An economical and environmentally benign synthesis method remains important in industrial processes. In this work, kaolin an abundant and non-toxic precursor is employed for mesoporous gamma-alumina synthesis. The study explored the consequence of varying polyoxyethylene (40) stearate (PS) surfactant concentration and aging time in the synthesis of mesoporous gamma-alumina with enhancing structural properties. The alumina was characterized using X-Ray Powder Diffraction (XRD), Thermogravimetric-Derivative Thermal Gravimetric (TG-DTG), Fourier Transform Infrared (FTIR), N2 Adsorption–Desorption, Field Emission Scanning Electron Microscopy (FESEM), Solid State 27Al MAS NMR, and X-Ray Fluorescence (XRF). Structural properties and morphology of the mesoporous γ-Al2O3 improved with increasing surfactant amount from 0.45 g to 1.8 g and aging time from 1 to 2 days, then dropped afterward. Accordingly, mesoporous γ-Al2O3 synthesized with 1.8 g PS and age for 2 days (Al-1.8-2) possessed the highest surface area of 319.2 m2/g, a narrow average pore diameter of 2.7 nm, and pore volume of 0.42 cm3/g. Thus, Al-1.8-2 was chosen and doped with 15% NaOH to obtained 15-Na/Al-1.8-2 catalyst. After modification, the basic sites increased, the surface area decreased, and the morphology changed, signifying Na incorporation into the alumina. On application in methanolysis reaction, up to 99.46% biodiesel yield was obtained, demonstrating a very high activity for the catalyst. The tremendous activity of the catalyst can be associated with the enhanced structural properties of the mesostructured gamma-alumina.