Measuring Protein-Ligand Binding by Hyperpolarized Ultrafast NMR.

Protein-ligand interactions can be detected by observing changes in the transverse relaxation rates of the ligand upon binding. The ultrafast NMR technique, which correlates the chemical shift with the transverse relaxation rate, allows for the simultaneous acquisition of R2 for carbon spins at different positions. In combination with dissolution dynamic nuclear polarization (D-DNP), where the signal intensity is enhanced by thousands of times, the R2 values of several carbon signals from unlabeled benzylamine are observable within a single scan. The hyperpolarized ultrafast chemical shift-R2 correlated experiment separates chemical shift encoding from the readout phase in the NMR pulse sequence, which allows it to beat the fundamental limit on the spectral resolution otherwise imposed by the sampling theorem. Applications enabled by the ability to measure multiple relaxation rates in a single scan include the study of structural properties of protein-ligand interactions.

Melting of aqueous NaCl solutions in porous materials: shifted phase transition distribution (SIDI) approach for determining NMR cryoporometry pore size distributions.

Nuclear magnetic resonance cryoporometry (NMRC) and differential scanning calorimetry thermoporometry (DSC-TPM) are powerful methods for measuring mesopore size distributions. The methods are based on the fact that, according to the Gibbs-Thomson equation, the melting point depression of a liquid confined to a pore is inversely proportional to the pore size. However, aqueous salt solutions, which inherently exist in a broad range of biological porous materials as well as technological applications such as electrolytes, do not melt at a single temperature. This causes artefacts in the pore size distributions extracted by traditional Gibbs-Thomson analysis of NMRC and DSC-TPM data. Bulk aqueous NaCl solutions are known to have a broad distribution of melting points between the eutectic and pure water phase transition points (252-273 K). Here, we hypothesize that, when aqueous NaCl solution (saline) is confined to a small pore, the whole melting point distribution is shifted toward lower temperatures by the value predicted by the Gibbs-Thomson equation. We show that this so-called shifted phase transition distribution (SIDI) approach removes the artefacts arising from the traditional Gibbs-Thomson analysis and gives correct pore size distributions for saline saturated mesoporous silica gel and controlled pore materials analyzed by NMR cryoporometry. Furthermore, we demonstrate that the method can be used for determining pore sizes in collagen-chondroitin sulphate hydrogels resembling the composition of the extracellular matrix of articular cartilage. It is straightforward to apply the SIDI analysis for DSC-TMP data as well.

CAT on MOUSE: Control and automation of temperature for single-sided NMR instruments such as NMR-MOUSE.

Temperature-dependent experiments are a rapidly growing area of interest for low-field NMR. In this work, we present a new device for wide-range temperature control for single-sided NMR instruments. The presented device, called CAT, is simple to build, inexpensive, and easy to modify to accommodate different samples. We present the capabilities of the device using a freezing temperature study of acetic acid/water mixtures. Additionally, we present the stability of the device over long measurement times. We believe that by introducing such a device with an open-source design, we allow researchers to use it in a wide range of applications and to fully incorporate variable-temperature studies in the world of single-sided instruments.

SPICY: a method for single scan rotating frame relaxometry.

T 1ρ is an NMR relaxation mode that is sensitive to low frequency molecular motions, making it an especially valuable tool in biomolecular research. Here, we introduce a new method, SPICY, for measuring T1ρ relaxation times. In contrast to conventional T1ρ experiments, in which the sequence is repeated many times to determine the T1ρ time, the SPICY sequence allows determination of T1ρ within a single scan, shortening the experiment time remarkably. We demonstrate the method using 1H T1ρ relaxation dispersion experiments. Additionally, we combine the sequence with spatial encoding to produce 1D images in a single scan. We show that T1ρ relaxation times obtained using the single scan approach are in good agreement with those obtained using the traditional experiments.

Ultrafast transverse relaxation exchange NMR spectroscopy.

