Summary: A Spectrophotometric method is described for the determination of 5'-nucleotidase.In combination with the enzymes nucleoside phosphorylase and xanthine oxidase, inosine, formed by hydrolysis of S'-IMP by S'-nucleotidase, is cleaved phosphorolytically to hypoxanthine, which is oxidized to uric acid. In the presence of ethanol, the hydrogen peroxide formed is reduced by catalase and equivalent amounts of acetaldehyde are produced. The aldehyde is dehydrogenated (NADP-dependent) by aldehyde dehydrogenase and the production ι rate of NADPH is recorded at 334 nm. The inhibition of the unspecific cleavage of 5'-IMP by phosphatases is examined critically. Eine neue spektrophotometrische Methode zur Bestimmung von 5'-Nucleotidase lZusammenfassung: Eine spektrophotometrische Methode f r die Bestimmung der S'-Nucleotidaseaktivit t wird be-! schrieben. ;In Kombination mit den Enzymen Nucleosidphosphorylase und Xanthinoxidase wird Inosin, das bei der Hydrolyse i von S'-IMP durch die S'-Nucleotidase entsteht, phosphorolytisch zu Hypoxanthin gespalten, das weiter zu Harns ure j oxidiert wird. Das entstandene Wasserstoffperoxid wird mit Katalase in Gegenwart von Ethanol reduziert. Dabei ent-! stehen quimol fe Mengen an Acetaldehyd. Der Aldehyd wird NADP-abh ngig durch die Aldehyddehydrogenase l dehydriert und das gebildete NADPH bei 334 nm automatisch registriert. Die Inhibierung der unspezifischen S'-IMP-} Hydrolyse wird kritisch untersucht.

A new method for the determination of xanthine oxidase activity with xanthine or hypoxanthine is described. The hydrogen peroxide produced by the oxidation of the substrates is reduced by catalase in the presence of high concentrations of ethanol. The acetaldehyde formed is further oxidized by aldehyde dehydrogenase NAD or NADP-dependent. The reduction rate of the coenzymes were measured at 334 nm and utilized as indicators for the xanthine oxidase. The sensitivity of the method with xanthine as substrate can be doubled by the addition of uricase, which oxidizes uric acid to allantoin.

A new spectrophotometric method for the determination of adenosine deaminase is described. Adenosine is deaminated to inosine, the latter is cleaved by an inosine-guanosine specific nucleoside phosphorylase to hypoxanthine and ribose-1-phosphate. Hypoxanthine can be oxidized further to uric acid by xanthine oxidase or to allantoin by xanthine oxidase and uricase. The hydrogen peroxide formed in these reactions is reduced by catalase to water. In the presence of high concentrations of ethanol, equivalent amounts of acetaldehyde are produced. The acetaldehyde is oxidized NAD(P) dependent and the production rate of NAD(P)H is recorded at 334 nm. The new method is suitable for the detection of adenosine deaminase in whole blood, lymphocytes, sera and tissues.

A new method for the determination of guanase is described. Xanthine, the product of the guanase reaction, is oxidized by xanthine oxidase, forming uric acid and hydrogen peroxide. Hydrogen peroxide is further reduced to water by catalase in the presence of ethanol. The acetaldehyde formed in this reaction step is dehydrogenated NAD or NADP dependent by aldehyde dehydrogenase. The NADH or NADPH production is measured and utilized for the calculation of the guanase activity. The sensitivity of the method can be doubled by the addition of uricase, which oxidizes uric acid to permit the formation of another mole of hydrogen peroxide.

A spectrophotometric method especially suitable for biological materials is described for the determination of purine nucleoside phosphorylase activity. In combination with the enzymes xanthine oxidase, catalase and aldehyde dehydrogenase, and in the presence of ethanol and NAD(P), the purines formed by phosphorylysis of purine nucleosides are oxidized and the absorption of the NAD(P)H formed is taken for the calculation of nucleoside phosphorylase activity.