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Research on renewable fuels has to assess possible adverse health and ecological risks as well as conflicts with global food supply. This investigation compares the two newly developed biogenic diesel fuels hydrotreated vegetable oil (HVO) and jatropha methyl ester (JME) with fossil diesel fuel (DF) and rapeseed methyl ester (RME) for their emissions and bacterial mutagenic effects. Samples of exhaust constituents were compared after combustion in a Euro III heavy duty diesel engine. Regulated emissions were analyzed as well as particle size and number distributions, carbonyls, polycyclic aromatic hydrocarbons (PAHs), and bacterial mutagenicity of the exhausts. Combustion of RME and JME resulted in lower particulate matter (PM) compared to DF and HVO. Particle numbers were about 1 order of magnitude lower for RME and JME. However, nitrogen oxides (NOx) of RME and JME exceeded the Euro III limit value of 5.0 g/kWh, while HVO combustion produced the smallest amount of NOx. RME produced the lowest emissions of hydrocarbons (HC) and carbon monoxide (CO) followed by JME. Formaldehyde, acetaldehyde, acrolein, and several other carbonyls were found in the emissions of all investigated fuels. PAH emissions and mutagenicity of the exhausts were generally low, with HVO revealing the smallest number of mutations and lowest PAH emissions. Each fuel showed certain advantages or disadvantages. As proven before, both biodiesel fuels produced increased NOx emissions compared to DF. HVO showed significant toxicological advantages over all other fuels. Since jatropha oil is nonedible and grows in arid regions, JME may help to avoid conflicts with the food supply worldwide. Hydrogenated jatropha oil should now be investigated if it combines the benefits of both new fuels.