Category Archives: Low Temperature

Cryogenic Platforms and Optimized DNP Sensitivity #DNPNMR

Matsuki, Yoh, and Toshimichi Fujiwara. “Cryogenic Platforms and Optimized DNP Sensitivity,” 7:16, 2018.

Modern high-field DNP NMR spectrometers are typically based on a cryogenic magic-angle sample spinning (MAS) capability. Conventionally, sample temperatures of T ∼100 K have been widely used, enabling substantial NMR signal enhancement with DNP at high external field conditions such as B0 =9.4 T. Today, however, the need for performing MAS DNP at much lower temperatures (T ≪100 K) is receiving growing attention for its ability to recover the rapidly degrading efficiency of the cross-effect (CE)-based DNP at even higher magnetic fields, B0 >10 T. In this article, we describe three contemporary cryogenic DNP MAS NMR probe systems: one is N2 based for T ∼100 K, and the other two are helium based for T ≪100 K. Principal requirements important in designing the cryogenic MAS NMR systems include long-term stability, cost efficiency, and readiness of operation. All the described setups incorporated various modifications and novel features to meet these challenges. In particular, the novel closed-cycle helium MAS system realizes all the requirements to a high standard, establishing an efficient and practical platform for ultralow sample temperature (T ∼30 K) MAS DNP. The resulting dramatic increase in sensitivity gain suggests the regained promise for the CE-based DNP at very high-field conditions (B0 >10 T). The experimental DNP data and effective sensitivity gain obtained with the described systems operating at 14.1 and 16.4 T are also discussed.

Magic angle spinning NMR below 6 K with a computational fluid dynamics analysis of fluid flow and temperature gradients

This article is not specifically about DNP spectroscopy. However, magic angle spinning at 6K is definitely of interest to DNP, especially when using low-power, solid-state microwave sources.

Sesti, E.L., et al., Magic angle spinning NMR below 6 K with a computational fluid dynamics analysis of fluid flow and temperature gradients. J. Magn. Reson., 2018. 286(Supplement C): p. 1-9.

We report magic angle spinning (MAS) up to 8.5 kHz with a sample temperature below 6 K using liquid helium as a variable temperature fluid. Cross polarization 13C NMR spectra exhibit exquisite sensitivity with a single transient. Remarkably, 1H saturation recovery experiments show a 1H T1 of 21 s with MAS below 6 K in the presence of trityl radicals in a glassy matrix. Leveraging the thermal spin polarization available at 4.2 K versus 298 K should result in 71 times higher signal intensity. Taking the 1H longitudinal relaxation into account, signal averaging times are therefore predicted to be expedited by a factor of >500. Computer assisted design (CAD) and finite element analysis were employed in both the design and diagnostic stages of this cryogenic MAS technology development. Computational fluid dynamics (CFD) models describing temperature gradients and fluid flow are presented. The CFD models bearing and drive gas maintained at 100 K, while a colder helium variable temperature fluid stream cools the center of a zirconia rotor. Results from the CFD were used to optimize the helium exhaust path and determine the sample temperature. This novel cryogenic experimental platform will be integrated with pulsed dynamic nuclear polarization and electron decoupling to interrogate biomolecular structure within intact human cells.

A sub-Kelvin cryogen-free EPR system

This article has nothing to do with DNP. However, the article nicely describes an impressive piece of isntrumentation to reach very low temperatures.

Melhuish, S.J., et al., A sub-Kelvin cryogen-free EPR system. J. Magn. Reson., 2017. 282(Supplement C): p. 83-88.

We present an EPR instrument built for operation at Q band below 1K. Our cryogen-free Dewar integrates with a commercial electro-magnet and bridge. A description of the cryogenic and RF systems is given, along with the adaptations to the standard EPR experiment for operation at sub-Kelvin temperatures. As a first experiment, the EPR spectra of powdered Cr12O9(OH)3(O2CCMe3)15 were measured. The sub-Kelvin EPR spectra agree well with predictions, and the performance of the sub-Kelvin system at 5K is compared to that of a commercial spectrometer.

Temperature dependence of cross-effect dynamic nuclear polarization in rotating solids: advantages of elevated temperatures #DNPNMR

Geiger, M.A., et al., Temperature dependence of cross-effect dynamic nuclear polarization in rotating solids: advantages of elevated temperatures. Phys. Chem. Chem. Phys., 2016. 18(44): p. 30696-30704.

