Probing Long‐Range Anisotropic Interactions: a General and Sign‐Sensitive Strategy to Measure 1H–1H Residual Dipolar Couplings as a Key Advance for Organic Structure Determination

Residual dipolar couplings (RDCs) are amongst the most powerful NMR parameters for organic structure elucidation. In order to maximize their effectiveness in increasingly complex cases such as flexible compounds, a maximum of RDCs between nuclei sampling a large distribution of orientations is needed, including sign information. For this, the easily accessible one‐bond ¹H–¹³C RDCs alone often fall short. Long‐range ¹H–¹H RDCs are both abundant and typically sample highly complementary orientations, but accessing them in a sign‐sensitive way has been severely obstructed due to the overflow of ¹H–¹H couplings. Here, we present a generally applicable strategy that allows the measurement of a large number of ¹H–¹H RDCs, including their signs, which is based on a combination of an improved PSYCHEDELIC method and a new selective constant‐time β‐COSY experiment. The potential of ¹H–¹H RDCs to better determine molecular alignment and to discriminate between enantiomers and diastereomers is demonstrated.     

Long‐Range Residual Dipolar Couplings: A Tool for Determining the Configuration of Small Molecules

Together with NOE and J coupling, one‐bond residual dipolar coupling (RDC), which reports on the three‐dimensional orientation of an internuclear vector in the molecular frame, plays an important role in the conformation and configuration analysis of small molecules in solution by NMR spectroscopy. When the molecule has few C? H bonds, or too many bonds are in parallel, the available RDCs may not be sufficient to obtain the alignment tensor used for structure elucidation. Long‐range RDCs that connect nuclei over multiple bonds are normally not parallel to the single bonds and therefore complement one‐bond RDCs. Herein we present a method for extracting the long‐range RDC of a chosen proton or group of protons to all remotely connected carbon atoms, including non‐protonated carbon atoms. Alignment tensors fitted directly to the total long‐range couplings (T=J+D) enabled straightforward analysis of both the long‐range and one‐bond RDCs for strychnine.     

Probing Long-Range Anisotropic Interactions: a General and Sign-Sensitive Strategy to Measure 1H–1H Residual Dipolar Couplings as a Key Advance for Organic Structure Determination

Residual dipolar couplings (RDCs) are amongst the most powerful NMR parameters for organic structure elucidation. In order to maximize their effectiveness in increasingly complex cases such as flexible compounds, a maximum of RDCs between nuclei sampling a large distribution of orientations is needed, including sign information. For this, the easily accessible one-bond ¹H–¹³C RDCs alone often fall short. Long-range ¹H–¹H RDCs are both abundant and typically sample highly complementary orientations, but accessing them in a sign-sensitive way has been severely obstructed due to the overflow of ¹H–¹H couplings. Here, we present a generally applicable strategy that allows the measurement of a large number of ¹H–¹H RDCs, including their signs, which is based on a combination of an improved PSYCHEDELIC method and a new selective constant-time β-COSY experiment. The potential of ¹H–¹H RDCs to better determine molecular alignment and to discriminate between enantiomers and diastereomers is demonstrated.     

Twist Grain Boundary Liquid‐Crystalline Phases under the Effect of the Magnetic Field: A Complete 2H and 13C NMR Study

A liquid crystal ( HZL 7/* ) containing an (S)‐2‐methylbutyl‐(S)‐lactate unit in the chiral chain, is investigated by means of ²H and ¹³C NMR spectroscopy in order to obtain information on its orientational order, its molecular structure and the effect of external magnetic fields on the supramolecular structure of its phases. This mesogen presents very peculiar mesomorphic properties and exhibits frustrated TGBA* and TGBC* phases in a wide temperature range up to 60 °C, as well as an additional phase transition from TGBC₁* to TGBC₂*. ²H NMR measurements show, for the first time, a peculiar magnetic field effect in unwinding the supramolecular structure of both the TGBA* and TGBC* phases. This effect is particularly evident at higher magnetic fields, while different behaviour is observed at lower magnetic fields. This indicates that the supramolecular structure is very sensitive to magnetic fields of the order of 1 Tesla. Moreover, the analysis of the ²H and ¹³C NMR spectra of HZL 7/* allow us to obtain several structural properties, such as the tilt angle of the TGBC* phases and the local orientational order parameters referred to the phenyl and biphenyl fragments. This is the first structural characterization of the frustrated phases of these complexes by means of NMR.     

