DETECTION OF SACCHARIDES USING DIAZO-COMP OUND AS A COLOR REAGENT

We investigated the reaction of saccharides with six diazo-compounds: sulfanilic acid, 4-aminobenzoic acid, 3-aminobenzoic acid, 4-iodoanili ne, 4-chloroaniline and 4-aminoacetophenone. Five reduced sugars, gluc ose, fructose, xylose, maltose and lactose, reacted with various diazo -compound to form colored products under alkaline conditions, a nonred uced sugar, sucrose, did not react. The absorption maximum of the colo red product varied with the diazo-compound, correlating with the Hamme tt substituent constant of the substituent attached to the benzene rin g on the aromatic primary amine, but there was little difference in th e absorption spectra of the five sugars reacted with the same diazo-co mpound. Their absorption maxima ranged from 530 to 540 nm and their mo lar absorptivities from 216 to 649 l mol(-1) cm(-1) in the case of the reaction of diazotized 4-aminobenzoic acid, which gave the colored pr oduct with the largest color intensity. The absorption spectrum of the colored product indicated a reversible change when hydrochloric acid or sodium hydroxide were added, like that of a pH indicator. In the co lor reaction of the reduced sugars, a lag time of 3 similar to 18 min was required before the color reaction began. The reaction rate of the ketose, fructose, was higher than that of glucose, xylose, maltose an d lactose, the reaction time being about 60 min for fructose and about 140 min for the other reduced sugars. The colored product was easily oxidized by air after completion of the reaction, but was stabilized b y addition of ascorbic acid. Since nonreduced sugar did not react with diazo-compound, it seems that the carbonyl groups formed in the reduc ed sugars under alkaline conditions take part in this color reaction. hs this color reaction of the sugars occured at room temperature, it m ay be of use for their detection.