Mutagenic activity of airborne particulate matter as an indicative measure of atmospheric pollution

Mutagenic activity of organic extracts of airborne particulate matter at four different sites within the urban area of the city of Porto Alegre, Brazil, was investigated using the Salmonella/microsome assay, with the Kado microsuspension method. The extracts were obtained by sonication, sequentially extracted according to polarity, with cyclohexane (CX) and dichloromethane (DCM) solvents. The different fractions were tested for mutagenicity with the Salmonella typhimurium strains TA98, TA98NR and TA98/1,8-DNP6, without S9 mix metabolic activation. A positive frameshift mutagenic response was observed for non-polar (CX) and/or moderately polar (DCM) compounds at the different sites. The responses varied at different seasons of the year, and the highest revertants per m3 (rev/m3) values were observed at the site subject to the strongest influence of automotive vehicles (site 3) in spring (17.13 rev/m3) in DCM fractions, and in summer (13.01 rev/m3) in CX fractions. The responses observed for the TA98NR and TA98/1,8-DNP6 strains suggest the contribution of nitrocompounds to the mutagenic activity observed. Although there appears to be an indicative association between the increased mass per unit volume of air (TSP) and the mutagenicity of organic extracts of airborne particulate matter in the present study, the Salmonella/microsome assay was a sensitive method to define areas contaminated by genotoxic compounds, even in samples that present TPS values acceptable by the environmental quality standards established by law.     

Mutagenicity of particulates from the laboratory combustion of plastics

Carcinogenic polycyclic aromatic hydrocarbons (PAHs) and nitropolycyclic aromatic hydrocarbons (nitro-PAHs) have been identified in airborne particulate organic matter extracts. The pollutant sources were generally contributed by motor vehicles and industrial activity. Massive quantities of urban solid wastes, containing plastic materials such as PVC, PET, PS, and PE, burnt in the open air in local garbage dumps are frequently found in developing countries. In this study, the smog particulates from the combustion of these synthetic polymers were produced in a laboratory combustion chamber. The mutagenicity of acetone extracts from the smog particulates was evaluated with Salmonella typhimurium TA98 and TA100 in the presence and absence of S9 mix. Four samples in TA98 exhibited higher mutagenicity than those in TA100. The greatest mutagenicity was observed from the extracts of particulates from combustion of PVC followed by that of PS, PET, and PE. To determine the major mutagenic compounds in these samples, mutagens were partially purified through TLC and their mutagenicity was monitored with TA98. 1-NP and DNPs in the above samples were also determined by HPLC. The amounts of 1-NP and DNPs generally corresponded with their mutagenicity. Higher levels of 1-NP and DNPS from the combustion of PVC, PET, and PS. the combustion of synthetic polymer wastes might be responsible for the presence of high levels of 1-NP and DNPs in Taiwan urban air.     

Genotoxicity of fractionated organic material in airborne particles from São Paulo, Brazil

Particulate matter less than 10 microns aerodynamic diameter (PM10) is associated with adverse health effects including increased respiratory problems and mortality. PM10 is also associated with increases in cancer in some urban areas. Identification of toxic compounds in PM10 is a step toward estimating exposure to these compounds and evaluating their public health risk. However, the toxic compounds on PM10 are part of a highly complex mixture of compounds that makes chemical characterization difficult. Before this study, there has been little investigation of genotoxic compounds in particulate matter from Latin American cities. Here, both bioassay (mutagenicity) and chemical analyses were conducted with organic solvent extracts of PM10 collected from S?o Paulo, a major Brazilian city. Sequential extraction in dichloromethane (DCM) followed by acetone (ACE) yielded 20.3% and 10.2% of the total mass, respectively. Non-polar and moderately polar organic material solubilized in DCM. ACE extracted more polar organic species and some inorganic ions. Both extracts were fractionated separately using cyanopropyl-bonded silica chromatography with organic solvents of increasing polarity. The mass distribution among the fractions was measured. The mutagenic activity of the fractions was assayed using the microsuspension procedure with the Salmonella typhimurium tester strain TA98, with and without addition of metabolic enzymes (S9). The DCM extract had about four times higher mutagenic activity than the ACE extract. In general, addition of S9 resulted in an increase in mutagenicity of DCM fractions, but a decrease for the ACE extract. Most of the activity was concentrated in fractions in the mid-range of polarity within both the DCM and ACE extracts. The fractions were analyzed by gas chromatography with mass selective detection (GC/MS) without derivatization. The most mutagenic fractions in the DCM extract contained ketones, aldehydes, and quinolines. The most mutagenic ACE fraction had ketones, carboxylic acids, and aldehydes.     

