RESUMO
Monoamine oxidase B has been purified from human blood platelets 185-fold to a specific activity of 113 nmole/min/mg protein by a combination of Triton X-100 solubilization and ion exchange chromatography. A protein fraction corresponding to 58,000 Da on sodium dodecyl sulfate-polyacrylamide gel electrophoresis was identified as monoamine oxidase by its ability to bind [3H]Pargyline.
Assuntos
Plaquetas/enzimologia , Monoaminoxidase/isolamento & purificação , Cromatografia por Troca Iônica , Eletroforese em Gel de Poliacrilamida , Humanos , Monoaminoxidase/sangue , Octoxinol , PolietilenoglicóisRESUMO
Monoamine oxidase is assayed in tissue by a colorimetric reaction using horse radish peroxidase and 2,2'-azinodi(ethylbenzthiazoline-6-sulfonic acid to measure H2O2 formed during oxidation of amines. The method has a coefficient of variation of approximately 2.5% and provides results comparable with those of radiometric assay. Monoamine oxidase activities in rat liver mitochondria and crude mitochondrial fraction from brain and with tyramine as a substrate were 18.9 +/- 0.4 and 4.61 +/- 0.15 nmol/min/mg of protein, respectively, using this method. Kinetic parameters of liver and brain monoamine oxidase with various substrates and inhibitors appeared to be the same when determined by either colorimetric or radiometric methods.
Assuntos
Compostos Cromogênicos , Isoenzimas , Monoaminoxidase/análise , Peroxidases , Ácidos Sulfônicos , Animais , Azidas , Benzotiazóis , Encéfalo/enzimologia , Colorimetria , Peróxido de Hidrogênio/análise , Mitocôndrias Hepáticas/enzimologia , Oxirredução , Peroxidase , Ratos , Azida Sódica , Especificidade por SubstratoRESUMO
A rapid and sensitive assay for the measurement of monoamine oxidase (MAO) activity was developed. This method utilizes high pressure liquid chromatography with fluorescence excitation at 280 nm and detection of 330 nm at pH 5.0 of indoleacetic acid and 5-hydroxyindoleacetic acid, the deaminated products of two substrates for MAO, tryptamine, and serotonin, respectively. The assay allows for the complete separation of metabolites from either of the two substrates. The method has been used to determine MAO activity in the frontal cortex and caudate nucleus of rat brain using tryptamine and serotonin as substrates. The ease and rapidity of this assay make it sutable for use where routine enzyme determinations are required.
Assuntos
Monoaminoxidase/análise , Animais , Encéfalo/enzimologia , Núcleo Caudado/enzimologia , Córtex Cerebral/enzimologia , Cromatografia Líquida de Alta Pressão , Ácido Hidroxi-Indolacético/análise , Masculino , Ratos , Serotonina/metabolismo , Espectrometria de Fluorescência , Triptaminas/metabolismoRESUMO
The michaelis constants (Vmax and Km) for platelet monoamine oxidase (MAO) with tyramine as substrate are found to be significantly lower in chronic schizophrenic patients than in normal controls. Furthermore, these kinetic parameters for the MAO of paranoid chronic schizophrenics are significantly lower than those for nonparanoid chronic schizophrenics. Paranoid chronic schizophrenia may be a separate biochemical disorder from other chronic schizophrenias.
