Designing broadband pulsed dynamic nuclear polarization sequences in static solids

The coherent transfer of electron spin polarization to nuclei by means of a microwave pulse sequence is a promising new approach to enhancing the sensitivity of solid-state nuclear magnetic resonance (NMR). The development of pulse sequences for dynamic nuclear polarization (DNP) of bulk nuclei is far from complete, as is the understanding of what makes a good DNP sequence. In this context, we introduce a new sequence, termed Two-Pulse Phase Modulation (TPPM) DNP. We provide a general theoretical description for electron-proton polarization transfer by periodic DNP pulse sequences and find it in excellent agreement with numerical simulations. In experiments at 1.2T, TPPM DNP generates a higher gain in sensitivity than existing sequences XiX (X-inverse-X) and TOP (Time-Optimized Pulsed) DNP but does so at relatively high nutation frequencies. In contrast, we find that the XiX sequence performs very well at nutation frequencies as low as 7MHz. A combination of theoretical analysis and experimental investigation makes clear that fast electron-proton polarization transfer, due to a well-preserved dipolar coupling in the effective Hamiltonian, correlates with a short build-up time of the dynamic nuclear polarization of the bulk. Experiments further show that the performances of XiX and TOP DNP are affected differently by the concentration of the polarizing agent. These results constitute important reference points for the development of new and better DNP sequences.

Dynamic nuclear polarization (DNP) is a nuclear magnetic resonance (NMR) hyperpolarization technique that mediates polarization transfer from unpaired electrons to nuclear spins. DNP performance can vary significantly depending on the types of polarizing agents employed, and the criteria for optimum DNP efficiency are not fully understood. Thus, a better understanding of the structure, electron paramagnetic resonance (EPR) line widths, and relaxation properties would aid in designing more efficient DNP polarizing agents. However, EPR characterizations of the polarizing agents are typically performed in different environments (e.g., strength of magnetic field and microwave power) than typical DNP experiments. Here, we demonstrate a low-cost and home-built setup that enables in situ EPR detection in a dual resonance DNP-NMR/EPR probe using an optical chopper. The optical chopper modulates the microwave irradiation, thereby modulating the longitudinal magnetization (Mz) of the electron spins, which results in a LOngitudinal-Detected (LOD) EPR spectrum. The EPR spectrum acquired for TEMPOL at 6.7 T/188 GHz using a mechanical modulation showed a good agreement with the spectrum acquired using an electrical modulation. Hence, our work provides a proof-of-principle setup that enables an optical chopper EPR spectrometer.

We describe a new triply tuned (e−, 1H, and 13C) resonance structure operating at an electron Larmor frequency of 139.5 GHz for dynamic nuclear polarization (DNP) and electron nuclear double-resonance (ENDOR) experiments. In contrast to conventional double-resonance structures, the body of the microwave cavity simultaneously acts as a NMR coil, allowing for increased efficiency of radiofrequency irradiation while maintaining a high quality factor for microwave irradiation. The resonator design is ideal for low-γ-nuclei ENDOR, where sensitivity is limited by the fact that electron spin relaxation times are on the order of the RF pulse lengths. The performance is demonstrated with 2H ENDOR on a standard perdeuterated bis-diphenylene-phenyl-allyl stable radical. In DNP experiments, we show that the use of this resonator, combined with a low microwave power setup (17 mW), leads to significantly higher 1H signal enhancement (ϵ ∼ 400 ± 50) than previously achieved at 5-T fields. The results emphasize the importance of optimizing the microwave B1 field by improving either the quality factor of the microwave resonator or the microwave power level.

Impact of non-polar solvents in dynamic nuclear polarization at high magnetic fields.

Dynamic nuclear polarization (DNP) combined with high magnetic fields and fast magic angle spinning (MAS) has opened up a new avenue for the application of exceptionally sensitive 1H NMR detection schemes to study protonated solids. Recently, it has been shown that DNP experiments at fast MAS rates lead to slower spin diffusion and hence reduced DNP enhancements for impregnated materials. However, DNP enhancements alone do not determine the overall sensitivity of a NMR experiment. Here we measure the overall sensitivity of one-dimensional 1H detected relayed DNP experiments as a function of the MAS rate in the 20–60 kHz regime using 0.7 mm diameter rotors at 21.2 T. Although faster MAS rates are detrimental for the DNP enhancement on the target material, due to slower spin diffusion, we find that with increasing spinning rates the gain in sensitivity due to 1H line-narrowing and the folding-in of sideband intensity compensates a large part of the loss of overall hyperpolarization. We find that sensitivity depends on the atomic site in the molecule, and is maximised at between 40 and 50 kHz MAS for the sample of L-histidine.HCl·H2O studied here. There is a 10–20 % difference in sensitivity between the optimum MAS rate and the fastest rate currently accessible (60 kHz).