In vitro and in vivo mutagenicity studies with airborne particulate extracts

The contribution of nitro compounds to airborne particulate mutagenicity was studied with Salmonella typhimurium strains TA98, TA98NR, TA98/1,8DNP6. The results obtained indicate that nitropyrenes play a minor role in air particulate mutagenicity. Seasonal variations indicate a relatively greater contribution of nitro compounds to the mutagenicity of spring and summer samples. Fractionation of extracts into acidic, neutral and basic components shows that neutral compounds account for about two-thirds of the total mutagenic activity. Attempts to extract mutagens adsorbed onto particulate matter with aqueous media were almost completely negative. No significant mutagenicity was detected in urine and faecal extracts and in plasma samples of Sprague-Dawley rats treated with air particulate extracts at 80 mg/kg either per os or by i.p. injection. Negative results were obtained in the micronucleus test with Swiss mice treated at 200 and 400 mg/kg (twice by i.p. injection). A significant decrease in liver aminopyrine-N-demethylase was observed in Swiss mice injected with air particulate extracts or its basic and neutral fractions. In vitro experiments suggest a direct interaction of test materials with microsomal cytochrome P-450.     

Detection of mutagenic activity in automobile exhaust

Using the Ames Salmonella-microsome system, we detected mutagenic activity in the exhaust from two kinds of 4-cycle gasoline engines of unregulated and regulated cars, and from diesel engines, as well as in the particulates from air collected in tunnels. The mutagenicity of particulates from a car equipped with a catalyst (regulated car), as compared with that from an unregulated car, was reduced very much (down to 500 from 4500 revertants/plate/m3 in tester strain TA98). However, the mutagenicity of the ether-soluble acid and neutral fractions from the condensed water of emissions from a regulated car was still high (down to 2880 from 10 900 revertants/plate/m3 in tester strain TA100). The mutagenic activity of emission exhaust from old diesel car engines was very high; the particulates showed 9140 and 19 600 revertants/plate/m3 from strain TA98 incubated with an activating rat-liver S9 fraction. A small diesel engine of the type used for the generation of electric power or in farm machinery also produced exhaust with highly mutagenic particulates. The mutagenic activity of a methanol extract of particulate air pollutants collected in a highway tunnel showed 39 revertants/plate/m3 toward strain TA98 and 87 toward strain TA100. The ether-soluble neutral fraction yielded 86 revertants/plate/m3 from strain TA98 and 100 from strain TA100. This fraction also contained carcinogenic compounds, including benzo[a]pyrene, benzo[e]pyrene, benz[a]anthracene, benzo[ghi]perylene and chrysene. Very high mutagenic activity was detected, especially in the particulate air pollutants collected at night, in another tunnel on a superhighway: 60-88 revertants/plate/m3 from strain TA100 for the sample collected by day, but 121-238, by night. Night traffic includes many more diesel-powered vehicles compared with gasoline-powered automobiles.     

