensional quantitative nuclear magnetic resonance (2D qNMR) technology with a high signal resolution and great application potential has become increasingly accessible for the quantitation of complex mixtures. However, the requirement that the relaxation recovery time be equal to at least five times T1 (longitudinal relaxation time) makes it difficult for 2D qNMR to simultaneously achieve high quantitative accuracy and high data acquisition efficiency. By comprehensively using relaxation optimization and nonuniform sampling, we successfully established an optimized 2D qNMR strategy for HSQC experiments at the half-hour level and then accurately quantified the diester-type C19-diterpenoid alkaloids in Aconitum carmichaelii. The optimized strategy had the advantages of high efficiency, high accuracy, good reproducibility, and low cost and thus could serve as a reference to optimize 2D qNMR experiments for quantitative analysis of natural products, metabolites, and other complex mixtures.

Preparation of ultracold ions in strong magnetic field and application to gas-phase NMR spectroscopy II

This paper presents a gas-phase nuclear magnetic resonance (NMR) apparatus, which may enable a tandem combination with an ion cyclotron resonance mass spectrometer. The NMR technique is widely used for analyzing the physical and chemical properties of materials; however, it is mostly limited to materials in condensed phases. Here, we construct a gas-phase NMR apparatus to extend this technique to gas-phase molecular ions. We outline the principle of NMR detection based on a Stern-Gerlach-type experiment and describe the experimental procedures and results for the preparation and manipulation of ultracold ions, which are fundamental for the proposed NMR detection. The development of the gas-phase NMR probe and radio-frequency excitation system and the features of the apparatus are presented. We also discuss the feasibility of combining this apparatus with an ion cyclotron resonance cell for realizing an NMR apparatus for mass-selected molecular ions briefly.

Site-specific protein backbone deuterium 2Hα quadrupolar patterns by proton-detected quadruple-resonance 3D 2HαcαNH MAS NMR spectroscopy

A novel deuterium-excited and proton-detected quadruple-resonance three-dimensional (3D) 2HαcαNH MAS nuclear magnetic resonance (NMR) method is presented to obtain site-specific 2Hα deuterium quadrupolar couplings from protein backbone, as an extension to the 2D version of the experiment reported earlier. Proton-detection results in high sensitivity compared to the heteronuclei detection methods. Utilizing four independent radiofrequency (RF) channels (quadruple-resonance), we managed to excite the 2Hα, then transfer deuterium polarization to its attached Cα, followed by polarization transfers to the neighboring backbone nitrogen and then to the amide proton for detection. This experiment results in an easy to interpret HSQC-like 2D 1H–15N fingerprint NMR spectrum, which contains site-specific deuterium quadrupolar patterns in the indirect third dimension. Provided that four-channel NMR probe technology is available, the setup of the 2HαcαNH experiment is relatively straightforward, by using low power deuterium excitation and polarization transfer schemes we have been developing. To our knowledge, this is the first demonstration of a quadruple-resonance MAS NMR experiment to link 2Hα quadrupolar couplings to proton-detection, extending our previous triple-resonance demonstrations. Distortion-free excitation and polarization transfer of ∼160–170 kHz 2Hα quadrupolar coupling were presented by using a deuterium RF strength of ∼20 kHz. From these 2Hα patterns, an average backbone order parameter of S = 0.92 was determined on a deuterated SH3 sample, with an average η = 0.22. These indicate that SH3 backbone represents sizable dynamics in the microsecond timescale where the 2Hα lineshape is sensitive. Moreover, site-specific 2Hα T1 relaxation times were obtained for a proof of concept. This 3D 2HαcαNH NMR experiment has the potential to determine structure and dynamics of perdeuterated proteins by utilizing deuterium as a novel reporter.

