14N Solid‐State NMR Spectroscopy of Amino Acids

¹⁴N ultra‐wideline solid‐state NMR (SSNMR) spectra were obtained for 16 naturally occurring amino acids and four related derivatives by using the WURST–CPMG (wideband, uniform rate, and smooth truncation Carr–Purcell–Meiboom–Gill) pulse sequence and frequency‐stepped techniques. The ¹⁴N quadrupolar parameters were measured for the sp³ nitrogen moieties (quadrupolar coupling constant, C_Q, values ranged from 0.8 to 1.5 MHz). With the aid of plane‐wave DFT calculations of the ¹⁴N electric‐field gradient tensor parameters and orientations, the moieties were grouped into three categories according to the values of the quadrupolar asymmetry parameter, η_Q: low (≤0.3), intermediate (0.31–0.7), and high (≥0.71). For RNH₃⁺ moieties, greater variation in N?H bond lengths was observed for systems with intermediate η_Q values than for those with low η_Q values (this variation arose from different intermolecular hydrogen‐bonding arrangements). Strategies for increasing the efficiency of ¹⁴N SSNMR spectroscopy experiments were discussed, including the use of sample deuteration, high‐power ¹H decoupling, processing strategies, high magnetic fields, and broadband cross‐polarization (BRAIN‐CP). The temperature‐dependent rotations of the NH₃ groups and their influence on ¹⁴N transverse relaxation rates were examined. Finally, ¹⁴N SSNMR spectroscopy was used to differentiate two polymorphs of l ‐histidine through their quadrupolar parameters and transverse relaxation time constants. The strategies outlined herein permitted the rapid acquisition of directly detected ¹⁴N SSNMR spectra that to date was not matched by other proposed methods.     

Semi‐analytical description of the S = 9/2 quadrupole nutation NMR experiment: multinuclear application to 113In and 115In in indium phosphide

The density matrix of a spin S = 9/2 excited by a radiofrequency pulse is calculated. The interaction involved during the excitation of the spin system is first‐order quadrupolar. Consequently, the results are valid for any ratio of the quadrupolar coupling ω_Q to the pulse amplitude ω₁. The behavior of the central transition intensities versus the pulse length is discussed. The ¹¹⁵In and ¹¹³In nuclei in a powdered sample of indium phosphide (InP) are used to illustrate the results. It is found that the ratio of the quadrupolar coupling constants determined in this work is in excellent agreement with the ratio of the quadrupole moments of the two nuclei. Copyright © 2015 John Wiley & Sons, Ltd.     

Ultra‐wideline 27Al NMR Investigation of Three‐ and Five‐Coordinate Aluminum Environments

Ultra‐wideline ²⁷Al NMR experiments are conducted on coordination compounds with ²⁷Al nuclei possessing immense quadrupolar interactions that result from exceptionally nonspherical coordination environments. NMR spectra are acquired using a methodology involving frequency‐stepped, piecewise acquisition of NMR spectra with Hahn‐echo or quadrupolar Carr–Purcell Meiboom–Gill (QCPMG) pulse sequences, which is applicable to any half‐integer quadrupolar nucleus with extremely broad NMR powder patterns. Despite the large breadth of these central transition powder patterns, ranging from 250 to 700 kHz, the total experimental times are an order of magnitude less than previously reported experiments on analogous complexes with smaller quadrupolar interactions. The complexes examined feature three‐ or five‐coordinate aluminum sites: trismesitylaluminum (AlMes₃), tris(bis(trimethylsilyl)amino)aluminum (Al(NTMS₂)₃), bis[dimethyl tetrahydrofurfuryloxide aluminum] ([Me₂‐Al(μ‐OTHF)]₂), and bis[diethyl tetrahydrofurfuryloxide aluminum] ([Et₂‐Al(μ‐OTHF)]₂). We report some of the largest ²⁷Al quadrupolar coupling constants measured to date, with values of C_Q(²⁷Al) of 48.2(1), 36.3(1), 19.9(1), and 19.6(2) MHz for AlMes₃ , Al(NTMS₂)₃ , [Me₂‐Al(μ‐OTHF)]₂ , and [Et₂‐Al(μ‐OTHF)]₂ , respectively. X‐ray crystallographic data and theoretical (Hartree–Fock and DFT) calculations of ²⁷Al electric field gradient (EFG) tensors are utilized to examine the relationships between the quadrupolar interactions and molecular structure; in particular, the origin of the immense quadrupolar interaction in the three‐coordinate species is studied via analyses of molecular orbitals.     

