Response surface methodology (RSM) examining the effects of five-level-three-factors and their mutual interactions was utilized to optimize the fermentation conditions to enhance capsular polysaccharide (CPS) production of Streptococcus pneumoniae type 3. Twenty experiments conducted in an 8-l lab-scale fermentor were designed to assess fermentation pH, supplemented glucose concentration, and stirring rate. The predicted highest CPS production by the obtained optimization model equation was 256.14 mg/l at optimal conditions [pH 7.5, stirring rate 180 rpm, and supplemented glucose concentration 1% (w/v)]. The validity of the response model was confirmed by the good agreement between the predicted and experimental results. The maximum amount of CPS obtained was 255.03+/-2.23 mg/l.