Showing papers in "Environmental Mutagenesis in 1981"
TL;DR: The results support the complementary and confirmatory nature of these tests for genotoxic chemicals and indicate the usefulness of the hepatocyte UDS system as a component in a battery of short-term predictive tests for mutagens/carcinogens.
Abstract: The autoradiographic identification of unscheduled DNA synthesis (UDS) in primary cultures of adult rat hepatocytes (HPC) has been proposed as a predictive test for mutagens/carcinogens. To assess the predictive value of this test, results in the hepatocyte UDS assay were compared with data for bacterial mutagenicity using a modified Ames test. Over 200 compounds representing a variety of chemical classes consisting of procarcinogens, ultimate carcinogens, and noncarcinogens were tested in each system. The accurate discrimination of many carcinogens/noncarcinogens was demonstrated by both systems. The induction of UDS in hepatocytes showed an excellent correlation with bacterial mutagenesis in response to polycyclic aromatic hydrocarbons, aromatic amines, biphenyls, nitrosamines, carbamates, azo-compounds, acridines, halogenated compounds, nitrosureas, quinolines, pyridines, purines, pyrimidines, esters and carbamates. Nitrocompounds, although active in bacteria, were poor inducers of UDS. The results support the complementary and confirmatory nature of these tests for genotoxic chemicals and indicate the usefulness of the hepatocyte UDS system as a component in a battery of short-term predictive tests for mutagens/carcinogens.
269 citations
TL;DR: The mutagenicities of 61 flavonoids and those of 11 compounds structurally related to flavonoid compounds were tested with Salmonella typhimurium strains TA100 and TA98 and it was found that quercetin was the strongest mutagen.
Abstract: The mutagenicities of 61 flavonoids (naturally occurring flavonoid aglycones and flavonal glycosides and synthetic flavonoids) and those of 11 compounds structurally related to flavonoids were tested with Salmonella typhimurium strains TA100 and TA98. Among the 22 flavone derivatives tested, only wogonin was strongly mutagenic, while five derivatives, apigenin triacetate, acacetin, chrysoeriol, pedalitin, and pedalitin tetraacetate, were only weakly mutagenic. Two bisflavonyl derivatives, neither of which has a 3-hydroxyl group, were not mutagenic. Of the 16 flavonol derivatives tested, all except 3-hydroxyflavone and the tetra- and penta-methyl ethers of quercetin were mutagenic. Of the five flavanone derivatives tested, only 7,4-dihydroxyflavanone was mutagenic, showing weak activity. Of the four flavanonol derivatives tested, hydrorobinetin and taxifolin were weakly mutagenic. Of the six isoflavone derivatives tested, tectorigenin was weakly mutagenic. Of the 11 compounds in the miscellaneous group structurally related to flavonoids, only iso-liquiritigenin was mutagenic, showing weak activity. For the emergence of strong mutagenicity, the double bond between positions 2 and 3 and the hydroxyl group at position 3 are required, except in wogonin, which does not have a hydroxyl group at position 3 but is strongly mutagenic to TA100. The 3-O-acetyl ester of flavonol, quercetin, was mutagenic with S9 mix, but 3-O-methyl ethers were not. Six flavonol glycosides, three quercetin glycosides and three kaempferol glycosides were mutagenic after preincubation with “hesperidinase,” a crude extract of Aspergillus niger. Of 66 flavonoid agylcones and compounds structurally related to flavonoids, quercetin was the strongest mutagen. The carcinogenicity of this compound should be clarified because it is ubiquitously found in vegetables.
177 citations
TL;DR: A derivative of Salmonella typhimurium TA98 which does not respond to the potent mutagenicity of 1,8-dinitropyrene is described and appears to be deficient in a nitroreductase which reduces nitrated pyrenes and possibly other nitrated polycyclic aromatic hydrocarbons to corresponding hydroxylamines, the penultimate mutagens.
