Nanoparticles of organic carbon (NOC) are formed in combustion of hydrocarbon-rich fuels and have been detected in vehicle exhausts, suggesting their presence in urban atmospheres. Epidemiological studies showed that some causal relationships exist between particle concentration in the air and a wide range of health effects, but no toxicological studies are reported on the potential health risk of particles smaller than 4 nm. The present study investigated the mutagenicity and the reactivity of NOC collected in water samples from the exhausts of diesel and gasoline engines. Mutagenicity was tested following the Ames Test, with and without metabolic activation. Reactivity was investigated by using a new approach aimed to identify electrophilic agents present in the sample material, which if introduced into the organism, could interact with nucleophilic sites of biological macromolecules (DNA and proteins), forming adducts. Given the large number of nucleophilc sites within biological macromolecules, the complexity of NOC, and the inexact knowledge of its chemical structure, this approach was simplified by examining in vitro interactions between NOC particles and model peptides through LCIMS analyses of incubation mixtures The results indicate a high reactivity and, in several cases, the mutagenicity of NOCs, thus calling for suitable biomarkers assess NOC exposure associated with vehicle emissions.