Levels Of Mutagenicity Research Articles

The characterization of the mutagenic activity of soil

The solvent extracts of non-agricultural soil were shown to cause a potent mutagenic response to Salmonella bacteria. The extract of soil from local sites had activities of 100 000–250 000 TA98 revertants per kg with S9 activation and 44 000–83 000 revertants per kg to TA100 with activation. The soil is mutagenic to both strains without S9, but at lower levels. Nitroreductase-deficient bacterial strains show that the soil extract may have mutagenic compound(s) containing a nitro group. The revertant numbers per gram of soil are lowest at the surface, increase at 4–10 cm and decrease in a linear fashion to a depth of 0.5 meter. HPLC fractionation shows that there is more than one separable component responsible for the mutagenicity of the soil. The source and consequences of the high levels of mutagenicity in ‘ordinary soil’ are unknown. Soil mutagens can be a significant complicating factor to those using the Ames/Salmonella assay for environmental sampling.

Electronic factors and acridine frameshift mutagenicity — a pattern recognition study

Using the ADAPT and CHEMLAB-II systems for structure-activity analysis, computer-calculated electronic properties of molecules were used to derive structure-activity relationships for predicting the mutagenicity of a set of substituted acridines in strain TA1537 of the Ames Salmonella assay. A collection of 40 acridines, with a variety of substituents, was examined. A set of 4 electronic desctriptors was found which could be used to correctly classify all but two of the compounds as mutagenic or nonmutagenic. A negative correlation was found between the sum of the Hammett aromatic substituent parameters and the level of mutagenicity of the structures, expressed as log(number of revertants/plate + 1) at a 20-μg dose. This correlation, however, was not high enough to allow precise estimation of the mutagenicity values.

Mutagen levels in urine from snuff users, cigarette smokers and non tobacco users — A comparison

The mutagenic activity of concentrates of urine from snuff users, cigarette smokers and non tobacco users has been investigated. A concentration procedure involving use of Sep-Pak C 18 columns and elution with methylene chloride was used. The concentrates were assayed for mutagenicity towards strain TA98 of Salmonella typhimurium, both in the presence and absence of a metabolic activation system, the postmitochondrial liver fraction (S9) from Aroclor 1254 induced rats. The mean mutagenic activity of smokers' urine concentrates was 8.6 × 10 3 revertants per 24 h and significantly higher than the corresponding values for snuff users, abstinent snuff users and non tobacco users, which were (1.3, 1.3 and 0.9) × 10 3, respectively. No significant difference in mutagenic activity was found between urine from snuff users, whether using or abstaining from snuff, and urine from non tobacco users. It could thus be concluded that the level of urinary mutagens, isolated by adsorption on Sep-Pak C 18 columns, is not elevated by habitual usage of Swedish wet snuff.

Interactions of chromium with nitrilotriacetic acid (NTA) in the induction of genetic effects in bacteria†

The genotoxic activity of NTA, which is a substitute for polyphosphates in household laundry detergents, as well as the interactions between NTA and soluble (potassium dichromate) and insoluble (lead chromate) Cr(VI) compounds in the induction of gene mutations and DNA damage in bacteria were investigated. NTA did not induce point mutations in S. typhimurium TA100 strain, independently of the presence of rat liver metabolic activation, nor did it modify the direct mutagenicity of potassium dichromate. Insoluble lead chromate was instead clearly mutagenic in the Salmonella/microsome assay only in the presence of NTA, and the levels of mutagenicity correlated very well with the amounts of Cr(VI) solubilized by NTA in the form of chromate ion, as determined by the colorimetric reaction with diphenylcarbazide. In a fluctuation test using E. coli WP2 uvrA, NTA was not mutagenic, but it significantly increased the mutagenicity of potassium dichromate besides that of lead chromate. Conversely, as assessed by means of three different procedures, NTA consistently yielded an increased lethality in repair‐deficient E. coli strains of the WP2 series, various genetic mechanisms being involved in repairing the specific DNA damage. Moreover, mixture of NTA with insoluble lead chromate resulted in the release of genotoxic Cr(VI) ion. Finally, NTA was negative in the SOS chromotest, evaluating the induction of SOS DNA repair mechanisms in E. coli PQ37. These results confirm the ability of NTA to elicit the genotoxicity of insoluble heavy metals in bacteria, indicate that also the interaction of NTA with soluble metals can result in a moderate increase of some genetic effects, and point to a possible direct genotoxicity of NTA itself, at least as far as the induction of DNA damage is concerned.

