Dynamic nuclear polarization (DNP) is a method that permits NMR signal intensities of solids and liquids to be enhanced significantly, and is therefore potentially an important tool in structural and mechanistic studies of biologically relevant molecules. During a DNP experiment, the large polarization of an exogeneous or endogeneous unpaired electron is transferred to the nuclei of interest (I) by microwave (μw) irradiation of the sample. The maximum theoretical enhancement achievable is given by the gyromagnetic ratios (γ e /γ l ), being ∼660 for protons. In the early 1950s, the DNP phenomenon was demonstrated experimentally, and intensively investigated in the following four decades, primarily at low magnetic fields. This review focuses on recent developments in the field of DNP with a special emphasis on work done at high magnetic fields (≥5 T), the regime where contemporary NMR experiments are performed. After a brief historical survey, we present a review of the classical continuous wave (cw) DNP mechanisms-the Overhauser effect, the solid effect, the cross effect, and thermal mixing. A special section is devoted to the theory of coherent polarization transfer mechanisms, since they are potentially more efficient at high fields than classical polarization schemes. The implementation of DNP at high magnetic fields has required the development and improvement of new and existing instrumentation. Therefore, we also review some recent developments in μw and probe technology, followed by an overview of DNP applications in biological solids and liquids. Finally, we outline some possible areas for future developments.
A new polarizing agent with superior performance in dynamic nuclear polarization experiments is introduced, and utilizes two TEMPO (2,2,6,6-tetramethylpiperidine-1-oxyl) moieties connected through a rigid spiro tether (see structure). The observed NMR signal intensities were enhanced by a factor of 1.4 compared to those of TOTAPOL, a previously described TEMPO-based biradical with a flexible tether.
Using dynamic nuclear polarization (DNP)/nuclear magnetic resonance instrumentation that utilizes a microwave cavity and a balanced rf circuit, we observe a solid effect DNP enhancement of 94 at 5 T and 80 K using trityl radical as the polarizing agent. Because the buildup rate of the solid effect increases with microwave field strength, we obtain a sensitivity gain of 128. The data suggest that higher microwave field strengths would lead to further improvements in sensitivity. In addition, the observation of microwave field dependent enhancements permits us to draw conclusions about the path that polarization takes during the DNP process. By measuring the time constant for the polarization buildup and enhancement as a function of the microwave field strength, we are able to compare models of polarization transfer, and show that the major contribution to the bulk polarization arises via direct transfer from electrons, rather than transferring first to nearby nuclei and then transferring to bulk nuclei in a slow diffusion step. In addition, the model predicts that nuclei near the electron receive polarization that can relax, decrease the electron polarization, and attenuate the DNP enhancement. The magnitude of this effect depends on the number of near nuclei participating in the polarization transfer, hence the size of the diffusion barrier, their T 1 , and the transfer rate. Approaches to optimizing the DNP enhancement are discussed.
ABSTRACT:A new biradical polarizing agent, bTbtk-py, for dynamic nuclear polarization (DNP) experiments in aqueous media is reported. The synthesis is discussed in light of the requirements of the optimum, theoretical, biradical system. To date, the DNP NMR signal enhancement resulting from bTbtk-py is the largest of any biradical in the ideal glycerol/water solvent matrix, ε = 230. EPR and X-ray crystallography are used to characterize the molecule and suggest approaches for further optimizing the biradical distance and relative orientation.
The synthesis and characterization of a biradical containing a 1,3-bisdiphenylene-2-phenylallyl (BDPA) free radical covalently attached to a 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) free radical are described. The synthesis of the biradical is a step towards improved polarizing agents for dynamic nuclear polarization (DNP).Biradicals are of considerable interest as polarizing agents for microwave driven dynamic nuclear polarization (DNP) NMR experiments. 1,2 Irradiation of the biradical's electron paramagnetic resonance (EPR) drives electron-nuclear transitions that transfer the large polarization of the electrons to the nuclear spins and thereby enhances the signal-to-noise ratio in NMR experiments. The enhancement factors can reach a theoretical maximum of ~660 for electron-1 H polarization transfer and ~2600 when 13 C is the nuclear spin of interest. 3 Thus, optimized biradical polarizing agents can dramatically decrease acquisition times. These signal enhancements are important for a variety of applications involving solid-state NMR, particularly for systems that are not amenable to crystallographic studies, such as amyloid 4 and membrane 5 proteins.In previous work, we demonstrated that bis-nitroxide biradicals, which contain two tethered TEMPO moieties, provide 1 H/ 13 C-signal enhancements ~200 fold in solid-state magic-anglespinning (MAS) NMR spectra. 2 We are interested in molecules that can produce DNP by a cross effect (CE) mechanism 3 that involves three spins, two dipolar coupled electrons and a nuclear spin (usually 1 H), denoted by S 1 and S 2 and I, with electron and nuclear Larmor frequencies, ω 0S 1 , ω 0S 2 , and ω 0 I , respectively. In a DNP experiment, microwave irradiation of the biradical's EPR spectrum induces the two electrons to undergo a spin flip-flop process, during which a nuclear spin is polarized if the electron Larmor frequencies are separated by the nuclear Larmor frequency and therefore satisfy the matching condition, ω 0S 1 -ω 0S 2 = ω 0 I . The high 1 H polarizaton is then transferred via cross-polarization to 13 C or 15 N, resulting in an enhanced MAS NMR spectrum. 6 The efficiency of the CE mechanism depends on how many pairs of electrons satisfy the matching condition. Thus, the ideal CE polarizing agent would be a biradical with an EPR spectrum consisting of two sharp lines separated by ω 0I . However, at the high magnetic fields (>5T) where contemporary NMR experiments are performed, only a few known radicals NIH Public Access exhibit narrow spectra. Among them are two stable species -trityl radical derivatives 7 and the BDPA radical 8 (Scheme 1), which have similar isotropic g-values (g iso (trityl)= 2.00307, 1 g iso (BDPA)= 2.00264). If trityl or BDPA serves as one of the lines in the EPR spectrum of a polarizing agent, then to satisfy the CE matching condition it is necessary to introduce another radical with a line separated from the first by ω 0I . There are no known stable radicals that provide a narrow line and meet this condition, however TEMPO ...
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