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Biomedical subjects

D R Doerge

Publications and source records attributed to D R Doerge.

At least 73 records · Page 4Linked to original sources
PubMed1991

Determination of ethylenethiourea in crops using particle beam liquid chromatography/mass spectrometry.

Several food crops were analyzed for residues of ethylenethiourea (ETU), a suspect thyroid and liver carcinogen present in EBDC fungicides, using a commercial particle beam (PB) LC/MS method. The PB/LC/MS detection limits for ETU in crops (5 ppb, 1.25 ng) are comparable to those obtained by LC with electrochemical detection. Spectra obtained from crop samples containing as little as 5 ng of ETU were matched with the NBS library reference EI spectrum. Isotopically labeled ETU was used as an internal standard for quantitation and determination of recoveries. No enhancement of molecular ion signal intensity from unlabeled ETU was observed upon coelution with the isotopically labeled variant. This MS method permits detection of ETU with increased selectivity without compromising sensitivity.

Chromatography, Liquid↗
PubMed1988

Mechanism-based inhibition of lactoperoxidase by thiocarbamide goitrogens. Identification of turnover and inactivation pathways.

Direct evidence is presented in support of mechanism-based (suicide) inactivation of lactoperoxidase by thiocarbamide thyroid inhibitors. The turnover of 1-methylbenzimidazolidine-2-thione was demonstrated by identifying the inhibitor-derived products 1-methylbenzimidazole and bisulfite ion that are formed concurrent to enzyme inactivation. The turnover of a hydroperoxide cosubstrate, 5-phenyl-4-pentenyl hydroperoxide, was quantitated from formation of the corresponding alcohol during enzyme inactivation. A specific inactivation pathway is suggested by the covalent binding of 1 mol of 14C- and 35S-labeled benzimidazolidine-2-thione and 1-methylbenzimidazolidine-2-thione per mole of inactivated lactoperoxidase. These results are explained by partitioning of inhibitor-derived S-oxygenated intermediates between turnover and inactivation pathways. The properties of the inactivation process are unique among thiono-sulfur compounds and suggest that benzimidazolinesulfenic acids are the reactive intermediates.

Antithyroid Agents↗
PubMed1988

Hydroperoxide-dependent metabolism of goitrogenic thiocarbamides by lactoperoxidase.

1. Inhibition of lactoperoxidase by thiocarbamides is consistent with a suicide mechanism whereby enzyme-catalysed S-oxygenation produces reactive intermediates which covalently modify the active site haem. 2. The reaction of thiocarbamide goitrogens with lactoperoxidase in the presence of hydroperoxides results in time-dependent and irreversible enzyme inactivation and an altered visible spectrum of the haem prosthetic group of the inactivated enzyme. 3. A mechanism of S-oxygenation for the inactivation is suggested by lactoperoxidase-catalysed formation of stable S-oxides from thioamide and organosulphur functional groups, and by a common dependence of substrate and inhibitor binding constants on their electrochemical oxidation potentials. 4. Hydroperoxide-dependent inactivation of lactoperoxidase by benzimidazoline-2-thiones occurs concomitantly to the covalent binding of stoichiometric amounts of 14C- or 35S-labelled inhibitors per mole of enzyme, and the formation of turnover products derived from the hydroperoxide cosubstrate and inhibitor.

Animals↗
PubMed1986

Mechanism-based inhibition of lactoperoxidase by thiocarbamide goitrogens.

The irreversible inactivation of bovine lactoperoxidase by thiocarbamide goitrogens was measured, and the kinetics were consistent with a mechanism-based (suicide) mode. Sulfide ion inactivated, 2-mercaptobenzimidazole-inactivated, and 1-methyl-2-mercaptoimidazole-inactivated lactoperoxidases have different visible spectra, suggesting different products were formed. The results support a mechanism in which reactive intermediates are formed by S-oxygenation reactions catalyzed by lactoperoxidase compound II. It is proposed that the reaction of electron-deficient intermediates with the heme prosthetic group is responsible for the observed spectral changes and inactivation by thiocarbamides.

Animals↗
PubMed1986

Oxygenation of organosulfur compounds by peroxidases: evidence of an electron transfer mechanism for lactoperoxidase.

The oxygenation of benzyl methylsulfide, thioanisole, and thiobenzamide to the respective sulfoxides was found to be catalyzed by chloroperoxidase, lactoperoxidase, and horseradish peroxidase. The activities of lactoperoxidase and horseradish peroxidase were similarly low toward benzyl methylsulfide and thioanisole but lactoperoxidase efficiently catalyzed the oxygenation of thiobenzamide while horseradish peroxidase showed low activity. Chloroperoxidase had high reactivity toward all three substrates tested in halide-independent reactions and only small differences in the rates of enzymatic sulfoxidation were observed. The logarithm of lactoperoxidase activity was found to linearly correlate with the voltammetric peak potentials for oxidation of the three substrates tested. The results of this study are consistent with a one-electron transfer mechanism for lactoperoxidase-mediated sulfoxidation.

Chloride Peroxidase↗
PubMed1985

The action of alpha-ketoglutarate dehydrogenase on 4-chloronitrosobenzene: evidence for species-dependent differences in active site properties.

The reaction of bovine (Bos taurus) and porcine (Sus scrufa) cardiac alpha-ketoglutarate dehydrogenase complex (alpha-KGD) with 4-chloronitrosobenzene (I) was shown to produce a hydroxamic acid (IV) and a product due to a Bamberger rearrangement as previously shown for Escherichia coli alpha-KGD. The conversion of I into an active site-bound electrophile was general among the three alpha-KGD enzymes tested, but quantitative differences in products and kinetics were shown. The reaction of I was specific for the resolved alpha-ketoglutarate decarboxylase subunit.

