Abstract: The aim of this study was to optimize the main media components that affect the production of lactic acid by Lactobacillus rhamnosus fermentation. In the experiment, the two variables (glucose and peptone) were optimized using the Central Composite Design (CCD) method by STATISTICA Software. The optimization studies were successfully carried out in shake flask experiments. The results indicated that the optimum concentration of glucose and peptone for optimum bacterial growth rate and lactic acid production in shake flask were 9.80 and 9.98 g L-1, respectively. The optimum productivity of the lactic acid was 0.630 g g-1 h which correspond to optimum growth rate of the bacteria at 0.341 h-1.