Gas chromatography/mass spectrometry (GC/MS) of diesel fuel aromatics detected polynuclear aromatic hydrocarbons from naphthalenes to phenanthrenes, but no four- or five-ring aromatics. This aromatic fraction treated with NO2 was found to contain nitro-aromatics, but only the naphthalene and biphenyl nitro-aromatics were detectable by direct GC/MS. By reduction of the nitro groups to amines, diazotization and reduction to yield aryl-iodides, it was possible to demonstrate that nitro-derivatives of most of the starting aromatics were present after NO2-treatment. Diesel fuel was separated into aliphatic and aromatic fractions by extraction with diethyl sulfoxide. These fractions were devoid of mutagenic activity in the Ames bioassay and exhibited low cytotoxicity to CHO cells in culture. However, after reaction with NO2, the products contained frameshift mutagens which did not require activation by S-9 microsomal enzymes. The biological activity of the NO2-treated aromatic fraction from fuel was more than 40 times greater in Salmonella TA100 than fuel aliphatics treated with NO2. The LC50 to CHO cells in culture increased more than fivefold for aromatics and more than tenfold for aliphatics. Similar to the diesel exhaust particulate extract, the cytotoxicity of the nitrated fractions was decreased by serum and glutathione. Reaction of fuel aromatics with NO2 may be one mechanism which contributes to the formation of cytotoxic and mutagenic activities in diesel exhaust.

The dichloromethane extract of a coal combustion fly ash sample obtained from an experimental fluidized bed coal combustor was tested for mutagenicity in Salmonella typhimurium and cultured Chinese hamster ovary (CHO) cells. The extract was directly mutagenic in S typhimurium strain TA98 and the nitroreductase deficient strains TA98NR and TA98/1,8DNP6. The mutagenicity observed in TA98NR and TA98/1,8DNP6 was lower than that in TA98. Addition of exogenous Aroclor 1254-induced rat liver supernatant (liver S9) decreased the bacterial mutagenicity of the extract. A different mutagenic response was observed in CHO cells. In the absence of liver S9, although the extract was cytotoxic to CHO cells, no significant mutagenicity was observed. Addition of exogenous liver S9 decreased the cytotoxicity and increased the mutagenicity at both Na+-K+-ATPase and hypoxanthine-guanine phosphoribosyl transferase (HGPRT) gene loci in CHO cells. Using gas chromatography/mass spectrometry (GC/MS) and tandem quadruple mass spectrometry, a number of polynuclear aromatic hydrocarbons (PAHs) and nitrated PAHs (nitro-PAHs) were tentatively identified and quantitated. A possible explanation of the difference in bacterial and mammalian mutagenicity of the extract is that the bacterial mutagenicity was induced by the nitro-PAHs that are potent bacterial mutagens and mammalian mutagenicity was induced by both PAHs and nitro-PAHs that are promutagens.

Chinese hamster ovary (CHO) cells were grown on hydrated collagen gels, the overlaying medium removed leaving the cells at an airkollagen interface, and the cells exposed to a dynamic flow of ethylene oxide. Increases in CHO cell mutant frequency and decreases in cell viability were observed. To establish if the exposure system could be simplified, cells were exposed in sealed bottles (static system) to ethylene oxide. No substantial changes in cytotoxicity, mutant frequency, or effective concentration were noted when comparing static versus dynamic exposure systems. The general usefulness of the exposure system using cells grown on collagen gels was evaluated in a static system using propylene oxide and 1,2dichloroethane, both of which were found to be mutagenic and cytotoxic. Comparatively, the exposure of cells by the collagen gel method was as effective in detecting genotoxic damage as were conventional methods (cells covered with medium) using cells grown on glass substrates. The exposure of CHO cells on collagen gels to highly volatile mutagens was simple and inexpensive, and may be generally useful in the detection of gaseous or volatile mutagens.

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