Mutagenicity tests of fabric-finishing agents in salmonella typhimurium: Fiber-reactive wool dyes and cotton flame retardants

Thirty-nine fabric-finishing agents were tested for mutagenic activity in Salmonella typhimurium. Twenty-four fiber-reactive wool dyes and three acid dyes (not fiber-reactive) were screened by spot tests in strains TA100, TA98, TA1535, and TA1537. Among these dyes, seven bromoacrylamide dyes and one vinyl sulfone dye were mutagenic. Additionally, one of the three acid dyes was mutagenic in spot tests. The mutagenicity of the acid dye was due to an impurity or breakdown product rather than to the dye itself; the origin of the activities of the other dyes is unknown. No mutagenicity was observed among five chlorotriazine or four sulfonyl-ethane sulfonic acid dyes. Eight phosphorus-containing flame retardants (phosphonium, phosphine, phosphine oxide, and phosphonic acid derivatives) and methyl-N-methylolcarbamate, which is employed to obtain a flame-retardant finish on cotton, were tested for mutagenicity in strains TA100, TA98, TA1535, and TA1537, using quantitative incorporation assays. All were nonmutagenic. Two of three bromoalkyl-substituted triazine flame retardants were mutagenic in strains TA100 and TA1535. It is unknown whether this activity is due to impurities or to the parent compound. The flame retardants tested were either in actual commercial use or in experimental development for potential commercial processes. These results indicate the need for early testing of potential fabric-finishing agents and processes.     

Mutagenicity evaluation of amino-oxazoline derivatives using in vitro and in vivo short-term tests

During routine investigation of potential drugs it was found that 1-methyl-4-(2-oxazoline-2-ylamino)-indazole (<1) compound 1 was mutagenic for Salmonella typhimurium TA 1535 and TA 100. The genotoxicity of compound 1 was confirmed by the following in vitro test systems: V79 Chinese hamster cells (HGPRT-locus), Saccharomyces cerevisiae D7 (mitotic gene conversion, mutations, aberrations), and Escherichia coli (rec-assay). On the other hand, when compound 1 was tested in vivo (micronucleus test in mice and sister chromatid exchange in Chinese hamsters) it did not show evidence of genotoxic activity. In order to study structure/activity relationships, different analogues of compound 1 were tested in Salmonella typhimurium TA 1535. The tests showed that (1) most 2-amino-oxazolines were mutagenic for the test organism; (2) the 2-amino-imidazoline and 2-amino-thiazolidine derivatives tested were not mutagenic; (3) substituents bound to the extranuclear nitrogen of the 2-aminooxazoline ring had only a weak influence on the mutagenic potential; and (4) methylation of the oxazoline ring, most conspicuously at the carbon in position 5, strongly reduced the mutagenic effect. From these observations it is concluded that the reaction of nucleophilic DNA sites with the most electrophilic site of the oxazoline ring, ie, the carbon in position 5, is responsible for the genotoxicity of amino-oxazoline compounds. Due to the genotoxicity observed in these in vitro tests this class of compounds was no longer developed, but dropped, even without performing a long-term carcinogenicity study or considering the negative in vivo findings.     

Structural basis of the mutagenicity in bacteria of nitrated naphthalene and derivatives

While 2-nitronaphthalene was a weak direct-acting base-substitution mutagen (1.4 revertants/nanomole) for Salmonella typhimurium, the analogous nitronaphthalic acid anhydride and imides were moderate frameshift mutagens (～20 rev/nanomole in strain TA98). Although imide derivatives are efficient DNA intercalators, mutagenicity data indicate that the bulk of the frameshift activity is derived from adduct formation between hydroxylamine intermediates and DNA. The low level of frameshift activity (～8% of total) resulting from simple intercalation (measured in strain TA1537) is not dependent upon reduction of the nitro function. Evidence is presented that suggests that the reduction of the nitro function to the corresponding hydroylamine might not involve a free nitroso intermediate. The introduction of a second nitrofunction into nitronaphthalenes results in great positional effects of the various isomers on mutagenic activity and specificity.     