Molecular exchange between different physical or chemical environments occurs due to either diffusion or chemical transformation. Nuclear magnetic resonance (NMR) spectroscopy provides a means of understanding the molecular exchange in a noninvasive way and without tracers. Here, we introduce a novel two dimensional, single-scan ultrafast Laplace NMR (UF LNMR) method to monitor molecular exchange using transverse relaxation as a contrast. The UF T2-T2 relaxation exchange spectroscopy (REXSY) method shortens the experiment time by one to two orders of magnitude compared to its conventional counterpart. Contrary to the conventional EXSY, the exchanging sites are distinguished based on T2 relaxation times instead of chemical shifts, making the method especially useful for systems including physical exchange of molecules. Therefore, the UF REXSY method offers an efficient means for quantification of exchange processes in various fields such as cellular metabolism and ion transport in electrolytes. As a proof of principle, we studied a halogen-free orthoborate based ionic liquid system and followed molecular exchange between molecular aggregates and free molecules. The results are in good agreement with the conventional exchange studies. Due to the single-scan nature, the method potentially significantly facilitates the use of modern hyperpolarization techniques to boost the sensitivity by several orders of magnitude.

High-purity lignin fractions and nanospheres rich in phenolic hydroxyl and carboxyl groups isolated with alkaline deep eutectic solvent from wheat straw.

A combined pretreatment based on alkaline deep eutectic solvent (DES) of K2CO3 and glycerol and sequential acid fractionation was developed to extract reactive lignin from wheat straw biomass. This process exhibited excellent purification performance in lignin isolation, and the lignin fractionated at low pH displayed high reactivity, having hydroxyl and carboxyl groups up to 9.60 and 2.52 mmol/g, respectively. Silica was selectively separated and removed during the precipitation stage, avoiding the "silica interference". Moreover, DES-lignin nanospheres created by self-assembly using lignin fractions obtained by acid precipitation possessed a high zeta potential, large particle size and high content of hydrophilic groups. Overall, the findings related to the dissociation mechanism and fractionation of reactive lignin during alkaline DES pretreatment and the acid sequence precipitation are crucial for facilitating lignin valorization in high-added value products.

Sensitive, Efficient and Portable Analysis of Molecular Exchange Processes by Hyperpolarized Ultrafast NMR.

Molecular exchange processes are ubiquitous in nature. Here, we introduce a method to analyze exchange processes by using low-cost, portable, single-sided NMR instruments. The inherent magnetic field inhomogeneity of the single-sided instruments is exploited to achieve diffusion contrast of exchange sites and spatial encoding of 2D data. This so-called ultrafast diffusion exchange spectroscopy method shortens the experiment time by two to four orders of magnitude. Furthermore, because full 2D data are measured in a single scan (in a fraction of a second), the sensitivity of the experiment can be improved by several orders of magnitude using so-called nuclear spin hyperpolarization methods (in this case, dissolution dynamic nuclear polarization). As the first demonstration of the feasibility of the method in various applications, we show that the method enables quantification of intra- and extracellular exchange of water in a yeast cell suspension.

Hyper-CEST NMR of metal organic polyhedral cages reveals hidden diastereomers with diverse guest exchange kinetics.

Guest capture and release are important properties of self-assembling nanostructures. Over time, a significant fraction of guests might engage in short-lived states with different symmetry and stereoselectivity and transit frequently between multiple environments, thereby escaping common spectroscopy techniques. Here, we investigate the cavity of an iron-based metal organic polyhedron (Fe-MOP) using spin-hyperpolarized 129Xe Chemical Exchange Saturation Transfer (hyper-CEST) NMR. We report strong signals unknown from previous studies that persist under different perturbations. On-the-fly delivery of hyperpolarized gas yields CEST signatures that reflect different Xe exchange kinetics from multiple environments. Dilute pools with ~ 104-fold lower spin numbers than reported for directly detected hyperpolarized nuclei are readily detected due to efficient guest turnover. The system is further probed by instantaneous and medium timescale perturbations. Computational modeling indicates that these signals originate likely from Xe bound to three Fe-MOP diastereomers (T, C3, S4). The symmetry thus induces steric effects with aperture size changes that tunes selective spin manipulation as it is employed in CEST MRI agents and, potentially, impacts other processes occurring on the millisecond time scale.

