Methylene blue formation

Ray et al. [77] used an indirect method based on AAS for the determination of sulfide in flooded acid sulfate soils. Hydrogen sulfide, evolved during the anaerobic metabolism of sulfate, is readily converted into insoluble metal sulfides, chiefly iron sulfide, in flooded acid sulfate soils. A method for determining sulfide is based on the precipitation of the sulfide as zinc sulfide and subsequent determination by methylene blue formation or iodine titrimetry. 

Because some amount of S2- may escape out as H2S during standard preparation and a small proportion may be oxidized to sulfate over a few hours, and also because methylene blue formation reaction may not thermodynamically go to completion, it is, therefore, always recommended that sulfide concentration should be determined by titrimetric iodide procedure and an average percent error of the methylene blue procedure be compared against this titrimetric procedure. 

The quantitative extraction of MB from the aqueous phase requires 10 -10" -fold excess of perchlorate and shaking with two or three portions of CHCI3. Equilibrium is attained after shaking for 20-30 s. The optimum acidity for the Methylene Blue formation is also suitable for the extraction with CHCI3. 

When reduced ferredoxin is used as the electron donor for polysulfide reduction, the reaction is monitored by the production of sulfide. The assays are performed in 8-ml serum-stoppered vials under Ar that are shaken at 150 rpm at 80°. The reaction mixture (2 ml) contains 100 vaM EPPS buffer, pH 8.0, 10 mM pyruvate, 2 toM coenzyme A, 150 p,g POR from P. furiosus, 25 lM ferredoxin, 40 p.g FNOR, and a source of elemental sulfur. This is polysulfide (1.5 mA/), sublimed elemental sulfur (0.5%, w/v J. T. Baker, Marietta, GA), or colloidal sulfur (0.05%, w/v Fluka, Ronkonkoma, NY). Note that sublimed elemental sulfur has a very low solubility, whereas colloidal sulfur generates a fine suspension. At 20 min intervals over 2 hr, aliquots of the reaction are removed with a syringe and the amount of sulfide produced is measured by methylene blue formation." One unit of activity is defined as 1 p.mol sulfide produced per min. When reactions are monitored by sulfide production, it is important that control assays be carried out without the addition of enzyme. This is particularly important when poly sulfide is used as the source of elemental sulfur. 

Irradiation of 4-hydroxy- and 4-alkoxy-3-pyrazolin-5-one derivatives (163 R = OH, OR) leads to ring cleavage with the formation of /3-diamides (165) 69TL271). The methylene blue sensitized rearrangement of the same pyrazolinone (R = H) to the oxindole (166) also... 

Warm a few of the crystals with yellow ammonium sulphide solution for a few minutes, acidify with hydrochloric acid, und finally add a little feiric chloride. A deep blue colouration Ls produced, due to the formation of methylene blue. 

Methylene blue plus light mediates 8-hydroxyguanine formation in DNA. Arch. Biochem. Biophys. 273, 106-111. 

The property of polyanions to give metachromatic reactions with basic dyes80 (see Section XI) has been widely exploited for the titration of glycosaminoglycans. Because of its high charge-density, heparin displays large shifts in the visible spectrum of the dye upon complex-formation, and can conveniently be titrated (most commonly, with Methylene Blue or Azure A), even in the presence of minor proportions of other glycosaminoglycans. It should be realized that quantitative responses with metachromatic dyes are different for different pure heparin... 

Methylene blue and other reducible dyes were shown to enhance the activity of NADPHa-linked MHbR (K8, K9). This is confirmed by the finding that intravenous injections of methylene blue in methemo-globinemic patients result in a striking decrease of MHb levels (e.g., B14, K9, K10). This seems to be paradoxical, since methylene blue is capable of reacting with Hb with formation of MHb, but the dye reacts much more effectively as an artificial electron carrier in the NADPH2-MHbR Systran (B14). It has been stated (K10) that methemoglobin reduction is associated with the formation of pyruvate in equivalent amounts, but that in reactions accelerated by reducible dyes no correlation between pyruvate formation and MHb reduction could be found. 

B2. Barron, E. S. G., and Harrop, G. A., The effect of methylene blue and other dyes upon the glycolysis and lactic acid formation of mammalian and avian erythrocytes. J. Biol. Chetn. 79, 65-87 (1028). 

The aqueous solution of methylene blue is not decolorised by sodium hydroxide the blue water-solubie ammonium base is formed. The dye is converted by reducing agents into the easily oxidisable leuco-base. The following experiment, which demonstrates the formation of methylene blue by the introduction of sulphur between the rings of Bind-schedler s green, is very instructive. 

Wasserman et al.9T> irradiated a methanolic solution of 41 a in the presence of oxygen and methylene blue for 18 h at 25 °C. After hydrogenation of the reaction mixture over palladium on charcoal for 24 h they were able to isolate three products 120 (26%), 121 (11%), and 5% of a substance which was assigned structure 122 on the basis of spectroscopic and X-ray crystallographic data. The mechanism of formation of this polycycle probably involves an epoxide of type 123... 

Photolytic. Acher et al. (1981) studied the dye-sensitized photolysis of terbacil in aerated aqueous solutions over a wide pH range. After a 2-h exposure to sunlight, terbacil in aqueous solution (pH range 3.0-9.2) in the presence of methylene blue (3 ppm) or riboflavin (10 ppm) decomposed to 3-ter/-5-butyl-5-acetyl-5-hydroxyhydantoin. Deacylation was observed under alkaline conditions (pH 8.0 or 9.2) affording 3-/er/-5-hydroxyhydantoin. In neutral or acidic conditions (pH 6.8 or 3.0) containing riboflavin, a mono-Wdealkylated terbacil dimer and an unidentified water-soluble product formed. Product formation, the relative amounts of products formed, and the rate of photolysis were all dependent upon pH, sensitizer, temperature, and time (Acher et ah, 1981). 

Diaryl- and tetraaryl-substituted furans such as tetraphenylfuran (384) for example, yield generally cw-diaroylethylenes such as 380, probably via intermediate ozonide formation.257 Secondary reactions seem to depend very much on the nature of the solvents. Thus, cis-dibenzoylstilbene (380) has been observed by direct photooxygenation of 384 in CS2 as well as by methylene blue-sensitized photooxygenation of 384 in methanol.257,258 However, when the latter reaction was carried out in acetone, epoxide 386 and the enolbenzoate 387 were obtained.258... 

Gruetter, C. A., Gruetter, D. Y., Lyon, J. E., Kadowitz, P. J., and Ignarro, L. J. (1981). Relationship between cyclic guanosine 3 5 -monophosphate formation and relaxation of coronary arterial smooth muscle by glyceryl trinitrate, nitroprusside, nitrite and nitric oxide Effects of methylene blue and methemoglobin. J. Pharmacol. Exp. Ther. 219, 181-186. 

A nonmitochondrial muscle diaphorase has also been described (Koizumi and Brown (1971), capable of forming NOMb in solutions containing nitrite and Mb, using methylene blue as a nonspecific electron carrier. NOMb formation by both the mitochondrial and mtnmitochondrial diaphorase enzyme systems requires the presence of reduced NADH. 

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