RESUMO
To test the hypothesis that a combination of high salivary nitrate and high nitrate-reducing capacity are protective against dental caries, 209 children attending the Dental Institute, Barts and The London NHS Trust were examined. Salivary nitrate and nitrite levels, counts of Streptococcus mutans and Lactobacillus spp., and caries experience were recorded. Compared with control subjects, a significant reduction in caries experience was found in patients with high salivary nitrate and high nitrate-reducing ability. Production of nitrite from salivary nitrate by commensal nitrate-reducing bacteria may limit the growth of cariogenic bacteria as a result of the production of antimicrobial oxides of nitrogen, including nitric oxide.
Assuntos
Cárie Dentária/prevenção & controle , Lactobacillus/crescimento & desenvolvimento , Nitrato Redutases/metabolismo , Nitratos/análise , Saliva/química , Streptococcus mutans/crescimento & desenvolvimento , Adolescente , Anti-Infecciosos/análise , Criança , Pré-Escolar , Contagem de Colônia Microbiana , Índice CPO , Cárie Dentária/microbiologia , Feminino , Análise de Alimentos , Humanos , Lactente , Masculino , Nitrato Redutase , Óxido Nítrico/análise , Nitritos/análise , Saliva/enzimologia , Saliva/microbiologiaRESUMO
Xanthine oxidoreductase (XOR) is progressively inactivated while catalysing the reduction of inorganic nitrite to NO by xanthine. Inactivation results from conversion of the enzyme into its desulpho-form. The rate of inactivation increases with nitrite concentration. Similar behaviour was shown when NADH replaced xanthine as reducing substrate. A kinetic model is proposed incorporating a 'suicide' inactivation involving an enzyme-substrate (product) complex, rather than inactivation by free NO. The model provides a good fit to progress curves of the reaction of xanthine or NADH with nitrite in the presence of the oxidase or dehydrogenase forms of the enzyme. Inorganic nitrate, like nitrite, was shown to be reduced at the molybdenum site of XOR. With xanthine as reducing substrate, nitrite was produced in essentially a 1:1 stoichiometric ratio with respect to urate. Unlike the case of nitrite, the enzyme was not significantly inactivated, implying that inactivation during nitrite reduction depends on the presence of nascent NO in its enzyme complex.
Assuntos
Óxido Nítrico/metabolismo , Nitritos/metabolismo , Xantina Desidrogenase/antagonistas & inibidores , Xantina Desidrogenase/metabolismo , Xantina Oxidase/antagonistas & inibidores , Xantina Oxidase/metabolismo , Animais , Sítios de Ligação , Catálise , Bovinos , Cinética , Modelos Químicos , Molibdênio/química , Oxirredução , Xantina/metabolismo , Xantina Desidrogenase/química , Xantina Oxidase/químicaRESUMO
Xanthine oxidoreductase catalyses the anaerobic reduction of glyceryl trinitrate (GTN), isosorbide dinitrate and isosorbide mononitrate to inorganic nitrite using xanthine or NADH as reducing substrates. Reduction rates are much faster with xanthine as reducing substrate than with NADH. In the presence of xanthine, urate is produced in essentially 1:1 stoichiometric ratio with inorganic nitrite, further reduction of which is relatively slow. Organic nitrates were shown to interact with the FAD site of the enzyme. In the course of reduction of GTN, xanthine oxidoreductase was progressively inactivated by conversion to its desulpho form. It is proposed that xanthine oxidoreductase is one of several flavoenzymes that catalyse the conversion of organic nitrate to inorganic nitrite in vivo. Evidence for its further involvement in reduction of the resulting nitrite to nitric oxide is discussed.
Assuntos
Nitratos/metabolismo , Xantina Desidrogenase/metabolismo , Xantina Oxidase/metabolismo , Anaerobiose/fisiologia , Animais , Catálise , Bovinos , Flavina-Adenina Dinucleotídeo/metabolismo , Molibdênio/metabolismo , Nitritos/análise , Oxirredução , Xantina/metabolismoRESUMO
Iodonium compounds, especially diphenylene iodonium and iodonium diphenyl are used extensively as inhibitors of NADH-ubiquinone reductase and NADPH oxidase activity. Here, the use of a new iodonium compound, phenoxaiodonium is reported. The IC(50) of neutrophil superoxide production, measured using the superoxide dismutase inhibitable rate of cytochrome c reduction, was approximately 0.75 microM, while 50% inhibition of mitochondrial respiration, measured by the rate of oxygen uptake using a Clark type oxygen electrode, was at approximately 20 microM. The inhibition of oxidation of xanthine to urate by xanthine oxidase was also studied, giving a K(i) of 0.2 microM. Inhibition of nitric oxidase synthase (NOS: from rat brain) by 0.2 microM phenoxaiodonium was equivalent to 1 mM N(G)-nitro-L-arginine methyl ester HCl (L-NAME), that is total abolition of activity. We conclude that phenoxaiodonium is an extremely good inhibitor of flavo-enzymes, but like diphenylene iodonium and iodonium diphenyl, will be of limited use as a pharmacological tool for the elucidation of the involvement of such enzymes in specific cellular functions.
