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Biomedical subjects

I I Stepuro

Publications and source records attributed to I I Stepuro.

At least 19 recordsLinked to original sources
PubMed2005

Role of thiamine thiol form in nitric oxide metabolism.

In alkaline media the thiamine cyclic form is converted into a thiol form (pK(a) 9.2) with an opened thiazole ring. The thiamine thiol form releases nitric oxide from S-nitrosoglutathione (GSNO). Thiamine disulfide, mixed thiamine disulfide with glutathione, and nitric oxide are produced in the reaction. Free glutathione was recorded in small amounts. The concentration of formed nitric oxide agreed well with the concentration of degraded GSNO. The concentration of released nitric oxide was determined under anaerobic conditions spectrophotometrically by production of nitrosohemoglobin. In air, the release of nitric oxide was recorded by the production of nitrite or the oxidation of oxyhemoglobin to methemoglobin. The concentration of the thiol form in the body under physiological pH values (7.2-7.4) did not exceed 1.5-2.0%. We believe that due to the exchange reactions between the thiamine thiol form and S-nitrosocysteine protein residues, nitric oxide can be released and mixed thiamine-protein disulfides are formed. The mixed thiamine disulfides (including thiamine ester disulfides) as well as the thiamine disulfide form are quite easily reduced by low molecular weight thiols to form the thiamine cyclic form with a closed thiazole ring. A possible role of the thiamine thiol form in releasing deposited nitric oxide from low-molecular-weight S-nitrosothiols and protein S-nitrosothiols and in regulation of blood flow in the vascular bed is discussed.

Anaerobiosis↗
PubMed2005

Thiamine and vasculopathies.

After peroxynitrite addition to aqueous solutions of thiamine at neutral and alkaline pH formation of thiamine disulfide and fluorescent products was observed. The fluorescent compounds were identified as thiochrome (TChr) and oxodihydrothiochrome (ODTChr) using spectral and fluorescent methods as well as paper chromatography and mass spectrometry. TChr and ODTChr are not the end products of thiamine oxidation and in neutral medium are unstable to peroxynitrite action and degrade rapidly to form non-fluorescent products. Thiamine, TChr, and ODTChr protects tyrosine from its modification by peroxynitrite. In the presence of TChr and ODTChr modification of tyrosinyl residues in human serum albumin and cytocrome c decreased. The prolonged thiamine incubation with glucose, amino acids and nitrite was accompanied by oxidative transformation of thiamine and formation of fluorescent products. We have shown that thiamine is also oxidized into TChr and ODTChr, i.e., it forms the same products as after thiamine oxidation by peroxynitrite. Moreover, thiamine (or its derivatives) appears as peroxynitrite scavenger leading to toxic effects lowering at diabetes mellitus.

Diabetic Angiopathies↗
PubMed2002

[Use of ascorbic acid for raising clinical response to long-acting nitrates].

The data of clinic observation and ECG-monitoring show that ascorbic acid significantly increases antianginal vasodilatory effects of nitroglycerine thus preventing from development of ischemic miocardial reaction in response to complamin injection. Incubation under anaerobe conditions of ascorbic acid solution with NO-donors (sodium nitrate, nitroglycerine) or with the blood of ischemic patients who had been treated for a long time with long-acting nitrovasodilators, results in liberation in Varburg vasculium of gas bubbles identified as nitrogen oxide according to hemoglobin nitrosylation. Activation of endogen NO-donors with ascorbic acid and taking antilogs of antianginal effects of exogenous nitroglycerine makes it possible to substantially increase the efficiency of nitratotherapy and nitratoprophylaxis of angina pectoris.

Adult↗
PubMed2000

Ultrasound-induced formation of S-nitrosoglutathione and S-nitrosocysteine in aerobic aqueous solutions of glutathione and cysteine.

