Isolation and chemical-structural identification of a novel aromatic aminemutagen in an ozonized solution of m-phenylenediamine

The mutagenicity of a m-phenylenediamine (m-PD) solution was markedly enhan ced by oxidation with ozone. The ethyl acetate extracts from a m-PD solutio n ozonized at pH 10.7 were fractionated by normal-phase and reversed-phase column chromatography to isolate mutagens by monitoring mutagenic activitie s on Salmonella typhimurium TA98 in the presence of a mammalian metabolic a ctivation system (S9 mix). From fraction 5-3-2, which exhibited the stronge st mutagenicity (308 000 revertants/mg), a major mutagenic compound was iso lated. On the basis of the high-resolution EI-mass, H-1 NMR and C-13 NMR sp ectral, and X-ray crystallography data, the structure of this compound was determined to be 2-amino-5-[(3-aminophenyl)amino]-4-[(3-aminophenyl)imino]- 2,5-cyclohexadien-1-one (PDT-1), which is a novel compound. PDT-1 is a newl y identified frame-shift type mutagen, inducing 65 400 revertants and 295 0 00 revertants of S. typhimurium TA98 and YG1024 per micromole, respectively , in the presence of S9 mix. When a m-PD solution was oxidized with 1 or 2 mol of ozone at pH 4.0, 7.0, and 10.7, the contribution of PDT-1 to the mut agenicity of ethyl acetate extracts from the ozonized m-PD solution was 5-2 3%.