Abstract: A derivative of Salmonella typhimurium TA98 which does not respond to the potent mutagenicity of 1,8-dinitropyrene is described. This novel strain also shows a lack of response to the mutagenic action of 1,3-dinitropyrene and a greatly reduced response to 1,6-dinitropyrene and 1-nitropyrene. The responses to 1,3,6-trinitropyrene and 1,3,6,8-tetranitropyrene are affected to a much lesser extent. This strain (TA98/1,8DNP6) is fully sensitive to the mutagenicity of 4-nitroquinoline-1-oxide, niridazole, nitroacridine, and nonnitrated frameshift mutagens. This strain appears to be deficient in a nitroreductase which reduces nitrated pyrenes and possibly other nitrated polycyclic aromatic hydrocarbons to corresponding hydroxylamines, the penultimate mutagens.
163 citations
TL;DR: The induction of SCE and chromosomal aberrations by metals reemphasizes the sensitivity of cytological assays and their importance for detecting genetic damage caused by carcinogens.
Abstract: Sister chromatid exchange (SCE) and chromosome aberration induction were determined for several inorganic metal salts. Arsenic, nickel, and beryllium salts at concentrations effective in causing transformation of Syrian hamster cells (HEC) induced SCE and chromosome aberrations of HEC and human lymphocytes, whereas sodium tungstate, a non-transforming chemical, neither induced SCE nor chromosome aberrations. Normal human and hamster cells exhibited equal sensitivity to SCE induction; nontoxic concentrations of sodium arsenite, beryllium sulfate, and nickel sulfate caused an increase of 8-10 SCE/cell over control values. Sodium arsenite, a trivalent arsenic, and sodium arsenate, a pentavalent arsenic, produced increases in SCE but the former was effective at lower concentrations. Both arsenic salts were less efficient in inducing SCE in human whole blood than in purified lymphocyte cultures. Sodium arsenite, sodium arsenate, nickel sulfate, and beryllium sulfate also caused damage consisting primarily of chromatid type of aberrations. In HEC, with doses most effective in SCE induction , all four metals produced aberrations in 16-21% of cells. In human lymphocytes, 34 and 30% of the cells had chromosome damage after sodium arsenite and sodium arsenate, respectively, whereas beryllium sulfate or nickel sulfate caused damage in about 10% of the cells. The induction of SCE andmore » chromosomal aberrations by metals reemphasizes the sensitivity of cytological assays and their importance for detecting genetic damage caused by carcinogens.« less
127 citations
TL;DR: Weighted least squares estimation is used for fitting nonlinear models to Ames test data that are biologically plausible, incorporating mutagenicity, toxicity, and/or saturation.
Abstract: Weighted least squares estimation is used for fitting nonlinear models to Ames test data. The models are biologically plausible, incorporating mutagenicity, toxicity, and/or saturation. Regressions are weighted to compensate for variance that changes with mean and to attain outlier resistance.
61 citations
TL;DR: The use of a battery of indicator strains lacking different repair systems offers the advantage of providing preliminary information concerning the mechanism of DNA damage induction by a test agent, and demonstrates the efficiency of the E coli microsuspension assay as both a qualitative and quantitative screen of DNA-modifying activity.
Abstract: We have devised a microsuspension assay utilizing E coli indicator strains WP2, WP2 uvrA, WP67, CM611, WP100, W3110 polA+, and p3478 pola- for the detection of chemically-induced preferential kill of repair-deficient strains. Data are presented from tests of 77 compounds representing a wide range of chemical classes which demonstrate the efficiency of the E coli microsuspension assay as both a qualitative and quantitative screen of DNA-modifying activity. Furthermore, the use of a battery of indicator strains lacking different repair systems offers the advantage of providing preliminary information concerning the mechanism of DNA damage induction by a test agent.
61 citations
TL;DR: Evidence is presented that suggests that the reduction of the nitro function to the corresponding hydroxylamine might not involve a free nitroso intermediate, and which results in great positional effects of the various isomers on mutagenic activity and specificity.
