Showing papers by "Franz Oesch published in 1977"
TL;DR: Using the recently developed sensitive assay with [3H] benzo [a] pyrene 4,5-oxide as substrate, epoxide hydratase was shown to be present in 26 rat organs and tissues investigated, supporting the assumption that in these organs a single enzyme is responsible for the hydration of both substrates—as was earlier shown by several methods for the rat liver.
141 citations
TL;DR: The findings indicate that when microsomes from untreated and phenobarbital-treated mice were used the main contributors to the mutagenicity were simple epoxides (or compounds arising non-enzymically from them) and the activation of dihydrodiols must, however, contribute to a significant extent when microSomes from methylcholanthrene-treated dogs were used.
Abstract: The effect of epoxide hydratase upon the mutagenicity of benzo(a)pyrene was investigated using two Salmonella typhimurium strains (TA 1537 and TA 98). These two bacterial strains were found to differ characteristically in their susceptibility to different mutagens biologically produced from benzo(a)pyrene providing a diagnostic tool to investigate which types of mutagenic metabolites were produced in various metabolic situations. The results showed that the pattern of mutagenic metabolites produced by microsomes from methylcholanthrene-treated mice was very different from that produced by microsomes from phenobarbital-treated or untreated mice. However in all cases at least two mutagenic metabolites were produced. Epoxide hydratase was very efficient at reducing the mutagenic effect when benzo(a)pyrene was activated by microsomes from untreated or phenobarbital-treated mice. However, when microsomes from methylcholanthrene-treated mice were used the effect of hydratase depended upon the benzo(a)pyrene concentration. At low concentrations the mutagenicity was increased by addition of epoxide hydratase and decreased by inhibition of the hydratase. At high concentrations the reverse was true. These findings indicate that when microsomes from untreated and phenobarbital-treated mice were used the main contributors to the mutagenicity were simple epoxides (or compounds arising non-enzymically from them). The activation of dihydrodiols must, however, contribute to a significant extent when microsomes from methylcholanthrene-treated mice were used. Thus the role of epoxide hydratase was determined by the monooxygenase form present in the microsomes in the activating system.
95 citations
TL;DR: The observation that benzo(a)pyrene 4,5-oxide was a very good substrate for the cytosol localized glutathione S-transferase, but that it was not inactivated by this system when bacteria and microsomes were in direct contact, indicates that a severe limitation for the inactivation of benzo (a) pyrene metabolites by this enzyme is imposed by its localization in the cytOSol.
Abstract: Benzo(a)pyrene was activated to metabolites mutagenic for Salmonella typhimurium TA 98 by liver microsomes from control and phenobarbital treated mice. Under these conditions benzo(a)pyrene 4,5-oxide accounts for most of the mutagenicity. We have therefore investigated (1) the conjugation of benzo(a)pyrene 4,5-oxide with glutathione and (2) the effect of glutathione on the mutagenicity of benzo(a)pyrene.
85 citations
59 citations
TL;DR: It is concluded that species such as mice which possess high monooxygenase activity but very low epoxide hydratase activity are much more susceptible than man to those toxic effects which are mediated by metabolically formed epoxides which are substrates of epoxideHYDRatase.
Abstract: Microsomal monooxygenases catalyze the biosynthesis of epoxides from olefinic and aromatic compounds whilst microsomal epoxide hydratase and cytoplasmic glutathione S-transferases are responsible for their further biotransformation. Although catalytically very efficient the cytoplasmic glutathione S-transferases play, due to their subcellular localization, a minor role in the inactivation of epoxides derived from large lipophilic compounds and were, therefore, not included in this study. It was shown with such a lipophilic compound, benzo(a)pyrene, as a model substance and with liver enzyme mediated bacterial mutagenesis as biological endpoint that species and strain differences in epoxide hydratase and monooxygenases are reflected in very dramatic differences in mutagenicity of benzo(a)pyrene which varied from extremely potent to a degree which could easily be overlooked. In order to investigate whether the differences in enzyme activities were causally linked to the observed differences in mutagenicity, the enzyme activities were modulated by inhibition and induction. These manipulations were always accompanied by the corresponding changes in mutagenicity. It is concluded that species such as mice which possess high monooxygenase activity but very low epoxide hydratase activity are much more susceptible than man to those toxic effects which are mediated by metabolically formed epoxides which are substrates of epoxide hydratase. In this regard, it is especially noteworthy that mice possess a much lower hepatic epoxide hydratase activity than man.
52 citations
01 Jan 1977
TL;DR: Previous studies had shown that the 4,5- (K-region-) epoxide metabolite was a potent mutagen for the frameshift-sensitive Salmonella strains TA 1537 and TA 1538, and that the mutagenic response was potentiated by the presence of epoxide hydratase inhibitors at concentrations where no interference with other systems has been observed.
