Category Archives: Research Article

Increased flow rate of hyperpolarized aqueous solution for dynamic nuclear polarization-enhanced magnetic resonance imaging achieved by an open Fabry–Pérot type microwave resonator #DNPNMR

Fedotov, Alexey, Ilya Kurakin, Sebastian Fischer, Thomas Vogl, Thomas F. Prisner, and Vasyl Denysenkov. “Increased Flow Rate of Hyperpolarized Aqueous Solution for Dynamic Nuclear Polarization-Enhanced Magnetic Resonance Imaging Achieved by an Open Fabry–Pérot Type Microwave Resonator.” Magnetic Resonance 1, no. 2 (November 18, 2020): 275–84.

A continuous flow dynamic nuclear polarization (DNP) employing the Overhauser effect at ambient temperatures can be used among other methods to increase sensitivity of magnetic resonance imaging (MRI). The hyperpolarized state of water protons can be achieved by flowing aqueous liquid through a microwave resonator placed directly in the bore of a 1.5 T MRI magnet. Here we describe a new open Fabry–Pérot resonator as DNP polarizer, which exhibits a larger microwave exposure volume for the flowing liquid in comparison with a cylindrical TE013 microwave cavity. The Fabry–Pérot resonator geometry was designed using quasi-optical theory and simulated by CST software. Performance of the new polarizer was tested by MRI DNP experiments on a TEMPOL aqueous solution using a blood-vessel phantom. The Fabry–Pérot resonator revealed a 2-fold larger DNP enhancement with a 4-fold increased flow rate compared to the cylindrical microwave resonator. This increased yield of hyperpolarized liquid allows MRI applications on larger target objects.

Tailored flavoproteins acting as light-driven spin machines pump nuclear hyperpolarization

Ding, Yonghong, Alexey S. Kiryutin, Ziyue Zhao, Qian-Zhao Xu, Kai-Hong Zhao, Patrick Kurle, Saskia Bannister, et al. “Tailored Flavoproteins Acting as Light-Driven Spin Machines Pump Nuclear Hyperpolarization.” Scientific Reports 10, no. 1 (December 2020): 18658.

The solid-state photo-chemically induced dynamic nuclear polarization (photo-CIDNP) effect generates non-Boltzmann nuclear spin magnetization, referred to as hyperpolarization, allowing for high gain of sensitivity in nuclear magnetic resonance (NMR). Well known to occur in photosynthetic reaction centers, the effect was also observed in a light-oxygen-voltage (LOV) domain of the bluelight receptor phototropin, in which the functional cysteine was removed to prevent photo-chemical reactions with the cofactor, a flavin mononucleotide (FMN). Upon illumination, the FMN abstracts an electron from a tryptophan to form a transient spin-correlated radical pair (SCRP) generating the photo-CIDNP effect. Here, we report on designed molecular spin-machines producing nuclear hyperpolarization upon illumination: a LOV domain of aureochrome1a from Phaeodactylum tricornutum, and a LOV domain named 4511 from Methylobacterium radiotolerans (Mr4511) which lacks an otherwise conserved tryptophan in its wild-type form. Insertion of the tryptophan at canonical and novel positions in Mr4511 yields photo-CIDNP effects observed by 15N and 1H liquidstate high-resolution NMR with a characteristic magnetic-field dependence indicating an involvement of anisotropic magnetic interactions and a slow-motion regime in the transient paramagnetic state. The heuristic biomimetic design opens new categories of experiments to analyze and apply the photo-CIDNP effect.

Assessment of the Role of 2,2,2-Trifluoroethanol Solvent Dynamics in Inducing Conformational Transitions in Melittin: An Approach with Solvent 19F Low-Field NMR Relaxation and Overhauser Dynamic Nuclear Polarization Studies

Chaubey, Bhawna, Arnab Dey, Abhishek Banerjee, N. Chandrakumar, and Samanwita Pal. “Assessment of the Role of 2,2,2-Trifluoroethanol Solvent Dynamics in Inducing Conformational Transitions in Melittin: An Approach with Solvent 19F Low-Field NMR Relaxation and Overhauser Dynamic Nuclear Polarization Studies.” The Journal of Physical Chemistry B 124, no. 28 (July 16, 2020): 5993–6003.

