1-nitropyrene as a marker for the mutagenicity of diesel exhaust-derived particulate matter in workplace atmospheres

The use of 1-nitropyrene (1-NP) as a marker for the occupational exposure to diesel exhaust (DE) mutagens was investigated in workplace atmospheres contaminated with DE from a variety of emission sources, such as power supplies, forklifts, trucks, caterpillar vehicles, trains, ships' engines, and vehicles in city traffic. Total suspended particulate matter was collected by area sampling. The 1-NP content of acetone extracts of these samples as determined by gas chromatography-high resolution mass spectrometry varied from 0.080 to 17 μg/g acetone extractable matter, corresponding to air concentrations of 0.012 to 1.2 ng/m³. A sample collected in a rural area contained 0.0017 ng/m³ 1-NP. The mutagenicity of the extracts was tested in the Salmonella typhimurium strains TA98 and TA1538, using the microsuspension assay with and without metabolic activation by an exogeneous metabolizing system (rat liver S9-fraction). In addition, the S. typhimurium strains YG1021 and YG1024 were used because of their high sensitivity towards the mutagenicity of nitro polycyclic aromatic hydrocarbons. When plotting the mutagenic potency of the air sample extracts as determined in the absence of liver S9 versus the particle-associated 1-NP level, a relatively high correlation (r = 0.80–0.91) was observed in all of the S. typhimurium strains. High correlations (r = 0.80–0.93) were also observed when plotting the results of mutagenicity testing after activation by S9 versus the outcome of chemical analysis. These results show that the 1-NP content of workplace air samples is associated with their mutagenic potency, suggesting that 1-NP may be used as a marker for occupational exposure to DE-de-rived particle-associated mutagens © 1995 Wiley-Liss, Inc.     

Mutagenicity profile of atmospheric particulate matter in a small urban center subjected to airborne emission from vehicle traffic and sugar cane burning

Atmospheric particulate matter (PM) is genotoxic and recently was classified as carcinogenic to humans by the International Agency for Research on Cancer. PM chemical composition varies depending on source and atmospheric conditions. The Salmonella/microsome assay is the most used mutagenicity test and can identify the major chemical classes responsible for observed mutagenicity. The objective of this work was to characterize the mutagenicity of PM samples from a countryside city, Limeira, Brazil, which is influenced by heavy traffic and sugar cane biomass burning. Six samples of total PM were collected. Air mass backward trajectories were calculated. Organic extracts were assayed using the Salmonella/microsome microsuspension mutagenicity assay using TA98, YG1041, and TA1538, with and without metabolic activation (S9). YG1041 was the most sensitive strain and mutagenicity reached 9,700 revertants per m³ without metabolic activation. Potency for TA1538 was higher than TA98, indicating that this strain should be considered in air mutagenicity studies. The increased response to YG1041 relative to TA98, and the decreased response with S9, suggests that nitroaromatics are the major contributors. Limeira is among the most mutagenic cities in the world. High mutagenicity in Limeira seems to occur when the air mass from the area of sugarcane production is mixed with air from the region impacted by anthropogenic activities such as traffic. An increase in the formation of nitro‐polycyclic aromatic hydrocarbons may result from longer contact time between the aromatic compounds and the atmosphere with high NOx and ozone concentration, although more studies are required to confirm this hypothesis. Environ. Mol. Mutagen. 57:41–50, 2016. © 2015 Wiley Periodicals, Inc.     

9-Methoxytariacuripyrone, a naturally occurring nitro-aromatic compound with strong mutagenicity in Salmonella typhimurium

9-Methoxytariacuripyrone, a nitro-aromatic compound isolatedfrom Aristolochia brevipesshowed strong mutagenic activity instrain TA98, TA100 and some YG strains of Salmonella typhimuriumwithand without S9 mix. Incubation with cytosol resulted in a heavyincrease in mutagenicity. When incubated with microsomes theactivity was dramatically decreased. The results are discussedin view of the enzymes possibly involved in activation and detoxificationof the compound. The role of the basic structure on the mutagenicitymediated through the nitro group was also considered.     

