14

CHAPTER 2

AMPEROMETRIC GLUCOSE BIOSENSORS: PAST,

PRESENT AND FUTURE

This chapter is about the past, present and future of technology development of glucose

biosensors.

2.1 Historical Perspectives of Glucose Sensors

Foundations of biosensor and its technological advancements thereof, was laid down by father

of the biosensor, Professor Leland C Clark Jnr., in 1956 with advent of Clark oxygen

electrode. The first biosensor dates back to 1962, when Clark and Lyons of the Cincinnati

Children’s Hospital proposed the first enzyme based electrode to measure blood glucose[26].

The underlying principle being monitoring the oxygen consumption, during the oxidation of

glucose, by glucose oxidase (GOx) entrapped between semipermeable membranes over an

oxygen electrode. Since the pioneering work of Clark and Lyons, although a variety of

techniques and methodologies focusing on improvements of signal transduction and

immobilizations of the biomolecule for glucose biosensor are reported, still it has changed

little in principle over several years.

The electrochemical and colorimetric glucose biosensor developed alongside. First

colorimetric biosensor was launched by Dextrostix in 1965 in the form of blood glucose-

sensing strip based on colorimetric detection of hydrogen peroxide produced during the

oxidation of glucose by glucose oxidase [27]. This was closely followed by first functional

electrochemical biosensor by Updike and Hicks in 1967 [28] for glucose and potentiometric

biosensor for urea by Guilbault and Montalvo in 1969 [29]. Yellow Spring Instrument (YSI)

Company in 1975, launched the first Model 23 YSI glucose analyzer, for direct estimation of

glucose levels in blood. 1974-75 marked a turning point in biosensor history with the

proposed usage of - thermal transducers in enzymatic biosensors [30] and bacteria in place of

enzymes for measurement of alcohol. Later, in 1980 an optical biosensor using alcohol

oxidase enzyme was reported for alcohol detection [31]. Since then different biomolecules –

enzymes, microorganism, DNA, antigen/antibody etc. have been used as bioreceptor element.

Hence, based on the type of biomolecule used biosensor is classified as enzymatic biosensor,

15

microbial or whole cell biosensor, DNA biosensor and immunosensor. In addition to this,

depending on the type of transducer used biosensors are further classified as amperometric,

potentiometric, optical, piezoelectric, calorimetric biosensors. Electrochemical biosensors in

general, and electrochemical glucose biosensor in particular have been studied extensively

presenting various technological advancements leading to improvement in biosensor

parameters like selectivity, response time, stability etc [32]. These technological

developments are discussed in the following sections.

2.2 Technological advancements of electrochemical glucose biosensors

Based on the technology improvement of electrochemical glucose biosensors, three

generations of glucose biosensors have been reported [33].

2.2.1 First-generation of Glucose Biosensors

The first generation glucose biosensors estimated glucose concentration in the sample based

on hydrogen peroxide production by glucose oxidase (GOx) utilizing dissolved oxygen as

given below

A negative potential is applied to the Pt working electrode for a reductive detection of the

oxygen consumption as

The key point of above reaction lies in the redox center of the GOx (FAD) which performs

the function of the initial electron acceptor. The interaction of glucose molecule with flavin

adenine dinucleotiede (FAD) of GOx results in its reduction.

The rejuvenation of the cofactor of enzyme GOx occurs in the presence of molecular oxygen,

resulting in the formation of hydrogen peroxide (H2O2) as,

GOx

Glucose + O2 Gluconic acid + H2O2

O2 + 4H+ + 4e

- 2H2O

Glucose + GOx (FAD) Gluconate + GOx (FADH2)

GOx (FADH2) + O2 GOx (FAD) + H2O2

16

Thus, the rate of reduction of oxygen is directly proportional to the glucose concentration that

is enumerated by either measuring the reduced oxygen concentration or increased

concentration of hydrogen peroxide.

Hydrogen peroxide thus produced as a byproduct is oxidized at platinum (Pt) anode. The

electrons transferred are recognized by electrode and thus the number of electrons transferred

is directly proportional to the number of glucose molecules present.

This glucose biosensing technology of Clark was transferred to Yellow Spring Instrument

Company and on the same principle they launched the first commercial glucose biosensor in

market (Model 23A YSI analyzer) for the direct measurement of glucose in 1975. The usage

of the most expensive metal platinum for fabrication of this electrode restricted the biosensor

to clinical laboratories only.

