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.

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.

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.