Aspergillus oryzae was co-cultured with Trichoderma reesei using acorn cups extract containing up to 62% ellagitannins as substrate to produce ellagic acid with relatively high levels of ellagitannin acyl hydrolase, cellulase and xylanase. Ellagitannins concentration, initial pH, T reesei and A. oryzae during the fermentation were identified as important process parameters effecting ellagic acid accumulation and the enzymes syntheses. These parameters were optimized by uniformity design to determine the optimum condition for ellagic acid production. Under optimum operational condition, ellagic acid yield could be arrived at 24%, when the fermentation run lasted 96 h with an initial pH of 4.5, an ellagitannins concentration of 4 g 1(-1), T reesei of 3 ml and A. oryzae of 3 ml. Meanwhile, it was found that the three enzymes, activities correlated very well with ellagic acid yield, resulting in model with high coefficient of determination (R-2 = 0.98). The results indicate that the mixed culture of T reesei and A. oryzae is an effective approach to produce an enzyme system of degrading ellagitannins for ellagic acid production. (c) 2007 Elsevier Ltd. All rights reserved.