NMR structural characterization from one‐bond 13C?13C couplings: Complete assignment of a hydrogen‐poor depsidone

The connectivity, conformation, tautomeric form, and dynamics of a new depsidone (perisalazinic acid) were characterized using one‐bond ¹³C? ¹³C NMR scalar couplings (¹J_CC) obtained from the INADEQUATE experiment. Characterization of perisalazinic acid using more conventional NMR techniques is problematic due to the extremely limited number of C? H protons present. In the present study, 81 candidate structures were considered and a best fit structure was selected by comparing computed ¹J_CC values for each candidate to 15 experimental values. Of the six flexible moieties in perisalazinic acid, three are adequately represented by a single orientation stabilized by intramolecular hydrogen bonding. The three remaining groups are present as mixtures of conformers with two sites consisting of a pair of conformations and another disordered over six orientations. This study demonstrates the feasibility of complete three‐dimensional structural characterization of an unknown using only theoretical and experimental ¹J_CC values.     

CHARMM36 all‐atom additive protein force field: Validation based on comparison to NMR data

Protein structure and dynamics can be characterized on the atomistic level with both nuclear magnetic resonance (NMR) experiments and molecular dynamics (MD) simulations. Here, we quantify the ability of the recently presented CHARMM36 (C36) force field (FF) to reproduce various NMR observables using MD simulations. The studied NMR properties include backbone scalar couplings across hydrogen bonds, residual dipolar couplings (RDCs) and relaxation order parameter, as well as scalar couplings, RDCs, and order parameters for side‐chain amino‐ and methyl‐containing groups. It is shown that the C36 FF leads to better correlation with experimental data compared to the CHARMM22/CMAP FF and suggest using C36 in protein simulations. Although both CHARMM FFs contains the same nonbond parameters, our results show how the changes in the internal parameters associated with the peptide backbone via CMAP and the χ₁ and χ₂ dihedral parameters leads to improved treatment of the analyzed nonbond interactions. This highlights the importance of proper treatment of the internal covalent components in modeling nonbond interactions with molecular mechanics FFs. © 2013 Wiley Periodicals, Inc.     

Residual-Chemical-Shift-Anisotropy-Based Enantiodifferentiation in Lyotropic Liquid Crystalline Phases Based on Helically Chiral Polyacetylenes

Anisotropic NMR spectroscopy, revealing residual dipolar couplings (RDCs) and residual chemical shift anisotropies (RCSAs) has emerged as a powerful tool to determine the configurations of synthetic and complex natural compounds. The deduction of the absolute in addition to the relative configuration is one of the primary goals in the field. Therefore, the investigation of the enantiodiscriminating capabilities of chiral alignment media becomes essential. While RDCs and RCSAs are now used for the determination of the relative configuration routinely, RCSAs have not been measured in chiral alignment media such as chiral liquid crystals. Herein, we present this application by measuring RCSAs for chiral analytes such as indanol and isopinocampheol in the lyotropic liquid crystalline phase of an L-valine derived helically chiral polyacetylenes. We have also demonstrated that a single 1D ¹³C−{¹H} NMR spectrum suffices to get the RCSAs circumventing the necessity to acquire two spectra at two alignment conditions.     