Major drawbacks of first generation glucose biosensor:

Interference from electroactive species present in blood, such as uric acid, ascorbic

acid and other constituents of blood, at the high operational potential (+0.6V) required

for amperometric measurement of hydrogen peroxide. This limits the high selectivity

of the analyzer and results in inaccurate measurements of glucose concentration.

Oxygen deficit – Sensors involving natural oxygen as the electron acceptor due to

presence of oxidase enzyme, generally face errors resulting from fluctuations in

oxygen tension due to the limited solubility of oxygen in biological fluids. This

reduces the linear range of the biosensor.

Number of approaches have been suggested for addressing this problem, Joseph Wang and

group introduced a biosensor with high oxygen solubility based on a fluorocarbon (Kel-F oil)

pasting liquid [34]. Thus the internal flux of oxygen supports the reaction catalyzed by the

enzyme, even in the absence of oxygen in glucose solution. Other approach was proposed by

Gough’s group, in which they designed a two dimensional cylindrical electrode in which

diffusion of glucose is allowed only from one direction while the oxygen is allowed to diffuse

from both directions into the region where enzyme is immobilized [35, 36]. The above

strategy was achieved by developing a two-dimensional sensor design containing a cylindrical

H2O2 O2 + 2H+ + 2e

-

17

gel with GOx and covering the outer part with a silicone rubber tube which does not allow

glucose but is highly permeable to oxygen.

2.2.2 Second Generation glucose biosensor

Search for an alternative for the natural oxygen acting as an electron acceptor in first

generation biosensors lead the electrochemists to second generation biosensors. Use of

synthetic electron mediators opens up new horizons in the field of biosensors. The synthetic

electron mediators eliminated the need of oxygen for recording the electron transfer at the

electrode surface overcoming the drawbacks of limited oxygen pressure observed in first

generation biosensor. Moreover, the lower redox potential of chosen mediators (-0.1V vs

Ag/AgCl for Pursian Blue) results in no interference from other electro active species such as

ascorbic acid and uric acid. Optimal applied potential for eliminating interference was found

to be between 0.0V to 0.2V. In addition to overcoming the above mentioned two drawbacks,

usage of mediators also ensured faster rate of shuttling of electrons from the redox center of

the enzyme to the surface of the electrode.

Electron transfer rates are affected by the structure of the enzyme and hence accessibility of

the active site. The active center of GOx, the flavin adenine dinucleotide (FAD), is buried

inside a deep pocket between the two subunits of the dimeric enzyme. Thus, the direct

electron transfer from Glucose-reduced GOx(red) to metal electrodes is not facilitated because

of the appreciably large distance between GOx redox centers and the electrode surface (>12-

17Aº), resulting in a much retarded diffusion controlled electron transfer rates.

Mechanism of action of mediators can be explained as:

Glucose from the bulk solution diffuses to the enzyme active site and is converted to gluconic

acid. The electrons released during the above conversion are picked by the mediator and is

reduced; finally at the applied potential oxidation of mediator releases electrons that are

transferred to the electrode. Role of mediators in facilitating electron transfer is further

explained by set of equations given below,

18

where, SM(red) and SM(ox) represents the reduced and oxidized forms of synthetic mediator,

respectively. As represented in above equations the reduction of SM(ox) to form SM(red)

facilitates the reoxidation of reduced form of GOx (FADH2). Further oxidation of SM(red) at

the electrode surface regenerating SM(ox) and two electrons. The number of electron

transferred to the electrode is proportional to the glucose concentration. Some of the common

synthetic electron mediators, which have been used to increase the electron transfer rate or

performance of the sensor, are listed in Table 2.1.

Table 2.1 List of synthetic mediators and their redox potential.

Enzyme Synthetic Mediator Redox potential(Versus

SCE) (mV)

Glucose oxidase Vinyl ferrocene 250

[Fe(CN)6]4- 180

Indigo Disulfonate 188

Methylene blue 217

1,1- dimethyl ferrocene 100

[Ru(CN)6]4- 685

TCNQ 127

Ferrocene carboxylic acid 275

Ferrocene carboxaldehyde 518

TTF 300

Benzyl viologen 370

Hydroxy methyl ferrocene 185

Ferrocene 165

Glucose dehydrogenase N-ethyl phenazene -172

Ferrocene carboxylic acid 275

TMPD -10

1,1-dimethyl ferrocene 100

The characteristics features leading to enhanced usage of mediators are

a low redox potential which helps in avoiding interfering current from coexisting

electro active species leading to false measurements,

their low molecular weight and insoluble nature which allows them to effectively

diffuse without complexing,

high stability in both reduced and oxidized forms and,

low toxicity.

