Strains YG1024 Research Articles

Urinary mutagenicity as a biomarker in workers exposed to benzidine: correlation with urinary metabolites and urothelial DNA adducts.

Urinary mutagenicity has been used in occupational and epidemiological studies for over two decades as a cost-effective, general biomarker of exposure to genotoxic agents. However, few studies have compared urinary mutagenicity to additional biomarkers determined among low- and high-exposed groups. To address this issue, we evaluated the relationship between urinary mutagenicity and other types of biomarkers in a cross-sectional study involving 15 workers exposed to the urinary bladder carcinogen benzidine (BZ, high exposure), 15 workers exposed to BZ-dyes (low exposure), and 13 unexposed controls in Ahmedabad, India. Urinary organics were extracted by C18/methanol and evaluated for mutagenicity in the presence of S9 in the Salmonella strain YG1024, which is a frameshift strain that overproduces acetyltransferase. The results were compared to biomarker data reported recently from the same urine samples (Rothman et al., Proc. Natl Acad. Sci. USA, 93, 5084-5089, 1996) that included a metabolite biomarker (the sum of the urinary levels of BZ + N-acetylbenzidine + N,N'-diacetylbenzidine) and a DNA adduct biomarker [a presumptive N-(3'-phosphodeoxyguanosin-8-yl)-N'-acetylbenzidine (C8dG-ABZ) DNA adduct in exfoliated urothelial cells]. The mean +/- SE urinary mutagenicity (revertants/micromol of creatinine) of the low-exposure (BZ-dye) workers was 8.2 +/- 2.4, which was significantly different from the mean of the controls (2.8 +/- 0.7, P = 0.04) as was that of the mean of the high-exposure (BZ) workers (123.2 +/- 26.1, P < 0.0001). Urinary mutagenicity showed strong, positive correlations with urinary metabolites (r = 0.88, P < 0.0001) and the level of the presumptive C8dG-ABZ urothelial DNA adduct (r = 0.59, P = 0.0006). A strong association was found between tobacco use (bidi smoking) and urinary mutagenicity among the controls (r = 0.68, P = 0.01) but not among the exposed workers (r = 0.18, P = 0.11). This study confirms the ability of a biomarker such as urinary mutagenicity to detect low-dose exposures, identify additional genotoxic exposures among the controls, and correlate strongly with urinary metabolites and DNA adducts in the target tissue (urinary bladder epithelia) in humans.

Antimutagenicity of xanthophylls present in Aztec Marigold ( Tagetes erecta) against 1-nitropyrene

The principal natural food colorants used in modern food manufacture are anthocyanins, betalains, carotenoids, chlorophylls, riboflavin and caramel. Carotenoids (carotenes and xanthophylls) occur naturally in some foods such as carrots, red tomatoes, butter, cheese, paprika, palm oil, corn kernels, marigold petals, annatto, and red salmon. Carotenoids ( α- or β-carotene and xanthophylls) are excellent antioxidants and inhibit some types of cancers. In the present study, we used the Salmonella typhimurium tester strain YG1024 in the plate-incorporation test to examine the antimutagenicity of xanthophylls extracted from Aztec Marigold ( Tagetes erecta) on 1-nitropyrene (1-NP) mutagenicity. Further, we investigated the effect of lutein on DNA-repair system of tester strain YG1024, using a preincubation test. The possible mechanism of lutein on 1-NP mutagenicity was studied by comparing the absorption spectrum of lutein, 1-NP and lutein plus 1-NP. In a dose-response curve of 1-NP, the mutagenic potency was 4 317 revertants/nmol, and the dose of 0.06 μg of 1-NP/plate was chosen for the antimutagenicity studies. Lutein and xanthophylls from Aztec Marigold (pigments for poultry and human use) inhibited mutagenicity of 1-NP in a dose-dependent manner. Lutein and the pigments were not toxic to the bacteria at the concentrations tested (0.002, 0.02, 0.2, 2.0 and 10 μg/plate). The percentages of inhibition of 1-NP mutagenicity were 72%, 92% and 66.2% for lutein (10 μg/plate), pigment for poultry use (10 μg/plate) and pigment for human use (2 μg/plate), respectively. Lutein had no effect on the DNA-repair system of strain YG1024. A new peak was detected at 429 nm when lutein was added at 1-NP, and it was stable throughout the incubation time. The results suggest that the major mechanisms of lutein against 1-NP mutagenicity is the potential formation of a complex between lutein and 1-NP, which could limit the bioavailability of 1-NP.

