Hyperpolarized 13C NMR Spectroscopy of Urine Samples at Natural Abundance by Quantitative Dissolution Dynamic Nuclear Polarization

Hyperpolarized nuclear magnetic resonance (NMR) offers an ensemble of methods that remarkably address the sensitivity issues of conventional NMR. Dissolution Dynamic Nuclear Polarization (d-DNP) provides a unique and general way to detect ¹³C NMR signals with a sensitivity enhanced by several orders of magnitude. The expanding application scope of d-DNP now encompasses the analysis of complex mixtures at natural ¹³C abundance. However, the application of d-DNP in this area has been limited to metabolite extracts. Here, we report the first d-DNP-enhanced ¹³C NMR analysis of a biofluid -urine- at natural abundance, offering unprecedented resolution and sensitivity for this challenging type of sample. We also show that accurate quantitative information on multiple targeted metabolites can be retrieved through a standard addition procedure.     

Carbon-13 Nuclear Magnetic Resonance Study of 5′-Substituted-5′-Oeoxyadenosine Derivatives

Abstract

A series of 5′-substituted-5′-deoxyadenosina derivatives, potential inhibitors of methyltransferases were synthesized and their carbon～13 nuclear magnetic resonance spectra investigated.     

Studies of Molecular Motions of Ocotillol-Type Saponins in Solution by ~(13)C Nuclear Magnetic Relaxation

In this paper, the fully anisotropicoverall tumbling motions and side groups internal rotation of ocotillol-type saponins separated from the leaves of Panax Quidquefolium L. are investigated by ~(13)C nuclear magnetic relaxation. The fully anisotropic overall tumbling motion model with methyl conformation jumps internal rotation among three equivalent sites is presented, and the spectral density function of this model is derived. The rotation rates for overall tumbling motions to ocotillol-type saponins (OTS) are computed by Woessner's fully anisotropic overall tumbling motion model, and the internal rotation rate and barrier for side groups in OTS are calculated using free diffusion internal rotation model, restriction diffusion internal rotation model and conformation jumps internal rotation model, respectively.     

Study of entangled network formation in concentrated solutions of polymer by ~(13)C NMR

Information about the exact location of topological and cohesional entanglements at molecular level has been obtained by 13C NMR relaxation analysis. The results show that about 20% of the carbon atoms in the main chain are entangled in the 25% (by weight) solution, which is independent of the content of the 1,2-segment in polybutadiene and of the kind of solvent. However, the entanglement of the carbon atone on the end group of the side chain is very weak, they behave as slipping freely at the junctions.     

Development of Macrocyclic Mn(II)−Bispyridine Complexes as pH-Responsive Magnetic Resonance Imaging Contrast Agents

In this study, macrocyclic complexes of Mn (II) with pH-responsive relaxivities were developed. The ligands were based on the rigid MPi platform, which was composed of a triazacyclononane (TACN) macrocycle and two picolyl pendant arms. An aryl sulfonamide moiety introduced in the skeleton of MPi endowed the final Mn complex (Mn−MPA) with pH-sensitive relaxivities. Mn−MPA showed a ∼3-fold increase in longitudinal relaxivity associated with the protonation and decoordination of the sulfonamide group in an acidic environment. We found that it was feasible to tune the pH window of the complexes by choosing proper substituents on the aromatic ring or modifying on the picolyl pendants. Considering the good kinetic inertness, rapid and efficient response to pH stimuli, and easy modulation characteristics, Mn−MPAs can be considered ideal candidates for use as acidic microenvironment-specific MRI contrast agents.     