Mutagenicity and cross-linking activity of chloroalkylnitrosamines,possible new antitumor lead compounds

The mutagenicity of chlorinated alpha-acetoxy nitrosamines was assayed using nine bacterial strains with various DNA repair abilities and the mechanism of their reaction with DNA was evaluated. Three alpha-acetoxy nitrosamines without a chloro group were used to investigate the effect of the chloro group on mutagenicity. Three nitrosamines having chloroethyl, chloropropyl and chlorobutyl groups were directly mutagenic in all tester strains used in this study, which showed that they damaged DNA in intact bacteria. Compared with Salmonella typhimurium TA1535, mutagenic activity was enhanced in ogt gene-deficient strains (YG7108 and YG7104), suggesting that an O(6)-alkylguanine adduct causes the mutation. The chlorinated nitrosamines showed stronger mutagenicity than non-chlorinated nitrosamines, indicating that alkylating activity was strengthened by the presence of a chloro group. The nitrosamines, especially the chloropropyl homolog, showed clear mutagenicity in the strains with an intact excision repair system, S.typhimurium TA92, TA1975 and G46. Further, chloropropyl and chlorobutyl homologs showed interstrand cross-linking activity towards plasmid DNA. These results suggest that some chlorinated nitrosamines can act on DNA to form DNA cross-links, as observed in antitumor chloroethylnitrosoureas. Environmental nitrosamines are usually dealt with as potential carcinogens, but introduction of a chloro group has added the possibility of in vivo cross-linking activity, which is a classical and essential mechanism for antitumor agents. Therefore, the novel chlorinated nitrosamines examined in this study are proposed as new bifunctional antitumor lead compounds.     

Propyldazine is mutagenic in Salmonella typhimurium and Escherichia coli: distinct specificity for strains TA1537 and TA97

The antihypertensive drug propyldazine (Atensil) was demonstrated to be mutagenic with auxotrophic mutants of Salmonella typhimurium and Escherichia coli. Addition of liver S9 mix (postmitochondrial supernatant fraction supplemented with an NADPH-generating system) had little, if any, effect on the mutagenicity. The mutagenicity showed an unusual pattern of strain specificity. Increased frequencies of reversion were observed with all strains whose auxotrophy was caused by frame-shift mutations: the number of revertant colonies per plate from S. typhimurium TA98, TA1538, TA97, and TA1537 was increased up to 5-, 9-, 43-, and 160-fold, respectively, above background. Among the strains that became auxotrophic by substitution mutations, S typhimurium TA102, E. coli WP2, and E coli WP2 uvrA yielded positive results (twofold above background). S. typhimurium TA1535 and TA100 were not reverted by propyldazine. It should be noted that propyldazine, due to its low toxicity and good solubility, could be tested up to very high doses. Hence, although quite impressive mutagenic effects occurred, the mutagenic potency was moderate even in the most responsive strains, TA1537 and TA97 (about 0.3 and 1.0 revertants per nmole, respectively). With the limitation that the strain specificities were different, the mutagenic potency of propyldazine was in the same order of magnitude as that of hydralazine and dihydralazine, two related antihypertensive drugs which were already known to be mutagenic. In our hands, both compounds were mutagenic in S typhimurium TA1535, TA100, TA1537, and TA98. These results differ from data in the literature in that we found clear but weak effects even with strains for which others have reported negative results.     

Mutagenic potentials of dental cements as detected by the Salmonella/microsome test

The potential mutagenicity of a zinc phosphate (Poscal), a polycarboxylate (Aqualox) and glass ionomer cements with (Argion) and without (Meron) silver reinforcement were characterized by employing the Ames Salmonella/microsome test. The materials were eluted in dimethyl sulphoxide or physiologic saline and the aliquots were used either immediately or after an incubation period of 24h at 37 degrees C. Mutagenic effects of the materials were tested on Salmonella typhimurium strains TA 98, TA 100, TA 102 and TA 1535 using the standard plate incorporation assay, and in the presence or absence of S9 fraction from rat liver. Poscal and Aqualox elicited mutagenic effects on S. typhimurium TA 98 and TA 1535, whereas Meron exhibited mutagenic effects on S. typhimurium TA 98. No mutagenic effects were detected for Argion. The type of solvent, dose of the material and incubation as well as the interactions between these factors exhibited varying degrees of influences on the mutagenic activities of the cements (P<0.05 and P<0.1). We conclude that zinc phosphate, polycarboxylate, and glass ionomer cements may have possible mutagenic activities.     

