Potentiometric study of urease kinetics over pH 5.36-8.21

摘要

To meet the demand for a convenient method of urease assay we present the potentiometric measurement of urease activity with use of an ammonium ion-selective electrode. The membrane of the electrode was plasticized poly(vinyl chloride) containing physically immobilized nonactine as an NH4+-sensitive ionophore and a lipophilic salt as an anionic additive. The calibration in buffers covering pH 5.36-9.97 showed that the electrode had a Nernstian response at pH < 9.17 down to about 0.06 mM NH4+ and that for each buffer the detection limit grew with an increase in pH, concentration and the presence of Na(2)EDTA. The response time of the electrode was 15 s. In the study of urease kinetics performed with use of the electrode over pH 5.36-8.21 we have shown that the Michaelis constant of the enzyme does not significantly change with pH and that the maximum reaction rate-pH profile, unlike those hitherto reported, has two maxima located at a short distance of 0.7 pH unit (7.2 and 6.5). Such a profile may pave the way for a new interpretation of the acid-base mechanism of urease action. The potentiometric method presented can be helpful in screening for urease inhibitors, the substances of importance in therapies of diseases induced by ureolytic bacteria and in agriculture for proper nitrogen management in soil. [References: 52]