2D-NMR strategy dedicated to the analysis of weakly ordered,fully deuterated enantiomers in chiral liquid crystals

Methods for the assignment of the quadrupolar doublets in the deuterium NMR spectra of weakly ordered, perdeuterated or partially deuterated enantiomers dissolved in chiral liquid crystals are described which use robust 2D correlation NMR experiments. To overcome a lack of resolution in deuterium tilted Q-COSY 2D spectra in such materials, we propose and explore a correlation 2D sequence which is based on deuterium-carbon 2D correlation spectroscopy. The technique results in a (13)C-(2)H contour plot and allows the full resonance assignment of overcrowded deuterium 1D spectra using carbon-deuterium correlations. The (2)H autocorrelation and (13)C-(2)H correlation experiments are applied in the case of a racemic mixture of 2-ethylhexanoic acid-d(15) dissolved in a polypeptidic chiral oriented solvent. The performance and the limits of both techniques are presented and discussed. For the last step of the assignment procedure, we propose a simple method for obtaining two coherent sets of quadrupolar splittings, one for each enantiomer.     

Studies on the Stereoselective Synthesis of Deuterated D‐Ribose Derivatives

In view of the importance of the site‐specific substitution of the H‐atom by its stable isotope ²H in a stereoselective/stereospecific manner at the pentose sugar residue, decreasing the spectral overcrowding in various regions of 1D and 2D homo‐ and heteronuclear correlation spectra of oligo‐DNA and ‐RNA, there is always a need for the development of new methods for the incorporation of ²H at different sites of a ribose. High‐yielding multistep syntheses of C(2)‐, and (5R)‐ and (5S)‐3,5‐deuterated ribose derivatives have been envisaged for the application of site‐specific incorporation of multilabeled nucleosides into oligomers to facilitate their structure elucidation by NMR spectroscopy. All syntheses started from D ‐glucose after proper derivatization. In the case of C(2), >97 atom‐% isotope was incorporated, employing an inversion of the configuration at C(2) as the key reaction. For C(5), two different routes were envisaged: on the one hand, deuterated achiral reagent was treated with a conformationally locked sugar moiety 15 , while, on the other, chiral protonated sources were used to transfer the H‐atom to a C(5)‐deuterated aldehyde 18 . In all cases, enantiomeric and isotopic purities were found to be as high as >97% as determined by NMR spectroscopy.     

Selektive Verbreiterung von 1H-NMR Restsignalen deuterierter Lösungsmittel

¹H {²H} off-resonance noise decoupling experiments with incompletely deuterated compounds result in a selective broadening of those residual proton signals which are split by H/D couplings. By this technique hidden resonance lines which are overlapped by solvent signals can be easily detected, as is demonstrated by the analysis of acetone/acetone-d₆ mixtures. Typical experimental details are given.     

2D NMR characterization of poly(ethylene-co-5-vinyl-2-norbornene)

Unsaturated poly(ethylene-co-5-vinyl-2-norbornene) was synthetized using the [Ph₂C(Flu)(Cp)]ZrCl₂ metallocene/methylaluminoxane (MAO) catalyst system. ¹H and ¹³C NMR spectra of the copolymer were assigned by means of DEPT, homonuclear 2D ¹H-¹H COSY, and heteronuclear 2D ¹H-¹³C correlation NMR experiments. The used catalyst system produces mainly isolated 5-vinyl-2-norbornene (VNB) sequences. VNB is incorporated selectively via the cyclic double bond. The unreacted double bond of the copolymer exists in the 5-endo: 5-exo positions (3 : 1). Both isomers of VNB are polymerized with the same propability.     