Animals↗
PubMed1984

Primary arylamine oxidation by a flavinhydroperoxide. A study of the basis for the substrate specificity of the flavoprotein monooxygenase.

4a-Hydroperoxy-5-ethyl-3,8,10-trimethylisoalloxazine (FlEtOOH) was prepared as a chemical model for the flavin-containing monooxygenase enzyme of mammalian liver. FlEtOOH was found to oxidize a series of para-substituted primary arylamines to the corresponding arylnitroso compound. The rates of arylamine oxidation were found to correlate with the Hammett substituent constant, sigma, as well as with amine basicity. These results suggest that amine nucleophilicity should be an important determinant of flavin monooxygenase reactivity toward primary arylamines; nevertheless, the enzyme demonstrates considerable substrate preference based on other factors.

Amines↗
PubMed1984

Hydroperoxyflavin-mediated oxidations of organosulfur compounds. Model studies for the flavin monooxygenase.

Kinetic and product studies were carried out for the reaction of a synthetic hydroperoxyflavin with a series of organosulfur compounds as a model for the flavin-containing monooxygenase of mammalian liver (FMO). S-Oxidized products were identified, and the kinetics of the oxidation reactions were consistent with a mechanism involving attack of a sulfur nucleophile on the terminal oxygen atom of the hydroperoxyflavin. These results provide information about the substrate oxygenation step of FMO not available from steady-state enzyme kinetics.

Animals↗
PubMed1983

The synthesis and mutagenicity of the N-formyl analog of N-hydroxyphenacetin.

The synthesis and purification of N-hydroxy-N-formyl-p-phenetidine (N-OH-FP) is described. This new compound was subjected to mutagenicity testing using Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 both in the presence and absence of the post-mitochondrial fraction of rat liver homogenate. Simultaneous mutagenicity testing of the known phenacetin metabolite, N-hydroxyphenacetin (N-OH-AP), was conducted with the same tester strains. The N-formyl derived hydroxamic acid (N-OH-FP) was found to be a much stronger mutagen than N-hydroxy-phenacetin (N-OH-AP). Furthermore, N-OH-FP also behaved as a direct-acting mutagen unlike N-OH-AP. The chemical stabilities of N-OH-AP and N-OH-FP were studied in phosphate buffer in the pH range of 3-8; and both the hydroxamic acids were found to be stable to the conditions employed. The results of this study support the hypothesis that enzymatic deacylation is an activation process for the expression of mutagenicity by hydroxamic acids.

Aminophenols↗
PubMed1982

Unsaturated fatty acid-dependent oxidation of epinephrine in homogenates of the gorgonian, Pseudoplexaura porosa.

A novel epinephrine oxidation system in homogenates of the gorgonian Pseudoplexaura porosa was discovered. The enzymatic reaction required an unsaturated fatty acid and molecular oxygen or hydrogen peroxide. Diphenylisobenzofuran was also oxidized by Ps. porosa homogenates in the presence of an unsaturated fatty acid. Hydroxyl radical and superoxide anion did not appear to be involved in either of these oxidative reactions. The production of lipid hydroperoxides was not necessary for epinephrine oxidation and, with the exception of arachidonic acid, lipid hydroperoxide production did not occur. Evidence is presented for the involvement of singlet oxygen or a similar activated oxygen intermediate in the reactions, and a possible mechanism was proposed. The use of arachidonate-dependent epinephrine oxidation as a measure of prostaglandin synthetase activity is criticized.

Animals↗
PubMedDate not supplied

Placental transfer of the soy isoflavone genistein following dietary and gavage administration to Sprague Dawley rats.

Genistein, the principal soy isoflavone, has estrogenic activity and is widely consumed by humans for putative beneficial health effects. The goal of the present study was to measure placental transfer of genistein in rats as a possible route of developmental exposure. Pregnant Sprague-Dawley rats were administered genistein orally, either by diet or by gavage. Concentrations of genistein aglycone and conjugates were measured in maternal and offspring serum and brain using HPLC with isotope dilution electrospray tandem mass spectrometry. Although fetal or neonatal serum concentrations of total genistein were approximately 20-fold lower than maternal serum concentrations, the biologically active genistein aglycone concentration was only 5-fold lower. Fetal brain contained predominately genistein aglycone at levels similar to those in the maternal brain. These studies show that genistein aglycone crosses the rat placenta and can reach fetal brain from maternal serum genistein levels that are relevant to those observed in humans.

Administration, Oral↗
PubMedDate not supplied

Suicide inactivation of lactoperoxidase by 3-amino-1,2,4-triazole.

Amitrole (3-amino-1,2,4-triazole) meets the criteria for a suicide (mechanism-based) inhibitor of lactoperoxidase. Amitrole causes rapid inactivation of lactoperoxidase only in the presence of hydrogen peroxide, and the kinetics are consistent with a suicide mechanism. Approximately 7 mol of radiolabeled amitrole binds covalently per equivalent of lactoperoxidase activity lost. The visible spectrum of lactoperoxidase inactivated by amitrole is unchanged, suggesting that covalent modification of the heme prosthetic group does not occur. The 13C NMR spectrum of lactoperoxidase inactivated by [13C]amitrole shows unique resonances which support the hypothesis that covalent binding occurs on the protein moiety. The similarities between lactoperoxidase and thyroid peroxidase suggest a similar mechanism for inhibition of thyroid hormone synthesis by amitrole.

Amitrole↗