muconaldehyde and muconic-acid

Benzene is myelotoxic and leukemogenic in humans. The mechanisms leading to these effects, however have not been fully elucidated. One of the underlying mechanisms is believed to be the oxidative damage caused by its metabolites. A comparative study was undertaken to examine the relationships between reactive oxygen species (ROS) production, lipid peroxidation and subsequent cytotoxicity induced by five major benzene metabolites. The generation of ROS by benzene metabolites was demonstrated by the significant and dose-dependent increase of intracellular ROS formation in HL60 human promyelocytic leukemia cells in vitro. 1,4-Benzoquinone (BQ) was found to be the most potent metabolite in induction of ROS formation, followed by 1,2,4-benzenetriol (BT) and to a lesser extent, phenol (PH) and trans, trans-muconaldehyde (MD). No significant effect was observed when the cells were treated with trans, trans-muconic acid (MA). The enhancement of ROS production by BQ was effectively inhibited by the addition of catalase, deferoxamine (DFO) and dimethyl sulfoxide (DMSO), but unchanged by superoxide dismutase (SOD), suggest that hydrogen peroxide (H2O2) and hydroxyl radicals (OH) are the two major forms of ROS involved. The results also demonstrate that the ability of benzene metabolites in enhancing ROS generation is closely correlated to their capacity in causing lipid peroxidation and subsequent cytotoxicity. These findings together with earlier parallel observations on DNA damage suggest that ROS play an important role in the mechanism of carcinogenesis induced by benzene metabolites.

Topics: Aldehydes; Benzene Derivatives; Benzoquinones; Carcinogens; Catalase; Deferoxamine; Dimethyl Sulfoxide; Dose-Response Relationship, Drug; Free Radical Scavengers; HL-60 Cells; Humans; Hydrogen Peroxide; Hydroquinones; Hydroxyl Radical; Lipid Peroxidation; Phenols; Reactive Oxygen Species; Sorbic Acid; Structure-Activity Relationship; Superoxide Dismutase

Perfusate from rat livers perfused with benzene (approximately 0.7-7 x 10(-4) M) or trans,trans-muconaldehyde (MUC) (10(-4) M) was extracted and analyzed by reverse-phase HPLC. Based on retention time and co-elution experiments, benzene was found to be metabolized to trans,trans-muconic acid, a urinary ring-opened metabolite of benzene and a major in vivo and in vitro metabolite of MUC. These data demonstrate that benzene ring-opening occurs in the liver. Following perfusion with MUC (a microsomal hematotoxic metabolite of benzene), trans,trans-muconic acid and three other MUC metabolites were detected in the perfusate extract, suggesting that these metabolites would be present in the circulation following metabolism of MUC.

Topics: Aldehydes; Animals; Benzene; Chromatography, High Pressure Liquid; Liver; Male; Organ Culture Techniques; Perfusion; Rats; Rats, Inbred F344; Rats, Sprague-Dawley; Sorbic Acid

t,t-Muconaldehyde and t,t-muconic acid have been investigated for the induction of gene mutations in Salmonella typhimurium (reversion of the his- strains TA97, TA98, TA100, TA102, TA104 and TA1535), Escherichia coli (reversion of the trp- strain WP2 uvrA) and Chinese hamster V79 cells (acquisition of resistance toward 6-thioguanine). t,t-Muconaldehyde proved weakly mutagenic in strain TA104 in the presence and absence of NADPH-fortified postmitochondrial fraction from rat liver homogenate (S9 mix). In strains TA97, TA100 and TA102, weak positive responses were observed only in the presence of S9 mix. In strains TA98, TA1535 and WP2 uvrA, the result was negative. In V79 cells, the mutation frequency was increased from approximately 7 X 10(-6) to 90 X 10(-6) in cultures exposed to t,t-muconaldehyde at optimal concentration (1.7-3 microM in separate experiments). The concentration-response curve showed pronounced hyperlinearity, with no mutagenic effect being observed at a third of the optimal concentration. t,t-Muconic acid was greater than 100 times less toxic than t,t-muconaldehyde in both bacteria and mammalian cells, and it did not show any mutagenic effect. These results complete a previous mutagenicity study, carried out on benzene and 13 metabolites. It is concluded that the newly investigated metabolites cannot account for the bacterial mutagenicity of bioactivated benzene and benzene-trans-1,2-dihydrodiol, since these compounds exhibited their strongest response in strain TA1535. t,t-Muconaldehyde showed similarities in its mutagenicity to p-benzoquinone and hydroquinone. All three compounds showed, at most, weak effects in bacteria, but were strongly mutagenic in V79 cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Aldehydes; Animals; Benzene; Cells, Cultured; Cricetinae; Cricetulus; Escherichia coli; Fatty Acids, Unsaturated; Male; Mutagenicity Tests; Mutagens; Mutation; Rats; Rats, Inbred Strains; Sorbic Acid

