Direct-acting mutagenic properties of some hair dyes used in New Zealand

Mutagenicity or carcinogenicity data are not publicly available on many hair dyes or dye components commonly used within New Zealand. Representative mid- to dark-warm brown hair dyes of 12 brands supplying more than 1% of the New Zealand market were tested for direct-acting mutagenicity using the bacterial 'Ames' test. Despite recent scientific advances in the development of non-mutagenic dyes, 23 of the 40 products tested gave positive results in one or both of the tester strains used. There appeared to be differences between distributors in the proportion of their hair dyes which were mutagenic. In the case of 6 out of 10 of the above dyes which had tested positive, in vitro mutagenicity or toxicity was enhanced in the presence of verapamil, suggesting that risks from hair-dye exposure may change in the case of persons using this or similar drugs. It is recognised that there are uncertainties regarding human risks from dyes which are shown to be mutagenic in in vitro tests. However, from the above results, it seems possible to produce non-mutagenic hair dyes in this color range; and in the interests of public reassurance, it may be prudent to require that such dyes be used.     

Mutagenic action of dimethyl sulfate and N-nitrosoethylurea on the mulberry silkworm

Mutagenic effect of chemical mutagens, dimethalsulphate (DMS) and N-nitrosoethyl urea (NEU), on silkworm is studied. The effect of these mutagens have never been studied on silkworm before. DMS is found to be completely insufficient as mutagen for the silkworm, because its low mutability is accompanied with strong cytotoxic effect. NEU appeared to be of "milder" effect. Injection of male-moth proved to be very efficient. The majority of exuded mutants showed mosaic colouring of the graine.     

Bioassay-directed fractionation of mutagenic PAH atmospheric photooxidation products and ambient particulate extracts.

Simulated atmospheric gas-phase reactions of naphthalene, fluorene and phenanthrene have been carried out in an environmental chamber with bioassay-directed chemical analysis of the reaction products. Nitro-PAH were found to be the most significant mutagens formed from the reactions of naphthalene and fluorene. The mutagram (bar graph of mutagenic activity versus HPLC fraction) of the phenanthrene reaction products closely resembled that of an ambient air particulate extract with the most mutagenic activity being in a fraction more polar than that in which the nitro-PAH elute. Nitrophenanthrene lactones (nitro-6H-dibenzo[b,d]pyran-6-ones) were found to account for the observed activity of this polar fraction of the phenanthrene reaction products. It has been shown that the utilization of an environmental chamber with a known PAH-starting material and the ability to produce sufficient product for isomer-specific identifications of mutagens is a promising complement to bioassay-directed fractionation of ambient air particulate extracts.     

Mutagenesis induced with dioxidin in the Allium-test

The influence of dioxidin in different concentrations (10-100 mg/l) on the cytogenetic parameters of Allium cepa L. has been studied. The mutagenic effect of dioxidin was shown within all the range of the studied concentrations. The curve "dose-effect" has been determined for the concentrations of 10, 20, 30, 40, 50, and 100 mg/l. The peak of the mutagenic effect and significant reduction of the mitotic index were revealed at the concentration of 100 mg/l. It was shown that the mutagenic efficiency of the dioxidin statistically correlated with reduction of the mitotic activities.     

Testing of some azo dyes and their reduction products for mutagenicity using Salmonella typhimurium TA 1538

A series of ten azo dyes as well as various single ring aromatic amines substituted on the benzene ring were tested for bacterial mutagenicity with Salmonella typhimurium TA 1538 using a soft-agar overlay method. Two dyes, sudan 2 and chrysoidin induced mutation but only in the presence of a rat liver preparation. Chrysoidin was the more active. Testing of its reduction products, aniline and 1,2,4-triaminobenzene showed a liver metabolite of the latter compound could be responsible for the mutagenic effect, having a comparable mutagenicity with 1,2-diamino-4-nitro-benzene, one of the mutagenic constituents of hair dyes. Structure-activity studies on a series of ring-substituted anilines indicated that mutagenic activity required at least two positions to be substituted with either amino or nitro groups, or one of each. The bacteria as well as the liver enzyme preparation may partake in the activation of these chemicals. The correlation between mutagenicity and carcinogenicity for this group of compounds is discussed.     

Mutagenic activity of 4 active-principle forms of pharmaceutical drugs. Comparative study in the Salmonella typhimurium microsome test, and the HGPRT and Na+/K+ ATPase systems in cultured mammalian cells

The nitroimidazole derivatives used in human therapy have shown a strong mutagenic activity in bacterial tests using Ames strains of Salmonella typhimurium. Our study compared the response of 4 products of this family on bacterial target cells as well as on mammalian target cells (Chinese hamster V79 cells). The strong positive response on TA100 was greatly reduced on the nitroreductase-deficient strain TA100 Frl. Furthermore, no mutagenic activity was found in V79 mammalian cells that we examined for ouabain and 6-thioguanine resistance.     

