Magic-angle Spinning NMR Experiments Research Articles

Ammonothermal Synthesis and Solid-State NMR Study of the Imidonitridosilicate Rb3Si6N5(NH)6.

The imidonitridosilicate Rb3Si6N5(NH)6, being only the second representative of this compound class, was synthesized ammonothermally at 870 K and 230 MPa. Its crystal structure was solved from single-crystal X-ray diffraction data. The imidonitridosilicate crystallizes isotypically with the respective potassium compound in space group P4132 with the lattice parameter a=10.9422(4) Å forming a three-dimensional imidonitridosilicate tetrahedra network with voids for the rubidium ions. The structure model and the presence of the imide groups were verified by Fourier-Transform infrared (FTIR) and magic-angle spinning (MAS) NMR spectroscopy, using cross polarization 15N{1H} and 29Si{1H} MAS NMR experiments. Rb3Si6N5(NH)6 represents a possible intermediate during the ammonothermal synthesis of nitridosilicates. The characterization of such intermediates improves the understanding of the reaction pathway from ammonothermal solutions to nitrides. Thus, the ammonothermal synthesis is an alternative approach to the well-established high-temperature synthesis leading to the compound class of nitridosilicates.

Chemical composition of soil humin in an organic soil profile

Humin is identified as the most recalcitrant fraction of soil organic matter, but its synthesis and depth variability in the soil profile are unknown. Here, we suggest that carboxyl-rich alicyclic structures (CRAM) and fused aromatic ring structures (FAR) can process significant components of the humin structure. We conducted molecular-level characterization of humin using advanced solid-state nuclear magnetic resonance spectroscopy (SSNMR) experiments and calculated the proportion of FAR and functionalities of CRAM (OCq, –C(CH)n, –CH3) structures. Dipolar dephased (dd) direct polarization magic angle spinning (DPMAS) NMR experiments provided insights into non-protonated carbon and mobile quaternary carbons. We calculated the proportions of non-protonated aromatic carbon and non-polar aliphatic C in the organic layers of the topsoil (Oa), mid-soil (Om), and mineral subsoil (Cg). It was evident that the total aliphatic carbon constitutes a significant proportion of humin, as is the non-protonated C attached to CH3. The 13C dd/DPMAS NMR spectrum of Cg humin clearly showed the formation and preservation of hydrophobic OCq, which is indicative of CRAM structures. CRAM structures could be attached to FAR, which makes the whole structure relatively more hydrophobic, a critical humin functionality in deeper soil. The findings from our research can be successfully utilized in terrestrial carbon sequestration studies to amplify the potential of humin as a recalcitrant carbon reservoir.

Origin of Oxygen in Graphene Oxide Revealed by 17O and 18O Isotopic Labeling.

Wet-chemical oxidation of graphite in a mixture of sulfuric acid with a strong oxidizer, such as potassium permanganate, leads to the formation of graphene oxide with hydroxyl and epoxide groups as the major functional groups. Nevertheless, the reaction mechanism remains unclear and the source of oxygen is a subject of debate. It could theoretically originate from the oxidizer, water, or sulfuric acid. In this study, we employed 18O and 17O labeled reagents to experimentally elucidate the reaction mechanism and, thus, determine the origin of oxo-functional groups. Our findings reveal the multifaceted roles of sulfuric acid, acting as a dispersion medium, a dehydrating agent for potassium permanganate, and an intercalant. Additionally, it significantly acts as a source of oxygen next to manganese oxides. Through 17O solid-state magic-angle spinning (MAS) NMR experiments, we exclude water as a direct reaction partner during oxygenation. With labeling experiments, we conclude on mechanistic insights, which may be exploited for the synthesis of novel graphene derivatives.