Mutagenic activity in synthetic pyrethroids in Salmonella typhimurium

Four pyrethroids, allethrin, resmethrin, permethrin and fen-valerate,were tested for mutagenicity in bacterial reversion assay systemswith seven strains (TA1535, TA100, TA1538, TA98, TA1537, TA97and TA104) of Salmonella typhimurium. Our results show thatthree pyrethroids, namely resmethrin, permethrin and fenvalerate,were not found to be mutagenic in S. typhimurium in the presenceor absence of a rat liver activation system. Allethrin was foundto be mutagenic with TA100, TA104 and TA97 strains and requiredmetabolic activation (S9 mix) in order to show its activity,mainly with TA100 and TA104 strains.     

Sensitivity of salmonella YG5161 for detecting PAH‐associated mutagenicity in air particulate matter

The Salmonella/microsome assay is the most used assay for the evaluation of air particulate matter (PM) mutagenicity and a positive correlation between strain TA98 responses and benzo[a]pyrene (B[a]P) levels in PM has been found. However, it seems that the major causes of PM mutagenicity in this assay are the nitro and oxy‐PAHs. Salmonella YG5161, a 30‐times more responsive strain to B[a]P has been developed. To verify if YG5161 strain was sufficiently sensitive to detect mutagenicity associated with B[a]P mutagenicity, PM samples were collected in Brazil and Sweden, extracted with toluene and tested in the Salmonella/microsome microsuspension assay. PAHs and B[a]P were determined and the extracts were tested with YG5161 and its parental strain TA1538. The extracts were also tested with YG1041 and its parental strain TA98. For sensitivity comparisons, we tested B[a]P and 1‐nitropyrene (1‐NP) using the same conditions. The minimal effective dose of B[a]P was 155 ng/plate for TA1538 and 7 ng/plate for YG5161. Although the maximum tested dose, 10 m³/plate containing 9 ng of B[a]P in the case of Brazilian sample, was sufficient to elicit a response in YG5161, mutagenicity was detected at a dose as low as 1 m³/plate (0.9 ng). This is probably caused by nitro‐compounds that have been shown to be even more potent than B[a]P for YG5161. It seems that the mutagenicity of B[a]P present in PM is not detectable even with the use of YG5161 unless more efficient separation to remove the nitro‐compounds from the PAH extract is performed. Environ. Mol. Mutagen. 55:510–517, 2014. © 2014 Wiley Periodicals, Inc.     

Evaluation of the genotoxicity of river sediments from industrialized and unaffected areas using a battery of short-term bioassays

The present investigation evaluated the capacity of the Salmonella mutagenicity test, the comet assay, and the micronucleus assay to detect and characterize the genotoxic profile of river sediments. Three stations were selected on an urban river (Bouches du Rhône, France) exposed to various sources of industrial and urban pollution (StA, StB, and StC) and one station on its tributary (StD). One station in a nonurban river was included (REF). The concentrations of 16 polycyclic aromatic hydrocarbons (PAHs) were determined by HPLC, and the genotoxicity of the sediments was monitored by the Salmonella mutagenicity test (TA98 + S9, YG1041 ± S9), the comet assay, and the micronucleus assay on CHO cells. Chemical analysis showed that the total PAH concentrations ranged from 23 μg kg^?1 dw (REF) to 1285 μg kg^?1 dw (StD). All the sediments were mutagenic in the Salmonella mutagenicity test. The mutagenicity was probably induced by the presence of nitroarenes (StA, StB, StC, and StD) and aromatic amines (REF) as deduced from the mutagenicity profiles of strains YG1041 ± S9 and TA98 + S9. The comet assay revealed direct DNA lesions in REF, StA, and StB sediments and metabolization‐dependent DNA damage in StC and StD. The micronucleus assay showed an absence of clastogenicity for StA ± S9 and StC‐S9, and a significant clastogenicity ± S9 for the three other stations. The genotoxicity ranking determined by the comet assay + S9 matched the ranking of total and carcinogenic PAH concentrations, and this assay was found to be the most sensitive. Environ. Mol. Mutagen., 2008. © 2008 Wiley‐Liss, Inc.     

