A review of the mutagenicity and rodent carcinogenicity of ambient air

Although ambient air was first shown to be carcinogenic in 1947 and mutagenic in 1975, no overarching review of the subsequent literature has been produced. Recently, Claxton et al. [L.D. Claxton, P.P. Matthews, S.H. Warren, The genotoxicity of ambient outdoor air, a review: Salmonella mutagenicity, Mutat. Res./Rev. Mutat. Res. 567 (2004) 347-399] reviewed the literature on the mutagenicity of urban air in the Salmonella mutagenicity assay. Here, we review the literature on the mutagenicity of urban air in other test systems and review the carcinogenicity of urban air in experimental systems. Urban air was carcinogenic in most of the reports involving rodents. Studies ascribed carcinogenic activity primarily to PAHs, nitroarenes, and other aromatic compounds. Atmospheric conditions, along with the levels and types of pollutants, contributed to the variations in carcinogenic and mutagenic activity of air from different metropolitan areas. The majority of the mutagenesis literature was in the Salmonella assay (50%), with plant systems accounting for most of the rest (31%). The present data give little support to the use of plant systems to compare air mutagenicity among multiple sites or studies. Studies in mice have shown that particulate air pollution causes germ-cell mutations. Air sheds contain similar types and classes of mutagens; however, the levels of these compounds vary considerably among air sheds. Combustion emissions were associated with much of the mutagenicity and carcinogenicity of urban air. Most studies focused on the particulate fraction; thus, additional work is needed on the volatile and semi-volatile fractions, metals, and atmospheric transformation. Smaller particles have greater percentages of extractable organic material and are more mutagenic than larger particles. Although hundreds of genotoxic compounds have been identified in ambient air, only a few (<25) are routinely monitored, emphasizing the value of coupling bioassay with chemistry in the monitoring of air for carcinogenic and mutagenic activities and compounds.     

Genotoxicity and mutagenicity of chromium(VI)/ascorbate-generated DNA adducts in human and bacterial cells

Reduction of carcinogenic Cr(VI) by vitamin C generates ascorbate-Cr(III)-DNA cross-links, binary Cr(III)-DNA adducts, and can potentially cause oxidative DNA damage by intermediate reaction products. Here, we examined the mutational spectrum and the importance of different forms of DNA damage in genotoxicity and mutagenicity of Cr(VI) activated by physiological concentrations of ascorbate. Reduction of Cr(VI) led to a dose-dependent formation of both mutagenic and replication-blocking DNA lesions as detected by propagation of the pSP189 plasmids in human fibroblasts. Disruption of Cr-DNA binding abolished mutagenic responses and normalized the yield of replicated plasmids, indicating that Cr-DNA adducts were responsible for both mutagenicity and genotoxicity of Cr(VI). The absence of DNA breaks and abasic sites confirmed the lack of a significant production of hydroxyl radicals and Cr(V)-peroxo complexes in Cr(VI)-ascorbate reactions. Ascorbate-Cr(III)-DNA cross-links were much more mutagenic than smaller Cr(III)-DNA adducts and accounted for more than 90% of Cr(VI) mutagenicity. Ternary adducts were also several times more potent in the inhibition of replication than binary complexes. The Cr(VI)-induced mutational spectrum consisted of an approximately equal number of deletions and G/C-targeted point mutations (51% G/C --> T/A and 30% G/C --> A/T). In Escherichia coli cells, Cr(VI)-induced DNA adducts were only highly genotoxic but not mutagenic under either normal or SOS-induced conditions. Lower toxicity and high mutagenicity of ascorbate-Cr(III)-DNA adducts in human cells may result from the recruitment of an error-prone bypass DNA polymerase(s) to the stalled replication forks. Our results suggest that phosphotriester-type DNA adducts could play a more important role in human than bacterial mutagenesis.     

