Abstract: Response surface methodology (RSM) based on a three-level three-factor Box-Behnken design of experiment was used to optimize the exopolysaccharide content (EPC) and the mycelium biomass in submerged cultivation by Clitocybe sp. AS 5.112. The critical factors selected for the investigation were temperature, time of cultivation and volume of medium, based on the results of previous Plackett-Burman design. By analyzing the response surface plots, the optimum ranges of cultivation temperature, time and medium volume for obtaining over 1 253.00 ?g·ml-1 of EPC were in 24.3－25.8℃, 9.7－10.2 d and 76.0－90.0 ml, respectively. While for obtaining over 8.32 mg·ml-1 of dry cell weight (DCW), the above variables would be in the range of 23.8－24.8℃, 9.6－10.3 d and 71.0－98.0 ml, respectively. By solving the inverse matrix from the quadratic regression equations, the optimal conditions for obtaining 1 265.45 ?g·ml-1 of EPC were 25.0℃, 9.9 d and 83.4 ml, to gain 8.50 mg·ml-1 of DCW were 24.4℃, 9.9 d and 87.1 ml. In order to obtain the maximum yield of EPC and DCW at the same time, the above conditions would be 24.5℃, 9.9 d and 84.7 ml, respectively, in this situation, the maximum predicted EPC and DCW were 1 261.60 ?g·ml-1 and 8.47 mg·ml-1, respectively. The experimental data under various conditions have validated the theoretical values.

Key words: Clitocybe sp., Optimization, Plackett-Burman design, Box-Behnken design, Response surface methodology