Reduction in the mutagenicity of synthetic dyes by successive treatment with activated sludge and the ligninolytic fungus, Irpex lacteus

Synthetic dyes are released in wastewater from textile manufacturing plants, and many of these dyes are genotoxic. In the present study, the mutagenicity of azo, anthraquinone, and triphenyl methane dyes was investigated before and after successive biodegradation with activated sludge and the ligninolytic fungus, Irpex lacteus. Two biodegradation systems were used to reduce the genotoxicity of dyes that were not efficiently inactivated by activated sludge alone. Mutagenicity was monitored with the Salmonella reversion assay conducted with the base-pair substitution detector strains, TA100 and YG1042, and the frame-shift detector strains, TA98 and YG1041, with and without rat liver S9. All dyes except for Congo Red (CR) were mutagenic with S9 activation. Assays conducted with the dyes indicated that only the azo dye Reactive Orange 16 (RO16) was mutagenic in both TA98 and TA100. Methyl Red and Disperse Blue 3 (DB3) were mutagenic in TA98, YG1041 and YG1042, while Reactive Black 5 was mutagenic in YG1041 and YG1042. Remazol Brilliant Blue R (RBBR), Crystal violet (CV) and Bromophenol Blue (BPB) were mutagenic only in TA98, but the toxicity of the latter two dyes complicated the evaluation of their mutagenicity. CR was not mutagenic in any of the tester strains. Biodegradation studies conducted with RO16 and DB3 indicated that the two-step biodegradation process reduced the mutagenic potential of RO16 and DB3 to a greater extent than activated sludge alone; the mutagenicity of the two dyes was reduced by 95.2% and 77.8%, respectively, by the two-step process. These data indicate that the combined biodegradation process may be useful for reducing the mutagenicity associated with wastewater from textile factories that contain recalcitrant dyes.     

Mutagenicity of blue rayon extracts of fish bile as a biomarker in a field study

Blue rayon (BR) in combination with the Salmonella/microsome assay was used to evaluate the mutagenicity of fish bile samples. Specimens of Mugil curema from two sites were collected over a 1‐year period. Piaçaguera channel contains high concentrations of total polycyclic aromatic hydrocarbons (PAHs) and other contaminants, while Bertioga channel was considered the reference sites in this study. Bile was extracted with BR and tested with TA98, TA100, and YG1041 strains with and without S9 in dose response experiments. PAH metabolite equivalents were analyzed using reverse‐phase high performance liquid chromatography /fluorescence. Higher mutagenic responses were observed for the contaminated site; YG1041 with S9 was the most sensitive strain/condition. Mutagenicity ranged from 3,900 to 14,000 rev./mg at the contaminated site and from 1,200 to 2,500 rev./mg of BR at the reference site. The responses of YG1041 were much higher in comparison with the TA98 indicating the presence of polycyclic compounds from the aromatic amine class that cause frameshift mutation. TA100 showed a positive mutagenic response that was enhanced following S9 treatment at both sites suggesting the presence of polycyclic compounds that require metabolic activation. benzo(a)pyrene, naphthalene, and phenanthrene metabolite equivalents were also higher in the bile of fish collected at the contaminated site. It was not possible to correlate the PAH metabolite quantities with the mutagenic potency. Thus, a combination of the Salmonella/microsome assay with YG1041 with S9 from BR bile extract seems to be an acceptable biomarker for monitoring the exposure of fish to mutagenic polycyclic compounds. Environ. Mol. Mutagen., 2010. © 2009 Wiley‐Liss, Inc.     