International Commission for Protection against Environmental Mutagens and Carcinogens. ICPEMC working paper No. 5. Genotoxicity tests as predictors of carcinogens: an analysis

Differences between the results of numerical validation studies comparing in vitro and in vivo genotoxicity tests with the rodent cancer bioassay are leading to the perception that short-term tests predict carcinogenicity only with uncertainty. Consideration of factors such as the pharmacokinetic distribution of chemicals, the systems available for metabolic activation and detoxification, the ability of the active metabolite to move from the site of production to the target DNA, and the potential for expression of the induced lesions, strongly suggests that the disparate sensitivity of the different test systems is a major reason why numerical validation is not more successful. Furthermore, genotoxicity tests should be expected to detect only a subset of carcinogens, namely genotoxic carcinogens, rather than those carcinogens that appear to act by non-genetic mechanisms. Instead of relying primarily on short-term in vitro genotoxicity tests to predict carcinogenic activity, these tests should be used in a manner that emphasizes the accurate determination of mutagenicity or clastogenicity. It must then be determined whether the mutagenic activity is further expressed as carcinogenicity in the appropriate studies using test animals. The prospects for quantitative extrapolation of in vitro or in vivo genotoxicity test results to carcinogenicity requires a much more precise understanding of the critical molecular events in both processes.     

Flavonoids and alkenylbenzenes: mechanisms of mutagenic action and carcinogenic risk

The present review focuses on the mechanisms of mutagenic action and the carcinogenic risk of two categories of botanical ingredients, namely the flavonoids with quercetin as an important bioactive representative, and the alkenylbenzenes, namely safrole, methyleugenol and estragole. For quercetin a metabolic pathway for activation to DNA-reactive species may include enzymatic and/or chemical oxidation of quercetin to quercetin ortho-quinone, followed by isomerisation of the ortho-quinone to quinone methides. These quinone methides are suggested to be the active alkylating DNA-reactive intermediates. Recent results have demonstrated the formation of quercetin DNA adducts in exposed cells in vitro. The question that remains to be answered is why these genotoxic characteristics of quercetin are not reflected by carcinogenicity. This might in part be related to the transient nature of quercetin quinone methide adducts, and suggests that stability and/or repair of DNA adducts may need increased attention in in vitro genotoxicity studies. Thus, in vitro mutagenicity studies should put more emphasis on the transient nature of the DNA adducts responsible for the mutagenicity in vitro, since this transient nature of the formed DNA adducts may play an essential role in whether the genotoxicity observed in vitro will have any impact in vivo. For alkenylbenzenes the ultimate electrophilic and carcinogenic metabolites are the carbocations formed upon degradation of their 1'-sulfooxy derivatives, so bioactivation of the alkenylbenzenes to their ultimate carcinogens requires the involvement of cytochromes P450 and sulfotransferases. Identification of the cytochrome P450 isoenzymes involved in bioactivation of the alkenylbenzenes identifies the groups within the population possibly at increased risk, due to life style factors or genetic polymorphisms leading to rapid metaboliser phenotypes. Furthermore, toxicokinetics for conversion of the alkenylbenzenes to their carcinogenic metabolites and kinetics for repair of the DNA adducts formed provide other important aspects that have to be taken into account in the high to low dose risk extrapolation in the risk assessment on alkenylbenzenes. Altogether the present review stresses that species differences and mechanistic data have to be taken into account and that new mechanism- and toxicokinetic-based methods and models are required for cancer risk extrapolation from high dose experimental animal data to low dose carcinogenic risks for man.     

Review of the genotoxicity of nitrogen oxides

Nitrogen oxides (NO_x) are formed in combustion processes and are major pollutants in urban air. Relatively few studies on the genotoxicity of NO₂ and NO have been performed. These studies indicate that NO₂ is genotoxic in vitro, but the effect of NO seems to be very slight.One in vivo study showed chromosome aberrations and mutations in lung cells after inhalation of NO₂ (and NO), but tests for chromosome aberrations in lymphocytes and spermatocytes or micronuclei in bone marrow were negative after inhalation of NO₂. Based on present studies, there is no clear evidence of a carcinogenic potential of NO₂, although lung adenomas were induced in the susceptible strain A/J mouse.The primary metabolites of NO_x are nitrite and nitrate. Nitrate seems to be devoid of genotoxic properties, but nitrite is genotoxic in vitro, and there are also positive in vivo results. Cancer studies have been mainly negative. However, carcinogenic nitrosamines have been shown to be formed in vivo after inhalation of NO₂.Nitrogen oxides are key components in atomospheric smog formation, which may lead to secondary effects. Strongly mutagenic nitro-PAH compounds are easily formed, and mutagenic reaction products may be formed photochemically from alkenes.     