Effect of albumin on frequency of spontaneous mutations in somatic mammalian cells in vitro

Human albumin is able to induce statistically significant increasing of the frequency of spontaneous mutations in the hprt locus in cultured somatic mammalian cells. The strait dependence of the mutagenic effect on the protein concentration has been shown. The products of albumin degradation reveal antimutagenic activity in the investigated test-system.     

Assessing the potential of a "color effect" for hair analysis of 11-nor-9-carboxy-delta(9)-tetrahydrocannabinol: analysis of a large sample of hair specimens

It has been hypothesized that hair color may play a role in the concentration of various drugs of abuse in hair. Several studies have shown that melanin in hair appears to play a binding role for at least some commonly abused drugs. However, these studies have been limited by a number of factors when assessing the clinical significance of a hypothesized melanin or color effect. This study evaluates the possible effect of hair color on the concentration of 11-nor-9-carboxy-Delta(9)-tetrahydrocannabinol (c-THC) in human hair. The analysis is based on 3886 positive c-THC hair specimens drawn from a universe of approximately 80000 specimens of scalp hair harvested from the posterior vertex of the head. Analysis of variance of color categorization by c-THC concentration shows that c-THC concentration does not have a significant association with hair color (Hair Color F = 1.148, p =.332) and therefore does not have a demonstrable "color effect".     

Bioassay-directed fractionation of mutagenic PAH atmospheric photooxidation products and ambient particulate extracts

Simulated atmospheric gas-phase reactions of naphthalene, fluorene and phenanthrene have been carried out in an environmental chamber with bioassay-directed chemical analysis of the reaction products. Nitro-PAH were found to be the most significant mutagens formed from the reactions of naphthalene and fluorene. The mutagram (bar graph of mutagenic activity versus HPLC fraction) of the phenanthrene reaction products closely resembled that of an ambient air particulate extract with the most mutagenic activity being in a fraction more polar than that in which the nitro-PAH elute. Nitrophenanthrene lactones (nitro-6H-dibenzo[b,d]pyran-6-ones) were found to account for the observed activity of this polar fraction of the phenanthrene reaction products. It has been shown that the utilization of an environmental chamber with a known PAH-starting material and the ability to produce sufficient product for isomer-specific identifications of mutagens is a promising complement to bioassaydirected fractionation of ambient air particulate extracts.     

The mutagenic potential of madder root in dyeing processes in the textile industry

The roots of Rubia tinctorum L. have a long tradition of being used in dyeing processes of textiles from centuries ago until the present time. The colouring principles belong to the class of hydroxyanthraquinones. Concern arose because several of these compounds were recognised as mutagenic in vitro and even carcinogenic in rodents. To assess the possible risk to humans caused by coloured textiles, mutagenicity was investigated with two madder root samples of different origin (Iran and Bhutan) along the entire dyeing process from root extracts to the dyed wool. The Salmonella/microsome test (Ames assay) with the strains TA98, TA100 and TA1537 was used. Significant mutagenic effects could be detected in madder root extracts and also in the final product, the dyed wool. Madder root from Iran showed considerably higher mutagenic responses than samples from Bhutan. Analytical investigations of the extracts by HPLC showed the presence of a spectrum of anthraquinones typical for madder root. Three mutagenic compounds, lucidine, rubiadine and purpuroxanthine, together with the non-mutagenic alizarine could be detected. The mutagenic response of the different samples was positively correlated with the concentration of the mutagenic anthraquinones, and with lucidine in particular. Based on these investigations a risk to dye-house workers and users of textiles dyed with R. tinctorum must be anticipated.     

A rise of mutation level in tissues of monkey goby and sturgeon fry under conditions of block by antibodies to the serotonin-modulated anticonsolidation protein

The work presents data on studies of effect of a decrease in the serotoninergic system activity on the level of mutagenic changes (the micronuclear test) in the goby Neogobius fluviatilis and the fry of sturgeon Acipenser güldenstädti persicus. It has been shown that the long exposure of the animals to conditions of industrial and oil pollution leads to a significant decrease in their liver of the level of serotonin-modulated anticonsolidation protein (SMAP) correlating directly with the serotonin level as well as to a sharp rise of the level of micronuclear in erythrocytes. The intramuscular administration of anti-SMAP polyclonal antibodies to the sturgeon fry produces a significant increase of the amount of micronuclear as compared with that in the animals injected with non-immune γ-globulin. The obtained results allow concluding that the decrease of activity of the serotoninergic system is the mechanism that is triggered with adverse environmental factors and realizes mutagenic damages in the modified genetic apparatus.     

