High-field Nuclear Magnetic Resonance Spectrometer Research Articles

Fragment Screening and Fast Micromolar Detection on a Benchtop NMR Spectrometer Boosted by Photoinduced Hyperpolarization.

Fragment-based drug design is a well-established strategy for rational drug design, with nuclear magnetic resonance (NMR) on high-field spectrometers as the method of reference for screening and hit validation. However, high-field NMR spectrometers are not only expensive, but require specialized maintenance, dedicated space, and depend on liquid helium cooling which became critical over the recurring global helium shortages. We propose an alternative to high-field NMR screening by applying the recently developed approach of fragment screening by photoinduced hyperpolarized NMR on a cryogen-free 80 MHz benchtop NMR spectrometer yielding signal enhancements of up to three orders in magnitude. It is demonstrated that it is possible to discover new hits and kick-off drug design using a benchtop NMR spectrometer at low micromolar concentrations of both protein and ligand. The approach presented performs at higher speed than state-of-the-art high-field NMR approaches while exhibiting a limit of detection in the nanomolar range. Photoinduced hyperpolarization is known to be inexpensive and simple to be implemented, which aligns greatly with the philosophy of benchtop NMR spectrometers. These findings open the way for the use of benchtop NMR in near-physiological conditions for drug design and further life science applications.

Continuous flow synthesis of pyridinium salts accelerated by multi-objective Bayesian optimization with active learning.

We report a human-in-the-loop implementation of the multi-objective experimental design via a Bayesian optimization platform (EDBO+) towards the optimization of butylpyridinium bromide synthesis under continuous flow conditions. The algorithm simultaneously optimized reaction yield and production rate (or space-time yield) and generated a well defined Pareto front. The versatility of EDBO+ was demonstrated by expanding the reaction space mid-campaign by increasing the upper temperature limit. Incorporation of continuous flow techniques enabled improved control over reaction parameters compared to common batch chemistry processes, while providing a route towards future automated syntheses and improved scalability. To that end, we applied the open-source Python module, nmrglue, for semi-automated nuclear magnetic resonance (NMR) spectroscopy analysis, and compared the acquired outputs against those obtained through manual processing methods from spectra collected on both low-field (60 MHz) and high-field (400 MHz) NMR spectrometers. The EDBO+ based model was retrained with these four different datasets and the resulting Pareto front predictions provided insight into the effect of data analysis on model predictions. Finally, quaternization of poly(4-vinylpyridine) with bromobutane illustrated the extension of continuous flow chemistry to synthesize functional materials.

Accurate measurements of self-diffusion coefficients with benchtop NMR using a QM model-based approach.

The measurement of self-diffusion coefficients using pulsed-field gradient (PFG) nuclear magnetic resonance (NMR) spectroscopy is a well-established method. Recently, benchtop NMR spectrometers with gradient coils have also been used, which greatly simplify these measurements. However, a disadvantage of benchtop NMR spectrometers is the lower resolution of the acquired NMR signals compared to high-field NMR spectrometers, which requires sophisticated analysis methods. In this work, we use a recently developed quantum mechanical (QM) model-based approach for the estimation of self-diffusion coefficients from complex benchtop NMR data. With the knowledge of the species present in the mixture, signatures for each species are created and adjusted to the measured NMR signal. With this model-based approach, the self-diffusion coefficients of all species in the mixtures were estimated with a discrepancy of less than 2 % compared to self-diffusion coefficients estimated from high-field NMR data sets of the same mixtures. These results suggest benchtop NMR is a reliable tool for quantitative analysis of self-diffusion coefficients, even in complex mixtures.

Correlation of high-field and zero- to ultralow-field NMR properties using 2D spectroscopy.

