Fluorescence line narrowing (FLN) was demonstrated for five benzo[a]pyrene (BP)-nucleoside adducts synthesized by one-electron oxidation of BP in the presence of guanosine, deoxyguanosine, and deoxyadenosine. The standard FLN spectra were used to prove that a major depurination adduct from the binding of BP to DNA in rat liver nuclei is 7-(benzo[a]pyren-6-yl)guanine (N7Gua). The structural characterization was performed with only 20 pg of the adduct. Metabolic activation of BP by one-electron oxidation in the horseradish peroxidase catalyzed reaction of BP with DNA (in vitro) was also investigated. The major adduct identified was 8-(benzo[a]pyren-6-yl)guanine (C8Gua).