Sequential optimization strategy, based on statistical experimental designs, was employed to enhance the production of uricase by Pseudomonas aeruginosa local isolate. Glucose supplementation to the basal production medium inhibits uricase production, perhaps by catabolic repression. For screening of bioprocess parameters significantly influencing uricase activity, the two-level Plackett-Burman design was used. Among fifteen variables tested; pH, CuSO4 and FeSO4 were selected based on their high significant effect on uricase activity. A near optimum medium formulation was obtained using this method with increased uricase yield by 15-folds. Response surface methodology (RSM) was adopted to acquire the best process conditions. In this respect, the three-level Box-Behnken design was employed. A polynomial model was created to correlate the relationship between the three variables and uricase activity. The optimal combination of the major constituents of media for uricase production evaluated from the non-linear optimization algorithm of EXCEL-Solver was as follows: pH, 5.5; CuSO4, 10(-3) M; and FeSO4, 10(-2) M. The predicted optimum uricase activity was 7.1 U/ml/min, which is 16.5 times than the basal medium. (c) 2004 Elsevier Ltd. All rights reserved.