Variable Angle Sample Spinning Research Articles

Variable angle NMR spectroscopy and its application to the measurement of residual chemical shift anisotropy

The successful measurement of anisotropic NMR parameters like residual dipolar couplings (RDCs), residual quadrupolar couplings (RQCs), or residual chemical shift anisotropy (RCSA) involves the partial alignment of solute molecules in an alignment medium. To avoid any influence of the change of environment from the isotropic to the anisotropic sample, the measurement of both datasets with a single sample is highly desirable. Here, we introduce the scaling of alignment for mechanically stretched polymer gels by varying the angle of the director of alignment relative to the static magnetic field, which we call variable angle NMR spectroscopy (VA-NMR). The technique is closely related to variable angle sample spinning NMR spectroscopy (VASS-NMR) of liquid crystalline samples, but due to the mechanical fixation of the director of alignment no sample spinning is necessary. Also, in contrast to VASS-NMR, VA-NMR works for the full range of sample inclinations between 0° and 90°. Isotropic spectra are obtained at the magic angle. As a demonstration of the approach we measure 13C-RCSA values for strychnine in a stretched PDMS/CDCl 3 gel and show their usefulness for assignment purposes. In this context special care has been taken with respect to the exact calibration of chemical shift data, for which three approaches have been derived and tested.

Simplification of the 1H NMR spectra of enantiomers dissolved in chiral liquid crystals, combining variable angle sample spinning and selective refocusing experiments

This work presents a technique to simplify overcrowded proton spectra in chiral liquid crystal solvents using rotation of the sample near the magic angle, VASS, combined with homonuclear selective refocusing 2D NMR experiments, SERF. This methodology provides a powerful tool to visualise enantiomers out of unresolved proton spectra. A modified SERF sequence is presented where the resulting 2D spectrum can be phased to increase the resolution. Accurate enantiomeric excesses are determined that are not possible to measure on static samples. Two examples are presented.

Applications of Variable-angle Sample Spinning Experiments to the Measurement of Scaled Residual Dipolar Couplings and 15N CSA in Soluble Proteins

NMR spectra of ubiquitin in the presence of bicelles at a concentration of 32% w/v have been recorded at 700 MHz under sample spinning conditions at the magic angle (54.7 degrees ) and at an angle of 45.5 degrees . At the magic angle, the 1H-15N HSQC spectrum of ubiquitin in bicelles is virtually indistinguishable from the one recorded on the protein in solution. Spinning the sample at the magic angle creates an isotropic environment with no preferred bicelle orientations, thus allowing the determination of scalar coupling constants. For an angle of rotation of 45.5 degrees , the bicelles orient with their normal perpendicular to the spinning axis leading to the observation of strong residual dipolar couplings and chemical shift variations of the 15N resonances.

Measurement of scaled residual dipolar couplings in proteins using variable-angle sample spinning.

NMR spectra of ubiquitin in the presence of bicelles at a concentration of 25% w/v have been recorded under sample spinning conditions for different angles of rotation. For an axis of rotation equal to the magic angle, the (1)H/(15)N HSQC recorded without any (1)H decoupling in the indirect dimension corresponds to the classical spectrum obtained on a protein in an isotropic solution and allows the measurement of scalar J-couplings (1) J (NH). For an angle of rotation smaller than the magic angle, the bicelles orient with their normal perpendicular to the spinning axis, whereas for an angle of rotation greater than the magic angle the bicelles orient with their normal along the spinning axis. This bicelle alignment creates anisotropic conditions that give rise to the observation of residual dipolar couplings in ubiquitin. The magnitude of these dipolar couplings depends directly on the angle that the rotor makes with the main magnetic field. By changing this angle in a controlled manner, residual dipolar couplings can be either scaled up or down thus offering the possibility to study simultaneously a wide range of dipolar couplings in the same sample.