Assay of ptaquiloside, the carcinogenic principle of bracken, Pteridium aquilinum, by mutagenicity testing in Salmonella typhimurium

The mutagenicity of ptaquiloside, the carcinogenic principleof Pteridium aquilinum, was tested in Salmonella typhimuriumTA100 and TA98. Under weakly basic conditions (pH 8.5), ptaquilosidedecomposed into a conjugated dienone (considered to be the ultimateform), which was mutagenic in both strains. A novel bioassay,using the pre-incubation method at pH 8.5 with S. typhimuriumtester strains was developed for the assay of ptaquiloside extractedfrom plants. By this bioassay the ptaquiloside content of fernscollected at different localities during various seasons, andin various parts of the plant was determined. The ubiquitouspresence of ptaquiloside in fresh plant materials was confirmed.Bracken processed in alkali was found not to contain the carcinogen. ¹Present address: Wako Pure Chemical Industries Ltd, Kawagoe-shi,Saitama 328, Japan. ²Present address: Tochigi Health Centre, Tochigi-shi, Japan. ³To whom correspondence should be addressed      

The effect of biotransformation of 2,4-dinitrotoluene on its mutagenic potential

Because both oxidative and reductive metabolism of the hepatocarcinogen2,4-dinitrotoluene (2,4-DNT) can occur in vivo; we have examinedthe mutagenicity of compounds which can be formed from 2,4-DNTin an attempt to establish which metabolic pathways contributeto the formation of genotoxic products. A quantitative reversionassay using Salmonella typhimurium TA98 was used to evaluatethe mutagenicity of these compounds. 2,4-Dinitrobenzyl alcohol,2-amino-4-nitro-toluene and 2-nitroso-4-nitrotoluene were foundto be more mutagenic to S. typhimurium than is 2,4-DNT and didnot require metabolic activation by post-mitochondrial super-natantsof Aroclor-induced rat liver homogenates (S9) for their effect.2-Amino-4-nitrobenzoic acid was also mutagenic to S. typhimuriumTA98 in the absence of S9, but its mutagenicity was enhancedwhen S9 was included in the incubation mixture. 2,4-Diaminotoluenerequired S9 for demonstration of mutagenicity and was approximatelyas effective, on a molar basis, as 2,4-DNT in inducing reversionto histidine prototrophy. These results suggest that both oxidativeand reductive metabolism may be involved in production of mutagenicmetabolites of 2,4-DNT. ¹Present address to which correspondence should be addressed:Department of Pharmacology and Toxicology, University of MississippiMedical Center, 2500 N. State Street, Jackson, MS 39216, USA      

Genetic differences between the standard Ames tester strains TA100 and TA98

The standard Ames tester strains of Salmonella typhimurium areseparated by many steps in their pedigree, some involving mutagentreatments, and contain independently isolated uvrB-bio-galdeletions and rfa mutations. In this work the araD531 mutationwas introduced into the Ames tester strains TA100 and TA98.The responsiveness of the resulting strains (BA15 and BA14)to a number of chemical mutagens was then assessed by monitoringthe induction of forward mutations to L-arabinose resistance(Ara test). Here we have shown that these two strains of theAmes test differ greatly in their responses to mutagens, inways that are not associated with the mutagenic specificitiesof the original his mutations. In general, the genetic backgroundof strain TA100 appears to be more sensitive to the killingeffects of chemicals than that of TA98. The greatest differenceswere found with nifurtimox (NFX) and its analogue, compound1K. The Ara test responded to the mutagenic effects of thesetwo nitrofurans when carried out in the genetic background ofstrain TA98 but not in that of TA100. A higher sensitivity tothe lethal effects of NFX and 1K together with the greater nitroreductioncapability of strain TA100 as compared with TA98 might explainthe differences. In conclusion, our results indicate that thestandard Ames S. typhimurium tester strains are not isogenicand that genetic differences at loci other than his might besignificant for mutagenicity testing. To this respect the routineuse of the isogenic set of S. typhimurium strains constructedby Popkin et al. (Mut. Res., 224, 453–464, 1989) and derivedfrom strain hisD3052 (as the standard TA98) seems advisable.     