Evaluation of the genotoxicity of gentian violet in bacterial and mammalian cell systems

Previous studies indicate that gentian violet (GV), a triphenylmethane dye used in agriculture and human medicine, is a clastogen in vitro and a carcinogen in chronically exposed mice and rats. Data on its genotoxic activity, however, have been incomplete and partly contradictory. Mutagenesis and DNA damage experiments were conducted to re-evaluate the genotoxic potential of GV in both bacterial and mammalian cell systems. GV was mutagenic in Salmonella typhimurium tester strains TA97 and TA104, but there was little mutagenic activity detected in strains TA98 and TA100. A rat liver homogenate fraction (S9) tended to increase mutagenicity. The major microsomal metabolites of GV, pentamethylpararosaniline and N,N,N',N'-tetramethylpararosaniline were less mutagenic in TA97 and TA104, while N,N,N',N"-tetramethylpararosaniline was a weak mutagen in Salmonella. GV was not mutagenic in Chinese hamster ovary (CHO) cell strain CHO-K1-BH4, and was a questionable mutagen in CHO-AS52 cells. While GV produced DNA damage as measured by sedimentation of nucleotids derived from B6C3F1 mouse lymphocytes treated in vitro, no damage was found in lymphocytes isolated from mice dosed with GV. GV was also a weak producer of gene amplification in an SV40-transformed Chinese hamster cell line. The results indicate that GV is a point mutagen in bacteria; however, since similar exposure conditions produced weak mutagenic activity in mammalian cells, GV may be carcinogenic by virtue of its clastogenic activity.     

Microbial mutagenicity of isomeric two-, three-, and four-ring amino polycyclic aromatic hydrocarbons

The isomers of various two-, three-, and four-ring amino polycyclic aromatic hydrocarbons were tested for mutagenic activity using a microbial plate incorporation test with four Salmonella typhimurium strains (TA98, TA100, TA1535, and TA1537). All compounds were assayed with an S9 metabolic activating enzyme system. The two-ring compounds were tested only with TA98. All were weakly mutagenic (1-10 rev/micrograms) except 2-aminobiphenyl, which was not mutagenic under these test conditions. All except two of the 13 fused three-ring compounds (aminofluorenes, aminoanthracenes, and aminophenanthrenes) were active frame shift mutagens; only the aminophenanthrenes were active base-pair mutagens. The potency of this group of isomeric compounds ranged from moderately (approximately 20 rev/microgram) to strongly (greater than 5,000 rev/microgram) mutagenic. As a group, the pericondensed four-ring amino compounds were the most mutagenic of the three groups tested. All of the aminofluoranthene and aminopyrene isomers showed significant mutagenic activity with TA98, TA100, and TA1537. In general, the mutagenic potency of the amino polycyclic aromatic compounds tested was highly dependent on the structural position of the amino group.     

Evidence that 4-aminobiphenyl, benzidine, and benzidine congeners produce genotoxicity through reactive oxygen species

4-Aminobyphenyl (4-Ab), benzidine (Bz), and Bz congeners were evaluated for their ability to induce genotoxicity through an oxidative mechanism. The mutagenicity of these compounds was tested in the presence and absence of Aroclor 1254-induced rat S9 mix using Salmonella typhimurium tester strain TA102, which is sensitive to agents producing reactive oxygen species (ROS). In the presence of S9, 4-Ab, Bz, N-acetyl-benzidine, and 3,3-dimethoxybenzidine were strongly mutagenic in TA102, whereas, 3,3,5,5-tetra-methylbenzidine, 3,3-dimethylbenzidine (O-tolidine), and N,N-diacetylbenzidine were not mutagenic. In addition, 3,3-dichlorobenzidine and 4,4-dinitro-2-biphenylamine were directly mutagenic in TA102. Incorporation of the free radical and metal scavengers, catalase, superoxide dismutase (SOD), butylated hydroxytolune (BHT), and ethylenediamine tetraacetic acid (EDTA) reduced the mutagenic responses of 4-Ab and Bz, whereas heat-inactivated catalase and SOD had no effect. 4-Ab and Bz also induced lipid peroxidation in the presence of S9 mix as shown using the thiobarbituric acid reactive substances assay. The results of this study indicate that 4-Ab and Bz induce mutations through the induction of ROS.     

