Imidazo-1,2,3-triazines as substrates and inhibitors for xanthine oxidase.

A new group of compounds analogous to the purines was recently described (1) in which carbon atom 2 of the purine ring was replaced by nitrogen. One of these imidazo-1 ,2,3-triazines (I), conveniently called 2-azaadenine, was shown to be an antagonist of hypoxanthine and of adenine in its toxic action against bacteria and mice. The corresponding hydroxy compound, 2-azahypoxanthine, was relatively inert. In this paper are reported the results of a study of these 2-axapurines as inhibitors of the enzyme, xanthine oxidase, in the oxidation of the normal substrates, hypoxanthine and xanthine. Milk xanthine oxidase, purified as described by Ball (2) and Kalckar (3), was used. The enzymatic reactions were followed spectroscopically in the ultraviolet region (4). The 2-azapurines themselves were attacked by xanthine oxidase with consequent changes in spectra. Therefore, in studying the competition of the azapurines with hypoxanthine or xanthine for xanthine oxidase, special care was taken to obtain measurements which permitted observation of the simultaneous oxidation of both normal substrate and analogue. By following the oxidation of mixtures at several selected wavelengths, it was found that with equimolar azaadenine and hypoxanthine the azaadenine was almost completely oxidized before the hypoxanthine was greatly changed. In the case of azaadenine and xanthine, the attack of both substrates was simultaneous at more nearly equivalent rates. W-Axapurines As Substrates for Xanthine Oxidase-When xanthine oxidase was added to a solution of azaadenine, the spectrum of the latter changed, indicating that it was being oxidized. When action ceased, new maxima were found, the most notable of these being the intense peaks at 270 and 224 rnp (Fig. 1). Azahypoxanthine was also attacked by the enzyme. The new absorption maxima formed in the oxidation of these 2-azapurines served as convenient points at which to measure the speed of their oxidations by following the changes in optical density with time. The relative rates of oxidation of equimolar amounts of azaadenine, azahypoxanthine, and hypoxanthine by the same quantity of xanthine oxidase