Biodiesel has become one of the essential fuels in the present and future, and it can be produced from vegetable oil and animal fat. However, current feedstocks to produce biodiesel slow down the growth of biodiesel implementation due to the high cost of feedstock. As a result, rice bran oil (RBO) is claimed to be a potential feedstock for biodiesel production. A non-catalytic in-situ biodiesel production from low cost feedstock (RBO) using subcritical ethanol-water mixture probably decrease the production cost further and environmentally benign. Therefore, in this work the influence of four independent variables, adding co-solvent/without co-solvent, ethanol concentration, temperature, and time of reactions on the yield of biodiesel was examined tentatively. The independent variables limitations were (a) co-solvent of n-hexane, ethyl acetate and chloroform, (b) ethanol concentration of 20 - 80 %, v/v, (c) temperature of reaction 120 - 200 o C, and (d) time of reaction 1 - 4 h employed to direct the trials. In this examination historical data design was utilized and a quadratic polynomial model was built up and after that at last optimized by using response surface methodology (RSM). It was found that the yield of biodiesel achieved an optimum value of 79.79 %, v/v using ethyl acetate as co-solvent with ethanol concentration of 78.83 % (v/v), under constant operation conditions of P = 8 Mpa, T = 160 o C, t = 2 h, N = 400 rpm, ratio of rice bran (RB): solvent: co-solvent= 10 (g): 80 mL: 20 mL. In the other hand (rice bran 2), it was found that the yield of biodiesel achieved an optimum value of 76.98 % for 3.2 h of reaction time and temperature of reaction 200 o C, under constant operation conditions of P = 8 Mpa, co-solvent = ethyl acetate, ethanol concentration = 80 %, v/v, N = 400 rpm, ratio of RB: solvent: co-solvent= 10 (g): 80 mL: 20 mL. The examination has likewise uncovered that authentic plan information with RSM is a well-organized statistical technique for forecasting the optimum operating conditions of a non-catalytic in-situ biodiesel production from rice bran using subcritical ethanol-water mixture assumes a crucial role of ethanol concentration, types of co-solvent, temperature and time of reactions.