19

All the above mentioned unique qualities lead to an improved linear response and thus a

prolonged lifetime of the biosensor, since the deactivation of the enzyme due to production of

hydrogen peroxide is eliminated. Unfortunately, usage of mediators have their own associated

problems which hinders the successful performance of the biosensors.

Major drawbacks of using mediators in second generation glucose biosensor:

High competition between synthetic mediator and oxygen: Although the

probability of reaction between synthetic mediators with the active center of GOx(red)

occurs at a faster rate than oxygen, still the possibility of competition for oxidation of

the reduced GOx by dissolved oxygen with the synthetic mediator is highly likely,

thus resulting in the accumulation of hydrogen peroxide near the electrode surface

leading to reduced bioactivity of enzyme and biosensor response.

Interference: The possibility of oxidation of coexisting electro active species such as

ascorbate even at low applied potential not only affects the accuracy of the sensor but

also enhances the chances of reaction of the synthetic mediator with interfering

species. Thus leading to further inaccurate or false measurements.

Stability of synthetic mediator near electrode surface: Small size and highly

diffusive nature of synthetic mediators poses problem of leaching of mediator from

intermediate region between enzyme and electrode surface. This limits their use in

applications where continuous operation of biosensor is required to avoid mediator

leaching.

During eighties strategies other than incorporation of synthetic electron mediators [37] to

facilitate electron transfer between the GOx redox center and the electrode surface were also

introduced, such as the concept of wired enzymes [38]. Wired enzymes involved redox

hydrogels (redox ions/mediators immobilized on to hydrogels) acting as electrical wires for

conducting the electron from GOx active center to the electrode surface [39].

2.2.3 Nanomaterials: A better platform for biosensor fabrication

The unique properties of nanostructures have been exploited to achieve parameters like fast

response time, high sensitivities, low detection limits, wide range linearity and low power

requirements necessary for highly precise and defined analyte sensing. The high sensitivity

(196nA/mM) and wide linear range (0.2-20 mM) demonstrated by glucose biosensor based on

modified sol-gel composite at the surface of a basal plane pyrolytic graphite electrode

20

decorated with MWCNT presented by Abdollah Salimi et al. shows the promising behavior of

nanomaterial as compared to the previous hydrogel or membrane based biosensors [40].

Glucose oxidase immobilized on gold nanowires based biosensor that could detect glucose in

just 8 seconds as opposed to few minutes of earlier membrane based biosensors was proposed

by Yashuang Lu et al [41]. The high conductivity and biocompatible behavior of elemental

gold at nanoscale makes them a potential platform form immobilization of Glucose oxidase.

Glucose biosensor based on gold nanoparticles proposed by Sylvain Thibault et al., represents

an extremely efficient system allowing even lower detection of glucose concentrations

(0.37mM) with wide linear range [42].

2.2.4 Third-Generation Glucose Biosensors

In order to avoid complications offered by synthetic or natural mediators in second generation

biosensors, a lot of work is being done for finding new strategies for direct electron transfer

between the electrode and active center of enzyme. This led to development of highly

selective and sensitive third-generation biosensors. However, there are only few reports in the

literature concerning the direct electron transfer (DET) between active center of GOx and

electrode surface, although DET for many enzymes have been achieved [43-45] by

immobilizing them within the thin films with different modifications. The intrinsic barrier to

electron flow is the globular structure of GOx with the active site, containing FAD/FADH2

redox cofactor, buried deep inside a cavity of ~13A◦ is a major hinderance for direct electron

transfer in case of thin film or hydrogels based electrodes. Unsuccessful efforts to obtain

direct electron transfer of GOx at conventional electrodes led to exploration of new electrode

materials. In the year 1987, Albery, Cranston and Bartlett suggested incorporation of organic

conducting salt electrodes in order to avoid protein denaturation and fast direct electron

transfer. These conducting salts can be modified into single crystals, as pressed pellet or a

paste with graphite powder in order to prepare electrode. The conducting organic salts like

tetracyanoquinodimethane (TCNQ) and tetrathiafulvane (TTF), have proved to be useful for

the above application [37, 46]. Different researchers exploited these materials in different

ways to achieve high sensitivities. A third generation glucose sensor based on the growing

tree-shaped crystal structure of TTF-TCNQ was proposed by Khan et al [47]. The reduced

distance between the enzyme active center and electrode and immobilization of enzyme in

correct orientation at the electrode surface allowed direct oxidation of the enzyme at a low

applied potential of 0.1 V, athough no explanation for direct oxidation of obtained results

21

were provided by the authors. Cenas and Kulys [48] presented number of arguments against

the direct electron transfer presented by Palmisano et al in a glucose biosensor fabricated

using growing TTF-TCNQ tree-like crystals through an anti-interference layer of a

nonconducting polypyrrole film [49]. Further, number of mediatorless glucose biosensors

based on different materials like polypyrrole system, oxidized boron-doped diamond

electrodes etc were proposed [50].