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 microM induced a concentration-response in Salmonella typhimurium. At concentrations above 5 microM, 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 microM 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-acetyltransferase to generate the ultimate mutagenic products.

Antimutagenic activity of natural xanthophylls against aflatoxin B1 inSalmonella typhimurium

Carotenoids (carotenes and xanthophylls) are excellent antioxidants with antimutagenic and anticarcinogenic properties. They occur naturally in some foods such as carrots, red tomatoes, butter, cheese, paprika, palm oil, corn kernels, Marigold petals, annatto, and red salmon. In the present study, we used the Salmonella plate incorporation test to examine the effect of xanthophylls extracted from Aztec Marigold (Tagetes erecta) on the AFB1 mutagenicity, using tester strain YG1024. The effect of lutein on the DNA-repair system in YG1024 was investigated by a pre-incubation test. In a dose-response curve of AFB1, the mutagenic potency was 1,031 revertants/nmol. The dose of 0.5 microgram AFB1/ plate was chosen for the antimutagenicity studies. Pure lutein and xanthophylls from Aztec Marigold flower (oleoresin and xanthophyll plus) inhibited the mutagenicity of AFB1 in a dose-dependent manner. The pigments were more efficient at inhibiting the AFB1 mutagenicity than pure lutein. The percentages of inhibition on AFB1 mutagenicity were 37, 66, and 76% for lutein, oleoresin, and xanthophyll plus at the dose of 2 micrograms/plate, respectively. Lutein had a modest effect on the DNA-repair system of YG1024. In spectrophotometric studies, a new absorption peak was detected at 378 nm when lutein and AFB1 were incubated together, and lutein reacted with AFB1 metabolites. The results suggest that the inhibitory mechanism of lutein against AFB1 mutagenicity is most probably the result of a combination of the following events: formation of a complex between lutein and AFB1, direct interaction between lutein and AFB1 metabolites, and finally that the lutein may also affect the metabolic activation of AFB1 by S9 and the expression of AFB1-modified Salmonella DNA.

Mutagenic synergy between paraoxon and mammalian or plant-activated aromatic amines

Paraoxon (diethyl-p-nitrophenylphosphate) is the toxic, but non-mutagenic metabolite of the organophosphorus ester (OP) insecticide parathion. Although this agent has been used as a deacetylase inhibitor in many studies, we discovered a mutagenic synergy with paraoxon and plant-activated m-phenylenediamine or with direct-acting 2-acetoxyacetylaminofluorene in Salmonella typhimurium cells [Gichner T et al. (1996): Environ Mol Mutagen 27; 59-66]. In the present study, mammalian-activated m-phenylenediamine, o-phenylenediamine, p-phenylenediamine, benzidine, 2,3-diaminophenazine or 2-aminofluorene, as well as plant-activated benzidine or 2-aminofluorene expressed an elevated mutagenic potency when assayed with S. typhimurium strain YG1024 in the presence of paraoxon. Under non-toxic conditions, paraoxon amplified the S. typhimurium mutant yield induced by these aromatic amines between 1.9-fold and 8.4-fold. Spectrophotometric analysis demonstrated that the rate of degradation of 2-acetoxyacetylaminofluorene was not significantly different in phosphate buffer with or without paraoxon or with S. typhimurium cytosol with or without paraoxon. Also paraoxon-mediated mutagenic synergy does not appear to be due to a direct reaction with aromatic amines. Mutagenic synergy between aromatic amines and OP oxon products may be a cause of concern because people are chronically exposed to environmental and dietary aromatic amines, and a significant segment of the U.S. population tested positive for OP insecticide residues.

Nitroarene concentrations and direct-acting mutagenicity of diesel exhaust particulates fractionated by silica-gel column chromatography.