Study of phase transitions in 4-n-alkoxybenzilidene-4′-n-alkylanilines by one and two dimensional 13C NMR

Abstract

The phase transition of a series of homologous liquid-crystalline compounds, nO.m (4-n-alkoxybenzilidene-4′-n-alkylanilines), from the nematic phase to the smectic A phase has been studied by ¹³C NMR. The order parameters, determined by a two dimensional technique called separated local field spectroscopy combined with off-magic angle spinning, of different molecular segments of these compounds are related linearly to the ¹³C chemical shifts. Changes in the order parameters of the phenyl rings as well as those of the chains during the S_A–N transition depend on the nature of the phase transition. These changes are quantitatively related to the McMillan ratio, which is defined as the ratio between the S_A–N transition temperature (T _SAN) and the nematic to isotropic transition temperature (T _NI), i.e. M = T _SAN/T _NI. The S_A–N transition is first order for M > M _TCP, and second order for M < M _TCP, where TCP is the tricritical point. The value of M _TCP was found to be 0·958 ± 0·004, in excellent agreement with that obtained from spin probe studies (0·959 ± 0·005) reported by Freed and co-workers [1].     

Synthesis and Evaluation of Aromatic DTPA-bis(amide) Gadolinium Complexes as Magnetic Resonance Imaging Contrast Agents

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Energetics of C–C Bond Scission in Ethane Hydrogenolysis: A Theoretical Study of Possible Intermediates and Reaction Pathways

C–C bond scission steps, which are often considered as rate-determining in ethane hydrogenolysis, are studied by the Unity Bond Index–Quadratic Exponential UBI–QEP method. The binding energies of atomic carbon with Group VIII and IB metal surfaces Ni(111), Pd(111), Pt(111), Rh(111), Ru(001), Ir(111), Fe(110), Cu(111), and Au(111) are estimated using experimental data on the adsorption of various species on these surfaces. These estimates are corrected using data from density functional theory (DFT) on the adsorption heats of the CH _x species. Metal surfaces are arranged in the following series according to the binding strength of a carbon atom: Cu(111) < Au(111) < Pd(111) < Ru(001) Pt(111) < Ni(111) Rh(111) < Ir(111) < Fe(110). The values of chemisorption heats range from 121 kcal/mol for Au(111) to 193 kcal/mol for Fe(110). The activity of these surfaces toward C–C bond scission increases in the same series. The results of this work suggest that the most probable C–C bond scission precursors are ethyl, ethylidyne, adsorbed acetylene, CH₂CH, CH₂C, and CHC. Theoretical data obtained by different methods are compared and found to agree well with each other. An overview of experimental data on ethane hydrogenolysis mechanisms is given.     

The Effect of Gadolinium Doping in [13C6,2H7]Glucose Formulations on 13C Dynamic Nuclear Polarization at 3.35 T

The promise of hyperpolarized glucose as a non-radioactive imaging agent capable of reporting on multiple metabolic routes has led to recent advances in its dissolution-DNP (dDNP) driven polarization using UV-light induced radicals and trityl radicals at high field (6.7 T) and 1.1 K. However, most preclinical dDNP polarizers operate at the field of 3.35 T and 1.4–1.5 K. Minute amounts of Gd³⁺ complexes have shown large improvements in solid-state polarization, which can be translated to improved hyperpolarization in solution. However, this Gd³⁺ effect seems to depend on magnetic field strength, metal ion concentration, and sample formulation. The effect of varying Gd³⁺ concentrations at 3.35 T has been described for ¹³C-labeled pyruvic acid and acetate. However, it has not been studied for other compounds at this field. The results presented here suggest that Gd³⁺ doping can lead to various concentration and temperature dependent effects on the polarization of [¹³C₆,²H₇]glucose, not necessarily similar to the effects observed in pyruvic acid or acetate in size or direction. The maximal polarization for [¹³C₆,²H₇]glucose appears to be at a Gd³⁺ concentration of 2 mM, when irradiating for more than 2 h at the negative maximum of the DNP intensity profile. Surprisingly, for shorter irradiation times, higher polarization levels were determined at 1.50 K compared to 1.45 K, at a [Gd³⁺]=1.3 mM. This was explained by the build-up time constant and maximum at these temperatures.     