Abstract: While 2-nitronaphthalene was a weak direct-acting base-substitution mutagen (1.4 revertants/nanomole) for Salmonella typhimurium, the analogous nitronaphthalic acid anhydride and imides were moderate frameshift mutagens (approximately 20 rev/nanomole in strain TA98). Although imide derivatives are efficient DNA intercalators, mutagenicity data indicate that the bulk of the frameshift activity is derived from adduct formation between hydroxylamine intermediates and DNA. The low level of frameshift activity (approximately 8% of total) resulting from simple intercalation (measured in strain TA 1537) is not dependent upon reduction of the nitro function. Evidence is presented that suggests that the reduction of the nitro function to the corresponding hydroxylamine might not involve a free nitroso intermediate. The introduction of a second nitrofunction into nitronaphthalenes results in great positional effects of the various isomers on mutagenic activity and specificity.
52 citations
TL;DR: A simple scheme has been developed for confirming the phenotype of the standard set of Salmonella typhimurium tester strains using a series of filter paper discs impregnated with diagnostic mutagens or bacterial toxins.
Abstract: A simple scheme has been developed for confirming the phenotype of the standard set of Salmonella typhimurium tester strains. This scheme employs a series of filter paper discs impregnated with diagnostic mutagens or bacterial toxins. Up to 6 diagnostic discs can be placed on a petri dish to test a single Salmonella strain. The Salmonellae are distinguished by their responses to ampicillin, crystal violet, nitrofurantoin, 9-aminoacridine, 4-nitro-o-phenylenediamine and sodium azide.
52 citations
TL;DR: Serum and lung cytosol were more effective than acellular lung lavage fluid in releasing mutagenic activity from diesel particles in a manner similar to that of physiological fluids in removing mutagens.
Abstract: The Ames Salmonella typhimurium plate incorporation assay was used to evaluate the mutagenicity of organics extracted from diesel exhaust particles. Organic solvents were more efficient than physiological fluids in removing mutagens from diesel particles, with dichloromethane extracts having the greatest mutagenic activity of the solvents examined. Serum and lung cytosol were more effective than acellular lung lavage fluid in releasing mutagenic activity from diesel particles. The mutagenic activity of diesel particle organics preextracted with dichloromethane is greatly reduced upon the addition of serum and lung cytosol to organics. Subsequently, incubation of protease with serum and lung cytosol-bound diesel organics increases the mutagenic activity. Fluorescence intensity was quantitated and found to correlate with mutagenic activity in the organic and serum extracts, but not the lung cytosol extracts. These studies suggest that substantial mutagenic activity is released from diesel particles upon incubation with serum and lung cytosol.
52 citations
TL;DR: This rat lymphocyte culture system provides a reliable means of investigating chemically induced SCE in the rat and is recommended to maintain low baseline SCE levels and to avoid cytotoxicity.
Abstract: Lymphocyte culture systems have the major advantage of permitting the analysis of in vivo cytogenetic damage with minimal injury to the animal under study. This paper describes a rat lymphocyte culture system designed for the study of sister chromatid exchange (SCE) induced by in vivo exposure to genotoxic agents. A standard protocol was established in which 1 to 2 ml of blood are removed from rats by cardiac puncture, washed three times with phosphate-buffered saline (pH 7.4), and grown in RPMI 1640. 5-Bromodeoxyuridine (BrdU) (1.0 microM) is added after 24 hr, and cells are harvested after 56 hr of culture. Critical steps for successful blast transformation include washing the blood in buffered saline and the adding of 2.0 to 4.0 microgram phytohemagglutinin/ml to the culture medium. The use of low concentrations of BrdU (less than or equal to 5.0 microM) is recommended to maintain low baseline SCE levels and to avoid cytotoxicity. The mutagenic carcinogen, ethyl methanesulfonate (EMS), was used as a positive control agent and at a dose of 300 mg/kg caused a fourfold increase in SCE frequency. Twenty-eight days after EMS administration (30, 100, 300 mg/kg), lymphocytes from treated animals still displayed SCE levels at least 50% above baseline. This system provides a reliable means of investigating chemically induced SCE in the rat.
47 citations
TL;DR: Factors determining the precision and variability of the Ames Salmonella test for mutagenicity were investigated, and the most important source of variability in the agar-overlay method was nonuniformity in the soft-agar layer thickness.