S-Nitrosocompounds are formed when aqueous solutions of cysteine or glutathione are exposed to ultrasound (880 kHz) in air. The yield of the S-nitrosocompounds was as high as 10% for glutathione and 4% for cysteine of the initial thiol concentrations (from 0.1 to 10 mM) in the aqueous solutions. In addition to the formation of S-nitrosocompounds, thiol oxidation to disulfide forms was observed. After the oxidation of over 70% of the sulfhydryl groups, formation of peroxide compounds as well as cysteic acid derivatives was recorded. The formation of the peroxide compounds and peroxide radicals in the ultrasound field reduced the yield of S-nitrosocompounds. S-Nitrosocompounds were not formed when exposing low-molecular-weight thiols to ultrasound in atmospheres of N(2) or CO. In neutral solutions, ultrasound-exposed cysteine or glutathione released NO due to spontaneous degradation of the S-nitrosocompounds. N(2)O(3), produced due to the spontaneous degradation of the S-nitrosocompounds in air, nitrosylated sulfhydryl groups of glutathione manifested in the appearance of new absorption bands at 330 and 540 nm. The nitrogen compounds formed in an ultrasound field modified the sulfhydryl groups of apohemoglobin and serum albumin. The main target for ultrasound-generated oxygen free radicals were cystine residues oxidized to cysteic acid residues.

Air↗
PubMed1999

Reduction of nitrite by glycosylated amino acids and glycosylated albumin.

Glycosylated amino acids and glycosylated human serum albumin reduce nitrite to nitric oxide under anaerobic conditions. The amount of nitric oxide produced was recorded by generation of nitrosoHb from deoxyHb. Without preincubation after the addition of sodium nitrite, glucose or a mixture of glucose with amino acid or serum albumin did not cause spectrophotometrically detectible transformation of deoxyHb into nitrosoHb. The generation of NO increased with an increase in content of colored "final" products of amino acid and serum albumin glycosylation in the incubation mixture. The incubation of blood plasma of patients with diabetes mellitus with nitrite also resulted in the increased production of NO as compared to blood plasma of healthy subjects. During the incubation of healthy subjects' blood plasma with nitrite a small amount of NO was produced. The removal of low-molecular-weight compounds was accompanied by a significantly decreased generation of NO by blood plasma.

Adult↗
PubMed1997

Generation of NO during oxidation of hemoglobin ferroforms by nitrite.

The oxidation of hemoglobin solutions or erythrocyte suspensions containing a mixture of deoxyHb and oxyHb by NaNO2 (under decreased partial pressure of dissolved O2) resulted in the generation of metHb and nitrosoHb. The maximum amount of nitrosoHb was generated during the oxidation of deoxyHb. An increase in oxygen content was accompanied with increased generation of metHb, which was the only hemoglobin form under aerobic conditions. In the presence of oxygen, GSH was oxidized by NaNO2 to GSSG either in solution of oxyHb or in the structure of erythrocytes. GSH decelerated the oxidation of Hb to metHb due to prolongation of the slow phase and suppression of the autocatalytic phase of the reaction. The oxidation of GSH to GSSG was induced by NO2 radicals and not by NO. The incubation of deoxyHb with S-nitrosoglutathione resulted in its complete conversion to nitrosoHb, and this indicated that NO was released during the spontaneous decomposition of GSNO. The addition of S-nitrosoglutathione to oxyHb resulted in the generation of metHb in the solution.

Erythrocytes↗
PubMed1997

Reduction of methemoglobin and ferricytochrome c by glycosylated amino acids and albumin.

The fraction of Amadori products gradually decreased during heavy glycosylation of amino acids and human serum albumin while the amount of a colored product with the maximum fluorescence at 420 nm decreased. The addition of the produced ketoamines of amino acids to the solution of native albumin quenched its own fluorescence due to generation of a Schiff base with amino groups of the protein. Carbonyl-containing Amadori products obtained during the early steps of glycosylation were less potent electron donors than amino acids more heavily modified by the carbohydrate. Under anaerobic conditions, glycosylated amino acids and human serum albumin reduced metHb and ferricytochrome c to ferroforms. In the presence of oxygen, the electron was transferred from glycosylated amino acids to ferriforms of the heme proteins and also to oxygen molecules with the generation of superoxide anions and hydrogen peroxide. Free oxygen radicals and hydrogen peroxide induced damage to the protein globule of Hb associated with the release of hemin, Fe(III) ions, and cleavage of the porphyrin ring.

Amino Acids↗
PubMed1997

Thiamine oxidative transformations catalyzed by copper ions and ascorbic acid.

