Crystal structure validation of verinurad via proton-detected ultra-fast MAS NMR and machine learning.

The recent development of ultra-fast magic-angle spinning (MAS) (>100 kHz) provides new opportunities for structural characterization in solids. Here, we use NMR crystallography to validate the structure of verinurad, a microcrystalline active pharmaceutical ingredient. To do this, we take advantage of 1H resolution improvement at ultra-fast MAS and use solely 1H-detected experiments and machine learning methods to assign all the experimental proton and carbon chemical shifts. This framework provides a new tool for elucidating chemical information from crystalline samples with limited sample volume and yields remarkably faster acquisition times compared to 13C-detected experiments, without the need to employ dynamic nuclear polarization.

Pure Isotropic Proton NMR Spectra in Solids using Deep Learning.

The resolution of proton solid-state NMR spectra is usually limited by broadening arising from dipolar interactions between spins. Magic-angle spinning alleviates this broadening by inducing coherent averaging. However, even the highest spinning rates experimentally accessible today are not able to completely remove dipolar interactions. Here, we introduce a deep learning approach to determine pure isotropic proton spectra from a two-dimensional set of magic-angle spinning spectra acquired at different spinning rates. Applying the model to 8 organic solids yields high-resolution 1 H solid-state NMR spectra with isotropic linewidths in the 50-400 Hz range.

Two-dimensional Pure Isotropic Proton Solid State NMR.

One key bottleneck of solid-state NMR spectroscopy is that 1 H NMR spectra of organic solids are often very broad due to the presence of a strong network of dipolar couplings. We have recently suggested a new approach to tackle this problem. More specifically, we parametrically mapped errors leading to residual dipolar broadening into a second dimension and removed them in a correlation experiment. In this way pure isotropic proton (PIP) spectra were obtained that contain only isotropic shifts and provide the highest 1 H NMR resolution available today in rigid solids. Here, using a deep-learning method, we extend the PIP approach to a second dimension, and for samples of L-tyrosine hydrochloride and ampicillin we obtain high resolution 1 H-1 H double-quantum/single-quantum dipolar correlation and spin-diffusion spectra with significantly higher resolution than the corresponding spectra at 100 kHz MAS, allowing the identification of previously overlapped isotropic correlation peaks.

Atomic-Level Structure of Zinc-Modified Cementitious Calcium Silicate Hydrate.

It has recently been demonstrated that the addition of zinc can enhance the mechanical strength of tricalcium silicates (C3S) upon hydration, but the structure of the main hydration product of cement, calcium silicate hydrate (C-S-H), in zinc-modified formulations remains unresolved. Here, we combine 29Si DNP-enhanced solid-state nuclear magnetic resonance (NMR), density functional theory (DFT)-based chemical shift computations, and molecular dynamics (MD) modeling to determine the atomic-level structure of zinc-modified C-S-H. The structure contains two main new silicon species (Q(1,Zn) and Q(2p,Zn)) where zinc substitutes Q(1) silicon species in dimers and bridging Q(2b) silicon sites, respectively. Structures determined as a function of zinc content show that zinc promotes an increase in the dreierketten mean chain lengths.

Pure Isotropic Proton Solid State NMR.

Resolution in proton solid state magic angle sample spinning (MAS) NMR is limited by the intrinsically imperfect nature of coherent averaging induced by either MAS or multiple pulse sequence methods. Here, we suggest that instead of optimizing and perfecting a coherent averaging scheme, we could approach the problem by parametrically mapping the error terms due to imperfect averaging in a k-space representation, in such a way that they can be removed in a multidimensional correlation leaving only the desired pure isotropic signal. We illustrate the approach here by determining pure isotropic 1H spectra from a series of MAS spectra acquired at different spinning rates. For six different organic solids, the approach is shown to produce pure isotropic 1H spectra that are significantly narrower than the MAS spectrum acquired at the fastest possible rate, with linewidths down to as little as 48 Hz. On average, we observe a 7-fold increase in resolution, and up to a factor of 20, as compared with spectra acquired at 100 kHz MAS. The approach is directly applicable to a range of solids, and we anticipate that the same underlying principle for removing errors introduced here can be applied to other problems in NMR spectroscopy.

Theory and simulations of homonuclear three-spin systems in rotating solids.

