Stationary interphases with extended alkyl chains: A comparative study on chain order by solid-state NMR spectroscopy

Stationary interphases with long n-alkyl chains (n = 18, 22, 30, 34) have been examined by solid-state NMR spectroscopy. The determination of the silane functionality and the degree of cross-linking of silane ligands on the silica surface was performed by ²⁹Si CP/MAS NMR spectroscopy. High-speed ¹H MAS and ¹³C CP/MAS NMR spectroscopy were utilized to assess alkyl chain order and mobility of the different bonded phases. For this purpose, ¹H NMR line widths and ¹³C chemical shifts have been evaluated. It is shown that stationary phase order and rigidity increase with alkyl chain length. In addition, the temperature-dependent trans/gauche conformational change occurs at higher temperatures for a polymeric C₃₄ phase compared with a C₃₀ sorbent. This behaviour is discussed in the context of previously reported Chromatographic (HPLC) shape selectivity differences.     

Optimal control in NMR spectroscopy: Numerical implementation in SIMPSON

We present the implementation of optimal control into the open source simulation package SIMPSON for development and optimization of nuclear magnetic resonance experiments for a wide range of applications, including liquid- and solid-state NMR, magnetic resonance imaging, quantum computation, and combinations between NMR and other spectroscopies. Optimal control enables efficient optimization of NMR experiments in terms of amplitudes, phases, offsets etc. for hundreds-to-thousands of pulses to fully exploit the experimentally available high degree of freedom in pulse sequences to combat variations/limitations in experimental or spin system parameters or design experiments with specific properties typically not covered as easily by standard design procedures. This facilitates straightforward optimization of experiments under consideration of rf and static field inhomogeneities, limitations in available or desired rf field strengths (e.g., for reduction of sample heating), spread in resonance offsets or coupling parameters, variations in spin systems etc. to meet the actual experimental conditions as close as possible. The paper provides a brief account on the relevant theory and in particular the computational interface relevant for optimization of state-to-state transfer (on the density operator level) and the effective Hamiltonian on the level of propagators along with several representative examples within liquid- and solid-state NMR spectroscopy.     

Advances in solid-state NMR of membrane proteins

Solid-state nuclear magnetic resonance (SSNMR) is an NMR spectroscopy applied to condensed-phase systems, including membrane proteins. Membrane protein fold and function are dependent upon interactions with surrounding bilayer components. Structural and functional analyses are thus challenging, and new approaches are needed to better characterise these systems. SSNMR is uniquely suited to the examination of membrane proteins in native environments, and has the capabilities to elucidate complex protein mechanisms and structures. Notable research implementing SSNMR is aimed at developing new strategies and technology to efficiently target membrane proteins within synthetic and biological membranes. Significant advances have been made: observation of protein function in native environments, emergence of in situ methods to examine integral proteins within natural membranes, sensitivity enhancement techniques and cutting-edge structure determination methods. We present how these advances are applied to answer outstanding questions in structural biology. Experiments have shown consistent results for protein investigations in biological membranes and synthetic lipid compositions, indicating that SSNMR is an innovative and direct approach for the study of these systems.     

Sensitivity and resolution in proton solid-state NMR at intermediate deuteration levels: quantitative linewidth characterization and applications to correlation spectroscopy

We present a systematic study of proton linewidths in rigid solids as a function of sample spinning frequency and proton density, with the latter controlled by the ratio of protonated and perdeuterated model compounds. We find that the linewidth correlates more closely with the overall proton density (rho(H)) than the size of local clusters of (1)H spins. At relatively high magic-angle spinning (MAS) rates, the linewidth depends linearly upon the inverse MAS rate. In the limit of infinite spinning rate and/or zero proton concentration, the linewidth extrapolates to a non-zero value, owing to contributions from scalar couplings, chemical shift dispersion, and B(0) field inhomogeneity. The slope of this line depends on the overall concentration of unexchangeable protons in the sample and the spinning rate. At up to 30% protonation levels ( approximately 2 (1)H/100A(3)), proton detection experiments are demonstrated to have a substantial (2- to 3-fold) sensitivity gain over corresponding (13)C-detected experiments. Within this range, the absolute sensitivity increases with protonation level; the optimal compromise between sensitivity and resolution is in the range of 20-30% protonation. We illustrate the use of dilute protons for polarization transfer to and from low-gamma spins within 5A, and to be utilized as both magnetization source and detection spins. The intermediate protonation regime enhances relaxation properties, which we expect will enable new types of (1)H correlation pulse sequences to be implemented with improved resolution and sensitivity.     