NMR Crystallography: Evaluation of Hydrogen Positions in Hydromagnesite by 13C{1H} REDOR Solid‐State NMR and Density Functional Theory Calculation of Chemical Shielding Tensors

Solid‐state NMR measurements coupled with density functional theory (DFT) calculations demonstrate how hydrogen positions can be refined in a crystalline system. The precision afforded by rotational‐echo double‐resonance (REDOR) NMR to interrogate ¹³C–¹H distances is exploited along with DFT determinations of the ¹³C tensor of carbonates (CO₃^2?). Nearby ¹H nuclei perturb the axial symmetry of the carbonate sites in the hydrated carbonate mineral, hydromagnesite [4 MgCO₃?Mg(OH)₂?4 H₂O]. A match between the calculated structure and solid‐state NMR was found by testing multiple semi‐local and dispersion‐corrected DFT functionals and applying them to optimize atom positions, starting from X‐ray diffraction (XRD)‐determined atomic coordinates. This was validated by comparing calculated to experimental ¹³C{¹H} REDOR and ¹³C chemical shift anisotropy (CSA) tensor values. The results show that the combination of solid‐state NMR, XRD, and DFT can improve structure refinement for hydrated materials.     

Mechanism Studies of the Conversion of 13C‐Labeled n‐Butane on Zeolite H‐ZSM‐5 by Using 13C Magic Angle Spinning NMR Spectroscopy and GC–MS Analysis

By using ¹³C MAS NMR spectroscopy (MAS=magic angle spinning), the conversion of selectively ¹³C‐labeled n‐butane on zeolite H‐ZSM‐5 at 430–470 K has been demonstrated to proceed through two pathways: 1) scrambling of the selective ¹³C‐label in the n‐butane molecule, and 2) oligomerization–cracking and conjunct polymerization. The latter processes (2) produce isobutane and propane simultaneously with alkyl‐substituted cyclopentenyl cations and condensed aromatic compounds. In situ ¹³C MAS NMR and complementary ex situ GC–MS data provided evidence for a monomolecular mechanism of the ¹³C‐label scrambling, whereas both isobutane and propane are formed through intermolecular pathways. According to ¹³C MAS NMR kinetic measurements, both pathways proceed with nearly the same activation energies (E_a=75 kJ mol^?1 for the scrambling and 71 kJ mol^?1 for isobutane and propane formation). This can be rationalized by considering the intermolecular hydride transfer between a primarily initiated carbenium ion and n‐butane as being the rate‐determining stage of the n‐butane conversion on zeolite H‐ZSM‐5.     

NMR “Crystallography” for Uniformly (13C, 15N)‐Labeled Oriented Membrane Proteins

In oriented‐sample (OS) solid‐state NMR of membrane proteins, the angular‐dependent dipolar couplings and chemical shifts provide a direct input for structure calculations. However, so far only ¹H–¹⁵N dipolar couplings and ¹⁵N chemical shifts have been routinely assessed in oriented ¹⁵N‐labeled samples. The main obstacle for extending this technique to membrane proteins of arbitrary topology has remained in the lack of additional experimental restraints. We have developed a new experimental triple‐resonance NMR technique, which was applied to uniformly doubly (¹⁵N, ¹³C)‐labeled Pf1 coat protein in magnetically aligned DMPC/DHPC bicelles. The previously inaccessible ¹H_α–¹³C_α dipolar couplings have been measured, which make it possible to determine the torsion angles between the peptide planes without assuming α‐helical structure a priori. The fitting of three angular restraints per peptide plane and filtering by Rosetta scoring functions has yielded a consensus α‐helical transmembrane structure for Pf1 protein.     

Evaluating the accuracy of theoretical one‐bond 13C─13C scalar couplings and their ability to predict structure in a natural product

