Efficient Dynamic Nuclear Polarization at 800 MHz/527 GHz with Trityl‐Nitroxide Biradicals

Abstract

Cross-effect (CE) dynamic nuclear polarization (DNP) is a rapidly developing technique that enhances the signal intensities in magic angle spinning (MAS) NMR spectra. We report CE DNP experiments at 211, 600 and 800 MHz using a new series of biradical polarizing agents referred to as TEMTriPols, in which a nitroxide (TEMPO) and a trityl radical are chemically tethered. The TEMTriPol molecule with the optimal performance yields a record 1 H NMR signal enhancement of 65 at 800 MHz at a concentration of 10 mM in a glycerol/water solvent matrix. The CE DNP enhancement for the TEMTriPol biradicals does not decrease as the magnetic field is increased in the manner usually observed for bis-nitroxides. Instead, the relatively strong exchange interaction between the trityl and nitroxide moieties determines the magnetic field at which the optimum enhancement is observed. Graphical abstractA series of biradicals consisting of a nitroxide chemically tethered to a trityl are employed for cross-effect DNP at 211, 600 and 800 MHz. The relatively strong exchange interaction between the trityl and nitroxide moieties determines the field strength at which the enhancement is optimized, and yields a record 1 H NMR signal enhancement of 65 at 800 MHz.Correspondence to: Guinevere Mathies; Yangping Liu; Robert G. Griffin. # Current address: Dr. Kan-Nian Hu: Vertex Pharmaceuticals Incorporated, 50 Northern Avenue, Boston, MA 02210 (USA); Dr. Marc A. Caporini: Amgen Inc., 360 Binney Street, Cambridge, MA 02142 (USA) Supporting information for this article is given via a link at the end of the document. In these experiments, unpaired electrons (introduced into the NMR sample in the form of polarizing agents) are irradiated with microwaves at or near the electron Larmor frequency, and induce mutual electron-nuclear spin flips effecting a transfer of the electron spin polarization to the surrounding nuclei. Since its first application to MAS NMR at high field, [11] the efficiency of the DNP method has been increasing steadily through the development of dedicated highpower, high-frequency gyrotrons as microwave sources, specialized NMR/DNP cryo-probes, and optimization of sample preparation. Moreover, major increases in the enhancement factors have been derived from the chemical synthesis of improved polarizing agents. HHS Public AccessThe DNP mechanism that is currently most successful in high-field MAS NMR experiments is the cross-effect (CE It requires two interacting electrons, one of which interacts with a nearby nucleus, commonly a proton. The difference between the electron Larmor frequencies of these two electrons must be equal to the 1 H Larmor frequency, ω 0e1 -ω 0e2 = ±ω 01H . At this simplified CE matching condition (vide infra), strong state mixing occurs and consequently there is a high probability that microwave radiation resonant with electron 1 will flip electron 2 and the coupled proton together. [18] Initially, CE DNP experiments were performed with samples containing high concentrations of nitroxide monoradicals,...

Molecular Rationale for Improved Dynamic Nuclear Polarization of Biomembranes

“…37 The latter has been observed in nitroxide-trityl biradicals, where the unpaired electron of trityl is highly delocalized over the triarylmethyl molecular scaffold and is able to interact with the localized unpaired electron of the N-O moiety of the nitroxide radical. 39, 40 If the exchange interaction is strong enough such that J » A, where J is the magnitude of the exchange interaction and A is the hyperfine coupling constant, then additional multiplet structures would be observed in the EPR spectrum. Upon a further increase in J the multiplet structure would begin to exchange narrow until only a single transition is observed in the EPR spectrum.…”

“…In a similar fashion to the electron-electron dipolar coupling the exchange interaction can lead to the state mixing required for cross effect DNP. 40 However, Mathies G., et al note that the interaction between electrons should not equal or exceed the larmor frequency of the target nuclei for hyperpolarization. 40 Therefore, we contend that as the SL-lipid concentration is increased beyond 3 mol% the electron-electron dipolar coupling starts to approach the 1 H larmor frequency and begins to interfere with the cross effect DNP mechanism.…”

New NMR tools for protein structure and function: Spin tags for dynamic nuclear polarization solid state NMR

“…Here, the nuclear polarization originates from two coupled electrons whose difference in Larmor frequency matches the electron frequency: ω 1e − ω 2e = ω n . Effective experimental conditions for the cross effect have been discussed in many reports [38–40]. Since successful cross effect DNP requires two paramagnetic centers, the development of DNP optimized biradicals has greatly extended the applicability of this mechanism.…”

NMR Signal Quenching from Bound Biradical Affinity Reagents in DNP Samples

“…8,9 This mechanism is efficient in favorable cases at high magnetic fields up to 800 MHz. 10 The use of biradicals such as TOTAPOL 11 and AMUPol, 12 in which two nitroxide radicals are tethered to increase cross effect efficiency, has enabled excellent enhancements on many systems. 13–15 …”