Allopurinol: Discrimination of antioxidant from enzyme inhibitory activities

This study was designed to quantitatively discriminate the specific xanthine oxidase (XO) inhibitory from the relatively nonspecific antioxidant activities of allopurinol, both in vitro and in vivo in the rat. XO activity, determined by the spectrophotometric assay for urate generation over time, was completely inhibited in vitro by allopurinol at concentrations ≥ 200 μM. Allopurinolʹs antioxidant activity was determined in vitro using a linolenic acid peroxidation (LAP) assay. Although the known antioxidant butylated hydroxytoluene effectively inhibited LAP (80% inhibition of malondialdehyde generation at 101 μM), allopurinol (101–103 μM) did not inhibit this LAP (p < .01). Rat serum obtained after oral administration of allopurinol (100 mg/kg × 2 doses) did not suppress LAP in vitro more than did control rat serum. Following oral administration of allopurinol (2–50 mg/kg × 2 doses), dose-dependent inhibition of XO activity was observed in the homogenates of the liver (to 5% of control level; p < .001) and the intestine (to 12% of control level; p < .001). We conclude that while 2–50 mg/kg of oral allopurinol effectively suppresses XO activity in the rat liver and intestine, antioxidant activity is not seen even in doses up to 100 mg/kg. The selective enzymatic inhibitory effect of allopurinol at these doses therefore should provide a useful tool to allow the discrimination of the effects of xanthine oxidase in particular from the effects of reactive oxygen metabolites in general.