A new amperometric urea biosensor based on gold nanoparticle embedded poly(propylene-co-imidazole) was developed for the determination of urea. The urease adsorbed on the polymeric film catalyzed the hydrolysis of urea to ammonium and bicarbonate ions and the ammonium was then electrooxidized on the gold electrode with the aid of gold nanoparticles at +0.2 V versus Ag/AgCl using differential pulse voltammetry. The biosensor provided a linear current response to urea concentration from 0.1 to 30 mM, a detection limit of 36 mu M, a relative standard deviation of 2.43% (n = 18), and excellent storage stability, as the current decrease was only 3% after 75 days. The operation of the biosensor was evaluated by the analysis of municipal sewage wastewater collected from the inlet pipe of the treatment plant of Zonguldak City in Turkey. The effects of possible interferants were also characterized.