Mutagenicity of hexachlorobutadiene, perchlorobutenoic acid and perchlorobutenoic acid chloride

Hexachloro(1,3)butadiene (HCBD) is a well known environmental contaminant. The nephrocarcinogenic potential of HCBD has been shown in long-term studies with rats. Experiments were performed to assist in determining whether this effect is mediated by epigenetic or genotoxic mechanisms and to compare the mutagenic properties of HCBD with those of its monooxidation products, perchloro-3-butenoic acid (PCBA) and perchloro-3-butenoic acid chloride (PCBAC), which are conceivable metabolites of HCBD. All 3 compounds are mutagenic to the Salmonella typhimurium tester strain TA100. The mutagenic effect is dose-dependent and parallels the chemical reactivity of the compounds. HCBD is only mutagenic in the presence of drug-metabolizing enzymes (S9 mix) with an increased protein content. The mutagenic response after incubation with PCBAC and PCBA is 2-3-fold that of HCBD. Additionally, both PCBAC and PCBA exert a mutagenic response in the absence of S9 mix. The experiments support the assumption of a genotoxic potential of HCBD.     

Mutagenic activity of amino acid pyrolyzates in Salmonella typhimurium TA 98.

Pyrolyzates of 25 amino acids and 5 indole derivatives were tested for mutagenicity in the histidine-requiring mutant Salmonella typhimurium TA 98. Significant mutagenic activity was detected with pyrolyzates of most of the amino acids. These pyrolyzates required a liver microsomal fraction, as representative of mammalian metabolism, to be detected as mutagens. Among the pyrolyzates tested, the highest mutagenic activity was observed with that of L-tryptophan. As little as 10 microgram of the pyrolyzate of L-tryptophan had detectable mutagenic activity toward TA 98. The optimal pyrolysis temperatures for the formation of mutagenic products were shown to be 500 degrees C for L-tryptophan and 600 degrees C for the other amino acids. The results from pyrolyses of some indole derivatives suggest that an amino group at the alpha-position to the carboxyl group of L-tryptophan plays an important role in the formation of mutagens.     

Synthesis and structure--mutagenicity relationship of benzo-annulated cyclopentaphenanthrenes.

The synthesis of 2,3-dihydro-1H-indeno[5,4-a]anthracene (2), the fluoreno[a]anthracenes 3 and 4, 2,3-dihydro-1H-cyclopenta[a]chrysene (6), 3,4-dihydro-2-vinylphenanthrene (10) and cyclopenta[c]chrysenes 11, 12 has been described. Structure analysis of the new products by (1)H and (13)C NMR spectroscopy is presented. Estimates of the mutagenic activity of compounds 2--4, 6 and 11--14 in Salmonella typhimurium determined by Ames' test indicate that all products are inactive for both TA 98 and TA 100 strains except 4,5-dihydro-3H-cyclopenta[c]chrysene (12). The mutagenic properties of these compounds have been compared with those shown by previously studied benzo[g]cyclopenta[a]phenanthrenes and cyclopenta[c]phenanthrenes and discussed. Some conclusions have been drawn about the effects of benzoannulation and of the carbonyl function on the mutagenicity of this class of compounds.     

DNA sequence of the Rhizobium leguminosarum nodulation genes nodAB and C required for root hair curling

A 3.2kb fragment of DNA cloned from Rhizobium leguminosarum has been shown to contain the genes necessary for the induction of root hair curling, the first observed step in the infection of leguminous plants by R. leguminosarum. The DNA sequence of this region has been determined and three open reading frames were identified: genes corresponding to these open reading frames have been called nodA, nodB and nodC and are transcribed in that order. Mutations within the nodC gene completely blocked root hair curling. However, a subcloned fragment containing only the nodC gene did not induce normal root hair curling (although some branching was observed), indicating that the nodA and B genes may also be required for normal root hair curling. From an analysis of the predicted amino acid sequences of the nodAB and C genes it appeared unlikely that their products are secreted; therefore it is concluded that the induction of root hair curling could be due to a secreted metabolite.     

Mutagenicity of four hair dyes in Drosophila melanogaster.

The hair dye constituents p-phenylenediamine, 2,4-diaminoanisole sulfate, 2,4-diaminotoluene and 4-nitro-0-phenylenediamine were tested for mutagenicity in Drosophila melanogaster. The compounds were given orally to adult males. The induction of sex-linked recessive lethal mutation was used as a measure of mutagenicity. All four of the dyes tested were mutagenic with a peak mutagenic activity in metabolically active germ cells (spermatids and spermatocytes).     

Mutagenic activity of south Indian food items.