2, 2, 2-Trifluoroethanol (TFE) is one of the fluoroalcohols that have been known to induce and stabilize open helical structure in many proteins and peptides. The current study has benchmarked low field 19F NMR relaxation and 19F Overhauser Dynamic Nuclear Polarization (DNP) by providing a brief account of TFE solvent dynamics in a model Melittin (MLT; an antimicrobial peptide) solution with TFE: D2O cosolvent mixture at pH 7.4. Further, this approach has been employed to reveal the solvation of MLT by TFE in a nonbuffered solution with a pH 2.8 for the first time. The structural transition of MLT has been elucidated via solvent dynamics by measuring 19F TFE relaxation rates at 0.34 T for various TFE: D2O compositions in absence (bulk TFE) and in presence of MLT at both the pH values. A complementary initial record of Circular Dichroism (CD) experiments on these aqueous MLT solutions with TFE as cosolvent at two different pH conditions demonstrated the structural transition from random coil to helical, or from folded helical to open helical structure. The molecular correlation time derived from corresponding relaxation rates shows that TFE resides on the MLT surface in both pH conditions. However, the trends in the variation of molecular correlation time ratio as a function of TFE concentration represent that the mechanism and the extent to which TFE affects the MLT structural integrity are different at different pH. The extraction of the DNP coupling parameter from steady state 19F ODNP experiments performed in presence of TEMPOL at 0.34 T revealed changes in solvation dynamics of TFE concomitant with MLT structural transition. In summary, 19F relaxation and ODNP measurements made at low field have allowed direct monitoring of TFE dynamics during MLTs structural transition in terms of preferential solvation. The choice of experiments performed at moderately low field (0.34 T) enabled us to exploit on the one hand almost 1200-fold mitigation of the strong contribution of 19F CSA at 11.76 T, while on the other hand the ODNP experiment offered a window for probing molecular dynamics on timescales of the order of 10-1000 picoseconds.

Diffusion measurements with continuous hydrogenation in PHIP

Bussandri, S., L. Buljubasich, and R.H. Acosta. “Diffusion Measurements with Continuous Hydrogenation in PHIP.” Journal of Magnetic Resonance 320 (November 2020): 106833.

DOSY is a powerful spectroscopic NMR technique that resolves components in mixtures through the evaluation of different diffusion coefficients. The application of DOSY to dilute mixtures is hampered by the low signal to noise ratios (SNR), leading to long acquisition times. The use of PHIP may resolve this issue as long as reproducible signals are obtained in order to perform 2D experiments. Here we show that the use of hollow membranes and adequate gas flow produce constant polarization for a time-span that enables the acquisition of 2D experiments. A pressure gradient is evidenced by the presence of convection, which is accounted for by using a DPGSE sequence. The influence of J-coupling evolution during the sequence is studied both numerically and experimentally, to determine the optimum echo-time. The applicability of the method for samples with poor SNR is explored by setting the reaction rate to achieve a low intensity of polarized signals.

SQUID-based ultralow-field MRI of a hyperpolarized material using signal amplification by reversible exchange

Lee, Seong-Joo, Keunhong Jeong, Jeong Hyun Shim, Hyun Joon Lee, Sein Min, Heelim Chae, Sung Keon Namgoong, and Kiwoong Kim. “SQUID-Based Ultralow-Field MRI of a Hyperpolarized Material Using Signal Amplification by Reversible Exchange.” Scientific Reports 9, no. 1 (December 2019): 12422.

