Prussian blue modified glassy carbon electrodes - Study on operational stability and its application as a sucrose biosensor

Stabilisation of electrochemically deposited Prussian blue (PB) films on glassy carbon (GC) electrodes has been investigated and an enhancement in the stability of the PB films is reported if the electrodes are treated with tetrabutyl ammonium toluene-4-sulfonate (TTS) in the electrochemical activation step following the electrodeposition. A multi-enzyme PB based biosensor for sucrose detection was made in order to demonstrate that PB films can be coupled with an oxidase system. A tri-enzyme system, comprising glucose oxidase, mutarotase and invertase, was crosslinked with glutaraldehyde and bovine albumin serum on the PB modified glassy carbon electrode. The deposited PB operated as an electrocatalyst for electrochemical reduction of hydrogen peroxide, the final product of the enzyme reaction sequence. The electrochemical response was studied using flow injection analysis for the determination of sucrose, glucose and H2O2. The optimal concentrations of the immobilisation mixture was standardised as 8 U of glucose oxidase, 8 U of mutarotase, 16 U of invertase, 0.5% glutaraldchyde (0.025 mul) and 0.5 % BSA (0.025 mg) in a final volume of 5 mul applied at the electrode surface (0.066 cm(2)). The biosensor exhibited a linear response for sucrose (4-800 muM), glucose (2-800 muM) and H2O2 (1-800 muM) and the detection limit was 4.5, 1.5 and 0.5 muM for sucrose, glucose and H2O2, respectively. The sample throughput was ca. 60 samples h(-1). An increase in the operational and storage stability of the sucrose biosensor was also noted when the PB modified electrodes were conditioned in phosphate buffer containing 0.05 M TTS during the preparation of the PB films. (C) 2004 Elsevier B.V. All rights reserved.