The field of zero- to ultralow-field (ZULF) nuclear magnetic resonance (NMR) is currently experiencing rapid growth, owing to progress in optical magnetometry and attractive features of ZULF-NMR such as low hardware cost and excellent spectral resolution achieved under ZULF conditions. In this work, an approach is proposed and demonstrated for simultaneous acquisition of ZULF-NMR spectra of individual 13C-containing isotopomers of chemical compounds in a complex mixture. The method makes use of fast field cycling such that the spin evolution takes place under ZULF conditions, whereas signal detection is performed in a high-field NMR spectrometer. This method has excellent sensitivity, also allowing easy assignment of ZULF-NMR spectra to specific analytes in the mixture. We demonstrate that the spectral information is the same as that given by ZULF-NMR, which makes the method suitable for creating a library of ZULF-NMR spectra of various compounds and their isotopomers. The results of the field-cycling experiments can be presented in a convenient way as 2D-NMR spectra with the direct dimension giving the high-field 13C-NMR spectrum (carrying the chemical-shift information) and the indirect dimension giving the ZULF-NMR spectrum (containing information about proton-carbon J-couplings). Hence, the method can be seen as a variant of heteronuclear J-resolved spectroscopy, one of the first 2D-NMR techniques.

Incorporating Benchtop NMR Spectrometers in the Undergraduate Lab: Understanding Resolution and Circumventing Second-Order Effects

Nuclear magnetic resonance (NMR) spectroscopy is commonly introduced to students in the classroom, but its hands-on use in undergraduate laboratories is far less common. The significant costs to purchase and maintain a traditional high-field NMR spectrometer means that many institutions cannot offer this service to their students. Benchtop NMR spectrometers represent a very powerful alternative at a fraction of the cost and laboratory footprint. However, concerns about resolution and managing second-order effects can hamper its incorporation into the undergraduate curricula. Herein, we describe how resolution at the benchtop level differs from traditional high-field instruments and provide a thorough, tabulated list of 204 molecules which exhibit little to no second-order effects at the lower magnetic fields utilized in benchtop NMR spectroscopy.

Discovery and validation of an NMR-based metabolomic profile in urine as TB biomarker

Despite efforts to improve tuberculosis (TB) detection, limitations in access, quality and timeliness of diagnostic services in low- and middle-income countries are challenging for current TB diagnostics. This study aimed to identify and characterise a metabolic profile of TB in urine by high-field nuclear magnetic resonance (NMR) spectrometry and assess whether the TB metabolic profile is also detected by a low-field benchtop NMR spectrometer. We included 189 patients with tuberculosis, 42 patients with pneumococcal pneumonia, 61 individuals infected with latent tuberculosis and 40 uninfected individuals. We acquired the urine spectra from high and low-field NMR. We characterised a TB metabolic fingerprint from the Principal Component Analysis. We developed a classification model from the Partial Least Squares-Discriminant Analysis and evaluated its performance. We identified a metabolic fingerprint of 31 chemical shift regions assigned to eight metabolites (aminoadipic acid, citrate, creatine, creatinine, glucose, mannitol, phenylalanine, and hippurate). The model developed using low-field NMR urine spectra correctly classified 87.32%, 85.21% and 100% of the TB patients compared to pneumococcal pneumonia patients, LTBI and uninfected individuals, respectively. The model validation correctly classified 84.10% of the TB patients. We have identified and characterised a metabolic profile of TB in urine from a high-field NMR spectrometer and have also detected it using a low-field benchtop NMR spectrometer. The models developed from the metabolic profile of TB identified by both NMR technologies were able to discriminate TB patients from the rest of the study groups and the results were not influenced by anti-TB treatment or TB location. This provides a new approach in the search for possible biomarkers for the diagnosis of TB.

NMR spectroscopy goes mobile: Using NMR as process analytical technology at the fume hood.