In aqueous solutions containing Cu(II) ions and ascorbic acid, thiamine was observed to be oxidized to the fluorescent products thiochrome and oxodihydrothiochrome in neutral and acid media. At high initial concentrations of thiamine, thiochrome was practically the only product of thiamine oxidation. Catalase inhibited the oxidation rate approximately by 30-fold, whereas superoxide dismutase reduced the rate by only 2.5-fold. Aliphatic alcohols, glucose, and high concentrations of ascorbic acid effectively inhibited the production of thiochrome. The yield of thiochrome was also decreased in the presence of aliphatic amino acids, histidine, and particularly human serum albumin (HSA). With complete binding of copper ions by HSA, no formation of fluorescent products was observed. In neutral and acidic media under the action of hydroxyl radicals, thiamine formed a tricyclic semiquinone form which was then oxidized to thiochrome by superoxide anion or H2O2. Ascorbic acid played the main role in the reduction of Cu(II), whereas the contribution of superoxide anions was less significant. Cu(I) interacted with H2O2 to form hydroxyl radicals. The addition of H2O2 both to thiamine and to the mixture of thiamine and Cu(II) ions did not lead to significant production of thiochrome in neutral and acidic media.

Ascorbic Acid↗
PubMed1990

[Study of pH-dependence of kinetic parameters of pyruvate kinase from bovine adrenal cortex].

The effect of pH on the main kinetic parameters of pyruvate kinase function was studied. The maximal rate of the reaction as well as the values of Km for ADP and Ki for phenylalanine depend on pH and show a well-defined extremum at pH 6.8-7.0. Spectrofluorimetric titration of pyruvate kinase results in pH dependencies of changes in the fluorescence spectra parameters (e.g., quantum yield, half-width and position of the maximum). This enabled to determine the pH regions corresponding to changes in the state of tryptophan residues. Data from the enzyme inhibition by phenylalanine suggest that acidification of the medium leads to the decrease of the catalytic activity due to the protonation of the ionogenic group of the enzyme. Within the pH range of 7.0-8.0, the decrease of the pyruvate kinase activity is due to structural shifts in the enzyme molecule, as a result of which the steric complementariness of the enzyme active center with respect to the substrate (Mg.ADP) is impaired.

Adrenal Cortex↗
PubMed1989

[Stabilization of hemoglobin by aldehydes in reactions of autooxidation to methemoglobin].

Aldehydes cause the retardation of hemoglobin autooxidation to methemoglobin. The protective effect is due to semimercaptal formation between aldehydes and sulfhydryl groups of cysteine++ 93 of the oxyhemoglobin beta-chain. The efficiency of hemoglobin protection of different aldehydes depends on the correlation between the equilibrium constants of carbonyl compound binding to protein sulfhydryl or amino groups and their electron accepting properties.

Aldehydes↗
PubMed1987

Interaction of ethanol with blood proteins.

At concentrations arising in alcohol intoxication ethanol was reversibly bound to human serum albumin and hemoglobin. Alcohol altered the conformation and stability of the proteins, reduced the domain-domain interactions in the molecule of albumin and changed the binding of such hydrophobic ligands as ANS and bilirubin.

Bilirubin↗
PubMed1987

[Interaction of prostaglandin E2 with human serum albumin].

Using equilibrium dialysis, protein fluorescence and fluorescent probing as well as chemical modification, the interaction of prostaglandin E2 with human serum albumin was studied. The serum albumin molecule has a highly specific prostaglandin E2-binding site. The binding of prostaglandin causes conformational rearrangements in the protein molecule. The amino group of serum albumin is involved in the interaction with prostaglandin E2. Prolonged exposure of prostaglandin E2 to serum albumin causes partial irreversible binding of prostaglandin molecules to the protein.

Binding Sites↗
PubMed1987

[Interaction of pyridoxal-5-phosphate with human serum albumin and pancreatic ribonuclease].

Pyridoxal-5-phosphate (in a lesser degree, pyridoxal) interacts with both non-protonated and protonated exposed epsilon-amino groups of lysine residues and with alpha-amino groups in human serum albumin and pancreatic ribonuclease A. The reaction of Schiff base formation proceeds within a wide pH range--from 3.0 to 12.0. At a great pyridoxal-5-phosphate excess in ribonuclease A in neutral or slightly acidic aqueous media all the ten epsilon-amino groups of lysine residues and the alpha-amino groups of Lys-1 become modified. The formation of aldimine bonds of pyridoxal-5-phosphate with protonated amino groups in acidic media is determined by ionization of its phenol hydroxyl and phosphate residues. Acetaldehyde, propionic aldehyde and pyridine aldehyde interact only with non-protonated amino groups of the proteins. The equilibrium constants of pyridoxal-5-phosphate and other aldehydes binding to proteins and amino acids were determined. The rate constants of Schiff base formation for pyridoxal-5-phosphates with some amino acids and primary sites of proteins for direct and reverse reactions were calculated.

