The aim of this study is to test the behaviour of sunflower biodiesel in a diesel engine after being treated with a natural antioxidant additive produced from bio-oil extraction (final dosage of bio-oil compounds in doped biodiesel of 1.9 wt%). The influence of this renewable additive in both the engine performance and the produced emissions was evaluated. Five more fuels were used for the sake of comparison: petroleum diesel, neat sunflower biodiesel without additives, commercial biodiesel, commercial B10 blend and another B10 blend prepared from petro-diesel and doped sunflower biodiesel. Brake power was found to be similar for the six fuels, while the brake specific fuel consumption and the brake thermal efficiency were higher for biodiesel fuels. Only slight differences ( < 1%) were observed between the doped biodiesel and the neat one, showing that the bio-oil based additive did not negatively affect the general performance of the engine. Regarding gas emissions (analysed according to the European Stationary Cycle), weighted average emissions of NOx, and CO2 were higher for biodiesel fuels, while CO and opacity factor were lower in that case. Incorporating the bio-oil based additive reduced NOx emissions and smoke opacity by 3.0% and 4.4% compared with neat biodiesel, respectively, whilst CO and HC emissions increased by 0.7 and 14.3% respectively, values still remaining below those of diesel.