High-efficiency low-power 13C-15N cross polarization in MAS NMR

Biomolecular solid-state magic angle spinning (MAS) NMR spectroscopy frequently relies on selective 13C-15N magnetization transfers, for various kinds of correlation experiments. Introduced in 1998, spectrally induced filtering in combination with cross polarization (SPECIFIC-CP) is a selective heteronuclear magnetization transfer experiment widely used for biological applications. At MAS frequencies below 20 kHz, commonly used for 13C-detected MAS NMR experiments, SPECIFIC-CP transfer between amide 15N and 13Cα atoms (NCA) is typically performed with radiofrequency (rf) fields set higher than the MAS frequency for both 13C and 15N channels, and high-power 1H decoupling rf field is simultaneously applied. Here, we experimentally explore a broad range of NCA zero-quantum (ZQ) SPECIFIC-CP matching conditions at the MAS frequency of 14 kHz and compare the best high- and low-power matching conditions with respect to selectivity, robustness, and sensitivity at lower 1H decoupling rf fields. We show that low-power NCA SPECIFIC-CP matching condition gives rise to 20% sensitivity enhancement compared to high-power conditions, in 2D NCA spectra of microcrystalline assemblies of HIV-1 CACTD-SP1 protein with inositol hexakis-phosphate (IP6).

A New Look at the Mechanism of Cocrystal Formation and Coformers Exchange in Processes Forced by Mechanical and/or Thermal Stimuli - ex situ and in situ Studies of Low-Melting Eutectic Mixtures.

Three different devices: ball mill, hot stage melting, and magic angle spinning (MAS) NMR rotor were used for the preparation of ethenzamide (ET) cocrystals with glutaric acid (GLU), ethylmalonic acid (EMA) and maleic acid (MAL) as coformers. In each case, well-defined binary systems (ET:EMA, ET:GLU, ET:MAL) were obtained. The common features of the two solvent free methods of cocrystal formation (grinding, melting) are presented on the basis of arguments obtained by solid state NMR spectroscopy. Thermal analysis (Differential Scanning Calorimetry) proved that the eutectic phase arises over a wide range of molar ratios of components for each of the binary systems. NMR techniques, supported by theoretical calculations, allowed to provide details about the pathway of the reaction mechanism with atomic accuracy. It was found that the formation of ET cocrystals is a complex process that requires five steps. Each step has been recognized and described. Variable temperature 1D and 2D MAS NMR experiments allowed to track physicochemical processes taking place in a molten state. Moreover, it was found that in a multicomponent mixture consisting of all four components, ET, EMA, GLU, and MAL, ET in the molten phase behaves as a specific selector choosing only one partner to form binary cocrystals according to energy preferences. The process of exchange of coformers in binary systems during grinding, melting, and NMR measurements is described. The stabilization energies (Estab ) and molecular electrostatic potential (MEP) maps computed for the cocrystals under discussion and their individual components rationalize the selection rules and explain the relationships between individual species.

Ba12 [BN2 ]6.67 H4 : A Disordered Anti-Skutterudite filled with Nitridoborate Anions.

Skutterudites are of high interest in current research due to their diversity of structures comprising empty, partially filled and filled variants, mostly based on metallic compounds. We herein present Ba12 [BN2 ]6.67 H4 , forming a non-metallic filled anti-skutterudite. It is accessed in a solid-state ampoule reaction from barium subnitride, boron nitride and barium hydride at 750 °C. Single-crystal X-ray and neutron powder diffraction data allowed to elucidate the structure in the cubic space group Im (no. 204). The barium and hydride atoms form a three-dimensional network consisting of corner-sharing HBa6 octahedra and Ba12 icosahedra. Slightly bent [BN2 ]3- units are located in the icosahedra and the voids in-between. 1 H and 11 B magic angle spinning (MAS) NMR experiments and vibrational spectroscopy further support the structure model. Quantum chemical calculations coincide well with experimental results and provide information about the electronic structure of Ba12 [BN2 ]6.67 H4 .

