Elena C. McCoy

The basis of the insensitivity of Salmonella typhimurium strain TA98/1,8-DNP6 to the mutagenic action of nitroarenes.

Salmonella typhimurium tester strain TA98/1,8-DNP6 is resistant to the mutagenicity of 1,8-dinitropyrene because it lacks an esterification enzyme which is needed for the formation of the ultimate mutagen, presumably the corresponding hydroxamic acid ester. This enzyme does not appear to be required for the activation of all nitroarenes and arylamines, as some of these are fully active in TA98/1,8-DNP. It is suggested that these form electrophilic arylnitrenium ions nonenzymatically from nitroso- and N-hydroxylamino-arenes intermediates. The esterification enzyme appears to be a transacetylase. An assay using 2-aminofluorene as the acetyl acceptor is described. Derivatives of S. typhimurium TA100 also lacking this enzyme were obtained by Tn5-mediated mutagenesis.

Esterification of arylhydroxylamines: Evidence for a specific gene product in mutagenesis

Characterization of a Salmonellatyphmurium mutant strain (TA98/1,8-DNP6) resistant to the mutagenicity of nitrated polycyclic aromatic hydrocarbons (nitroarenes) revealed that it was also non-responsive to the mutagenic action of nitroso- and N-hydroxylaminoarenes. The mutant strain was fully sensitive to the mutagenic action of the corresponding hydroxamic acid ester. These results suggest that TA98/1,8-DNP6 is deficient in a specific esterifying enzyme and that esterification of the penultimate mutagenic metabolites of nitro- and aminoarenes (e.g., arylhydroxylamines) to form potent electrophiles is controlled by a specific gene.

Cigarette smoking may yield nitroarenes

Cigarette smoke condensates are readily nitrated to mutagenic substances exhibiting the properties expected of nitroarenes. In addition, nitroarenes also appear to be generated during the smoking of cigarettes enriched with nitrates.

Photochemical instability of 1-nitropyrene, 3-nitrofluoranthene, 1,8-dinitropyrene and their parent polycyclic aromatic hydrocarbons

The environmental contaminants pyrene, 1-nitropyrene, 1,8-dinitropyrene, fluoranthene, and 3-nitrofluoranthene were exposed to light (greater than or equal to 310 nm) either in DMSO, or following coating onto silica. Under all conditions tested the pyrenyl were less stable than the fluoranthenyl compounds. During irradiation in DMSO or on silica, 1-nitropyrene had half-lives of 1.2 and 6 days, while those of 3-nitrofluoranthene were 12.5 and greater than 20 days, respectively. The photodecomposition of 1,8-dinitropyrene resembled that of 1-nitropyrene with half-lives of 0.7 and 5.7 days. A principle photodecomposition product of 1,8-dinitropyrene was identified as 1-nitropyren-8-ol. It was also found that when the nitroarenes were exposed to light, the loss of compound was associated with a concomitant loss of mutagenicity in Salmonella typhimurium strain TA98. The mechanism of nitrated polycyclic aromatic hydrocarbon decomposition and 1-nitropyren-8-ol formation, and the relevance to the atmospheric disposition of these compounds are discussed.

Airplane emissions: A source of mutagenic nitrated polycyclic aromatic hydrocarbons

Organic solvent extracts from airplane emission particulates are mutagenic for Salmonella typhimurium strain TA98. Using Salmonella tester strains deficient in enzymes required for the bioactivation of various nitroarenes, the mutagenicity present in these emissions was ascribed to the presence of nitrated polycyclic aromatic hydrocarbons. Based on the known aircraft particulate emission rates at U.S. airports, and using 1-nitropyrene (1-NP) and 1,8-dinitropyrene (1,8-DNP) as surrogates, it is calculated that at a minimum 7 kg 1-NP and 20 g, 1,8-DNP are emitted daily at a typical U.S. airport.