Efforts to achieve DET using nanoparticles of different types and size were not very fruitful,

however, SWCNTs and MWCNTs were found to be good candidate. SWCNTs immobilized

vertically on the electrode surface provide suitable orientation for enzyme immobilization and

establishing connection between electrode surface and deeply buried active site [51, 52]. This

enables electron transfer over much longer distances of approximately 150 nm in shorter time

(few seconds) while a diffusion based electron transfer over length scales greater than 8-17A◦

results in much longer time of few minutes. Depending on the length of CNTs and efficient

connectivity with redox center the interfacial electron transfer rate varies, e.g. for 50 nm long

CNTs it is 42s-1

[53] while in another report with PLL-SWCNT-GOx electrode with 23 nm

long SWCNT much higher electron transfer rates of 70-100 s-1

were observed [54]. The

distance between the electrode surface is responsible for the large over potential requirement,

i.e. potential greater than the thermodynamic redox potential of the enzyme. To decrease the

working potential, better connectivity leading to DET is desired. This not just improves the

electron transfer rates but also takes care of the problems of interference from electroactive

species. Recently, research efforts are directed at achieving the same. Although substantial

progress has been made on the electronic coupling of GOx, further improvements in the

charge transport between its FAD redox center and electrodes are desired.

2.3 Glucose Biosensors: Research Efforts 1962-2012

More than 80,000 research articles related to various biosensors have been published since

1962. Out of which ~10% (>8020) of the papers are related to glucose biosensors alone and

greater than 66% of the glucose biosensors are enzymatic glucose biosensors. Recent interest

in nanomaterials is evident from the fact that ~80% of the reported glucose biosensors

research exploit the properties of nanomaterials for improved biosensing. Among

electrochemical, optical, piezoelectric and impedimetric glucose biosensors, amperometric

glucose biosensors (>92%) are most widely studied ones while optical glucose biosensors

22

contributes ~5% and potentiometric being only 2.5%. There are only two research articles on

piezoelectric and one on impedimetric glucose biosensors (see Figure 2.1). Probable reason

behind the above statistics being the ease of fabrication and cost effectivenesss of

amperometric biosensor. Table 2.2 below shows biosensor performance characteristics in

chronological order of various biosensors developed till date.

However, the research ideas are not effectively translated into product as evident from

comparatively much lower number of patents filed (see Table 2.3). Table 2.3 shows the

number of glucose biosensor patents filed and granted by different patent offices – US Patent

Office (USPTO), European Patent Office (EPO) and other countries patent offices (Others).

Table 2.3: Number of patents for glucose biosensor.

The commercial availability of the glucose biosensors confirms the dominating behavior of

the device (> 90% of commercial biosensors are glucose biosensors) in the biosensor market.

Immobilization matrices USPTO EPO Others

Membrane based 306 290 141

Hydrogel based 105 29 94

Nanomaterials based 52 9 49

Application in fermentation

industry

8 5 12

Figure 2.1: Percentage distribution of reported research articles based on

different types of glucose biosensors

23

The above table shows that out of approximately 1000 patents filed in the field only 25

glucose biosensor patents are applied in the field of fermentation industry. Thus more than

95% of the consumer market is occupied by blood glucose monitoring devices and just 3% of

the available technology is applied in the fermentation industry.

Table 2.2: Performance characteristics of amperometric glucose biosensors in chronological order.

Type of support for

Immobilization

Sensitivity Interference Detectio

n limit

Linear

range

Respons

e time

Stability Reference

GOx was immob. on

graphite followed by

adsorption of N-methyl-

phenazinium ion (PMS+)

- Low

interference

from

Galactose and

Mannose

0.5, to

150 μm

2 mM 20-60 s Aleast 9

months

Gunilla

Jönsson et

al., (1985)

[55]

GOx was incorporated into

polypyrrole films that were

electrochemically

deposited on PE.

- - - - 20-40 s 21 days Nicola C.