Characterization of organic extracts from standard reference materials 1649, ‘urban dust/organics,’ and 1650, ‘diesel particulate matter’, using a microsuspension assay. A WHO/IPCS/CSCM study

Mutagenicity associated with replicate organic extracts from standard reference materials 1649 ‘urban dust/organics’ (air particles), and 1650, ‘diesel particulate matter’ (diesel particles), was determined using a Salmonella microsuspension assay. The results indicate that the mutagenicity of samples such as these can readily be determined using the microsuspension assay with only 5% of the mass required for the standard plate incorporation asssay.In general, 80% of the variation in mutagenic activity was due to the bioassay procedure and 20% to the extraction process. Extracts from both samples had primarily direct-acting mutagenicity as there were no significant differences in responses with and without metabolic activation (S9). The TA98 - S9 mean air particles mutagenic activities (C.V., %) based on mass of extractable organics or particles were 4.4 (4.7%) and 0.29 (3.6%) revertants/μg, respectively, and for the diesel particles were 66 (44%) and 12 (29%) revertants/μg, respectively. More of the observed direct-acting mutagenicity in the diesel particles extracts was due to nitro-substituted compounds because there were significant reductions in activity with TA98NR (45% of TA98 -S9) and TA98-1,8-DNP6 (21% of TA98 -S9). In the air particles extracts, the TA98NR activities were not significantly different from TA98 - S9 but the TA98-1,8-DNP6 levels were.     

Mutagenic activity of airborne particulate matter in a petrochemical industrial area

Exposure to airborne particulate matter has adverse effects on human health and ecosystem. Mutagenic activity of airborne particulate organic matter extracts in three time periods from total suspended particles (TSP) and particles less than 10 microm (PM10) was evaluated in an area under the influence of a petrochemical industry located in the town of Triunfo, Brazil. The extracts were investigated using the Salmonella/microsome assay, with the microsuspension method. The extracts were obtained by sonication extracted using dichloromethane (DCM) solvent. The fractions were tested for mutagenicity with the Salmonella typhimurium strains TA98 (with and without metabolic activation), TA98NR and TA98/1,8DNP(6); or YG1021 and YG1024. A positive frameshift mutagenic response was observed for the environmental samples during the different periods. The responses according to percentage of extractable organic matter (EOM%), EOM/m(3), revertants/microg (rev/microg) and revertants/m(3) (rev/m(3)) were lower for TSP than for PM10 extracts. The highest rev/m(3) values were observed in PM10 extract samples collected in winter, July 2005, in the presence (13.79 rev/m(3)) or absence (6.87 rev/m(3)) of S9 fraction. Similarly in the first (1995) or second period (2000) the highest values for TSP were observed in winter, but with lower activity (3.00 and 0.89 rev/m(3) respectively). The responses observed for the nitrosensitive strains suggest the contribution of nitro, amino and/or hydroxylamino derivatives of PAHs to the total mutagenicity of matter extracted from airborne particles. The Salmonella/microsome assay was a sensitive method to define areas contaminated by genotoxic compounds, even in samples with TSP or PM10 values that are acceptable according to legal environmental quality standards, favoring environmental control measures with an effective response seen in the population's improved quality of life.     

Results of the IPCS collaborative study on complex mixtures

The International Programme on Chemical Safety (IPCS) sponsored a collaborative study to examine the intra- and inter-laboratory variation associated with the preparation and bioassay of complex chemical mixtures. The mixtures selected were National Institute of Standards and Technology (NIST) Standard Reference Materials (SRMs). 20 laboratories worldwide participated in the collaborative trial. The participating laboratories extracted the organic portion of two particulate samples — an air-particulate sample and a diesel-particulate sample - and bioassayed the extracts. The laboratories simultaneously bioassayed a NIST-prepared extract of coal tar and two control compounds (benzo[a]pyrene, and 1-nitropyrene). The bioassay method used was the Salmonella/mammalian microsome plate-incorporation test using strains TA98 and TA100. Study design also allowed for a comparison of sonication and Soxhlet extraction techniques. The mean extractable masses for the air particles and diesel particles were approximately 5% and 1.75%, respectively. The particulate samples were mutagenic in both strains with and without activation in all 20 laboratories. For TA100 the with and without activation slope values for the air particulate were 162 and 137 revertants per mg particles, respectively. For TA98 the respective diesel slope values were 268 and 269. The mutagenicity slope values for the diesel particles ranged from 3090 (TA98, + S9) to 6697 (TA100, + S9) revertants per mg particles. The coal tar solution was negative for both strains when exogenous activation was not used but was mutagenic in both strains with exogenous activation. The benzo[a]pyrene and 1-nitropyrene were used as positive controls and gave results consistent with the literature. This paper provides a complete summary of the data collected during the collaborative study. Companion papers provide further analysis and interpretation of the results.     

