Abstract
A potentiometric enzyme biosensor is a convenient detector for quantification of urea concentrations in industrial processes, or for monitoring patients with diabetes, kidney damage or liver malfunction. In this work, poly(3-hexylthiophene-co-3-thiopheneacetic acid) (P(3HT-co-3TAA)) was chemically synthesized, characterized and spin-coated onto conductive indium tin oxide (ITO) glass electrodes. Urease (Urs) was covalently attached to the smooth surface of this copolymer via carbodiimide coupling. The electrochemical behavior and stability of the modified Urs/P(3HT-co-3TAA)/ITO glass electrode were investigated by cyclic voltammetry, and the bound enzyme activity was confirmed by spectrophotometry. Potentiometric response studies indicated that this electrode could determine the concentration of urea in aqueous solutions, with a quasi-Nernstian response up to about 5 mM. No attempt was made to optimize the response speed; full equilibration occurred after 10 min, but the half-time for response was typically <1 min.
Highlights
Enzyme-based electrochemical biosensors have achieved great commercial importance since the first use of glucose oxidase in an amperometric sensor for glucose [1] in 1962
The carbodiimide coupling reaction provided by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) has previously been used to create a peptide bond between amides and carboxylic acids in aqueous solution at room temperature [16]
The semiconducting thiophene copolymer P(3HT-co-3TAA) (1:1) was synthesized and used successfully in a urea biosensor, since it acts as a matrix that can immobilize urease on its surface
Summary
Enzyme-based electrochemical biosensors have achieved great commercial importance since the first use of glucose oxidase in an amperometric sensor for glucose [1] in 1962 All such systems require the incorporation of the biocatalytic element onto (or into) the sensing electrode structure, and this has been achieved by physical adsorption [2] or entanglement [3], DNA intercalation [4] and a wide variety of covalent bonding techniques [5,6,7,8]. The carbodiimide coupling reaction provided by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) has previously been used to create a peptide bond between amides and carboxylic acids in aqueous solution at room temperature [16] This method has been applied for the immobilization of enzymes on modified conducting polymers under moderate conditions [17]
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