This study explores the feasibility of using a combination of experimental and theoretical 1‐bond ¹³C─¹³C scalar couplings (¹J_CC) to establish structure in organic compounds, including unknowns. Historically, ⁿJ_CC and ⁿJ_CH studies have emphasized 2 and 3‐bond couplings, yet ¹J_CC couplings exhibit significantly larger variations. Moreover, recent improvements in experimental measurement and data processing methods have made ¹J_CC data more available. Herein, an approach is evaluated in which a collection of theoretical structures is created from a partial nuclear magnetic resonance structural characterization. Computed ¹J_CC values are compared to experimental data to identify candidates giving the best agreement. This process requires knowledge of the error in theoretical methods, thus the B3LYP, B3PW91, and PBE0 functionals are evaluated by comparing to 27 experimental values from INADEQUATE. Respective errors of ±1.2, ±3.8, and ±2.3 Hz are observed. An initial test of this methodology involves the natural product 5‐methylmellein. In this case, only a single candidate matches experimental data with high statistical confidence. This analysis establishes the intramolecular hydrogen‐bonding arrangement, ring heteroatom identity, and conformation at one position. This approach is then extended to hydroheptelidic acid, a natural product not fully characterized in prior studies. The experimental/theoretical approach proposed herein identifies a single best‐fit structure from among 26 candidates and establishes, for the first time, 1 configuration and 3 conformations to complete the characterization. These results suggest that accurate and complete structural characterizations of many moderately sized organic structures (<800 Da) may be possible using only ¹J_CC data.     

hNCOcanH pulse sequence and a robust protocol for rapid and unambiguous assignment of backbone (1HN, 15N and 13C′) resonances in 15 N/13C‐labeled proteins

A three‐dimensional nuclear magnetic resonance (NMR) pulse sequence named as hNCOcanH has been described to aid rapid sequential assignment of backbone resonances in ¹⁵N/¹³C‐labeled proteins. The experiment has been derived by a simple modification of the previously described HN(C)N pulse sequence [Panchal et al., J. Biomol. NMR 20 (2001) 135–147]; t₂ evolution is used to frequency label ¹³C′ rather than ¹⁵N (similar trick has also been used in the design of hNCAnH pulse sequence from hNcaNH [Frueh et al., JACS, 131 (2009) 12880–12881]). The modification results in a spectrum equivalent to HNCO, but in addition to inter‐residue correlation peaks (i.e. H_i, C_i?1), the spectrum also contains additional intra‐residue correlation peaks (i.e. H_i?1, C_i?1) in the direct proton dimension which has maximum resolution. This is the main strength of the experiment and thus, even a small difference in amide ¹H chemical shifts (5–6 Hz) can be used for establishing a sequential connectivity. This experiment in combination with the HNN experiment described previously [Panchal et al., J. Biomol. NMR 20 (2001) 135–147] leads to a more robust assignment protocol for backbone resonances (¹H^N, ¹⁵N) than could be derived from the combination of HNN and HN(C)N experiments [Bhavesh et al., Biochemistry, 40 (2001) 14727–14735]. Further, this new protocol enables assignment of ¹³C′ resonances as well. We believe that the experiment and the protocol presented here will be of immense value for structural—and functional—proteomics research by NMR. Performance of this experiment has been demonstrated using ¹³C/¹⁵N labeled ubiquitin. Copyright © 2011 John Wiley & Sons, Ltd.     

RDC‐enhanced structure calculation of a β‐heptapeptide in methanol

Residual dipolar couplings (RDCs) are a rich source of structural information that goes beyond the range covered by the nuclear Overhauser effect or scalar coupling constants. They can only be measured in partially oriented samples. RDC studies of peptides in organic solvents have so far been focused on samples in chloroform or DMSO. Here, we show that stretched poly(vinyl acetate) can be used for the partial alignment of a linear β‐peptide with proteinogenic side chains in methanol. ¹D_CH, ¹D_NH, and ²D_HH RDCs were collected with this sample and included as restraints in a simulated annealing calculation. Incorporation of RDCs in the structure calculation process improves the long‐range definition in the backbone of the resulting 3₁₄‐helix and uncovers side‐chain mobility. Experimental side‐chain RDCs of the central leucine and valine residues are in good agreement with predicted values from a local three‐state model. Copyright © 2016 John Wiley & Sons, Ltd.     

