Color on Addition of Reagents

Hydantoin itself can be detected ia small concentrations ia the presence of other NH-containing compounds by paper chromatography followed by detection with a mercury acetate—diphenylcarba2one spray reagent. A variety of analytical reactions has been developed for 5,5-disubstituted hydantoias, due to their medicinal iaterest. These reactions are best exemplified by reference to the assays used for 5,5-diphenylhydantoiQ (73—78), most of which are based on their cycHc ureide stmcture. Identity tests iaclude the foUowiag (/) the Zwikker reaction, consisting of the formation of a colored complex on treatment with cobalt(II) salts ia the presence of an amine (2) formation of colored copper complexes and (3) precipitation on addition of silver(I) species, due to formation of iasoluble salts at N. ... 

Traut s reagent is fully water-soluble and reacts with primary amines in the pH range 7—10. The cyclic imidothioester is stable to hydrolysis at acid pH values, but its half-life in solution decreases as the pH increases beyond neutrality. However, even at pH 8 in 25 mM triethanolamine the rate of sulfhydryl formation without added primary amine was found to be negligible. On addition of dipeptide amine, the reagent reacted quickly as evidenced by the production of Ellman s reagent color. The rate of reaction also can be followed by 2-iminothiolane s absorbance at 248 nm (Xmax e = 8 840 M 1 cm 1). As the cyclic imidate reacts with amines, its absorbance at... 

Qualitatively, the presence of the nonulosaminic acids is best indicated by the brilliant-red coloration formed on addition of an acidic solution of p-dimethylaminobenzaldehyde (the so-called direct Ehrlich reaction ), and by the bright-purple coloration which develops upon boiling with Bial s reagent for several minutes at 100°. In addition, the nonulosaminic acids give a blue-violet coloration with Dische s diphenylamine reagent for deoxypentoses and a positive reaction in the tryptophan-perchloric acid test. Since no single one of these color reactions is absolutely specific for a nonulosaminic acid, it is advisable to carry out at least two of these for a qualitative analysis. For quantitative determinations, all four reactions have been employed, using either A/ -acetylneuraminic acid (m. p., 183-185° [a] —32.0°) or methoxyneuraminic acid [m. p., 200° (dec.) [a]o —55.0°] as colorimetric standards. 

Acidity, alkalinity, buffer capacity, and pH are all related and are usually readily controllable within limits by the addition of reagents. The pH must generally be kept on the acid side to prevent alkaline degradation with the attendant development of bitterness and brown color. When this reaction takes place, the pH is slowly reduced by the acidic end products until a stable pH is reached. For practical purposes, products are available from pH 3.5 to 5.5 with any reasonable quantity of suitable edible buffer salts, such as acetate, lactate, and citrate. 

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