Lactoperoxidase-catalyzed metabolism of N-hydroxy-N-2-fluorenylacetamide (N-OH-2-FAA) may be via one-electron oxidation to nitroxyl free radical which dismutates to equimolar N-acetoxy-N-2-fluorenylacetamide and 2-nitrosofluorene (2-NOF) and/or a Br(-)-dependent oxidative cleavage to 2-NOF. Hence, the 2-NOF:N-acetoxy-N-2-fluorenylacetamide ratios reflect the relative contributions of the two peroxidative pathways to the metabolism of N-OH-2-FAA. Peroxidative activities of rat uterus (UT) and mammary gland (MG) were extracted with a cationic detergent, cetyltrimethylammonium bromide (Cetab). MG extracts had 1 to 5% the specific activity of UT extracts when assayed with guaiacol as hydrogen donor. At 0.004% Cetab, which in the incubation media corresponds to the approximate physiological levels of 0.1 mM Br(-), oxidation of N-OH-2-FAA by UT extracts yielded a product ratio indicative of both peroxidative pathways with Br(-)-dependent oxidation prevailing. At 0.4% Cetab, one-electron oxidation was negligible and Br(-)-dependent conversion of N-OH-2-FAA to 2-NOF was markedly enhanced. At both 0.004 and 0.4% Cetab, MG extracts yielded only 2-NOF, suggesting solely Br(-)-dependent oxidation. With equivalent guaiacol units of peroxidative activities, MG extracts produced much lower amounts of 2-NOF than did UT extracts. The low specific activities of MG extracts necessitated the use of larger amounts of protein, which might have interfered with peroxidative metabolism. At 0.004% Cetab, formation of 2-NOF by the Br(-)-dependent pathway was greater at pH 5.5 than at 7.4. At acid pH, small amounts of 2-nitrofluorene were also formed by UT and MG extracts and could be attributed to further oxidation of 2-NOF. Peroxidative activities of the UT and MG extracts may be of granular leukocyte origin and their potential role in carcinogen activation and tumorigenesis is discussed.