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The priority faced by energy systems in road transport is to develop and implement clean technologies. These actions are expected to reduce emissions and slow down climate changes. An alternative in this case may be the use of biodiesel produced from microalgae. However, its production and use need to be justified economically and technologically. The main objective of this study was to determine the emissions from an engine powered by biodiesel produced from the bio-oil of Chlorella protothecoides cultured with different methods, i.e., using a pure chemical medium (BD-ABM) and a medium based on the effluents from an anaerobic reactor (BD-AAR). The results obtained were compared to the emissions from engines powered by conventional biodiesel from rapeseed oil (BD-R) and diesel from crude oil (D-CO). The use of effluents as a medium in Chlorella protothecoides culture had no significant effect on the properties of bio-oil nor the composition of FAME. In both cases, octadecatrienoic acid proved to be the major FAME (50% wt/wt), followed by oleic acid (ca. 22%) and octadecadienoic acid (over 15%). The effluents from UASB were found to significantly reduce the biomass growth rate and lipid content of the biomass. The CO2 emissions were comparable for all fuels tested and increased linearly along with an increasing engine load. The use of microalgae biodiesel resulted in a significantly lower CO emission compared to the rapeseed biofuel and contributed to lower NOx emission. Regardless of engine load tested, the HC emission was the highest in the engine powered by diesel. At low engine loads, it was significantly lower when the engine was powered by microalgae biodiesel than by rapeseed biodiesel. [ABSTRACT FROM AUTHOR]