Mutagenicity of human urine after the consumption of fried salted salmon

Mutagenicity in the urine of four non-smoking individuals who had eaten salted salmon cooked at home for both lunch and supper was monitored by means of Salmonella/microsome mutagenicity tests. Extracts from fresh and salted salmon had the same level of mutagenicity after being cooked for 10 min at 200°C, but no activity was detected before cooking. Salmonella strains TA98 and TA1538 were equally sensitive to the mutagens and required metabolic activation. No mutagenicity was shown with TA100 and TA1535. Urine samples were tested using a concentrate prepared by means of an XAD-2 resin column. Mutagenicity was detected mainly in urine excreted during 4–5 hr after the ingestion of cooked salmon, but only weak mutagenicity, or none at all, was detected in the urine after the ingestion of vegetables. The levels of urinary mutagenicity due to salmon consumption were not affected when cabbage was eaten simultaneously. The excretion of mutagenic substances was completed within about 20 hr, and there were almost no mutagens in the urine 24 hr after the ingestion of cooked salmon.

Mutagenicity of airborne particles related to meteorological and air pollution parameters

Many studies indicate that aerosols in the ambient atmosphere may have mutagenic properties. Most of these investigations were conducted in industrial or urban areas or near specific sources. The purpose of our studies was to assess the variation in mutagenicity of airborne particulate matter using the Salmonella/microsome test, over a prolonged period of time. In addition to 30 samples collected simultaneously at Wageningen (rural location) and Terschelling (background location), a further 95 samples were collected in Wageningen. At both locations mutagenicity varied markedly with time. Our studies indicate that the level of mutagenicity follows a yearly cycle, with the highest activity being found in winter. In winters of different years prevalent mutagenicity differed considerably. Mutagenicity was positively correlated between locations, pointing to large-scale processes and not to local sources. In the present study a relation between wind direction, air trajectories and mutagenicity was established. Differences within a short period of time could be explained by the trajectory of the sampled air mass. Furthermore, mutagenic activity was correlated with commonly registered air pollution components; multiple regression shows that the air pollution parameters SO 2, NO 2, NO, CO and O 3 together account for 70% of the variation in direct mutagenicity and 80% of the variation in indirect mutagenicity. SO 2 and NO 2, and SO 2, NO 2 and CO were significantly associated with the variation in direct and indirect mutagenicity.

Ｆｌｕｃｔｕａｔｉｏｎ ｔｅｓｔによるエポキシ樹脂硬化剤の変異原性についての検討

Mutagenicity of nine epoxy resin hardeners was examined by a fluctuation test modified by Gatehouse. The test was performed by using Salmonella typhimurium TA98 with a metabolic activation system. In our laboratory, the results of the fluctuation test were compared with the results obtained by the previously mentioned Ames preincubation method. Six out of nine epoxy resin hardeners showed mutagenic activity in both the fluctuation test and Ames preincubation method, but one out of the nine was negative in both test systems. Two out of the nine were positive by either of the two testing systems. The fluctuation test is disadvantageous in that it is marginally slower and requires slightly more labor than the Ames test and furthermore it is difficult to increase the amount of microsome because of background interference. These disadvantages, however, are somewhat offset by the advantages that small organs such as urinary bladder can be used instead of liver cells and that a small amount of microsome can be employed for metabolic activation. This test is also suitable for testing aqueous samples containing low levels of mutagen.

Mutagen formation during commercial processing of foods.