Assignment of 1H and 13C NMR data for diethyl 2‐ and 8‐quinolylmethylphosphonates and their palladium(II) dihalide complexes

¹H and ¹³C NMR spectral data for diethyl 2‐ and 8‐quinolylmethylphosphonates (L) and their palladium(II) dihalide complexes, trans‐[PdL₂X₂] (L = 2‐dqmp, 8‐dqmp; X = Cl, Br), are presented. The NMR analysis was performed on the basis of one‐ and two‐dimensional homo‐ and heteronuclear experiments including ¹H, ¹³C, APT, ¹H–¹H COSY, ¹H–¹³C COSY, HMQC and HMBC techniques. Copyright © 2003 John Wiley & Sons, Ltd.     

“Direct” 13C Hyperpolarization of 13C‐Acetate by MicroTesla NMR Signal Amplification by Reversible Exchange (SABRE)

Herein, we demonstrate “direct” ¹³C hyperpolarization of ¹³C‐acetate via signal amplification by reversible exchange (SABRE). The standard SABRE homogeneous catalyst [Ir‐IMes; [IrCl(COD)(IMes)], (IMes=1,3‐bis(2,4,6‐trimethylphenyl), imidazole‐2‐ylidene; COD=cyclooctadiene)] was first activated in the presence of an auxiliary substrate (pyridine) in alcohol. Following addition of sodium 1‐¹³C‐acetate, parahydrogen bubbling within a microtesla magnetic field (i.e. under conditions of SABRE in shield enables alignment transfer to heteronuclei, SABRE‐SHEATH) resulted in positive enhancements of up to ≈100‐fold in the ¹³C NMR signal compared to thermal equilibrium at 9.4 T. The present results are consistent with a mechanism of “direct” transfer of spin order from parahydrogen to ¹³C spins of acetate weakly bound to the catalyst, under conditions of fast exchange with respect to the ¹³C acetate resonance, but we find that relaxation dynamics at microtesla fields alter the optimal matching from the traditional SABRE‐SHEATH picture. Further development of this approach could lead to new ways to rapidly, cheaply, and simply hyperpolarize a broad range of substrates (e.g. metabolites with carboxyl groups) for various applications, including biomedical NMR and MRI of cellular and in vivo metabolism.     

First 13C‐NMR Assignments of Betaxanthins

Due to their inherent liability towards highly acidic conditions previously considered to be a prerequisite for data acquisition, betaxanthin structure dereplication by NMR spectroscopy has been scarcely reported and was, hitherto, exclusively based on ¹H‐NMR data interpretation. Applying only slightly acidic conditions, we herein report the first ¹³C‐NMR data of two betaxanthins, i.e., indicaxanthin ( 1 ), isolated from yellow‐orange cactus pear fruits (Opuntia ficus‐indica [L.] Mill . cv. ‘Gialla’), and of miraxanthin V ( 2 ) from yellow Swiss chard petioles (Beta vulgaris L. ssp. cicla [L.] Alef . cv. ‘Bright Lights’), as derived by gHSQC‐ and gHMQC‐NMR experiments and inverse detection.     

The 1H NMR spectrum of pyrazole in a nematic phase

The experimental ¹H nuclear magnetic resonance (NMR) spectrum of 1H‐pyrazole was recorded in thermotropic nematic liquid crystal N‐(p‐ethoxybenzylidene)‐p‐butylaniline (EBBA) within the temperature range of 299–308 K. Two of three observable dipolar D_HH‐couplings appeared to be equal at each temperature because of fast prototropic tautomerism. Analysis of the Saupe orientational order parameters using fixed geometry determined by computations and experimental dipolar couplings results in a situation in which the molecular orientation relative to the magnetic field (and the liquid crystal director) can be described exceptionally by a single parameter. Copyright © 2016 John Wiley & Sons, Ltd.     