Formaldehyde is a bacterial mutagen in a range of Salmonella and Escherichia indicator strains

Formaldehyde was examined for bacterial mutagenicity using Escherichiacoli WP2(pKM101) and WP2uvrA(pKM101), and Salmonella typhimuriumTA1535, TA1537, TA1538, TA98, TA100 and TA102, in the absenceof any exogenous source of metabolic activation. Using pre-incubationexposure, clear mutagenicity was seen for TA98, TA100 and TA102,and both E.coli strains. In standard plate-incorporation assays,consistent mutagenicity was seen only for TA100 and WP2uvrA(pKM101).No evidence of mutagenicity was seen for TA1535, TA1537 or TA1538using either method of exposure. These data confirm the enhancedability of the pre-incubation method to detect the mutagenicityof formaldehyde both quantitatively, as expressed by numbersof revertant colonies, and qualitatively, in terms of the rangeof indicator strains reverted. The relatively greater sensitivityof the pre-incubation assay is probably due to better containmentof a volatile agent and/or lack of interaction with agar duringthe initial period of exposure. The findings are consistentwith the suggestion that formaldehyde induces lesions in bacteriawhich are, at least to some extent, excision-repairable, andindicate that the presence of the R-factor plasmid may be requiredfor the expression of its mutagenicity in excision repair-deficientSalmonella.     

Plant-activation of the bicyclic aromatic amines benzidine and 4-aminobiphenyl

Benzidine and 4-aminobiphenyl (4-ABP) are promutagenic bicyclic aromatic amines that are activated into frameshift and base pair substitution mutagens by plant systems. Using the plant cell/microbe coincubation assay, plant-activated benzidine from 0 to 50 μM induced a concentration-response in Salmonella typhimurium. At concentrations above 5 μM, plant-activated benzidine induced frameshift and base pair substitution mutations in the N- or O-acetyltransferase over-expressing strains, DJ460, YG1024, and YG1029. With plant-activated 4-ABP, concentrations above 250 μM induced a significant mutagenic response in strains YG1024 and YG1029. A tobacco cell-free mixture, TX1MX, activated benzidine and 4-ABP into mutagenic metabolites in S. typhimurium strains YG1024, YG1029, and DJ460. The mutagenic sensitivities of plant-activated benzidine and 4-ABP were the same with two different types of plant activation systems, TX1 suspension cells and TX1MX cell-free medium. The plant activation of these aromatic amines is mediated by tobacco cell peroxidase. Plant-activated benzidine and 4-ABP are converted into intermediates that serve as substrates for bacterial or humanacetylCoA: N-hydroxyarylamine N-acetyl-transferase to generate the ultimate mutagenic products. Environ. Mol. Mutagen. 29:81–90, 1997 © 1997 Wiley-Liss, Inc.     

Evaluation of the genotoxicity of stevioside and steviol using six in vitro and one in vivo mutagenicity assays

Stevioside, a constituent of Stevia rebaudiana, is commonlyused as a non-caloric sugar substitute in Japan. The genetictoxicities of stevioside and its aglycone, steviol, were examinedwith seven mutagenicity tests using bacteria (reverse mutationassay, forward mutation assay, umu test and rec assay), culturedmammalian cells (chromosomal aberration test and gene mutationassay) and mice (micronucleus test). Stevioside was not mutagenicin any of the assays examined. The aglycone, steviol, however,produced dose-related positive responses in some mutagenicitytests, i.e. the forward mutation assay using Salmonella typhimuriumTM677, the chromosomal aberration test using Chinese hamsterlung fibroblast cell line (CHL) and the gene mutation assayusing CHL. Metabolic activation systems containing 9000 g supernatantfraction (S9) of liver homogenates prepared from polychlorinatedbiphenyl or phenobarbital plus 5, 6-benzoflavone-pretreatedrats were required for mutagenesis and clastogenesis. Steviolwas weakly positive in the umu test using S.typhimurium TA1535/pSK1002either with or without the metabolic activation system. Steviol,even in the presence of the S9 activation system, was negativein other assays, i.e. the reverse mutation assays using S.typhimuriumTA97, TA98, TA100, TA102, TA104, TA1535, TA1537 and Escherichiacoli WP2 uvrA/pKM101 and the rec-assay using Bacillus subtilis.Steviol was negative in the mouse micronucleus test The genotoxicrisk of steviol to humans is discussed. ⁹To whom correspondence should be addressed     

