Field-frequency Lock Research Articles

Correction of field instabilities in biomolecular solid-state NMR by simultaneous acquisition of a frequency reference.

With the advent of faster magic-angle spinning (MAS) and higher magnetic fields, the resolution of biomolecular solid-state nuclear magnetic resonance(NMR) spectra has been continuously increasing. As a direct consequence, the always narrower spectral lines, especially in proton-detected spectroscopy, are also becoming more sensitive to temporal instabilities of the magnetic field in the sample volume. Field drifts in the order of tenths of parts per million occur after probe insertion or temperature change, during cryogen refill, or are intrinsic to the superconducting high-field magnets, particularly in the months after charging. As an alternative to a field-frequency lock based on deuterium solvent resonance rarely available for solid-state NMR, we present a strategy to compensate non-linear field drifts using simultaneous acquisition of a frequency reference (SAFR). It is based on the acquisition of an auxiliary 1D spectrum in each scan of the experiment. Typically, a small-flip-angle pulse is added at the beginning of the pulse sequence. Based on the frequency of the maximum of the solvent signal, the field evolution in time is reconstructed and used to correct the raw data after acquisition, thereby acting in its principle as a digital lock system. The general applicability of our approach is demonstrated on 2D and 3D protein spectra during various situations with a non-linear field drift. SAFR with small-flip-angle pulses causes no significant loss in sensitivity or increase in experimental time in protein spectroscopy. The correction leads to the possibility of recording high-quality spectra in a typical biomolecular experiment even during non-linear field changes in the order of 0.1 ppm h without the need for hardware solutions, such as stabilizing the temperature of the magnet bore. The improvement of linewidths and peak shapes turns out to be especially important for H-detected spectra under fast MAS, but the method is suitable for the detection of carbon or other nuclei as well.

The Field-Frequency Lock for Fast Field Cycling Magnetic Resonance: From NMR to MRI

Magnetic field stability plays a fundamental role in Nuclear Magnetic Resonance (NMR) and Magnetic Resonance Imaging (MRI) experiments, guaranteeing accuracy and reproducibility of results. While high levels of stabilization can be achieved for standard NMR techniques, this task becomes particularly challenging for Fast Field Cycling (FFC) NMR and MRI, where the main magnetic field is switched to higher or lower levels during the pulse sequence, and field stabilization must be guaranteed within a very short time after switching. Recent works have addressed the problem with rigorous tools from control system theory, proposing a model based approach for the synthesis of magnetic field controllers for FFC-NMR. While an experimental proof of concept has underlined the correctness of the approach for a complete FFC-NMR setup, the application of the novel, model based Field-Frequency Lock (FFL) system to a FFC-MRI scanner requires proper handling of field encoding gradients. Furthermore, the proof of concept work has also stressed how further advances in the hardware and firmware could improve the overall performances of the magnetic field control loop. The main aim of this perspective paper is then discussing the key challenges that arise in the development of the FFL system suitable for a complete MRI scanner, as well as defining possible research directions by means of preliminary, simulated experiments, with the final goal of favoring the development of a novel, model based FFL system for FFC-MRI.

Overhauser dynamic nuclear polarization (ODNP)-enhanced two-dimensional proton NMR spectroscopy at low magnetic fields.

The majority of low-field Overhauser dynamic nuclear polarization (ODNP) experiments reported so far have been 1D NMR experiments to study molecular dynamics and in particular hydration dynamics. In this work, we demonstrate the application of ODNP-enhanced 2D J-resolved (JRES) spectroscopy to improve spectral resolution beyond the limit imposed by the line broadening introduced by the paramagnetic polarizing agent. Using this approach, we are able to separate the overlapping multiplets of ethyl crotonate into a second dimension and clearly identify each chemical site individually. Crucial to these experiments is interleaved spectral referencing, a method introduced to compensate for temperature-induced field drifts over the course of the NMR acquisition. This method does not require additional hardware such as a field-frequency lock, which is especially challenging when designing compact systems.

