High-resolution NMR Spectroscopy Research Articles

Electrochemical Depolymerization of Lignin in a Biomass-based Solvent.

Breaking down lignin into smaller units is the key to generate high value‐added products. Nevertheless, dissolving this complex plant polyphenol in an environment‐friendly way is often a challenge. Levulinic acid, which is formed during the hydrothermal processing of lignocellulosic biomass, has been shown to efficiently dissolve lignin. Herein, levulinic acid was evaluated as a medium for the reductive electrochemical depolymerization of the lignin macromolecule. Copper was chosen as the electrocatalyst due to the economic feasibility and low activity towards the hydrogen evolution reaction. After depolymerization, high‐resolution mass spectrometry and nuclear magnetic resonance spectroscopy revealed lignin‐derived monomers and dimers. A predominance of aryl ether and phenolic groups was observed. Depolymerized lignin was further evaluated as an anti‐corrosion coating, revealing enhancements on the electrochemical stability of the metal. Via a simple depolymerization process of biomass waste in a biomass‐based solvent, a straightforward approach to produce high value‐added compounds or tailored biobased materials was demonstrated.

NMR-Based Metabolomics Identify Metabolic Change in Spleen of Idiopathic Thrombocytopenic Purpura Patients.

Idiopathic thrombocytopenic purpura (ITP) is a common hematological disease and the abnormal platelet destruction in the spleen is a critical pathological mechanism for ITP. However, the metabolomic change in the spleen caused by ITP is still unclear. In the present study, the metabolomic information of 18 ITP and 20 normal spleen samples were detected by using 1H high-resolution magic angle spinning NMR spectroscopy (1H MAS NMR). Compared with normal spleen, the concentrations of acetate, alanine, glutamine, glycerol, isoleucine, lysine, valine, phenylalanine, leucine, and methanol in ITP spleen tissue were elevated and 3-hydroxybutyric acid, ascorbate, asparagine, ethanol, glycogen, low-density lipoprotein, malonate, myo-inositol, glycerophosphocholine, pyroglutamate, and taurine were decreased. Amino acids metabolic pathways, such as branched-chain amino acids pathway, were identified as the main involved pathways based on enrichment analysis. The decrease in taurine level in the spleen was the most obvious metabolic signature involving ITP with high sensitivity and specificity to distinguish the spleen of ITP from the normal (CI: 0.825–0.982). Notably, the level of taurine in the spleen was negatively correlated with the efficacy of splenectomy (r = 0.622, p = 0.006). Collectively, the data from our study revealed previously unknown ITP-related metabolomic changes in the spleen and found a potential diagnostic and efficacy-predictive biomarker for ITP treatment.

N-[3-(Chloromethyl)-1,2-benzisoxazol-5-yl]acetamide

Functionally substituted 1,2-benzisoxazoles are very important and promising heterocycles with various pharmacological activities. Benzoxazoles containing reactive 3-chloromethyl and 5-amino groups are practically unexplored derivatives in this series. In this communication, the simple method for the synthesis of N-[3-(chloromethyl)-1,2-benzisoxazol-5-yl]acetamide which is an interesting precursor for the preparation of a series of 3,5-disubstituted benzoxazoles was described. The structure of the synthesized compound was established by elemental analysis, high-resolution mass spectrometry, 1H, 13C NMR and IR spectroscopy, and mass spectrometry.

Multiphase Drug Distribution and Exchange in Oil-in-Water Nanoemulsion Revealed by High-Resolution 19F qNMR.