Hyperpolarised NMR to aid molecular profiling of electronic cigarette aerosols.

Signal amplification by reversible exchange (SABRE) hyperpolarisation is used to enhance the NMR signals of nicotine and acrolein in methanol-d4 solutions of electronic cigarette aerosols. Consequently, detection of 74 µM nicotine is possible in just a single scan 1H NMR spectrum. The first example of an aldehyde hyperpolarised using SABRE is demonstrated and we work towards novel real-world applications of SABRE-hyperpolarised NMR for chemical analysis.

Diffusion measurements of hydrocarbons in H-MCM-41 extrudates with pulsed-field gradient nuclear magnetic resonance spectroscopy.

Mesoporous materials are promising catalysts for production of biofuels. Herein, H-MCM-41 catalysts with different concentrations of the silica Bindzil binder (10-50 wt%) were prepared and characterized using pulsed-field gradient (PFG) NMR in the powder form and as extrudates. Effective diffusion coefficients (De) are measured in all cases. Diffusivities of n-hexadecane were found smaller for extrudates as compared to the powder catalysts. The estimates of diffusive tortuosity were also determined. PFG NMR data showed one major component that reveals diffusion in interconnected meso- and micropores and one other minor component (1-2%) that may correspond to more isolated pores or may represent complex effects of restricted diffusion. Therefore, several approaches including initial slope analysis of spin-echo attenuation curves, two-component fitting and Laplace inversion were used to discuss different aspects of diffusional transport in the studied H-MCM-41 materials. Correlations between De and the amount of Bindzil, the specific surface area, the micropore volume, the particle size, the total acid sites and the Lewis acid sites are discussed.

Parahydrogen-Induced Polarization in Hydrogenation Reactions Mediated by a Metal-Free Catalyst.

We report nuclear spin hyperpolarization of various alkenes achieved in alkyne hydrogenations with parahydrogen over a metal-free hydroborane catalyst (HCAT). Being an intramolecular frustrated Lewis pair aminoborane, HCAT utilizes a non-pairwise mechanism of H2 transfer to alkynes that normally prevents parahydrogen-induced polarization (PHIP) from being observed. Nevertheless, the specific spin dynamics in catalytic intermediates leads to the hyperpolarization of predominantly one hydrogen in alkene. PHIP enabled the detection of important HCAT-alkyne-H2 intermediates through substantial 1 H, 11 B and 15 N signal enhancement and allowed advanced characterization of the catalytic process.

Ultrafast methods for relaxation and diffusion.

Relaxation and diffusion NMR measurements offer an approach to studying rotational and translational motion of molecules non-invasively, and they also provide chemical resolution complementary to NMR spectra. Multidimensional experiments enable the correlation of relaxation and diffusion parameters as well as the observation of molecular exchange phenomena through relaxation or diffusion contrast. This review describes how to accelerate multidimensional relaxation and diffusion measurements significantly through spatial encoding. This so-called ultrafast Laplace NMR approach shortens the experiment time to a fraction and makes even single-scan experiments possible. Single-scan experiments, in turn, significantly facilitate the use of nuclear spin hyperpolarization methods to boost sensitivity. The ultrafast Laplace NMR method is also applicable with low-field, mobile NMR instruments, and it can be exploited in many disciplines. For example, it has been used in studies of the dynamics of fluids in porous materials, identification of intra- and extracellular metabolites in cancer cells, and elucidation of aggregation phenomena in atmospheric surfactant solutions.

Ultrafast Laplace NMR to study metal-ligand interactions in reversible polarisation transfer from parahydrogen.