Nuclear magnetic resonance (NMR) is potentially a very powerful process analytical technology (PAT) tool as it gives an atomic resolution picture of the reaction mixture without the need for chromatography. NMR is well suited for interrogating transient intermediates, providing kinetic information via NMR active nuclei, and most importantly provides universally quantitative information for all species in solution. This contrasts with commonly used PAT instruments, such as Raman or Flow-infrared (IR), which requires a separate calibration curve for every component of the reaction mixture. To date, the large footprint of high-field (≥400 MHz) NMR spectrometers and the immobility of superconducting magnets, coupled with strict requirements for the architecture for the room it is to be installed, have been a major obstacle to using this technology right next to fume hoods where chemists perform synthetic work. Here, we describe the use of a small, lightweight 60 MHz Benchtop NMR system (Nanalysis Pro-60) located on a mobile platform, that was used to monitor both small and intermediate scale Grignard formation and coupling reactions. We also show how low field NMR can provide a deceptively simple yes/no answer (for a system that would otherwise require laborious off-line testing) in the enrichment of one component versus another in a kilogram scale distillation. Benchtop NMR was also used to derive molecule specific information from Flow-IR, a technology found in most manufacturing sites, and compare the ease at which the concentrations of the reaction mixtures can be derived by NMR versus IR.

Recent Contributions of Nuclear Magnetic Resonance in Organocatalysis Mechanism Elucidation

Background: Nuclear Magnetic Resonance (NMR) is one of the most employed techniques in structural elucidation of organic compounds. In addition to its use in structural characterization, it has been widely employed in the investigation of reaction mechanisms, especially those involving catalysis. Objective: In this review, we aim to provide recent examples of the interface of NMR and organocatalysis reaction mechanism. Methods: Selected examples of different approaches for mechanism elucidation will be presented, such as isotopic effect, catalyst labelling and online reaction monitoring. A discussion involving the use of solid-state NMR will also be disclosed. Conclusion: NMR consists of a non-destructive technique, extremely useful in the real-time identification of intermediates in crude reaction mixtures. With the advent of two-dimensional experiments and high field NMR spectrometers, the reports of studies involving mechanistic elucidation were greatly enhanced. In this context, nowadays NMR appears as a powerful tool for the comprehension of reaction mechanisms, including the possibility of the proposal of unknown reaction mechanisms within organocatalysis.

In-Vivo NMR Spectroscopy: A Powerful and Complimentary Tool for Understanding Environmental Toxicity.

Part review, part perspective, this article examines the applications and potential of in-vivo Nuclear Magnetic Resonance (NMR) for understanding environmental toxicity. In-vivo NMR can be applied in high field NMR spectrometers using either magic angle spinning based approaches, or flow systems. Solution-state NMR in combination with a flow system provides a low stress approach to monitor dissolved metabolites, while magic angle spinning NMR allows the detection of all components (solutions, gels and solids), albeit with additional stress caused by the rapid sample spinning. With in-vivo NMR it is possible to use the same organisms for control and exposure studies (controls are the same organisms prior to exposure inside the NMR). As such individual variability can be reduced while continual data collection over time provides the temporal resolution required to discern complex interconnected response pathways. When multidimensional NMR is combined with isotopic labelling, a wide range of metabolites can be identified in-vivo providing a unique window into the living metabolome that is highly complementary to more traditional metabolomics studies employing extracts, tissues, or biofluids.

Determination of Protein ps-ns Motions by High-Resolution Relaxometry.

Many of the functions of biomacromolecules can be rationalized by the characterization of their conformational energy landscapes: the structures of the dominant states, transitions between states and motions within states. Nuclear magnetic resonance (NMR) spectroscopy is the technique of choice to study internal motions in proteins. The determination of motions on picosecond to nanosecond timescales requires the measurement of nuclear spin relaxation rates at multiple magnetic fields. High sensitivity and resolution are obtained only at high magnetic fields, so that, until recently, site-specific relaxation rates in biomolecules were only measured over a narrow range of high magnetic fields. This limitation was particularly striking for the quantification of motions on nanosecond timescales, close to the correlation time for overall rotational diffusion. High-resolution relaxometry is an emerging technique to investigate picosecond-nanosecond motions of proteins. This approach uses a high-field NMR spectrometer equipped with a sample shuttle device, which allows for the measurement of the relaxation rate constants at low magnetic fields, while preserving the sensitivity and resolution of a high-field NMR spectrometer. The combined analysis of high-resolution relaxometry and standard high-field relaxation data provides a more accurate description of the dynamics of proteins, in particular in the nanosecond range. The purpose of this chapter is to describe how to perform high-resolution relaxometry experiments and how to analyze the rates measured with this technique.

