Biodiesel is considered an alternative replacement for petroleum diesel, and enzymatic reaction is one method for biodiesel production. Understanding the kinetics of this reaction is important to achieve higher production rates. The kinetics of transesterification of castor oil using Novozym 435 was investigated in this study. The genetic algorithm was employed for estimation of kinetic parameters including Vmax, KmTG, and KmA in the Ping Pong kinetic model at different temperatures. In most enzymatic reactions that follow the Ping Pong mechanism, inhibition by alcohol has been taken into consideration. Here, we examined a competitive inhibition mechanism and found no inhibition effect at high alcohol‐to‐oil molar ratios of 12.6, 15.6, and 18.9 mol/mol. Arrhenius expressions were used to fit the kinetic parameters for reaction temperatures of 30–50 °C. An acceptable agreement with experimental data at 45 °C was observed when biodiesel yields were predicted using the temperature‐dependent parameters, thus validating the proposed model.