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Molecules (Basel, Switzerland) Dec 2021The effect of a reduced deuterium (D) content in the incubation medium on the survival of cultured neurons in vitro and under glucose deprivation was studied. In...
The effect of a reduced deuterium (D) content in the incubation medium on the survival of cultured neurons in vitro and under glucose deprivation was studied. In addition, we studied the effect of a decrease in the deuterium content in the rat brain on oxidative processes in the nervous tissue, its antioxidant protection, and training of rats in the T-shaped maze test under hypoxic conditions. For experiments with cultures of neurons, 7-8-day cultures of cerebellar neurons were used. Determination of the rate of neuronal death in cultures was carried out using propidium iodide. Acute hypoxia with hypercapnia was simulated in rats by placing them in sealed vessels with a capacity of 1 L. The effect on oxidative processes in brain tissues was assessed by changes in the level of free radical oxidation and malondialdehyde. The effect on the antioxidant system of the brain was assessed by the activity of catalase. The study in the T-maze was carried out in accordance with the generally accepted methodology, the skill of alternating right-sided and left-sided loops on positive reinforcement was developed. This work has shown that a decrease in the deuterium content in the incubation medium to a level of -357‱ has a neuroprotective effect, increasing the survival rate of cultured neurons under glucose deprivation. When exposed to hypoxia, a preliminary decrease in the deuterium content in the rat brain to -261‱ prevents the development of oxidative stress in their nervous tissue and preserves the learning ability of animals in the T-shaped maze test at the level of the control group. A similar protective effect during the modification of the H/H internal environment of the body by the consumption of DDW can potentially be used for the prevention of pathological conditions associated with the development of oxidative stress with damage to the central nervous system.
Topics: Adaptation, Biological; Animals; Antioxidants; Biomarkers; Cell Death; Cells, Cultured; Culture Media; Deuterium; Glucose; Hypoxia; Lipid Peroxidation; Neuroglia; Neurons; Oxidation-Reduction; Oxidative Stress; Rats
PubMed: 35011474
DOI: 10.3390/molecules27010243 -
NMR in Biomedicine Mar 2020Deuterium metabolic imaging (DMI) is a novel MR-based method to spatially map metabolism of deuterated substrates such as [6,6'- H ]-glucose in vivo. Compared with...
Deuterium metabolic imaging (DMI) is a novel MR-based method to spatially map metabolism of deuterated substrates such as [6,6'- H ]-glucose in vivo. Compared with traditional C-MR-based metabolic studies, the MR sensitivity of DMI is high due to the larger H magnetic moment and favorable T and T relaxation times. Here, the magnetic field dependence of DMI sensitivity and transmit efficiency is studied on phantoms and rat brain postmortem at 4, 9.4 and 11.7 T. The sensitivity and spectral resolution on human brain in vivo are investigated at 4 and 7 T before and after an oral dose of [6,6'- H ]-glucose. For small animal surface coils (Ø 30 mm), the experimentally measured sensitivity and transmit efficiency scale with the magnetic field to a power of +1.75 and -0.30, respectively. These are in excellent agreement with theoretical predictions made from the principle of reciprocity for a coil noise-dominant regime. For larger human surface coils (Ø 80 mm), the sensitivity scales as a +1.65 power. The spectral resolution increases linearly due to near-constant linewidths. With optimal multireceiver arrays the acquisition of DMI at a nominal 1 mL spatial resolution is feasible at 7 T.
Topics: Animals; Brain; Carbon-13 Magnetic Resonance Spectroscopy; Deuterium; Humans; Magnetic Fields; Magnetic Resonance Imaging; Phantoms, Imaging; Rats; Signal-To-Noise Ratio
PubMed: 31879985
DOI: 10.1002/nbm.4235 -
Molecular & Cellular Proteomics : MCP Dec 2019Amide hydrogen-deuterium exchange mass spectrometry (HDX-MS) has become widely popular for mapping protein-ligand interfaces, for understanding protein-protein...
Amide hydrogen-deuterium exchange mass spectrometry (HDX-MS) has become widely popular for mapping protein-ligand interfaces, for understanding protein-protein interactions, and for discovering dynamic allostery. Several platforms are now available which provide large data sets of amide hydrogen/deuterium exchange mass spectrometry (HDX-MS) data. Although many of these platforms provide some down-stream processing, a comprehensive software that provides the most commonly used down-stream processing tools such as automatic back-exchange correction options, analysis of overlapping peptides, calculations of relative deuterium uptake into regions of the protein after such corrections, rigorous statistical analysis of the significance of uptake differences, and generation of high quality figures for data presentation is not yet available. Here we describe the Deuterium Exchange Correction and Analysis (DECA) software package, which provides all these downstream processing options for data from the most popular mass spectrometry platforms. The major functions of the software are demonstrated on sample data.