Genotoxic potency of monofunctional alkylating agents in E. coli: comparison with carcinogenic potency in rodents

A quantitative correlation between carcinogenicity and genotoxicity was investigated by a comparison between the carcinogenic potency in rodents and the mutagenic (M), recombinogenic (R) and SOS-inducing (I) potencies in a bacterial test (E. coli multitest) for 9 monofunctional alkylating agents: N-nitroso-N-methylurethane, N-nitroso-N-ethylurea, epichlorohydrin, N-nitroso-N-methylurea, N-nitroso-N-methyl-N'-nitroguanidine, methyl methanesulfonate, diethylsulfate, dimethylsulfate, ethyl methanesulfonate. A significant positive correlation between the carcinogenic potency and the product of the mutagenic and recombinogenic potencies was found for all tested compounds. Thus, the E. coli multitest may be used as a simple test to search for correlations between carcinogenicity and genotoxicity of DNA-damaging agents.     

Quantitative comparative mutagenesis in bacteria, mammalian cells, and animal-mediated assays A convenient way of estimating genotoxic activity in vivo?

The accumulation of environmental compounds which exhibit genotoxic properties in short-term assays and the increasing lag of time for obtaining confirmation or not in long-term animal mutagenicity and carcinogenicity tests, makes it necessary to develop alternative, rapid methodologies for estimating genotoxic activity in vivo. In the experimental approach used here, it was assumed that the genotoxic activity of foreign compounds in animals, and ultimately humans, is determined among others by exposure level, organ distribution of (DNA) dose, and genotoxic potency per unit of dose, and that knowledge about these 3 parameters may allow to rapidly determine the expected degree of genotoxicity in various organs of exposed animals. In view of the high degree of qualitative correlation between mutagenic activity of chemicals in bacteria and in cultured mammalian cells, and their mutagenic and carcinogenic properties in animals, and in order to be able to distinguish whether mutagenic potency differences were due to differences in (DNA) dose rather than other physiological factors, the results of mutagenicity tests obtained in the present experiments using bacteria and mammalian cells were compared on the basis of DNA dose rather than exposure concentrations, with the following questions in mind: Is there an absolute or a relative correlation between the mutagenic potencies of various ethylating agents in bacteria (E. coli K12) and in mammalian cells (V79 Chinese hamster) after treatment in standardized experiments, and can specific DNA adducts be made responsible for mutagenicity? Is the order of mutagenic potency of various ethylating agents observed in bacteria in vitro representative of the ranking of mutagenic potency found in vivo? Since the answer to this last question was negative, a further question addressed to was whether short-term in vivo assays could be developed for a rapid determination of the presence (and persistence) of genotoxic factors in various organs of mice treated with chemicals. In quantitative comparative mutagenesis experiments using E. coli K12 and Chinese hamster cells treated under standardized conditions in vitro with 5 ethylating agents, there was no indication of an absolute correlation between the number of induced mutants per unit of dose in the bacteria and the mammalian cells. The ranking of mutagenic potency was, however, identical in bacteria and mammalian cells, namely, ENNG greater than ENU greater than or equal to DES greater than DEN congruent to EMS, the mutagenic activity of DEN being dependent on the presence of mammalian liver preparations.(ABSTRACT TRUNCATED AT 400 WORDS)     

Genotoxicity and genotoxic enhancing effect of tetrandrine in Salmonella typhimurium

Tetrandrine has been used for the treatment of silicosis in China. The potential genotoxic and carcinogenic hazards of this drug were studied using the Salmonella/histidine reversion assay and the SOS/Umu test. The results show that tetrandrine was weakly mutagenic to Salmonella typhimurium TA98 with metabolic activation and did not induce SOS response. However, tetrandrine increased the mutagenic activity of benzo[alpha]pyrene, trinitrofluorenone (TNF), 2-aminoanthracene (2AA), diesel emission particles, airborne particles, and cigarette smoke condensate by more than 100%; the activity of aflatoxin B1 and fried beef was increased by over 75%. It also increased the 2AA and TNF-induced SOS response by more than 300%. These results indicated that tetrandrine was a weak promutagen inducing frameshift mutations and was a potent genotoxic enhancer. The mechanism for the genotoxic enhancement is not known. However, the fact that the increase in mutagenicity was noted only in TA98 and not in TA1538 suggested that the enhancement of genotoxicity by tetrandrine may result from an increase in error-prone DNA repair.     