Towards a Model-Based Field-Frequency Lock for Fast-Field Cycling NMR

Fast-field cycling nuclear magnetic resonance (FFC NMR) relaxometry allows to investigate molecular dynamics of complex materials. FFC relaxometry experiments require the magnetic field to reach different values in few milliseconds and field oscillations to stay within few ppms during signal acquisition. Such specifications require the introduction of a novel field-frequency lock (FFL) system. In fact, control schemes based only on current feedback may not guarantee field stability, while standard FFLs are designed to handle very slow field fluctuations, such as thermal derives, and may be ineffective in rejecting faster ones. The aim of this work is then to propose a methodology for the synthesis of a regulator that guarantees rejection of field fluctuations and short settling time. Experimental trials are performed for both model validation and evaluation of the closed-loop performances. Relaxometry experiments are performed to verify the improvement obtained with the new FFL. The results highlight the reliability of the model and the effectiveness of the overall approach.

Field-frequency lock approach for 21.3-MHz high-performance NMR relaxation analyzer

Low-field NMR (LF-NMR) relaxation analyzers have been more and more widely used in food science, biomedicine, and petroleum exploration in recent years. An LF-NMR device analyzes various interactions between sample molecules through spin relaxation, diffusion, flow, and imaging experiments. However, temporal field fluctuations in the magnet limit the scope of application. A stable magnetic field is the basic guarantee for detecting weak signals with shorter relaxation time. This paper presents an approach involving a field-frequency lock for a 21.3-MHz high-performance NMR bipolar permanent magnet. The diameter spherical volume (DSV) of the magnet is only 60 mm. The field-frequency lock system uses 19F in a micro coil integrated into the main probe as a lock detector. The required magnetic field compensation can be calculated from the lock free induction decay (FID) signal frequency, which is measured by lock spectroscopy. The compensation coils used to produce the magnetic field were designed based on Helmholtz coils. The system determines the signal frequency by detecting the lock FID signal and calculates the required compensation-coil current to stabilize the main magnetic field of the analyzer. The results of practical locking experiments in a 21.3-MHz high-performance NMR relaxation analyzer showed that this new approach helps to reduce magnetic field fluctuations from 11 ppm/3h (11 × 10–6) to 0.4 ppm/3h (0.4 × 10–6), which meets the application requirements. This approach is especially viable and effective for a permanent magnet with large field fluctuations. This paper also provides observations of the effect of these fluctuations on NMR measurements before and after installation of the field-frequency lock system.

Towards a Model-Based Field-Frequency Lock for NMR

Nuclear Magnetic Resonance (NMR) relaxometry is a powerful technique that allows to investigate the properties of materials. More advanced NMR relaxometry techniques such as Fast Field-Cycling (FFC) require the magnetic field to reach any desired value in a very short time (few milliseconds) and field oscillations to stay within few ppms. Such specifications call for the introduction of a suitable Field Frequency Lock (FFL) system. FFL relies on an indirect measure of the magnetic field which can be obtained by performing a parallel NMR experiment with a known sample. In this paper we propose a PID controller to guarantee field fluctuations to stay below the desired level and short settling time. The tuning of the controller is based on a mathematical description of the entire process, which is validated by performing real experiments. Numerical simulations show promising results that we expect to be confirmed by real experiments.

Improving accuracy in DOSY and diffusion measurements using triaxial field gradients

NMR measurements of diffusion in solution, whether primarily quantitative, or, (as in DOSY, Diffusion-Ordered Spectroscopy) qualitative, can be particularly demanding. Here we show how the use of appropriate transverse (x, y) pulsed field gradients, orthogonal to the more usual z axis pulsed field gradient applied along the long axis of the sample, can greatly reduce two important sources of systematic error in diffusion experiments. These are the extra signal attenuation caused by sample convection, and gradient-dependent signal phase shifts caused by the magnetic field and field-frequency lock disturbances generated by field gradient pulses.

Distinction between a transgenic and a conventional common bean genotype by 1H HR-MAS NMR

In this work, it is proposed a methodology which allows to differentiate a conventional and a specific transgenic common beans, grown in greenhouse or under field conditions, based on modifications in chemical composition using 1H HR-MAS NMR. It is demonstrated that the influence of typical variables from field planting conditions had no significant influence on the ability to set apart transgenic from conventional. This methodology was corroborated by multivariate data analysis of the 1H NMR and IR spectra. This study also points out the simplicity of using the HR-MAS NMR technique for food analyses. The measurement is highly simplified because it does not require any pretreatment of the sample apart from the addition of a small amount of D2O necessary to produce homogeneous dough and a field frequency lock. Moreover, due to the high concentration of the sample, measurement time in HR-MAS NMR is very short.