Our laboratory recently identified trans,trans-muconaldehyde (MUC), a six-carbon diene dialdehyde, as a hematotoxic microsomal metabolite of benzene (Latriano et al., Proc Natl Acad Sci USA 83: 8356-8360, 1986). We also showed that MUC is metabolized in vitro to trans,trans-muconic acid (MA), a six-carbon diene dicarboxylic acid and known urinary metabolite of benzene. To elucidate further the role of ring-opened metabolites in benzene toxicity, the metabolism of benzene and MUC was examined in the benzene sensitive DBA/2N mouse strain and the less benzene sensitive C57BL/6 strain. A sensitive assay for urinary MA analysis was developed. The percent of benzene dose excreted as urinary MA within the first 24 hr after treatment decreased with an increase in benzene dose, i.e. from 9.8 to 0.4% in DBA/2N mice and from 17.6 to 0.2% in C57BL/6 mice treated with 0.5 to 880 mg/kg benzene. DBA/2N mice excreted significantly (P less than or equal to 0.05) more MA compared with C57BL/6 mice after treatment with hematotoxic benzene doses (220-880 mg/kg). At low benzene doses (0.5 to 2.5 mg/kg), C57BL/6 mice excreted significantly (P less than or equal to 0.05) more MA compared with DBA/2N mice. There were no significant differences in the metabolism of MUC to MA between the two strains after treatment with 0.5 to 3.0 mg/kg. Furthermore, mice from both strains excreted similar amounts of muconic acid when treated with 0.7 to 7.1 mg/kg MA. These results are consistent with the hypothesis that reactive ring-opened metabolites such as trans,trans-muconaldehyde play a role in benzene hematotoxicity. Sensitivity towards benzene may be due, in part, to increased metabolism to ring-opened compounds.

Topics: Aldehydes; Animals; Benzene; Chromatography, High Pressure Liquid; Fatty Acids, Unsaturated; Male; Mice; Mice, Inbred C57BL; Mice, Inbred DBA; Sorbic Acid; Species Specificity

The mechanism of the genotoxicity and metabolism of benzene (BZ) was investigated by using a free-radical scavenger, dimethyl sulfoxide (DMSO), to investigate the free radical mechanism in BZ metabolism. The presence of chromosomal breakage expressed as micronuclei (MN) in bone marrow polychromatic erythrocytes (PCE) and the presence of several BZ metabolites in the urine were monitored. Adult male ICR mice were exposed orally to DMSO after oral exposure to BZ (440 mg/kg b.w.). DMSO was administered either in different concentrations (1.25, 3.75 or 12.5% given at a volume of 0.01 ml/gm b.w.) or at different intervals after BZ exposure (1, 3 or 5 h). Each group consisted of five mice. It was found that the BZ-induced MN frequency was reduced by DMSO from 48.8 +/- 5.6 (SEM) to 2.6 +/- 0.7 per 1000 PCE when DMSO (12.5%) was administered at 1 h after BZ exposure (P less than 0.01), to 3.4 +/- 0.8 at 3 h (P less than 0.01) and to 36.2 +/- 12.1 at 5 h (P less than 0.01). The reduction of the clastogenic effect of BZ by DMSO was also dependent upon the DMSO doses. The MN frequency was significantly reduced from 48.8 +/- 5.6 to 29.4 +/- 10.9 with 1.25% DMSO (P less than 0.01) to 20 +/- 7.6 with 3.75% (P less than 0.01) and to 2.6 +/- 0.7 with 12.5% DMSO (P less than 0.01). The presence of different metabolites of BZ such as hydroquinone, catechol, trans-trans muconic acid (MA, the oxidized form of trans-trans muconaldehyde, ttM), and total and conjugated phenol was evaluated in the urine of the exposed mice using HPLC. Among these metabolites, the quantity of MA was found to have the closest positive correlation with the MN frequency (P less than 0.007). Phenol but not the other monitored metabolites was also positively correlated with MN frequency (P less than 0.03). Thus, our data show that the formation of genotoxic metabolites from BZ probably involves hydroxyl radicals and ttM as well as phenol are likely to be responsible for the clastogenic effect of benzene in vivo.

Topics: Aldehydes; Animals; Benzene; Dimethyl Sulfoxide; Male; Mice; Mice, Inbred ICR; Micronucleus Tests; Mutagens; Sorbic Acid