Genotoxicity of aniline derivatives in various short-term tests

Various substituted aniline derivatives were tested for genotoxicity in several short-term tests in order to examine the hypothesis that a substitution at both ortho positions (2,6-disubstitution) could prevent genotoxicity due to steric hindrance of an enzymatic activation to electrophilic intermediates. In the Salmonella/microsome assay, 2,6-dialkylsubstituted anilines and 2,4,6-trimethylaniline (2,4,6-TMA) were weakly mutagenic in strain TA100 when 20% S9 mix was used, although effects were small compared to those of 2,4-dimethylaniline and 2,4,5-trimethylaniline (2,4,5-TMA). In Drosophila melanogaster, however, 2,4,6-TMA and 2,4,6-trichloroaniline (TCA) were mutagenic in the wing spot test at 2-3 times lower doses than 2,4,5-TMA. In the 6-thioguanine resistance test in cultured fibroblasts, 2,4,6-TMA was again mutagenic at lower doses than 2,4,5-TMA. Two methylene-bis-aniline derivatives were also tested with the above methods: 4,4'-methylene-bis-(2-chloroaniline) (MOCA) was moderately genotoxic in all 3 test systems whereas 4,4'-methylene-bis-(2-ethyl-6-methylaniline) (MMEA) showed no genotoxicity at all. DNA binding studies in rats, however, revealed that both MOCA and MMEA produced DNA adducts in the liver at levels typically found for moderately strong genotoxic carcinogens. These results indicate that the predictive value of the in vitro test systems and particularly the Salmonella/microsome assay is inadequate to detect genotoxicity in aromatic amines. Genotoxicity seems to be a general property of aniline derivatives and does not seem to be greatly influenced by substitution at both ortho positions.     

Chromosomal aberrations and micronuclei induced in rat and mouse bone marrow cells by sodium nitrate

The genotoxicity of several anthraquinone compounds metabolically related to aflatoxin B1 was examined by means of the hepatocyte primary culture (HPC)/DNA repair test and the Salmonella microsome mutagenesis test, and compared to versicolorins A and B which are potent mutagenic and genotoxic intermediates of the aflatoxin biosynthetic pathway. 6,8-O-Dimethyl-versicolorins A, B and 6-deoxyversicolorin A were found to be strongly mutagenic and genotoxic. Genotoxicity of versicolorin A and 6,8-O-dimethylversicolorin A was stronger than that of versicolorin B and 6,8-O-dimethylversicolorin B, respectively, in the HPC/DNA repair test. Nidurufin and norsolorinic acid, which do not possess a bisfuran ring, exhibited questionable activities for mutagenicity and no genotoxicity. It is suspected that 6,8-O-dimethylversicolorins A, B and 6-deoxyversicolorin A as well as versicolorins A and B are genotoxic carcinogens.     

Cytotoxic and genotoxic effects of energetic compounds on bacterial and mammalian cells in vitro.

The mutagenicity and toxicity of energetic compounds such as 2,4, 6-trinitrotoluene (TNT), 1,3,5-trinitrobenzene (TNB), hexahydro-1,3, 5-trinitro-1,3,5-triazine (RDX) and octahydro-1,3,5,7-tetranitro-1,3, 5,7-tetrazocine (HMX), and of amino/nitro derivatives of toluene were investigated in vitro. Mutagenicity was evaluated with the Salmonella fluctuation test (FT) and the V79 Chinese hamster lung cell mutagenicity assay. Cytotoxicity was evaluated using V79 and TK6 human lymphoblastic cells. For the TK6 and V79 assays, TNB and 2, 4,6-triaminotoluene were more toxic than TNT, whereas RDX and HMX were without effect at their maximal aqueous solubility limits. The primary TNT metabolites (2-amino-4,6-dinitrotoluene, 4-amino-2, 6-dinitrotoluene, 2,4-diamino-6-nitrotoluene and 2, 6-diamino-4-nitrotoluene) were generally less cytotoxic than the parent compound. The FT results indicated that TNB, TNT and all the tested primary TNT metabolites were mutagenic. Except for the cases of 4-amino-2,6-dinitrotoluene and 2,4-diamino-6-nitrotoluene in the TA98 strain, addition of rat liver S9 resulted in either no effect, or decreased activity. None of the tested compounds were mutagenic for the V79 mammalian cells with or without S9 metabolic activation. Thus, the FT assay was more sensitive to the genotoxic effects of energetic compounds than was the V79 test, suggesting that the FT might be a better screening tool for the presence of these explosives. The lack of mutagenicity of pure substances for V79 cells under the conditions used in this study does not preclude that genotoxicity could actually exist in other mammalian cells. In view of earlier reports and this study, mutagenicity testing of environmental samples should be considered as part of the hazard assessment of sites contaminated by TNT and related products.     

