Development of Amperometric Biosensors with Carbon Nanotube … · Development of Amperometric...
Transcript of Development of Amperometric Biosensors with Carbon Nanotube … · Development of Amperometric...
Development of Amperometric Biosensors with Carbon Nanotube Composite Materials
YAO Yanli
A thesis submitted in partial fulfillment of the requirements
for the degree of
Doctor of Philosophy
Principal Supervisor: Dr. SHIU Kwok-Keung
Hong Kong Baptist University
May 2008
Abstract
Carbon nanotubes (CNTs) have attracted much attention since their discovery. CNTs
offer unique mechanical, chemical, physical and electronic properties. Many investigations
have demonstrated that CNTs possess good electrocatalytic activities. This thesis work
mainly focuses on the development of amperometric glucose biosensors utilizing carbon
nanotubes and CNT composite materials. The electrochemical behaviors of the CNT
composite materials have been investigated and the biosensor performance was explored.
CNT-modified electrodes with different types of CNT materials have been prepared.
Electrochemical reactions involving ferricyanide and hydrogen peroxide were examined at
the CNT modified electrodes. Glucose biosensors have been prepared with different types of
CNT materials and the amperometric behaviors for glucose determination have been
examined. Single walled carbon nanotube biosensor usually exhibited a wider dynamic range
and higher sensitivity for glucose determination.
Carbon nanotubes have been combined with different materials, including Prussian
blue inorganic nanoparticles, copper nanoparticles and gold colloid, to prepare different
CNT-based electrode substrates. Different polymeric materials were utilized as enzyme
entrapment agents. Chitosan, PDDA and poly(toluidine blue O) films have been employed.
Amperometric glucose biosensors can be prepared by using different CNT composite
materials and the biosensor performance was investigated.
A highly sensitive glucose biosensor has been fabricated by electrochemical
deposition of chitosan (CHIT) and glucose oxidase (GOD) on single-walled carbon nanotube
(SWCNT) modified glassy carbon electrode with copper nanoparticles. The presence of
SWCNT and copper nanoparticles offered improvement in electron transfer at the electrode
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surface. Electrodeposited chitosan film provided biocompatible matrix for the
immobilization of enzymes. The biosensor displayed linear response to glucose from 0.02 to
5 mM with a sensitivity of 22.4 mA M–1 cm–2 at an applied potential of +0.65 V (vs.
Ag|AgCl). On the other hand, Au colloid was prepared and assembled on the CNT surface by
a positively charged polymer PDDA. The direct electrochemistry of GOD on the
GC/CNT+Au/PDDA modified electrode was observed.
A mediator glucose biosensor has been constructed by immobilizing glucose oxidase
at electropolymerized poly(toluidine blue O) film on carbon nanotube modified glass carbon
electrode. Toluidine blue O moieties served as mediators for direct oxidation of glucose
oxidase and as polymeric network to maintain the biosensor activity. The detection potential
could be decreased to –0.1 V (vs. Ag|AgCl), where common interferences such as ascorbic
acid, uric acid and Acetaminophen were not oxidized to cause interferences. The
amperometric glucose biosensor offered a sensitivity of 5.09 mAM–1cm–2 and a wider linear
range of 1-18 mM.
Utilizing the unique properties of multi-walled carbon nanotubes, an amperometric
biosensor based on horseradish peroxidase (HRP) was fabricated. Experimental results
indicated that there was direct electron transfer between HRP enzyme and CNT-modified
glassy carbon electrode. Electrocatalytic reduction of hydrogen peroxide at the
GC/CNT/HRP/Nafion electrode was investigated. Additionally, a mediator-free bienzyme
glucose biosensor based on horseradish peroxidase and glucose oxidase was constructed. A
zero applied potential offered an improvement in the selectivity of the biosensor. Under the
optimized conditions, the bienzyme biosensor exhibited a high sensitivity for glucose
detection.