Aptamer–Target Interaction: A Comprehensive Study by Microchip Electrophoresis in Frontal Mode

An original methodology based on microchip electrophoresis in a continuous frontal analysis mode (FACMCE) was employed to provide new insights into the interaction between an aptamer and its target (lysozyme), i.e., the influence of various experimental conditions on possible conformation change of the aptamer. The parameters evaluated were: background electrolyte (BGE) nature and ionic strength, nature and concentration of an added divalent cation, use of an aptamer thermal treatment, conditions classically used when employing aptamers. Increasing the BGE ionic strength led to a decrease in the dissociation constant highlighting the role played by the non-ionic interactions, and a decrease in the number of binding sites due to a change of binding mode and/or an amplification of selectivity. Divalent cation addition in the BGE improved the binding affinity. We demonstrated that this is not exclusively related to an increase in ionic strength but also certainly to an aptamer conformational change. Furthermore, changing the BGE nature permitted to modulate the binding parameters. Finally, we showed that an initial heating of the aptamer solution has been proved critical to stabilize the optimal conformation and thus to get a high binding affinity as well as a smaller standard deviation on the binding parameters values. This study has evidenced the influence of a wide range of parameters so as to better grasp the target/aptamer interaction, and thus highlight some phenomena involved in such an interacting system. This work will therefore help for further applications of this binding system for selectivity improvement in bioanalytical development.     

seco-Labdanes: A Study of Terpenoid Structural Diversity Resulting from Biosynthetic C−C Bond Cleavage

The cleavage of a C−C bond is a complexity generating process, which complements oxidation and cyclisation events in the biosynthesis of terpenoids. This process leads to increased structural diversity in a cluster of related secondary metabolites by modification of the parent carbocyclic core. In this review, we highlight the diversifying effect of C−C bond cleavage by examining the literature related to seco-labdanes—a class of diterpenoids arising from such C−C bond cleavage events.     

Solute-Solvent Interactions in Solutions of Solvatochromic Phenoxides: A Dynamics Simulation Study

Solute-solvent interactions in protic (water, methanol and 2-propanol) and aprotic (DMSO) solvents of four solvatochromic phenoxides, the 4- and 2-pyridiniophenoxides, Brooker’s merocyanine and the N-methyloxyquinolinium betaine, were investigated with the aid of molecular dynamics simulations. Although the size of the first solvation shell of the phenoxide oxygen of all betaines remains constant in the three protic solvents, it comprises increasingly fewer solvent molecules as the volume of the hydroxylic solvent increases. In DMSO, the donor phenoxide group of the 4-pyridiniophenoxide betaine is loosely solvated, leading to an internal charge-transfer with smaller transition energies than in protic media.     

Inclusion of Polyphenol Oxidase Substrates in β-Cyclodextrin: A 1 H-NMR Study

A ¹H-NMR study of the interactionsbetween -cyclodextrin (-CD) and included phenolic molecules (chloragenic acid and caffeic acid ) in aqueous medium is reported. The results confirm that inclusion occurs. Data analysis by the continuous variation method shows that all the complexes have 1 : 1 stoichiometries. Values for the apparent association constants of the inclusion compounds are estimated and compared with previously reported values.     

Study of early stages of amyloid Aβ13-23 formation using molecular dynamics simulation in implicit environments

β-amyloid aggregation and formation of senile plaques is one of the hallmarks of Alzheimer’s disease (AD). It leads to degeneration of neurons and decline of cognitive functions. The most aggregative and toxic form of β-amyloid is Aβ_1-42 but in experiments, the shorter forms able to form aggregates are also used. The early stages of amyloid formation are of special interest due to the influence of this peptide on progression of AD. Here, we employed nine helices of undecapeptide Aβ_13-23 and studied progress of amyloid formation using 500 ns molecular dynamics simulation and implicit membrane environment. The small β-sheets emerged very early during simulation as separated two-strand structures and a presence of the membrane facilitated this process. Later, the larger β-sheets were formed. However, the ninth helix which did not form paired structure stayed unchanged till the end of MD simulation. Paired helix–helix interactions seemed to be a driving force of β-sheet formation at early stages of amyloid formation. Contrary, the specific interactions between α-helix and β-sheet can be very stable and be stabilized by the membrane.     