Laplace Nuclear Magnetic Resonance (NMR) can determine relaxation parameters and diffusion constants, giving valuable information about molecular structure and dynamics. Information about relaxation times (T1 and T2) and the self-diffusion coefficient (D) can be extracted from exponentially decaying NMR signals by performing a Laplace transform, which is a different approach to traditional NMR involving Fourier transform of a free induction decay. Ultrafast Laplace NMR uses spatial encoding to collect the entire data set in just a single scan which provides orders of magnitude time savings. In this work we use ultrafast Laplace NMR D-T2 correlation sequences to measure key relaxation (T2) and diffusion (D) parameters of methanolic solutions containing pyridine. For the first time we combine this technique with the hyperpolarisation technique Signal Amplification By Reversible Exchange (SABRE), which employs an iridium catalyst to reversibly transfer polarisation from parahydrogen, to boost the 1H NMR signals of pyridine by up to 300-fold. We demonstrate use of ultrafast Laplace NMR to monitor changes in pyridine T2 and D associated with ligation to the iridium SABRE catalyst and kinetic isotope exchange reactions. The combined 1440-fold reduction in experiment time and 300-fold 1H NMR signal enhancement allow the determination of pyridine D coefficients and T2 values at 25 mM concentrations in just 3 seconds using SABRE hyperpolarised ultrafast Laplace NMR.

Identification of extracellular nanoparticle subsets by nuclear magnetic resonance.

Exosomes are a subset of secreted lipid envelope-encapsulated extracellular vesicles (EVs) of 50-150 nm diameter that can transfer cargo from donor to acceptor cells. In the current purification protocols of exosomes, many smaller and larger nanoparticles such as lipoproteins, exomers and microvesicles are typically co-isolated as well. Particle size distribution is one important characteristics of EV samples, as it reflects the cellular origin of EVs and the purity of the isolation. However, most of the physicochemical analytical methods today cannot illustrate the smallest exosomes and other small particles like the exomers. Here, we demonstrate that diffusion ordered spectroscopy (DOSY) nuclear magnetic resonance (NMR) method enables the determination of a very broad distribution of extracellular nanoparticles, ranging from 1 to 500 nm. The range covers sizes of all particles included in EV samples after isolation. The method is non-invasive, as it does not require any labelling or other chemical modification. We investigated EVs secreted from milk as well as embryonic kidney and renal carcinoma cells. Western blot analysis and immuno-electron microscopy confirmed expression of exosomal markers such as ALIX, TSG101, CD81, CD9, and CD63 in the EV samples. In addition to the larger particles observed by nanoparticle tracking analysis (NTA) in the range of 70-500 nm, the DOSY distributions include a significant number of smaller particles in the range of 10-70 nm, which are visible also in transmission electron microscopy images but invisible in NTA. Furthermore, we demonstrate that hyperpolarized chemical exchange saturation transfer (Hyper-CEST) with 129Xe NMR indicates also the existence of smaller and larger nanoparticles in the EV samples, providing also additional support for DOSY results. The method implies also that the Xe exchange is significantly faster in the EV pool than in the lipoprotein/exomer pool.

Seaweed-Derived Alginate-Cellulose Nanofiber Aerogel for Insulation Applications.

The next generation of green insulation materials is being developed to provide safer and more sustainable alternatives to conventional materials. Bio-based cellulose nanofiber (CNF) aerogels offer excellent thermal insulation properties; however, their high flammability restricts their application. In this study, the design concept for the development of a multifunctional and non-toxic insulation material is inspired by the natural composition of seaweed, comprising both alginate and cellulose. The approach includes three steps: first, CNFs were separated from alginate-rich seaweed to obtain a resource-efficient, fully bio-based, and inherently flame-retardant material; second, ice-templating, followed by freeze-drying, was employed to form an anisotropic aerogel for effective insulation; and finally, a simple crosslinking approach was applied to improve the flame-retardant behavior and stability. At a density of 0.015 g cm-3, the lightweight anisotropic aerogels displayed favorable mechanical properties, including a compressive modulus of 370 kPa, high thermal stability, low thermal conductivity (31.5 mW m-1 K-1), considerable flame retardancy (0.053 mm s-1), and self-extinguishing behavior, where the inherent characteristics were considerably improved by crosslinking. Different concentrations of the crosslinker altered the mechanical properties, while the anisotropic structure influenced the mechanical properties, combustion velocity, and to some extent thermal conductivity. Seaweed-derived aerogels possess intrinsic characteristics that could serve as a template for the future development of sustainable high-performance insulation materials.