Dynamic nuclear polarization exploits electron spin polarization to boost signal-to-noise in magic-angle-spinning (MAS) NMR, creating new opportunities in materials science, structural biology, and metabolomics studies. Since protein NMR spectra recorded under DNP conditions can show improved spectral resolution at 180-200 K compared to 110 K, we investigate the effects of AMUPol and various deuterated TOTAPOL isotopologues on sensitivity and spectral resolution at these temperatures, using proline and reproducibly prepared SH3 domain samples. The TOTAPOL deuteration pattern is optimized for protein DNP MAS NMR, and signal-to-noise per unit time measurements demonstrate the high value of TOTAPOL isotopologues for Protein DNP MAS NMR at 180-200 K. The combined effects of enhancement, depolarization, and proton longitudinal relaxation are surprisingly sample-specific. At 200 K, DNP on SH3 domain standard samples yields a 15-fold increase in signal-to-noise over a sample without radicals. 2D and 3D NCACX/NCOCX spectra were recorded at 200 K within 1 and 13 hours, respectively. Decreasing enhancements with increasing 2H-content at the CH2 sites of the TEMPO rings in CD3-TOTAPOL highlight the importance of protons in a sphere of 4-6 A around the nitroxyl group, presumably for polarization pickup from electron spins.

Nuclear magnetic resonance studies of DNP-ready trehalose obtained by solid state mechanochemical amorphization #DNPNMR

Filibian, M., et al., Nuclear magnetic resonance studies of DNP-ready trehalose obtained by solid state mechanochemical amorphization. Phys Chem Chem Phys, 2016. 18(25): p. 16912-20.

(1)H nuclear spin-lattice relaxation and Dynamic Nuclear Polarization (DNP) have been studied in amorphous samples of trehalose sugar doped with TEMPO radicals by means of mechanical milling, in the 1.6-4.2 K temperature range. The radical concentration was varied between 0.34 and 0.81%. The highest polarization of 15% at 1.6 K, observed in the sample with concentration 0.50%, is of the same order of magnitude of that reported in standard frozen solutions with TEMPO. The temperature and concentration dependence of the spin-lattice relaxation rate 1/T1, dominated by the coupling with the electron spins, were found to follow power laws with an exponent close to 3 in all samples. The observed proportionality between 1/T1 and the polarization rate 1/Tpol, with a coefficient related to the electron polarization, is consistent with the presence of Thermal Mixing (TM) and a good contact between the nuclear and the electron spins. At high electron concentration additional relaxation channels causing a decrease in the nuclear polarization must be considered. These results provide further support for a more extensive use of amorphous DNP-ready samples, obtained by means of comilling, in dissolution DNP experiments and possibly for in vivo metabolic imaging.

Ultra-low temperature MAS-DNP #DNPNMR

Lee, D., et al., Ultra-low temperature MAS-DNP. J. Magn. Reson., 2016. 264: p. 116-124.

Since the infancy of NMR spectroscopy, sensitivity and resolution have been the limiting factors of the technique. Regular essential developments on this front have led to the widely applicable, versatile, and powerful spectroscopy that we know today. However, the Holy Grail of ultimate sensitivity and resolution is not yet reached, and technical improvements are still ongoing. Hence, high-field dynamic nuclear polarization (DNP) making use of high-frequency, high-power microwave irradiation of electron spins has become very promising in combination with magic angle sample spinning (MAS) solid-state NMR experiments. This is because it leads to a transfer of the much larger polarization of these electron spins under suitable irradiation to surrounding nuclei, greatly increasing NMR sensitivity. Currently, this boom in MAS-DNP is mainly performed at minimum sample temperatures of about 100 K, using cold nitrogen gas to pneumatically spin and cool the sample. This Perspective deals with the desire to improve further the sensitivity and resolution by providing “ultra”-low temperatures for MAS-DNP, using cryogenic helium gas. Different designs on how this technological challenge has been overcome are described. It is shown that stable and fast spinning can be attained for sample temperatures down to 30 K using a large cryostat developed in our laboratory. Using this cryostat to cool a closed-loop of helium gas brings the additional advantage of sample spinning frequencies that can greatly surpass those achievable with nitrogen gas, due to the differing fluidic properties of these two gases. It is shown that using ultra-low temperatures for MAS-DNP results in substantial experimental sensitivity enhancements and according time-savings. Access to this temperature range is demonstrated to be both viable and highly pertinent.

Low-temperature dynamic nuclear polarization with helium-cooled samples and nitrogen-driven magic-angle spinning

Thurber, K. and R. Tycko, Low-temperature dynamic nuclear polarization with helium-cooled samples and nitrogen-driven magic-angle spinning. J Magn Reson, 2016. 264: p. 99-106.

We describe novel instrumentation for low-temperature solid state nuclear magnetic resonance (NMR) with dynamic nuclear polarization (DNP) and magic-angle spinning (MAS), focusing on aspects of this instrumentation that have not been described in detail in previous publications. We characterize the performance of an extended interaction oscillator (EIO) microwave source, operating near 264GHz with 1.5W output power, which we use in conjunction with a quasi-optical microwave polarizing system and a MAS NMR probe that employs liquid helium for sample cooling and nitrogen gas for sample spinning. Enhancement factors for cross-polarized (13)C NMR signals in the 100-200 range are demonstrated with DNP at 25K. The dependences of signal amplitudes on sample temperature, as well as microwave power, polarization, and frequency, are presented. We show that sample temperatures below 30K can be achieved with helium consumption rates below 1.3l/h. To illustrate potential applications of this instrumentation in structural studies of biochemical systems, we compare results from low-temperature DNP experiments on a calmodulin-binding peptide in its free and bound states.