Abstract: Factors determining the precision and variability of the Ames Salmonella test for mutagenicity were investigated. The most important source of variability in the agar-overlay method was nonuniformity in the soft-agar layer thickness. Solution of this problem resulted from application of an agar-leveling table described in this paper. Several other procedural elements also contribute to improved precision, including temperature uniformity during incubation, incubation interval, consistency of plate agar volume, completeness of mixing the soft-agar overlay, peculiarities in the interaction of mutagens and mammalian liver microsomal extract (S9), and methods of storage and controls for tester strains. When these and other effects were well-controlled, variability of the test results was reduced from 200 or 300% to only +/- 10% or less. The significance of the factors affecting precision are discussed and an improved experimental protocol is presented.
TL;DR: It is shown here that the repair of the DNA damage converted into chromosome aberrations is more rapid in Down cells than normal cells, and this more rapid repair results in a higher probability of producing chromosomesAberrations, and hence higher aberration frequencies in Down thannormal cells.
Abstract: Unstimulated lymphocytes from individuals with Down Syndrome (trisomy 21) are more sensitive to the induction of dicentric and ring aberrations by X rays than normal lymphocytes. Several explanations involving the more rapid rejoining of X-ray--induced lesions in Down cells have been offered. It is shown here that the repair of the DNA damage converted into chromosome aberrations is more rapid in Down cells than normal cells. This more rapid repair results in a higher probability of producing chromosomes aberrations, and hence higher aberration frequencies in Down than normal cells.
TL;DR: In this paper, the authors compared the mutagen-activating capabilities of liver S-9 preparations from both Syrian golden hamsters and Sprague-Dawley rats, and found that the hamster liver was consistently more effective than rat liver for the activation of 2-acetylaminofluorene (AAF), 2-aminoanthracene (AA), 3-methylcholanthrene (MCA), and diethylnitrosamine (DEN) to their mutagenic forms.
Abstract: Before exerting a carcinogenic or mutagenic effect, many carcinogens must undergo metabolic activation. Liver S-9 preparations from rats treated with Aroclor 1254 (AC) have been widely used to extend the ability of the Salmonella mutagenicity assay to detect such nondirect-acting agents. We compared the mutagen-activating capabilities of liver S-9 preparations from both Syrian golden hamsters and Sprague-Dawley rats. The comparison was made between S-9 fractions from untreated, and phenobarbital (PB)- and AC-treated animals of both sexes. Also compared was the effect of varying the amount of S-9 protein. The preparations from hamster liver were consistently more effective than preparations from rat liver for the activation of 2-acetylaminofluorene (AAF), 2-aminoanthracene (AA) (except for AC-induced preparations), 3-methylcholanthrene (MCA), and diethylnitrosamine (DEN) to their mutagenic forms. Induced preparations from rats were more active than those from hamsters when using benzo(a)pyrene (BP), while with uninduced preparations, the opposite was true.
There was also a relationship between the amount of S-9 protein and the number of revertant colonies obtained. With the aromatic amines (AAF and AA), a partial inhibition of mutagenicity occurred at the highest protein concentrations tested. With BP, an inverse relationship was observed between protein concentration and the number of revertants occurring in the presence of the preparation from PB- and AC-induced male and female rats.
The relative order of activities of aryl hydrocarbon hydroxylase (AHH) and BP-4,5-epoxide hydrase (EH) in the preparations was AC-induced > PB-.
TL;DR: The results of these studies show that DES is a clastogenic agent in vivo which may relate to its carcinogenicity.
Abstract: Diethylstilbestrol diphosphate (DES-dp) clastogenesis was examined in the bone marrow of C57B1/6 male and female mice. Significant and sex-related dose effects were observed for the induction of chromatid-type chromosomal aberrations and for the inhibition of cellular proliferation. Females were more sensitive to the effects of DES-dp than males when assessed for either induced chromosomal aberrations or proliferative inhibition. Contrary to other published results, we did not observe either an increase in sister chromatid exchanges or an increased incidence of aneuploidy. Ovariectomy reduced the ability of DES-dp to inhibit cellular proliferation and decreased the high degree of variability between animals at high doses of DES-dp. The results of our studies show that DES is a clastogenic agent in vivo which may relate to its carcinogenicity.