Pilot study of the effect of diet on the mutagenicity of human faeces

A healthy non-smoking man, consuming a normal ‘western’diet (meat, vegetables, bread and alcohol) collected five consecutivecomplete bowel movements. He then added an extra 150 g of fat(from butter, cheese, milk, chocolate, peanuts, bacon and eggs)to his daily diet for 2 weeks, and collected four further consecutivebowel movements in the second week. After 6 months on his normaldiet, he added 30 g of wheat bran to his daily diet for 3 weeks,and collected four complete stool samples in the third week.Aqueous faecal extracts were prepared and assayed for auxotrophicgrowth-enhancement and bacterial mutagenicity (using fluctuationtests with Salmonella typhimurium TA100 and Escherichia coliWP2uvrApKM101). There was no significant difference in faecalwet weight between ‘normal’ and ‘high-fat’collections, but addition of 30 g bran was associated with a1.8-fold increase in stool weight, in good agreement with publisheddata. Fluctuation tests showed that normal and high-fat sampleswere mutagenic to S. typhimurium TA100 and to E. coli WP2uvrApKM101.There was considerable variation in mutagenic activity betweenconsecutive bowel movements. However, there was no significantdifference in mutagenicity between normal and high-fat samples.Faecal samples collected during the course of the high-fibrediet were significantly less mutagenic to both bacterial strains.Changes in auxotrophic growth-enhancing activity could not accountfor these changes in mutagenicity, since the pattern of changein growth-enhancement was very different from that seen formutagenicity. ¹To whom correspondence should be addressed     

Re-evaluation of the effect of ellagic acid on dimethylnitrosamine mutagenicity

Dimethylnitrosamine (DMN) is activated to mutagenic speciesin the Ames test (Salmonella typhimurium strain TA 100) by hamsterhepatic S9 preparation. This S9 activity is induced by administrationof ethanol to the animals. The organic solvents dimethylsulphoxide(DMSO) and N-methyl-2-pyrrolidinone (MP) inhibit this mutagenicity,apparently because they inhibit DMN demethylase activity (assayedas formaldehyde production). Ellagic acid, dissolved in DMSOor MP, had no inhibitory effect on DMN mutagenicity, beyondthe effect of the solvent vehicle. ²To whom correspondence should be addressed     

Mutagenicity testing of rutacridone epoxide and rutacridone, alkaloids in Ruta graveolens L., using the Salmonella/microsome assay

The mutagenicity of rutacridone and rutacridone epoxide wasinvestigated using Salmonella typhimurium without as well aswith different metabolic activation systems. Rutacridone epoxidewas found to be a direct acting mutagen in S. typhimurium strainsTA98, TA100 and TA1538; addition of rat liver preparations resultedin a marked decrease of mutagenicity. In contrast, rutacridonerequired metabolic conversion to exhibit mutagenic activity.Neither of the compounds had any effect on tester strain TA1978.S9 mixes as well as microsomal and cytosolic preparations fromuntreated, phenobarbital-treated and 3-methylcholanthrene-treatedrats were used to study the activation and deactivation capacitiesof the enzyme mixtures. In addition, the influence of enzymeinhibitors on the activation and deactivation of the furoacridoneswere tested. Evidence is presented that rutacridone is metabolizedby rat liver enzymes to the corresponding epoxide as the ultimatemutagen. ^*Dedicated to Professor Dr C.-G.Arnold on the occasion of his60th birthday.      