Solid-state NMR spectroscopy at ultrahigh resolution for structural and dynamical studies of MOFs

To characterize the structure and dynamics of metal–-organic frameworks (MOFs) in-depth at the molecular level, it is necessary to pursue high-resolution solid-state magic angle spinning (MAS) nuclear magnetic resonance (NMR) spectroscopy. Spectral resolution is usually affected by the quality of materials and various experimental conditions, of which magic angle (MA) accuracy is a crucial determinant. The current industrial criteria for MA calibration based on the common standard of KBr were found insufficient in guaranteeing optimal resolution MAS NMR for highly ordered MOFs. To drive towards higher-resolution MAS NMR spectroscopy, we propose a calibration protocol for more accurate MA with a higher-precision criterion based on 79Br MAS NMR of KBr, where the linewidth ratio of the fifth-order spinning sideband to the central band of KBr should be less than 1.00. As a result, ultrahigh-resolution 13C cross-polarization (CP) MAS NMR of MOF-5 is achieved with minimal linewidths as low as 4 Hz, and therefore MOF-5 can be used as a new standard convenient for verifying MA accuracy and also optimizing 13C CP conditions. Maintaining high-precision MA under variable temperature (VT) was found challenging on certain commercial MAS NMR probes, as was systematically investigated by VT NMR using KBr and MOF-5. Nevertheless, ultrahigh-resolution MAS NMR spectroscopy with stable MA under VT is employed to reveal fine structures and linker dynamics of a series of Zn-based MOFs with highly regulated structures. The ultrahigh-resolution NMR methodcan be generally applied to study a broad range of MOFs and other materials.

Differentiation of Superoxide Radical Anion and Singlet Oxygen and Their Concurrent Quantifications by Nuclear Magnetic Resonance.

While there have been various techniques, assays, and commercial kits developed to measure reactive oxygen species (ROS) with varying degrees of success, there is a lack of innovative methods to differentiate and quantify them simultaneously. In this work, we demonstrate a 19F nuclear magnetic resonance (NMR)-based method to differentiate two important types of ROS, superoxide radical anion and singlet oxygen, and to quantify them concurrently. By taking advantage of the unique chemical reactivity of two fluorine-containing molecules, 4-fluoro-3-methylphenyl boronic acid and 4-fluoro-3-methylphenol, serving as 19F NMR probes, we are able to differentiate and quantify, for the first time, superoxide radical anion and singlet oxygen generated by photosensitizers (PSs) concurrently. The results reveal that relative amounts of superoxide radical anion and singlet oxygen generated by a PS under light illumination are oftentimes sensitive to the environment, such as the presence or absence of electron donors. This method provides a means to identify the type of mechanism by which a PS functions under a given condition. We envision that this relatively simple, yet robust, method would be beneficial to a broad range of ROS-pertinent studies, such as photodynamic therapy and photoredox reactions.

Probing Ligand Binding Sites on Large Proteins by Nuclear Magnetic Resonance Spectroscopy of Genetically Encoded Non-Canonical Amino Acids.

N6-(((trimethylsilyl)-methoxy)carbonyl)-l-lysine (TMSK) and N6-trifluoroacetyl-l-lysine (TFAK) are non-canonical amino acids, which can be installed in proteins by genetic encoding. In addition, we describe a new aminoacyl-tRNA synthetase specific for N6-(((trimethylsilyl)methyl)-carbamoyl)-l-lysine (TMSNK), which is chemically more stable than TMSK. Using the dimeric SARS-CoV-2 main protease (Mpro) as a model system with three different ligands, we show that the 1H and 19F nuclei of the solvent-exposed trimethylsilyl and CF3 groups produce intense signals in the nuclear magnetic resonance (NMR) spectrum. Their response to active-site ligands differed significantly when positioned near rather than far from the active site. Conversely, the NMR probes failed to confirm the previously reported binding site of the ligand pelitinib, which was found to enhance the activity of Mpro by promoting the formation of the enzymatically active dimer. In summary, the amino acids TMSK, TMSNK, and TFAK open an attractive path for site-specific NMR analysis of ligand binding to large proteins of limited stability and at low concentrations.

Implementation of a dipole magnet power supply control system to improve magnetic field stability at the CSRe storage ring facility for precision mass measurement

Stable magnetic field is crucial for the precision experiments conducted at the heavy ion storage rings. Besides the current stability of the power supply, the magnetic field is also influenced by varying ambient factors such as temperatures. This paper proposed a dual-loop hysteresis control method to passively change the output current of the power supply according to the magnetic field monitored by the nuclear magnetic resonance (NMR) probes. As a result, the long-term relative changes of magnetic field were reduced from dB/B≈5.75× 10−5 to 2.3 × 10−5 within three days.