An oil-in-water (o/w) nanoemulsion (NE), composed of oil globules, stabilized by a surfactant, and dispersed in an aqueous phase, is increasingly developed in complex drug formulation. Kinetically stable NEs are used to formulate hydrophobic drugs and typically provide higher dosage strengths and better content uniformity. However, little is known accurately about drug distribution in its multiphase solution, especially for the possible drug presence in the surfactant (s) phase, the interface layer between the dispersed oil (o) and the continuous water (w) phases. Here, high-resolution 19F quantitative NMR spectroscopy was applied directly and noninvasively on an o/w NE drug product containing difluprednate (DFPN). The well-resolved 19F peaks of DFPN depended on the shielding molecules in each phase, which revealed mass-balanced DFPN distribution in multiple phases of (w), (s), and (o) of NE globules at a quantity of 1.8 ± 0.1, 35 ± 2, and 59 ± 3% per labeled content, respectively. Furthermore, the dilution-dependent 19F peak line broadening and shift suggested a millisecond dynamic exchange between the NE and the less-noticed smaller but thermodynamically stable microemulsion (ME) globules in NE solution. The high-resolution NMR result revealed that the drug availability could be quickly achieved using an o/w NE formulation because of the drug multiphase distribution and the ME-assisted fast drug exchange among globules.

Recent advances on the stability of dietary polyphenols

Recent advances on the stability of dietary polyphenols

New chromone analog and pyrrole alkaloid produced by Penicillium sclerotiorum and their antibacterial activity

A new chromone analog (1) and a new pyrrole alkaloid (2), together with four known compounds, were isolated from the endophytic fungus Penicillium sclerotiorum MPT-250 obtained from the stems of Taxus wallichiana var. chinensis (Pilger) Florin. The structural elucidation of these metabolites was performed by high-resolution mass spectrometry and NMR spectroscopy. Compounds 1 and 5 exhibited significant antibacterial activity against carbapenems-resistant Pseudomonas aeruginosa and multidrug-resistant Enterococcus faecium with an minimum inhibitory concentration (MIC) value of 3.13 μg/ml respectively.

5,5′-Thiobis(4-chloro-3H-1,2-dithiole-3-thione)

3H-1,2-Dithiole-3-thiones are important compounds with many types of significant pharmacological activity. Although many derivatives of this class have been described in the literature, their thioethers have not previously been obtained. In this communication, it is shown that the reaction of 4,5-dichloro-3H-1,2-dithiole-3-thione with potassium isocyanate unexpectedly gave 5,5′-thiobis(4-chloro-3H-1,2-dithiole-3-thione). The structure of the synthesized compound was established by elemental analysis, high-resolution mass spectrometry, 13C NMR and IR spectroscopy, and mass spectrometry.

Metabolomic analysis of endometrial cancer by high-resolution magic angle spinning NMR spectroscopy.

To analyze endometrial metabolite profiles between patients with endometrial cancer and controls. Seventeen (17) women with endometrium cancer and 18 controls were enrolled in this study. 1H HR-MAS (High Resolution-Magic Angle Spinning) NMR (Nuclear Magnetic Resonance) spectroscopy data obtained from endometrial tissue samples of patients with endometrial cancer and control group were analyzed with bioinformatics methods. Principal component analysis (PCA) and the partial least squares discriminant analysis (PLS-DA) score plots obtained with the multivariate statistical analysis of pre-processed spectral data shows a separation between the samples from patients with endometrial cancer and controls. Analysis results suggest that the levels of lactate, glucose, o-phosphoethanolamine, choline, glycerophosphocholine, phosphocholine, leucine, isoleucine, valine, glutamate, glutamine, n-acetyltyrosine, methionine, taurine, alanine, aspartate and phenylalanine are increased in patients with endometrial cancer compared to the controls. The metabolomics signature of patients with endometrial cancer is different from that of benign endometrial tissue.

High-Resolution Magic-Angle Spinning NMR Spectroscopy for Evaluation of Cell Shielding by Virucidal Composites Based on Biogenic Silver Nanoparticles, Flexible Cellulose Nanofibers and Graphene Oxide.