TL;DR: Data presented in this paper from tests of 61 additional compounds indicate that the B subtilis H17 and M45 strains provide an effective microbial system for identification of DNA damage susceptible to postreplicational repair.
Abstract: We have demonstrated the utility of an Escherichia coli microsuspension assay to detect and characterize chemical mediation of DNA damage by a wide variety of mutagens and carcinogens. The assay have been improved by the development of a microsuspension modification to the Bacillus subtilis ''rec'' assay. The addition of these gram-positive organisms has allowed detection of DNA damage induced by benzo(a)pyrene (B(a)P), 3-aminopyrene (3-AP), 7, 12-dimethylbenz(a)anthrancene (DMBA), 3-methylcholanthrene (3-MC), and 4-nitrobiphenyl (4-NBP). Data presented in this paper from tests of 61 additional compounds, including a representative number of direct and promutagenic agents, indicate that the B subtilis H17 and M45 strains provide an effective microbial system for identification of DNA damage susceptible to postreplicational repair. The results of this study further suggests that the inclusion of these strains in the microsuspension assay for DNA damage will markedly enhance the detection of agents which cannot readily penetrate the intact cell wall of E coli.
TL;DR: It is proposed that aniline may exert its tumorigenic potential in rats through the production of both genotoxic and cytotoxic metabolites as well as a less clear-cut, dose-related increase in SCE frequency with a 1.4-fold elevation.
Abstract: Sister chromatid exchange (SCE) and cell cycle analyses in human fibroblasts were used to ascertain the relative genotoxicity and cytotoxicity of aniline and its metabolites. Fibroblasts were allowed to attach to plastic petri dishes for 4 hr and exposed in serum-free medium for 2 hr to the following: aniline HCl (0.05–10.0 mM), acetanilide (0.1–10.0 mM), o-hydroxyacetanilide (0.01–2.0 mM), p-hydroxyacetanilide (0.1–10.0 mM), o-aminophenol (0.01–0.3 mM), p-aminophenol (0.005–0.2 mM), N-phenylhydroxylamine (0.001–0.5 mM). Triethylenemelamine (TEM; 0.25 and 0.49 μM) was used as a positive control. The treatment medium was replaced with complete medium containing 5-bromo-deoxyuridine (10 μM), and cells were allowed to grow for 48 hr with Colcemid present for the final 4.5 hr. Cell cycle analyses (percentage of cells in first, second, or third division) revealed that p-hydroxyacetanilide (10mM), p-aminophenol (≥0.1 mM), o-aminophenol (≥0.1 mM), N-phenylhydroxylamine (≥ mM), and TEM (0.49 μM) inhibited cell proliferation. Cell death was seen at doses of 0.5 mM N-phenylhydroxylamine, 0.2 mM p-aminophenol, and 0.3 mM o-aminophenol. Significant increases (P0.05) in SCE frequencies were found with aniline HCl, o-aminophenol, N-phenylhydroxylamine, and TEM. On an SCE/mmole basis at the highest concentrations examined, o-aminophenol was 270 times more potent than aniline in inducing SCE, whereas TEM was about 390 times more potent than o-aminophenol. Furthermore, fibroblasts treated with o-aminophenol responded in a dose-dependent fashion and exhibited a 2-fold increase in SCE frequency. N-phenylhydroxylamine induced a less clear-cut, dose-related increase in SCE frequency with a 1.4-fold elevation. Only marginal increases in SCE were observed with aniline at the highest doses. Using these data, we propose that aniline may exert its tumorigenic potential in rats through the production of both genotoxic and cytotoxic metabolites.
TL;DR: Two in vitro short-term tests that possess intrinsic capability for xenobiotic biotransformation detected the genotoxicity of a variety of carcinogenic polycyclic aromatic hydrocarbons and are useful for the evaluation of this class of carcinogens.