Proton Dynamic Nuclear Polarization (DNP) experiments were conducted on a 3.4 T homebuilt hybrid pulsed-EPR-NMR spectrometer, on static samples containing 10 mM or 40 mM TEMPOL in frozen glassy solutions of DMSO/water. During DNP experiments proton-NMR signals are enhanced with the help of microwave (MW) irradiation on or close to the Electron Paramagnetic Resonance (EPR) spectrum of the free radicals in the sample, transferring polarization from the free electrons to the nuclei. In the solid state a distinction is made between three DNP enhancement mechanisms: the Solid Effect (SE), the Cross Effect (CE) and Thermal Mixing (TM). In an effort to determine the dominant DNP mechanisms responsible for the enhancement of the nuclear signals, electron and nuclear spin-lattice relaxation rates, enhancement buildup times and microwave (MW) swept DNP spectra were measured as a function of temperature and MW irradiation strength. We observed lineshape variations of the DNP spectra that indicated changes in the relative contributions of SE-DNP and CE-DNP with temperature and MW power. Using a theoretical model describing the SE-DNP and CE-DNP the DNP spectra could be analyzed without involving the TM-DNP mechanism and the relative SE-DNP and CE-DNP contributions to the nuclear enhancement could be determined. From this analysis it follows that lowering the temperature beyond 20 K increases the SE-DNP and decreases the CE-DNP contributions. Possible explanations for this behavior are suggested.

In a previous communication [Hu et al., J. Am. Chem. Soc. 126, 10844 (2004)], an approach was demonstrated that improves the efficiency of the cross-effect polarization mechanism employed in high field dynamic nuclear polarization (DNP) experiments. Specifically, it was shown that tethering two TEMPO (2,2,6,6-tetramethylpiperidin-1-oxyl) radicals increases the electron-electron dipole coupling from approximately 1 MHz in solutions of monomeric TEMPO to approximately 25 MHz in a tethered biradical. The larger coupling resulted in an increase in the DNP enhancements by a factor of approximately 3-4, from 45-50 to approximately 165. Here, a second approach to improving the efficiency of the polarization process is described that involves approximately satisfying the matching condition |omega(2e)-omega(1e)|=omega(n), where omega(2e) and omega(1e) are two frequencies in the electron paramagnetic resonance (EPR) spectrum and omega(n) is the Larmor frequency of the nuclear spins being polarized. Specifically, in a mixture of TEMPO and trityl [tris (8-carboxy-2,2,6,6-tetramethyl(-d3)-benzo[1,2d:4,5-d']bis(1,3)dithiol-4-yl) methyl] radicals, the intensity maxima in the EPR spectra of these two species are approximately separated by the (1)H NMR frequency. In this case the frequency difference between the g(yy) value of TEMPO and the narrow pseudo-isotropic g-value of trityl is approximately 224 MHz and the (1)H Larmor frequency is 211 MHz. The optimal magnetic field for DNP using the mixtures was found to coincide with the trityl EPR resonance. At 90 K and 5 T, a mixture of 20 mM TEMPO and 20 mM trityl enhanced the (1)H polarization by a factor of approximately 160, an improvement over the enhancement of approximately 50 with 40 mM TEMPO. The reasons for the improvement are discussed and evidence is presented suggesting that DNP enhancement can be improved further by tethering TEMPO and trityl or two similar radicals.

An apparatus for pulsed ESR and DNP experiments using optically excited triplet states down to liquid helium temperatures

Despite extensive research conducted on plating and dendrite formation in lithium batteries, the molecular formation processes are not yet fully understood. Electron paramagnetic resonance (EPR) sensitively detects metallic Li species but misses non-paramagnetic ones. Nuclear magnetic resonance (NMR) is chemically selective, yet exhibits low sensitivity under low-field conditions. Dynamic nuclear polarization (DNP) overcomes this by transferring electron spin polarization to nuclei. Here, correlative EPR and DNP-enhanced NMR of lithium on copper is demonstrated using a custom setup operating at 0.34 T with a sweepable electromagnet. DNP experiments were conducted in pulsed mode to minimize sample heating. The resulting enhanced NMR signal allows the observation of electrochemically deposited lithium on copper, harvested from a Cu vs. Li cell, with an enhancement 400$$\\end{document}]]>. By changing the magnetic field strength by a few Gauss, the saturation of the conduction EPR transition was varied, leading to an altered Knight shift of metallic . The corresponding change of the DNP-polarized chemical shifts in the range from 240 ppm to 80 ppm allowed an indirect, saturation-based distinction of EPR species. Moreover, an enhancement by a factor of about 2 of the H signal from the surrounding electrolyte of electrochemically deposited lithium was observed, indicating the potential to investigate the solid–electrolyte interface (SEI). The setup employed a battery cell housing developed for EPR, demonstrating its suitability for in operando experiments in the future.