Diesel exhaust particulates were extracted with benzene-ethanol (3:1, v/v) and separated into five fractions by silica-gel column chromatography. Direct-acting mutagenic activity was assayed by the Ames test using the Salmonella typhimurium YG1024 strain. The total activity of five fractions was about four times greater than that of the crude extract, suggesting that the activities in the fractions were suppressed in the crude extract. Strong activity was observed in fraction 4 which was eluted with dichloromethane (61.5% of the total activity) and fraction 5 which was eluted with ethanol (35.3%). Nitropolycyclic aromatic hydrocarbons (NPAHs) were determined by high-performance liquid chromatography with chemiluminescence detection. They were found mainly in fraction 4, although one NPAH was in fraction 3 which was eluted with n-hexane-dichloromethane (3:1, v/v). Based on these results, 53.1% of the activity in fraction 4 was attributed to NPAHs. The contribution of 1-nitropyrene and 1,3-, 1,6- and 1,8-dinitropyrenes was great and that of the other NPAHs was small. The mutagenic compound in fraction 5 was not identified. Fractions 1 and 2, which were eluted with n-hexane, and fraction 3 suppressed the activity of fraction 4. Polycyclic aromatic hydrocarbons in fractions 2 and 3 were considered as possible suppressors of NPAHs.

Human urine mutagenicity study comparing cigarettes which burn or primarily heat tobacco

Cigarette smokers have been reported to void urine which is more mutagenic, as measured in the Ames assay, than urine voided by non-smokers. Condensate from the mainstream smoke of a cigarette which primarily heats tobacco (test cigarette) has shown significantly reduced mutagenicity in a battery of in vitro genotoxicity assays compared with tobacco-burning cigarettes. The objective of this study was to determine whether the reduction in mutagenic activity observed in the in vitro assays would be reflected in the urine of smokers of the test cigarette. Twenty smokers were enroled in a 4-week crossover study, with each smoker consuming test cigarettes ad libitum for a week and their usual brand of tobacco-burning cigarettes the other 3 weeks. Diet was strictly controlled throughout the study, and broiled and pan-fried meat was not served to minimize ingestion of mutagenic protein pyrolysis products. There was no statistically significant difference ( p = 0.06) in consumption of tobacco-heating and tobacco-burning cigarettes. There were no statistically significant differences ( p = 0.22) in salivary cotinine concentrations for smokers when smoking either tobacco-burning or tobacco-heating cigarettes. Urinary nicotine (ng/mg creatinine) was not different ( p = 0.31) for smokers when smoking either tobacco-burning or tobacco-heating cigarettes. Urinary cotinine (ng/mg creatinine) was 32% lower ( p = 0.0004) when smoking tobacco-heating cigarettes as compared with smoking tobacco-burning cigarettes. Twenty-four-hour urine samples were collected twice weekly, concentrated using XAD-2 resin and tested in Ames strains TA98 and YG1024 with metabolic activation. Tobacco-burning cigarette smokers experienced a 79% reduction in urinary mutagenicity as measured in strain YG1024 and a 72% reduction as measured in strain TA98 during the week that they smoked the tobacco-heating cigarette while maintaining a fixed dietary regimen. The results of this study indicate that smokers of tobacco-heating cigarettes void urine which is significantly less mutagenic than urine voided by smokers of tobacco-burning cigarettes.

Urinary mutagenicity and thioethers in nonsmokers: Role of environmental tobacco smoke (ETS) and diet

The urinary excretion of mutagens and thiothers was investigated in a controlled diet study and in two field studies. A diet containing charcoal-broiled meat and other items rich in mutagenic compounds increased the urinary mutagenicity as assessed in Salmonella typhimurium strain TA98 with metabolic activation ∼ 46-fold compared to a diet low in mutagens. The excretion of thiothers after ingestion of the diet rich in mutagens also increased significantly when compared to the diet low in mutagens. The increase was associated with the content of preformed thiothers in the diet. In the first field study with 21 nonsmokers, urinary mutagenicity as assessed in Salmonella typhimurium strain TA98 and excretion of thiothers showed no relation to either the self-reported exposure to environmental tobacco smoke (ETS) or to serum cotinine concentrations used as an objective marker for ETS exposure. In the second field study, urinary mutagenicity was determined with a tobacco-smoke sensitive Salmonella typhimurium strain YG1024 with metabolic activation. No correlation was found between the mutagenic activity in urine and ETS exposure duration, nicotine on the personal sampler, cotinine in saliva and cotinine in urine. Our results suggest that real-life ETS exposure does not measurably increase either urinary mutagen or urinary thioether excretion. Furthermore, diet seems to be the most important source for both urinary mutagen and thioether excretion in nonsmokers.