Full Correlations across Broad NMR Spectra by Two-Field Total Correlation Spectroscopy.

Total correlation spectroscopy (TOCSY) is a key experiment to assign nuclear magnetic resonance (NMR) spectra of complex molecules. Carbon-13 TOCSY experiments are essential to assign signals of protein side chains. However, the performance of carbon-13 TOCSY deteriorates at high magnetic fields since the necessarily limited radiofrequency irradiation fails to cover the broad range of carbon-13 frequencies. Here, we introduce a new concept to overcome the limitations of TOCSY by using two-field NMR spectroscopy. In two-field TOCSY experiments, chemical shifts are labelled at high field but isotropic mixing is performed at a much lower magnetic field, where the frequency range of the spectrum is drastically reduced. We obtain complete correlations between all carbon-13 nuclei belonging to amino acids across the entire spectrum: aromatic, aliphatic and carboxylic. Two-field TOCSY should be a robust and general approach for the assignment of uniformly carbon-13 labelled molecules in high-field and ultra-high field NMR spectrometers beyond 1000 MHz.

Photos of NMR pizza oven and grill, photo of laboratory lamp

Firing up the NMR Back in 1984, when Rich Haack first laid eyes on a high-field nuclear magnetic resonance spectrometer, he had a vision for the instrument that went beyond analyzing chemical compounds. “I thought if you could hollow it out, it would make a neat smoker, outdoor oven, or fire pit,” he tells Newscripts. Would that make it an NMR-B-Q ? Haack, a chemist who works in industry, never gave up on the idea. He just thought he would never get his hands on an NMR to hack. That all changed two years ago when he had the opportunity to acquire a defunct 400-MHz instrument from Open Technologies, a company that buys, sells, and services NMRs. Don Frank, the company’s chief executive officer, says the instrument was going to be scrapped. He knew of Haack’s plans, so he drove the old NMR from Open Technologies’ warehouse in Indiana to

In situ measurement of liquid-liquid equilibria by medium field nuclear magnetic resonance

A new method for non-invasive measurement of liquid-liquid equilibria (LLE) using a compact medium field nuclear magnetic resonance (NMR) spectrometer is presented. Mixing of all components, phase separation, and analysis of the composition of the coexisting phases is performed in situ in an NMR glass tube. Thus, the experimental effort is reduced and errors caused by sampling are eliminated. Furthermore, calibration of the analysis method is not necessary as quantitative information is obtained directly from the NMR spectra. The proposed method for studying LLE in situ can be swiftly conducted in standard chemical laboratories as medium field NMR spectrometer do not require dedicated laboratory infrastructure but enable convenient handling and fast analysis of the samples. In the present work, four non-reactive ternary solvent systems with closed miscibility gap (toluene + acetone + water, diethyl ether + acetone + water, diethyl ether + methanol + water, and acetonitrile + ethanol + cyclohexane) and one reactive ternary system (water + acetic acid + acetic anhydride) were investigated at a temperature of 22.0 °C using 1H medium field NMR spectroscopic measurements. For comparison, the composition of the coexisting phases is also examined for one non-reactive system (acetonitrile + ethanol + cyclohexane) using 13C medium field NMR spectroscopy as well as spatially resolved spectroscopy in a conventional high field NMR spectrometer. The comparison of the results of the present work to literature data shows that the new proposed method enables swift and reliable investigations of LLE.