Towards a beyond 1 GHz solid-state nuclear magnetic resonance: External lock operation in an external current mode for a 500 MHz nuclear magnetic resonance

Achieving a higher magnetic field is important for solid-state nuclear magnetic resonance (NMR). But a conventional low temperature superconducting (LTS) magnet cannot exceed 1 GHz (23.5 T) due to the critical magnetic field. Thus, we started a project to replace the Nb(3)Sn innermost coil of an existing 920 MHz NMR (21.6 T) with a Bi-2223 high temperature superconducting (HTS) innermost coil. Unfortunately, the HTS magnet cannot be operated in persistent current mode; an external dc power supply is required to operate the NMR magnet, causing magnetic field fluctuations. These fluctuations can be stabilized by a field-frequency lock system based on an external NMR detection coil. We demonstrate here such a field-frequency lock system in a 500 MHz LTS NMR magnet operated in an external current mode. The system uses a (7)Li sample in a microcoil as external NMR detection system. The required field compensation is calculated from the frequency of the FID as measured with a frequency counter. The system detects the FID signal, determining the FID frequency, and calculates the required compensation coil current to stabilize the sample magnetic field. The magnetic field was stabilized at 0.05 ppm∕3 h for magnetic field fluctuations of around 10 ppm. This method is especially effective for a magnet with large magnetic field fluctuations. The magnetic field of the compensation coil is relatively inhomogeneous in these cases and the inhomogeneity of the compensation coil can be taken into account.

Real Time Magnetic Field Sensing and Imaging Using a Single Spin in Diamond

The Zeeman splitting of a localized single spin can be used to construct a highly sensitive magnetometer offering almost atomic spatial resolution. While sub-μT sensitivity can be obtained in principle using pulsed techniques and long measurement times, a fast and easy method without laborious data postprocessing is desirable for a scanning-probe approach with high spatial resolution. In order to measure the resonance frequency in real time, we applied a field-frequency lock to the optically detected magnetic resonance signal of a single electron spin in a nanodiamond. We achieved a sampling rate of up to 100 readings per sec with a sensitivity of 6 μT/sqrt[Hz]. Images of the field distribution around a magnetic wire were acquired with ∼30 μT resolution and 4096 submicron sized pixels in 10 min. The response of several spins was used to reconstruct the field orientation.

Operation of a 500 MHz high temperature superconducting NMR: Towards an NMR spectrometer operating beyond 1 GHz

We have begun a project to develop an NMR spectrometer that operates at frequencies beyond 1GHz (magnetic field strength in excess of 23.5T) using a high temperature superconductor (HTS) innermost coil. As the first step, we developed a 500MHz NMR with a Bi-2223 HTS innermost coil, which was operated in external current mode. The temporal magnetic field change of the NMR magnet after the coil charge was dominated by (i) the field fluctuation due to a DC power supply and (ii) relaxation in the screening current in the HTS tape conductor; effect (i) was stabilized by the 2H field-frequency lock system, while effect (ii) decreased with time due to relaxation of the screening current induced in the HTS coil and reached 10−8(0.01ppm)/h on the 20th day after the coil charge, which was as small as the persistent current mode of the NMR magnet. The 1D 1H NMR spectra obtained by the 500MHz LTS/HTS magnet were nearly equivalent to those obtained by the LTS NMR magnet. The 2D-NOESY, 3D-HNCO and 3D-HNCACB spectra were achieved for ubiquitin by the 500MHz LTS/HTS magnet; their quality was closely equivalent to that achieved by a conventional LTS NMR. Based on the results of numerical simulation, the effects of screening current-induced magnetic field changes are predicted to be harmless for the 1.03GHz NMR magnet system.

