A series of nitro compounds (18 aromatic and one aliphatic) was evaluated using a modification of the standard (Salmonella typhimurium mutagenicity assay. A preincubation protocol was used with flavin mononucleotide (FMN) incorporated into the assay mixture to facilitate nitro reduction. Several aromatic nitro compounds (m-nitroaniline, p-nitroaniline, 2,6-dinitrotoluene, 2,4-dinitrotoluene, 2,3-dinitrotoluene, 1,8-dinitronaphthalene), which were negative or only weakly mutagenic when tested in the standard plate incorporation assay, showed FMN-dependent mutagenic responses with this procedure. For some nitro compounds, the addition of FMN was not needed for the detection of mutagenicity in the modified protocol. Not all nitro compounds were positive using the preincubation procedure with FMN. The lack of mutagenicity, however, does not appear to be the result of the inability of the modified method to reduce nitro compounds, since it was found that reduction does occur under the assay conditions for the two nonmutagens evaluated for nitro reduction (nitrobenzene and p-nitrophenol). It is suggested that the modified protocol may be useful for evaluating the mutagenicity of many nitro compounds.