This study evaluated the effect of biomass (soybean oil, eucalyptus sawdust, and coffee grounds) pyrolysis oil on the formulation of diesel fuels. The parameters analyzed were ignition delay time, emission of particulate matter and unburned hydrocarbons, and specific fuel consumption. The fraction of pyrolysis oil used as fuel was obtained by vacuum distillation at 80-240 degrees C. The use of this fraction resulted in a decrease in the ignition delay time in the combustion process, with the resulting increase in the cetane number due to the presence of phenolic groups in the pyrolysis oil, which modify the formation mechanism of peroxyl radicals by altering the temperature of the flame front. Additionally, particulate matter emissions are reduced significantly by up to 30% when compared with the base fuel. This is probably due to the high solubility of water in pyrolysis oil, which leads to the formation of an azeotropic mixture that lowers the boiling point and contributes to vaporize the fuel inside the combustion chamber, reducing the formation of particulate matter. These results indicate the promising potential of this fraction for use in the formulation of diesel fuel, decreasing ignition delay and increasing the cetane number, as well as significantly reducing particulate matter emissions. The main difficulty in using this fraction of pyrolysis oil is its chemical stability, since it has a strong tendency to form oligomers.