Towards beyond 1 GHz NMR: Mechanism of the long-term drift of screening current-induced magnetic field in a Bi-2223 coil

The screening current-induced magnetic field in the (Bi,Pb) 2Sr 2Ca 2Cu 3O x (Bi-2223) insert coil proposed for a beyond 1 GHz nuclear magnetic resonance (NMR) spectrometer may generate a long-term field drift, resulting in a loss of field-frequency lock operation and an inability to make high resolution NMR measurements. The measured screening current-induced magnetic field of a Bi-2223 double-pancake coil exhibits a hysteresis effect at 4.2 K that is reproduced by a numerical simulation based on a finite thickness rectangular superconductor bar model. The screening current-induced field at the coil center is of opposite polarity to that generated by the coil current, and thus the apparent field intensity shows a positive drift with time. On the contrary, the field at a coil end is of the same polarity as the coil field, and the apparent field intensity decreases with time. If we wait for ∼1000 h after coil excitation, the field drift rate approaches the field decay rate of the persistent current of 10 −8 h −1, suitable for a long-term NMR measurement in a beyond 1 GHz NMR spectrometer.

Organization and Assembly of Metal-Thiolate Clusters in Epithelium-specific Metallothionein-4

Mammalian metallothionein-4 (MT-4) was found to be specifically expressed in stratified squamous epithelia where it plays an essential but poorly defined role in regulating zinc or copper metabolism. Here we report on the organization, stability, and the pathway of metal-thiolate cluster assembly in MT-4 reconstituted with Cd(2+) and Co(2+) ions. Both the (113)Cd NMR studies of (113)Cd(7)MT-4 and the spectroscopic characterization of Co(7)MT-4 showed that, similar to the classical MT-1 and MT-2 proteins, metal ions are organized in two independent Cd(4)Cys(11) and Cd(3)Cys(9) clusters with each metal ion tetrahedrally coordinated by terminal and bridging cysteine ligands. Moreover, we have demonstrated that the cluster formation in Cd(7)MT-4 is cooperative and sequential, with the Cd(4)Cys(11) cluster being formed first, and that a distinct single-metal nucleation intermediate Cd(1)MT-4 is required in the cluster formation process. Conversely, the absorption and circular dichroism features of metal-thiolate clusters in Cd(7)MT-4 indicate that marked differences in the cluster geometry exist when compared with those in Cd(7)MT-1/2. The biological implication of our studies as to the role of MT-4 in zinc metabolism of stratified epithelia is discussed.

Efficient compensation of low-frequency magnetic field disturbances in NMR with fluxgate sensors

A simple stabilization scheme of B 0 magnetic field fluctuations is described. The method is based on external measurements of time dependent magnetic field fluctuations by fluxgate sensors and generation of a compensating correction current in a coil mounted directly on an NMR magnet. It is shown that such an approach efficiently eliminates relatively slow magnetic field variations with frequency up to approximately 100 Hz. In combination with a standard 2H field-frequency lock system, the method enables acquisition of reproducible lineshapes and dramatically improves overall performance of a high resolution NMR spectrometer. The presented solution might substitute for the internal lock system in these case where deuterium lock is not available.

Convection-compensating diffusion experiments with phase-sensitive double-quantum filtering

We present a design scheme for phase-sensitive, convection-compensating diffusion experiments with gradient-selected homonuclear double-quantum filtering. The scheme consists of three blocks: a 1/2 J evolution period during which antiphase single-quantum coherences are created; a period of double-quantum evolution; and another 1/2 J period, during which antiphase single-quantum coherences are converted back into an in-phase state. A single coherence transfer pathway is selected using an asymmetric set of gradient pulses, and both diffusion sensitization and convection compensation are built into the gradient coherence transfer pathway selection. Double-quantum filtering can be used either for solvent suppression or spectral editing, and we demonstrate examples of both applications. The new experiment performs well in the absence of a field-frequency lock and does not require magnitude Fourier transformation. The proposed scheme may offer advantages in diffusion measurements of spectrally crowded systems, particularly small molecules solubilized in colloidal solutions or bound to macromolecules.