Antiviral and non-toxic effects of silver nanoparticles onto in vitro cells infected with coronavirus were evaluated in this study using High-Resolution Magic-Angle Spinning Nuclear Magnetic Resonance (HR-MAS NMR) spectroscopy. Silver nanoparticles were designed and synthesized using an orange flavonoid—hesperetin (HST)—for reduction of silver(I) and stabilization of as obtained nanoparticles. The bio-inspired process is a simple, clean, and sustainable way to synthesize biogenic silver nanoparticles (AgNP@HST) with diameters of ∼20 nm and low zeta potential (−40 mV), with great colloidal stability monitored for 2 years. The nanoparticles were used for the fabrication of two types of antiviral materials: colloids (AgNP@HST spray) and 3D flexible nanostructured composites. The composites, decorated with AgNP@HST (0.05 mmol L−1), were made using cellulose nanofibers (CNF) obtained from orange peel and graphene oxide (GO), being denominated CNF@GO@AgNP@HST. Both materials showed high virucidal activity against coronaviruses in cell infection in vitro models and successfully inhibited the viral activity in cells. HR-MAS 1H-NMR technique was used for determining nanomaterials’ effects on living cells and their influences on metabolic pathways, as well as to study viral effects on cells. It was proven that none of the manufactured materials showed toxicity towards the intact cells used. Furthermore, viral infection was reverted when cells, infected with the coronavirus, were treated using the as-fabricated nanomaterials. These significant results open possibilities for antiviral application of 3D flexible nanostructured composite such as packaging papers and filters for facial masks, while the colloidal AgNP@HST spray can be used for disinfecting surfaces, as well as a nasal, mouth, and eye spray.

Temporal profile of serum metabolites and inflammation following closed head injury in rats is associated with HPA axis hyperactivity.

Closed head injury (CHI) causes neurological disability along with systemic alterations that can activate neuro-endocrine response through hypothalamic-pituitary-adrenal (HPA) axis activation. A dysregulated HPA axis function can lead to relocation of energy substrates and alteration in metabolic pathways and inflammation at the systemic level. Assessment of time-dependent changes in serum metabolites and inflammation after both mild and moderate CHI. Along with this, serum corticosterone levels and hypothalamic microglial response were observed. Rats underwent mild and moderate weight-drop injury and their serum and hypothalamus were assessed at acute, sub-acute and chronic timepoints. Changes in serum metabolomics were determined using high resolution NMR spectroscopy. Serum inflammatory cytokine, corticosterone levels and hypothalamic microglia were assessed at all timepoints. Metabolites including lactate, choline and branched chain amino acids were found as the classifiers that helped distinguish between control and injured rats during acute, sub-acute and chronic timepoints. While, increased αglucose: βglucose and TMAO: choline ratios after acute and sub-acute timepoints of mild injury differentiated from moderate injured rats. The injured rats also showed distinct inflammatory profile where IL-1β and TNF-α levels were upregulated in moderate injured rats while IL-10 levels were downregulated in mild injured rats. Furthermore, injury specific alterations in serum metabolic and immunologic profile were found to be associated with hyperactive HPA axis, with consistent increase in serum corticosterone concentration post injury. The hypothalamic microglia showed a characteristic activated de-ramified cellular morphology in both mild and moderate injured rats. The study suggests that HPA axis hyperactivity along with hypothalamic microglial activation led to temporal changes in the systemic metabolism and inflammation. These time dependent changes in the metabolite profile of rats can further strengthen the knowledge of diagnostic markers and help distinguish injury related outcomes after TBI.

Cembranoids From the Leaves of Nicotiana tabacum and Their Neuroprotective Activities

A new cembrane-type diterpenoid (1) and 9 known cembranoids (2-10) were isolated from the leaves of Nicotiana tabacum. The structure of the new metabolite was confirmed by extensive analyses of their spectroscopic data, including high-resolution electrospray ionization mass spectrometry (HR-ESI-MS), 1D- and 2D nuclear magnetic resonance (NMR) spectroscopy, and infrared spectroscopy (IR). All of the isolated compounds were tested for their protective effects against oxygen-glucose deprivation (OGD)-induced neurotoxicity in SH-SY5Y cells. The new compound 1 exhibited a moderate protective effect against OGD-induced neurotoxicity at 10 μM.

Magnetic Resonance Imaging (MRI) and MR Spectroscopic Methods in Understanding Breast Cancer Biology and Metabolism.

