Method for the quantitative enzymatic determination of ADP

US4923796A · kind A · utility

Assignee

• Boehringer Mannheim GmbH · DE

Inventors

• Ulfert Deneke · Fürth, DE
• Gerhard Micharl · Tutzing, DE
• Hans-Otto Beutler · Tutzing, DE

Key dates

• Filing date: Jan 17, 1989
• Grant date: May 8, 1990
• Expiry date: Jan 17, 2009

Classification

• Technology area (CPC Y): Emerging Cross-Sectional Technologies
• CPC primary: Y10S435/81
• WIPO field: Biotechnology
• WIPO sector: Chemistry

Abstract

In a method for the quantative enzymatic determination of adenosine diphosphate (ADP) in aqueous solution by phosphorylation of the ADP with a kinase in the presence of a phosphorylated substrate therefor, with the formation of ATP and a dephosphorylated substrate, such as galactose, formate, pyruvate or nucleoside diphosphate, the dephosphorylated substrate is reacted, optionally with the insertion of a further adjuvant reaction, such as a decarboxylation, transamination or hexosidiation, with NAD(P) in the presence of a dehydrogenase and the NAD(P)H formed is either measured directly or reacted with a tetrazolium salt in the presence of an electron carrier, with the formation of a formazane, which latter is then measured. A reagent suitable for carrying out this method contains a kinase, a phosphorylated substrate therefor different from ADP, a specific dehydrogenase for the dephosphorylated form of the substrate or for a transaminated, decarboxylated or hexosidized secondary product thereof, the co-enzyme of the dehydrogenase and a buffer, as well as optionally a tetrazolium salt, an electron carrier suitable therefor and optionally a surface-active agent.