Studies on the structures and interactions of glutathione in aqueous solution by molecular dynamics simulations and NMR spectroscopy

All-atom molecular simulations and temperature-dependent NMR have been used to investigate the conformations and hydrogen bonds of glutathione (GSH) in aqueous solution. The simulations start from three different initial conformations. The properties are characterized by intramolecular distances, radius of gyration, root-mean-square deviation, and solvent-accessible surface. GSH is highly flexible in aqueous solutions in the simulations. Moreover, conformations can covert between “extended” and “folded” states. Interestingly, the two different hydrogen atoms in cysteine (H_N2) and glycin (H_N3) show different capabilities in forming NH?OW hydrogen bonds. The temperature-dependent NMR results of the different amide hydrogen atoms also show agreements with the MD simulations. Competing formation of GSH hydrogen-bonding interactions in aqueous solutions leads to hydrogen-bonding networks and the distribution of conformations. These changes will affect the activity of GSH under physiological conditions.     

Rotor-synchronized dipolar-filter sequence at fast MAS in solid-state NMR

Dipolar filters are of considerable importance for eliminating the ¹H NMR signal of the rigid components of heterogeneous compounds while selecting the signal of their mobile parts. On the basis of such filters, structural and dynamical information of these compounds can often be acquired through further manipulations (e.g. spin diffusion) on the spin systems. To overcome the destructive interferences between the magic angle spinning (MAS) speed and the cycle-time of the widely-used Rotor-Asynchronized Dipolar Filter (RADF) sequence, we introduce a new method called Rotor-Synchronized Dipolar Filter (RSDF). This communication shows that this sequence does not present any interference with the spinning speed and is more compatible than RADF with high MAS frequencies (ν_R > 12 kHz). This new pulse sequence will potentially contribute to future researches on heterogeneous materials, such as multiphase polymer and membrane systems.     

Correlating fast and slow chemical shift spinning sideband patterns in solid-state NMR

An experiment is presented that enables the measurement of small chemical shift anisotropy tensors under fast magic-angle spinning (MAS). The two-dimensional spectra obtained give a fast MAS sideband pattern in the directly observed dimension with the spinning sideband intensities equivalent to the chemical shift anisotropy scaled by a factor of N, or equivalently the sample spinning frequency scaled by 1/N, in the indirectly observed dimension. The scaling factor may be arbitrarily varied by changing the number and timings of the rotor synchronized pi-pulses used. Desirable features of the experiment include a fixed length pulse sequence and efficient sampling of the indirectly observed dimension. In addition, neither quadrature detection in the indirect dimension nor storage periods are required, consequently no signal intensity is discarded by the pulse sequence. The experiment is demonstrated using (31)P NMR of sodium phosphate and (13)C NMR of fumaric acid monoethyl ester for which a scaling factor of N=10.2 was employed.     

Interference of homonuclear decoupling and exchange in the solid-state NMR of perfluorocyclohexane

We observe an interference between RF irradiation used for homonuclear decoupling of 19F and conformational exchange in the 13C spectrum of perfluorocyclohexane. We show that these effects can be readily reproduced in simulation, and characterise their dependence on the various NMR and experimental parameters. Their application to observing exchange rates on the kHz timescale is evaluated with respect to T(1rho) measurements and the connections between the two approaches established. The implications for experiments that use homonuclear decoupling of 1H to resolve 1J(CH)couplings in the solid-state are also evaluated in detail.     