Structure-activity relationships in the mutagenicity of N-substituted derivatives of phenanthrene-9,10-imine

A series of K-region, N-substituted phenanthrene imines were tested for mutagenicity in Salmonella typhimurium TA100. All chemicals were mutagenic in the absence of an exogenous metabolic activation system. The apparent decay times of the mutagenic species in diffusion plates and their alkylating activities were also measured. The unsubstituted phenanthrene-9,10-imine was approximately 70-fold more mutagenic than the corresponding phenanthrene-9,10-oxide. N-substitution with electron-releasing groups resulted in chemicals that were more mutagenic than those substituted with electron-withdrawing groups. The mutagenic activity of the latter group of chemicals was comparable with that of phenanthrene-9,10-oxide. Except for N-chlorophenanthrene imine, both alkylation of p-nitrothiophenol and apparent decay times in diffusion plates were inversely correlated with mutagenicity. It is hypothesized that reactivity towards p-nitrothiophenol (alkylating activity) and mutagenicity reflect different reactions, in contrast to other chemical mutagens. The results suggest that the high potency of phenanthrene imines as mutagens is possibly due to DNA binding via an aziridinium ion rather than a carbonium ion.     

Antibacterial activity and mutagenicity studies of water-soluble phosphazene high polymers.

Eight water-soluble phosphazene high polymers, [NPR2]n (R, organic, water-solubilizing side-group; n, approx: 15,000) and the small-molecule counterparts of the polymers were examined for antibacterial activity against six different strains of bacteria (Escherichia coli, Salmonella typhimurium (TA 100), Salmonella pullorum, Streptococcus faecalis, Bacillus subtilis and Pseudomonas aeruginosa). Antibacterial testing was carried out by measuring zones of inhibition and changes in solution turbidity over time. In addition, the antibacterial activity of the surfaces of cross-linked poly[di(methoxyethoxyethoxy)phosphazene] (MEEP) hydrogels were investigated. A number of the high polymers, as well as the MEEP hydrogels, impeded bacterial growth. Only E. coli was unaffected by the phosphazenes. A possible explanation for the antibacterial character of the polymers is presented. The same compounds were monitored for potential mutagenic activity using the Salmonella typhimurium tester strains TA 100 and TA 98. None of the high polymers or their small-molecule analogues showed mutagenic activity in either strain of Salmonella at the concentrations tested. The use of these materials as coatings for artificial implants is discussed.     

Mutagenic activity of the 4,5- and 9,10-dihydrodiols of benzo[J]fluoranthene and their syn- and anti-dihydrodiol epoxides in salmonella typhimurium

