Mutagenicity and genotoxicity of nitroarenes

Directly active mutagens are formed on exposure of the promutagen benzo[a]pyrene to gaseous pollutants in smog. In simulated atmospheres containing 1 part per million nitrogen dioxide and traces of nitric acid, directly mutagenic nitro derivatives are readily formed from both benzo[a]pyrene and perylene, a non-mutagen in the Ames reversion assay. Possible formation of direct mutagens by such reactions on sample collection filters, in exhaust effluents, and in the atmosphere should be recognized.

The soluble organic fraction (SOF) of particulate matter from diesel exhaust (from point sources, ambient air, etc.) contains hundreds of organic constituents. Norman-phase high pressure liquid chromatography (HPLC) has been used to separate the SOF into subfractions suitable for subsequent chemical analysis and bioassays. These fractions consist of non-polar(PAH), moderately polar (transition) and highly polar constituents. The non-polar fractions have been well characterized and consist of PAH and aliphatic hydrocarbons. The specific compounds present in the transition and polar fractions are for the most part unknown. This analytical information has been difficult to obtain since these compounds are highly labile, polar, of low volatility and in very low concentrations when compared to the bulk of material found in the SOF. Mutagenicity tests using the Ames Salmonella typhimurium assay indicate that the transition fraction accounts for most of the mutagenicity when compared to the non-polar (PAH) and polar fractions. A variety of chromatographic and mass spectrometric techniques are described that have been used to determine the composition of the HPLC fractions. More than one hundred species have been identified in the transition fraction of diesel particulate matter using high resolution gas chromatography (HRGC)/high resolution mass spectrometry (HRMS), HPLC and direct-probe high resolution mass spectrometry. It has been found that the transition fraction contains mostly PAH derivatives consisting of hydroxy, ketone, quinone, carboxaldehyde, acid anhydride and dihydroxy derivatives of PAH. Three nitro-PAH species have been tentatively identified and 1-nitropyrene positively identified in the transition fraction. The 1-nitropyrene was found to account for approximately 45% and 30% of the direct-acting mutagenicity observed for the transition fraction and total extract, respectively. The HPLC separation procedure was shown to give better than 95% recovery of the mass and mutagenic activity. The problem of PAH oxidation during the analytical procedures and possible effect on bioassay results are discussed.

A number of tests have been described which are thought to be capable of identifying carcinogens without using the actual induction of cancer as an endpoint. This study compared the performance of 6 such tests on a selection of 120 organic chemicals. The tests studies were: (1) mutation of Salmonella typhimurium; (2) cell transformation; (3) degranulation of endoplasmic reticulum; (4) sebaceous gland suppression; (5) tetrazolium reduction and (6) subcutaneous implant. A further 4 tests were examined briefly, but were not included in the complete evaluation. The chemicals were classified into carcinogens (58) and non-carcinogens (62) on the basis of published experimental data, and into 1 of 4 broad chemical classes. There was considerable variation between tests in their ability to predict carcinogenicity, with the cell-transformation test and the bacterial-mutation test being the most accurate (94% and 93% accurate respectively). These 2 tests were considered to be of general use in screening, since they were clearly more accurate than the others. Statistical consideration of various combinations of these tests showed that the use of cell transformation and bacterial mutation together, provide an advantage over the use of either test alone. The inclusion of the other 4 tests in a screening battery predictably resulted in a great increase in overall inaccuracy and loss of discrimination, even though the detection of carcinogens is improved. All the tests were shown to generate both false positive and false negative results, a situation which may be controlled by the use, where possible, of appropriate chemical-class controls, to identify the test which is optimal for the class of chemical under test. Structural analogy may have a part to play in the rapid detection of environmental carcinogens, and some general guidelines for its use are given.