Dietary components and food dishes commonly consumed in South India were screened for their mutagenic activity. Kesari powder, calamus oil, palm drink, toddy and Kewra essence were found to be strongly mutagenic; garlic, palm oil, arrack, onion and pyrolysed portions of bread toast, chicory powder were weakly mutagenic, while tamarind and turmeric were not. Certain salted, sundried and oil fried food items were also mutagenic. Cissus quadrangularis was mutagenic, while 'decoctions' of cumin seeds, aniseeds and ginger were not. Several perfumes, essential oils and colouring agents, which are commonly used were also screened and many of them exhibited their mutagenic potential by inducing the 'reverse mutation' in Salmonella typhimurium tester strains.     

Mutagenicity of benzyl chloride in the Salmonella/microsome mutagenesis assay depends on exposure conditions

Benzyl chloride (BCl) is a clear yellowish, volatile liquid that is widely used as an intermediate for the production of benzyl alcohol and benzyl compounds used in perfumery, dyes and pharmaceuticals. In previous studies BCl has shown weak and inconsistent mutagenicity in the Salmonella/microsome mutagenesis assay (Ames test). The aim of the present study was to investigate the potential mutagenic activity of BCl using modifications of the standard Ames test in order to adapt the method to the volatile nature of the test compound. Tests were performed using (a) the standard plate-incorporation method, (b) incubation of the treated plates in closed containers, (c) a vaporization-diffusion method to expose Ames test plates to volatilised BCl and (d) the pre-incubation method. Using the standard plate-incorporation method, BCl showed no (in the absence of metabolic activation) or very weak (in the presence of metabolic activation) mutagenic activity in Salmonella typhimurium tester strain TA 100. The use of the pre-incubation method did not improve detection of mutagenic activity of BCl. The use of closed containers significantly increased the response, but the most marked response was obtained by testing BCl in volatilised form in the vaporization-diffusion method. Using the latter approach there appeared to be less toxicity of the BCl treatments to the tester bacteria. Our findings suggest that BCl may show greater mutagenic activity in the gaseous phase. This work underlines the importance of using appropriate methods for the evaluation of volatile compounds. The modifications described here are easy to realize in practice and should prove useful for the investigation of other volatile materials or atmospheric contaminants.     

A comparative analysis of mutagenic and SOS-induced activity in three classes of chemical compounds

Mutagenic and SOS-inducing potential of 23 derivatives of fluorenone, phenanthrenequinone and biphenyl have been studied in tester strains of Salmonella typhimurium and in Escherichia coli strain PQ 37. 14 of these compounds revert the mutation hisD3052 (much less than -1 much greater than type), but none of them induce mutations in the strain TA 1535. Maximal mutagenic activity has been shown in strain TA 1538 for amide of 2,7-dinitrofluorenone-4-carbonic acid (580 revertants per nmol), 2,7-dinitrophenanthrenequinone (308 revertants per nmol), 2,4,7-trinitrophenanthrenequinone (306 revertants per nmol) and 2',4,4'-trinitrobiphenyl-2-carbonic acid (251 revertants per nmol). In plasmid-containing strain TA 98 the mutagenic potential of the compounds tested is lower than in the TA 1538 strain. It has been suggested that mutagenic activity of these compounds can be attributed to their acceptor properties, namely, the ability to form charge transfer complexes with DNA. SOS-inducing activity has been shown for 5 compounds, also positive in mutation induction. Mutagenic and SOS-inducing activities positively correlate in fluorenone derivatives. Among phenanthrenequinone derivatives, compounds with high mutagenic activity only can induce SOS response. None of the biphenyls tested induce SOS functions. The compounds giving the positive result in the SOS-chromotest have rigid co-planar structure.     

Structure-activity relationships for unsaturated dialdehydes. 1. The mutagenic activity of 18 compounds in the Salmonella/microsome assay

A considerable number of terpenes that contain an "unsaturated dialdehyde" functionality, and possess various biological activities, such as antimicrobial activity, pungency, antifeedant activity, and/or mutagenicity, have been isolated from natural sources. However, large qualitative and quantitative activity differences have been observed for the natural unsaturated dialdehydes, and small structural changes (e.g., stereoisomerization) seem to dramatically affect the biological activity. As part of a general attempt to study structure-activity relationships for unsaturated dialdehydes, the activity of compounds 1-18 (Table 1) in the Salmonella/microsome assay (strains TA98, TA2637 and TA100) has been investigated. 10 of the compounds were found to possess direct-acting mutagenic activity, although the mutagenic potencies vary considerably in this group (from 430 to 0.32 revertants per nmole in the Salmonella strain TA2637). Some structural features that appear to moderate the activity are discussed. The necessity of an intact unsaturated dialdehyde functionality for the mutagenic activity of isovelleral (1) (see Scheme 1 for names, numbers, and chemical structures) in the Salmonella/microsome assay was demonstrated by chemical conversions: modification of either aldehyde group or reduction of the double bond led to loss of activity.