Keeping PASE with WEFT: SHWEFT–PASE pulse sequences for 1H NMR spectra of highly paramagnetic molecules

Metalloproteins are a category of biomolecules in which the metal site is usually the locus of activity or function. In many cases, the metal ions are paramagnetic or have accessible paramagnetic states, many of which can be studied using NMR spectroscopy. Extracting useful information from ¹H NMR spectra of highly paramagnetic proteins can be difficult because the paramagnetism leads to large resonance shifts (~400 ppm), extremely broad lines, extreme baseline nonlinearity, and peak shape distortion. It is demonstrated that employing polychromatic and adiabatic shaped pulses in simple pulse sequences, then combining existing sequences, leads to significant spectral improvement for highly paramagnetic proteins. These sequences employ existing technology, with available hardware, and are of short duration to accommodate short nuclear T₁ and T₂. They are shown to display uniform excitation over large spectral widths (~75 kHz), accommodate high repetition rates, produce flat baselines over 75 kHz while maintaining peak shape fidelity, and can be used to reduce spectral dynamic range. High‐spin (S = 5/2) metmyoglobin, a prototypical highly paramagnetic protein, was used as the test molecule. The resulting one‐dimensional (1D) pulse sequences combine shaped pulses with super‐water elimination Fourier transform, which can be further combined with paramagnetic spectroscopy to give shaped pulses with super‐water elimination Fourier transform–paramagnetic spectroscopy. These sequences require, at most, direct current offset correction and minimal phasing. The performance of these sequences in simple ¹H 1D, 1D NOE, and two‐dimensional NOESY experiments is demonstrated for metmyoglobin and Paracoccus denitrificans Co²⁺‐amicyanin (S = 3/2), and employed to make new heme hyperfine resonance assignments for high‐spin metBjFixLH_151–256, the heme sensing domain of Bradyrhizobium japonicum FixL. Copyright © 2013 John Wiley & Sons, Ltd.     

Single‐Scan Multidimensional NMR Analysis of Mixtures at Sub‐Millimolar Concentrations by using SABRE Hyperpolarization

Signal amplification by reversible exchange (SABRE) is a promising method to increase the sensitivity of nuclear magnetic resonance (NMR) experiments. However, SABRE‐enhanced ¹H NMR signals are short lived, and SABRE is often used to record 1D NMR spectra only. When the sample of interest is a complex mixture, this results in severe overlaps for ¹H spectra. In addition, the use of a co‐substrate, whose signals may obscure the ¹H spectra, is currently the most efficient way to lower the detection limit of SABRE experiments. Here, we describe an approach to obtain clean, SABRE‐hyperpolarized 2D ¹H NMR spectra of mixtures of small molecules at sub‐millimolar concentrations in a single scan. The method relies on the use of para‐hydrogen together with a deuterated co‐substrate for hyperpolarization and ultrafast 2D NMR for acquisition. It is applicable to all substrates that can be polarized with SABRE.     

The structure and conformation of a mesogenic compound between almost zero and almost complete orientational order

The conformational distributions in molecules that form liquid crystalline phases are predicted to depend strongly on orientational order. Results are presented here to test this hypothesis. The mesogen 4‐hexyloxy‐4′‐cyanobiphenyl (6OCB) has been studied by NMR spectroscopy in the isotropic phase and in the nematic phase. In the isotropic phase the field‐induced orientational ordering produces small dipolar couplings between ¹³C and ¹H nuclei, which were determined from the ¹³C spectra. Couplings between ¹H nuclei were also obtained using 2D selective refocusing experiments. In the nematic phase, both ¹H–¹H dipolar couplings and quadrupolar splittings for deuterium nuclei were measured for partially‐deuterated samples. Both proton and deuterium spectra were also obtained for 6OCB in an equimolar mixture with 4‐(ethoxybenzylidene)‐4′‐butylaniline (EBBA). This mixture exhibits SmA and SmB phases. The data obtained from these experiments has been analysed to yield the probability distribution of the conformations in this molecule generated by rotations about bonds. It is found that there is a substantial influence of the orientational order of the molecules on these distributions.     

