For improving the biodiesel yield from a transesterification reaction, an experimental validation of the optimal control is essential. An experimental setup involving the control of biodiesel production process is developed. The activation energies (Ea) involved in the kinetic model are estimated from the obtained experimental data. The hot water flow rate through the reactor coil is optimized by solving a dynamic optimization problem (DOP) with estimated Ea values for maximization of the biodiesel concentration at the final time. The DOP produces marginal improvement in the endpoint biodiesel concentration by 2.88% and reduces the reactor energy cost by 3.58% compared to the use of the published Ea values and the base case, respectively. Using the estimated Ea values, the batch time decreases by 44.44% compared to the literature study, for achieving the optimal biodiesel concentration. Therefore, the optimal control established through our experimental work improves the biodiesel yield while satisfying energy -efficiency. (C) 2020 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.