Levels of bacterial mutagenicity 3-17 times above spontaneous are generated during commercial thermal processing (canning) of foods, particularly foods high in protein. The potential for other processing operations, including pasteurization, dehydration, and concentration, to produce substances active in the Ames Salmonella assay was also examined. Two heated fish model systems, canned salmon and fried sole, were established by extracting mutagen precursors from fish tissues with water. The model system studies suggest that the limiting reactants for mutagen formation differ from one food product to another, and that Maillard type browning reactions are involved in mutagen production. Bisulfite treatment was found to inhibit mutagen formation in modal systems and whole food products. Isolation and partial characterization of the mutagens in both fried and canned pink salmon showed that at least three distinct mutagens were present. These mutagens exhibited HPLC retention time patterns on C18, cyano, and amino columns different than the major mutagens present in other cooked and grilled meats and fish.

Activation and detoxication of promutagens by toadfish ( Opsanus tau) hepatic postmitochondrial fractions in the Salmonella assay

Three groups of experiments were conducted to characterize the hepatic postmitochondrial fraction (S9) from the oyster toadfish ( Opsanus tau) as an activation system for promutagens in the Salmonella assay and to provide an initial evaluation of the extent to which data from standard in vitro assays with mammalian activation systems are predictive of possible genotoxic effects in this marine fish. In the first group of experiments the effects of increasing the concentration of S9 from untreated and 3-methylcholanthrene (MC)- or Aroclor 1254 (AC)-pretreated toadfish and Sprague-Dawley rats on the mutagenicities of different concentrations of 2-aminoanthracene (2AA) and benzo[ a]pyrene (BAP) were examined in Salmonella (TA98) plate assays. The maximum levels of 2AA mutagenicity attained by S9 from untreated (UI S9) toadfish and rats were comparable, but UI S9 from toadfish was moreeffective than UI S9 from rats in mediating BAP mutagenicity. MC pretreatment decreased maximum levels of 2AA mutagenicity and increased maximum levels of BAP mutagenicity mediated by S9 from both species. MC pretreatment also altered the pattern of dependence of 2AA mutagenicity on the concentration of S9 protein for S9 from both species. A similar alteration in the pattern of dependence of BAP mutagenicity on the concentration of S9 protein was also observed with S9 from MC-pretreated toadfish. Although AC pretreatment of rats effected changes in the mutagenicities of both test chemicals similar to those effected by MC pretreatment, AC pretreatment of toadfish effected little or no change in the mutagenicities of either test chemical. The changes in the pattern of dependence of 2AA and BAP mutagenicities on the concentration of S9 protein effected by MC pretreatment of toadfish were confirmed in a separate group of experiments. A third group of experiments was designed to examine the effects of α-naphthoflavone (ANF) on the mutagenicities of 2AA and BAP mediated by UI and MC S9 from toadfish. Although ANF did not affect the 2AA mutagenicity mediated by UI S9, a significant decrease in 2AA mutagenicity and a significant increase in BAP mutagenicity mediated by MC S9 and a significant decrease in BAP mutagenicity mediated by UI S9 were observed. These results indicate that 2AA and BAP are effectively activated by toadfish S9 and that, as in rats, these two test chemicals are activated and/or detoxicated by different cytochrome P-450-dependent pathways. These results also support the contention that cytochrome P-450-dependent detoxication pathways can be an important determinant of the mutagenic potency of some promutagens in vitro.

Mutagenicity of benzylic acetates, sulfates and bromides of polycyclic aromatic hydrocarbons