Determination of Alcohol Content in Alcoholic Beverages Using 45 MHz Benchtop NMR Spectrometer

Alcohol or ethanol is considered the most widely used recreational drug worldwide, and its production, consumption, and sale are strictly regulated by laws. Alcohol content of alcoholic beverages (wine, beers, and spirits) is about 3–50% v/v. Analytical methods to determine the alcohol content must be reliable, precise, and accurate. In this study, the amount of ethanol in several alcoholic beverages was determined using a 45 MHz low-field benchtop NMR (nuclear magnetic resonance) spectrometer. Internal standard and standard addition analytical methods were utilized to quantify ethanol. For both methods, acetic acid or acetonitrile was used as internal standard to quantify alcohol content by using the peak area corresponding to the methyl peaks of ethanol, acetic acid, or acetonitrile. Results showed that internal standard method gave values of percent alcohol that are in close agreement with the indicated label as confirmed by running the samples in a 400 MHz high-field NMR spectrometer using acetic acid as internal standard. This study demonstrates the utility of a benchtop NMR spectrometer that can provide an alternative technique to analyze percent alcohol in alcoholic products.

Optimized Droplet Sizing of Water-in-Crude Oil Emulsions Using Nuclear Magnetic Resonance

Water-in-crude oil emulsions are an increasing problem during production. Essential to any emulsion breaking method is an ability to accurately measure droplet size distributions; this is rendered extremely difficult given that the samples are both concentrated and opaque. Here, we systematically consider the use of a standard, low-field benchtop nuclear magnetic resonance (NMR) apparatus to accurately measure the droplet size distributions. Such measurements are challenging because the NMR signal from the oil phase erroneously contributes to the measured water droplet size distribution. Conventionally, the oil-phase signal is nulled-out based on differences in the NMR T1 relaxation parameter between water and oil. However, in the case of crude oil, the oil presents a broad T1 distribution, rendering this approach infeasible. On the basis of this oil T1 distribution, we present an optimization routine that adjusts various NMR measurement timing parameters [observation time (Δ) and inversion time (Tinv)] to effectively eliminate this erroneous crude oil contribution. An implementation of this optimization routine was validated against measurements performed using unambiguous chemical-shift selection of the water (droplet) signal, as would conventionally be provided by high-field superconducting NMR spectrometers. We finally demonstrate successful droplet sizing of a range of water-in-crude oil emulsions.

Design and performance of a microsecond time-resolved photo-chemically induced dynamic nuclear polarization add-on for a high-field nuclear magnetic resonance spectrometer

We describe the design and operation of microsecond time-resolved photo-chemically induced dynamic nuclear polarization hardware, designed to go with a commercial 600 MHz nuclear magnetic resonance (NMR) spectrometer. A frequency-tripled neodymium-doped yttrium/aluminum garnet (Nd:YAG) laser is used as the light source, with a system of mirrors or prisms to route light to the NMR sample from above, removing the need for NMR probe modifications. The experiment has been designed for a shared NMR spectrometer and is straightforward and inexpensive to assemble and operate.

An analytical perspective of NMR spectroscopy of biological fluids and cells.

Nuclear magnetic resonance (NMR) spectroscopy, first used to study a biological system 40 years ago,I,2 is the forerunner of both magnetic resonance imaging (MRI) and magnetic resonance spectroscopy (MRS). However, its use in clinical science has been minimal until recently. Both MRI and MRS instruments are variants of the MNR spectrometers found in many university chemistry laboratories.' These high-field spectrometers can provide detailed chemical information of a high quality on small sample volumes (0'5 mL) of fluids and cells4,s and are capable of identifying species in solution in a quantitative manner. Wide-bore magnetic resonance techniques for imaging have been accepted and integrated into the panoply of clinical tests as one of the new range of 'scanners' [e.g. positron emission tomography (PET) and computer assisted tomography (CAT)] as an adjunct to X-rays.6--8 The advent of ethical approval of higher field magnets (1,5 T or 30 MHz) allows the use of MRS of imaged regions of the body for the study of in situ, in vivo metabolism.v This review sets out to highlight how NMR spectroscopy of biologic~l samples such as fluids and cells could be used 10 routine clinical biochemistry. It is extremely uncommon to find narrow bore (5 rom diameter) high-field NMR spectrometers in clinical chemistry departments. As a direct result many of the formative studies reviewed here arose out of collaborative studies between clinical biochemists and chemists. Inevitably such work has been performed in vitro. During the following discussion, where possible, we have included