The homonuclear dipolar coupling is the internal spin interaction that contributes the most to the line shapes in magic-angle-spinning (MAS) 1H NMR spectra of solids, and linewidths typically extend over several hundred Hertz, limiting the 1H resolution. Understanding and reducing this contribution could provide rich structural information for organic solids. Here, we use average Hamiltonian theory to study two- and three-spin systems in the fast MAS regime. Specifically, we develop analytical expressions to third order in the case of two and three inequivalent spins (I = ½). The results show that the full third-order expression of the Hamiltonian, without secular approximations or truncation to second order, is the description that agrees the best, by far, with full numerical calculations. We determine the effect on the NMR spectrum of the different Hamiltonian terms, which are shown to produce both residual shifts and splittings in the three-spin systems. Both the shifts and splittings have a fairly complex dependence on the spinning rate with the eigenstates having a polynomial ωr dependence. The effect on powder line shapes is also shown, and we find that the anisotropic residual shift does not have zero average so that the powder line shape is broadened and shifted from the isotropic position. This suggests that in 1H MAS spectra, even at the fastest MAS rates attainable today, the positions observed are not exactly the isotropic shifts.

The Atomic-Level Structure of Cementitious Calcium Aluminate Silicate Hydrate Determined by NMR.

We review our recent paper which resolves the long-standing dilemma of the location and nature of the six-fold coordinated aluminum in calcium aluminate silicate hydrate (C-A-S-H) samples. First principles calculations predict that at high Ca:Si and H2O ratios, aluminum is incorporated into the bridging sites of the linear silicate chains and that the stable coordination number is six. We confirm this hypothesis experimentally by one- and two-dimensional dynamic nuclear polarization enhanced 27 Al and 29 Si solid-state NMR experiments in which we correlate the distinctive six-fold coordinated aluminum NMR signal at 5 ppm to 29 Si NMR signals from silicates in C-A-S-H.

The Atomic-Level Structure of Cementitious Calcium Aluminate Silicate Hydrate.

Despite use of blended cements containing significant amounts of aluminum for over 30 years, the structural nature of aluminum in the main hydration product, calcium aluminate silicate hydrate (C-A-S-H), remains elusive. Using first-principles calculations, we predict that aluminum is incorporated into the bridging sites of the linear silicate chains and that at high Ca:Si and H2O ratios, the stable coordination number of aluminum is six. Specifically, we predict that silicate-bridging [AlO2(OH)4]5- complexes are favored, stabilized by hydroxyl ligands and charge balancing calcium ions in the interlayer space. This structure is then confirmed experimentally by one- and two-dimensional dynamic nuclear polarization enhanced 27Al and 29Si solid-state NMR experiments. We notably assign a narrow 27Al NMR signal at 5 ppm to the silicate-bridging [AlO2(OH)4]5- sites and show that this signal correlates to 29Si NMR signals from silicates in C-A-S-H, conflicting with its conventional assignment to a "third aluminate hydrate" (TAH) phase. We therefore conclude that TAH does not exist. This resolves a long-standing dilemma about the location and nature of the six-fold-coordinated aluminum observed by 27Al NMR in C-A-S-H samples.

Improving the Sensitivity of FESTA Methods for the Analysis of Fluorinated Mixtures.

The analysis of complex mixtures is an important but often intractable problem. When species contain sparse fluorine atoms, NMR spectra of fluorine-containing spin systems can be efficiently extracted from an intact mixture using the recently proposed FESTA (Fluorine-Edited Selective TOCSY Acquisition) methodology. Here an alternative approach to the existing selective reverse INEPT FESTA (SRI-FESTA) experiment is described, based on the use of a modulated spin echo for the initial excitation. MODO-FESTA (modulated echo FESTA) is simpler and has a significant sensitivity advantage over SRI-FESTA. Comparisons are presented of the relative sensitivity and spectral purity of the two types of methods.

Homonuclear Decoupling in 1 H†NMR of Solids by Remote Correlation.

The typical linewidths of 1 H NMR spectra of powdered organic solids at 111 kHz magic-angle spinning (MAS) are of the order of a few hundred Hz. While this is remarkable in comparison to the tens of kHz observed in spectra of static samples, it is still the key limit to the use of 1 H in solid-state NMR, especially for complex systems. Here, we demonstrate a novel strategy to further improve the spectral resolution. We show that the anti-z-COSY experiment can be used to reduce the residual line broadening of 1 H NMR spectra of powdered organic solids. Results obtained with the anti-z-COSY sequence at 100 kHz MAS on thymol, ß-AspAla, and strychnine show an improvement in resolution of up to a factor of two compared to conventional spectra acquired at the same spinning rate.

