Abstract The methanolysis of Simarouba Glauca DC oil, was studied using ZnCuAl layered double hydroxides catalysts prepared through urea hydrolysis. The influence of calcination temperature was investigated over the textural properties and catalytic activity of the ZnCuAl catalysts. The solids were pretreated at different calcination temperatures, followed by their characterization by means of XRD, SEM, FT-IR, CO2-TPD, decomposition of 2-propanol and their textural properties assessed with the aid of N2 adsorption and elemental analysis with ICP-MS. The reaction products were studied through FT-IR and gas chromatography. The XRD results showed formation of a hydrotalcite-type material after drying at 100 °C, although thermal treatment at temperatures greater than 200 °C led to formation of a metal oxides mixture. FT-IR allowed checking the evolution of the OH–, CO32– and NO3– groups after thermal treatment, which caused main changes on the basic propeties of the catalysts, evidenced through the CO2-TPD and decomposition of 2-propanol results. The catalyst calcined at 300 °C rendered a larger yield toward biodiesel production during methanolysis of the Simarouba Glauca DC oil, which is atributed to the presence of a larger concentration of medium strength basic sites. The ZnCuAl-300 catalyst showed a 6% decrease in the yield toward biodiesel production after being subjected to three reaction cycles, which was attributed to the partial segregation of metallic Cu to the catalyst surface, corroborated by XRD.