Detection of oxidative mutagens in an urban air-particulate extract: a preliminary study.

The Ames assays strains TA98 and TA100 have been useful in characterizing complex mixtures from organic solvent extracts of particles from diesel-powered vehicles, ambient air, and other sources. In this paper we report preliminary experiments using TA102, a bacterial strain that detects compounds that can oxidize DNA, to characterize the mutagenicity of an ambient air sample collected in Ann Arbor, MI. Four sets of ambient air filters were collected in duplicate over a period of several days. The mutagenicities of methylene chloride extracts of these filters were compared using strains TA98, TA100 and TA102. The concentration-mutagenicity data for TA98 and TA100 were linear over the concentration range 0-200 micrograms extract/plate. The mutagenicity of the extracts using TA102 was much lower than the other two strains and was non-linear over the concentration range tested. These results suggest that it would be difficult to use TA102 to identify the oxidative mutagens present in an ambient air particulate extract.     

Mutagenicity in Salmonella typhimurium mutants of serum extracts from airborne particulates

Airborne particulates collected from urban and non-urban air were extracted with calf serum or benzene, and their mutagenic potencies were evaluated in the Salmonella reversion assay. The serum extracts were mutagenic to strains TA98 and TA100 and contained both direct- and indirect-acting mutagens. Mutagenic activities for TA98 recovered from the particulates by serum or benzene extraction were much less in the serum extracts than in the benzene extracts. There was no significant difference in mutagenic potencies of the extracts between the urban and non-urban particulates, irrespective of the presence of S9 mix. The calculated mutagenic activities per m3 of air, however, were greater for urban air than for non-urban air, because of higher concentration of particulates in urban air than in non-urban air. Serum effectively reduced both direct and indirect mutagenic activities of the benzene extracts except for an insufficient reduction in direct mutagenicity at a high dose of benzene extracts. These findings suggest that serum could contribute greatly to decrease the mutagenicity of airborne particulates by mechanisms such as less efficient solubilization of mutagenic components and inactivation by protein binding. Biological availability of mutagens, therefore, should be considered for evaluation of actual mutagenic hazard by airborne particulates.     

Salmonella mutagenicity of three complex mixtures assayed with the microsuspension technique. A WHO/IPCS/CSCM study.

In a collaborative study on complex mixtures, three complex mixtures and two pure compounds were assayed with the Salmonella microsuspension technique. The two pure compounds were benzo[a]pyrene (BaP) and 1-nitropyrene (1-NP). The three complex mixtures were standard reference materials (SRMs) from the U.S. National Institute of Standards and Technology, SRM 1649, SRM 1650 and SRM 1597. The two samples SRM 1649, an urban dust particulate matter, and SRM 1650, a diesel particulate matter, were sonicated with dichloromethane. Sample SRM 1597 was an extract of a coal tar sample with a complex mixture of polycyclic aromatic hydrocarbons. The microsuspension assay was performed with Salmonella strains TA98 and TA100 according to Kado et al. (1983) with minor modifications (L?froth et al., 1988). The results showed that the microsuspension technique is a more sensitive assay than the plate incorporation method. Depending on sample, strain and metabolic condition the mutagenic responses were 3-37 times higher in the microsuspension assay than in the conventional plate incorporation assay. The microsuspension method is thus useful for environmental samples which are often available in only small amounts.     

Detection of mutagenic activity in particulate air pollutants.

