NMR Signal Enhancement by Effective SABRE Labeling of Oligopeptides

Signal amplification by reversible exchange (SABRE) can enhance nuclear magnetic resonance signals by several orders of magnitude. However, until now this was limited to a small number of model target molecules. Here, a new convenient method for SABRE activation applicable to a variety of synthetic model oligopeptides is demonstrated. For the first time, a highly SABRE‐active pyridine‐based biocompatible molecular framework is incorporated into synthetic oligopeptides. The SABRE activity is preserved, demonstrating the importance of such earmarking. Finally, a crucial exchange process responsible for SABRE activity is identified and discussed.     

Heteronuclear Cross‐Relaxation Effects in the NMR Spectroscopy of Hyperpolarized Targets

Dissolution dynamic nuclear polarization (DNP) enables high‐sensitivity solution‐phase NMR experiments on long‐lived nuclear spin species such as ¹⁵N and ¹³C. This report explores certain features arising in solution‐state ¹H NMR upon polarizing low‐γ nuclear species. Following solid‐state hyperpolarization of both ¹³C and ¹H, solution‐phase ¹H NMR experiments on dissolved samples revealed transient effects, whereby peaks arising from protons bonded to the naturally occurring ¹³C nuclei appeared larger than the typically dominant ¹²C‐bonded ¹H resonances. This enhancement of the satellite peaks was examined in detail with respect to a variety of mechanisms that could potentially explain this observation. Both two‐ and three‐spin phenomena active in the solid state could lead to this kind of effect; still, experimental observations revealed that the enhancement originates from ¹³C→¹H polarization‐transfer processes active in the liquid state. Kinetic equations based on modified heteronuclear cross‐relaxation models were examined, and found to well describe the distinct patterns of growth and decay shown by the ¹³C‐bound ¹H NMR satellite resonances. The dynamics of these novel cross‐relaxation phenomena were determined, and their potential usefulness as tools for investigating hyperpolarized ensembles and for obtaining enhanced‐sensitivity ¹H NMR traces was explored.     

13C PHIP NMR spectra and polarization transfer during the homogeneous hydrogenation of alkynes with parahydrogen

Homogeneously catalyzed hydrogenations of unsaturated substrates with parahydrogen not only lead to strong polarization signals in ¹H NMR spectra, but also can give rise to strong heteronuclear polarization, especially if the hydrogenations are carried out in low magnetic fields. As a typical example, the polarization transfer from protons to carbon nuclei during the hydrogenation of alkynes is outlined for several substrates. In systems containing easily accessible triple bonds, e.g. phenylethyne or 3,3‐dimethyl‐1‐butyne, polarization transfer occurs to all carbon nuclei in the molecule. Accordingly, in NMR spectra recorded in situ all ¹³C resonances can be observed with good to excellent signal‐to‐noise ratios using only a single transient. The qualitative influence of symmetry and electronic aspects of the substrate and its hydrogenation product on the efficiency of the transfer of polarization to the ¹³C‐nuclei are discussed. Copyright © 2001 John Wiley & Sons, Ltd.     

Progress in Hyperpolarized Ultrafast 2D NMR Spectroscopy

An important development in the field of NMR spectroscopy has been the advent of hyperpolarization approaches, capable of yielding nuclear spin states whose value exceeds by orders‐of‐magnitude what even the highest‐field spectrometers can afford under Boltzmann equilibrium. Included among these methods is an ex situ dynamic nuclear polarization (DNP) approach, which yields liquid‐phase samples possessing spin polarizations of up to 50 %. Although capable of providing an NMR sensitivity equivalent to the averaging of about 1 000 000 scans, this methodology is constrained to extract its “superspectrum” within a single—or at most a few—transients. This makes it a poor starting point for conventional 2D NMR acquisition experiments, which require a large number of scans that are identical to one another except for the increment of a certain t₁ delay. It has been recently suggested that by merging this ex situ DNP approach with spatially encoded “ultrafast” methods, a suitable starting point could arise for the acquisition of 2D spectra on hyperpolarized liquids. Herein, we describe the experimental principles, potential features, and current limitations of such integration between the two methodologies. For a variety of small molecules, these new hyperpolarized ultrafast experiments can, for equivalent overall durations, provide heteronuclear correlation spectra at significantly lower concentrations than those currently achievable by conventional 2D NMR acquisitions. A variety of challenges still remain to be solved before bringing the full potential of this new integrated 2D NMR approach to fruition; these outstanding issues are discussed.     