Topics: Datasets as Topic; Deuterium; Electronic Data Processing; Hydrogen Deuterium Exchange-Mass Spectrometry; Software; User-Computer Interface
PubMed: 31594786
DOI: 10.1074/mcp.TIR119.001731 -
Molecular and Cellular Biochemistry Dec 2021Deuterium (D) is a stable isotope of hydrogen (H) with a mass number of 2. It is present in natural waters in the form of HDO, at a concentration of 16.8 mmol/L,...
Deuterium (D) is a stable isotope of hydrogen (H) with a mass number of 2. It is present in natural waters in the form of HDO, at a concentration of 16.8 mmol/L, equivalent to 150 ppm. In a phase II clinical study, deuterium depletion reduced fasting glucose concentration and insulin resistance. In this study, we tested the effect of subnormal D-concentration on glucose metabolism in a streptozotocin (STZ)-induced diabetic rat model. Animals were randomly distributed into nine groups to test the effect of DO (in a range of 25-150 ppm) on glucose metabolism in diabetic animals with or without insulin treatment. Serum glucose, fructose amine-, HbA1c, insulin and urine glucose levels were monitored, respectively. After the 8-week treatment, membrane-associated GLUT4 fractions from the soleus muscle were estimated by Western blot technique. Our results indicate that, in the presence of insulin, deuterium depletion markedly reduced serum levels of glucose, -fructose amine, and -HbA1c, in a dose-dependent manner. The optimal concentration of deuterium was between 125 and 140 ppm. After a 4-week period of deuterium depletion, the highest membrane-associated GLUT4 content was detected at 125 ppm. These data suggest that deuterium depletion dose-dependently enhances the effect of insulin on GLUT4 translocation and potentiates glucose uptake in diabetic rats, which explains the lower serum glucose, -fructose amine, and -HbA1c concentrations. Based on our experimental data, deuterium-depleted water could be used to treat patients with metabolic syndrome (MS) by increasing insulin sensitivity. These experiments indicate that naturally occurring deuterium has an impact on metabolic regulations.
Topics: Animals; Blood Glucose; Deuterium; Diabetes Mellitus, Experimental; Disease Models, Animal; Glucose Transporter Type 4; Hypoglycemic Agents; Insulin; Male; Muscle, Skeletal; Rats; Rats, Wistar; Water
PubMed: 34510301
DOI: 10.1007/s11010-021-04231-0 -
MAbs 2019High concentration biotherapeutic formulations are often required to deliver large doses of drugs to achieve a desired degree of efficacy and less frequent dose....
High concentration biotherapeutic formulations are often required to deliver large doses of drugs to achieve a desired degree of efficacy and less frequent dose. However, highly concentrated protein-containing solutions may exhibit undesirable therapeutic properties, such as increased viscosity, aggregation, and phase separation that can affect drug efficacy and raise safety issues. The characterization of high concentration protein formulations is a critical yet challenging analytical task for therapeutic development efforts, due to the lack of technologies capable of making accurate measurements under such conditions. To address this issue, we developed a novel dilution-free hydrogen/deuterium exchange (HDX) mass spectrometry (MS) method for the direct conformational analysis of high concentration biotherapeutics. Here, we particularly focused on studying phase separation phenomenon that can occur at high protein concentrations. First, two aliquots of monoclonal antibodies (mAbs) were dialyzed in either hydrogen- or deuterium-containing buffers at low salt and pH. Phases that separated were then discretely sampled and subjected to dilution-free HDX-MS analysis through mixing the non-deuterated and deuterated protein aliquots. Our HDX-MS results analyzed at a global protein level reveal less deuterium incorporation for the protein-enriched phase compared to the protein-depleted phase present in high concentration formulations. A peptide level analysis further confirmed these observed differences, and a detailed statistical analysis provided direct information surrounding the details of the conformational changes observed. Based on our HDX-MS results, we propose possible structures for the self-associated mAbs present at high concentrations. Our new method can potentially provide useful insights into the unusual behavior of therapeutic proteins in high concentration formulations, aiding their development.
Topics: Antibodies, Monoclonal; Biological Products; Deuterium; Deuterium Exchange Measurement; Dialysis; Humans; Hydrogen; Mass Spectrometry; Peptides; Protein Conformation
PubMed: 30890021
DOI: 10.1080/19420862.2019.1589850 -
Journal of the American Society For... May 2022Experimental measurement of time-dependent spontaneous exchange of amide protons with deuterium of the solvent provides information on the structure and dynamical...
Experimental measurement of time-dependent spontaneous exchange of amide protons with deuterium of the solvent provides information on the structure and dynamical structural variation in proteins. Two experimental techniques are used to probe the exchange: NMR, which relies on different magnetic properties of hydrogen and deuterium, and MS, which exploits the change in mass due to deuteration. NMR provides residue-specific information, that is, the rate of exchange or, analogously, the protection factor (i.e., the unitless ratio between the rate of exchange for a completely unstructured state and the observed rate). MS provides information that is specific to peptides obtained by proteolytic digestion. The spatial resolution of HDX-MS measurements depends on the proteolytic pattern of the protein, the fragmentation method used, and the overlap between peptides. Different computational approaches have been proposed to extract residue-specific information from peptide-level HDX-MS measurements. Here, we demonstrate the advantages of a method recently proposed that exploits self-consistency and classifies the possible sets of protection factors into a finite number of alternative solutions compatible with experimental data. The degeneracy of the solutions can be reduced (or completely removed) by exploiting the additional information encoded in the shape of the isotopic envelopes. We show how sparse and noisy MS data can provide high-resolution protection factors that correlate with NMR measurements probing the same protein under the same conditions.
