CONDENSATION ALPHA-CYCLODEXTRIN POLYMER MEMBRANE WITH COVALENTLY IMMOBILIZED GLUCOSE-OXIDASE AND MOLECULARLY INCLUDED MEDIATOR FOR AMPEROMETRIC GLUCOSE BIOSENSOR

A highly permeable film of the condensation alpha-cyclodextrin polymer (alpha-CDP) was used as a membrane for fabricating a ''second-generation'' glucose amperometric biosensor. Glucose oxidase (GOD) was covalently immobilized, whereas 1,4-benzoquinone (BQ), or tetrathiafulvalene (TTF) mediator molecules were included in the membrane by the alpha-cyclodextrin sites. A simple one-step procedure for the biosensor preparation was developed. It consisted of casting the membrane onto the surface of a glassy carbon, gold, or platinum disk substrate from an aqueous solution, pH 2.0 (HCl), containing equimolar quantities of a water-soluble alpha-cyclodextrin prepolymer, a glutaric dialdehyde cross-linking reactant, GOD, and the mediator. Cyclic voltammetry (CV) limiting currents for glucose biocatalytic electrooxidation were measured for different enzyme loadings and mediator dosings inside the membrane as well as for different glucose concentrations in a (0.1 M phosphate buffer + 0.1 M NaClO4) pH 7.0 solution. The limiting-electrocatalytic currents at the (alpha-CDP)-GOD-TTF electrode were pH independent and occurred at E(1/2) = 0.03 V (vs. SCE), i.e., at a favorably low potential value. The detectability of glucose was 10 mu M for the (alpha-CDP)-GOD-BQ electrode at the 2 mg (18.5 unit) GOD loading and a signal-to-noise ratio of 3. The values of apparent. Michaelis constant for glucose and the maximum limiting electrocatalytic current density, determined from the Michaelis-Menten analysis for the 1 mg (18.5 unit) GOD loading, were (4.5 +/- 0.5) mM and (190 +/- 20) mu A cm(-2) for the (alpha-CDP)-GOD-BQ electrode, and (5.6 +/- 0.8) mM and (137 +/- 16) mu A cm(-2) for the (alpha-CDP)-GOD-TTF electrode, respectively. The sensor performance was examined with respect to the method of membrane preparation, membrane composition and long-term stability. Miniaturization of the biosensor is facile if, for instance, a 10 mu m Pt disk microelectrode substrate is used.