Application Of Pulsed Field Gradients Research Articles

H(C)P and H(P)C triple‐resonance experiments at natural abundance employing long‐range couplings

Modified two-dimensional (2D) triple-resonance H(C)P and H(P)C experiments based on INEPT/HMQC and double-INEPT schemes are applied to the study of organophosphorus compounds at natural abundances. The implementation of effective (1)H--(13)C gradient selection, additional purging pulsed field gradients, spinlock pulses, and improved phase cycling is demonstrated to allow weak correlation signals based on long-range couplings to be readily observed. Through the combination of two heteronuclear long-range coupling constants, (n)J(CH) and (n)J(PC) in H(C)P experiments or (n)J(PH) and (n)J(PC) in H(P)C experiments, protons can be correlated to a second heteronucleus through 4-7 chemical bonds. These experiments thus overcome the inherit limitations of classical (1)H-X HMBC experiments, which require a nonzero value of the heteronuclear coupling constant (n)J(XH). Ultra-broadband inversion composite pulses are successfully employed in the H(P)C INEPT/HMQC and H(P)C double-INEPT pulse sequences to increase the utility of the experiments and the quality of obtained spectra. This work extends and completes a set of 2D phase-sensitive triple-resonance experiments applicable at natural abundances, and also offers insight into the methodology of triple-resonance experiments and the application of pulsed field gradients. A one-dimensional triple-resonance experiment employing carbon detection is suggested for accurate determination of small (n)J(PC).

Characterisation of the supramolecular structure of malonamides by application of pulsed field gradients in NMR spectroscopy

Malonamides are known and extensively studied for their lanthanide and actinide extracting properties. Those studies have also highlighted aggregated phenomena and a splitting of the organic phase, in some particular experimental conditions. To explain this behaviour of extractants, (1)H NMR was used to study micellar phenomena by the determination of the self-diffusion coefficients of two malonamides only different by the length of their alkyl chain (DMDBTDMA and DMDBPMA), in presence of n-dodecane and for systems saturated with water or anhydrous. Several information on the aggregates and on the malonamide supramolecular structure were obtained by fitting the curves of self-diffusion coefficient vs. concentration and by conjugated NMR experimental data to potentiometric titrations and physical measurements.

A fiber optic-based system for behavioral eyeblink measurement in a MRI environment

We previously described a reliable system to control the timing of multiple stimuli and to detect behavioral eyeblink responses in fMRI studies of learning in animals. Here, we report a significant advancement of the original system, which incorporates a fiber-optic probe in order to avoid the interference associated with the application of pulsed field gradients during MR imaging, particularly echo planar imaging. Eyeblink responses recorded with our fiber-optic probe and modified detection circuit demonstrate the ability of our new system to acquire behavioral data free of gradient-induced artifacts, thereby eliminating the need for low-pass filtering. This fiber optic-based system should be applicable to both animal and human imaging experiments.

Use of Long-Range C-H Heteronuclear Multiple Bond Connectivity in the Assignment of the 13 C NMR Spectra of Complex Organic Molecules

Use of Long-Range C-H Heteronuclear Multiple Bond Connectivity in the Assignment of the 13 C NMR Spectra of Complex Organic Molecules

Signal Selection in High-Resolution NMR by Pulsed Field Gradients: I. Geometrical Analysis

A geometrical description for the selection of coherence transfer pathways in high resolution NMR by the application of pulsed field gradients along three orthogonal directions in space is presented. The response of the spin system is one point of the three-dimensional Fourier transform of the sample volume affected by a sequence of field gradients. The property that a pathway is retained (or suppressed) when a sequence of field gradients is applied is expressed by the property of vectors, representing the pathway and the sequence, respectively, to be orthogonal (or not orthogonal). Ignoring imperfections of RF pulses, and with the exception of sensitivity enhanced experiments and experiments where the relevant coherence order is zero while field gradients are applied, it is shown that at most only half of the relevant pathways, as compared to a phase cycled experiment, are retained when field gradients are used for signal selection.