FESTA: An Efficient Nuclear Magnetic Resonance Approach for the Structural Analysis of Mixtures Containing Fluorinated Species.

In complex mixtures, proton nuclear magnetic resonance (NMR) spectra are often very crowded, making spectral analysis complicated or even impossible, particularly when detailed structural information about the mixture components is needed. A new 1D NMR method (fluorine-edited selective TOCSY acquisition, FESTA) is introduced that facilitates the structural analysis of mixtures of species that contain fluorine. It allows simplified 1H spectra to be obtained that show only those protons that are in a spin system coupled to fluorine of interest. The new method is illustrated by factorizing a complex 1H spectrum into subspectra for individual spin systems involving different 19F sites.

13C Satellite-Free 1H NMR Spectra.

A new NMR experiment (Destruction of Interfering Satellites by Perfect Echo Low-pass filtration, DISPEL) is introduced that facilitates the analysis of low-level components in high dynamic range mixtures by suppressing one-bond 13C satellite signals in 1H spectra. Since the natural abundance of 13C is around 1.1%, these satellites appear at 0.54% of the intensity of a parent peak, mimicking and often masking impurity signals. The new experiment suppresses one-bond 13C satellite signals, with high efficiency, at negligible cost in signal-to-noise ratio, and over a wide range of one-bond coupling constants, without the need for broadband 13C decoupling.

Barriers to resolution in 1H NMR of rotating solids.

The role of 1H solid-state NMR in structure elucidation of solids is becoming more preponderant, particularly as faster magic-angle spinning rates (MAS) become available which improve 1H detected assignment strategies. However, current 1H spectral resolution is still relatively poor, with linewidths of typically a few hundred Hz, even at the fastest rates available today. Here we detail and assess the factors limiting proton linewidths and line shapes in MAS experiments with five different samples, exemplifying the different sources of broadening that affect the residual linewidth. We disentangle the different contributions through one- and two-dimensional experiments: by using dilution to identify the contribution of ABMS; by using extensive deuteration to identify the dipolar contributions; and by using variable MAS rates to determine the ratio between homogeneous and inhomogeneous components. We find that the overall widths and the nature of the different contributions to the linewidths can vary very considerably. While we find that faster spinning always yields narrower lines and longer coherence lifetimes, we also find that for some resonances the dipolar contribution is no longer dominant at 100 kHz MAS. When the inhomogeneous sources of broadening, such as ABMS and chemical shift disorder, are dominant, two-dimensional 1H-1H correlation experiments yield better resolution for assignment. Particularly the extraction of the antidiagonal of a 2D peak will remove any correlated inhomogeneous broadening, giving substantially narrower 1H linewidths.

Atomic-level structure determination of amorphous molecular solids by NMR.

Structure determination of amorphous materials remains challenging, owing to the disorder inherent to these materials. Nuclear magnetic resonance (NMR) powder crystallography is a powerful method to determine the structure of molecular solids, but disorder leads to a high degree of overlap between measured signals, and prevents the unambiguous identification of a single modeled periodic structure as representative of the whole material. Here, we determine the atomic-level ensemble structure of the amorphous form of the drug AZD4625 by combining solid-state NMR experiments with molecular dynamics (MD) simulations and machine-learned chemical shifts. By considering the combined shifts of all 1H and 13C atomic sites in the molecule, we determine the structure of the amorphous form by identifying an ensemble of local molecular environments that are in agreement with experiment. We then extract and analyze preferred conformations and intermolecular interactions in the amorphous sample in terms of the stabilization of the amorphous form of the drug.

1H Hyperpolarization of Solutions by Overhauser Dynamic Nuclear Polarization with 13C-1H Polarization Transfer.

Dynamic nuclear polarization (DNP) is a method that can significantly increase the sensitivity of nuclear magnetic resonance. The only effective DNP mechanism for in situ hyperpolarization in solution is Overhauser DNP, which is inefficient for 1H at high magnetic fields. Here we demonstrate the possibility of generating significant 1H hyperpolarization in solution at room temperature. To counter the poor direct 1H Overhauser DNP, we implement steady-state 13C Overhauser DNP in solutions and then transfer the 13C hyperpolarization to 1H via a reverse insensitive nuclei enhanced by polarization transfer scheme. We demonstrate this approach using a 400 MHz gyrotron-equipped 3.2 mm magic angle spinning DNP system to obtain 1H DNP enhancement factors of 48, 8, and 6 for chloroform, tetrachloroethane, and phenylacetylene, respectively, at room temperature.

