Petrol (gasoline) contains a number of toxicants. This study used human biomonitoring to evaluate the genotoxic effects of exposure to benzene in petrol fumes in 100 Indian petrol-pump workers (PPWs) and an equal number of controls. The study was corroborated with in silico assessments of the Comet assay results from the human biomonitoring study. An in vitro study in human lymphocytes was also conducted to understand the genotoxicity of benzene and its metabolites. In a subset of the population studied, higher blood benzene levels were detected in the PPWs (n = 39; P < 0.01) than the controls (n = 18), and 100-250 ppb benzene was also detected in air samples from the petrol pumps. PPWs had higher levels of DNA damage than the controls (P < 0.01). In addition, the micronucleus assay was performed on lymphocytes from a subset of the subjects, and the micronucleus frequency for PPWs was significantly higher (n = 39; 14.79 +/- 3.92 per thousand) than the controls (n = 18; 7.54 +/- 3.00 per thousand). Human lymphocytes were treated in vitro with benzene and several of its metabolites and assayed for DNA damage with the Comet assay. Benzene and its metabolites produced significant (P < 0.05) levels of DNA damage at and above concentrations of 10 microM. The metabolite, p-benzoquinone, produced the greatest amount of DNA damage, followed by hydroquinone > benzene > catechol > 1,2,4,-benzenetriol > muconic acid. This study demonstrates that, using sensitive techniques, it is possible to detect human health risks at an early stage when intervention is possible. possible.