Determination of sample temperature in unstable static fields by combining solid-state 79Br and 13C NMR

This is not an article about DNP-NMR spectroscopy, however, it deals with the measurements of temperatures in solid-state NMR experiments using KBR and referencing its chemical shift to 13C of adamantane in unstable magnetic fields.

Purusottam, R.N., G. Bodenhausen, and P. Tekely, Determination of sample temperature in unstable static fields by combining solid-state (79)Br and (13)C NMR. J Magn Reson, 2014. 246(0): p. 69-71.

Monitoring the isotropic chemical shifts to calibrate the sample temperature presupposes a perfect stability of the static magnetic field. It can be difficult to satisfy this requirement in solid-state NMR measurements. This paper describes a simple way to recover the accurate temperature dependence of the (79)Br resonance after subtracting changes of resonance frequency due to variations of the static field, monitored by the (13)C resonance.

Remote sensing of sample temperatures in nuclear magnetic resonance using photoluminescence of semiconductor quantum dots

This article is not exclusively about DNP-NMR spectroscopy, but describes an interesting approach of measuring cryogenic temperatures using a simple fiber optic-based setup. It is particularly useful for low-temperature MAS NMR experiments, including DNP-NMR spectroscopy.

Tycko, R., Remote sensing of sample temperatures in nuclear magnetic resonance using photoluminescence of semiconductor quantum dots. J Magn Reson, 2014. 244C(0): p. 64-67.

Knowledge of sample temperatures during nuclear magnetic resonance (NMR) measurements is important for acquisition of optimal NMR data and proper interpretation of the data. Sample temperatures can be difficult to measure accurately for a variety of reasons, especially because it is generally not possible to make direct contact to the NMR sample during the measurements. Here I show that sample temperatures during magic-angle spinning (MAS) NMR measurements can be determined from temperature-dependent photoluminescence signals of semiconductor quantum dots that are deposited in a thin film on the outer surface of the MAS rotor, using a simple optical fiber-based setup to excite and collect photoluminescence. The accuracy and precision of such temperature measurements can be better than +/-5K over a temperature range that extends from approximately 50K (-223 degrees C) to well above 310K (37 degrees C). Importantly, quantum dot photoluminescence can be monitored continuously while NMR measurements are in progress. While this technique is likely to be particularly valuable in low-temperature MAS NMR experiments, including experiments involving dynamic nuclear polarization, it may also be useful in high-temperature MAS NMR and other forms of magnetic resonance.

Magic-Angle Spinning NMR of Cold Samples

Solid-state DNP experiments are often performed at cryogenic temperatures and this article is an excellent review about the current technologies to spin samples at (very) low temperatures.

ConcistrÈ, M., et al., Magic-Angle Spinning NMR of Cold Samples. Acc. Chem. Res., 2013.

Magic-angle-spinning solid-state NMR provides site-resolved structural and chemical information about molecules that complements many other physical techniques. Recent technical advances have made it possible to perform magic-angle-spinning NMR experiments at low temperatures, allowing researchers to trap reaction intermediates and to perform site-resolved studies of low-temperature physical phenomena such as quantum rotations, quantum tunneling, ortho-para conversion between spin isomers, and superconductivity. In examining biological molecules, the improved sensitivity provided by cryogenic NMR facilitates the study of protein assembly or membrane proteins. The combination of low-temperatures with dynamic nuclear polarization has the potential to boost sensitivity even further. Many research groups, including ours, have addressed the technical challenges and developed hardware for magic-angle-spinning of samples cooled down to a few tens of degrees Kelvin. In this Account, we briefly describe these hardware developments and review several recent activities of our group which involve low-temperature magic-angle-spinning NMR. Low-temperature operation allows us to trap intermediates that cannot be studied under ambient conditions by NMR because of their short lifetime. We have used low-temperature NMR to study the electronic structure of bathorhodopsin, the primary photoproduct of the light-sensitive membrane protein, rhodopsin. This project used a custom-built NMR probe that allows low-temperature NMR in the presence of illumination (the image shows the illuminated spinner module). We have also used this technique to study the behavior of molecules within a restricted environment. Small-molecule endofullerenes are interesting molecular systems in which molecular rotors are confined to a well-insulated, well-defined, and highly symmetric environment. We discuss how cryogenic solid state NMR can give information on the dynamics of ortho-water confined in a fullerene cage. Molecular motions are often connected with fundamental chemical properties; therefore, an understanding of molecular dynamics can be important in fields ranging from material science to biochemistry. We present the case of ibuprofen sodium salt which exhibits different degrees of conformational freedom in different parts of the same molecule, leading to a range of line broadening and line narrowing phenomena as a function of temperature.

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