Assessment of the potential germ cell mutagenicity of industrial and plant protection chemicals as part of an integrated study of genotoxicity in vitro and in vivo

An approach is described that enables the germ cell mutagenicity of chemicals to be assessed as part of an integrated assessment of genotoxic potential. It is recommended, first, that the genotoxicity of a chemical be defined by appropriate studies in vitro. This should involve use of the Salmonella mutation assay and an assay for the induction of chromosomal aberrations, but supplementary assays may be indicated in specific instances. If negative results are obtained from these 2 tests there is no need for the conduct of additional tests. Agents considered to be genotoxic in vitro should then be assessed for genotoxicity to rodents. This will usually involve the conduct of a bone marrow cytogenetic assay, and in the case of negative results, a genotoxicity test in an independent tissue. Agents found to be non-genotoxic in vivo are regarded as having no potential for germ cell mutagenicity. Agents found to be genotoxic in vivo may either be assumed to have potential as germ cell mutagens, or their status in this respect may be defined by appropriate germ cell mutagenicity studies. The basis of the approach, which is supported by the available experimental data, is that germ cell mutagens will be evident as somatic cell genotoxins in vivo, and that these will be detected as genotoxins in vitro given appropriate experimentation. The conduct of appropriate and adequate studies is suggested to be of more value than the conduct of a rigid set of prescribed tests.     

Review of the genotoxicity of styrene in humans

Styrene (CAS No. 100-42-5) is an important industrial chemical for which positive results have been reported in in vitro and in vivo genotoxicity assays. Styrene-exposed workers have been studied extensively over two decades for the induction of various types of genotoxic effects. The outcomes of these studies have been conflicting, and where positive responses have been reported, it has proved difficult to demonstrate clear relationships between levels of damage reported and exposure levels. In this review, we have assessed studies addressing mutagenicity (chromosome aberrations, micronuclei and gene mutations) and other endpoints (sister chromatid exchanges, DNA breaks and DNA adducts) using criteria derived from the IPCS guidelines for the conduct of human biomonitoring studies. Based on the re-evaluated outcomes, the data are not convincing that styrene induces gene mutations. The evidence for induction of clastogenicity in occupationally exposed workers is less clear, with a predominant lack of induction of micronuclei in different studies, but conflicting responses in chromosome aberration assays. The results of numerous studies on sister chromatid exchanges do not provide evidence of a clear positive response, despite these being induced in animals exposed to styrene at high concentrations. However, there is evidence that both DNA adducts and DNA single strand breaks are induced in styrene workers. These types of damage are considered indicative of exposure of the target cells and interaction with cellular DNA but do not necessarily result in heritable changes. There is evidence that the metabolism of styrene in humans is affected by genetic polymorphisms of metabolizing genes and that these polymorphisms affect the outcome of in vitro mutagenicity studies on styrene. Therefore, studies that have addressed the potential of this factor to affect in vivo responses were considered. To date, there are no consistent relationships between genetic polymorphisms and induction of genotoxicity by styrene in humans, but further work is warranted on larger samples. The analyses of individual studies, together with a consideration of dose-response relationships and the lack of a common profile of positive responses for the various endpoints in different studies, provide no clear evidence that styrene exposure in workers results in detectable levels of mutagenic damage. However, evidence of exposure to genotoxic metabolites is demonstrated by the formation of DNA adducts and strand breaks.     