Labeling Strategy and Signal Broadening Mechanism of Protein NMR Spectroscopy in Xenopus laevis Oocytes

We used Xenopus laevis oocytes, a paradigm for a variety of biological studies, as a eukaryotic model system for in‐cell protein NMR spectroscopy. The small globular protein GB1 was one of the first studied in Xenopus oocytes, but there have been few reports since then of high‐resolution spectra in oocytes. The scarcity of data is at least partly due to the lack of good labeling strategies and the paucity of information on resonance broadening mechanisms. Here, we systematically evaluate isotope enrichment and labeling methods in oocytes injected with five different proteins with molecular masses of 6 to 54 kDa. ¹⁹F labeling is more promising than ¹⁵N, ¹³C, and ²H enrichment. We also used ¹⁹F NMR spectroscopy to quantify the contribution of viscosity, weak interactions, and sample inhomogeneity to resonance broadening in cells. We found that the viscosity in oocytes is only about 1.2 times that of water, and that inhomogeneous broadening is a major factor in determining line width in these cells.     

Shaped Ceria Nanocrystals Catalyze Efficient and Selective Para‐Hydrogen‐Enhanced Polarization

Intense para‐hydrogen‐enhanced NMR signals are observed in the hydrogenation of propene and propyne over ceria nanocubes, nano‐octahedra, and nanorods. The well‐defined ceria shapes, synthesized by a hydrothermal method, expose different crystalline facets with various oxygen vacancy densities, which are known to play a role in hydrogenation and oxidation catalysis. While the catalytic activity of the hydrogenation of propene over ceria is strongly facet‐dependent, the pairwise selectivity is low (2.4 % at 375 °C), which is consistent with stepwise H atom transfer, and it is the same for all three nanocrystal shapes. Selective semi‐hydrogenation of propyne over ceria nanocubes yields hyperpolarized propene with a similar pairwise selectivity of (2.7 % at 300 °C), indicating product formation predominantly by a non‐pairwise addition. Ceria is also shown to be an efficient pairwise replacement catalyst for propene.     

Biomolecular DNP‐Supported NMR Spectroscopy using Site‐Directed Spin Labeling

Dynamic nuclear polarization (DNP) has been shown to greatly enhance spectroscopic sensitivity, creating novel opportunities for NMR studies on complex and large molecular assemblies in life and material sciences. In such applications, however, site‐specificity and spectroscopic resolution become critical factors that are usually difficult to control by current DNP‐based approaches. We have examined in detail the effect of directly attaching mono‐ or biradicals to induce local paramagnetic relaxation effects and, at the same time, to produce sizable DNP enhancements. Using a membrane‐embedded ion channel as an example, we varied the degree of paramagnetic labeling and the location of the DNP probes. Our results show that the creation of local spin clusters can generate sizable DNP enhancements while preserving the intrinsic benefits of paramagnetic relaxation enhancement (PRE)‐based NMR approaches. DNP using chemical labeling may hence provide an attractive route to introduce molecular specificity into DNP studies in life science applications and beyond.     

2D NMR Trace Analysis by Continuous Hyperpolarization at High Magnetic Field

Nuclear magnetic resonance is often the technique of choice in chemical analysis because of its sensitivity to molecular structure, quantitative character, and straightforward sample preparation. However, determination of trace analytes in complex mixtures is generally limited by low sensitivity and extensive signal overlap. Here, we present an approach for continuous hyperpolarization at high magnetic field that is based on signal amplification by reversible exchange (SABRE) and can be straightforwardly incorporated in multidimensional NMR experiments. This method was implemented in a 2D correlation experiment that allows detection and quantification of analytes at nanomolar concentration in complex solutions.