Construction of a Fluorescent H2O2 Biosensor with Chitosan 6-OH Immobilized β-Cyclodextrin Derivatives

In the present work, a fluorescent H2O2 biosensor was constructed by encapsulating fluorescent probe Rhodamine B (RhmB) in the hydrophobic cavity of the cyclodextrin (beta-CD) and immobilizing catalase (CAT) on the 2-NH2 of chitosan (CTS) in a chitosan 6-OH immobilized beta-cyclodextrin derivative (CTS-6-CD). The inclusion complex of CTS-6-CD to RhmB (CTS-6-CD-RhmB) was prepared by a solution method. Its structure and inclusion efficiency were determined by Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), and fluorescence spectroscopy (FL). CAT was immobilized on CTS-6-CD-RhmB to eventually form the functional membrane, CTS-6-CD-RhmB-CAT, via glutaraldehyde crosslinking, which was further characterized by FTIR and FL, and used as a H2O2 biosensor. The functional membrane was used to simultaneously oxidize and detect H2O2. The detection condition was optimized as pH 8, a reaction temperature of 25 degrees C, and an immobilized enzyme concentration of 2 x 10(-4) mol/L. The fluorescence response of the biosensor exhibited a good linear relationship with the concentration of H2O2 in the range of 20 mM-300 mu M and the detection limit of 10(-8) mol/L.