Ba12[BN2]6.67H4: A Disordered Anti‐Skutterudite filled with Nitridoborate Anions

AbstractSkutterudites are of high interest in current research due to their diversity of structures comprising empty, partially filled and filled variants, mostly based on metallic compounds. We herein present Ba12[BN2]6.67H4, forming a non‐metallic filled anti‐skutterudite. It is accessed in a solid‐state ampoule reaction from barium subnitride, boron nitride and barium hydride at 750 °C. Single‐crystal X‐ray and neutron powder diffraction data allowed to elucidate the structure in the cubic space group Im (no. 204). The barium and hydride atoms form a three‐dimensional network consisting of corner‐sharing HBa6 octahedra and Ba12 icosahedra. Slightly bent [BN2]3− units are located in the icosahedra and the voids in‐between. 1H and 11B magic angle spinning (MAS) NMR experiments and vibrational spectroscopy further support the structure model. Quantum chemical calculations coincide well with experimental results and provide information about the electronic structure of Ba12[BN2]6.67H4.

Simplified Preservation of Equivalent Pathways Spectroscopy.

Inspired by the recently proposed transverse mixing optimal control pulses (TROP) approach for improving signal in multidimensional magic-angle spinning (MAS) NMR experiments, we present simplified preservation of equivalent pathways spectroscopy (SPEPS). It transfers both transverse components of magnetization that occur during indirect evolutions, theoretically enabling a √2 improvement in sensitivity for each such dimension. We compare SPEPS transfer with TROP and cross-polarization (CP) using membrane protein and fibril samples at MAS of 55 and 100 kHz. In three-dimensional (3D) (H)CANH spectra, SPEPS outperformed TROP and CP by factors of on average 1.16 and 1.69, respectively, for the membrane protein, but only a marginal improvement of 1.09 was observed for the fibril. These differences are discussed, making note of the longer transfer time used for CP, 14 ms, as compared with 2.9 and 3.6 ms for SPEPS and TROP, respectively. Using SPEPS for two transfers in the 3D (H)CANCO experiment resulted in an even larger benefit in signal intensity, with an average improvement of 1.82 as compared with CP. This results in multifold time savings, in particular considering the weaker peaks that are observed to benefit the most from SPEPS.

Frequency-swept dynamic nuclear polarization

Dynamic nuclear polarization (DNP) improves the sensitivity of NMR spectroscopy by the transfer of electron polarization to nuclei via irradiation of electron-nuclear transitions with microwaves at the appropriate frequency. For fields > 5 T and using g ∼ 2 electrons as polarizing agents, this requires the availability of microwave sources operating at >140 GHz. Therefore, microwave sources for DNP have generally been continuous-wave (CW) gyrotrons, and more recently solid state, oscillators operating at a fixed frequency and power. This constraint has limited the DNP mechanisms which can be exploited, and stymied the development of new time domain mechanisms. We report here the incorporation of a microwave source enabling facile modulation of frequency, amplitude, and phase at 9 T (250 GHz microwave frequency), and we have used the source for magic-angle spinning (MAS) NMR experiments. The experiments include investigations of CW DNP mechanisms, the advantage of frequency-chirped irradiation, and a demonstration of an Overhauser enhancement of ∼25 with a recently reported water-soluble BDPA radical, highlighting the potential for affordable and compact microwave sources to achieve significant enhancement in aqueous samples, including biological macromolecules. With the development of suitable microwave amplifiers, it should permit exploration of multiple new avenues involving time domain experiments.

Great Offset Difference Internuclear Selective Transfer.

Carbon-carbon dipolar recoupling sequences are frequently used building blocks in routine magic-angle spinning NMR experiments. While broadband homonuclear first-order dipolar recoupling sequences mainly excite intra-residue correlations, selective methods can detect inter-residue transfers and long-range correlations. Here, we present the great offset difference internuclear selective transfer (GODIST) pulse sequence optimized for selective carbonyl or aliphatic recoupling at fast magic-angle spinning, here, 55 kHz. We observe a 3- to 5-fold increase in intensities compared with broadband RFDR recoupling for perdeuterated microcrystalline SH3 and for the membrane protein influenza A M2 in lipid bilayers. In 3D (H)COCO(N)H and (H)CO(CO)NH spectra, inter-residue carbonyl-carbonyl correlations up to about 5 Å are observed in uniformly 13C-labeled proteins.