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Table of contents
Declaration…………………………………..……………………………………………….i
Abstract……………………………………………………………………………................ii
Acknowledgements..………………………………………………………………...............vi
Table of Contents………………………………………………………………….................v
List of Tables………………………………………………………………………………..xii
List of Figures……………………………………………………………………………....xiii
List of Symbols and Abbreviations…………………………………………………………xxi
Chapter 1 Introduction……………………………………………………………...................1
1.1 Carbon nanotubes…………………………………………………………..................1
1.1.1 Synthesis and purification of CNTs…………………………………………3
1.1.2 Characterization of CNTs……………………………………........................5
1.1.3 Properties of CNTs………………………………………………..................6
1.1.3.1 Electronic properties………………………………………………….....6
1.1.3.2 Thermal stability and mechanical properties……………………………6
1.1.3.3 Electrochemical properties…………………………………...................7
1.1.4 Defects of CNTs…………………………………………………..................9
1.1.5 Functionalization of carbon nanotubes……………………………………....9
1.1.6 Applications of CNT materials…………………………………..................10
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1.1.6.1 Energy storage…………………………………………………………10
1.1.6.2 Nanoprobes………………………………………………………........11
1.1.6.3 Biosensors and chemical sensors………………………………………12
1.1.6.4 Metal supports and templates…………………………….....................12
1.2 Biosensors……………… ………………………………………………..................13
1.2.1 Immobilization of enzymes on electrode…………………………………...14
1.2.2 Biosensor development…………………………………………..................16
1.2.2.1 First-generation biosensors……………………………….....................16
1.2.2.2 Second-generation biosensors................................................................17
1.2.2.3 Third-generation biosensors…………………………………………...18
1.2.3 Carbon nanotube based biosensors……………………………....................18
1.3 Objectives………………………………………………………………...................20
References ………………………………………………………………….. …………23
Chapter 2 Electron transfer properties of different carbon nanotube materials and their
applications in glucose biosensors………………………………………………...35
2.1 Introduction………………………………………………………………………....35
2.2 Experimental……………………………………………………………..................37
2.2.1 Reagents and instruments…………………………………………..................37
2.2.2 Preparation of CNT-modified electrodes and biosensors…………..................38
2.2.3 Amperometric determination of glucose……………………………………...38
2.3 Results and discussion……………………………………………………………....39
2.3.1 Electrochemical behaviors of CNT-modified electrodes…………..................39
2.3.2 Electron transfer properties of CNT-modified electrodes…………………….41
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2.3.3 Oxidation/Reduction of hydrogen peroxide at CNT-modified
electrode………………………………………………………………………44
2.3.4 Preparation of CNT-modified glucose biosensors…………………………....48
2.3.5 Electrocatalytic oxidation of glucose at CNT-modified biosensors.................49
2.4 Conclusions……………………………………………………………….................52
References…………………………………………………………………….................53
Chapter 3 Spontaneous deposition of prussian blue on multi-walled carbon nanotubes and
their application in amperometric biosensor…………………………………….55
3.1 Introduction………………………………………………………………………….55
3.2 Experimental…………………………………………………………………………57
3.2.1 Chemicals and instruments………………………………………………........57
3.2.2 Preparation of CNT-PB modified electrode……………………………….....58
3.2.3 Spectroscopic characterization of electrode substrates……………................59
3.2.4 Preparation of CNT-PB modified glucose biosensor………………………...59
3.2.5 Amperometric determination………………………………………………....60
3.3 Results and discussion……………………………………………………………....60
3.3.1 Deposition of PB on different electrode substrates…………………………..60
3.3.2 Electrochemical characterization of PB……………………………………...65
3.3.3 Ultraviolet-visible absorption characteristics of CNT/PB modified electrode.66
3.3.4 Reduction of hydrogen peroxide at different electrodes………………………67
3.3.5 Preparation of amperometric glucose biosensor based on the CNT/PB/GOD
modified electrode……………………………………………………………69
3.3.6 Effects of pH on the electro-oxidation of glucose………………………….....71
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3.3.7 Effects of detection potential…………………………………………….......72
3.3.8 Detection of glucose and the stability of biosensor……………….................74
3.4 Conclusions………………………………………………………………………....76
References………………………………………………………………………………77
Chapter 4 Preparation of SWCNT-nanocopper-chitosan composites and its application in
the development of amperometric biosensors…………………………………...79
4.1 Introduction……………………………………………………………....................79
4.2 Experimental……………………………………………………………...................81
4.2.1 Reagents and instruments…………………………………………...................81
4.2.2 Preparation of GC/SWCNT/Cu/CHIT-GOD biosensor……………………….83