Molecular Orientation and Conformation of Phosphatidylinositides in Membrane Mimetics Using Variable Angle Sample Spinning (VASS) NMR

For many biological molecules, determining their geometry as they exist in a membrane environment is a crucial step in understanding their function. Variable angle sample spinning (VASS) NMR provides a new route to obtaining geometry information on membrane-associating molecules; it has been used here to scale and separate anisotropic contributions to phosphorus chemical shifts in NMR spectra of phosphatidylinositol phosphates. The procedure allows spectral assignment via correlation with isotropic chemical shifts and determination of a family of probable headgroup orientations via interpretation of anisotropic shift contributions. The molecules studied include phosphtidylinositol-4-phosphate (PI(4)P) and phosphatidylinositol 4,5-bisphosphate (PI(4,5)P 2). A membrane-like environment is provided by a dispersion of alkyl-poly(ethylene) glycols and n-alcohols that forms a field-orienting liquid crystal with a director that can be manipulated by varying the sample spinning axis. The experiments presented indicate that the variable angle sample spinning method will provide a direct approach for assignment and extraction of structural information from membrane-associating biomolecules labeled with a wider variety of NMR active isotopes.

NMR of bicelles: orientation and mosaic spread of the liquid-crystal director under sample rotation.

Model-membrane systems composed of liquid-crystalline bicellar phases can be uniaxially oriented with respect to a magnetic field, thereby facilitating structural and dynamics studies of membrane-associated proteins. Here we quantitatively characterize a method that allows the manipulation of the direction of this uniaxial orientation. Bicelles formed from DMPC/DHPC are examined by (31)P NMR under variable-angle sample-spinning (VAS) conditions, confirming that the orientation of the liquid-crystalline director can be influenced by sample spinning. The director is perpendicular to the rotation axis when Theta (the angle between the sample-spinning axis and the magnetic field direction) is smaller than the magic angle, and is parallel to the rotation axis when Theta is larger than the magic angle. The new (31)P NMR VAS data presented are considerably more sensitive to the orientation of the bicelle than earlier (2)H studies and the analysis of the sideband pattern allows the determination of the orientation of the liquid-crystal director and its variation over the sample, i.e., the mosaic spread. Under VAS, the mosaic spread is small if Theta deviates significantly from the magic angle but becomes very large at the magic angle.

NMR spectra of mixed liquid crystals of opposite diamagnetic susceptibility anisotropies near critical point under variable angle sample spinning

The deuterium NMR spectra of molecules oriented in mixtures of nematic liquid crystalline phases for which the macroscopic diamagnetic susceptibility anisotropy (Δ χ) is nearly zero reveal the dynamics of the director for different angles the spinning axis makes with the magnetic field and for different spinning speeds. It is observed that a switch over of the director orientation occurs as a function of the spinning speed and the angle the spinning axis makes with the magnetic field. The farther the angle between the spinning axis and the magnetic field from the magic angle, the higher the spinning speed required to effect such a switch over of the director orientation. The switch over takes place gradually as indicated by the appearance of the ‘powder pattern’ in the 2H NMR spectra.

Manipulation of the director in bicellar mesophases by sample spinning: a new tool for NMR spectroscopy.

It is shown that bicellar nematic liquid-crystalline phases can be oriented with the director (the normal to the bicellar plane) at an arbitrary angle to the applied magnetic field by sample rotation around one axis (variable-angle sample spinning) or around two axes successively (switched-angle spinning). This promises to open novel possibilities for NMR studies of bicelles and proteins incorporated into bicelles or dissolved in a solution containing bicelles, including the correlation of several orientations in a two-dimensional NMR experiment.

Sign determination of dipolar couplings in field-oriented bicelles by variable angle sample spinning (VASS).

Residual dipolar couplings are being increasingly used as structural constraints for NMR studies of biomolecules. A problem arises when dipolar coupling contributions are larger than scalar contributions for a given spin pair, as is commonly observed in solid state NMR studies, in that signs of dipolar couplings cannot easily be determined. Here the sign ambiguities of dipolar couplings in field-oriented bicelles are resolved by variable angle sample spinning (VASS) techniques. The director behavior of field-oriented bicelles (DMPC/DHPC, DMPC/CHAPSO) in VASS is studied by 31P NMR. A stable configuration occurs when the spinning angle is smaller than the magic angle, 54.7 degrees, and the director (or bicelle normal) of the disks is mainly distributed in a plane perpendicular to the rotation axis. Since the dipolar couplings depend on how the bicelles are oriented with respect to the magnetic field, it is shown that the dipolar interaction can be scaled to the same order as the J-coupling by moving the spinning axis from 0 degree toward 54.7 degrees. Thus the relative sign of dipolar and scalar couplings can be determined.