Abstract: Aromatic and olefinic compounds can be metabolized by microsomal monooxygenases to epoxides which chemically represent electrophilic species (for reviews, see refs. 1–5). Spontaneous binding of such epoxides to DNA, RNA, and protein has been observed (6–10). Accordingly, such metabolites have been suggested and, in some instances, shown to disturb the normal functions of cells, leading to such effects as mutagenesis (11–14), malignant transformation (15–19), or cell necrosis (20). However, aromatic and olefinic compounds are biotransformed to a vast array of metabolites (cf. refs. 21–27), possibly including a considerable number of reactive metabolites other than epoxides. The relative importance of epoxides among other reactive metabolites is at present unknown. With respect to the model compound used in this study, benzo[a]pyrene, our previous studies had shown that the 4,5- (K-region-) epoxide metabolite was a potent mutagen for the frameshift-sensitive Salmonella strains TA 1537 and TA 1538 (28), that the premutagenic hydrocarbon required a NADPH-supported microsomal monooxygenase system to become mutagenically active, and that the mutagenic response was potentiated by the presence of epoxide hydratase inhibitors at concentrations where no interference with other systems has been observed (28). Yet no conclusion could be reached whether the relative contribution of epoxide metabolites to the overall muta-genic effect of bioactivated benzo[a]pyrene was of any significance since the potentiation of the mutagenic effect by epoxide hydratase inhibitors could simply mean that blocking this pathway led to an accumulation of epoxides, making them important in this situation, while in absence of such inhibitors their contribution to the overall mutagenic effect may have been negligible.
25 citations
TL;DR: In this article, a 3.6-bis-trimethylsilyl-1,4-cyclohexadiene with tritiated water was used to obtain benzene oxide.
Abstract: Benzene oxide -[U-14C] was prepared from benzene -(U-14C) by modifications of methods described for the inactive compound.
Benzene oxide-[3.6–3H] was prepared by decomposition of 3.6-bis-trimethylsilyl-1,4-cyclohexadiene with tritiated water. bromination of the 1,4-cyclohexadiene-[3,6-3H] so obtained. epoxidation and dehydrobromination.
With the latter method benzene oxide-[3,6–3H] can be prepared at a much lower cost and higher specific activity than benzene oxide-[U-14C].
5 citations
01 Jan 1977
TL;DR: Benzo(a)pyrene was activated by liver microsomes to mutagens detected by the reversion of histidine dependent Salmonella typhimurium TA 1537, and it was shown that epoxide hydratase was a critical enzyme for the inactivation of these Mutagens.
Abstract: SUMMARY Benzo(a)pyrene was activated by liver microsomes to mutagens detected by the reversion of histidine dependent Salmonella typhimurium TA 1537. Using pure epoxide hydratase or epoxide hydratase inhibitors, comparing animal species with high and low epoxide hydratase activity, or inducing monooxygenase activity, it was shown that epoxide hydratase was a critical enzyme for the inactivation of these mutagens. Many clinically used drugs are metabolized to epoxides. Epoxides are not necessarily mutagenic, but since epoxide hydratase has a very low substrate specificity, such epoxides may competitively inhibit the hydration of mutagenic epoxides, as demonstrated in the present study for the metabolically produced epoxide from the clinically used drug cyproheptadine. Interestingly the structurally closely related epoxide derived from carbamazepine did not significantly inhibit epoxide hydratase. In a therapeutic situation it would be expected that the concentration of the epoxides metabolically produced from the drug would be much greater than that of the epoxides produced from polycyclic hydrocarbons which are present ubiquitously, but at very low levels. Thus, competitive inhibition by the former may be very effective and may potentiate adverse biological effects of the latters.
01 Jan 1977
TL;DR: The findings suggest that the enzyme isolated is the only one present in the rat liver microsomal fractions which is responsible for the hydration of such substrates.
Abstract: The microsomal enzyme epoxide hydratase has been purified to homogeneity as judged by electrophoretical, ultracentrifugal and immunological criteria and by C- and N-terminal analysis. The preparation procedure consisted of solubilisation using the non-ionic detergent cutscum, (NH4)2SO4 precipitation, ion-exchange chromatography on DEAE-cellulose and cellulose phosphate and hydrophobic chromatography on butyl-sepharose. The product was detergent-free, had a relatively high content of hydrophobic amino acids and tended to aggregate in aqueous solutions. The protein had a minimum molecular weight of 49,000 ± 500 with a sedimentation coefficient of S20w≃ 3. Antibodies raised against the homogeneous preparation precipitated the entire hydratase activity in solubilised microsomes towards benzo(a)pyrene 4,5-(K-region-)oxide and styrene oxide as substrates. Moreover, the pure enzyme had a very broad substrate specificity, hydrating both arene and alkene oxides. The same general relationship between the hydration velocities of 6 K-region epoxides of polycyclic hydrocarbons was obtained with rat liver microsomal fractions and pure epoxide hydratase. These findings suggest that the enzyme isolated is the only one present in the rat liver microsomal fractions which is responsible for the hydration of such substrates.