Laccases were produced and characterized during the process of cultivating Pleurotus sajor-caju P5-2001 in stirred-tank bioreactor. The mycelial biomass reached 5.2 g L(-1) on the fifth day, while sucrose was fully consumed after eight days and a high laccase activity peak (40 U mL(-1)) was reached on the sixth day of cultivation. Previous studies carried out using the enzymatic broth in an activity gel with ABTS as the substrate indicated the presence of three different laccase proteins. The optimal pH values for the activity of the laccases were 2.4, 3.2, and 4.4, respectively, using ARTS as the substrate. The optimal temperature was determined using various buffers and pH (sodium acetate pH 5.0, sodium citrate pH 5.0, and Mc'llvaine pH 3.2). The best results were obtained at 50 degrees C for all conditions tested with ABTS as the substrate. Notable thermal stability, with high levels of relevant enzymatic activities, was seen at 20 degrees C and 30 degrees C over a period of 168 h. At 40 degrees C, activity showed a considerable decrease after 36 h, and at 50 degrees C and 60 degrees C, there were sharp decreases after only a few hours. (C) 2010 Elsevier Ltd. All rights reserved.