Theoretical and 27Al NMR Spectroscopic Investigations of Binary Intermetallic Alkaline-Earth Aluminides.

The binary alkaline-earth aluminides AEAl2 (AE = Ca and Sr) and AEAl4 (AE = Ca-Ba) have been synthesized from the elements and investigated via powder X-ray diffraction experiments. CaAl2 adopts the cubic MgCu2-type structure (Fd3̅m), while SrAl2 crystallizes in the orthorhombic KHg2-type (Imma). LT-CaAl4 crystallizes with the monoclinic CaGa4-type (C2/m), while HT-CaAl4, SrAl4, and BaAl4 adopt the tetragonal BaAl4-type structure (I4/mmm). The close structural relation of the two CaAl4 polymorphs was established using a group-subgroup relation in the Bärnighausen formalism. In addition to the room-temperature and normal pressure phase of SrAl2, a high-pressure/high-temperature phase has been prepared using multianvil techniques, and its structural and spectroscopic parameters were determined. Elemental analysis by inductively coupled plasma mass spectrometry showed that no significant impurities with other elements besides the weighed ones are present and the chemical compositions match the synthesized ones. The title compounds have been furthermore investigated by 27Al solid-state magic angle spinning NMR experiments to validate the crystal structure and to gain information about the influence of the composition on the electron transfer and the NMR characteristics. This has also been investigated from a quantum chemical point of view using Bader charges, while the stabilities of the binary compounds in the three phase diagrams (Ca-Al, Sr-Al and Ba-Al) have been studied by calculations of formation energies per atom.

Windowed cross polarization at 55 kHz magic-angle spinning

Cross polarization (CP) transfers via Hartmann-Hahn matching conditions are one of the cornerstones of solid-state magic-angle spinning NMR experiments. Here we investigate a windowed sequence for cross polarization (wCP) at 55 kHz magic-angle spinning, placing one window (and one pulse) per rotor period on one or both rf channels. The wCP sequence is known to have additional matching conditions. We observe a striking similarity between wCP and CP transfer conditions when considering the flip angle of the pulse rather than the rf-field strength applied during the pulse. Using fictitious spin-1/2 formalism and average Hamiltonian theory, we derive an analytical approximation that matches these observed transfer conditions. We recorded data at spectrometers with different external magnetic fields up to 1200 MHz, for strong and weak heteronuclear dipolar couplings. These transfers, and even the selectivity of CP were again found to relate to flip angle (average nutation).

Fluorine-Labeled N-Boc-l-proline as a Marker for Solid-State NMR Characterization of Biofunctionalizations on Paper Substrates

An efficient approach employing 4-dimethylaminopyridine and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride as the coupling reagent is presented, which enables the functionalization of cotton linter paper substrates with the 19F spin label N-boc-cis-4-fluoro-l-proline. This spin label can be easily quantified by 19F magic angle spinning (MAS) NMR experiments to determine its loading on paper substrates. During the functionalization, the spin label stays intact, as confirmed by the 1H–19F heterocorrelation (1H → 19F CP MAS FSLG HETCOR) experiments. In combination with dynamic nuclear polarization (19F MAS DNP), the N-boc-cis-4-fluoro-l-proline spin label allows us to inspect 1 μmol/g and even lower molecule loadings on paper substrates, providing a highly sensitive local probe to analyze the structure of biofunctionalizations at the nanoscale on paper substrates in the future.

