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A novel method using Plackett-Burman design (PBD) coupled with Draper-Lin small composite design (SCD) and desirability function (DF) was developed to optimize Tuber melanosporum fermentation medium. Firstly, sucrose, yeast extract, peptone and Mg(2+) were identified to be key medium components by PBD. Secondly, in order to evaluate the relationships between responses and the identified key components, mathematical models were developed by SCD. Finally, by using DF, the responses were optimized simultaneously and the optimal concentration was located to be 73 g/L sucrose, 11 g/L yeast extract, 8 g/L peptone, and 46 mM Mg(2+). Under the identified optimal conditions, the maximal biomass, the production of extracellular (EPS) and intracellular (IPS) polysaccharides was 25.10 + or - 0.12 g/L, 3.88 + or - 0.23 g/L and 2.87 + or - 0.32 g/L, respectively, which agreed with the predicted values well. Compared with the basic medium, the biomass, the production of EPS and IPS was enhanced by 54.4%, 71.7% and 124.2%, respectively.

For the first time, a fed-batch fermentation process of Tuber melanosporum was developed for the efficient production of bioactive mycelia and Tuber polysaccharides. Each 1.67 g/L of peptone and 8.33 g/L of yeast extract were added on day 3, 6, and 9, respectively, and sucrose was fed to maintain its concentration around 35-5 g/L when its residual level decreased to 10-5 g/L. Then, the maximal biomass, the production of extracellular polysaccharides (EPS) and intracellular polysaccharides (IPS) reached 53.72 +/- 2.57 g DW/L, 7.09 +/- 0.62 and 4.43 +/- 0.21 g/L, respectively. Compared with the batch culture conducted in the enriched medium, the biomass, the production of EPS and IPS were enhanced by 55.8%, 222.3% and 103.2%, respectively. Not only the cell density but also the production of EPS and IPS were the highest ever reported in truffle fermentation, and the biomass was also the highest as ever reported in mushroom fermentation. (C) 2009 Elsevier Ltd. All rights reserved.