The objective of this study was to determine the relative mutagenic activities of the major dihydrodiol metabolites of benzo[j]fluoranthene (B[j]F) and their corresponding syn- and anti-dihydrodiol epoxides. Salmonella typhimurium tester strains TA97a, TA98, and TA100 were used to evaluate the mutagenic potencies of the parent hydrocarbon and these suspect proximate and ultimate mutagenic metabolites. B[j]F and the trans-dihydrodiol metabolites were active only in the presence of an external metabolic activation system (S9) with the exception of the B[j]F-4,5-diol, which was weakly active in TA98 and TA100 in the absence of S9. The B[j]F-4,5-diol was more mutagenic than the B[j]F-9,10-diol in tester strains TA98 and TA100, whereas the opposite effect was observed in TA97a. In the absence of S9, the anti-B[j]F-4,5-diol epoxide was more mutagenic than the syn-B[j]F-4,5-diol epoxide and the syn- and anti-B[j]F-9,10-diol epoxides in tester strains TA97a and TA100. The exceptional mutagenic potency of the anti-B[j]F-4,5-diol epoxide in TA100 resembles that observed by epoxides located within a fjord, or by the anti-diol epoxides of bay region methylated polycyclic aromatic hydrocarbons. In contrast, the mutagenicity of the pseudo bay region dihydrodiol epoxides arising from the B[j]F-9,10-diol more closely resembles that observed with the classical bay region dihydrodiol epoxides of chrysene. In summary, both dihydrodiol metabolites of B[j]F are mutagenic in S. typhimurium, and the relative potency varies among the tester strains. The highest mutagenic response was achieved in tester strain TA100, which detects base-pair substitutions. The most potent direct-acting dihydrodiol epoxide in this tester strain was the anti-B[j]F-4,5-diol epoxide, which agrees with the results of mouse skin painting studies that indicate that the B[j]F-4,5-diol is more tumorigenic that the parent hydrocarbon or the B[j]F-9,10-diol. A cova-lent DNA adduct formed between the anti-B[j]F-4,5-diol epoxide and deoxyguanosine was the major species of DNA adduct formed in S. typhimurium. This adduct corresponds to the major DNA adduct formed in mouse skin following application B[j]F.     

Genotoxic activity in microorganisms of tetryl, 1,3-dinitrobenzene and 1,3,5-trinitrobenzene

N-Methyl-N,2,4,6-tetranitroaniline (tetryl), 1,3-dinitrobenzene, and 1,3,5-trinitrobenzene were subjected to DNA repair assays using the Escherichia coli W3110/polA⁺, p3478/polA^? system, reverse mutation assays with His^? Salmonella typhimurium strains TA1535, TA1537, TA1538, TA98, and TA100, and mitotic recombinogenic tests with the yeast Saccharomyces cerevisiae D5. Tests were carried out in the absence of an exogenous activation system and in tissue homogenate-mediated assays using Aroclor 1254-induced, male rat-liver-derived S9 mix. Mutagenic activity of tetryl was demonstrated with S typhimurium strains TA1537, TA1538, TA98, and TA100. The responses were particularly strong in the absence of S9 mix. Tetryl also induced increases in recombinant numbers and frequencies in the S cerevisiae test without the S9 mix, but not in its presence. 1,3-Dinitrobenzene was demonstrated to be a mutagen with S typhimurium strains TA1538, TA98, and TA100. Slight activity was also seen with TA1537. The S9 mix reduced the magnitude of the responses. 1,3,5-Trinitrobenzene was also demonstrated to be mutagenic with S typhimurium strains TA1535, TA1537, TA1538, TA98, and TA100. Again, the S9 mix reduced the magnitude of the responses. In this segment of a programme initiated by military authorities, the genotoxic potential of three nitroaromatic compounds, which have found significant use in explosive preparations, has been demonstrated. Twelve other compounds used in ordnance were not active in any of the test systems. These were octahydro-1-acetyl-3,5,7-trinitro-S-tetramine (SEX), hexahydro-1,3-dinitro-5-acetyl-S-triamine (TAX), ethyl centralite, 2-nitrodiphenylamine, N-nitrosodiphenylamine, diphenylamine, diethyleneglycoldinitrate, nitroguanidine, lead salicylate, lead resorcylate, red phosphorus, and zinc chloride.     

Electronic considerations in the mutagenesis of some 4,5-bridged chrysenes

The mutagenic activity of four 4,5-bridged chrysene derivatives, benz(a)aceanthrylene, and 5-methylchrysene was examined using histidine auxotrophic strains TA98 and TA100 of Salmonella typhimurium. All compounds showed a positive mutagenic response with both TA100 and TA98 in the presence of S-9. A correlation between the electronic character of the bridging group and mutagenic activity for the chrysene derivatives is proposed.     

Examination of Alternaria alternata mutagenicity and effects of nitrosylation using the Ames Salmonella test.