Studies were performed to determine the direct mutagenicity of the acetates and some bromides and sulfates of hydroxymethyl polycyclic aromatic hydrocarbons in S. typhimurium strains TA98 and TA100. Benzylic acetates, bromides and sulfates were synthesized and characterized. The compounds tested were benzyl alcohol, 5-hydroxymethylchrysene, 1-hydroxymethylpyrene, 6-hydroxymethylbenzo[a]pyrene, 6-(2-hydroxyethyl)benzo[a]pyrene, 6-hydroxymethylanthanthrene, 9-hydroxymethylanthracene, 9-hydroxymethyl-10-methylanthracene, 7-hydroxymethylbenz[a]anthracene, 7-(2-hydroxyethyl)benz[a]anthracene, 12-hydroxymethylbenz[a]anthracene, 7-hydroxymethyl-12-methylbenz[a]anthracene, 12-hydroxymethyl-7-methylbenz[a]anthracene, 1-hydroxy-3-methylcholanthrene, 2-hydroxy-3-methylcholanthrene, 3-hydroxy-3, 4-dihydrocyclopental[cd]pyrene and 4-hydroxy-3, 4-dihydrocyclopental[cd]pyrene. The benzylic sulfate esters of 6-hydroxymethylbenzo[a]pyrene and 7-hydroxymethylbenz[a]anthracene were the most mutagenic compounds, whereas the aliphatic sulfate ester of 7-hydroxyethylbenz[a]anthracene did not cause an increase in mutations above background. All meso-anthracenic benzylic acetate esters were mutagenic in both strains with various degrees of activity, whereas the corresponding non-benzylic esters were inactive, as expected. Of the non-meso-benzylic acetate esters, only the 3-acetoxy-3, 4-dihydrocyclopenta[cd]pyrene was mutagenic. In the benzylic bromide series, only the eight mesoanthracenic were mutagenic, whereas benzyl bromide and 5-bromomethylchrysene were inactive. The aliphatic bromides, 6-(2-bromoethyl)benzo[a]pyrene and 7-(2-bromoethyl)benz[a]anthracene did not display significant activity. The potencies of the acetate esters more accurately reflect the mutagenicity because the rate of solvolysis did not compete with the reactivity of the esters with bacterial DNA. In the case of benzylic sulfates and bromides, the rate of solvolysis was very rapid and could have diminished the level of mutagenicity, depending on the assay conditions. These results demonstrate that meso-anthracenic benzylic acetates, sulfates and bromides are mutagenic, whereas benzylic acetate esters attached to other carbon atoms are inactive.

Mutagens in human urine: effects of cigarette smoking and diet

Human urine from smokers and nonsmokers on strictly controlled diets was assayed for mutagenic activity. Two distinct diets were employed in this study. Diet study A consisted of a high-meat, high-fat diet, observed for 5 days, followed by a vegan diet, adhered to for the next 5 days. The vegan diet contained no meat, fish, eggs, or dairy products. It was comprised of soy products, prepackaged vegan dinners, seeds, nuts, fruits, vegetables, beans and herbal teas. Diet study B consisted of 3 days on a typical western diet followed by a macrobiotic diet of grains and fresh vegetables for 5 days. Portions of 24-h urine samples were assayed in Salmonella typhimurium TA1538. The levels of urinary creatinine and cotinine were measured. Mutagenic activity was observed in the urine of most smokers. However, the levels of mutagens in the urine of light smokers were similar to those of nonsmokers. For both nonsmokers and smokers there was a significant increase in urine mutagenicity when volunteers were on the vegan diet. Several nonsmokers on the vegan diet in diet study A had pronounced mutagenic activity in their urine samples, in some instances at higher levels than that in the urine of smokers on a meat diet. In diet study B no clear differences were observed between the meat diet and the macrobiotic diet. In diet studies A and B the mutagenic potency of smokers' urine could not be correlated with cotinine levels alone or with urinary pH. These data suggest that dietary factors can play a dominant role in the mutagenicity of urine concentrates.

The problem of safe levels of environmental mutagens with regard to dose-effect dependence

The problem of safe levels of environmental mutagens with regard to dose-effect dependence

Identification and quantification of highly mutagenic nitroacetoxypyrenes and nitrohydroxypyrenes in diesel-exhaust particles