Paramagnet induced signal quenching in MAS–DNP experiments in frozen homogeneous solutions

Dynamic nuclear polarization experiments were performed to study the solutions of the stable free radical α,γ‐Bisdiphenylene‐β‐phenyl allyl complex with benzene (1∶1) in some highly fluorinated aliphatic and aromatic solvents. In solutions examined in this study, the Overhauser effect, which normally arises due to both dipolar and scalar interactions between the unpaired electrons of the free radical molecules and fluorine nuclei of solvent molecules occurs mainly. 1‐Iodotridecafluorohexane, 2,2,3,4,4,4‐Hexafluoro‐1‐butanol, N‐methyl‐bis‐trifluoroacetamide, hexafluoroacetylacetone, octafluorotoluene, and hexafluorobenzene were used as the solvents. The experiments were performed at a low field double resonance NMR spectrometer, which operates at 1.53 mT. The NMR enhancements depend on competition between intermolecular magnetic interactions. The measurements were performed at four different temperatures to test the dipolar and the scalar part of the coupling between the fluorine nucleus (19F) and the unpaired electron. It was found that the dipolar interactions are more effective for the aliphatic solvents, while the scalar interactions are more effective for the aromatics. The nuclear‐electron coupling parameter varies between 0.018 and 0.157 in all aliphatic solvents and between −0.063 and −0.035 in aromatic solvents.

Nuclear magnetic resonance (NMR) has become the method of choice for many types of applications. Still, sensitivity is a limiting factor in the applicability of NMR, leading to long measurement times in advanced multidimensional experiments, and becoming prohibitive when very limited sample quantities are available. This low sensitivity is mostly an intrinsic consequence of the low energy scale of the nuclear moment in a static field, when compared to other thermodynamic energies like k B T. The commercial developments are mostly aimed at an increase in the static field and simultaneously a reduction of the noise using cryocooled detection coils. Current research shows a number of interesting developments toward the enhancement of the nuclear polarization by optical pumping or by transfer from the electronic bath in dynamic nuclear polarization (DNP) experiments. A more technological approach is based on the miniaturization of the RF coils. In the next decade, one may expect the advent of the “lab on a chip” with in situ chemical processing and NMR analysis capabilities. A brave new method to improve detection sensitivity is based on very sensitive micromechanical force detectors. Recently, it was demonstrated that the low-temperature force detection sensitivity is sufficient to detect the magnetic moment of a single (electron) spin. These developments show that the NMR detection limits in terms of absolute sensitivity or imaging resolution are still open to significant improvements.

Biological Applications of Hyperpolarized 13C NMR

Solid state nuclear magnetic resonance (NMR) enables atomic-resolution characterization of the molecular structure and dynamics within complex heterogeneous samples, but it is typically insensitive. Dynamic nuclear polarization (DNP) increases the NMR signal intensity by orders of magnitude and can be performed in combination with magic angle spinning (MAS) for sensitive, high-resolution spectroscopy. Here we report MAS DNP experiments, for the first time, within intact human cells with >40-fold DNP enhancement and a sample temperature of <6 K. In addition to cryogenic MAS results at <6 K, we also show in-cell DNP enhancements of 57-fold at 90 K. In-cell DNP is demonstrated using biradicals and sterically shielded monoradicals as polarizing agents. A novel trimodal polarizing agent is introduced for DNP, which contains a nitroxide biradical, a targeting peptide for cell penetration, and a fluorophore for subcellular localization with confocal microscopy. The fluorescent polarizing agent provides in-cell DNP enhancements of 63-fold at a concentration of 2.7 mM. These experiments pave the way for structural characterization of biomolecules in an endogenous cellular context.

Nuclear Magnetic Resonance (NMR) spectroscopy would be a method of choice to follow biochemical events in cells because it can analyze molecules in complex environments. However, the intrinsically low sensitivity of NMR makes in-cell measurements challenging. Dynamic Nuclear Polarization (DNP) has emerged as a method to circumvent this limitation, but most polarizing agents developed for DNP are unstable in reducing cellular environments. Here, we introduce the use of Gd(III)-based DNP polarizing agents for in-cell NMR spectroscopy. Specifically, we show their persistent stability in cellular formulations, and we investigate the DNP performance of the Gd(III)-based complex [Gd(tpatcn)] in human embryonic kidney cell lysates and intact cells. For cell lysates, DNP enhancements up to -27 are obtained on the cellular signals, reproducible even after storage at room temperature for days. Mixing the [Gd(tpatcn)] solution with intact cells enables the observation of cellular signals with DNP, and DNP enhancement factors of about -40 are achieved.