A common malignancy that affects women is breast cancer. It is the second leading cause of cancer-related death among women. Metabolic reprogramming occurs during cancer growth, invasion, and metastases. Functional magnetic resonance (MR) methods comprising an array of techniques have shown potential for illustrating physiological and molecular processes changes before anatomical manifestations on conventional MR imaging. Among these, in vivo proton (1H) MR spectroscopy (MRS) is widely used for differentiating breast malignancy from benign diseases by measuring elevated choline-containing compounds. Further, the use of hyperpolarized 13C and 31P MRS enhanced the understanding of glucose and phospholipid metabolism. The metabolic profiling of an array of biological specimens (intact tissues, tissue extracts, and various biofluids such as blood, urine, nipple aspirates, and fine needle aspirates) can also be investigated through in vitro high-resolution NMR spectroscopy and high-resolution magic angle spectroscopy (HRMAS). Such studies can provide information on more metabolites than what is seen by in vivo MRS, thus providing a deeper insight into cancer biology and metabolism. The analysis of a large number of NMR spectral data sets through multivariate statistical methods classified the tumor sub-types. It showed enormous potential in the development of new therapeutic approaches. Recently, multiparametric MRI approaches were found to be helpful in elucidating the pathophysiology of cancer by quantifying structural, vasculature, diffusion, perfusion, and metabolic abnormalities in vivo. This review focuses on the applications of NMR, MRS, and MRI methods in understanding breast cancer biology and in the diagnosis and therapeutic monitoring of breast cancer.

Magic angle spinning dynamic nuclear polarization solid-state NMR spectroscopy of γ-irradiated molecular organic solids

In the past 15 years, magic angle spinning (MAS) dynamic nuclear polarization (DNP) has emerged as a method to increase the sensitivity of high-resolution solid-state NMR spectroscopy experiments. Recently, γ-irradiation has been used to generate significant concentrations of homogeneously distributed free radicals in a variety of solids, including quartz, glucose, and cellulose. Both γ-irradiated quartz and glucose previously showed significant MAS DNP enhancements. Here, γ-irradiation is applied to twelve small organic molecules to test the applicability of γ-irradiation as a general method of creating stable free radicals for MAS DNP experiments on organic solids and pharmaceuticals. Radical concentrations in the range of 0.25 ​mM–10 ​mM were observed in irradiated glucose, histidine, malic acid, and malonic acid, and significant 1H DNP enhancements of 32, 130, 19, and 11 were obtained, respectively, as measured by 1H→13C CPMAS experiments. However, concentrations of free radicals below 0.05 ​mM were generally observed in organic molecules containing aromatic rings, preventing sizeable DNP enhancements. DNP sensitivity gains for several of the irradiated compounds exceed that which can be obtained with the relayed DNP approach that uses exogeneous polarizing agent solutions and impregnation procedures. In several cases, significant 1H DNP enhancements were realized at room temperature. This study demonstrates that in many cases γ-irradiation is a viable alternative to addition of stable exogenous radicals for DNP experiments on organic solids.

Chemical shielding and electric field gradient tensors of disodium 5′-adenosine triphosphate trihydrate determined by solid-state NMR spectroscopy and ab initio calculations

The tri-phosphate and sodium ion environments in well crystallized disodium 5′adenosine triphosphate trihydrate (ATP•3H2O) are investigated using high-resolution solid-state NMR spectroscopy and ab initio calculations. The six inequivalent phosphorus sites resolved in 31P MAS spectra have been assigned on the basis of experimentally obtained results and theoretical ab initio quantum chemical calculations of 31P chemical shielding tensors with increased numerical accuracy. Aided by 31P-31P dipolar connectivity established in DQ-SQ correlation experiment and DFT calculations carried out at the B3LYP/(aug-cc-pDVZ(3s,2p,1d), 6–31G(d,p)) level, phosphorus resonance assignments in MAS spectra have been made and the 31P chemical shielding anisotropies and tensor orientations in the molecule-fixed frame have been determined. 31P shielding tensors are found to be oriented with the most shielded direction nearly perpendicular to the O-P-O plane involving the two adjoining oxygens. The complete resolution of the four sodium sites has been achieved from 3Q-MAS experiments performed at 7.05 T and this has enabled 23Na quadrupole parameters to be determined experimentally and compared with theoretical calculations. Besides aiding a partial resonance assignment of the 23Na 3Q-MAS spectrum, our HF/6–311++G(2d,2p) calculations of 23Na EFG tensors show that for two sodium sites which are tightly coordinated the orientation of the EFG tensor is distinct. These exhibit unique directions along which the field gradient is the smallest and largest.