Intensity and mosaic spread analysis from PISEMA tensors in solid-state NMR

The solid-state NMR experiment PISEMA, is a technique for determining structures of proteins, especially membrane proteins, from oriented samples. One method for determining the structure is to find orientations of local molecular frames (peptide planes) with respect to the unit magnetic field direction, B0. This is done using equations that compute the coordinates of this vector in the frames. This requires an analysis of the PISEMA function and its degeneracies. As a measure of the sensitivity of peptide plane orientations to the data, we use these equations to derive a formula for the intensity function in the powder pattern. With this function and other measures, we investigate the effect of small changes in peptide plane orientations depending on the location of the resonances in the powder pattern spectrum. This gives us an indication of the change in lineshape due to mosaic spread and a way to interpret these in terms of an orientational error bar.     

Criteria to average out the chemical shift anisotropy in solid-state NMR when irradiated with BABA I,BABA II,and C7 radiofrequency pulse sequences

Floquet–Magnus expansion is used to study the effect of chemical shift anisotropy in solid-state NMR of rotating solids. The chemical shift interaction is irradiated with two types of radiofrequency pulse sequences: BABA and C7. The criteria for the chemical shift anisotropy to be averaged out in each rotor period are obtained.     

Resolution effects and scaling in numerical simulations of passive scalar mixing in turbulence

The effects of finite grid resolution on the statistics of small scales in direct numerical simulations of turbulent mixing of passive scalars are addressed in this paper. Simulations at up to 2048³ grid points with grid spacing Δx varied from about 2 to 1/2 Batchelor scales (η_B) show that most conclusions on Schmidt number (Sc) dependence from prior work at less stringent resolution remain qualitatively correct, although simulations at resolution Δx≈η_B are preferred and will give adequate results for many important quantities including the scalar dissipation intermittency exponent and structure functions at moderately high orders. For Sc≥1, since η_B=ηSc^−1/2 (where η is the Kolmogorov scale), the requirement Δx≈η_B is more stringent than the corresponding criterion Δx≈η for the velocity field, which is thus well resolved in simulations aimed at high Schmidt number mixing. A simple argument is given to help interpret the effects of Schmidt and Reynolds numbers on trends towards local isotropy and saturation of intermittency at high Schmidt number. The present results also provide evidence for a trend to isotropy at high Reynolds number with fixed Sc=1.0. This is a new observation apparently not detected in less well resolved simulations in the past, and will require further investigation in the future.     

Enhanced resolution in proton solid-state NMR with very-fast MAS experiments

We present a new smooth amplitude-modulated (SAM) method that allows to observe highly resolved ¹H spectra in solid-state NMR. The method, which works mainly at fast or ultra-fast MAS speed (ν_R > 25 kHz) is complementary to previous methods, such as DUMBO, FSLG/PMLG or symmetry-based sequences. The method is very robust and efficient and does not present line-shape distortions or fake peaks. The main limitation of the method is that it requires a modern console with fast electronics that must be able to define the cosine line-shape in a smooth way, without any transient. However, this limitation mainly occurs at ultra-fast MAS where the rotation period is very short.     

The influence of the molecular system on the performance of heteronuclear decoupling in solid-state NMR

The intensity of the carbon signal in a CPMAS experiment has been measured for two CH and three CH(2) moieties in four test molecules under different phase-modulated proton decoupling conditions and as a function of the spinning rate. The proton decoupling schemes investigated were the golden standard TPPM and three of the GTn family. Aim of this analysis was to better describe experimentally the impact and limitations of phase-modulated decoupling. Sizeable differences in the response to decoupling were observed in otherwise chemically identical molecular fragments, such as the CHCH(2) found in tyrosine, phenyl-succinic acid or 9-Anthrylmethyl-malonate, probably due to differences in spin-diffusion rates. In keeping with known facts, the efficiency of the decoupling was observed to decrease with the MAS rate, but with somewhat different trends for the tested systems.     

