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P D Josephy

Publications and source records attributed to P D Josephy.

At least 73 records · Page 4Linked to original sources
PubMed1985

Peroxidase-catalyzed benzidine binding to DNA and other macromolecules.

[14C]Benzidine is rapidly oxidized by a peroxidase/H2O2 system to products which bind irreversibly to DNA. The presence of exogenous DNA also prevented benzidine polymerization to 'benzidine brown' and azobenzidine. Two molar equivalents of H2O2 were required to oxidize the benzidine and achieve maximal DNA binding. Furthermore, 95% of the benzidine was trapped and 36 nmol benzidine was bound per mg DNA. Polyriboguanylic acid was as effective as DNA in binding benzidine, but polyriboadenylic acid, polyribouridylic acid and polyribocytidylic acid were much less effective. Binding of [14C]benzidine correlated well with the absorbance at 295 nm and 390 nm of the modified DNA or various synthetic homopolymers of ribonucleotides isolated from the reaction mixture. The peroxidase/H2O2 system also catalyzed the binding of dichlorobenzidine, o-tolidine and o-dianisidine to DNA but 3,5,3',5'-tetramethylbenzidine, a non-carcinogen, did not bind. The binding could be prevented by various biological hydrogen donors, thiols, or phenolic antioxidants. The mechanisms for DNA protection were investigated; the oxidized benzidine species involved in binding can be reduced with ascorbate, NADPH, or thiols, and trapped by thiols or phenolic antioxidants to form conjugates or adducts.

Animals↗
PubMed1985

Oxidative activation of benzidine and its derivatives by peroxidases.

Benzidine (4,4'-diaminobiphenyl) is a known human carcinogen; exposure to this substance resulted in an epidemic of bladder cancer among workers in the dye industry in Europe and North America. The chemical or enzymatic oxidation of benzidine proceeds via a racial cation detectable by electron spin resonance. Peroxidase-catalyzed oxidation of benzidine generates reactive electrophiles which readily form adducts with phenol and thiol compounds. The structures of these novel metabolites are described. Peroxidases, including prostaglandin synthase, catalyze benzidine binding to protein and nucleic acid; the nature of the resulting adducts is unknown. The relevance of these processes to benzidine carcinogenesis in vivo is the subject of research and debate. A central question remains: is benzidine activated in extra-hepatic target tissues such as bladder epithelium, or transported to these tissues following hepatic oxidative metabolism?

Animals↗
PubMed1983

Oxidation of p-aminophenol catalyzed by horseradish peroxidase and prostaglandin synthase.

p-Aminophenol oxidation is catalyzed by horseradish peroxidase. ESR spectroscopy demonstrates the formation of the p-aminophenoxy free radical, the one-electron oxidation product of p-aminophenol. The same radical is formed by alkaline autoxidation of this compound. The ultimate products of p-aminophenol oxidation are mainly polymeric; however, indophenol was isolated in low yield. Oxidation of p-aminophenol is also catalyzed by the hydroperoxidase activity of prostaglandin synthase, using ram seminal microsomal preparations as the enzyme source.

Aminophenols↗
PubMed1983

An electron spin resonance study of the activation of benzidine by peroxidases.

The oxidation of benzidine, a carcinogenic aromatic amine, by H2O2 is catalyzed by horseradish peroxidase or lactoperoxidase. The resulting cation free radical is moderately stable at pH 5.0, and was identified by electron spin resonance spectroscopy. Two-electron oxidation yields the benzidine di-imine. This species reacts with phenol or catechol derivatives to give colored adducts. Monoacetylbenzidine is a relatively poor peroxidase substrate, and the biological implications of this difference are discussed.

Benzidines↗
PubMed1982

The horseradish peroxidase-catalyzed oxidation of 3,5,3',5'-tetramethylbenzidine. Free radical and charge-transfer complex intermediates.

Benzidine and related compounds are well known substrates for horseradish peroxidase/H2O2 oxidation. Typically, two different colored products are formed. In this paper, we study the oxidation of 3,5,3',5'-tetramethylbenzidine. The first colored product is a blue charge-transfer complex of the parent diamine and the diimine oxidation product. This species exists in rapid equilibrium with the radical cation. The radical was observed by ESR spectroscopy, and hyperfine splitting constants were determined. Addition of equimolar hydrogen peroxide yields the yellow diimine, which is stable at acid pH. At less than equimolar peroxide, all four species (diamine, radical cation, charge-transfer complex, and diimine) exist in equilibrium. A theoretical analysis of this redox system is presented, including a determination of the extinction coefficients and equilibrium constant for the nonradical species.

