INHIBITION OF XANTHINE OXIDASE 285 



formed from 6-chloropurine by the action of xanthine oxidase, K^ being 

 0.00006 mif (Duggan and Titus, 1959). It inhibits urate degradation in 

 the rat 75% at a dose of 20 mg/kg (Duggan et al., 1961). The over all AF 

 for the binding to uricase is approximately 10 kcal/mole and this would 

 indicate the formation of some type of stable bond. It was pointed out that 

 this analog is very stable and might be useful in studying the metabolism 

 and disposition of urate. It should induce urate accumulation in the tissues 

 and the effects of this might have some bearing on the manifestations of 

 gout. The inhibition of uricase by xanthine (2,6-dihydroxypurine) in the 

 tabulation above is interesting because it illustrates a novel effect in a multi- 

 enzyme system. In the sequence: 



xanthine 



oxidase uricase 

 Xanthine > urate > allantoin 



the initial substrate inhibits the second enzyme in the series, causing ac- 

 cumulation of urate in increasing amounts as the xanthine concentration 

 rises (e.g., in rat liver homogenates). At high concentrations the urate con- 

 centration falls due to the substrate inhibition of xanthine oxidase (Van 

 Pilsun, 1953). 



Further studies on uricase have been reported by Bergmann et al. (1963 a) 

 and some of the results, including calculations of the approximate apparent 

 relative binding energies, are presented in the following tabulation. The 

 xV-methylpurines are relatively weak inhibitors and are not included in 

 the tabulation. It is clear that the best inhibitors contain a 2-OH group 

 (designated as OH for convenience but the keto form is probably dominant), 

 and this position is the most important in the binding; substitution of the 

 2-0II with a 2-SH group lowers the inhibitory activity markedly. The 

 weakening of the binding by A^-substitution points to the importance of 

 the imino group for attachment. The inhibitions are generally" sensitive to 

 the pH and for most analogs increase with a rise in pH, although for 6- 

 SH-8-OH-purine there is a decrease, and with 8-OH-purine there is no 

 effect. The pH probably influences the tautomerism, which is quite impor- 

 tant since the binding depends on the states of the N and OH groups. The 

 K^ for 2,8-diOH-6-SH-purine is 0.0026 niM and for 2,6-diOH-8-SH-purine 

 is 0.00039 mM (Bergmann et al, 1963 b). The 2-OH-6,8-diSH-purine (which 

 is 6,8-dithiourate) is a much more potent inhibitor and, although its K^ 

 was not given, it must be at least around 0.00004 mil/. 



Pteridines 



The inhibition of xanthine oxidase by synthetic folate, reported by 

 Kalckar and Klenow in 1948, was soon found to be due to some impurity, 

 and the simultaneous observation, by Lowry and Bessey, of the very po- 



