A sensitive fluorometric assay for measuring xanthine dehydrogenase and oxidase in tissues.
Free Radic Biol Med
; 6(6): 607-15, 1989.
Article
em En
| MEDLINE
| ID: mdl-2753392
ABSTRACT
The conversion of xanthine dehydrogenase to a free radical producing oxidase is an important component of oxygen-mediated tissue injury. Current assays for these enzymes are of limited sensitivity, making it difficult to analyze activities in organ biopsies or cultured cells. The xanthine oxidase-catalyzed conversion of pterin (2-amino-4-hydroxypteridine) to isoxanthopterin provides the basis for a fluorometric assay which is 100-500 times more sensitive than the traditional spectrophotometric assay of urate formation from xanthine. Enzyme activity as low as 0.1 pmol min-1 ml-1 can be measured with the fluorometric pterin assay. Xanthine oxidase is assayed in the presence of pterin only, while combined xanthine dehydrogenase plus oxidase activity is determined with methylene blue which replaces NAD+ as an electron acceptor. The relative proportions and specific activities of xanthine oxidase and dehydrogenase determined by the fluorometric pterin assay are comparable with the spectrophotometric measurement of activities present in rat liver, intestine, kidney, and plasma. The assay has been successfully applied to brain, human kidney, and cultured mammalian cells, where xanthine dehydrogenase and oxidase activities are too low to detect spectrophotometrically.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Xantina Desidrogenase
/
Xantina Oxidase
/
Cetona Oxirredutases
Tipo de estudo:
Diagnostic_studies
Limite:
Animals
/
Female
/
Humans
Idioma:
En
Revista:
Free Radic Biol Med
Assunto da revista:
BIOQUIMICA
/
MEDICINA
Ano de publicação:
1989
Tipo de documento:
Article