A composite pulse sequence for the broadband excitation in overtone N NMR spectroscopy

In this Letter, a composite π/2 pulse sequence,  135, for the broadband excitation in overtone NMR spectroscopy of spin S=1 quadrupolar nuclei is proposed. The performances of single and composite π/2 pulses against resonance offset and rf field inhomogeneity are compared based on the results from ¹⁴N overtone experiments on a single crystal sample of N-acetyl-D,L-valine. The results reveal that the  135 composite π/2 pulse is less sensitive to the pulse imperfections and it will be useful in the design of multidimensional overtone NMR experiments.     

Selective J‐resolved‐HMQC‐1 and ‐2, new methods for measuring proton–proton coupling constants in strongly coupled spin systems

Efficient pulse sequences for measuring ¹H–¹H coupling constants (J_HH) in strongly coupled spin systems, named selective J‐resolved‐HMQC‐1 and ‐2, have been developed. In the strongly coupled spin systems such as ‐CH₂‐CH_A(OH)‐CH_B(OH)‐CH₂‐, measurements of ³J_HAHB are generally difficult owing to the complicated splitting caused by the adjacent CH₂ protons. For easier and accurate measurements of ³J_HAHB in such a spin system, a selective excitation pulse is incorporated into the J‐resolved HMQC pulse sequence. In the proposed methods, only two strongly coupled protons, H_A and H_B which are excited by a selective pulse, are observed as J‐resolved HMQC signals. The cross peaks of H_A and H_B appear as doublets owing to ³J_HAHB along the F₁ dimension in the selective J‐resolved HMQC‐1 and ‐2 experiments. The efficiency of the proposed pulse sequences has been demonstrated in application to the stereochemical studies of the complicated natural product, monazomycin. Copyright © 2011 John Wiley & Sons, Ltd.     

Conformation of 4-ethoxy-4′-biphenylyl cyanide in the isotropic and anisotropic liquid-crystalline state — a rotational isomeric state simulation of 2H NMR spectra

The conformation of the ethoxy tail flanking the cyanobiphenyl core has been studied in the liquid-crystalline phase as well as in the isotropic solution. In solution, the vicinal coupling constant ³J_CH for the moiety C^phO CH (C^ph: phenyl carbon) provided the information regarding the rotation around the O C bond. Phenetole was mostly used as a model compound. The conventional rotational isomeric state (RIS) analysis yielded the energy difference between the gauche and trans state to be of the order of 1,6 to 1,4 kcal/mol in CDCl₃, C₆D₆ and dimethyl-d₆ sulfoxide. In this treatment, the rotational state around the neighboring bond (C^ph O) was taken to occur at 0 or π. The deuterated compound, 4-ethoxy-4′-biphenylyl-d₇ cyanide, was prepared and studied in the liquid-crystalline state by ²H NMR. The orientational order parameters of the mesogenic core were determined by the analysis of dipolar and quadrupolar splitting data due to aromatic deuterons. The quadrupolar splitting data were also obtained for the deuterated ethoxy tail. The conformation of the tail was elucidated according to the RIS simulation scheme previously proposed. The fraction of the trans conformation f_t was estimated to be 0,79 around the nematic-isotropic transition temperature (T_NI = 91,9°C). It was suggested that the conformation of the ethoxy tail remains nearly unaffected by the phase transition in the immediate vicinity of T_NI. The fraction f_t increases moderately toward an asymptotic value as temperature decreases.     