Characterization of the CPY isozyme profile induced by cyclohexanol

In a previous report we described the ability of cyclohexanolto induce CYP activity. In order to characterize this inductionwe tested the capacity of liver S9 from rats orally treatedwith cyclohexanol for 5 days, to activate several carcinogenicnitrosamines into mutagens in the Salmonella typhimurium TA100test system. Additionally, Western blot analysis of hepaticmicrosomes from the same treated animals were analysed withspecific antibodies against P450 protein families 1A1/A2, 2B1/B2and 2E1. Cyclohexanol-S9 mixture was more efficient in activatingthe following nitrosamines: N-nitrosodimethylamine (NDMA), N-nitrosodipropylamine(NDPA), N-nitrosomethylpropylamine (NMPA), N-nitrosodibutylamine(NDBA), and N-nitrosopyrrolidine (NPYR) into bacterial mutagensthan S9 from non-treated animals. The mutagenicity of N-nitrosodiethylamine(NDEA) was not modified in the presence of S9 from cyclohexanol-treatedanimals. Since the main metabolic pathway leading to the productionof mutagenic intermediates of NDMA and NPYR is catalysed byisozyme CYP2E1 and that of NDPA, NMPA and NDBA by CYP2B1/B2,mutagenicity experiments predicted that cyclohexanol inducesthese two P450 isozyme families. Western blot analysis confirmedthe results of the mutagenicity assay, showing an increase inthe intensity of CYP2E1 and CYP2B1/B2 protein bands in hepaticmicrosomes from cyclohexanol treated rats in comparison withnon-treated controls. Bacterial mutagenicity tests with specificpro-mutagens were good predictors of the P450 induction propertiesof cyclohexanol. ³To whom correspondence should be addressed at: Instituto de Investigaciones Biomédicas, Universidad Nacional Autonoma de Mexico, Apartado Postal 70228, 04510 Mexico, D.F., México     

Mutagenic synergy between paraoxon and plant-activated m-phenylenediamine or 2-acetoxyacetylaminofluorene

Paraoxon (diethyl-p-nitrophenylphosphate) is the toxic, but non-mutagenic metabolite of the organo-phosphorus ester insecticide parathion. Although this agent has been used as a deacetylase inhibitor in many studies, we discovered a mutagenic synergy when paraoxon was incubated with plant-activated m-phenylenediamine (mPDA) or with direct-acting 2-acetoxyacetylaminofluorene (2AAAF) and S. typhimurium tester strains. Using non-toxic concentrations of plant-activated mPDA and paraoxon a 10-fold increase in the mutant yield of S. typhimurium was observed. The mutagenicity of the plant-activated mPDA product required that O-acetyltransferase (OAT) be expressed by the S. typhimurium tester strain. However, the paraoxon-dependent mutagenic synergy was observed using the direct-acting arylamine metabolite, 2AAAF, with strains YG1024, TA98 and TA98/1,8-DNP₆ regardless of their OAT activity. This mutagenic synergy is dependent upon the presence of an activated acetylated form of the arylamine. The data presented here demonstrate that this mutagenic synergy is limited to paraoxon and not to the parent compound (parathion) or to a major metabolite of parathion (p-nitrophenol). © 1996 Wiley-Liss, Inc.     

Investigations into the genotoxic potential of loxtidine, a long-acting H2-receptor antagonist

Loxtidine, a potent, non-competitive histamine H₂-receptor antagonistwas evaluated for genotoxic potential using a range of short-termmutagenicity assays. Unequivocally negative results were obtainedin a Salmonella/plate incorporation assay and a liquid pre-incubationassay (using S.typhimurium strains TA1535, TA100, TA1537, TA1538and TA98), a fluctuation assay [using Escherichia coli strainsWP2, WP2 uvrA (R46) and 343/113 lys60 (R46)], a gene conversionassay (using Saccharomyces cerevisiae JD1) and a human peripherallymphocyte cytogenetic assay. All of these in vitro tests werecarried out in the presence and absence of rat liver S9 mix.In addition, the major metabolites of loxtidine in the rat werealso negative in the same range of microbial mutagenicity assays.Loxtidine was inactive in the mouse micronucleus test afteroral administration. The potential nitrosatability of loxtidinewas investigated using an expanded version of the WHO NitrosationAssay Procedure, and detectable quantities of mutagenic nitroso-specieswere not formed. The subsequent appearance of carcinoid tumourswithin the gastric fundus of rodents treated orally with loxtidinefor most of their natural lifespan, led to additional assaysbeing carried out on this compound to determine whether thetumorigenic effects were due to alternative mutagenic mechanisms.Negative results were obtained in an in vitro unscheduled DNAsynthesis assay using primary rat hepatocytes, and an assayfor spindle damaging agents using Muntjac skin fibroblasts.It can be concluded from these results that loxtidine is unlikelyto be a genotoxic carcinogen. The increase in carcinoid tumourincidence observed in rats and mice after loxtidine treatmentwas probably related to the prolonged achlorhydria producedby this potent unsurmountable histamine H₂-receptor antagonist.