13CNMR of methane in anAlPO4−11molecular sieve: Exchange effects and shielding anisotropy

${}^{13}\mathrm{C}$ NMR spectra of ${}^{13}{\mathrm{CH}}_{4}$ in an ${\mathrm{AlPO}}_{4}\ensuremath{-}11$ molecular sieve reveal exchange effects between adsorbed and nonadsorbed methane gas. An application of pulsed field gradients is introduced to decrease nonadsorbed and exchanging gas signals in order to extract the chemical shift anisotropy line shape of the adsorbed gas. The resulting ${}^{13}\mathrm{C}$ shielding anisotropy of methane is compared to existing value for methane in related SAPO-11 material. Less anisotropic shielding is observed in ${\mathrm{AlPO}}_{4}\ensuremath{-}11,$ most likely due to the lack of charge-compensating cations.

α/β-ge-SELINCOR-TOCSY, a New Method for the Determination of H,C Coupling Constants

Long-range H,C coupling constants provide important a specific carbon atom which is in the a or in the b spin state. Two soft pulses are used: The first one selects the structural information. There are many methods known for the determination of JH,C coupling constants (1) . One may carbon resonance under proton decoupling, and a second soft pulse determines the spin state. The suppression of the divide them into different approaches. Many experiments use an evolution time, which is adjusted to the heteronuclear main signal arising from protons bound to C is improved by the application of pulsed field gradients. Obtaining the coupling constant (2–12) . There are also several carbonor proton-selective experiments. Since one is looking for H,C coupling constants in high resolution is no problem, since this is a one-dimensional method. coupling constants smaller than 10 Hz, one must find a compromise between a sufficiently long evolution delay and reThe new method consists of two parts: a spin-selective geSELINCOR and a subsequent TOCSY transfer. It is already laxation. Unfortunately, very small couplings may not evolve, and the consequence is the absence of magnetization known that the carbon-selective inverse correlation called SELINCOR (26) is significantly improved when pulsed field transfer. An interesting way to avoid these difficulties was progradients are used for coherence-pathway selection (27) . In this experiment, a carbon-selective pulse is used under proposed by Zhu et al. (13, 14) . They compared peak intensities of a reference spectrum (an H,C correlation without carbon ton decoupling to choose a special carbon resonance. This is done in an INEPT-like fashion and a CW spin lock is pulses) with cross peaks in the HMQC experiment adjusted for JCH couplings. They conclude that the coupling is small used for decoupling (28) . Thus, the experiment starts with the proton magnetization of the directly bonded proton of when there is a small signal present in the H,C correlation in comparison to the reference spectrum. Of course, this is the chosen carbon atom. To complete the INEPT segment, a 907 hard proton pulse is applied. Then the carbon magnetinot satisfying for very small coupling constants (o1 Hz), but for large molecules, this method seems to be useful. zation created in antiphase to the proton is spatially labeled by the third pulsed field gradient. The two weak gradients Another group of experiments is related to the application of heteronuclear-coupled TOCSY or NOESY experiments. surrounding the 1807 pulses are needed to retain the 1807 character of these two pulses (29) . In Fig. 1, the pulse For labeled compounds, the standard TOCSY, for example, is sufficient (15, 16) , and E. COSY-like signals are obtained. sequence of a /b-ge-SELINCOR is presented. The last proton pulse creates multiple-quantum coherFor complicated molecules, it is advisable to apply a 3DHSQC-TOCSY or NOESY (17) . ences (IS , IS , IS , and IS) as shown in the coherence-pathway diagram at the bottom of Fig. 1. For a methine For unlabeled compounds, a heteronuclear filter is necessary (18–22) . Recently, carbon-selective one-dimensional carbon, we apply a 1807 carbon soft pulse shifted with the maximum of its exitation profile about half of the JCH couexperiments were proposed, which have the advantage of high digital resolution in contrast to the multidimensional pling constant, because in this experiment the carbon offset is set on the resonance of the chosen carbon. This is easily experiments. Nuzillard and Freeman use a combination of a heteronuclear filter with (multiple) selective Hartmann– done by the method of Patt (30) . The second soft pulse transforms IS to ( i /2)IS or ( i /2)IS as shown in Hahn transfers to select a single carbon satellite in the proton spectrum (23) . Similar approaches with an unselective TOCSY spin lock were also proposed (24, 25) . Here, we suggest a new method, which relies on the same IS 1807 I (1,2) x i 2 IS . [1] idea that one may select magnetization of protons bound to