Simultaneous recoupling of chemical shift tensors of two nuclei by R-symmetry sequences

Chemical shift tensors (CSTs) are sensitive probes of structure and dynamics. R-symmetry pulse sequences (RNCSA) can efficiently recouple CSTs of varying magnitudes in magic angle spinning (MAS) NMR experiments, for a broad range of conditions and MAS frequencies. Herein, we introduce dual-channel R-symmetry pulse sequences for simultaneously recording CSTs of two different nuclei in a single experiment (DORNE-CSA). We demonstrate the performance of DORNE-CSA sequences for simultaneous measurement of 13C and 15N CSTs, on a U-13C,15N-labeled microcrystalline l-histidine. We show that the DORNE-CSA method is robust, provides accurate CST parameters, and takes only half of the measurement time compared to a pair of RNCSA experiments otherwise required for recording the CSTs of individual nuclei. DORNE-CSA approach is broadly applicable to a wide range of biological and inorganic systems.

Quantifying quadrupole effects in the NMR spectra of spin-1/2 nuclei in rotating solids.

Quantifying the NMR spectra of spin I = 1/2 nuclei coupled to quadrupolar spins (nuclei with spin quantum number, S > 1/2) has remained an arduous task in solid-state magic angle spinning (MAS) NMR experiments. In particular, extraction of chemical shift anisotropy (CSA) tensors from line-shapes of spin I = 1/2 nuclei coupled to quadrupolar spin (S = 1) in MAS experiments has remained challenging owing to the simultaneous presence of both heteronuclear dipolar interactions and quadrupolar interactions. Unlike experiments that involve only spin-1/2 nuclei, both faster spinning frequencies and stronger decoupling field strengths on the quadrupolar spins are essential to average/minimize the contributions from heteronuclear dipolar interactions. To this end, a quantitative theory based on the concept of "effective fields" is proposed to deduce optimal conditions in experiments that involve simultaneous recoupling and decoupling of heteronuclear dipolar interactions. Through analytic expressions, the spectral frequencies and intensities observed in experiments are quantified and rigorously verified. Since extraction of molecular constraints in NMR experiments involves iterative fitting of experimental data, we believe that the analytic expressions derived would speed up and be beneficial in quantifying such experiments.

The effect of methyl group rotation on 1H-1H solid-state NMR spin-diffusion spectra.

Fast magic-angle spinning (MAS) NMR experiments open the way for proton-detected NMR studies and have been explored in the past years for a broad range of materials, comprising biomolecules and pharmaceuticals. Proton-spin diffusion (SD) is a versatile polarization-transfer mechanism and plays an important role in resonance assignment and structure determination. Recently, the occurrence of negative cross peaks in 2D 1H-1H SD-based spectra has been reported and explained with higher-order SD effects, in which the chemical shifts of the involved quadruple of nuclei need to compensate each other. We herein report negative cross peaks in SD-based spectra observed for a variety of small organic molecules involving methyl groups. We combine experimental observations with numerical and analytical simulations to demonstrate that the methyl groups can give rise to coherent (SD) as well as incoherent (Nuclear Overhauser Enhancement, NOE) effects, both in principle manifesting themselves as negative cross peaks in the 2D spectra. Analytical calculations and simulations however show that higher-order coherent contributions dominate the experimentally observed negative peaks in our systems. Methyl groups are prone to the observation of such higher order coherent effects. Due to their low-frequency shifted 1H resonances, the chemical-shift separation relative to for instance aromatic protons in spatial proximity is substantial (>4.7 ppm in the studied examples) preventing any sizeable second-order spin-diffusion processes, which would mask the negative contribution to the peaks.

1H Detected Relayed Dynamic Nuclear Polarization.

Recently, it has been shown that methods based on the dynamics of 1H nuclear hyperpolarization in magic angle spinning (MAS) NMR experiments can be used to determine mesoscale structures in complex materials. However, these methods suffer from low sensitivity, especially since they have so far only been feasible with indirect detection of 1H polarization through dilute heteronuclei such as 13C or 29Si. Here we combine relayed-DNP (R-DNP) with fast MAS using 0.7 mm diameter rotors at 21.2 T. Fast MAS enables direct 1H detection to follow hyperpolarization dynamics, leading to an acceleration in experiment times by a factor 16. Furthermore, we show that by varying the MAS rate, and consequently modulating the 1H spin diffusion rate, we can record a series of independent R-DNP curves that can be analyzed jointly to provide an accurate determination of domain sizes. This is confirmed here with measurements on microcrystalline l-histidine·HCl·H2O at MAS frequencies up to 60 kHz, where we determine a Weibull distribution of particle sizes centered on a radius of 440 ± 20 nm with an order parameter of k = 2.2.