Ames's strains of Salmonella typhimurium were used to evaluate the mutagenic activity of airbone particulate materials collected at six different points in the industrial area of Ohmuta and the residential area Fukuoka. Tests were done in presence of rat-liver S-9 fraction isolated from rats that had been treated with Aroclor 1254. When the number of revertant colonies per plate was plotted against the amount of methanol extract of particulate air pollutants, using strain TA98, approximately linear relationships were observed for active samples. Generally, mutagenic activity of the samples increased in proportion to the density of air pollutants. In our system, 38--349 microng of methanol extract, from 0.225--4.51 m3 of air from the factory districts in Ohmuta City gave 100 his+ revertants per plate. On the other hand, 54--2300 microng of air pollutants, from 1.29--14.1 m3 of air from the residential districts in Fukuoka City, gave a comparable activity. Every sample from each area had mutagenic activity. Chemicals in air pollutants were fractionated by alumina column chromatography and identified by gas chromatography and mass spectrometry. More than 28 compounds, including 12 unknown substances were identified as polycyclic hydrocarbons. Twelve of these compounds are already known to be carcinogens and to induce reversions to histidine independence in strain TA98 of Salmonella.     

Results of the IPCS collaborative study on complex mixtures.

The International Programme on Chemical Safety (IPCS) sponsored a collaborative study to examine the intra- and inter-laboratory variation associated with the preparation and bioassay of complex chemical mixtures. The mixtures selected were National Institute of Standards and Technology (NIST) Standard Reference Materials (SRMs). 20 laboratories worldwide participated in the collaborative trial. The participating laboratories extracted the organic portion of two particulate samples--an air-particulate sample and a diesel-particulate sample--and bioassayed the extracts. The laboratories simultaneously bioassayed a NIST-prepared extract of coal tar and two control compounds (benzo[a]pyrene, and 1-nitropyrene). The bioassay method used was the Salmonella/mammalian microsome plate-incorporation test using strains TA98 and TA100. Study design also allowed for a comparison of sonication and Soxhlet extraction techniques. The mean extractable masses for the air particles and diesel particles were approximately 5% and 17.5%, respectively. The particulate samples were mutagenic in both strains with and without activation in all 20 laboratories. For TA100 the with and without activation slope values for the air particulate were 162 and 137 revertants per mg particles, respectively. For TA98 the respective diesel slope values were 268 and 269. The mutagenicity slope values for the diesel particles ranged from 3090 (TA98, +S9) to 6697 (TA100, +S9) revertants per mg particles. The coal tar solution was negative for both strains when exogenous activation was not used but was mutagenic in both strains with exogenous activation. The benzo[a]pyrene and 1-nitropyrene were used as positive controls and gave results consistent with the literature. This paper provides a complete summary of the data collected during the collaborative study. Companion papers provide further analysis and interpretation of the results.     

Characterization of organic extracts from standard reference materials 1649, 'urban dust/organics,' and 1650, 'diesel particulate matter', using a microsuspension assay. A WHO/IPCS/CSCM study.

Mutagenicity associated with replicate organic extracts from standard reference materials 1649 'urban dust/organics' (air particles), and 1650, 'diesel particulate matter' (diesel particles), was determined using a Salmonella microsuspension assay. The results indicate that the mutagenicity of samples such as these can readily be determined using the microsuspension assay with only 5% of the mass required for the standard plate incorporation assay. In general, 80% of the variation in mutagenic activity was due to the bioassay procedure and 20% to the extraction process. Extracts from both samples had primarily direct-acting mutagenicity as there were no significant differences in responses with and without metabolic activation (S9). The TA98-S9 mean air particles mutagenic activities (C.V., %) based on mass of extractable organics or particles were 4.4 (4.7%) and 0.29 (3.6%) revertants/micrograms, respectively, and for the diesel particles were 66 (44%) and 12 (29%) revertants/microgram, respectively. More of the observed direct-acting mutagenicity in the diesel particles extracts was due to nitro-substituted compounds because there were significant reductions in activity with TA98NR (45% of TA98 -S9) and TA98-1,8-DNP6 (21% of TA98 -S9). In the air particles extracts, the TA98NR activities were not significantly different from TA98 -S9 but the TA98-1,8-DNP6 levels were.     

