Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) ; A Plackett-Burman experimental design was used to evaluate seven medium components added to molasses (corn steep liquor, sodium acetate, magnesium sulfate, manganese sulfate, ammonium citrate, potassium phosphate and Tween 80). Corn steep liquor (CSL), K2HPO4 and Tween 80 increased lactic acid production. The concentrations of these three components as well as the molasses were further optimized using the response surface method. A maximal lactic acid production of 94.8 g L-1 was obtained when the concentrations of molasses, CSL, K2HPO4 and Tween 80 were 193.50 g L-1, 37.50 mL L-1, 2.65 g L-1 and 0.83 mL L-1, respectively. However, in both shaker and bioreactor, approximately one fourth of the sugar added initially was not utilized after 48 hours of fermentation. Future studies that consider high conversion of sugar into final product as well as high volumetric productivity are necessary to improve the fermentation process and to reduce the downstream costs.