KINETIC ENZYME ASSAY FOR DETERMINING THE CO2 CONTENT OF BODY FLUIDS USING PEP CARBOXYLASE WITH INHIBITOR

摘要

An improved kinetic assay is provided for spectrophotometrically determining dissolved carbon dioxide (CO2) in body fluid wherein at pH 8.0 the CO2 is present substantially as bicarbonate ion (HCO3). The sample is subjected to a reaction mixture containing phosphoenolpyruvate (PEP) and phosphoenolpyruvate carboxylase (PEPC) which enzymatically catalyzes conversion of HCO3 to oxaloacetate (OA). The OA produced is subject to a second reaction mixture containing NADH and malate dehydrogenase (MDH) which reduces the OA to malate and oxidizes the NADH to NAD+. The CO2 level in the sample may be indirectly determined by measuring the change in NADH concentration. An inhibitor is added to the reaction mixture which renders the reaction kinetics of the system essentially first order over the entire bicarbonate concentration range of interest. Best results have been obtained through use of an inhibitor which provides an anion selected from the group consisting of Cl03, Cl04, SCN, or I.