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Title: Electrochemical enzyme electrodes for glucose and creatinine
Author: Lad, Umesh
Awarding Body: University of Leeds
Current Institution: University of Leeds
Date of Award: 2012
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Amperometric enzyme electrodes have been developed for the detection of glucose, sarcosine and creatinine, and tested at "in-house" fabricated graphite rod electrodes. Determination of these analytes was primarily performed in the presence of ferrocenemonocarboxylic acid (FMCA) but also by the oxidation of enzymatically generated hydrogen peroxide (H202) for comparison. Cyclic voltammetry (CV) and amperometric responses were used to analyse the enzyme electrode performances. Colloidal silver/gold-(Ag/Au)-nanoparticles (NPs) and Ag/Au-NPs composite films developed in this study were characterized by ultraviolet-visible (UV -Vis) and Fourier transform infrared (FfIR) spectroscopy, scanning electron microscopy (SEM) and transmission electron microscopy (TEM), and energy-dispersive X-ray (EDX) analysis. In order to determine creatinine, the amperometric enzyme electrode was prepared by immobilizing creatininase (CA), creatinase (Cl) and sarcosine oxidase (SOD) in the optimized ratio of 2:3:5units (U) respectively, onto the surface of graphite working electrodes (WEs) by cross-linking with glutaraldehyde (GA) in the presence of bovine serum albumin (BSA). The linear working range of the electrode was up to -1100f,lM with a sensitivity of,lM-1, the response time was 29s (t95% at 100 ---+ 200f,lM creatinine change) and the detection limit was 190f,lM (SIN = 3). Glucose oxidase (GOD) was also immobilized onto the surface of the graphite WE by cross-linking with GA in the presence of BSA to measure glucose. The ratio amounts between, GOD, BSA and GA was also optimized. The linear working range of the electrode was 1-8.5mM with a sensitivity of, the response time was 6s (t95% at 0.5 ---+ ImM glucose change) and the detection limit was 0.45mM (SIN = 3). Ag-NPs in the enzyme layer had a negative effect on response however; the presence of Au-NPs improved the sensitivity by almost 20%. Secondly, GOD was immobilized into partially prehydrolysed silica sol-gel (pphTEOS) and polyvinyl alcohol (PV A) composite film on the surface of the graphite electrodes. For comparison, Ag- and Au-NPs were incorporated into the immobilization process by preliminary preparing Ag/Au-NPs using PVA as a reducing agent. The linear working range of the electrode was 1.5-6mM with a sensitivity of 4.322 1 2 mM-I h . ,t e response time was 5s (t95% at 0.5 ---+ ImM glucose change) and the detection limit was 0.35mM (SIN = 3). PVA loaded with Au-NPs was capable of extending the linear range by a further 3mM. 11 This immobilization method was also employed for SOD to determine its activity towards sarcosine. The linear working range of the electrode was 1.S-SrnM with a sensitivity of, the response time was 17s (t95% at O.S ~ 1rnM sarcosine change) and the detection limit was 0.S3rnM (SIN = 3). Investigations on stability, reproducibility, selectivity and effect of pH on the enzyme electrodes have also been studied. In addition, electro-deposition of polypyrrole (PPy) films on graphite electrode surfaces was also preliminary examined for possible use as modified electrode surfaces for enzyme immobilization.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available