Mutagenic activity as a parameter to assess ambient air quality for protection of the environment and human health

Atmospheric pollution has significant effects on maintaining the integrity of ecosystems and on the population's quality of life. Epidemiological studies have clearly associated related health problems, especially respiratory diseases, with exposure to air pollution. Organic compounds adsorbed to the airborne particulate matter are mutagenic in the Salmonella/microsome assay, and a considerable number of them are known to be carcinogenic to rodents. Studies performed at four sites within the urban area of Porto Alegre, capital of the state of Rio Grande do Sul, identified higher mutagenic activity at the sites with heavier vehicle traffic in assays without and with metabolic activation. The responses varied at different seasons of the year, and the highest revertants per cubic meter (rev/m(3)) values were observed in spring for moderately polar compounds, and in summer for non-polar ones. A pilot study was also performed in the region under the influence of a industrial petrochemical area. Most of the sites studied within the industrial area, as compared to others sampled in the nearby environment, presented higher levels of mutagenic activity independent of total suspended particulates (TSP) concentration in the sample. In the urban and industrial regions, the observed mutagenic activities were strongly associated with the presence of polycyclic aromatic hydrocarbons (PAHs). The responses observed in the TA98NR and TA98/1,8-DNP(6) strains suggest the activity of nitrocompounds in both studies. The Salmonella/microsome assay is a sensitive method to define areas contaminated by these compounds, even in samples with TSP values that are consistent with the legal environmental quality standards.     

Genotoxicity and PAC analysis of particulate and vapour phases of environmental tobacco smoke

Samples of indoor air were collected from an office room (88 m3) both before smoking and during experimental smoking of 96 cigarettes by 10 persons within 6 h. The particulates were collected on glass-fibre filters and the vapour-phase compounds on XAD-2 resin. The samples were extracted with acetone and analysed quantitatively for polycyclic aromatic compounds and qualitatively with GC-MS. The extracts of filters and XAD-2 resins were fractionated into neutral/acidic and 2 basic (strong and weak bases) fractions; all these fractions were tested with the sister-chromatid exchange (SCE) assay in Chinese hamster ovary (CHO) cells and with the Salmonella/microsome test (strain TA98). Total concentrations of PAC were 205 ng/m3 in the background sample and 1207 ng/m3 after contamination by cigarette smoking. The total PAC concentrations were 4-6 times higher in the vapour phase than in the particulate phase. The fractions of the particulate samples collected before smoking showed mainly marginal genotoxic activity, whereas after smoking their genotoxicity increased dramatically. The fractions of the vapour phase samples were not genotoxic before smoking, but after smoking the neutral/acidic and strong basic fractions induced responses in both assays. The SCE assay was more sensitive towards the vapour-phase mutagens of environmental tobacco smoke (ETS). The relative responses of the two basic fractions, whereas the fraction containing neutral and acidic compounds was the most potent in the SCE assay. In the Salmonella test, the mutagenic activity was mainly detected with metabolic activation, while the induction of SCE in CHO cells was also seen without an exogenous metabolic activation system.     

Results of a comparative study on the Salmonella pre-incubation and plate incorporation assays using test samples from the IPCS collaborative study.

Three complex mixtures (air particles, diesel particles and a coal tar fraction) and two pure compounds (benzo[a]pyrene and 1-nitropyrene) were tested in both the pre-incubation and the plate incorporation assay employing Salmonella typhimurium TA98 and TA100. Each experiment was conducted independently 2 or 4 times in duplicate in the presence and absence of metabolic activation. The mutagenic activities were calculated by least squares linear regression from the slope of the linear portion of each dose-response curve. Although slightly higher mutagenic activity was observed in the pre-incubation assay for the two pure compounds and with the plate incorporation assay for the diesel particulate sample, the overall data from both assays gave similar values and good correlations in TA100 and TA98. The results indicate that the pre-incubation assay could be used for these samples instead of the plate incorporation assay.     