Platelet Factor 4 and Interleukin-8 CXC Chemokine Heterodimer Formation Modulates Function at the Quaternary Structural Level

The apparent complexity of biology increases as more biomolecular interactions that mediate function become known. We have used NMR spectroscopy and molecular modeling to provide direct evidence that tetrameric platelet factor-4 (PF4) and dimeric interleukin-8 (IL8), two members of the CXC chemokine family, readily interact by exchanging subunits and forming heterodimers via extension of their antiparallel beta-sheet domains. We further demonstrate using functional assays that PF4/IL8 heterodimerization has a direct and significant consequence on the biological activity of both chemokines. Formation of heterodimers enhances the anti-proliferative effect of PF4 on endothelial cells in culture, as well as the IL8-induced migration of CXCR2 vector-transfected Baf3 cells. These results suggest that CXC chemokine biology, and perhaps cytokine biology in general, may be functionally modulated at the molecular level by formation of heterodimers. This concept, in turn, has implications for designing chemokine/cytokine variants with modified biological properties.

External field-frequency lock probe for high resolution solid state NMR

A transmission line lock probe is described for magnetic field stabilization of a high field superconducting magnet with a significant drift rate for high resolution solid state NMR experiments employing a conventional magic angle spinning (MAS) probe. The lock probe connects to the standard spectrometer lock channel and employs a spherical sample bulb containing D2O doped such that the peak is homogeneously broadened despite the sample being displaced from the homogeneous central region of the static magnetic field. Use of a separate lock probe makes it possible to remove the MAS probe for changing samples while maintaining field-frequency lock.

Abstracts

Abstracts

L -band electron paramagnetic resonance spectrometer for use in vivo and in studies of aqueous biological samples

The development of L-band (∼1.2GHz) frequency EPR spectrometers has made feasible many in vivo studies in laboratory animals and, recently, in human volunteers. The lower dielectric and eddy current losses that occur at L-band balance the lower Zeeman splitting so useful measurements can be made in conductive aqueous samples. We describe typical resonators used in such studies and provide details on the construction of the spectrometer, including the bridge, the automatic frequency control subsystem, the low-noise high-stability tunable L-band frequency source, as well as the low-frequency components—the signal receiver and the modulation unit. The application of EPR spectroscopy to larger subjects requires special care in the design of an appropriate magnet with sufficient homogeneity and stability, yet with dimensions that allow operation with a wide range of subject sizes. We describe our solution, which involves a permanent magnet, air-core scan coils to provide the field sweep and offset, and field stabilization by means of a field-frequency lock. We also describe the magnetic field modulation system, which operates at 25 kHz to avoid distortion in spectra from materials with narrow lines (such as lithium phthalocyanine). We refer to recent reviews to illustrate the range of in vivo studies and the clinical applications of the type of spectrometer described here.

Chemical shift referencing in MAS solid state NMR

Solid state 13C magic angle spinning (MAS) NMR spectra are typically referenced externally using a probe which does not incorporate a field frequency lock. Solution NMR shifts on the other hand are more often determined with respect to an internal reference and using a deuterium based field frequency lock. Further differences arise in solution NMR of proteins and nucleic acids where both 13C and 1H shifts are referenced by recording the frequency of the 1H resonance of DSS (sodium salt of 2,2-dimethyl-2-silapentane-5-sulphonic acid) instead of TMS (tetramethylsilane). In this note we investigate the difficulties in relating shifts measured relative to TMS and DSS by these various approaches in solution and solids NMR, and calibrate adamantane as an external 13C standard for solids NMR. We find that external chemical shift referencing of magic angle spinning spectra is typically quite reproducible and accurate, with better than ±0.03 ppm accuracy being straight forward to achieve. Solid state and liquid phase NMR shifts obtained by magic angle spinning with external referencing agree with those measured using typical solution NMR hardware with the sample tube aligned with the applied field as long as magnetic susceptibility corrections and solvent shifts are taken into account. The DSS and TMS reference scales for 13C and 1H are related accurately using MAS NMR. Large solvent shifts for the 13C resonance in TMS in either deuterochloroform or methanol are observed, being +0.71 ppm and −0.74 ppm from external TMS, respectively. The ratio of the 13C resonance frequencies for the two carbons in solid adamantane to the 1H resonance of TMS is reported.