Abstract: The hepatocyte primary culture (HPC)--DNA repair test and the adult rat liver epithelial cell (ARL)--hypoxanthine-guanine phosphoribosyl transferase (HGPRT) mutagenesis assay are two in vitro short-term tests that possess intrinsic capability for xenobiotic biotransformation. Both assays detected the genotoxicity of a variety of carcinogenic polycyclic aromatic hydrocarbons. Thus, these two tests, which embody intact cellular metabolism, are useful for the evaluation of this class of carcinogens and provide results that strengthen those obtained in tests dependent upon subcellular metabolism.
TL;DR: It is concluded that the SCE assay in human lumphocytes may be a useful indicator of the potential risks to humans of airborne particulate pollutants, as it utilizes human cells recently taken from the host, is rapid and economical, and requires small quantities of test materials.
Abstract: The genotoxicities of organic extracts of airborne particles have been studied extensively in the Salmonella/mammalian microsome (Ames) test, but in few other bioassays. In these studies, we tested benzene-acetone extracts of particulate pollutants collected in Lexington, Kentucky, for capacity to induce increases in sister chromatid exchanges (SCE) in human lumphocytes and V79 cells, as well as in the Ames assay. Extracts induced linear dose-related increases in SCE in human lumphocytes and in bacterial revertants.However, variable responses were observed in SCE assays in V79 cells with and without activation by rat liver S9 or feeder layers of irradiated Syrian hamster fetal cells. We conclude that the SCE assay in human lumphocytes may be a useful indicator of the potential risks to humans of airborne particulate pollutants, as it utilizes human cells recently taken from the host, is rapid and economical, and requires small quantities of test materials. However, thorough studies of the quantitative relationships between SCE induction and mutagenicity in human cells are needed.
TL;DR: Unstimulated human lymphocytes in whole blood were treated with lead, cadmium, and zinc acetate separately and in combinations of two or three metal salts with different concentrations of between 10(-3) and 10(-5) moles, respectively.
Abstract: Unstimulated human lymphocytes in whole blood were treated for three hours with lead, cadmium, and zinc acetate separately and in combinations of two or three metal salts with different concentrations of between 10(-3) and 10(-5) moles, respectively. Untreated cultures and sodium acetate-treated samples served as controls. Chromosome analysis from 48 cultures revealed higher incidences of chromatid-type aberrations and gaps only for cultures exclusively treated with cadmium. The results are discussed under the respect of heavy metal metabolism in human lymphocytes.
TL;DR: Chemical analysis of the highly mutagenic tap water concentrate from May revealed the presence of a number of organic contaminants that were absent from control concentrates prepared from deionized and distilled treated-well water.
Abstract: We have studied the mutagenicity of the water from Lake Bloomington and of the tap water that is made from the lake water. The lake, which is the source of drinking water for Bloomington, Illinois (pop. 44,000), is surrounded by land that is farmed intensively--being mainly in maize and soybeans. Samples were collected monthly from May through October 1979 and concentrated 3,000X with XAD-2 resin. Nearly all of the lake and tap water concentrates were mutagenic in Salmonella typhimurium TA98 and TA100 in the presence of S-9 mix, and the May tap water concentrate was highly mutagenic. In addition, many of the concentrates were toxic to the bacteria in the absence of S-9 mix. Chemical analysis of the highly mutagenic tap water concentrate from May revealed the presence of a number of organic contaminants that were absent from control concentrates prepared from deionized and distilled treated-well water. In addition, unconcentrated lake and tap water were tested in a reverse-mutation test in maize (Zea mays); no mutagenicity was detected. This study indicates that the contamination of drinking water with agricultural and/or industrial chemicals may result in a potential health hazard.
TL;DR: The response of strain TA100 (mostly a base-substitution detector) is opposite that of TA98, showing antagonism and additivity in similar concentration ranges.
Abstract: Binary mixtures of benzo(a)pyrene (B(a)P) and benzo(e)pyrene (B(e)P) produce synergistic mutagenic (comutagenic) responses in Salmonella typhimurium strain TA98 (a frameshift detector). The optimum enhancement (25 X) was found at B(a)P concentration of 0.3 microgram/plate and B(e)P concentration of 1.5 microgram/plate. The response of strain TA100 (mostly a base-substitution detector) is opposite that of TA98, showing antagonism and additivity in similar concentration ranges.