Alkylation↗
PubMed1986

[Deuterium exchange of the methyl group of pyruvate catalyzed by human serum albumin].

Human serum albumin catalyzes proton exchange of the methyl group of the pyruvate molecule in heavy water. The exchange process is mainly due to the formation of bonds of a Schiff base type between six deprotonated protein amino groups and pyruvate. Both hydrated and non-hydrated forms of pyruvate interact with positively charged side amino acid residues of the polypeptide chain (most probably, with arginine) located in the hydrophobic "pockets" of the protein globule. The value of equilibrium association constants with serum albumin exceeds that for the hydrated form of pyruvate.

Biological Transport↗
PubMedDate not supplied

[Localization of pyridoxal-5-phosphate, covalently bound with human hemoglobin. Spectrofluorimetric studies].

Human apohemoglobin tryptophan residues were localized in the regions of the protein globule with restricted mobility. By the method of dynamic quenching of phosphopyridoxyl chromophore fluorescence, the heterogeneity of pyridoxal-5-phosphate molecules covalently bound to the human hemoglobin molecules was determined from the accessibility to solvent. The first four pyridoxal-5-phosphate molecules are localized in the hydrophobic regions of the hemoglobin molecule; at the same time, they have a high mobility. One of these molecules is situated at the site inaccessible to the solvent, which coincides with the anion-binding center of the oxyhemoglobin molecule. The next pyridoxal-5-phosphate molecules modify the surface amino groups of the protein. In the apohemoglobin molecule, the pyridoxal-5-phosphate binding sites are more exposed to the solvent, as compared to hemoglobin. In the hemoglobin molecule modified by pyridoxal-5-phosphate, an effective electron excitation energy transfer from tryptophan residues to phosphopyridoxyl chromophores occurs. The effective distances between tryptophanyls of single subunits of hemoglobin and the covalently bound pyridoxal-5-phosphate molecule were estimated to be 19 A for the alpha-subunit and 17 A for the beta-subunit.

Hemoglobins↗
PubMedDate not supplied

[Interaction of riboflavin and hemoproteins with organic free radicals and superoxide anions generated in the ultrasound field].

The reduction of ferricytochrome c to the ferro form in aqueous alcohol solutions in air by the action of ultrasound and the complete inhibition of this process in the presence of superoxide dismutase indicate the generation of superoxide anions. Further exposure to the ultrasonic field leads to a reverse process of oxidation of the cytochrome c ferro form to the ferri form by hydrogen peroxides and organic peroxides. The addition of catalase protects the cytochrome c ferro form from oxidation to the ferri form. The oxidized form of riboflavin effectively interacts with organic free radicals and superoxide anions to produce a leuko form, which is easily oxidized by air oxygen or the ferri forms of hemoglobin and cyt c to form riboflavin and hydrogen peroxide or the ferro forms of heme-containing proteins, respectively. The recurrence of redox reactions in the presence of riboflavin, organic free radicals, and O2 and the ferri forms of heme-containing proteins suggests that riboflavin can play a role of an antioxidant in the organism. It is supposed that, due to interaction with superoxide anions, riboflavin stabilizes the NO level in the organism under conditions of increased superoxide anion generation and (or) decreased superoxide dismutase activity. A possible role of riboflavin in the modulation of toxic and signal pathways of nitrogen oxide is discussed.

Air↗
PubMedDate not supplied

[Nonenzymatic reduction of Met-Hb under the action of NAD radicals generated in the presence of HAD.H and Fe(II) salts. Antioxidant properties of Met-Hb].

It has been found that superoxide radicals formed at autooxidation of Fe(II) ions or of their complexes with EDTA in the presence of NAD.H cause reduction of met-Hb into deoxy or oxy-form. Under the effect of hydroxyl radicals generated in the ultrasonic field in the presence of NADH reduction of met-Hb into the ferro-form (carboxy-Hb) was observed in Co atmosphere and not in O2 atmosphere. It was induced by a high oxidation rate of oxy-Hb by hydroxyl radicals into met-Hb as compared to carboxy-Hb. Reduction of met-Hb was shown to be accompanied with the formation of NAD+. The role of NAD. radicals in the reduction of met-Hb which acts as an antioxidant in respect to organic free radicals was discussed.

Antioxidants↗