The N-heterocyclic aromatic pollutant, 7H-dibenzo[c,g]carbazole (DBC), is a potent carcinogen having both local and systemic effects. The overall objective of this research was to investigate the nature of the covalent binding of DBC with nucleic acids in vitro. DBC was shown to bind to polynucleotides, RNA and DNA in an in vitro rat or hamster microsomal enzyme assay, exhibiting a preferential binding to polyguanylic acid (poly[G]). Benzo[a]pyrene (BaP) binding to these same nucleic acids was determined simultaneously and was approximately 10-fold higher than DBC binding under identical experimental conditions. DBC-nucleic acid binding was shown to be dependent upon the presence of a microsomal activating system, the results being similar for rat or hamster liver microsomes. This microsome-dependent binding was unaffected by the addition of epoxide hydrase activity modifiers but was almost completely inhibited by alpha-naphthoflavone. The nature of DBC-nucleic acid binding was investigated using fluorescence spectroscopy. Benzo[c]carbazole and 5,5,6,6-tetrahydrodibenzo[c,g]carbazole were synthesized as representatives of the effect of disruption of the DBC pi-electron system on fluorescence excitation and emission. DBC-poly[G] adducts were isolated from binding assay mixtures and separated by HPLC. Results indicated that there are at least three different DBC-poly[G] adducts formed in vitro. The emission spectra of isolated adducts were similar in shape to that of DBC; however, the adduct spectra were shifted 5-10 nm toward longer wavelengths. This suggests that the bound DBC species have intact pi-electron systems. Results are consistent with binding through the nitrogen position as well as binding through the 1,2,3,4-ring of the molecule.