Evaluation of genotoxity and mutagenicity of DL-p-chlorophenylalanine, its methyl ester and some N-acyl derivatives

DL-p-chlorophenylalanine (PCPA) and its derivatives were evaluated for genotoxic effects using Escherichia coli and Bacillus subtilis strains lacking various DNA-repair mechanisms in spottest and in suspension test. The mutagenic activity of studied compounds was determined by the Ames test. Reverse mutation test was performed with Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 without S9 mix. 0.02 M nitrosomethylurea (NMU) standard mutagen was used as a positive control. The results showed that the parent nonessential amino acid PCPA had no detectable genotoxic and mutagenic activities in bacteria. The methyl ester of this amino acid and its N-phenylacetyl derivative possessed weak genotoxicity. Meanwhile N-sec-butyloxycarbonyl, N-benzyloxycarbonyl, N-(p-nitrophenylacetyl) and N-(p-nitrophenoxyacetyl) derivatives of DL-p-chlorophenylalanine exhibited appreciable genotoxicity. Among the seven tested compounds only N-benzyloxycarbonyl and N-(p-nitrophenoxyacetyl) derivatives of DL-p-chlorophenylalanine have been found to be mutagenic. Only parent PCPA possessed antimutagenic properties in respect of nitrosomethylurea. The structural modification, which strongly affects genotoxicity and mutagenicity perhaps may be due to steric hydrance of the substituents, causing interference with enzyme and DNA interactions.     

The pathogen reduction treatment of platelets with S-59 HCl (Amotosalen) plus ultraviolet A light: genotoxicity profile and hazard assessment

Despite restrictive donor criteria and screening procedures, infections resulting from the transfusion of bacterially contaminated platelet products continue to occur. Pathogen reduction technologies targeting nucleic acids have been developed. However, concerns about the safety of these procedures exist; the main concern being the possible mutagenic and carcinogenic effects of the pathogen-inactivated preparation in the recipient. This report reviews the genotoxicity profile of the S-59 (Amotosalen) plus long wavelength ultraviolet light (UVA) pathogen reduction technology, and assesses the mutagenic and carcinogenic hazards in recipients of treated platelets. S-59, a synthetic heterocyclic psoralen, non-covalently intercalates into the nucleic acids of pathogens and forms crosslinks when UVA photoactivated. Before clinical use, the levels of residual S-59 and free photoproducts are greatly reduced using a 'compound adsorption device' (CAD). In vitro, S-59 is mutagenic in Salmonella typhimurium and mouse lymphoma L5178Y TK(+/-) cells, and is clastogenic in CHO cells. There is reduced activity (Salmonella, CHO cells) or no activity (mouse lymphoma cells) with metabolic activation (S9 mix). When tested up to toxic dose levels, S-59 was negative in the mouse bone marrow micronucleus assay and the rat hepatocyte unscheduled DNA synthesis (UDS) test. Based on comparative studies conducted with S-59 plus UVA-treated platelets (up to 25 times without CAD), any genotoxic effects can be attributed to residual S-59. Considering (1) the known genotoxic mechanism of action for S-59, (2) the negative in vivo studies for S-59 at multiples >40,000x over clinical peak plasma levels, and (3) the fact that the positive in vitro genotoxicity effects for the end product seem due to residual S-59, any mutagenic hazard to a recipient of S-59 plus UVA-treated platelets is negligible and there is no concern about a carcinogenic potential as a consequence of a mutagenic activity. This conclusion is supported by a negative p53(+/-) mouse carcinogenicity study.     

Activation of mutagens by hepatocytes and liver 9000 X g supernatant from human origin in the Salmonella typhimurium mutagenicity assay. Comparison with rat liver preparations

The mutagenicity of 10 known genotoxic compounds, of several chemical classes, was measured in Salmonella typhimurium mutagenicity assays comprising isolated human hepatocytes or human liver 9000 X g supernatant (S9) from 4 different individuals, as activating system. The mutagenic activity of several compounds as determined with the Salmonella/hepatocyte suspension assay showed obvious differences when compared with the values obtained in the Salmonella/S9 plate assay. For instance, the mutagenic activity of BZ, DMN and DEN appeared to be much higher in the hepatocyte assay than in the S9 assay. However, 2-AF and 2-AAF were activated more effectively into mutagens in the S9 assay than in the hepatocyte assay. 2-AF was slightly more mutagenic than 2-AAF in the hepatocyte assay, whereas it was far more mutagenic than 2-AAF in the S9 assay. DMN was found more mutagenic than DEN in the hepatocyte assay, whereas in the S9 assay DEN appeared to be slightly more mutagenic. Furthermore, great interindividual differences in the metabolic activation of certain compounds, e.g. BZ and DMN, were observed in the hepatocyte suspension assay, whereas these variations were less evident in the S9 plate assay. Comparison of the mutagenicity data obtained with the human liver preparations, with those obtained with rat liver preparations, showed great interspecies differences in the capacity to activate certain chemicals into mutagens. The use of human liver preparations, in particular isolated human hepatocytes, may be of great value in studies on inter- and intraspecies variations in metabolic activation of genotoxic agents.     