Molds of the genus Alternaria are common food pathogens responsible for the spoilage of fruits, vegetables, grains, and nuts. Although consumption of Alternaria alternata-contaminated foodstuffs has been implicated in an elevated incidence of esophageal carcinogenesis, the mutagenic potencies of several A. alternata toxins seem unable to account for the levels of activity found using crude mycelial extracts. In this study, the mutagenic effects of nitrosylation were examined with the major Alternaria metabolites Altenuene (ALT), Alternariol (AOH), Alternariol Monomethyl Ether (AME), Altertoxin I (ATX I), Tentoxin (TENT), Tenuazonic Acid (TA), and Radicinin (RAD) using the Ames Salmonella strains TA98 and TA100. In the absence of nitrosylation, ATX I was mutagenic when tested from 1 to 100 microg/plate in TA98 with rat liver S9 for activation, while AOH and ATX I were weakly mutagenic +/- S9 in TA100. Incubation with nitrite generally increased mutagenic potencies with ATX I strongly mutagenic +/- S9 in both TA98 and TA100, while ALT, AOH, AME, and RAD responses were enhanced in TA100 + S9. However, subsequent examination of three extracts made from A. alternata culture broth, acetone-washed mycelia, and the acetone washes showed a different mutagenic response with both broth and acetone washes directly mutagenic in TA98 and TA100 but with a reduced response + S9. The acetone-washed mycelial extract was found to have the lowest mutagenic activity of the three extracts tested. Nitrosylation had little effect on the mutagenicity of any of the extracts. Thus, while nitrosylation increases the mutagenicity of ATX I, and to a lesser extent that of several other Alternaria toxins, the results demonstrate that Alternaria produces a major mutagenic activity with a S. typhimurium response different from that found with the purified toxins. Efforts are currently underway to chemically identify this mutagenic species. Published 2001 Wiley-Liss, Inc.     

Rat nasal tissue activation of benzo(a)pyrene and 2-aminoanthracene to mutagens in Salmonella typhimurium

Cytochrome P-450-dependent monooxygenase activity has been measured in the nasal turbinates of dogs and rats. The capacity of male Fischer-344 rat nasal tissue to bioactivate benzo(a)pyrene (BaP) and 2-aminoanthracene (2-AA) to mutagens in Salmonella typhimurium was investigated. 2-AA was mutagenic in strains TA98 and TA100 when nasal tissue S-9 was utilized as the activating enzyme system and BaP was mutagenic in strain TA100. At all doses and protein concentrations tested, 2-AA displayed nearly 500-1000 times greater bacterial mutagenicity than BaP. In strain TA-100, nasal tissue S-9 was approximately twice as active toward 2-AA as lung S-9 and 75% as active as liver S-9. Aryl hydrocarbon hydroxylase activity was detected in rat nasal tissue when 14C-BaP was used as a substrate. Rat nasal tissue metabolized BaP to several oxidized metabolites which included dihydrodiols, quinones, and phenols. 3-Hydroxybenzo(a)pyrene and BaP-3, 6-quinone were the major metabolites detected (150 pmoles/mg protein/30 min). These results indicate that rat nasal tissue can metabolize promutagens to reactive species which may play an important role in xenobiotic-induced nasal tumors.     

Hydroxymethyl-substituted furans: mutagenicity in Salmonella typhimurium strains engineered for expression of various human and rodent sulphotransferases

5-Hydroxymethylfurfural (HMF) and furfuryl alcohol (FFA) are present in numerous foodstuffs at high levels. FFA is also used for the production of polymers. Both compounds had demonstrated some evidence of carcinogenic activity in 2-year bioassays. We tested these compounds and four congeners for mutagenicity in Salmonella typhimurium TA100 and TA100-derived strains expressing human or rodent sulphotransferases (SULTs). 5-Hydroxymethylfuroic acid, a metabolite of HMF, was not mutagenic in any strain. 3-Hydroxymethylfuran was weakly mutagenic in all strains independently of SULT expression. HMF, 2,5-(bishydroxymethyl)furan (metabolite of HMF), FFA and 5-methyl-FFA were inactive in TA100 but strongly mutagenic when human SULT1C2 was expressed. This form has been detected in ovary, kidney and foetal tissues. Human SULT1A1, SULT1A2 and SULT1A3 as well as murine Sult1a1 and Sult1d1 also activated some hydroxymethyl-substituted furans to varying degrees. Whereas chemically synthesised 5-sulphooxymethylfurfural was mutagenic in TA100, furfuryl sulphate was bacteriotoxic, only leading to marginal increases in the number of revertants. Furfuryl acetate, an uncharged ester of FFA, used as fragrance and food flavouring, was clearly mutagenic. We determined half-life times of 120 min, 20 s and 10 h, respectively, for 5-sulphooxymethylfurfural, furfuryl sulphate and furfuryl acetate at 37°C in water. It is likely that the short lifespan of furfuryl sulphate, together with its charge, led to insufficient penetration of the bacteria when added externally, although it was mutagenic when generated by appropriate SULTs from FFA within the cell.     