Heavy-duty diesel-exhaust particles were collected, extracted and fractionated into diethyl ether-soluble neutral, acidic and basic fractions. The mutagenicity of these fractions was measured with Salmonella typhimurium strains TA100, TA98, TA98NR and TA98/1,8-DNP 6 in the presence and absence of a 9000 × g post-mitochondrial supernatant from Aroclor-induced rat liver (S9 mix). The neutral and acidic fractions showed high mutagenicity with TA98 in the absence of S9 mix, the acidic fraction having the highest specific activity. In the absence of S9 mix, the mutagenicity of crude, neutral and acidic fractions was greater in TA98 than in TA98NR and TA98/1,8-DNP 6. Chemically-synthesized nitroacetoxypyrenes and nitrohydroxypyrenes were fractionated into the neutral and acidic fractions, respectively. These nitroarenes were purified by high-performance liquid chromatography and their mutagenicity was measured with the 4 strains. With TA98 in the absence of S9 mix, 1-nitro-3-acetoxypyrene, 1-nitro-6/8-acetoxypyrene, 1-nitro-3-hydroxypyrene, 1-nitro-6/8-hydroxypyrene induced 16 700, 336, 992, 94 His + revertants per plate per nmole, respectively. In the absence of S9 mix, the level of mutagenicity of these nitroarenes was highest in TA98, lowest in TA98/1,8-DNP 6 and intermediate in TA98NR. The neutral and acidic fractions of diesel-exhaust particles were analyzed by gas chromatography-mass spectrometry and gas chromatography-mass fragmentography. The neutral fraction was found to contain nitroacetoxypyrenes, 1-nitropyrene, 1,6-dinitropyrene, while nitrohydroxypyrenes were detected in the acidic fraction. The amounts of 1-nitro-3-acetoxypyrene, 1-nitropyrene, 1,6-dinitropyrene and 1-nitro-3-hydroxypyrene were 6.3, 62, 0.81, and 70 ng per mg of crude extract, and accounted for 12, 3.6, 8.0, and 9.0%, respectively, of mutagenicity of the crude extract in TA98 in the absence of S9 mix.

Identification of mutagenic compounds formed during chlorination of humic acid

Humic acid chlorination products are being studied in an effort to identify the chemicals responsible for the mutagenicity formed during water chlorination. In the present report, 19 chlorinated organic compounds have been identified and quantified in ether extracts of chlorinated humic acid solutions. 10 of these compounds, including a number of chlorinated propanones and chlorinated propenals, are direct-acting mutagens in the Salmonella/microsome mutagenicity assay. The position of the chlorine substituent has been found to be an important factor in the mutagenic activity of these two classes of compounds. The total mutagenicity of the compounds identified thus far, when tested either individually or as a composite, accounts for only 7–8% of the total TA100 mutagenicity, and less than 2% of the TA98 mutagenicity formed during humic acid chlorination. The addition of bromide to the humic acid chlorination reaction results in up to a 2-fold increase in the level of mutagenicity formed.

DNA-damaging activity in ethanol-soluble fractions of feces from New Zealand groups at varying risks of colorectal cancer.

Using repair-proficient and repair-deficient strains of E. coli, we investigated the application of a liquid incubation assay to measure the DNA-damaging activity of ethanol-soluble fecal extracts. This method appears to be suitable for the study of a wide range of sample types. It was used to measure the DNA-modifying activity of ethanol-soluble fecal extracts from a group of European colorectal cancer patients. Data were compared with those from Europeans of similar age and sex distribution who did not have bowel cancer. We also studied groups of Maoris, Samoans, and European Seventh-Day Adventists who followed an ovo-lacto vegetarian diet. There are significant levels of DNA-modifying materials in the feces of many Europeans on a mixed diet, regardless of whether or not they have cancer. The number of positive samples was less in the Polynesian groups, and there were no samples that could be unequivocally scored as positive in the Seventh-Day Adventist groups. We conclude that diet can significantly reduce the level of ethanol-soluble mutagens, at least in New Zealand Europeans. The data may provide an explanation for the reduced incidence of bowel cancer in Seventh-Day Adventist groups.

Role of tumor promotion in affecting the multi-hit nature of carcinogenesis.