Hyperpolarization transfer pathways in inorganic materials.

Dynamic nuclear polarization can be used to hyperpolarize the bulk of proton-free inorganic materials in magic angle spinning NMR experiments. The hyperpolarization is generated on the surface of the material with incipient wetness impregnation and from there it is propagated towards the bulk through homonuclear spin diffusion between weakly magnetic nuclei. This method can provide significant gains in sensitivity for MAS NMR spectra of bulk inorganic compounds, but the pathways of the magnetization transfer into the material have not previously been elucidated. Here we show how two-dimensional experiments can be used to study spin diffusion from the surface of a material towards the bulk. We find that hyperpolarization can be efficiently relayed from surface sites to multiple bulk sites simultaneously, and that the bulk sites also engage in rapid polarization exchange between themselves. We also show evidence that the surface peaks can exchange polarization between different sites in cases of disorder.

Advanced characterization of regioselectively substituted methylcellulose model compounds by DNP enhanced solid-state NMR spectroscopy.

Dynamic Nuclear Polarization MAS NMR is introduced to characterize model methylcellulose ether compounds at natural isotopic abundance. In particular an approach is provided to determine the position of the methyl ether group within the repeating unit. Specifically, natural abundance 13C-13C correlation experiments are used to characterize model 3-O-methylcellulose and 2,3-O-dimethylcellulose, and identify changes in chemical shifts with respect to native cellulose. We also probe the use of through space connectivity to the closest carbons to the CH3 to identify the substitution site on the cellulose ether. To this end, a series of methylcellulose ethers was prepared by a multistep synthesis approach. Key intermediates in these reactions were 2,6-O-diprotected thexyldimethylsilyl (TDMS) cellulose and 6-O-monoprotected TDMS cellulose methylated under homogeneous conditions. The products had degrees of substitution of 0.99 (3-O-methylcellulose) and 2.03 (2,3-O-dimethylcellulose) with exclusively regioselective substitution. The approaches developed here will allow characterization of the substitution patterns in cellulose ethers.

Fast remote correlation experiments for 1H homonuclear decoupling in solids.

In 1H MAS spectra, the residual homogeneous broadening under MAS is due to a combination of higher-order shifts and splittings. We have recently shown how the two-dimensional anti-z-COSY experiment can be used for the removal of the splittings. However, this requires spectra with high resolution in the indirect dimension (t1), leading to experiment times of hours. Here, we show how anti-z-COSY can be adapted to be combined with the two-dimensional one pulse (TOP) transformation which leads to significantly reduced experimental time while retaining the line narrowing effect. The experiment is demonstrated on a powdered sample of L-histidine monohydrochloride monohydrate, where the new TAZ-COSY sequence at 100 kHz MAS, yields between a factor 1.6 and 2.3 increase in resolution compared with the equivalent one-pulse experiment, in just 20 min. The same methodology is also adapted for the acquisition of liquid state 1H homodecoupled data, and an example is given for testosterone.

Suppression of 13C satellites in 1H DOSY spectra.

Diffusion-ordered spectroscopy (DOSY) is a valuable tool for the analysis of intact mixtures, since it can separate the signals of components according to their apparent diffusion coefficients. However, DOSY experiments are acutely sensitive to spectral quality, and especially to signal overlap, which can lead to misleading apparent diffusion coefficients. Here, we introduce a new NMR experiment to reduce signal overlap in mixtures with a wide range of concentrations, by removing one-bond 13C satellites. In such high dynamic range mixtures, 13C isotopomer signals from major components can overlap with signals from minor components, causing problematic distortions in the diffusion domain of a DOSY spectrum. The new method, Oneshot-iDISPEL, is a combination of the Oneshot and DISPEL experiments, and its performance has been demonstrated on a Greek alcoholic beverage, ouzo, which contains small amounts of anise flavour components and sucrose. Ethanol is a major component, and the suppression of its 13C satellites reduces signal overlap with minor components, offering significant improvement in DOSY spectra.

Ultraclean pure shift NMR.

"Pure shift" methods can greatly improve the resolution of proton NMR spectra. However, current pure shift spectra show small periodic artefacts that prevent their use for studying dilute mixture components. A new technique, compatible with all current pure shift methods, is presented that suppresses such sidebands to arbitrary order, allowing ultraclean spectra to be obtained.