TL;DR: Frequencies of sister chromatid exchanges (SCE) were analyzed in bone marrow cells of mice injected with mitomycin C both before and during infusion with BrdU, indicating that most or all MMC-induced DNA damage that results in SCE formation is removed in a single cell cycle after its administration.
Abstract: Frequencies of sister chromatid exchanges (SCE) were analyzed in bone marrow cells of mice injected with mitomycin C (MMC) both before and during infusion with bromodeoxyuridine (BrdU). Administration of MMC at 1, 6.5, and 13 hours after the onset of BrdU infusion resulted in the induction of approximately 45 SCE/cell, independent of time of administration. When MMC was injected 26 hours prior to BrdU infusion, only baseline levels of SCE were noted. The effects of multiple doses of MMC (chronic administration) were examined in mice treated with 1--5 mg/kg on a weekly or bimonthly basis. SCE analysis was performed one week after the final injection. At all doses and with all treatment regimes, SCE frequencies did not differ from control levels. The results indicate that most or all MMC-induced DNA damage that results in SCE formation is removed in a single cell cycle after its administration.
TL;DR: In this article, the mutagenicity of ambient air particles from Morgantown, West Virginia, has been monitored for 6 months using the Ames Salmonella assay system, where airborne particles were extracted with dichloromethane (DCM) and/or ethyl acetate plus methanol (E + M) in sequence.
Abstract: The mutagenic activity of ambient air particles from Morgantown, West Virginia, has been monitored for 6 months using the Ames Salmonella assay system. Airborne particles, collected on glass fiber filters using a Hi-Vol sampler, were extracted with dichloromethane (DCM) and/or ethyl acetate plus methanol (E + M) in sequence. A dose-dependent mutagenic response was observed in Salmonella typhimurium TA 98 for DCM extracts from all samples. E + M extracts were mutagenic only when samples were extracted with E + M before DCM extraction. The mutagenic activity of samples collected in June and July was independent of S-9 in vitro activation, whereas the mutagenicity of those collected from October to December increased in the presence of S-9 activation. The class fractionation of extracts showed that only acidic and polynuclear aromatic fractions were mutagenic. The mutagenicity of particles from Morgantown air was also detected with the Salmonella arabinose-resistant assay system.
TL;DR: Reaction of fuel aromatics with NO2 may be one mechanism which contributes to the formation of cytotoxic and mutagenic activities in diesel exhaust.
Abstract: 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 dimethyl 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.
TL;DR: The use of the established mutagenesis assay in Paramecium as a prescreen for hazardous environmental particles provided evidence for both heat-stable, heat-labile and acid extractable mutagenic agents.
Abstract: The use of the established mutagenesis assay in Paramecium as a prescreen for hazardous environmental particles is described. Since these protozoans ingest particles of the size respired by animals and man, the biological effects of the respirable fraction of fly ash particles were monitored in particle-feeding eukaryotic cells. Fly ash from coal combustion was utilized for these studies and was found to be mutagenic. The effects of physical and chemical treatment of the particle mutagenicity provided evidence for both heat-stable, heat-labile and acid extractable mutagenic agents.
TL;DR: These mutants are all stable, and should be useful for the study of mammalians DNA repair processes and mechanisms of mutagenesis.