Overcoming Solvent Saturation-Transfer Artifacts in Protein NMR at Neutral pH. Application of Pulsed Field Gradients in Measurements of 1H- 15N Overhauser Effects

Artifacts due to solvent saturation-transfer effects result in incorrect measurements of 1H- 15N heteronuclear NOE (HNOE). These artifacts are commonly observed in aqueous protein solutions at neutral pH. We describe the application of pulsed field gradients (PFGs) together with long recycle delays in overcoming errors in HNOE measurements which arise from H 2O solvent preirradiation and solvent saturation transfer. Even in the absence of explicit solvent irradiation, the HNOE pulse sequence itself results in a nonequilibrium spin-state distribution of solvent nuclei which can then be transferred by chemical exchange into amide-proton sites. This effect can be avoided by using PFGs for suppression of solvent H 2O together with a recycle delay sufficiently long for the magnetization of water to relax back to equilibrium values during the preparation period. These effects were studied in 15N-enriched human type α transforming growth factor at pH 7.1. Comparisons of PFG-HNOE experiments with and without selective H 2O irradiation and with different recycle times provide estimates of the effects of solvent irradiation on HNOE measurements, which are different for different amide nitrogen-15 nuclei. The amplitudes of these artificial HNOE enhancements are roughly correlated with solvent accessibilities of amide sites in the three-dimensional structure of hTGFα.

Rapid and simple approach for the NMR resonance assignment of the carbohydrate chains of an intact glycoprotein Application of gradient-enhanced natural abundance 1H- 13C HSQC and HSQC-TOCSY to the α-subunit of human chorionic gonadotropin

The structure assessment of an intact glycoprotein in solution requires an extensive assignment of the carbohydrate NMR resonances. However, assignment of homonuelear spectra is very complicated because of the severe overlap of protein and carbohydrate signals. Application of pulsed field gradients allowed high quality natural abundance 1H- 13C HSQC and HSQC-TOCSY spectra to be recorded of the α-subunit of human chorionic gonadotropin. Most carbohydrate 1H- 13C correlations appear in a distinct region between the aromatic region and the protein C α-H α region. The enormous reduction in overlap led to fast and unambiguous assignment of the anomeric 1H- 13C correlations. Subsequently, correlations of the monosaccharide skeleton atoms were readily assigned in the HSQC-TOCSY spectrum.

Modern multi-dimensional protein NMR spectroscopy: II. New experimental approaches

The first part of this review, published in the previous issue of Trends in Analytical Chemistry, dealt mainly with the new heteronuclear three-dimensional and four-dimensional NMR methods for the assignment of 1H, 13C and 15N resonances in larger proteins. In this part we will describe three recent developments in NMR techniques that still have to see their full impact in practice: (a) heteronuclear cross polarization as an alternative to INEPT for heteronuclear coherence transfer, (b) principles of the measurement of scalar couplings that are (much) smaller than the resonance linewidths, and (c) the application of pulsed field gradients in high-resolution NMR.

Quadrature detection in F1 induced by pulsed field gradients

The improved understanding and control of induced eddy current effects has led to the increased application of pulsed field gradients in high-resolution 2D NMR spectroscopy. Of particular interest is the ability to select specific coherences in a single acquisition with concomitant “quadrature detection” in the F 1 dimension. A product-operator analysis of this effect for an AX spin system which describes the selection of N-type or P-type signals using appropriate gradient episodes in the COSY and 2D double-quantum experiments is presented.