Mutagenicity of chloroalkene epoxides in bacterial systems

6 alpha-chloroepoxides have been tested for in vitro activity in a variety of systems. The epoxides were cis- and trans-1-chloropropene oxide, cis- and trans-1,3-dichloropropene oxide, trichloroethylene oxide and tetrachloroethylene oxide. The epoxides were assayed for mutagenicity in the absence of metabolic activation in S. typhimurium TA1535 and E. coli WP2 uvrA and for preferential inhibition of growth of DNA-repair-deficient E. coli. All 6 epoxides possessed DNA-modifying activity as evidenced by their ability to preferentially inhibit DNA polymerase-deficient E. coli. All of the test chemicals except trichloroethylene oxide were mutagenic for S. typhimurium and all except trichloroethylene oxide and tetrachloroethylene oxide were mutagenic for E. coli Wp2 uvrA. Cis- and trans-1,3-dichloropropene oxide were the most potent mutagens and DNA modifiers. For all cases, the cis isomers were more active than the corresponding trans isomers. alpha-Chloroepoxides are considered likely to be the active intermediates of the carcinogenic parent halo-olefins. These mutagenicity studies are considered relevant in assessing the carcinogenicity of the parent hydrocarbons.

The chemical activation of non-mutagenic nitrated polycyclic aromatic hydrocarbons to mutagens.

Nitrated polycyclic aromatic hydrocarbons, including carcinogens, may be non-mutagenic in microorganisms because bacterial nitroreductases are unable to reduce their nitro function to proximate mutagenic hydroxylamines. This reduction of the nitro moiety can be accomplished chemically in situ using zinc dust. The procedure, which is compatible with the Salmonella mutagenicity assay, was used to generate mutagens from chemicals which otherwise are non-mutagenic even in the presence of microsomal preparations.

Carcinogenic N-hydroxylaminopurine derivatives do not act as base analog mutagens in Salmonella typhimurium

N-Hydroxylaminopurines are highly mutagenic for growing as well as resting Salmonella typhimurium strain TA100 and to a lesser extent for strain TA98. Aminopurines, under similar conditions, are not mutagenic. N-Methylhydroxylaminopurine, under similar conditions, exhibits only minimal activity. The results are taken to indicate that unlike non-hydroxylated aminopurines, N-hydroxylaminopurines exert their mutagenicity not by acting as base analogs but by direct covalent binding with DNA-guanine.

Evidence that nitroarene metabolites form mutagenic adducts with DNA-adenine as well as with DNA-guanine

Nitropyrenes as well as several other nitroarenes and their metabolites exhibit considerable mutagenicity for Salmonella tester strains (TA102 and TA96) which have adenine-thymine base pairs at the mutational target. This finding is unexpected as previous biochemical studies had shown that arylation at the C8 position of DNA-guanine is the only chemically and biologically significant reaction. This conclusion is supported by the extraordinary mutagenic potency of these chemicals in Salmonella strains with guanine at the mutational site (e.g., TA98). The present results indicate that a minor reaction with DNA-adenine may result in the formation of an unusually potent promutagenic DNA adduct.

Genetic and quantum chemical basis of the mutagenicity of nitroarenes for adenine-thymine base pairs

The mutagenicity of nitroarenes for Salmonella typhimurium strains with adenine-thymine base pairs at the mutational site is dependent upon enzymic reduction of the nitro function. Although the electrophilic metabolites of nitroarenes are capable of mutating adenine-thymine base pairs, they show a marked preference for guanine-cytosine pairs when given a choice. Quantum chemical calculations indicate the reactivity order for nucleophilic sites in an AT run of base pairs to be the N-7 of adenine (N7(A)) first, followed by an approximately equal reactivity for C-8 of adenine (C8(A)) and O4 of thymine (O4(T)). Given the low probability of reaction of electrophilic metabolites of nitroarenes with adenine-thymine base pairs, the mutagenic potency of nitroarenes for strains with adenine-thymine base pairs at the mutational site is remarkable.