Recently, rubber seed oil (RSO) has been considered as a promising potential oil source for biodiesel production. However, RSO is a non-edible feedstock with a significant high free fatty acid (FFA) content which has an adverse impact on the process of biodiesel production. In this study, ultrasonic-assisted esterification process was conducted as a pre-treatment step to reduce the high FFA content of RSO from 40.14% to 0.75%. Response surface methodology (RSM) using central composite design (CCD) was applied to the design of experiments (DOE) and the optimization of esterification process. The result showed that methanol to oil molar ratio was the most influential factor for FFA reduction whereas the effect of amount of catalyst and the reaction were both insignificant. The kinetic study revealed that the activation energy and the frequency factor of the process are 52.577 kJ/mol and 3.53 x 10(8) min(-1), respectively.