Date of Award

2001

Document Type

Master Thesis

Degree Name

Masters of Science (Research)

Department

Department of Chemical and Life Sciences

First Advisor

Dr. Marie O'Dowd

Abstract

This thesis is concerned with the immobilisation of enzymes on electrosynthesised poly (o- phenylenediamine) membranes for use on amperometric electrodes. The thesis opens with a discussion of the basic principles of enzymology and a review of a diverse range of biosensors, followed by a consideration of amperometry as governed by the Cottrell equation.

Glucose oxidase and horseradish peroxidase were co-immobilised in poly (o-phenylenediamine) on platinum electrodes at + 0.6 V versus Ag/AgCl. Using this approach the enzymes are entrapped in a strongly adherent, highly reproducible thin membrane, of thickness approximately 10 nm. This one-step procedure produces a glucose sensor with a response time less than 1 second, a high sensitivity of 419.63 nA/mM/cm2, a linear range up to 7 mM and a limit of detection of 0.03 nM glucose. From kinetic analyses, the response of the electrode is rate limited by the enzyme kinetics, exhibiting an apparent Michaelis-Menton constant km =17.03 mM and a limiting current density Imax of 12.8 µA/cm2. The activation energy for the enzymatic reaction was calculated as 7.6 kJ/mol. Due to permselectivity characteristics of the membrane, the access of ascorbate, urate, and acetaminophen, common interfering species, to the electrode surface is blocked. The electrode was found to be stable for more than 30 days.

Using the glucose electrode as a model, a hypoxanthine electrode was prepared by the co-immobilisation of xanthine oxidase and horseradish peroxidase in poly (o-phenylenediamine). This electrode again exhibited a fast response, a sensitivity of 54.2 nA/µM/cm2, a linear range up to 62 µM hypoxanthine and a limit of detection of 0.25 µM hypoxanthine. The kinetic parameters determined for the optimised biosensor were 196.1 µM for the km and 19.9 µA/cm2 for the Imax. The temperature dependence of the amperometric response indicated an activation energy of 22.8 kJ/mol.

Access Level

info:eu-repo/semantics/openAccess

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