General Features of the X Part of an ABX Spin System in Isotropic and Liquid Crystalline Phases as Illustrated by the13C–{1H} Spectra of 2,2′-Difluorobiphenyl

The13C–{1H} NMR spectra of 2,2′-difluorobiphenyl dissolved in isotropic and liquid crystalline solvents have been obtained and analyzed. They are examples of the X part of an ABX spectrum. It is shown that the spectrum of the isotropic solution yieldsJAX,JBX,JAB, δAB, and δX, but only if all the transitions are detected, and that intensities as well as frequencies of the transitions are used in the analysis. It is demonstrated that for 2,2′-difluorobiphenyl this requires that for some of the carbons it is necessary to detect very weak transitions. For the spectra of liquid crystalline solutions of ABX systems it is shown that the dipolar couplingsDAX,DBX, andDABare obtained only if these couplings are in a certain sensitive range of relative values. The sensitive range can be adjusted by using variable angle sample spinning (VASS). It is demonstrated that VASS spectra taken near the magic angle can be used to obtain the absolute signs of the scalar couplings.

A Study of the Order Parameters of Two Liquid Crystals by One and Two Dimensional C-13 NMR

The orientational ordering of six liquid crystalline compounds in two homologous series, namely 4-n-alkoxybenzilidene-4′-n-alkylanilines (nO. m) and 4, 4′-di-n-alkyldiphenyldiacetylenes (PTTP), has been studied at different temperatures by 2D C-13 NMR with variable angle sample spinning. The carbon-proton dipolar coupling constants were obtained and the order parameters of the aromatic cores were calculated. The C-13 chemical shifts for each carbon nucleus in the aromatic cores were also measured as a function of temperature by 1D C-13 NMR. A linear correlation between the chemical shifts and the order parameters was observed for each homologous series. The results furnish a useful and convenient database for the evaluation of order parameters for these two series of liquid crystals from simple measurements of C-13 chemical shifts.

Analytical treatment of the variable-angle sample spinning nuclear magnetic resonance spectra in the presence of the second-order quadrupole interaction determination of the quadrupole parameters and the chemical shift tensor

The analytical expressions for the transition probability of the central transition of the variable-angle sample spinning nuclear magnetic resonance (VAS NMR) spectrum for non-integral spins in the presence of second-order quadrupole interactions are derived on the basis of the stationary perturbation theory. The modulating parameters of the VAS NMR spectrum due to the time-dependent part of the quadrupole energy are found to be different (asymmetry parameter η > 0.1) from those obtained from the average Hamiltonian theory. Using the conditions for saddle points and local maxima, it has been observed that there is a range of values for sample spinning angle β around 32, 44 and 72 ° to obtain the single peak; the range width can indicate the value of η. Analytical expressions for the single-peak positions corresponding to β around 32, 44 and 72 ° are derived in terms of quadrupole parameters and chemical shift tensor. Further, we have described an analytical method to find the principal values of the chemical shift tensor in addition to the quadrupole parameters, using VAS NMR. As an illustration, the quadrupole parameters are computed for Na 2MoO 4 and Na 2SO 4 and compared with the values available in the literature (computed from wide-line NMR and magic-angle sample spinning (MAS) NMR data using the position of peaks and shoulders). For Na 2MoO 4 the principal values of the chemical shift tensor are found to be + 17.5, − 12 and − 6.5 ppm. The present method is accurate and easy to apply as it involves only the positions of peaks and not of shoulders. Moreover, it is suitable for all values of the chemical shift tensor.

