Reagentless chemiluminescence biosensor for determination of hydrogen peroxide based on the immobilization of horseradish peroxidase on biocompatible chitosan membrane

A reagentless chemiluminescence (CL) flow sensor is developed for the determination of hydrogen peroxide. The analytical reagent luminol was immobilized on anion-exchange resins and packed in a glass tube, the horseradish peroxidase (HRP) was immobilized on biocompatible chitosan (CHIT) membrane which was formed on the glass coil to construct a transparent sensing flow cell. The two segments are integrated to fabricate a reagentless flow biosensor. The properties of CHIT, HRP and CHIT-HRP films have been carefully characterized by Fourier transform infrared (FT-IR). The results indicate that HRP retains the essential feature of its native structure in the CHIT film. The sample throughput is 120 h(-1) with small sample volumes (30 mul). The two linear working regions are 1.0 x 10(-7) to 1.0 x 10(-5) mol l(-1) and 1.0 x 10(-5) to 2.0 x 10(-4) mol l(-1). The detection limit is 4.0 x 10(-8) mol l(-1) (S/N = 3) with a relative standard deviation of 1.1% for the 4.0 x 10(-6) mol l(-1) H2O2 solution in 31 repeated measurements. The sensor could be used for 3 months. (C) 2002 Elsevier Science B.V. All rights reserved.