The assessment of risk from low-level exposure to radiation and chemicals is hindered by the basic lack of scientific understanding of the complex nature of the multiple levels of protective or synergistic interactions. Radiation and chemicals have the potential of inducing mutations, cell death and altered gene expression. The biological consequences of each of these effects is again complex, since many factors can enhance or mask the effect of a single mutation, cytotoxic or gene modulatory change. Carcinogenesis, representing but one chronic disease state, is a multi-step process, involving the clonal expansion of a single altered cell (that is, the initiation event). Radiation and chemicals appear to contribute to the initiation process by their ability to induce viable mutations. Insufficient theoretical and empirical knowledge precludes a determination of whether mutagenesis and, hence, initiation exhibit a threshold phenomenon. Because of a variety of redundancy mechanisms on the genetic and cellular levels, the physiological impact of a single dysfunctional cell is felt only after it is amplified to a large number (the promotion phase of carcinogenesis). Radiation and chemical-induced cytotoxicity, as well as noncytotoxic chemical induction of mitogenesis, can induce surviving single dysfunctional stem cells to multiply. If during this multiplication of dysfunctional cells, the initiated cells are further exposed chronically to low levels of mutagens, there is an enhanced probability of additional genetic changes. The accumulation of foci of dysfunctional cells can occur in any stem cell population of any tissue. As with mutagens, the existence of threshold levels for promoting conditions and chemicals is still not yet scientifically validated. However, specific examples of the actions of a few promoters does seem to be consistent with that idea. The concepts of initiation and promotion imply the existence of anti-initiation and antipromotion conditions. Together with genetic factors, the complex and unpredictable interactions of radiation and chemicals as initiators, anti-initiators, promoters and antipromoters, defy prospects of an easy means to predict the consequences of exposure to chronic low levels of radiation and chemicals.

Mutagenicity in gastric juice.

Mutagenicity has been measured in the gastric juice of 228 patients using the Ames bacteriological test system; while mutagenicity in control and duodenal ulcer patients did not differ from saline controls, mutagenicity was significantly increased compared with controls in patients suffering gastric ulcer (p less than 0.002), carcinoma (p less than 0.002), and in patients after gastric resection (p less than 0.01). A transient rise in mutagenicity was seen following H2 antagonist ingestion (p less than 0.002). Increased levels of mutagenicity were found to correlate closely with gastric juice pH and bacterial count. Histidine concentrations in gastric juice did not explain the mutagenicity results.

Mutagen content of table wines made from various grape species and hybrid cultivars

The mutagen content of wines produced from the European Vitis vinifera grapes was compared with that of wines produced from hybrids of V. vinifera and species indigenous to North America. Mutagens were extracted on an XAD-2 Amberlite resin column and activated with S-9 and/or faecalase in the Salmonella/microsomal mutagen assay. All white wines had insignificant mutagen levels. The only red wine to produce statistically significant reversion frequencies was that made from the Concord grape. The mutagens were shown to be extracted from the grape skins during fermentation.

Mutagens in Foods

SummaryThe application of short‐term tests, particularly those involving the use of bacteria, has demonstrated the presence in foods of several hundred chemicals with mutagenic activity. The pyrolysis of proteins (heating above 30(fC) and the reaction at lower temperatures between carbohydrates and amino‐acids has been shown to result in the formation of a range of potent mutagens. Despite the fact that substances identified as mutagens in short‐term tests are usually carcinogens in animal feeding tests, the value of short‐term tests for predicting human health hazards remains problematical. Foods contain a variety of substances that protect against the effects of some carcinogens and a rational perspective is required when attempting to assess the risk associated with the low levels of mutagens in foods.

OCCUPATIONAL EXPOSURE TO CYCLOPHOSPHAMIDE

Several lines of research have demonstrated the risk of occupational exposure of healthcare personnel to anticancer drugs. Urine samples from two nurses working in a cancer clinic were analysed for cyclophosphamide (CP) by gas chromatography after they had prepared the drug for treatment. Since the drug might be absorbed through the skin, urine samples from five volunteers were also examined after a solution of CP had been applied topically to the cubital fossa area. Variable quantities of intact CP were identified in samples from the volunteers collected over 24 h, but in most cases the drug was evident only in urine samples given more than 6 h after application. CP was found in several urine samples from the nurses, but the quantities of CP in these samples were not related to the amounts of drug handled. The identity of CP was confirmed by gas-chromatography/mass-spectrometry. CP appeared sooner in the urine samples from the nurses than in those from the volunteers, suggesting a faster route of absorption, perhaps through inhalation of aerosols generated during dissolution of the drug. These findings suggest that the increased levels of mutagenicity observed in urine samples of nurses who worked on oncology wards might have arisen, in part, from metabolites of CP.