Order parameters of the mesogenic core in two nematic liquid crystals containing a four-unit link

Abstract A series of compounds containing a four-unit link, 4′-n-alkylphenyl-4-n-hexyloxycinnamates, has been synthesized, and their mesomorphic behaviour has been determined. The orientational ordering of two compounds in this series has been studied in detail by 2D carbon-13 nuclear magnetic resonance with variable angle sample spinning. The carbon-proton dipolar coupling constants were obtained and the order parameters were calculated for different molecular segments (two phenyl rings, the central link containing an ethylenic motif, and the methylene carbons of the alkoxy chain). The temperature dependence of the local order parameters of each fragment were determined using these data and C-13 chemical shifts which were measured as a function of temperature. In these non-rigid molecules, the ratio between the order parameters of the two phenyl rings was found to be temperature dependent. The ring bearing the double bond is more ordered at low temperatures compared to the ring bearing the short alkyl ch...

A study of ferroelectric liquid crystals by one and two dimensional nuclear magnetic resonance

Abstract The orientational ordering of three ferroelectric liquid crystals has been studied by 2D carbon-13 nuclear magnetic resonance with variable angle sample spinning. These liquid crystals are esters of α-chloro acids and 4-heptoxy-4′-hydroxybiphenyl, and have very large spontaneous polarization. Since the helical structures of the optically active compounds are retained at a magnetic field of 7.05 T, their racemates were studied. The 2D NMR technique used was separated local field (SLF) spectroscopy with an efficient proton-proton dipolar decoupling sequence BLEW-48 in the evolution period. The carbon-proton dipolar coupling constants were obtained and the order parameters for different molecular segments of the liquid crystal were calculated. The C-13 chemical shifts were measured as functions of temperature and the data were used to obtain the temperature dependence of order parameters. The relations between the order parameters, molecular structures, and phase behavior are discussed.

Coherent reduction of dipolar interactions in molecules dissolved in liquid-crystal solvents using a new multiple-pulse technique during acquisition

A new pulse sequence, MET 6, synchronized to the sampling of the magnetization is proposed with special application to molecules in liquid crystals. It acts coherently on the dipolar interactions to produce a coherent reduction of the couplings in a way which compares to the variable angle sample spinning technique. The theory is developed in terms of average Hamiltonian theory and a full simulation of the sequence is presented together with an experimental study of its effects on an A 2 spin system dissolved in a nematic solvent. An analysis of the propagation of pulse imperfections is also given. A theoretical and experimental comparison is made between METE and other common multiple-pulse techniques used to average dipolar interactions in solids.

Side‐band analysis in variable‐angle sample‐spinning (VASS) NMR for the central transition of quadrupolar nuclei (half‐integer spin)

AbstractVariable‐angle sample‐spinning (VASS) NMR was investigated for the case of limited rotation speed of the spinner. Side‐band intensities and line‐shapes for the central transition of half‐integer quadrupolar spin nuclei were calculated as a function of the parameter \documentclass{article}\pagestyle{empty}\begin{document}$ X = \frac{1}{4}[I(I + 1) - \frac{3}{4}]v_{\rm Q} ^2 (1 + \eta ^2 /3)/v_{\rm L} v_{\rm R}$\end{document}. It is shown that complex sideband patterns can be analysed using a very fast computational program.

Carbon-13 nuclear magnetic resonance of ferroelectric liquid crystals with off-magic-angle spinning

The orientational ordering of three ferroelectric liquid crystals 4′-(3-methyl-2-chloropentanoyloxy)-4-alkyloxybiphenyls has been studied by 2D carbon-13 nuclear magnetic resonance (NMR) with variable angle sample spinning. The three homologs have hexyloxy (C6), heptyloxy (C7), and octyloxy (C8) chains, respectively, at the 4 position. They exhibit smectic A (SA) and chiral smectic C (S*C; ferroelectric) phases and have unusually large spontaneous polarization of the electric dipoles. In the SA phase, macroscopic alignment of the molecular director along the spinning axis can be achieved by cooling the sample rapidly from the isotropic liquid. For the S*C phase, the molecular alignment is different for the three compounds. At a magnetic field of 7.05 T, the helical structures of C6 and C8 unwind and the molecular directors align along the spinning axis at a spinning rate of 1 kHz. However, under the same conditions, the helical structure of C7 is retained and its carbon-13 NMR spectrum shows a partial powder pattern. On the other hand, the racemate of C7 has a normal smectic C (SC) phase without a helical structure, and the molecular director can be easily aligned for the NMR study. The racemate of C8 was also synthesized and studied for comparison. The 2D NMR technique used was separated local field (SLF) spectroscopy with an efficient proton–proton dipolar decoupling sequence BLEW-48 in the evolution period. The carbon–proton dipolar coupling constants were determined and the order parameters for different molecular segments of the liquid crystal were calculated. The carbon-13 chemical shifts were also measured as functions of temperature and the data were used to probe the temperature dependence of order parameters. The results show that the C–H bond at the first chiral center in these compounds has an unusually large negative order parameter. This could be due to restricted rotation of the molecular segment at the chiral center, which would be related to the high spontaneous polarization of these compounds.