A novel C2 symmetric chiral stationary phase with N‐[(4‐Methylphenyl)sulfonyl]‐l‐leucine as chiral side chains

In this study, a series of chiral stationary phases based on N‐[(4‐methylphenyl)sulfonyl]‐l ‐leucine amide, whose enantiorecognition property has never been studied, were synthesized. Their enantioseparation abilities were chromatographically evaluated by 67 enantiomers. The chiral stationary phase derived from N‐[(4‐methylphenyl)sulfonyl]‐l ‐leucine showed much better enantioselectivities than that based on N‐(4‐methylbenzoyl)‐l ‐leucine amide. The construction of C₂ symmetric chiral structure greatly improved the enantiorecognition performance of the stationary phase. The C₂ symmetric chiral stationary phase exhibited superior enantioresolutions to other chiral stationary phases for most of the chiral analytes, especially for the chiral analytes with C₂ symmetric structures. By comparing the enantioseparations of the enantiomers with similar structures, the importance of hydrogen bond interaction, π–π interaction, and steric hindrance on enantiorecognition was elucidated. The enantiorecognition mechanism of trans‐N,N′‐(1,2‐diphenyl‐1,2‐ethanediyl)bis‐acetamide, which had an excellent separation factor on the C₂ symmetric chiral stationary phase, was investigated by ¹H‐NMR spectroscopy and 2D ¹H‐¹H nuclear overhauser enhancement spectroscopy.     

Double quantum modulation NMR spectroscopy on spins with I = 1 in solids

NMR experiments with amplitude modulated rf pulses are introduced for the detection of double quantum coherences from spin I = 1 nuclei with large quadrupolar frequencies v_Q in solids. rf pulses of intensity v₁ and with modulation frequency v_m create effective irradiation fields of intensity $\text{1}\text{4}$v²₁/(v_Q-v_m) on the double quantum transition when v₁<¦v_Q-v_m¦?v_Q+v_m. ²D-NMR measurements on an oriented single crystal of deuterated malonic acid are presented.     

Enantiomeric Separations in Capillary Electrochromatography with Crown Ether‐Capped β‐Cyclodextrin‐Bonded Silica Particles as Chiral Stationary Phase

Two novel types of crown ether capped β‐cyclodextrin (β‐CD) bonded silica, namely, 4′‐aminobenzo‐X‐crown‐Y (X=15, 18 and Y=5, 6, resp.) capped [3‐(2‐O‐β‐cyclodextrin)‐2‐hydroxypropoxy] propylsilyl‐appended silica, have been prepared and used as stationary phases in capillary electrochromatography (CEC) to separate chiral compounds. The two stationary phases have a chiral selector with two recognition sites: crown ether and β‐CD. They exhibit excellent enantioselectivity in CEC for a wide range of compounds. After inclusion of metal ions (Na⁺ or K⁺) from the running buffer into the crown ether units, the stationary phases become positively charged and can provide extra electrostatic interaction with ionizable solutes and enhance the dipolar interaction with polar neutral solutes. This enhances the host‐guest interaction with the solute and improves chiral recognition and enantioselectivity. Due to the cooperation of the anchored β‐CD and the crown ether, this kind of crown ether capped β‐CD bonded phase shows better enantioselectivity than either β‐CD‐ or crown ether bonded phases only. These new types of stationary phases have good potential for fast chiral separation with CEC.     

Separation of catechin epimers by complexation using ion mobility mass spectrometry

Ion mobility coupled with mass spectrometry provides a fast and repeatable method to separate catechin epimers by previous complexation with selected chiral modifiers and transition metals. Several combinations with chiral ligands such as D‐ and L‐amino acids and/or additional metal cations, chiral crown ethers, tartaric acid and heptakis(2,6‐di‐O‐methyl)‐β‐cyclodextrin were screened for their ability to affect the separation efficiency. The clusters having the form of [2M + D‐amino acid + Cu²⁺ ? 3H]^? (M stands for (?)‐epicatechin or (+)‐catechin) showed improvement in stereodifferentiation between two epimeric catechins in comparison to the analysis of pure epimers, where no separation was observed or the separation was hampered by the formation of mixed dimer complexes. Among various examined D‐amino acids only those possessing hydrophobic side chains induced the improvement of separation efficiency. The best peak‐to‐peak resolution (R_p–p) was determined to be 0.71 for [2M + D‐Leucine + Cu²⁺ ? 3H]^? clusters. Copyright © 2015 John Wiley & Sons, Ltd.     