DNP-enhanced solid-state NMR spectroscopy of chromatin polymers

Chromatin is a DNA-protein polymer that represents the functional form of the genome. The main building block of chromatin is the nucleosome, a structure that contains 147 base pairs of DNA and two copies each of the histone proteins H2A, H2B, H3 and H4. Previous work has shown that magic angle spinning (MAS) NMR spectroscopy can capture the nucleosome at high resolution although studies have been challenging due to low sensitivity, the presence of dynamic and rigid components, and the complex interaction networks of nucleosomes within the chromatin polymer. Here, we use dynamic nuclear polarization (DNP) to enhance the sensitivity of MAS NMR experiments of nucleosome arrays at 100 K and show that well-resolved 13C-13C MAS NMR correlations can be obtained much more efficiently. We evaluate the effect of temperature on the chemical shifts and linewidths in the spectra and demonstrate that changes are relatively minimal and clustered in regions of histone-DNA or histone-histone contacts. We also compare samples prepared with and without DNA and show that the low temperature 13C-13C correlations exhibit sufficient resolution to detect chemical shift changes and line broadening for residues that form the DNA-histone interface. On the other hand, we show that the measurement of DNP-enhanced 15N-13C histone-histone interactions within the nucleosome core is complicated by the natural 13C abundance network in the sample. Nevertheless, the enhanced sensitivity afforded by DNP can be used to detect long-range correlations between histone residues and DNA. Overall, our experiments demonstrate that DNP-enhanced MAS NMR spectroscopy of chromatin samples yields spectra with high resolution and sensitivity and can be used to capture functionally relevant protein-DNA interactions that have implications for gene regulation and genome organization.

In situ 13 C solid-state polarization transfer NMR to follow starch transformations in food.

Convenience food products tend to alter their quality and texture while stored. Texture-giving food components are often starch-rich ingredients, such as pasta or rice. Starch transforms depending on time, temperature and water content, which alters the properties of products. Monitoring these transformations, which are associated with a change in mobility of the starch chain segments, could optimize the quality of food products containing multiple ingredients. In order to do so, we applied a simple and efficient in situ 13 C solid-state magic angle spinning (MAS) NMR approach, based on two different polarization transfer schemes, cross polarization (CP) and insensitive nuclei enhanced by polarization transfer (INEPT). The efficiency of the CP and INEPT transfer depends strongly on the mobility of chain segments-the time scale of reorientation of the CH-bond and the order parameter. Rigid crystalline or amorphous starch chains give rise to CP peaks, whereas mobile gelatinized starch chains appear as INEPT peaks. Comparing 13 C solid-state MAS NMR experiments based on CP and INEPT allows insight into the progress of gelatinization, and other starch transformations, by reporting on both rigid and mobile starch chains simultaneously with atomic resolution by the 13 C chemical shift. In conjunction with 1 H solid-state MAS NMR, complementary information about other food components present at low concentration, such as lipids and protein, can be obtained. We demonstrate our approach on starch-based products and commercial pasta as a function of temperature and storage.

3D-printed stators & drive caps for magic-angle spinning NMR

3D printing has evolved into an invaluable tool for rapid and cost-effective production of intricate parts. In this paper we describe 3D printing and other rapid prototyping methods to fabricate 3.2 mm stators and drive caps for use in magic angle spinning (MAS) NMR experiments. These components can be fabricated with the assistance of computer-aided design (CAD) software and at a fraction of the cost of commercial parts. Additionally, we show that the performance of these 3D printed stators and drive caps is comparable to commercially available systems and that they have significant advantages over their machined counterparts.