Benzidines↗
PubMed1982

Ocular absorption and toxicity of a radiosensitizer and its effect on hypoxic cells.

The recurrence of retinoblastoma after radiation treatment may be related to hypoxic cell radioresistance. Radiosensitizing drugs acting on hypoxic cells without affecting the response of oxygenated cells could improve treatment of ocular tumors while minimizing complications of radiotherapy. A desmethyl derivative of misonidazole is as effective a radiosensitizer as misonidazole (as measured in vitro) and is better suited to ocular administration, since it is more soluble than misonidazole. We studied the ocular toxic effects of a desmethyl derivative of misonidazole after subconjunctival administration of 140 and 70 mg. The higher dose produced an intolerable ocular toxic effect, but at the lower dose, the toxic effect was moderate and reversible. We compared ocular pharmacokinetics of the desmethyl derivative of misonidazole after subconjunctival and intravenous (IV) injections. Subconjunctival administration yielded vitreous and anterior chamber concentrations of the radiosensitizer sufficient to produce a notable dose-modifying effect (as high as 1.8 in the anterior chamber and 1.25 in the vitreous at 70-mg doses). In contrast, even at doses of 140 mg, IV injections of the desmethyl derivative of misonidazole did not result in therapeutically useful ocular levels.

Animals↗
PubMed1982

Chemical structure of the adducts formed by the oxidation of benzidine in the presence of phenols.

Bioactivation of carcinogens by peroxidases has received increasing attention since the discovery of the oxidation of carcinogens by prostaglandin hydroperoxidase. Benzidine and 3,5,3',5'-tetramethylbenzidine are oxidized by horseradish peroxidase and prostaglandin synthase to two-electron oxidation products (di-imines). Di-imines readily react with the phenolic anti-oxidant butylated hydroxyanisole to form adducts. In this paper, we have studied the oxidation of benzidine by horseradish peroxidase in the presence of phenolic compounds and characterized the resultant benzidine/phenol adducts. A benzidine/2,6-dimethylphenol adduct was isolated and characterized by mass spectrometry and high field n.m.r. The reaction of [14C]benzidine in the presence of horseradish peroxidase and phenol yielded only the benzidine/phenol adduct. Our results indicate that the benzidine/phenol adducts are analogous to the indoaniline dyes, differing only in substitution of a biphenyl group for a benzene ring. The reaction of benzidine di-imine with endogenous phenols may represent a new pathway for detoxication, removing potentially harmful metabolites of benzidine.

Benzidines↗
PubMed1982

Cooxidation of the clinical reagent 3,5,3'5'-tetramethylbenzidine by prostaglandin synthase.

Prostaglandin synthase catalyzes the oxidation of arachidonic acid to prostaglandin H2 via a hydroperoxide intermediate, prostaglandin G2. The prostaglandin synthase system cooxidizes 3,5,3'5'-tetramethylbenzidine (TMB), a derivative of the human carcinogen benzidine, to colored products. This process is arachidonic acid dependent and indomethacin sensitive. The reaction is also supported by hydroperoxides, and, in this case, indomethacin has no effect. This suggests that the cooxidation is mediated by the hydroperoxidase activity of prostaglandin synthase. The products of TMB oxidation by this system are the same as those obtained with lactoperoxidase and H2O2 or with horseradish peroxidase and H202. The initial products of TMB oxidation are the TMB radical cation and a charge-transfer complex composed of TMB and its two-electron (diimine) oxidation product. The TMB radical cation was identified by electron spin resonance spectroscopy. Ascorbic acid reduces the products, regenerating the parent compound.

Benzidines↗
PubMed1981

In vitro metabolism of misonidazole.

Misonidazole (MISO) is a nitroimidazole drug currently undergoing clinical trials as a radiosensitizer of hypoxic tumour cells. The drug is also toxic to such cells, probably because of metabolic activation of the nitro group under hypoxia. The metabolic fate of 14C-labelled MISO is examined, using hypoxic mammalian (CHO) cells in vitro. Organic-soluble and acid-soluble metabolites are formed, and radioactivity is bound to macromolecules. The organic-soluble products are separated by TLC and HPLC. Evidence is presented to show that one of the metabolites is hydroxylamino-misonidazole. The significance of metabolic nitroreduction is discussed.

Animals↗