Enzyme biosensors for urea determination based on an ionophore free pH membrane electrode

A new type of enzymatic sensors for the determination of urea based on an ionophore-free pH membrane electrode has been constructed. The membrane material was polyvinyl chloride modified with 1,4-diaminobutane. Chemical urease immobilization enabled the formation of a mono- and bimolecular enzyme layer directly on the pH sensor surface. The influence of the immobilization method modifications on the biosensor properties was investigated. The most important features of the obtained sensors were high sensitivity, long lifetime (over 1 month) and short response time (2–5 min for steady state). The characteristics of the sensors were examined at different buffer concentrations, pH values and stirring rates. The results obtained were in good agreement with theoretical predictions. Optimal measurement conditions were evaluated (a buffer concentration of 5 mM and pH 6) for the urea concentration working range from 0.1 to 30 mM.