5 Applications of Modern-High-Field NMR Spectroscopy in Medicinal Chemistry

Publisher Summary This chapter discusses the applications of modern-high-field nuclear magnetic resonance (NMR) spectroscopy in medicinal chemistry. One of the most useful aspects of NMR compared with other types of spectroscopy is the elucidation of spatial geometry. Modern high-field instruments give spectra in which the fine structure of the signals is more clearly resolved but may still not always show first-order coupling. The analysis of coupling constants can yield stereochemical and conformational data, examples of which are given in the chapter. The difficulty of interpreting crowded one-dimensional spectra has been tackled by the development of experiments that display spectral data in two dimensions. Some of these experiments can be difficult to appreciate fully other than in theoretical terms. However, this should not preclude the medicinal chemist from making use of the spectra obtained for the purposes of structure elucidation. The most dramatic developments have been in the field of biological NMR. The introduction of very high-field NMR spectrometers has increased the resolution of complex spectra of biomolecules, such as peptides, proteins, and polysaccharides while the application of two-dimensional experiments has aided spectral assignment.

NMR Studies of Biophysical Problems

Nuclear magnetic resonance (NMR) has become more and more actively used in various biophysical and biochemical studies. Under such circumstances it seems timely to publish a special number of this Journal containing papers which introduce recent attainments in this field to non-specialist readers.The importance of NMR in the studies of biological molecules and higher-order systems stems from the following features. (1) Thanks to the developments of high-field NMR spectrometers with superconducting magnets, it is now possible to perform detailed investigations of the microstructure of biopolymers. (2) Possibilities of gaining a deeper insight into molecular motions are opened by the advent of pulse. Fourier transform techniques through accurate measurements of the spin-lattice relaxation times. (3) New aspects of biomolecular structures are being revealed by the application of recent experimental methods such as the use of paramagnetic probes. (4) The observation of resonances of various nuclei other than 1H (e. g., 13C, 15N, 31P, 19F, and 2H) provides a powerful tool for solving many problems which have not been so far tractable.In Japan also, NMR studies of biological systems are steadily increasing in number and advancing in quality. We find that quite a high percentage of the papers reported at the domestic meetings and conferences during 1974-1975 are concerned with either measurements of the spin-lattice relaxation times or the use of paramagnetic probes. Therefore, The papers published in this special number are also more or less related to these topics. The title and author (s) of each paper are as follows.1. Introduction (M. Tasumi & T. Iizuka)2. Molecular motions and spectral assignments based on the NMR relaxation mea- surements (R. Chujõ)3. Use of nuclear magnetic relaxation times to the determination of molecular conformation in solution (K. Akasaka & S. Shibata)4. Side-chain conformation of polyamino acids (K. Nagayama)5. NMR studies on the phospholipids from an unsaturated fatty-acid auxotroph of E. Coli (H. Akutsu & Y. Kyogoku)6. Structural studies on membrane models by NMR spectroscopy (M. Kainosho)7. Studies on the structure and function of hemoproteins and heme-enzymes by FT-NMR with superconducting magnet (I. Morishima, T. Inubushi & T. Iizuka)8. Structural studies of biomolecules by the lanthanide-ion probe method (M. Tasumi & F. Inagaki)