Using CW DNP techniques we have recently obtained large signal enhancements in magic-angle spinning (MAS) solid-state NMR (SSNMR) spectra of 15N-alanine labeled T4 lysozyme and """"""""C-Glycine in frozen aqueous solutions of 40:60 water/glycerol with the free radical 4-amino TEMPO as the source of electron polarization. Although these CW-DNP techniques provide large enhancements (E-10), they requires long polarization transfer times (on the order of the spin-lattice relaxation time, TJ. This has motivated us to consider pulsed polarization transfer techniques for DNP, which might be more efficient than the CW DNP methods. We have performed a Nuclear Rotating Frame DNP (NRF-DNP) experiment in which a proton NMR signal enhancement per unit time of elt = 90 has been obtained at high field (BO = 5 T, Vp, = 139.5 GlIz) using 15 mM trityl radical in a 40:60 water/glycerol frozen solution at 11 K. The electron-nuclear polarization transfer is performed in the nuclear rotating frame with microwave/RF irradiation times of 100 ms. A majority of the enhancement is attributed to the thermal mixing mechanism. The growth of the signal enhancement is governed by the rotating frame nuclear spin-lattice relaxation time (TP), typically 10- 100 ms at I I K. Due to the rapid polarization transfer' * the experiment can be recycled at a rate of approximately 11T,P and is not limited by the much longer lab frame nuclear spin-lattice relaxation time (T,,,), typically many minutes at low temperatures. The NRF-DNP experiment does not require high microwave power; significant signal enhancements were obtained with a low power (20 mW) Gunn diode microwave source and no microwave resonant structure. Finally, the symmetric trityl radical is an ideal polarization agent for pulsed DNP studies of biological systems since it is water-soluble and has a narrow EPR line width of 10 G at a field of 5 Tesla.
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