The release of mutagens from airborne particles in the presence of physiological fluids

Airborne particulates collected indoors in residences and outdoors were extracted by soxhlet extraction and sonication with methanol. In a comparative study in which mutagenic activity was evaluated in the Salmonella typhimurium reversion assay both soxhlet extraction and sonication proved to be suitable extraction methods. First, the residue, obtained by sonication of loaded filters with methanol followed by evaporation to dryness (tar), was sonicated with newborn calf serum and lung lavage fluid from pigs. All serum extracts of the tar were mutagenic to Salmonella typhimurium TA98, and contained direct- and indirect-acting mutagens. However, the mutagenic activity recovered by serum was only about half of the total mutagenic activity of the tar. The other part of the mutagenic activity remained in the tar. Lung lavage fluid was only able to remove 5-10% of direct-acting mutagens from the tar of all samples. About 20% of indirect-acting mutagens from indoor air were recovered in lung lavage fluid, while the lung lavage fluid extract from outdoor air did not show indirect mutagenic activity. Second, mutagenic activity recovered by direct sonication of the filters with physiological fluids was comparable with the recovery obtained by sonication of the tar. However, after sonication of the filter with lung lavage fluid hardly any mutagenic activity remained on the filter, whereas after sonication of the tar a clear mutagenic activity was observed in the non-soluble residue.     

Mutagenic activity of airborne particulate matter in a petrochemical industrial area

Exposure to airborne particulate matter has adverse effects on human health and ecosystem. Mutagenic activity of airborne particulate organic matter extracts in three time periods from total suspended particles (TSP) and particles less than 10 μm (PM10) was evaluated in an area under the influence of a petrochemical industry located in the town of Triunfo, Brazil. The extracts were investigated using the Salmonella/microsome assay, with the microsuspension method. The extracts were obtained by sonication extracted using dichloromethane (DCM) solvent. The fractions were tested for mutagenicity with the Salmonella typhimurium strains TA98 (with and without metabolic activation), TA98NR and TA98/1,8DNP₆; or YG1021 and YG1024. A positive frameshift mutagenic response was observed for the environmental samples during the different periods. The responses according to percentage of extractable organic matter (EOM%), EOM/m³, revertants/μg (rev/μg) and revertants/m³ (rev/m³) were lower for TSP than for PM10 extracts. The highest rev/m³ values were observed in PM10 extract samples collected in winter, July 2005, in the presence (13.79 rev/m³) or absence (6.87 rev/m³) of S9 fraction. Similarly in the first (1995) or second period (2000) the highest values for TSP were observed in winter, but with lower activity (3.00 and 0.89 rev/m³ respectively). The responses observed for the nitrosensitive strains suggest the contribution of nitro, amino and/or hydroxylamino derivatives of PAHs to the total mutagenicity of matter extracted from airborne particles. The Salmonella/microsome assay was a sensitive method to define areas contaminated by genotoxic compounds, even in samples with TSP or PM10 values that are acceptable according to legal environmental quality standards, favoring environmental control measures with an effective response seen in the population's improved quality of life.     

Results of a comparative study on the Salmonella pre-incubation and plate incorporation assays using test samples from the IPCS collaborative study

Three complex mixtures (air particles, diesel particles and a coal tar fraction) and two pure compounds (benzo[a]pyrene and 1-nitropyrene) were tested in both the pre-incubation and the plate incorporation assay employing Salmonella typhimurium TA98 and TA100. Each experiment was conducted independently 2 or 4 times in duplicate in the presence and absence of metabolic activation. The mutagenic activities were calculated by least squares linear regression from the slope of the linear portion of each dose-response curve. Although slightly higher mutagenic activity was observed in the pre-incubation assay for the two pure compounds and with the plate incorporation assay for the diesel particulate sample, the overall data from both assays gave similar values and good correlations in TA100 and TA98. The results indicate that the pre-incubation assay could be used for these samples instead of the plate incorporation assay.