Potentiometric biosensor for determination of urea in milk using immobilized Arthrobacter creatinolyticus urease

The extracellular urease from Arthrobacter creatinolyticus was partially purified by ammonium sulfate precipitation and immobilized on PAN [poly(acrylonitrile-methylmethacrylate-sodium vinylsulfonate)] membrane. The urease immobilized PAN membrane exhibited an activity of 97.92U/cm(2) under the optimum conditions of 1.0% enzyme concentration, 15% glutaraldehyde, 24h immobilization time and temperature of 4°C. The changes in surface morphology of the membrane after immobilization were studied by SEM and ATR-FTIR analysis. Immobilized membrane was associated with potentiometric electrode for calibration of biosensor and the results showed a linear response for wide range of urea concentration from 1 to 100mM. The immobilized urease had good storage stability for a period of 70days at 4°C and could be effectively reused for 13cycles. Urease immobilized membrane was also employed in analysis of urea spiked milk samples.