Design of a cryogen-free high field dual EPR and DNP probe

We present the design and construction of a cryogen free, dual electron paramagnetic resonance (EPR) and nuclear magnetic resonance (NMR) probe for novel dynamic nuclear polarization (DNP) experiments and concurrent “in situ” analysis of DNP mechanisms. We focus on the probe design that meets the balance between EPR, NMR, and low temperature performance, while maintaining a high degree of versatility: allowing multi-nuclear NMR detection as well as broadband DNP/EPR excitation/detection. To accomplish high NMR/EPR performance, we implement a novel inductively coupled double resonance NMR circuit (1H-13C) in a solid state probe operating at cryogenic temperatures. The components of the circuit were custom built to provide maximum NMR performance, and the physical layout of this circuit was numerically optimized via magnetic field simulations to allow maximum microwave transmission to the sample for optimal EPR performance. Furthermore this probe is based around a cryogen free gas exchange cryostat and has been designed to allow unlimited experiment times down to 8.5 Kelvin with minimal cost. The affordability of EPR/DNP experiment is an extremely important aspect for broader impact with magnetic resonance measurements. The purpose of this article is to provide as complete information as we have available for others with interest in building a dual DNP/EPR instrument based around a cryogen-free cryostat.

From hemolymph to in-vivo: The potential of a 1 mm microlitre flow probe with separate lock chamber for NMR metabolomics in mass limited environmental samples

Metabolomics is one of the leading approaches for understanding the toxic-mode-of-action of environmental contaminants. Nuclear Magnetic Resonance (NMR) spectroscopy has been commonly used in metabolomic studies; however, its main drawback is its relatively low sensitivity, making it challenging to study mass limited but environmentally crucial samples. In this work a 1 mm microlitre probe modified with a separate lock chamber to address this challenge, provided substantial improvements in mass sensitivity relative to conventional 5 mm NMR probes. The 1 mm probe is used to analyze various components of the model organism Daphnia magna, including hemolymph, parthenogenetic eggs, dormant eggs, and neonates. A μL volume flow system is designed for the 1 mm probe to perform an in-vivo exposure of neonates to high salt concentrations. The metabolic investigation of these samples was only achieved due to the minimum sample requirements and high salt tolerance of the probe, demonstrating that the 1 mm microlitre probe modified with a separate lock chamber holds significant potential for future metabolomic studies of mass limited samples.

Molecular identification and quantification of defect sites in metal-organic frameworks with NMR probe molecules

The defects in metal-organic frameworks (MOFs) can dramatically alter their pore structure and chemical properties. However, it has been a great challenge to characterize the molecular structure of defects, especially when the defects are distributed irregularly in the lattice. In this work, we applied a characterization strategy based on solid-state nuclear magnetic resonance (NMR) to assess the chemistry of defects. This strategy takes advantage of the coordination-sensitive phosphorus probe molecules, e.g., trimethylphosphine (TMP) and trimethylphosphine oxide (TMPO), that can distinguish the subtle differences in the acidity of defects. A variety of local chemical environments have been identified in defective and ideal MOF lattices. The geometric dimension of defects can also be evaluated by using the homologs of probe molecules with different sizes. In addition, our method provides a reliable way to quantify the density of defect sites, which comes together with the molecular details of local pore environments. The comprehensive solid-state NMR strategy can be of great value for a better understanding of MOF structures and for guiding the design of MOFs with desired catalytic or adsorption properties.

Direct measurement of the 3He+ magnetic moments

Helium-3 has nowadays become one of the most important candidates for studies in fundamental physics1–3, nuclear and atomic structure4,5, magnetometry and metrology6, as well as chemistry and medicine7,8. In particular, 3He nuclear magnetic resonance (NMR) probes have been proposed as a new standard for absolute magnetometry6,9. This requires a high-accuracy value for the 3He nuclear magnetic moment, which, however, has so far been determined only indirectly and with a relative precision of 12 parts per billon10,11. Here we investigate the 3He+ ground-state hyperfine structure in a Penning trap to directly measure the nuclear g-factor of 3He+{g}_{I}^{{prime} }=-,4.2550996069(30{)}_{{rm{stat}}}(17{)}_{{rm{sys}}}, the zero-field hyperfine splitting {E}_{{rm{HFS}}}^{exp }=-,8,,665,,649,,865.77{(26)}_{{rm{stat}}}{(1)}_{{rm{sys}}} Hz and the bound electron g-factor {g}_{e}^{{rm{exp }}}=-,2.00217741579(34{)}_{{rm{stat}}}(30{)}_{{rm{sys}}}. The latter is consistent with our theoretical value {g}_{e}^{{rm{theo}}}=-,2.00217741625223(39) based on parameters and fundamental constants from ref. 12. Our measured value for the 3He+ nuclear g-factor enables determination of the g-factor of the bare nucleus {g}_{I}=-,4.2552506997(30{)}_{{rm{stat}}}(17{)}_{{rm{sys}}}(1{)}_{{rm{theo}}} via our accurate calculation of the diamagnetic shielding constant13{sigma }_{{}^{3}{mathrm{He}}^{+}}=0.00003550738(3). This constitutes a direct calibration for 3He NMR probes and an improvement of the precision by one order of magnitude compared to previous indirect results. The measured zero-field hyperfine splitting improves the precision by two orders of magnitude compared to the previous most precise value14 and enables us to determine the Zemach radius15 to {r}_{Z}=2.608(24) fm.

