The quantitative effects of fermentation time, temperature and broth content on mycelial growth in terms of dry cell weight (DCW) and extracellular polysaccharide content (EPC) produced by the fungus Boletus spp. ACCC 50328 in submerged fermentation were investigated separately using response surface methodology (RSM). The three factors chosen for the present investigation were based on the results of a previous Plackett-Burman (PB) design. The experimental data obtained were fitted to a second-order polynomial equation using multiple regression analysis and also analyzed by appropriate statistical methods. By solving the regression equation and also by analyzing the response surface contour plots, the optimal batch fermentation conditions were determined: under conditions of fermentation time, 10.7 days; temperature. 24.23 degreesC: and broth content, 50.0 ml, EPC reached 1053.987 mug ml(-1) of fermentation liquor. When fermentation time, temperature and broth content were 10.9 days, 26.77 degreesC and 64.9 ml, respectively, DCW reached 4.216 mg ml(-1). In order to obtain simultaneously the maximum yield of EPC and DCW, the above conditions would be 10.5 days, 26.23 degreesC and 58.70 ml; and under these conditions, the maximum predicted yield of EPC and DCW were 992.258 mug ml(-1) and 4.163 mg ml(-1), respectively. These predicted values were also verified by validation experiments. (C) 2004 Elsevier Ltd. All rights reserved.