To minimize the use of volatile organic solvents in biodiesel preparation, a deep eutectic solvent (DES) consisting of choline chloride and glycerol (1:2 M ratio) was prepared and used as the co-solvent for transesterification of rapeseed oil to biodiesel catalyzed by sodium hydroxide. This eutectic solvent is inexpensive, biodegradable and non-toxic; therefore, it is a benign solvent when comparing with volatile and usually toxic organic solvents. To optimize the yield of fatty acid methyl ester (FAME), response surface methodology (RSM) and Box-Behnken Design (BBD) were employed to evaluate several key reaction parameters such as methanol/oil molar ratio, catalyst concentration and DES concentration. This study suggests that up to 98% FAME yields could be obtained at optimum conditions of 6.95 methanol/oil molar ratio, 1.34 wt% catalyst concentration and 9.27 wt% DES concentration. Data analysis and verification results further confirmed the suitability of BBD and RSM methods for optimizing the NaOH-catalyzed biodiesel synthesis. In comparison with the reaction without DES, the addition of DES in the transesterification reaction substantially improved the FAME yield, reduced the side reactions (such as saponification), and enabled a straightforward biodiesel separation and purification. (C) 2014 Elsevier B.V. All rights reserved.