Comparing the Potential of Helmholtz and Planar NMR Microcoils for Analysis of Intact Biological Samples.

Nuclear magnetic resonance (NMR) spectroscopy has played an integral role in medical and environmental metabolic research. However, smaller biological entities, such as eggs and small tissue samples, are becoming increasingly important to better understand toxicity, biological growth/development, and diseases. Unfortunately, their small sizes make them difficult to study using conventional 5 mm NMR probes due to limited sensitivity. The use of microcoil NMR holds great potential for the analysis of such samples, where the coil can be designed to match the sample size to significantly improve NMR mass sensitivity and the filling factor. Here, we compare the potential of planar and Helmholtz microcoil designs to execute complex experiments for the analysis of intact, mass-limited biological samples. The planar coil offers the advantage of an open access design, potentially allowing flow systems to be incorporated and varying sample sizes to be studied; however, its relatively inhomogeneous B1 field leads to reduced NMR performance. The Helmholtz microcoil overcomes this drawback with its symmetrical design, improving B1 homogeneity across the sample but with the caveat that the size and shape of the sample is limited to the spacing between the two parallel coils. The line shape, sensitivity, and RF performance are compared on both coils using standard samples and biological samples. This study found that the Helmholtz microcoil used here considerably outperforms the planar coil in multipulse experiments and has great potential to study complex biological samples in the 50-200 nL range.

Application of Counter-wound Multi-arm Spirals in HTS Resonator Design.

Significant sensitivity improvements have been achieved by utilizing high temperature superconducting (HTS) resonators in nuclear magnetic resonance (NMR) probes. Many nuclei such as 13C benefit from strong excitation fields which cannot be produced by traditional HTS resonator designs. We investigate the use of double-sided, counter-wound multi-arm spiral HTS resonators with the aim of increasing the excitation field at the required nuclear Larmor frequency for 13C. When compared to double-sided, counter-wound spiral resonators with similar geometry, simulations indicate that the multi-arm spiral version develops a more uniform current distribution. Preliminary tests of a two-arm resonator indicate that it may produce a stronger excitation field.

Effect of Ligands and Transducers on the Neurotensin Receptor 1 Conformational Ensemble.

Using a discrete, intracellular 19F nuclear magnetic resonance (NMR) probe on transmembrane helix 6 of the neurotensin receptor 1 (NTS1), we aim to understand how ligands and transducers modulate the receptor's structural ensemble in a solution. For apo NTS1, 19F NMR spectra reveal an ensemble of at least three conformational substates (one inactive and two active-like) in equilibrium that exchange on the millisecond to second timescale. Dynamic NMR experiments reveal that these substates follow a linear three-site exchange process that is both thermodynamically and kinetically remodeled by orthosteric ligands. As previously observed in other G protein-coupled receptors (GPCRs), the full agonist is insufficient to completely stabilize the active-like state. The inactive substate is abolished upon coupling to β-arrestin-1 (βArr1) or the C-terminal helix of Gαq, which comprises ≳60% of the GPCR/G protein interface surface area. Whereas βArr1 exclusively selects for pre-existing active-like substates, the Gαq peptide induces a new substate. Both transducer molecules promote substantial line broadening of active-like states, suggesting contributions from additional microsecond to millisecond exchange processes. Together, our study suggests that (i) the NTS1 allosteric activation mechanism may be alternatively dominated by induced fit or conformational selection depending on the coupled transducer, and (ii) the available static structures do not represent the entire conformational ensemble observed in a solution.