The mode of action of Trp-P-1 in DNA excision repair

To determine the mutagenicity of nitrophenanthrenes, three mononitrophenanthrenes (NPhs), 11 dinitrophenanthrenes (diNPhs) and eight trinitrophenanthrenes (tiNPhs) were synthesized, and their mutagenicity was investigated by using Salmonella typhimurium his — strains TA98, TA100, and TA98NR, nitroreductase-deficient, and TA98/1,8-DNP6, O-acetyltransferase-deficient mutants, and strains YG1021 and YG1026, nitroreductase-overproducing mutants of TA98 and TA100, respectively, and strains YG1024 and YG1029, O-acetyltransferase-overproducing mutants of TA98 and TA100, respectively. 1-, 3- and 9-NPhs induced 329, 620 and 438 revertants per nmol in strain TA100, respectively, and 4,839, 11,309 and 16728 revertants per nmol, respectively, in strain YG1029. Mutagenicity of 1,6-, 2,6-, 2,9-, 2,10-, 3,5-, 3,6- and 3,10-diNPh was elevated in strains YG1021, YG1024, YG1026 and YG1029. Among these derivatives, 1,6-, 2,6-, 3,6- and 3,10-diNPhs were more mutagenic in strains YG1024 and YG1029 than YG1021 and YG1026, and they showed a structure-activity relationship between mutagenicity and NO2-substitution. Nitro derivatives substituted at the 3 and 6 positions of their chemical structure strongly mutated both strains YG1024 and YG1029, whereas those substituted at the 9 and 10 positions showed weak mutagenicity. In addition, nitro substituents at positions 4 and 5 were perpendicular while those on positions 2,3,6 and 7 were nearly coplanar to the aromatic ring. Furthermore, 2,6,9-, 3,6,9- and 1,6,9-trinitrophenanthrenes (triNPhs) were mutagenic for strain TA100, and their mutagenicity was more enhanced in YG1024 and YG1029 than in YG1021 and YG1026. Of the eight triNPhs all except 1,5,10-triNP were mutagenic in TA98 and TA100, and their mutagenicity was more enhanced in YG1024 and YG1029 than in YG1021 and YG1026. These results suggest that these compounds are mutagens that are activated by O-acetyltransferase esterification following nitroreductase. The nitrated derivatives substituted at the 2(7) and 3(6) positions of the phenanthrene ring were highly mutagenic. The relationship between chemical structure and the mutagenicity of NPh derivatives is discussed.

Mutagenicity of nitrophenanthrene derivatives for Salmonella typhimurium: effects of nitroreductase and acetyltransferase

To determine the mutagenicity of nitrophenanthrenes, three mononitrophenanthrenes (NPhs), 11 dinitrophenanthrenes (diNPhs) and eight trinitrophenanthrenes (tiNPhs) were synthesized, and their mutagenicity was investigated by using Salmonella typhimurium his — strains TA98, TA100, and TA98NR, nitroreductase-deficient, and TA98/1,8-DNP 6, O-acetyltransferase-deficient mutants, and strains YG1021 and YG1026, nitroreductase-overproducing mutants of TA98 and TA100, respectively, and strains YG1024 and YG1029, O-acetyltransferase-overproducing mutants of TA98 and TA100, respectively. 1-, 3- and 9-NPhs induced 329, 620 and 438 revertants per nmol in strain TA100, respectively, and 4,839, 11,309 and 16728 revertants per nmol, respectively, in strain YG1029. Mutagenicity of 1,6-, 2,6-, 2,9-, 2,10-, 3,5-, 3,6- and 3,10-diNPh was elevated in strains YG1021, YG1024, YG1026 and YG1029. Among these derivatives, 1,6-, 2,6-, 3,6- and 3,10-diNPhs were more mutagenic in strains YG1024 and YG1029 than YG1021 and YG1026, and they showed a structure-activity relationship between mutagenicity and NO 2-substitution. Nitro derivatives substituted at the 3 and 6 positions of their chemical structure strongly mutated both strains YG1024 and YG1029, whereas those substituted at the 9 and 10 positions showed weak mutagenicity. In addition, nitro substituents at positions 4 and 5 were perpendicular while those on positions 2,3,6 and 7 were nearly coplanar to the aromatic ring. Furthermore, 2,6,9-, 3,6,9- and 1,6,9-trinitrophenanthrenes (triNPhs) were mutagenic for strain TA100, and their mutagenicity was more enhanced in YG1024 and YG1029 than in YG1021 and YG1026. Of the eight triNPhs all except 1,5,10-triNP were mutagenic in TA98 and TA100, and their mutagenicity was more enhanced in YG1024 and YG1029 than in YG1021 and YG1026. These results suggest that these compounds are mutagens that are activated by O-acetyltransferase esterification following nitroreductase. The nitrated derivatives substituted at the 2(7) and 3(6) positions of the phenanthrene ring were highly mutagenic. The relationship between chemical structure and the mutagenicity of NPh derivatives is discussed.