Development of a CE‐MS2 method for the enantiomeric separation of L/D‐carnitine: Application to the analysis of infant formulas

A new chiral analytical method based on CE‐MS is proposed for the identification and simultaneous quantification of D /L ‐carnitine in infant formulas. Previous derivatization of carnitine with FMOC enabled the optimization of the chiral separation using CE with UV detection. An optimization of electrospray‐MS parameters using a partial filling of the non‐volatile chiral selector (succinyl‐γ‐CD) was performed. A selective fragmentation using MS² experiments with an ion trap analyser was carried out to confirm the identity of D /L ‐carnitine according to the current legislation. Satisfactory results were obtained in terms of linearity, precision, and accuracy. Interestingly, the CE‐MS² method developed allowed a sensitivity enhancement with respect to UV detection of 100‐fold, obtaining an LOD of 100 ng/g for D ‐carnitine. The determination of L ‐carnitine and its enantiomeric purity in 14 infant formulas supplemented with carnitine was successfully achieved, sample preparation only requiring an ultrafiltration with centrifugal filter devices to retain the components with the highest molecular weights.     

Change of Quadrupole Moment upon Excitation and Symmetry Breaking in Multibranched Donor-Acceptor Dyes

Upon photoexcitation, a majority of quadrupolar dyes, developed for large two-photon absorption, undergo excited-state symmetry breaking (ES-SB) and behave as dipolar molecules. We investigate how the change of quadrupole moment upon S₁←S₀ excitation, ΔQ, influences the propensity of a dye to undergo ES-SB using a series of molecules with a A -π- D -π- A motif where D is the exceptionally electron-rich pyrrolo[3,2-b]pyrrole and A are accepting groups. Tuning of ΔQ is achieved by appending a secondary acceptor group, A’ , on both sides of the D core and ES-SB is monitored using a combination of time-resolved IR and broadband fluorescence spectroscopy. The results reveal a clear correlation between ΔQ and the tendency to undergo ES-SB. When A is a stronger acceptor than A’ , ES-SB occurs already in non-dipolar but quadrupolar solvents. When A and A’ are identical, ES-SB is only partial even in highly dipolar solvents. When A is a weaker acceptor than A’ , the orientation of ΔQ changes, ES-SB is observed in dipolar solvents only and involves major redistribution of the excitation over the D -π- A and D-A’ branches of the dye.     

Hydrogen bonds and local symmetry in the crystal structure of gibbsite

First‐principles quantum mechanical calculations of NMR chemical shifts and quadrupolar parameters have been carried out to assign the ²⁷Al MAS NMR resonances in gibbsite. The ²⁷Al NMR spectrum shows two signals for octahedral aluminum revealing two aluminum sites coordinated by six hydroxyl groups each, although the crystallographic positions of the two Al sites show little difference. The presence of two distinguished ²⁷Al NMR resonances characterized by rather similar chemical shifts but quadrupolar coupling constants differing by roughly a factor of two is explained by different character of the hydrogen bonds, in which the hydroxyls forming the corresponding octahedron around each aluminum site, are involved. The Al‐I site characterized by a C_Q = 4.6 MHz is surrounded by OH? groups participating in four intralayer and two interlayer hydrogen bonds, while the Al‐II site with the smaller quadrupolar constant (2.2 MHz) is coordinated by hydroxides, of which two point toward the intralayer cavities and four OH‐bonds are aligned toward the interlayer gallery. In high‐resolution solid‐state ¹H CRAMPS (combination of rotation and multiple‐pulse spectroscopy) four signals with an intensity ratio of 1:2:2:1 are resolved which allow to distinguish six nonequivalent hydrogen sites reported in the gibbsite crystal structure and to ascribe them to two types of structural OH groups associated with intralayer and interlayer hydrogen bonds. This study can be applied to characterize the gibbsite‐like layer—intergallery interactions associated with hydrogen bonding in the more complex systems, such as synthetic aluminum layered double hydroxides. Copyright © 2010 John Wiley & Sons, Ltd.