Mutagenic lipid peroxides from edible oils

Weak mutagenic activity was detected in several commercially available edible palm and corn oils using liquid incubation bioassays with Salmonella typhimurium TA1537. Chromatographic fractionation of unrefined palm oil established that mutagenic activity was present in three fractions that also contained fatty acyl hydroperoxides. Similar weak mutagenic activity was also demonstrated for linoleic and linolenic acid hydroperoxides. In all cases, the mutagenicity was abolished by exogenous catalase, implying that the observed activity was moderated by hydrogen peroxide.     

Mutagenicity of N- and O-acetyl derivatives of methyl 3,4-diphenyl-5-hydroxylamino-2-furoate and N-hydroxy-4-aminobiphenyl in Salmonella typhimurium

Methyl 3,4-diphenyl-5-hydroxylamino-2-furoate (N-OH-MDPF) (I), methyl 3,4-diphenyl-5-acetoxyamino-2-furoate (N-OAc-MDPF) (II), methyl 3,4-diphenyl-N-hydroxy-5-acetylamino-2-furoate (N-OH-MDPAF) (III), and methyl 3,4-diphenyl-N-acetoxy-5-acetylamino-2-furoate (N-OAc-MDPAF) (IV) were synthesized and tested for mutagenic activity for Salmonella typhimurium TA98 and TA100. The hydroxylamine (I) and acetyl derivatives (II-IV) did not show mutagenic activity in TA98 or TA100. In contrast, the parent nitro compound, methyl 3,4-diphenyl-5-nitro-2-furoate (MDPNF) (V) was found to be equally active in TA98 and TA98-DNP, and more active in TA100 and TA104. The mutagenic activity in TA100 and TA104 decreased significantly under anaerobic conditions. Additionally, MDPNF was previously shown to be less mutagenic in the nitroreductase-deficient derivatives TA100NR and TA98NR, suggesting a requirement for nitro reduction. Incubation of V with NADPH and bacterial lysates of TA98 or TA98NR yielded a metabolite which was identified as I based on chromatographic and mass spectral characteristics. The rate of reduction by the lysate of TA98NR was about one-third that of TA98, showing a correlation between mutagenicity and nitroreductase activity. The lysates of TA98 did not reduce N-OH-MDPF further to the amine. In contrast to the lack of mutagenic activity of I-IV, N-hydroxy-4-aminobiphenyl (N-OH-ABP) and its acetyl derivatives were active in TA98, but less so in TA98-DNP. These data suggest that mechanisms involving O-acetylation of N-hydroxylamine to the acetoxyamine or acyl transfer reactions are not involved in the generation of mutagen from MDPNF. Furthermore, the differential mutagenic response of V in TA98 and TA98NR, its reduction to I, and the lack of activity of I suggest that the intermediates of reduction between the nitro and hydroxylamine, such as nitro or nitroso free radical anions, may be involved in mutagenesis. The decreased response of V under anaerobic conditions and increased response in TA104 suggest that secondary oxygen radicals generated from reduction intermediates may be responsible for the mutagenicity of MDPNF.     

Mutagenicity testing of di(2-ethylhexyl)phthalate and related chemicals in Salmonella

Di(2-ethylhexyl)phthalate and 33 other phthalates, ethylhexanol derivatives, and related chemicals were tested for mutagenicity in Salmonella typhimurium strains TA98, TA100, TA1535, and TA1537 without metabolic activation and in the presence of rat and hamster liver S-9 metabolic activation systems. No mutagenic activity was seen with any of the chemicals tested.