Mutagenic synergy between paraoxon and plant-activatedm-phenylenediamine or 2-acetoxyacetylaminofluorene

Paraoxon (diethyl-p-nitrophenylphosphate) is the toxic, but non-mutagenic metabolite of the organophosphorus 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).

Mutagenicity of 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid treated with nitrite in the presence of alcohols

The mutagenicity of a product produced from 1-mehtyl,1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid (MTCCA), which is a component in soy sauce, after treatment with 50 mM nitrite at pH 3, 37°C, for 60 min in the presence of 7.5% ethanol was much higher than that in the absence of ethanol during the nitrite treatment. The enhancement of the mutagenicity of nitrite-treated MTCCA by ethanol required simultaneous treatment of MTCCA with nitrite and ethanol. The mutagenicity of MTCCA treated with nitrite in the presence and absence of ethanol was detected in the same fractions on HPLC and was highest for Salmonella typhimurium strain YG1029 possessing elevated O-acetyltransferase activity among the several Salmonella test strains, suggesting that the same mutagen belonging to aromatic compounds was produced both in the presence and absence of ethanol. Methanol, n-propanol and isopropanol as well as ethanol were also observed to have an augmenting effect. However, the sugars glucose and sucrose had no effect. When MTCCA was treated with nitrite in the presence of commercial alcoholic beverages equivalent to 1.25–10% ethanol, Japanese ‘sake’ and ‘shochu’ were demonstrated to have a highly augmenting effect and beer, wine, whisky and brandy to have a mildly augmenting effect.

Mutagenic activation of aromatic amines by a human hepatoma cell (Hep G2) supernatant tested by means of Salmonella typhimurium strains with different acetyltransferase activities

The study was carried out to characterize hepatoma cells (Hep G2) as activation system relevant to man and to investigate which are the tester strains most suitable for the mutagenic assay of aromatic amines. A supernatant prepared from the human hepatoma cell line Hep G2 was used to activate benzidine, 2-aminofluorene (2-AF) and 2-acetylaminofluorene (2-AAF) in the Salmonella typhimurium reversion assay. Activation by Hep G2 supernatant was studied with increasing concentrations of the three compounds, in tester strains TA98, YG1024, DJ400 and DJ460. Benz[ a]anthracene (BA) pretreatment of cells increases the mutagenicity of benzidine in strains YG1024, DJ460 and DJ400. Activation of 2-AAF and 2-AF was observed in strains YG1024, DJ400 and, at the highest tested dose, in DJ460. These results were compared with those obtained with S9 from control and Aroclor 1254 (Aro)-pretreated rat liver. With strain TA98 comparable responses were obtained except for 2-AF which was better activated using rat liver S9. The use of strain YG1024 greatly increases the sensitivity of the response. Strain DJ460 makes it possible to detect activation of 2-AF and 2-AAF by Aro-induced rat liver. These results indicate that Hep G2 supernatant is a useful metabolic activation system of human origin that can be used to replace rat liver S9. An appropriate choice of the Salmonella strain not only can increase the sensitivity of the response, but may also help to overcome certain metabolic shortcomings of the Hep G2 cell line and rat liver S9.