Topics: Deuterium; Deuterium Exchange Measurement; Hydrogen; Magnetic Resonance Spectroscopy; Mass Spectrometry; Peptides; Proteins
PubMed: 35385652
DOI: 10.1021/jasms.2c00005 -
Biochimica Et Biophysica Acta Jun 2013Amide hydrogen/deuterium exchange detected by mass spectrometry (HXMS) is seeing wider use for the identification of intrinsically disordered parts of proteins. In this... (Review)
Review
Amide hydrogen/deuterium exchange detected by mass spectrometry (HXMS) is seeing wider use for the identification of intrinsically disordered parts of proteins. In this review, we discuss examples of how discovery of intrinsically disordered regions and their removal can aid in structure determination, biopharmaceutical quality control, the characterization of how post-translational modifications affect weak structuring of disordered regions, the study of coupled folding and binding, and the characterization of amyloid formation. This article is part of a Special Issue entitled: Mass spectrometry in structural biology.
Topics: Amino Acid Sequence; Deuterium; Deuterium Exchange Measurement; Humans; Hydrogen; Mass Spectrometry; Molecular Sequence Data; Protein Conformation; Protein Folding; Proteins
PubMed: 23099262
DOI: 10.1016/j.bbapap.2012.10.009 -
Analytical Chemistry Jan 2015
Review
Topics: Animals; Deuterium; Deuterium Exchange Measurement; Humans; Hydrogen; Mass Spectrometry
PubMed: 25398026
DOI: 10.1021/ac5040242 -
Biochimica Et Biophysica Acta Nov 2015Oxidation of alcohols and amines is catalyzed by multiple families of flavin- and pyridine nucleotide-dependent enzymes. Measurement of solvent isotope effects provides... (Review)
Review
Oxidation of alcohols and amines is catalyzed by multiple families of flavin- and pyridine nucleotide-dependent enzymes. Measurement of solvent isotope effects provides a unique mechanistic probe of the timing of the cleavage of the OH and NH bonds, necessary information for a complete description of the catalytic mechanism. The inherent ambiguities in interpretation of solvent isotope effects can be significantly decreased if isotope effects arising from isotopically labeled substrates are measured in combination with solvent isotope effects. The application of combined solvent and substrate (mainly deuterium) isotope effects to multiple enzymes is described here to illustrate the range of mechanistic insights that such an approach can provide. This article is part of a Special Issue entitled: Enzyme Transition States from Theory and Experiment.
Topics: Alcohols; Amines; Biocatalysis; Deuterium; Enzymes; Isotopes; Models, Chemical; Molecular Structure; Solvents; Substrate Specificity
PubMed: 25448013
DOI: 10.1016/j.bbapap.2014.10.020 -
European Journal of Pharmaceutical... May 2022Despite being a first-line clinical drug, thienopyridines have many unsatisfactory aspects, including the low bioavailability of clopidogrel(CLP) and the high bleeding...
BACKGROUND AND PURPOSE
Despite being a first-line clinical drug, thienopyridines have many unsatisfactory aspects, including the low bioavailability of clopidogrel(CLP) and the high bleeding risk of prasugrel. We synthesized deuterium clopidogrel(D-CL, patented in China) to alleviate the deficiency of CLP in clinical, such as a slow onset, a greater influence of gene polymorphism, and a high frequency of drug-drug interaction.
EXPERIMENTAL APPROACH
Molecular docking was used to analyze the affinity between D-CL and the P2Y receptor. The levels of active metabolites of D-CL were detected using HPLC/MS-MS and the activities of main metabolic enzymes were analyzed; Subsequently, platelet aggregation function, thrombus model were used to evaluate the pharmacodynamics of D-CL. Finally, the safety of D-CL were evaluated through examination of blood routine, PT, APTT, bleeding time, serological tests, liver pathological biopsy, liver cell apoptosis and detection of apoptosis-related proteins.
KEY RESULTS
The introduction of deuterium made the binding of CLP to P2Y receptor more stable, improved the concentration of active metabolites, and substantially reduced the inhibition of major metabolic enzymes, including CYP2B6, CYP2C9, and CYP2C19, thereby, exerting better antiplatelet effects without increasing the risk of bleeding, along with a concomitant decrease in the apoptosis of hepatocytes.
Topics: Clopidogrel; Deuterium; Formic Acid Esters; Hydrogen; Molecular Docking Simulation; Platelet Aggregation; Platelet Aggregation Inhibitors; Thiophenes
PubMed: 35257876
DOI: 10.1016/j.ejps.2022.106157