Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.2.1.31 (beta-glucuronidase)
7,680 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The metabolism of benzo[a]pyrene (BP) by hamster embryo cells was studied. The production of water-soluble metabolites, shown to be conjugates with glucuronic acid, depended on BP concentration. With increased BP concentration the amount of glucuronic acid conjugates increased, but the proportion of conjugates in BP or its metabolites present in the medium decreased. The metabolites extracted with ethylacetate were trans-7,8-dihydrodiol-BP (7,8-dihydrodiol) and trans-9,10-dihydrodiol-BP (9,10-dihydrodiol), but large peaks of phenolic metabolites were found by high pressure liquid chromatography (HPLC) after digesting the medium with beta-glucuronidase. Therefore, BP is metabolized to oxygenated forms, and of these, most of the phenolic metabolites and parts of the dihydrodiols are conjugated with glucuronic acid. The proportions of dihydrodiols to phenols, estimated by HPLC after beta-glucuronidase digestion, decreased when the BP concentration was decreased. The results suggest that dihydrodiols are less readily glucuronidated than phenols and so may be metabolized further to metabolites other than glucuronic acid conjugates.
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PMID:Metabolism of benzo[A]pyrene in hamster embryo cells. Effect of the concentration of benzo[A]pyrene on its metabolism. 46 31

Various in vitro-inhibitors were added with 3H-benzo(a)pyrene (BP) into the perfusion fluids in isolated rat lung perfusions to see whether their effects are dependent on the integrity of tissue. 3H-BP and its metabolites were measured by thin-layer chromatography and radiometry from both samples of perfusion medium and homogenates of lung tissue. The total covalent binding to lung tissue was used as a measure of the formation of reactive metabolites. In methylcholanthrene-induced rat lung, the metabolism of BP was inhibited by alpha-naphthoflavone, an inhibitor of monooxygenase, and less with diethylmaleate, a depletor of glutathione, with salicylamide, an inhibitor of conjugases, and, astonishingly, with D-saccharo-1,4-lactone, an inhibitor of beta-glucuronidase. With trichloropropene oxide, which inhibits epoxide hydratase, the metabolism was either decreased or unchanged. Nicotine had no effect on BP-metabolism. Nicotine and diethylmaleate increased statistically significantly and alpha-naphthoflavone and salicylamide decreased the covalent binding of radioactivity to lung tissue. In most cases, the changes in BP metabolism observed during perfusion can be explained on the basis of effects of modifiers on the enzyme systems.
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PMID:Effects of various in vitro--inhibitors of benzo(a)pyrene metabolism in isolated rat lung perfusion. 47 52

beta-Glucuronidase catalyzes the hydrolysis of benzo[a]pyrene-3-glucuronide to 3-hydroxybenzo[a]pyrene. During the enzymatic hydrolysis, a benzo[a]pyrene derivative is formed which binds to DNA to a far greater extent than either the 3-hydroxybenzo[a]pyrene or its glucuronide. These results suggest that conjugates of benzo(a)pyrene may be converted by beta-glucuronidase at intracellular and organ sites distal to the initial sites of oxygenation and conjugation of benzo(a)pyrene to activated intermediates that are possibly carcinogenic.
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PMID:beta-Glucuronidase catalyzed hydrolysis of benzo(a)pyrene-3-glucuronide and binding to DNA. 61 59

Human lung explants have been maintained in vitro for a period of 25 days. Autoradiographic studies indicated that the broncholar epithelial cells, type 2 alveolar epithelial cells, and stromal fibroblasts incorporated 3H-thymidine during the culture. After 7 to 10 days, type 2 cells were the predominant alveolar epithelial cell type. Lamellar inclusion bodies were released from the type 2 cells and accumulated in the alveolar spaces. The metabolism of benzo[alpha]pyrene (BP) in human lung explants cultured for up to 7 days was investigated. Human lung explants had measurable aryl hydrocarbon hydroxylase activity and could metabolize BP into forms that were bound to cellular DNA and protein. Peripheral lung had significantly lower aryl hydrocarbon hydroxylase activity than cultured bronchus but both tissues had similar binding levels of BP to DNA. Radioautographic studies indicated that all cell types in the peripheral lung can metabolize BP. The major ethylacetate extractable metabolites of BP formed by peripheral lung were tetrols and trans-7,8-diol. The primary water-soluble metabolite released with arylsulfatase and beta-glucuronidase was 3-hydroxybenzo[alpha]pyrene.
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PMID:Explant culture of human peripheral lung. I. Metabolism of benzo[alpha]pyrene. 66 Dec 25