H-Nuclear Magnetic Resonance and Phase Solubility Studies of the Stoichiometries in 2,4-D: α- and β-Cyclodextrins Inclusion Complexes

The interaction in solution between2,4-dichlorophenoxyacetic acid with - and-cyclodextrins was evaluated by phasesolubility studies. Association constants werecalculated by this technique. The stoichiometries were1 : 2 and 1 : 1 for the - and -cyclodextrincomplexes, respectively. In order to corroborate thecomplexation and the knowledge of structural aspectsof the host : guest interaction, proton nuclearmagnetic resonance (¹H-NMR) spectroscopy wasemployed. The application of the continuous variationtechnique corroborated the calculated complexstoichiometries by solubility assays. ComplementaryNOE studies were applied in order to corroborate theproposed complex structures.     

Revealing protein structures in solid-phase peptide synthesis by 13C solid-state NMR: evidence of excessive misfolding for Alzheimer's β

Solid-phase peptide synthesis (SPPS) is a widely used technique in biology and chemistry. However, the synthesis yield in SPPS often drops drastically for longer amino acid sequences, presumably because of the occurrence of incomplete coupling reactions. The underlying cause for this problem is hypothesized to be a sequence-dependent propensity to form secondary structures through protein aggregation. However, few methods are available to study the site-specific structure of proteins or long peptides that are anchored to the solid support used in SPPS. This study presents a novel solid-state NMR (SSNMR) approach to examine protein structure in the course of SPPS. As a useful benchmark, we describe the site-specific SSNMR structural characterization of the 40-residue Alzheimer's β-amyloid (Aβ) peptide during SPPS. Our 2D (13)C/(13)C correlation SSNMR data on Aβ(1-40) bound to a resin support demonstrated that Aβ underwent excessive misfolding into a highly ordered β-strand structure across the entire amino acid sequence during SPPS. This approach is likely to be applicable to a wide range of peptides/proteins bound to the solid support that are synthesized through SPPS.     

Identification of Indigo Dyes in Painting Layers by Pyrolysis Methylation and Silylation. A Case Study: “The Dinner of Emmaus” by G. Preti

Reactive pyrolysis, under methylating or silylating conditions, in combination with gas chromatography-mass spectrometry (GC-MS) was evaluated as an analytical method for the detection of indigo dyes in painting layers. Samples with the addition of tetramethylammonium hydroxide (TMAH) or hexamethyldisilazane (HMDS), for pyrolysis/methylation and pyrolysis/silylation experiments, respectively, were pyrolysed at 600 °C by means of a heated filament pyrolyser and the evolved products were analysed on-line by GC-MS. Methyl and silylated pyrolytic markers, related to 2-aminobenzoic acid and the indoxyl moiety, were established from the analysis of synthetic indigo, neat or as a pigment of artificial painting layers containing a siccative oil. The occurrence of these markers was investigated in a real sample, taken from the painting The dinner of Emmaus by G.Preti (XVII century), in order to identify the blu pigment. Positive identification of indigo was achieved by pyrolysis/silylation and confirmed by RAMAN spectroscopy.     

A New Approach to C60-Containing Polyphosphazenes by Polymerization of Phosphonitrile Chloride Trimer in the Presence of C60

A novel synthetic strategy was developed to prepare polyphosphazenes containing C60 moieties. Thus, the phosphonitrile chloride trimer underwent thermal ring-opening polymerization （ROP） in the presence of C60 molecules to yield the reactive macromolecular intermediate, C60-containing poly（dichlorophosphazene）. And then, the other groups could be linked to the phosphazene backbone by nucleophilic substitution reaction of the chlorine atoms in this intermediate to produce a series of C60-containing polyphosphazene. The polymer exhibits good solubility in common organic solvents and is thermally stable.     

Study of the copper(II)-Aztreonam™ system by potentiometry and spectrophotometry,and structural characterization by 13C NMR relaxation

Potentiometric and spectrophotometric results obtained for the Cu(II)-Aztreonam system give a clear indication of the cheating properties of this antibiotic toward the Cu²⁺ ion, coordinated already at pH ≌ 2. Only one single complex of 1:1 stoichiometry is formed. Measurements of ¹³C relaxation rates allow us to attribute a coordination scheme via the carboxylic and aminic groups.