Abstract: Through a new approach, we have sought to isolate ultraviolet light (UV)-sensitive and DNA repair mutant Chinese hamster fibroblasts. The procedure consisted of 1) mutation induction by 5-bromodeoxyuridine (Brd U)-blacklight and UV treatments; 2) incorporation of 3H-thymidine in repair-proficient cells at high temperature (38.5 degrees C) following UV damage; 3) cold holding (4.0 degrees C) of these cells to induce tritium killing; and 4) recovery and testing of repair-deficient and UV-sensitive cells which have survived and formed colonies at low temperature (34.0 degrees C). In our initial attempt at this protocol, we isolated 72 surviving colonies from 2 x 10(7) cells plated for selection. Of the 72 colonies, 20 demonstrated potential interest and four were selected for extensive study. One, identified as UVs-7, is slightly more sensitive to UV, but not sensitive to X rays or N-acetoxy-2-acetylaminofluorene (NAc-AAF). The mutant exhibits a highly reduced level of unscheduled DNA synthesis (UDS), as compared to the parental line. Two additional lines, UVs-40 UVs-44, are sensitive to UV, X ray, N-methyl-N-nitro-N-nitrosoguanidine (MNNG), and NAc-AAF, but exhibit normal UDS. A fourth line, UVr-23, has enhanced UDS, is resistant to UV, but exhibits no difference in sensitivity to x ray or NAc-AAf. These mutants aremore » all stable, and should be useful for the study of mammalians DNA repair processes and mechanisms of mutagenesis.« less
TL;DR: The cytotoxicity and mutagenicity of the fungicides captan and folpet were determined in the CHO/HGPRT system which utilizes Chinese hamster ovary cells and resistance to 6-thioguanine to estimate mutation induction at the hypoxanthine-guanine phosphoribosyl transferase locus.
Abstract: The cytotoxicity and mutagenicity of the fungicides captan and folpet were determined in the CHO/HGPRT system which utilizes Chinese hamster ovary cells and resistance to 6-thioguanine to estimate mutation induction at the hypoxanthine-guanine phosphoribosyl transferase locus. Treatment of cultures with each compound for 5 hr in serum-free medium resulted in reproducible, significant, concentration-dependent increases in the frequency of 6-thioguanine-resistant mutants.
TL;DR: It is concluded that low temperature storage of activating systems is an acceptable procedure, however, the results indicate that certain enzyme activities change during storage, suggesting that aberrant results may be obtained when stored activating systems are used in in vitro tests to screen for mutagens.
Abstract: Activating systems for in vitro mutagenesis assays are commonly prepared and stored at low temperature until required. The objective of the studies reported here was to determine the long-term stability of activating systems stored at - 85 degrees C. A broad range of microsomal enzymes in the postmitochondrial supernatant (PMS) and the microsomal fraction of livers from Aroclor 1254 treated rats were studied in conjunction with the ability of these fractions to catalyse the conversion of dimethylnitrosamine (DMN) and benzo(a)pyrene (B(a)P) to products mutagenic to Chinese hamster ovary (CHO) cells and Salmonella typhimurium TM677. Biphenyl-2- and biphenyl-4-hydroxylase showed a rapid decline in activity on storage, epoxide hydratase activity increased with storage and other enzyme activities studied were relatively stable for up to 32 weeks. No consistent trends in the ability of either the microsomes or the PMS to catalyze DMN or B(a)P induced mutation were observed for up to 12 weeks with CHO cells and 24 weeks with bacteria. It is concluded that low temperature storage of activating systems is an acceptable procedure. However, the results also indicate that certain enzyme activities change during storage, suggesting that aberrant results may be obtained when stored activating systems are used in in vitro tests to screen for mutagens.
TL;DR: The Ames test was performed with a mixture of rat/hamster S-9, and except for an increased mutagenic activation by the mixture with nitrosopiperidine the mixture was comparable to the rat S- 9 alone, indicating that replacing ratS-9 with a rat/Hamster S9 mixture in the standard Ames test could increase the results of the test without interfering with rat S -9 activity.
Abstract: Based on the findings of Nagao et al [1978] that phenacetin is negative in the standard Ames test with Aroclor induced rat S-9 and positive with hamster S-9, the test was performed with a mixture of rat/hamster S-9. Phenacetin was mutagenic with the mixture. The activity of the mixture was compared to the rat S-9 alone with low concentrations of 2-aminoanthracene (a strong promutagen for Salmonella typhimurium TA 1535, TA 100, TA 1537, and TA 98), nitrosopiperidine (a weak promutagen), and 1,2 epoxybutane (a weak, direct-acting mutagen). Except for an increased mutagenic activation by the mixture with nitrosopiperidine the mixture was comparable to the rat S-9 alone, indicating that replacing rat S-9 with a rat/hamster S9 mixture in the standard Ames test could increase the sensitivity of the test without interfering with rat S-9 activity.