The formation of water-soluble metabolites of tritium-labeled benzo[a]pyrene (BP) by cultured hamster embryo cells was studied. The ratio of the radioactivity in the aqueous phase to that in the organic phase increased with the incubation period. After incubation for 48 h with 3.75 nmol/ml of [3H] BP in the medium more than 90% of the 3H-radioactivity was found in the aqueous phase, whereas with 10-fold more BP about half the radioactivity remained in the organic phase. The main metabolites extracted from the medium at 37.5 nmol/ml BP with ethyl acetate by high pressure liquid chromatography (HPLC) were 9,10-diol and 7,8-diol; but after treatment of the medium with beta-glucuronidase the main oxygenated metabolites were phenols, the amount of 9-OH BP being more than that of 3-OH BP. beta-Glucuronidase also released 9,10-diol and 7,8-diol, but most of these diols were in the free form in the medium. The medium from cells treated with 3.75 nmol/ml BP has a quantitatively different profile, and most of the radioactivity obtained by extraction with organic solvent and digestion with beta-glucuronidase was eluted in the regions of phenols. These results show that in hamster embryo cells BP is mainly metabolised to conjugates of phenols with glucuronic acid.
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PMID:Glucuronidation of benzo[a]pyrene in hamster embryo cells. 68 21

Dihydrodiol dehydrogenase (DD; EC 1.3.1.20) purified to homogeneity from rat liver cytosol will catalyze the NAD(P)(+)-dependent oxidation of (+/-)-trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (B[a]P-diol) to yield benzo[a]pyrene-7,8-dione (BPQ). To verify that BPQ is a metabolite of B[a]P-diol in rat liver, an S100 fraction was supplemented with NAD+ and NADP+, and the formation of BPQ was followed by reverse-phase HPLC. The identity of BPQ was established by co-chromatography with an authentic standard (under different solvent conditions) and by RP-HPLC using a diode-array detector which established that the metabolite shared spectral identity with BPQ. The formation of BPQ in the S100 fraction was blocked by either a competitive inhibitor (indomethacin) or a suicide substrate [1-(4-nitrophenyl)-propen-1-ol] for DD, indicating that BPQ was being formed by this enzyme. To assess the contribution of DD to the metabolism of [3H]B[a]P-diol, subcellular fractions obtained from uninduced rat liver were fortified with co-factors to optimize the activity of enzymes that would compete for this proximate carcinogen. Under these conditions, S100 fractions fortified with NAD+ and NADP+ metabolized 25% of the B[a]P-diol, producing 731 +/- 154 pmol of BPQ. In contrast, rat liver microsomes fortified with an NADPH generating system metabolize 75% of the B[a]P-diol producing 2614 +/- 379 pmoles of benzo[a]pyrene-tetrahydrotetrols. Rat liver homogenates (S10) fortified with either uridine diphosphoglucuronic acid or phosphoadenosine phosphosulfate produced 180 +/- 56 and 95 +/- 31 pmoles of conjugates respectively, which were recovered as B[a]P-diol after treatment of the aqueous phase with either beta-glucuronidase or aryl sulfatase. Of the metabolites analyzed BPQ was formed in the second largest amount. These studies show that in uninduced rat liver DD may play a significant role in the metabolism of B[a]P-diol. The metabolic fate of BPQ remains to be determined.
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PMID:Contribution of dihydrodiol dehydrogenase to the metabolism of (+/-)-trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene in fortified rat liver subcellular fractions. 139 42

1. Metabolites and DNA adducts of 3H-benzo(a)pyrene (BaP) formed by isolated hepatocytes from English sole (Parophrys vetulus) in vitro were compared to those in bile and liver of sole exposed i.m. to 3H-BaP. 2. English sole liver was perfused with a collagenase solution and hepatocytes were isolated with greater than 95% viability. Determination of kinetic parameters for metabolism of 3H-BaP showed a Km of 29 +/- 10 microM and an apparent Vmax of 1300 pmol BaP metabolized/10(6) cells per h. 3. Analysis of medium from hepatocyte cultures and bile by ion-pair h.p.l.c. showed significant amounts of radioactivity in regions where glucuronide and glutathione conjugates of BaP metabolites elute. No sulphate conjugates of BaP metabolites were detected. The major unconjugated metabolite formed by hepatocytes was the BaP-9,10-dihydrodiol. 4. Hydrolysis of glucuronide conjugates by beta-glucuronidase and reversed-phase h.p.l.c. analysis of chloroform-soluble metabolites showed the presence of BaP-7,8-dihydrodiol, 1-hydroxyBaP and 3-hydroxyBaP. The identities of these metabolites were confirmed by comparing their fluorescence spectra with those of standard BaP metabolites. 5. Analysis by 32P-postlabelling of the BaP-DNA adducts formed in isolated hepatocytes and liver revealed that major adducts detected are derived from the anti-7,8-dihydrodiol-9,10-epoxideBaP (anti-BaPDE) and syn-BaPDE. 6. Results show that the types of conjugated metabolites and BaP-DNA adducts formed in primary hepatocyte culture were similar to those in bile and liver of English sole exposed to BaP. Thus, isolated hepatocytes from English sole afford a reliable alternative to live fish for studies of the mechanisms of hepatic xenobiotic metabolism and DNA adduct formation in a species shown to be susceptible to induction of hepatocarcinogenesis by PAHs.
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PMID:The metabolism of benzo(a)pyrene by English sole (Parophrys vetulus): comparison between isolated hepatocytes in vitro and liver in vivo. 141 84