Identification of Pigmented Substances in Black Honey from Leucosceptrum canum: Novel Quinonoids Contribute to Honey Color.

Leucosceptrum canum (Lamiaceae) honey (LCH) is a rare and high-value honey with an unusual dark brown color. Its pigments may be related to the purported health benefits of the honey and could serve as indicators of authentic LCH. Here, we purified the pigmented substances contributing to LCH's color and identified them using high-resolution mass spectrometry and nuclear magnetic resonance spectroscopy. The major pigmented substances were 1-[4-(3-carboxy-propylamino)-3,6-dioxo-cyclohexa-1,4-dienyl]-pyrrolidine-2-carboxylic acid (GPBQ), 1-[4-(3-carboxy-propylamino)-3,6-dioxo-cyclohexa-1,4-dienyl]-pyrrolidine-2-carboxylic acid. (GAPBQ), and 2,5-di-(N-prolyl)-para-benzoquinone (DPBQ). According to UHPLC-DAD data recorded at 370 nm, GPBQ, GAPBQ, and DPBQ contents ranged from 2.58 to 5.13, 2.93 to 4.96, and 4.26 to 9.22 mg/kg, respectively. These amino acid-quinone conjugates, found here for the first time, are pigments that can serve as markers to control the quality of LCH.

Interfacing Liquid State Hyperpolarization Methods with NMR Instrumentation

Advances in liquid state hyperpolarization methods have enabled new applications of high-resolution NMR spectroscopy. Utilizing strong signal enhancements from hyperpolarization allows performing NMR spectroscopy at low concentration, or with high time resolution. Making use of the high, but rapidly decaying hyperpolarization in the liquid state requires new techniques to interface hyperpolarization equipment with liquid state NMR spectrometers. This article highlights rapid injection, high resolution NMR spectroscopy with hyperpolarization produced by the techniques of dissolution dynamic nuclear polarization (D-DNP) and para-hydrogen induced polarization (PHIP). These are popular, albeit not the only methods to produce high polarization levels for liquid samples. Gas and liquid driven sample injection techniques are compatible with both of these hyperpolarization methods. The rapid sample injection techniques are combined with adapted NMR experiments working in a single, or small number of scans. They expand the application of liquid state hyperpolarization to spins with comparably short relaxation times, provide enhanced control over sample conditions, and allow for mixing experiments to study reactions in real time.

FUNCTIONAL MATERIALS FROM WASTE PAPER. IV. COMPARATIVE STUDY OF FUNCTIONAL COMPOSITION, SUPRAMOLECULAR STRUCTURE AND MORPHOLOGY OF POWDER CELLULOSES ISOLATED FROM WASTE PAPER, PLANT CELLULOSES AND UNCONVENTIONAL RAW MATERIALS

A comparative study of the supramolecular structure, physico-chemical properties and morphology of powder celluloses (PCs) obtained from the waste paper of newsprint and cardboard, plant cellulose and unconventional raw materials was conducted using the methods of FTIR spectroscopy, high-resolution 13C NMR spectroscopy in the solid phase, wide-angle X-ray scattering and scanning electron microscopy (SEM). According to the results of FTIR and 13C NMR spectroscopy, the functional composition of PCs is similar to that of wood PCs and matches highly purified cellulose samples. The crystallographic structure of powder samples corresponds to the structure of cellulose I. The crystallinity and transverse dimensions of crystallites are 39.4% and 40.2% and 45 Å [110] and 54 Å [110] for PC from newsprint paper and cardboard, respectively. These values considerably exceed the analogous values for the initial waste paper samples. A comparative analysis of the SEM results showed a significant similarity of PCs fibers of different origin. The results of the study of the physico-chemical properties of PCs isolated from waste paper make it possible to predict their competitive application in comparison with plant PCs.