1H, 13C and 31P NMR spectral analysis of monoalkyl (α‐anilinobenzyl)phosphonates and their dipalladium(II) metallocyclic complexes

A ¹H, ¹³C and ³¹P NMR study of monoethyl (HL1) and monobutyl (HL2) esters of (α‐anilinobenzyl)phosphonic acid and their metallocyclic dipalladium complexes (Pd₂L₄,L = L1, L2) in DMSO‐d₆ was performed, based on 1D and 2D homo‐ and heteronuclear experiments including ¹H,¹³C,³¹P,APT,¹H–¹H COSY, ¹H–¹³C COSY, gs‐HMQC and gs‐HMBC NMR techniques. The results obtained are discussed with respect to those for some palladium(II) complexes reported for various anilinobenzylphosphonate derivatives. Copyright © 2002 John Wiley & Sons, Ltd.     

Unprecedented Reaction Pathway of Sterically Crowded Calcium Complexes: Sequential C−N Bond Cleavage Reactions Induced by C−H Bond Activations

Five bis(quinolylmethyl)‐(1H ‐indolylmethyl)amine (BQIA) compounds, that is, {(quinol‐8‐yl‐CH₂)₂NCH₂(3‐Br‐1H ‐indol‐2‐yl)} ( L¹H ) and {[(8‐R³‐quinol‐2‐yl)CH₂]₂NCH(R²)[3‐R¹‐1H ‐indol‐2‐yl]} ( L^2–5H ) ( L²H : R¹=Br, R²=H, R³=H; L³H : R¹=Br, R²=H, R³=i Pr; L⁴H : R¹=H, R²=CH₃, R³=i Pr; L⁵H : R¹=H, R²=n Bu, R³=i Pr) were synthesized and used to prepare calcium complexes. The reactions of L^1–5H with silylamido calcium precursors (Ca[N(SiMe₂R)₂]₂(THF)₂, R=Me or H) at room temperature gave heteroleptic products ( L^{1, 2} )CaN(SiMe₃)₂ ( 1 , 2 ), ( L^{3, 4} )CaN(SiHMe₂)₂ ( 3 a , 4 a ) and homoleptic complexes ( L^{3, 5} )₂Ca ( D3 , D5 ). NMR and X‐ray analyses proved that these calcium complexes were stabilized through Ca⋅⋅⋅C−Si, Ca⋅⋅⋅H−Si or Ca⋅⋅⋅H−C agostic interactions. Unexpectedly, calcium complexes (( L^3–5 )CaN(SiMe₃)₂) bearing more sterically encumbered ligands of the same type were extremely unstable and underwent C−N bond cleavage processes as a consequence of intramolecular C−H bond activation, leading to the exclusive formation of (E )‐1,2‐bis(8‐isopropylquinol‐2‐yl)ethane.     

A tridentate CNO‐donor palladium(II) complex as efficient catalyst for direct C―H arylation: Application in preparation of imidazole‐based push–pull chromophores

A series of imidazolium chlorides for the formation of tridentate CNO‐donor palladium(II) complexes featuring N‐heterocyclic carbene moieties have been developed from cheap and readily available starting materials with high yields. Their palladium complexes were prepared by reactions between the ligand precursors and PdCl₂ using K₂CO₃ as base in pyridine with reasonable yields. These air‐stable metal complexes were characterized using ¹H NMR and ¹³C{¹H} NMR spectroscopy and elemental analyses. Heteronuclear multiple bond correlation experiments were performed to identify key NMR signals of these compounds. The structures of two of the complexes were also established by single‐crystal X‐ray diffraction analysis. One of these complexes was successfully applied in the direct C―H functionalization reactions between heterocyclic compounds and aryl bromides, producing excellent yields of coupled products. The coupling reactions were scalable, allowing grams of coupled products to be obtained with a mere 2 mol% of Pd loading. The catalyst system developed allowed the large‐scale preparation of several push–pull chromophores straightforwardly. Photophysical properties based on UV–visible and fluorescence spectroscopy for these chromophores were investigated. Deep blue photoluminescence with moderate quantum efficiency and twisted intramolecular charge transfer excited state were observed for these chromophores. Density functional theory (DFT) and time‐dependent DFT calculations were performed to support the experimental results.