Murexide reaction

Purine derivatives (e. g. xanthine) are oxidized by chloramine T in the presence of hydrochloric acid and form purple-red anunonium salts of purpuric acid (murexide) with ammonia. Whether the murexide reaction is also the cause of the fluorescence is open to question. 

In the concentration range above 1 ng substance per spot, red-colored chromatogram zones (murexide reaction) could be seen on a pale background these could be excited to blue (caffeine, h/ f 75- 80 theobromine, hRf 55-60) or yellow (theophylline h/ f 35-40) fluorescence on a dark background in long-wavelength UV light (X = 365 nm). 

Murexide Reaction.—A few centigrams of uric acid are evaporated to dryness in a small porcelain basin on the water bath with some drops of slightly diluted nitric acid. When ammonia is added to the residue an intense purple colour is produced. 

The relaxation times for the complex formation of Li+ and Na+ with murexide have been determined. In these studies the electric field pulse technique was used, which has a resolution time of about 30 nanoseconds. The method is described in detail elsewhere 15). Fig. 10 shows a t q)ical relaxation curve for the Na-murexide reaction. The effect for K-murexide complex formation (requiring higher concentration, due to the lower stability constant) was already beyond the resolution of this method. 

Heparinoids and mucopolysaccharides react with, and modify, many of the plasma proteins. Heparin combines with fibrinogen, globulins and albumin. As judged by electrophoresis and various types of analysis and staining, the particular plasma protein components with which heparin combines are dependent upon the concentration of protein, concentration of heparin, pH value, and salts present. This explains the somewhat contradictory statements in literature about combinations of heparin with plasma proteins. The combination may result in change of solubility of the protein and reverse protein tests . Heparin can modify the murexide reaction for calcium in serum by affecting the calcium-protein-heparin complex. Many heparinoids... 

White, odorless, tasteless crystals dec by heat without melting and with evolution of HCN, d ] 89. One gram dissolves in about 15,000 parts cold water, about 2000 parts boiling water sol in glycerol, in so]ns of alkali hydroxides, their carbonates, sodium acetate and sodium phosphate inso] in alcohol, ether. Gives murexide reaction. 

A fairly large number of structurally-related compoxmds give a positive murexide reaction. They are useful as an identification reaction for uric acid (Figure 3.34.5) and other purine derivates. These substances all contain the same structural feature as the six-membered heterocyclic ring of xanthine. 

The compound responsible for the purple color, or at least one of the compounds responsible, has been isolated. A larger number of compounds, which is believed to be responsible for the yellow reaction product of the first step of analysis, have also been isolated. But still, it is evident that a very complex chain of chemical reactions takes place during the procedure. The core of the reaction is, apparently, that the compound in question, through a number of steps, is oxidized into the intermediate alloxane that ultimately condenses to murexoin. The ammonium salt of murexoin is the principal contributor to the purple color of the final test solution. The murexide reaction of caffeine can therefore, extremely simplified, be written as above (Figure 3.34.6). 

Whereas the exact reaction mechanisms of the test are not easily elucidated in detail, some knowledge has been gathered about the structural prerequisites a compound must meet before it can give a positive murexide reaction. It appears that some rules can be set up about the substituents on the 6-member heterocyclic ring, including the nature of the substituents, which in the case of the xanthines includes a 5-membered heterocyclic ring. 

Kozuka, H., Koyama, M. and Okitsu, T., Murexide reaction of caffeine with hydrogen peroxide and hydrochloric acid, Chem. Pharmacal. Bull., 29,1981,433. 

Detection, analysis A. precipitation reagents that effect formation of poorly water soluble salts are, e.g., Dragendorff s, FrOhde s, and Mayer s reagents, picric acid, picrolonic acid, dipicrylamine, phosphomolyb-dic acid, Reinecke s salt, and sodium tetraphenylbo-rate. Special color reactions are used for some groups of A., e.g., Vitali s reaction to detect tropane A., the thalleioquine reaction for Cinchona A., the murexide reaction for purines. 

Theobromine (3,7-dimethylxanthine, 3,7-dihydro-3,7-dimethyl-lW-purine-2,6-dione). C7H,N402, Mr 180.17 formula, see under theophylline. Monoclinic, bitter tasting needles, mp. 357 °C, sublimes at 290-295 °C, soluble in hot water, alkali hydroxides, concentrated acids, moderately soluble in ammonia, poorly soluble in cold water and alcohol. With acids T. forms salts which decompose in water detection by the murexid reaction. T. is the main alkaloid of cocoa (Theobroma cacao, 1.5-3 wt.-%), from which it is obtained - especially from the husks in which it accumulates during fermentation. The typical bitter taste of cocoa is the result of interactions between T. and the pip-erazinediones formed in the roasting process. T. has diuretic, vasodilatory, and stimulating effects on cardiac muscle. The activities are weaker than those of the structurally related caffeine (a methylation product of T.) with which it co-occurs in cola nuts. For further pharmacological properties, see table under theophylline. 

A more specific method of detecting barbituric acids is through the murexide reaction [86a, 173] and of thiobarbituric acids, through the iodine-azide reaction (Rgt. No. 142) [173]. Bromine-containing barbituric acids may be characterised by oxidation yielding bromine which then converts fluorescein into eosin (Rgt. No. 119). Larger amounts of barbituric acids and ureides, separated by TLC, can be removed from the layer by microsublimation and identified through their crystal form [8]. 

Uric acid is only sparingly soluble in water and a weak dibasic acid (pK i = 5.4, pf A2 = 10.6). A violet color appears when uric acid is heated with HNO3, cooled and treated with NH3 (Murexide reaction). The action of POCI3 on uric acid produces 2,6-dichloro-8-hydroxypurine, and under drastic conditions yields 2,6,8-trichloropurine. 

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