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

The extracellular urease from Arthrobacter creatinotyticus 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 97.92 U/cm(2) under the optimum conditions of 1.0% enzyme concentration, 15% glutaraldehyde, 24 h immobilization time and temperature of 4 degrees 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 100 mM. The immobilized urease had good storage stability for a period of 70 days at 4 degrees C and could be effectively reused for 13 cycles. Urease immobilized membrane was also employed in analysis of urea spiked milk samples. (C) 2015 Elsevier B.V. All rights reserved.