The signal amplification by reversible exchange (SABRE) technique is a very promising method for increasing magnetic resonance (MR) signals. SABRE can play a particularly large role in studies with a low or ultralow magnetic field because they suffer from a low signal-to-noise ratio. In this work, we conducted real-time superconducting quantum interference device (SQUID)-based nuclear magnetic resonance (NMR)/magnetic resonance imaging (MRI) studies in a microtesla-range magnetic field using the SABRE technique after designing a bubble-separated phantom. A maximum enhancement of 2658 for 1H was obtained for pyridine in the SABRE-NMR experiment. A clear SABRE-enhanced MR image of the bubble-separated phantom, in which the para-hydrogen gas was bubbling at only the margin, was successfully obtained at 34.3 μT. The results show that SABRE can be successfully incorporated into an ultralow-field MRI system, which enables new SQUID-based MRI applications. SABRE can shorten the MRI operation time by more than 6 orders of magnitude and establish a firm basis for future low-field MRI applications.

Scalable dissolution-dynamic nuclear polarization with rapid transfer of a polarized solid #DNPNMR

Kouřil, Karel, Hana Kouřilová, Samuel Bartram, Malcolm H. Levitt, and Benno Meier. “Scalable Dissolution-Dynamic Nuclear Polarization with Rapid Transfer of a Polarized Solid.” Nature Communications 10, no. 1 (December 2019): 1733.

In dissolution-dynamic nuclear polarization, nuclear spins are hyperpolarized at cryogenic temperatures using radicals and microwave irradiation. The hyperpolarized solid is dissolved with hot solvent and the solution is transferred to a secondary magnet where strongly enhanced magnetic resonance signals are observed. Here we present a method for transferring the hyperpolarized solid. A bullet containing the frozen, hyperpolarized sample is ejected using pressurized helium gas, and shot into a receiving structure in the secondary magnet, where the bullet is retained and the polarized solid is dissolved rapidly. The transfer takes approximately 70 ms. A solenoid, wound along the entire transfer path ensures adiabatic transfer and limits radical-induced low-field relaxation. The method is fast and scalable towards small volumes suitable for high-resolution nuclear magnetic resonance spectroscopy while maintaining high concentrations of the target molecule. Polarization levels of approximately 30% have been observed for 1-13C-labelled pyruvic acid in solution.

Lignin-polysaccharide interactions in plant secondary cell walls revealed by solid-state NMR #DNPNMR

Kang, Xue, Alex Kirui, Malitha C. Dickwella Widanage, Frederic Mentink-Vigier, Daniel J. Cosgrove, and Tuo Wang. “Lignin-Polysaccharide Interactions in Plant Secondary Cell Walls Revealed by Solid-State NMR.” Nature Communications 10, no. 1 (December 2019): 347.

Lignin is a complex aromatic biopolymer that strengthens and waterproofs plant secondary cell walls, enabling mechanical stability in trees and long-distance water transport in xylem. Lignin removal is a key step in paper production and biomass conversion to biofuels, motivating efforts to re-engineer lignin biosynthesis. However, the physical nature of lignin’s interactions with wall polysaccharides is not well understood. Here we show that lignin selfaggregates to form highly hydrophobic and dynamically unique nanodomains, with extensive surface contacts to xylan. Solid-state NMR spectroscopy of intact maize stems, supported by dynamic nuclear polarization, reveals that lignin has abundant electrostatic interactions with the polar motifs of xylan. Lignin preferentially binds xylans with 3-fold or distorted 2-fold helical screw conformations, indicative of xylans not closely associated with cellulose. These findings advance our knowledge of the molecular-level organization of lignocellulosic biomass, providing the structural foundation for optimization of post-harvest processing for biofuels and biomaterials.

Nuclear spin-hyperpolarization generated in a flavoprotein under illumination: experimental field-dependence and theoretical level crossing analysis

Ding, Yonghong, Alexey S. Kiryutin, Alexandra V. Yurkovskaya, Denis V. Sosnovsky, Renad Z. Sagdeev, Saskia Bannister, Tilman Kottke, et al. “Nuclear Spin-Hyperpolarization Generated in a Flavoprotein under Illumination: Experimental Field-Dependence and Theoretical Level Crossing Analysis.” Scientific Reports 9, no. 1 (December 2019): 18436.