Distribution of nitropyrenes and mutagenicity in airborne particulates collected with an Andersen sampler

Distributions of 1,3-dinitropyrene (1,3-DNP), 1,6-DNP, 1,8-DNP, 1-nitropyrene (1-NP) and mutagenicity in airborne particulates collected in downtown Kanazawa, Japan with an Andersen high-volume air sampler were examined. Mutagenicities of benzene-ethanol extract from particulates were determined by the Ames test using S. typhimurium strains with S9 mix, while concentrations of DNPs and 1-NP were determined by high-performance liquid chromatography (HPLC) using chemiluminescence detection. In the finest particulate fraction (smaller than 1.1 μm), 68% and 75% of the total mutagenicities were observed in TA98 and YG1024 strains, respectively. In the same fraction, 65–82% of three DNPs as well as 84% of 1-NP were observed. Mutagenic contributions of 1,3-DNP, 1,6-DNP, 1,8-DNP and 1-NP in the extract were respectively 0.6, 1.2, 1.8 and 1.6% in the TA98 stain, and 2.5, 5, 9 and 2.1% in the YG1024 strain.

A simple technique for monitoring mutagenicity and benzo(a)pyrene content in mussels and their ambient water

AbstractBy hanging blue rayon, an adsorbent specific for chemicals with polycyclic aromatic structures, in seawater for one day, the mutagenicity and the benzo(a)pyrene pollution of the water were monitored. Mussels living in the sites of water sampling were also subjected to the blue rayon extraction process and assayed. The mutagenicity was evaluated by the Salmonella/microsome reversion assay of Ames, with the use of a recently developed sensitive strain YG1024. The monitoring was performed in the Seto Inland Sea at 5 sites near Okayama, in the year 1991–1992. All the water and mussel samples gave positive benzo(a)pyrene contents and many of them showed positive mutagenicity (as assayed with metabolic activation). The mutagenicity and the benzo(a)pyrene contents were found to be significantly correlated both for the water samples and for the mussel samples. In contrast, there were no correlations between the water and the mussel samples regarding either the mutagenicity or benzo(a)pyrene contents. This lack of correlations may be attributed to the fact that the water samples gave results for a one‐day exposure whereas the mussel samples gave those of a long‐term exposure. This simple technique has advantages over the conventional sampling method in terms of easy handling, high sensitivity, and feasibility of monitoring distant sites. © by John Wiley &amp; Sons, Inc.

Characterization of mutagenic activity in instant hot beverage powders.

Extracts of several grain-based coffee-substitute blends and instant coffees were mutagenic in the Ames/Salmonella test using TA98, YG1024, and YG1029 with metabolic activation. The beverage powders induced 150 to 500 TA98 and 1,150 to 4,050 YG1024 revertant colonies/g, respectively. Increased sensitivity was achieved using strain YG1024. No mutagenic activity was found in instant hot cocoa products. The mutagenic activity in the beverage powders was shown to be stable to heat and the products varied in resistance to acid nitrite treatment. Differential bacterial strain specificity, and a requirement for metabolic activation suggest that aromatic amines are present. Characterization of the mutagenic activity, using HPLC and the Ames test of the collected fractions, showed the coffee-substitute blends and instant coffees contain several mutagenic compounds. Known heterocyclic amines are not responsible for the major part of the mutagenic activity. The main mutagenic activity in grain-based coffee-substitute blends and instant coffees is due to several unidentified compounds, which are most likely aromatic amines.