4.3 Results and discussion…………………………………………………....................84
4.3.1 Electrodeposition of copper and chitosan-GOD composites on SWCNT-
modified electrodes…………………………………………………………….84
4.3.2 Electrocatalytic oxidation of hydrogen peroxide and glucose at the modified
electrodes…………………………………………………………….................87
4.3.3 Effects of copper deposition time on glucose oxidation………….....................94
4.3.4 Effects of the detection potential on the biosensor performance……………...95
4.3.5 Effects of interfering species………………………………………..................97
4.3.6 Amperometric detection of glucose ………………………………....................99
4.4 Conclusions……………………………………………………………....................101
References……………………………………………………………………………....102
Chapter 5 Direct electrochemistry of glucose oxidase at carbon nanotube-gold colloid
modified electrode with poly(diallyldimethylammonium chloride) coating…105
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5.1 Introduction……………………………………………………………...................105
5.2 Experimental section……………………………………………………………….108
5.2.1 Reagents and instruments………………………………………......................108
5.2.2 Synthesis of gold colloid nanoparticles………………………….....................109
5.2.3 Fabrication of glucose biosensors…………………………………..................109
5.3 Results and discussion……………………………………………………………...110
5.3.1 TEM and SEM characterization of gold nanoparticle composites…………....110
5.3.2 UV-visible spectral characteristics………………………………....................112
5.3.3 Electrochemical behaviors of the biosensors………………………………....113
5.3.4 Electrochemical detection of glucose at the GC/CNT/Au/PDDA-GOD modified
electrode………………………………………………………….....................119
5.3.5 Detection of glucose employing an electron mediator……………..................125
5.3.6 Effects of interferences……………………………………………..................128
5.3.7 Stability and reproducibility of the biosensor……………………………….....130
5.4 Conclusions……………………………………………………………....................131
References…………………………………………………………………………….....133
Chapter 6 Low potential detection of glucose at carbon nanotube modified glassy carbon
electrode with electropolymerized poly(toluidine blue O) film…………………137
6.1 Introduction……………………………………………………………...................137
6.2 Experimental……………………………………………………………………….139
6.2.1 Reagents and Instruments…………………………………………………….139
6.2.2 Preparation of the glucose biosensor………………………………………....140
6.2.3 Amperometric measurement of glucose………………………………………141
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6.3 Results and Discussion…………………………………………………………....141
6.3.1 Electropolymerization of toluidine blue O. ………………………………....141
6.3.2 AFM characterization of the modification procedure…………………….....144
6.3.3 Redox response of hydrogen peroxide at CNT-modified electrode with PTBO
coating………………………………………………………………………146
6.3.4 Electrochemical characterization of the glucose biosensor………..................147
6.3.5 Effects of PTBO film thickness on the biosensor performance……………. .152
6.3.6 Effects of detection potential on the biosensor performance………………...153
6.3.7 Amperometric determination of glucose…………………………………......156
6.4 Conclusions…………………………………………………………….................158
References…………………………………………………………………………......159
Chapter 7 A mediator free bienzyme amperometric biosensor based on horseradish
peroxidase and glucose oxidase immobilized on carbon nanotube modified
electrode………………………………………………………………….......162
7.1 Introduction…………………………………………………………………….....162
7.2 Experimental……………………………………………………………………...163
7.2.1 Materials and reagents………………………………………………….........163
7.2.2 Fabrication of biosensor…………………………………………………......164
7.2.3 Electrochemical measurements……………………………………………...164
7.3 Results and discussion.…………………………………………………...............165
7.3.1 Electrochemical behavior of HRP…………………………………………...165
7.3.2 Electrochemical behavior of GC/CNT/Nafion-HRP for hydrogen
peroxide…………………………………………………………………….168
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7.3.3 Characteristics of GC/CNT/Nafion-HRP, GOD bienzyme biosensor……......170
7.3.4 Effects of enzyme composition…………………………………….................173
7.3.5 The effect of the detection potential………………………………..................174
7.3.6 Amperometric detection of glucose……………………………………….......175
7.3.7 Influence of interferences……………………………………………………..178
7.4 Conclusions…………………………………………………………………………179
References………………………………………………………………………………180
Chapter 8 Summary and future work……………………………………………………....183
8.1 Summary…………………………………………………………………………...183
8.2 Future work…………………………………………………………….................187
8.2.1 Fabrication of miniature biodevices………………………………………….187
8.2.2 Preparation of Au colloid…………………………………………………….187
References………………………………………………………………………...........189
CURRICULUM VITAE…………………………………………………………………...190
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