Long-chain fatty acids inhibit glucuronidation of benzo(a)pyrene phenols in perfused liver; therefore, this study was designed to investigate interactions of fatty acids with beta-glucuronidase, glucuronosyl transferase, and energy supply. In beta-glucuronidase-deficient C3H/He mice, infusion of oleate (250 microM) increased the release of free benzo(a)pyrene phenols from 14 to 33 nmol/g/h and decreased release of glucuronides into the perfusate from 25 to 17 nmol/g/h. Rates of accumulation of glucuronides in the liver were also diminished from 11 to 4 nmol/g/h after infusion of oleate (250 microM). Fatty acids did not affect the release of benzo(a)pyrene metabolites into bile, and the ratio of free phenol to glucuronide production was increased from 0.57 to 1.30. A similar trend was observed in livers from DBA/2 mice that have beta-glucuronidase. Rates of hydrolysis of benzo(a)pyrene-O-glucuronide were not altered in isolated microsomes by addition of oleoyl coenzyme A (CoA) or octanoyl CoA (10- approximately 100 microM). Thus, we conclude that fatty acids do not alter glucuronidation by acting on beta-glucuronidase. The concentration of cofactors (UDP-glucuronic acid, UDP-glucose, and adenine nucleotides) involved in hepatic conjugation was not altered by infusion of concentrations of oleate (300 microM) that inhibited glucuronidation in perfused livers. When oleate concentrations were increased to 600 microM, UDP-glucuronic acid and UDP-glucose decreased 44 and 49%, respectively, and the ATP:ADP ratio declined concomitantly. Oleoyl CoA inhibited UDP-glucuronosyl transferase noncompetitively (half-maximal inhibition, 10 microM) in microsomes with 3-hydroxy-benzo(a)pyrene or p-nitrophenol as substrate. In contrast, octanoyl CoA was a very poor inhibitor of transferase activity. Inhibition of the transferase by oleoyl CoA was increased markedly by treatment with detergents (Triton X-100), i.e., half-inhibition of glucuronosyl transferase was obtained with about 2 microM oleoyl CoA. Inhibition of UDP-glucuronosyl transferase by oleoyl CoA was also increased in a dose-dependent manner by albumin, possibly due to increasing access of the CoA derivative to the enzyme. Collectively, these data indicate that fatty acids diminish glucuronidation via the formation of acyl CoA compounds that inhibit UDP-glucuronosyl transferase noncompetitively.
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PMID:Inhibition of glucuronidation of benzo(a)pyrene phenols by long-chain fatty acids. 190 48

1. Orally administered 3H-benzo[a]pyrene (3H-BaP) was excreted in the bile of White Suckers predominantly as water soluble metabolites some of which were hydrolyzed by arylsulfatase or beta-glucuronidase. 2. Non-hydrolysible polar metabolites comprised a substantial proportion of biliary metabolites. 3. HPLC analysis revealed fluorescent and 3H-labelled peaks which co-eluted with standards of the glucuronide and sulfate conjugates of BaP. 4. The most polar peak co-chromatographed with a double-radiolabelled metabolite produced in vitro with 3H-BaP and 35S-glutathione. 5. Inhibition of epoxide hydrolase in vitro reduced all water soluble metabolites except the glutathione conjugate of BaP. 6. Glutathione conjugation represents a major hepatic detoxication pathway of BaP in White Suckers.
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PMID:The role of glutathione S-transferases in the hepatic metabolism of benzo[a]pyrene in white suckers (Catostomus commersoni) from polluted and reference sites in the Great Lakes. 197 53

Two monoclonal antibodies (10C10 and 4D5) have been developed from the spleen cells of Balb/c mice immunized with 6-aminobenzo[a]pyrene covalently coupled to bovine serum albumin. These antibodies have been used in an immunoassay for the detection of benzo[a]pyrene and its metabolites in mouse urine. The antibodies were characterized in terms of sensitivity and specificity by competitive enzyme-linked immunosorbent assay (ELISA). With both antibodies, 50% inhibition of antibody binding is at 4 pmol of BP. The antibodies also cross-react with a number of BP metabolites as well as with several other polycyclic aromatic hydrocarbons (PAHs) including pyrene, 1-aminopyrene, and 7,12-dimethylbenz[a]anthracene but with different sensitivities. These results suggest that this assay will detect multiple PAH metabolites in urine. To test the assay on biological samples, mice were treated with [3H]BP, and urine was collected and digested with beta-glucuronidase and aryl sulfatase. Several methods were used to isolate BP and its metabolites from the urine, including ethyl acetate extraction, Sep-pak C18 cartridge chromatography, XAD2 resin chromatography, and immunoaffinity chromatography with antibody 4D5. Analysis of the urine extracts with antibody 4D5 gave 50% inhibition at 12-15 pmol of metabolites. Thus, quantitation of metabolites in this sample by competitive ELISA against a standard curve of BP would have underestimated actual metabolite levels by about 70%. This assay will be applied to the analysis of urines from individuals with environmental or occupational exposure. Since humans are usually exposed to BP in complex mixtures of PAHs, multiple metabolites may be present in the urine, making absolute quantitation difficult. This assay should thus serve as a general indicator of exposure to this class of chemicals.
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PMID:Immunologic methods for the detection of benzo[a]pyrene metabolites in urine. 213 77


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