Bacterial mutagenicity of cigarette smoke and its interaction with ethanol

The mutagenicities of mainstream cigarette smoke (CS), a cigarette smoke condensate (CSC) and smokers' urines were investigated by using batteries of Salmonella typhimurium and Escherichia coli strains. The S9-mediated mutagenicity of CSC was remarkably enhanced when using nitroreductase- and especially O-acetyltransferase-overproducing derivatives of the classical strains TA98 and TA100, with the following rank of sensitivity: YG1024 > YG1029 > YG1021 > TA98NR > YG1026 > TA98 > TA100 > TA100-DNP6 > TA100NR > TA98-1,8-DNP6. With YG1024, a doubling of spontaneous revertants was observed with as little as 1/110 of the smoke condensate recovered from one cigarette under our experimental conditions. Similarly, the S9-mediated mutagenicity of mainstream CS was considerably increased in YG1024 and YG1029, the O-acetyltransferase-overproducing derivatives of TA98 and TA100, respectively. In the absence of S9 mix, the concentrates of 23 urine specimens from five smokers failed to revert S. typhimurium TA98 and YG1024, and were equitoxic in E. coli WP2 and its repair-deficient counterpart CM871 (uvrA-, recA-, lexA-). In the presence of S9 mix, all specimens were mutagenic, with an average YG1024:TA98 ratio of 6.6:1. These patterns suggest that the bacterial mutagenicity of smoke-associated complex mixtures is mainly due to aromatic amines. The mutagenicities of other typical constituents of CS, i.e. the polycyclic aromatic hydrocarbon benzo[a]pyrene (BP) and the tobacco specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), were not appreciably enhanced in the O-acetyltransferase-overproducing strain YG1029, compared to its parental strain TA100. Moreover, BP and NNK induced less than additive mutagenic responses when combined at high doses.(ABSTRACT TRUNCATED AT 250 WORDS)

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 micrograms/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-derived particle-associated mutagens.

Biotransformation of genotoxic agents in marine sponges. Mechanisms and modulation

Marine sponges do not appear to suffer from neoplastic diseases, in spite of possible high exposures resulting from their nature as sessile bottom filter feeders which pump large volumes of sea water. The assessment of several parameters related to the biotransformation of mutagens/carcinogens showed that the metabolic machinery of sponge medulla cells is mainly oriented towards detoxification, with some differences depending on species (Geodia cydonium or Tethya aurantium). Glutathione (GSH) levels were unexpectedly high in these cells, especially in Geodia, in which the concentration of this tripeptide was more than twice that measured in liver preparations from untreated rats, at least when related to the protein content. The oxidoreductive enzyme activities involved in the glutathione cycle were balanced in such a way as to favour a high GSH: oxidized glutathione (GSSG) ratio. GSH S-transferase activity was conversely rather low, compared to that of rat liver, and the dehydrogenases involved in the hexose monophosphate shunt were high in Tethya but low in Geodia. The metabolism of mutagens was investigated by using the Salmonella typhimurium his- strains TA100, TA98 and YG1024. Sponge S12 fractions failed to activate aflatoxin B1, benzo[a]pyrene and the two heterocyclic amines 3-amino-1-methyl-5H-pyrido[4,3-b]indole and 2-amino-3,4-dimethyl-imidazo[4,5-f]quinoline. Although far less efficiently than untreated rat liver S12 fractions, Geodia and especially Tethya preparations weakly activated the three aromatic amines 2-acetyl-aminofluorene, 2-aminofluorene and 2-aminoanthracene. On the other hand, sponge S12 fractions were remarkably efficient in decreasing the mutagenic potency of the direct-acting mutagens 4-nitroquinoline 1-oxide and sodium dichromate.(ABSTRACT TRUNCATED AT 250 WORDS)

Genotoxicity of m-phenylenediamine and 2-aminofluorene in Salmonella typhimurium and human lymphocytes with and without plant activation.

The promutagenic arylamines, m-phenylenediamine (mPDA) and 2-aminofluorene (2-AF), were evaluated for their genotoxicity in Salmonella typhimurium strain YG1024 and in human lymphocytes. These agents were assayed with and without TX1MX plant activation mix. Both arylamines without activation were refractory in S. typhimurium, demonstrating that plant activation was required for the generation of their ultimate mutagenic metabolites. However, using the alkaline single-cell gel/Comet assay, both mPDA and 2-AF directly induced DNA damage in human lymphocytes. This effect was reduced when the human cells were treated with the arylamine plus TX1MX. mPDA with or without plant activation was not toxic to the exposed cells. However, at concentrations over 80 microM, 2-AF was toxic to lymphocytes. This toxic response was eliminated by incubation with TX1MX. mPDA and 2-AF were plant-activated into mutagens for S. typhimurium. However, these plant-activated products had a reduced genotoxic potency in human lymphocytes.