The use of pressurized carbon dioxide for the purification of biodiesel was demonstrated. A given amount of carbon dioxide was injected in a high pressure equilibrium cell containing a crude biodiesel mixture, which consisted of non-reacted glycerides (diacylglycerols and monoacylglycerols), fatty acid methyl esters, soaps (sodium carboxylates), glycerol and methanol. The cell was then pressurized in the range of 6-12 MPa at temperatures varying from ambient (298 K) to 323 K. The amount of carbon dioxide injected in the crude biodiesel varied from 20 to 50 in mass percentage. Two phases were present at equilibrium under all conditions. By means of one-way valve, the lighter phase was sampled and its composition was analyzed using gel permeation chromatography (GPC). Results were then compared to conventional biodiesel purification steps based on water washing and adsorption. The characterization was carried out in accordance to the specification standards given by ANP (National Agency of Petroleum, Natural Gas and Biofuels, Brazil). An experimental design was performed to investigate the influence of carbon dioxide composition, pressure and temperature. The results have shown that the process is able to produce a biodiesel fraction with significantly decreased levels of diacylglycerol, monoacylglycerol, sodium, glycerol and methanol. The temperature is the variable that most affects the purification of biodiesel, while the carbon dioxide concentration and pressure have minor effects. The best result was obtained at room temperature. (C) 2014 Published by Elsevier B.V.