IDENTIFICATION OF AN N-HYDROXYGUANIDINE REDUCING ACTIVITY OF XANTHINE-OXIDASE

A guanoxabenz (2.6-dichlorobenzylideneamino)-3-hydroxyguanidine; an N- hydroxyguanidine] reducing enzymatic activity of rat spleen cytosol wa s investigated By means of protein purification and N-terminal amino a cid sequencing, the reducing activity was shown to reside in xanthine oxidase. The action of the enzyme on guanoxabenz resulted in the forma tion of guanoxabenz [1-(2,6-dichlorobenzylidene-amino)-3-guanidine] : the product formation could be monitored by HPLC and its identity was confirmed by NMR,analysis. The reduction of guanoxabenz required xanth ine or NADH as reducing substrates, while the process could be blocked by allopurinol, a selective inhibitor of xanthine oxidase. By using b ovine milk xanthine oxidase, the guanoxabenz reducing activity of the enzyme was also verified. We conclude that guanoxabenz is a novel elec tron acceptor structure fur xanthine oxidase.