Genome-Mining-Guided Discovery and Characterization of the PKS-NRPS-Hybrid Polyoxyperuin Produced by a Marine-Derived Streptomycete

The azinothricin family comprises several cyclic hexadepsipeptides with diverse pharmacological bioactivities, including antimicrobial, antitumoral, and apoptosis induction. In this work, using a genome mining approach, a biosynthetic gene cluster encoding an azinothricin-like compound was identified from the Streptomyces sp. s120 genome sequence (pop BGC). Comparative MS analysis of extracts from the native producer and a knockout mutant led to the identification of metabolites corresponding to the pop BGC. Furthermore, regulatory elements of the BGC were identified. By overexpression of an LmbU-like transcriptional activator, the production yield of 1 and 2 was increased, enabling isolation and structure elucidation of polyoxyperuin A seco acid (1) and polyoxyperuin A (2) using high-resolution mass spectrometry and NMR spectroscopy. Compound 1 exhibited a low antibiotic effect against Micrococcus luteus, while 2 showed a strong Gram-positive antibiotic effect in a micro-broth-dilution assay.

A Simple and Feasible Quantification of Metabolites in the Human Follicular Fluid Using 1H HR-MAS NMR Spectroscopy

This study presents a reliable method for the quantification of metabolite concentrations in follicular fluid with the high-resolution magic angle spinning nuclear magnetic resonance (HR-MAS NMR) spectroscopy and the ERETIC2 (Electronic REference To access In vivo Concentrations) based on PULCON (pulse length based concentration determination) principle. The positive effect of the HR-MAS probe technology on spectral quality and its ability to perform analyses with very low sample amounts were the most important factors of proposing this method. In evaluating the performance of the proposed method, standard creatine solutions in different concentrations containing DSS (2,2-dimethyl-2-silapentane-5-sulfonate sodium salt) as an internal reference standard were analyzed using different pulse programs (cpmgpr1d and zg30). The results obtained with the ERETIC2 were compared with the classical internal standard NMR quantification method (DSS method). The relative standard deviation (RSD) values for ERETIC2 were in the range of 0.3% - 5.7% and recovery values were calculated as minimum 90.3%, while RSD values for DSS method were in the range of 0.1% - 3.1% and recovery values were minimum 97.0%. Besides, it was observed that the metabolite concentration values calculated using the ERETIC2 procedure of follicular fluid samples obtained from the women with endometriosis and healthy controls were compatible with the values those obtained using different methodologies. The obtained results showed that the proposed quantification method based on the HR-MAS spectroscopy can easily be used in biological fluids and therefore it can be utilized as a good alternative to the internal standard method considering its accuracy and precision.

Direct Production of a Hyperpolarized Metabolite on a Microfluidic Chip.

Microfluidic systems hold great potential for the study of live microscopic cultures of cells, tissue samples, and small organisms. Integration of hyperpolarization would enable quantitative studies of metabolism in such volume limited systems by high-resolution NMR spectroscopy. We demonstrate, for the first time, the integrated generation and detection of a hyperpolarized metabolite on a microfluidic chip. The metabolite [1-13C]fumarate is produced in a nuclear hyperpolarized form by (i) introducing para-enriched hydrogen into the solution by diffusion through a polymer membrane, (ii) reaction with a substrate in the presence of a ruthenium-based catalyst, and (iii) conversion of the singlet-polarized reaction product into a magnetized form by the application of a radiofrequency pulse sequence, all on the same microfluidic chip. The microfluidic device delivers a continuous flow of hyperpolarized material at the 2.5 μL/min scale, with a polarization level of 4%. We demonstrate two methods for mitigating singlet–triplet mixing effects which otherwise reduce the achieved polarization level.