Sensitivity and resolution enhancement in solid-state NMR spectroscopy of bicelles

Magnetically aligned bicelles are becoming attractive model membranes to investigate the structure, dynamics, geometry, and interaction of membrane-associated peptides and proteins using solution- and solid-state NMR experiments. Recent studies have shown that bicelles are more suitable than mechanically aligned bilayers for multidimensional solid-state NMR experiments. In this work, we describe experimental aspects of the natural abundance (13)C and (14)N NMR spectroscopy of DMPC/DHPC bicelles. In particular, approaches to enhance the sensitivity and resolution and to quantify radio-frequency heating effects are presented. Sensitivity of (13)C detection using single pulse excitation, conventional cross-polarization (CP), ramp-CP, and NOE techniques are compared. Our results suggest that the proton decoupling efficiency of the FLOPSY pulse sequence is better than that of continuous wave decoupling, TPPM, SPINAL, and WALTZ sequences. A simple method of monitoring the water proton chemical shift is demonstrated for the measurement of sample temperature and calibration of the radio-frequency-induced heating in the sample. The possibility of using (14)N experiments on bicelles is also discussed.     

Recent progress in solid-state NMR studies of drugs confined within drug delivery systems

Recent progress in the application of solid-state NMR (SS NMR) spectroscopy in structural studies of active pharmaceutical ingredients (APIs) embedded in different drug carriers is detailed. This article is divided into sections. The first part reports short characterization of the nanoparticles and microparticles that can be used as drug delivery systems (DDSs). The second part shows the applicability of SS NMR to study non-steroidal anti-inflammatory drugs (NSAIDs). In this section, problems related to API–DDS interactions, morphology, local molecular dynamics, nature of inter- or intramolecular connections, and pore filling are reviewed for different drug carriers (e.g. mesoporous silica nanoparticles (MSNs), cyclodextrins, polymeric matrices and others). The third and fourth sections detail the recent applications of SS NMR for searching for antibiotics and anticancer drugs confined in zeolites, MSNs, amorphous calcium phosphate and other carriers.     

Use of SPAM and FAM pulses in high-resolution MAS NMR spectroscopy of quadrupolar nuclei

The merits of SPAM and FAM pulses for enhancing the conversion of triple- to single-quantum coherences in the two-dimensional MQMAS experiment are compared using (87)Rb (spin I=3/2) and (27)Al (I=5/2) NMR of crystalline and amorphous materials. Although SPAM pulses are more easily optimized, our experiments and simulations suggest that FAM pulses yield greater signal intensity in all cases. In conclusion, we argue that, as originally suggested, SPAM and FAM pulses are best implemented in phase-modulated whole-echo MQMAS experiments and that the use of SPAM pulses to record separate echo and antiecho data sets, which are then combined, generally yields lower signal-to-noise ratios.     

A Modified Alderman–Grant Coil makes possible an efficient cross-coil probe for high field solid-state NMR of lossy biological samples

The design, construction, and performance of a cross-coil double-resonance probe for solid-state NMR experiments on lossy biological samples at high magnetic fields are described. The outer coil is a Modified Alderman–Grant Coil (MAGC) tuned to the ¹H frequency. The inner coil consists of a multi-turn solenoid coil that produces a B₁ field orthogonal to that of the outer coil. This results in a compact nested cross-coil pair with the inner solenoid coil tuned to the low frequency detection channel. This design has several advantages over multiple-tuned solenoid coil probes, since RF heating from the ¹H channel is substantially reduced, it can be tuned for samples with a wide range of dielectric constants, and the simplified circuit design and high inductance inner coil provides excellent sensitivity. The utility of this probe is demonstrated on two electrically lossy samples of membrane proteins in phospholipid bilayers (bicelles) that are particularly difficult for conventional NMR probes. The 72-residue polypeptide embedding the transmembrane helices 3 and 4 of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) (residues 194–241) requires a high salt concentration in order to be successfully reconstituted in phospholipid bicelles. A second application is to paramagnetic relaxation enhancement applied to the membrane-bound form of Pf1 coat protein in phospholipid bicelles where the resistance to sample heating enables high duty cycle solid-state NMR experiments to be performed.