A deep eutectic solvent (DES) consisting of choline chloride and glycerol (1:2 molar ratio) was prepared and used as the solvent in CaO-catalyzed transesterification of rapeseed oil to produce biodiesel. In addition to its solvent role, DES activated commercial CaO to be a catalyst for the transesterification reaction. The yield of fatty acid methyl ester (FAME) with the addition of DES reached 91.9%, while the FAME yield with no DES was only 4.0%. The commercial CaO samples before and after reaction were characterized by Powder X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), and Scanning Electronic Microscopy (SEM). These measurements indicate that the surface of commercial CaO is typically covered by a layer of CaCO3 and Ca(OH)(2) which inhibits the activity of CaO, while the addition of DES can eliminate this inhibiting layer leading to CaO activation.