Abstract: In the present study, efforts have been made to statistically optimize the formulation parameters of sustained-release lipospheres of Enalapril maleate, a water-soluble peptide, using wax and polar lipid combination. A 3-factor, 3-level Box-Behnken design was used to derive a second order polynomial equation and construct 3D surface plots to predict the effect on responses. The peptide amount (X1), Tween® 80 concentration (X2) and stirring speed (X3) were selected as independent variables while the mean diameter (Y1) and the entrapment efficiency (Y2) of lipospheres were chosen as the dependent variables in the present investigation. Lipospheres were prepared by Water-in-oil-in-water double emulsion (w/o/w) method. The Response Surface Methodology (RSM) and multiple response optimization were used to select optimal formulation with maximum entrapment and particle size in range. The optimal formulation was subsequently characterized in terms of morphology, release kinetics and stability studies. The maximum entrapment (80.62±2.54%) was achieved with 10mg EM, 0.1% (v/v) Tween® 80 and 1000 rpm stirring speed. The observed responses coincided well with the predicted values from the RSM optimization technique. Scanning electron microscopy confirmed the smooth spherical microspheres in the size range of 23.00±0.82-34.57±1.04 μm. Kinetic models revealed that drug release followed non-Fickian release pattern. Finally, drug bioactivity was found to remain intact after microencapsulation. Thus, Box-Behnken design demonstrated the role of the derived equation and 3D surface plots in predicting the values of dependent variables for the preparation and optimization of Enlapril maleate loaded lipospheres.