Mutagenicity of stereochemical configurations of 1,2-epoxybutene and 1,2:3,4-diepoxybutane in human lymphblastoid cells

The carcinogenicity of 1,3-butadiene (BD) is related to its bioactivation to several DNA-reactive metabolites; accumulating evidence suggests that the stereochemistry of these BD intermediates may play a significant role in the mutagenic and carcinogenic actions of the parent compound. The objective of this study was to evaluate the cytotoxicity and mutagenicity of stereochemical forms of 1,2-epoxybutene (EB) and 1,2:3,4-diepoxybutane (DEB), two genotoxic BD metabolites, in a human lymphoblastoid cell line, TK6. Cytotoxicity was measured by comparing cloning efficiencies in chemical-exposed cells versus those in control cells. The hypoxanthine-guanine phosphoribosyltransferase (HPRT) and thymidine kinase (TK) mutant frequencies (MFs) were measured using a cell cloning assay. HPRT mutants collected from cells exposed to the three forms of DEB were analyzed by PCR to characterize large genetic alterations. All the three stereoisomers of DEB caused increased HPRT and TK MFs compared to the concurrent control samples. There were no significant differences in cytotoxicity or mutagenicity among the three isomers of DEB in TK6 cells. Molecular analysis of HPRT mutants revealed similar distributions of types of mutations among the three isomers of DEB. There were also no statistically significant differences in mutagenic efficiencies between the two isomers of EB in TK6 cells. These results were consistent with the in vivo findings that there was little difference in the mutagenic efficiencies of racemic-DEB versus meso-DEB in rodents. Thus, in terms of mutagenic efficiency, stereochemical configurations of EB and DEB are not likely to play a significant role in the mutagenicity and carcinogenicity of BD.     

Comparative genotoxicity of nitrosamine drinking water disinfection byproducts in Salmonella and mammalian cells

Nitrosamine water disinfection byproducts (DBPs) are an emerging class of non-halogenated, nitrogen-containing water contaminants. Five nitrosamine DBPs were analyzed for genotoxicity (N-nitrosodimethylamine (NDMA), N-nitrosopiperidine (NPIP), N-nitrosopyrrolidine (NPYR), N-nitrosomorpholine (NMOR) and N-nitrosodiphenylamine (NDPhA). Using Salmonella typhimurium strain YG7108 the descending rank order of mutagenicity was NDMA>NPIP>NMOR>NPYR; NDPhA was not mutagenic. We developed and calibrated an exogenous S9 mix that was highly effective in activating NDMA in Chinese hamster ovary (CHO) cells using the SCGE (Comet) assay. The descending rank order for genotoxicity was NDMA>NPIP>NMOR. NDPhA was genotoxic only at one concentration and NPYR was not genotoxic. The genotoxic potencies in S. typhimurium and CHO cells were highly correlated. Based on their comparative genotoxicity attention should be focused on the generation and occurrence of NDMA, NPIP and NMOR. Current drinking water disinfection processes may need to be modified such that the generation of nitrosamine DBPs is effectively limited in order to protect the environment and the public health.     

Rec effect of certain textile dyes in Bacillus subtilis

A large number of compounds are toxic, genotoxic, mutagenic, teratogenic and/or carcinogenic. The genotoxicity of four textile dyes commonly used in India namely Sulphur Red Brown 360 (SRB), Jade Green 2G (JG), Reactofix Turquoise Blue 5GFL (RTB) and Direct Scarlet 4BS (DS) was determined by Bacillus subtilis spore Rec assay, both in the presence and absence of metabolizing activation mixture (S9 mix). Each dye was toxic at higher dose levels. A dose-dependent increase in the depth of growth inhibition zones was observed for all dyes. Zones of inhibition were usually clearer at higher doses of the dyes and with Rec- bacteria, but were translucent with Rec+ bacteria. SRB and DS were toxic to Rec+ and Rec- bacteria. JG was less genotoxic in the absence of S9 mix, however, its genotoxic potential increased in the presence of S9 mix. Reactofix T blue was more genotoxic in the absence of S9 mixture.