In the present study, Annona squamosa seed oil has been evaluated as a potential feedstock for biodiesel production. The response surface methodology was used to determine the optimal conditions for the biodiesel production using a central composite design. A quadratic polynomial was developed to predict the response as a function of independent variables and their interactions and only the significant factors affecting the yield were fitted to a second-order response surface reduced 2 factor interaction (2FI) model. Four process variables were assessed at five levels. A biodiesel yield of 98.19% was obtained at optimum conditions: 7.53:1 methanol to oil molar ratio, 1.18 wt% catalyst concentration, reaction temperature of 59.55 degrees C, and reaction time of 47.29 min.