Real-Time Monitoring of Hydrolysis Reactions of Pyrophosphates with Dissolution Dynamic Nuclear Polarization.

Dissolution dynamic nuclear polarization (d-DNP) has enabled applications such as the real-time monitoring of chemical reactions. Such applications are mainly for 13C and 15N spins with long spin-lattice relaxation times in the molecules of interest. However, the only applications for phosphorus using d-DNP are pH imaging and nucleation during crystallization due to the short relaxation times. Here we show that it is possible to observe enzyme reactions using d-DNP with phosphorus. Hyperpolarized 31P spins in pyrophosphate were obtained using bullet-DNP, which requires less dilution of highly polarized solid samples. Real-time monitoring of the hydrolysis reaction of pyrophosphate by inorganic pyrophosphatase from baker's yeast at physiological pH and was successfully achieved and the reaction rate was determined. This is an important reaction for a wide range of applications related to medicine, agriculture, and quantum life science.

Efficiency analysis of helium-cooled MAS DNP: case studies of surface-modified nanoparticles and homogeneous small-molecule solutions.

Despite the growing number of successful applications of dynamic nuclear polarization (DNP)-enhanced magic-angle spinning (MAS) NMR in structural biology and materials science, the nuclear polarizations achieved by current MAS DNP instrumentation are still considerably lower than the theoretical maximum. The method could be significantly strengthened if experiments were performed at temperatures much lower than those currently widely used (∼100 K). Recently, the prospects of helium (He)-cooled MAS DNP have been increased with the instrumental developments in MAS technology that uses cold helium gas for sample cooling. Despite the additional gains in sensitivity that have been observed with He-cooled MAS DNP, the performance of the technique has not been evaluated in the case of surfaces and interfaces that benefit the most from DNP. Herein, we studied the efficiency of DNP at temperatures between ∼30 K and ∼100 K for organically functionalized silica material and a homogeneous solution of small organic molecules at a magnetic field B0 = 16.4 T. We recorded the changes in signal enhancement, paramagnet-induced quenching and depolarization effects, DNP build-up rate, and Boltzmann polarization. For these samples, the increases in MAS-induced depolarization and DNP build-up times at around 30 K were not as severe as anticipated. In the case of the surface species, we determined that MAS DNP at 30 K provided ∼10 times higher sensitivity than MAS DNP at 90 K, which corresponds to the acceleration of experiments by multiplicative factors of up to 100.

Advanced instrumentation for DNP-enhanced MAS NMR for higher magnetic fields and lower temperatures.

Sensitivity enhancement of MAS NMR using dynamic nuclear polarization (DNP) is gaining importance at moderate fields (B0<9T) and temperatures (T>90K) with potential applications in chemistry and material sciences. However, considering the ever-increasing size and complexity of the systems to be studied, it is crucial to establish DNP under higher field conditions, where the spectral resolution and the basic NMR sensitivity tend to improve. In this perspective, we overview our recent efforts on hardware developments, specifically targeted on improving DNP MAS NMR at high fields. It includes the development of gyrotrons that enable continuous frequency tuning and rapid frequency modulation for our 395 GHz-600 MHz and 460 GHz-700 MHz DNP NMR spectrometers. The latter 700 MHz system involves two gyrotrons and a quasi-optical transmission system that combines two independent sub-millimeter waves into a single dichromic wave. We also describe two cryogenic MAS NMR probe systems operating, respectively, at T ∼ 100K and ∼ 30K. The latter system utilizes a novel closed-loop helium recirculation mechanism, achieving cryogenic MAS without consuming any cryogen. These instruments altogether should promote high-field DNP toward more efficient, reliable and affordable technology. Some experimental DNP results obtained with these instruments are presented.

Post-processing of individual signals for de-noising.

In this work, we examine the fluctuation of the intensity and the phase of an NMR signal during repetition of experiments and investigate possibilities of using these information to judge suspicious peaks, whose true colors may be noises or genuine signals. We firstly analyze the intensity and the phase of an NMR signal separately, and show that for the accumulated spectral profile the contribution of the intensity is less than that of the phase. Secondly we show that we can de-noise a noisy spectrum by using the standard deviation of phase at each spectral point. We then compare the de-noising effect of the present approach and that of the phase-covariance method proposed recently, which is an alternative method of appreciating phase distribution. Finally, effects of the dispersion component are discussed.

Phase covariance in NMR signal.

To examine noisy nuclear-magnetic resonance (NMR) spectra, we have developed a novel signal analysis method based on phase correlation between the NMR signal and the excitation pulse. The new phase-covariance analysis is compatible with the conventional signal accumulation, and successful de-noising is demonstrated.

(14)N quadrupolar coupling of amide nitrogen and Peptide secondary structure as studied by solid-state NMR spectroscopy.

To establish a relationship between the secondary structure of a peptide and the quadrupolar coupling of its amide (14)N, we examined (14)N quadrupolar couplings for eight different polypeptide samples, each of whose secondary structure (alpha-helix or beta-sheet) is known. The (14)N quadrupolar coupling is estimated from indirect observation of a (14)N overtone resonance under magic-angle spinning. From the observed indirect (14)N overtone spectra and calculated (14)N quadrupolar couplings for model molecules by using ab initio calculation (Gaussian03), it is shown that the quadrupolar coupling for the alpha-helix is larger than that for the beta-sheet by a few 100 kHz irrespective of the kind of amino acid residues examined (Ala, Val, Leu).