Assuntos
Inibidores Enzimáticos/farmacologia , Flavoproteínas/antagonistas & inibidores , Oniocompostos/farmacologia , Animais , Bovinos , Relação Dose-Resposta a Droga , Transporte de Elétrons/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase/metabolismo , Oniocompostos/química , Oxigênio/farmacocinética , Superóxidos/metabolismo , Suínos , Fatores de Tempo , Xantina Oxidase/antagonistas & inibidores , Xantina Oxidase/metabolismoRESUMO
Peroxynitrite, a potent oxidising, nitrating and hydroxylating agent, results from the reaction of nitric oxide with superoxide. We show that peroxynitrite can be produced by the action of a single enzyme, xanthine oxidoreductase (XOR), in the presence of inorganic nitrite, molecular oxygen and a reducing agent, such as pterin. The effects of oxygen concentration on peroxynitrite production have been examined. The physiologically predominant dehydrogenase form of the enzyme is more effective than the oxidase form under aerobic conditions. It is proposed that XOR-derived peroxynitrite fulfils a bactericidal role in milk and in the digestive tract.
Assuntos
Nitratos/metabolismo , Xantina Desidrogenase/fisiologia , Xantina Oxidase/fisiologia , Animais , Bovinos , Relação Dose-Resposta a Droga , Compostos Inorgânicos/metabolismo , Cinética , Leite/enzimologia , Modelos Químicos , Nitratos/síntese química , Nitritos/metabolismo , Oxirredução , Oxigênio/metabolismo , Pterinas/metabolismo , Superóxido Dismutase/farmacologiaRESUMO
Xanthine oxidase (XO) was shown to catalyze the reduction of isoamyl and isobutyl nitrites to nitric oxide (NO) in the presence of xanthine under anaerobic conditions. NO was produced at a stoichiometric ratio of 2:1 versus urate generation, steady-state analysis of which showed Michaelis-Menten kinetics with xanthine as varied substrate and substrate inhibition with varied organic nitrite. Under the conditions of NO generation from isoamyl nitrite, XO was progressively inactivated by a mechanism involving conversion of Mo=S to Mo=O, yielding "desulfo" enzyme. It is proposed that XO is involved in the metabolism of organic nitrites to NO in vivo and that the observed inactivation serves to explain the phenomenon of tolerance.
Assuntos
Óxido Nítrico/metabolismo , Nitritos/metabolismo , Vasodilatadores/farmacocinética , Xantina Oxidase/química , Xantina Oxidase/metabolismo , Animais , Biotransformação , Bovinos , Cinética , Leite/enzimologia , Ácido Úrico/análiseRESUMO
Xanthine oxidase (XO) was shown to catalyze the reduction of nitrite to nitric oxide (NO), under anaerobic conditions, in the presence of either NADH or xanthine as reducing substrate. NO production was directly demonstrated by ozone chemiluminescence and showed stoichiometry of approximately 2:1 versus NADH depletion. With xanthine as reducing substrate, the kinetics of NO production were complicated by enzyme inactivation, resulting from NO-induced conversion of XO to its relatively inactive desulfo-form. Steady-state kinetic parameters were determined spectrophotometrically for urate production and NADH oxidation catalyzed by XO and xanthine dehydrogenase in the presence of nitrite under anaerobic conditions. pH optima for anaerobic NO production catalyzed by XO in the presence of nitrite were 7.0 for NADH and =6.0 for xanthine. Involvement of the molybdenum site of XO in nitrite reduction was shown by the fact that alloxanthine inhibits xanthine oxidation competitively with nitrite. Strong preference for Mo=S over Mo=O was shown by the relatively very low NADH-nitrite reductase activity shown by desulfo-enzyme. The FAD site of XO was shown not to influence nitrite reduction in the presence of xanthine, although it was clearly involved when NADH was the reducing substrate. Apparent production of NO decreased with increasing oxygen tensions, consistent with reaction of NO with XO-generated superoxide. It is proposed that XO-derived NO fulfills a bactericidal role in the digestive tract.