Investigation of variable angle sample spinning (VASS) NMR of quadrupolar nuclei. I. Theory

Variable angle sample spinning NMR of quadrupolar nuclei in the solid state has been theoretically investigated in order to determine the optimal angles to achieve high-resolved and quantitative spectra. The earlier literature contributions have been included in a more general model. Complete analytical expressions were derived and are reported for all possible cases. It is concluded that magic angle spinning (at 54.7°) is to be used whenever the dipolar interactions and/or the chemical shift anisotropy dominate the spectra. For the other cases, θ=43.5° (which would allow an easy discrimination of the quadrupolar frequency and the asymmetry parameter), and θ=60°–70° (where the linewidth is half of that obtained for MAS conditions) are proposed as preferred spinning angles.

13C chemical shielding anisotropy studied by variable-angle sample spinning

Spinning the sample at angles other than the magic angle is shown to have interesting applications for obtaining chemical shielding anisotropies for systems where severe overlapping makes the analysis of static powder pattern either cumbersome or impossible. Results for 1,3,5-trimethoxybenzene and p-dimethoxybenzene are in excellent agreement with those obtained via single-crystal studies.

Solid state aluminum-27 NMR studies of tridecameric Al-OxoHydroxy clusters in basic aluminum selenate, sulfate, and the mineral zunyite

Aluminum forms a number of tridecameric (Al,,) cluster compounds in which 02-, OH-, FP, and HZ0 are ligands. Examples are the basic aluminum salts (chloride, selenate, and sulfate, Refs. (1, 2)) and the m ineral zunyite (3). The basic aluminum salts are formed by partial hydrolysis of A1(H20)63’ by base, have commercial utility, and have recently been the subject of several studies using aluminum-27 NMR spectroscopy, both in solution (4, 5) and in the crystalline solid state (I, 6). The basic aluminum salts all contain a central AlO unit, surrounded by twelve six-coordinate Al atoms. Early solution NMR studies readily detected signals arising from AD4 in the tr idecamer (4), but only under certain conditions (at high temperature) could resonances attributable to the octahedral aluminums be observed, due presumably to their large quadrupole coupling constants (e2qQ/h). Similarly, detection of the octahedral aluminums in the basic chlorides and sulfates in the solid state has until now not been successful (6). In contrast, solid state NM studies using “mag ic-angle” sample spinning (MASS) (7) of the m ineral zunyite, a basic aluminum silicate having the structure [Ali,(OH, F)i6F2]Si502&1, have recently revealed quite narrow lines for both fourand six-coordinate aluminum (I). In this paper, we present 27A1 MASS, variable-angle sample-spinning (VASS) (8) and static NMR spectra of basic aluminum selenate, sulfate, and the m ineral zunyite, obtained at several magnetic field strengths, which has allowed us to determine e2qQ/h and isotropic chemical-shift values for both Al(4) and Al(6) sites in all three systems. We discuss the likely origins for the large differences in e2qQ/ h between Al(6) sites in zunyite (-2.5 MHz) and the basic aluminum selenate and sulfate (10 MHz). Availability of e*qQ/h (and asymmetry parameter, 7) information will be shown elsewhere to permit evaluation of rotational correlation times for the Ali3 clusters in solution, and thereby to permit estimation of oxygen-l 7 e’qQ/h values in such systems (9). The basic aluminum selenate and sulfate were obtained as Na[AI04 a Ali2(OH)24(OH2)12] (X0&.YH20 (X = Se or S and Y 13, Ref. (2)) from