The solid-state photo-chemically induced dynamic nuclear polarization (photo-CIDNP) effect generates non-equilibrium nuclear spin polarization in frozen electron-transfer proteins upon illumination and radical-pair formation. The effect can be observed in various natural photosynthetic reaction center proteins using magic-angle spinning (MAS) nuclear magnetic resonance (NMR) spectroscopy, and in a flavin-binding light-oxygen-voltage (LOV) domain of the blue-light receptor phototropin. In the latter system, a functionally instrumental cysteine has been mutated to interrupt the natural cysteineinvolving photochemistry allowing for an electron transfer from a more distant tryptophan to the excited flavin mononucleotide chromophore. We explored the solid-state photo-CIDNP effect and its mechanisms in phototropin-LOV1-C57S from the green alga Chlamydomonas reinhardtii by using fieldcycling solution NMR. We observed the 13C and, to our knowledge, for the first time, 15N photo-CIDNP signals from phototropin-LOV1-C57S. Additionally, the 1H photo-CIDNP signals of residual water in the deuterated buffer of the protein were detected. The relative strengths of the photo-CIDNP effect from the three types of nuclei, 1H, 13C and 15N were measured in dependence of the magnetic field, showing their maximum polarizations at different magnetic fields. Theoretical level crossing analysis demonstrates that anisotropic mechanisms play the dominant role at high magnetic fields.

Glyoxalase activity in human erythrocytes and mouse lymphoma, liver and brain probed with hyperpolarized 13C-methylglyoxal #DNPNMR #dDNP

Shishmarev, Dmitry, Philip W. Kuchel, Guilhem Pagès, Alan J. Wright, Richard L. Hesketh, Felix Kreis, and Kevin M. Brindle. “Glyoxalase Activity in Human Erythrocytes and Mouse Lymphoma, Liver and Brain Probed with Hyperpolarized 13C-Methylglyoxal.” Communications Biology 1, no. 1 (December 2018): 232.

Methylglyoxal is a faulty metabolite. It is a ubiquitous by-product of glucose and amino acid metabolism that spontaneously reacts with proximal amino groups in proteins and nucleic acids, leading to impairment of their function. The glyoxalase pathway evolved early in phylogeny to bring about rapid catabolism of methylglyoxal, and an understanding of the role of methylglyoxal and the glyoxalases in many diseases is beginning to emerge. Metabolic processing of methylglyoxal is very rapid in vivo and thus notoriously difficult to detect and quantify. Here we show that 13C nuclei in labeled methylglyoxal can be hyperpolarized using dynamic nuclear polarization, providing 13C nuclear magnetic resonance signal enhancements in the solution state close to 5,000-fold. We demonstrate the applications of this probe of metabolism for kinetic characterization of the glyoxalase system in isolated cells as well as mouse brain, liver and lymphoma in vivo.

Zero-field nuclear magnetic resonance of chemically exchanging systems

Barskiy, Danila A., Michael C. D. Tayler, Irene Marco-Rius, John Kurhanewicz, Daniel B. Vigneron, Sevil Cikrikci, Ayca Aydogdu, et al. “Zero-Field Nuclear Magnetic Resonance of Chemically Exchanging Systems.” Nature Communications 10, no. 1 (December 2019): 3002.

Zero- to ultralow-field (ZULF) nuclear magnetic resonance (NMR) is an emerging tool for precision chemical analysis. In this work, we study dynamic processes and investigate the influence of chemical exchange on ZULF NMR J-spectra. We develop a computational approach that allows quantitative calculation of J-spectra in the presence of chemical exchange and apply it to study aqueous solutions of [15N]ammonium (15NHfl4 ) as a model system. We show that pH-dependent chemical exchange substantially affects the J-spectra and, in some cases, can lead to degradation and complete disappearance of the spectral features. To demonstrate potential applications of ZULF NMR for chemistry and biomedicine, we show a ZULF NMR spectrum of [2-13C]pyruvic acid hyperpolarized via dissolution dynamic nuclear polarization (dDNP). We foresee applications of affordable and scalable ZULF NMR coupled with hyperpolarization to study chemical exchange phenomena in vivo and in situations where high-field NMR detection is not possible to implement.

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