High-Resolution Reconstruction for Multidimensional Laplace NMR.

As a perfect complement to conventional NMR that aims for chemical structure elucidation, Laplace NMR constitutes a powerful technique to study spin relaxation and diffusion, revealing information on molecular motions and spin interactions. Different from conventional NMR adopting Fourier transform to deal with the acquired data, Laplace NMR relies on specially designed signal processing and reconstruction algorithms resembling the inverse Laplace transform, and it generally faces severe challenges in cases where high spectral resolution and high spectral dimensionality are required. Herein, based on the tensor technique for high-dimensional problems and the sparsity assumption, we propose a general method for high-resolution reconstruction of multidimensional Laplace NMR data. We show that the proposed method can reconstruct multidimensional Laplace NMR spectra in a high-resolution manner for exponentially decaying relaxation and diffusion data acquired by commercial NMR instruments. Therefore, it would broaden the scope of multidimensional Laplace NMR applications.

Comparison of Lignin Fractions Isolated from Wheat Straw Using Alkaline and Acidic Deep Eutectic Solvents.

This study aims to examine the characteristics of two solid lignin fractions isolated from wheat straw using alkaline and acidic deep eutectic solvents (DESs). The chemical properties and morphological characteristics of the two lignin fractions were evaluated by measuring their purity, elemental composition, molecular weight and particle size distributions, and microstructure. Their chemical structure was evaluated using DRIFT (diffuse reflectance infrared Fourier transform) spectroscopy, GPC (gel permeation chromatography), TGA (thermogravimetric analysis), 13C NMR (nuclear magnetic resonance), 31P NMR, and HSQC NMR. Our findings showed that the lignin isolated using alkaline DESs was less pure and had a smaller particle size, higher molecular weight, and thermal stability compared to the lignin isolated using acidic DESs. Their lignin structure was also determined to be different due to varying selective fractures on the linkages of lignin. These results suggest that the DES treatments could selectively extract lignin from wheat straw with different yields, compositions, morphologies, and structures, which could then provide a theoretical basis for the selection of DESs for specially appointed lignin extraction.

Ultrafast diffusion exchange nuclear magnetic resonance.

The exchange of molecules between different physical or chemical environments due to diffusion or chemical transformations has a crucial role in a plethora of fundamental processes such as breathing, protein folding, chemical reactions and catalysis. Here, we introduce a method for a single-scan, ultrafast NMR analysis of molecular exchange based on the diffusion coefficient contrast. The method shortens the experiment time by one to four orders of magnitude. Consequently, it opens the way for high sensitivity quantification of important transient physical and chemical exchange processes such as in cellular metabolism. As a proof of principle, we demonstrate that the method reveals the structure of aggregates formed by surfactants relevant to aerosol research.

Accelerating Restricted Diffusion NMR Studies with Time-Resolved and Ultrafast Methods.

Restricted diffusion of fluids in porous materials can be studied by pulsed field gradient nuclear magnetic resonance (NMR) non-invasively and without tracers. If the experiment is repeated many times with varying diffusion delays, detailed information about pore sizes and tortuosity can be recorded. However, the measurements are very time-consuming because numerous repetitions are needed for gradient ramping and varying diffusion delays. In this paper, we demonstrate two different strategies for acceleration of the restricted diffusion NMR measurements: time-resolved diffusion NMR and ultrafast Laplace NMR. The former is based on time-resolved non-uniform sampling, while the latter relies on spatial encoding of two-dimensional data. Both techniques allow similar 1-2 order of magnitude acceleration of acquisition, but they have different strengths and weaknesses, which we discuss in detail. The feasibility of the methods was proven by investigating restricted diffusion of water inside tracheid cells of thermally modified pine wood.

Correction to "Comprehensive NMR Analysis of Pore Structures in Superabsorbing Cellulose Nanofiber Aerogels".

[This corrects the article DOI: 10.1021/acs.jpcc.9b08339.].