Peroxidase and toluidine blue-incorporated DNA bio-polyion complex film as biosensing layer for hydrogen peroxide determination

摘要

Horseradish peroxidase (HRP) and toluidine blue (TB) were incorporated in polyion complex membrane composed of double stranded DNA(dsDNA) and chitosan prepared on the surface of an glassy carbon (GC) disk electrode to fabricate highly sensitive and selective reagentless H_2O_2 biosensor. The embedded-TB in the DNA/chitosan membrane exhibited excellent electrochemical redox property with an electron transfer rate constant of 3.12 ± 0.5 sec~(-1), and shuttled electron effectively from the base GC electrode to catalytic center of the HRP. Under the applied potential of -0.22V (versus Ag/AgCl) and pH 7.0, the resulting electrode (HRP/DNA-TB/chitosan/GCE) exhibited rapid (<10 s) and sensitive response to H_2O_2. The calibration curve of H2O_2, plotting steady-state cathodic current versus H_2O_2 concentration, was linear up to 0.1 mM with a detection limit of 1 μM H_2O_2 (S/N = 3). The H_2O_2 response was scarcely interfered by ascorbic acid and uric acid, which potentially reduce oxidized intermediate of the HRP and interfere with the response of peroxidase-based electrodes.