Polyelectrolyte-coated alginate microspheres for optical urea sensing

An optical biosensor based on urease immobilized alginate microspheres and its application for determination of urea levels in the dialysate is described. Ammonium ions liberated in urease catalyzed hydrolysis of urea lead to an increase in the pH of the medium, thereby causing the color of a pH sensitive dye to change. In this work, cresol red, immobilized in polyelectrolyte multilayers assembled on alginate microspheres via layer-by-layer self-assembly technique, has been utilized for pH sensing. Measurements were made at wavelengths 434 and 572 nm allowing ratiometric analysis. The response characteristics of the microspheres with the pH sensitive dye and the effects of experimental conditions (on stability of the enzyme) have been studied in detail. Urease enzyme displayed an average residual activity of ~65 % at end of 4 weeks of storage upon immobilization in alginate microspheres. The resulting biosensor allows rapid determination of urea over a range of 0.01 to 6.7 mM and display a response time of ~2 min. Analysis of urea in dialysate samples with this optical sensor yielded results that were comparable to that determined by Nessler´s reagent method (one of the standard colorimetric methods for urea determination). These results demonstrate the potential to develop an optical urea sensor that is stable over a month.