Foulds et

al., (1986)

[56]

Cellulose acetate, GOx

(crosslinked with

glutaraldehyde) and

polyurethane are placed on

surface of central platinum

wire surrounded by a

stainless steel tubing

- - - 500 mg/dl 100 sec 6 days Kerner W

et al.,

(1988) [57]

Polysiloxanes are used for

interaction between GOx &

CPE

- None by

thiocynates

- 16-71 mM <10 s 2 years

(20%)

Paul D.

Hale et al.,

(1991)

[58]

Incorporation of GOx into

graphite paste modified

with

tetracyanoquinodimethane

(TCNQ)

- - 0.5 mM 5-50mM 15-50

sec

35 days PC Pandey

et al.,

(1992) [59]

Immobilizing GOx and

coating Nafion membrane

7.1 +/- 0.5

nA/mM

- - 0.5-40 mM 30 s 6 days Jianzhong

Zhu et al.,

24

(1994) [8]

Glutaraldehyde & BSA is

used for crosslinking GOx

on CPE

- Interference

from ascorbic

acid, uric acid

and

paracetamol

- 2–20 mM 60-120 s 6 days Miloslav

Pravda et

al., (1995)

[60]

Poly (o-aminophenol) film - - - 0.001-1.0

mM

More

than 4

sec

30 days

(30%

reductio

n)

Z.Zhang et

al.(1996)[6

1]

Sol−gel organic−inorganic

hybrid material was used

for immob. of GOx

600 nA

mmol-1

L-1

Interference by

L-Ascorbate

- 0 to 16 mM 11s 5

months

Bingquan

Wang et

al., (1998)

[62]

Immobilization of GOx

into poly(o-

phenylenediamine) (POPD)

on Pt electrode. Additional

layer of Prussian blue (PB)

was also placed.

0.2 to 0.7

μA mM−1

cm−2

Diminished

ascorbate

interference

- 8 to 14 mM 4 to 8 s - R

Garjonyte

et al.,

(1999) [63]

GOx and poly(p-

phenylenediamine) (poly-

PPD) were coimmobilized

at the surface of a platinum

microdisk electrode

160 μA

cm-2

mM-1

No

interefernce to

ascorbic acid,

uric acid &

cysteine

- 5.0 x 10-5

to

3.0 x 10-3

M

<2 s 2 month Xu Jing-

Juan et al.,

(2000) [64]

3- APTES, Nafion®, GOX,

and perfluorocarbon

polymer (PFCP)

2.21

nA/mM

No

interference

- 2.8 to 167

mM

- 66 days T.

Matsumoto

et al.,

(2001) [65]

Polymerization of p- 5.2 × 10–3

No 10–4

M 2.5 x 10-4

to <2 s 90 days Jing-Juan

25

chlorophenol (4-CP) at a Pt

electrode.

nA L mol–1

interference 1.5 x 10-2

mol/ L

Xu et al.,

(2001) [66]

Co-electrodeposition of a

poly(vinylimidazole)

complex of

[Os(bpy)2Cl]+/2+ (PVI-

Os).

349

nA/mM

No

interference

upto 2 mM

glucose by

ascorbic acid

0.03mM 0 - 30 mM 5 s 64 hours

(50%)

Junjei Fei

et al.,

(2003) [67]

GOx was immob. into a

copper dispersed sol-gel

derived ceramic-

graphite composite

- No

interference to

ascorbic acid

& uric acid.

1.8 × 10-

5

M

4.0×10-5

to 5.6×10-3

M

6-9 s 60 days

(78%)

D. Ravi

Shankaran,

et al.,

(2003) [68]

GOx was immob. into a

sol-gel composite at the

surface of a basal plane

pyrolytic graphite electrode

modified with MWCNT.

196

nA/mM

- 50 μM 0.2-20 mM <5 s 3 weeks. Abdollah

Salimi et

al., (2004)

[40]

Adsorption of GOx at the

platinum nanoparticle-

modified CNT

91mA M-

1cm

-2

Weak

interference of

Ascorbic acid

and Uric acid

- 0.1-

13.5mM

5 s 22 days

(73.5 %)

Hao Tang

et al.,

(2004) [69]

Polyelectrolyte & chitosan

was used to immob. GOx

on Pt electrode

21 mA M−1

cm−2

No

interference

10 μM 0.05–15

mM

<8 s 3

months

(65%)

Minghui

Yang et al.,

(2004) [70]

Microdeposition of

PEDOT/PSS and GODon

ITO-glass

6.43 μA

M−1

cm−2

- - 60 mM - - L. Setti et

al., (2005)

[71]

GOx was immobilized in

sol–gel chitosan/silica

hybrid composite film

420 nA

mM–1

No

interference

8.0×10–6

M

5.0×10–5

to

2.6×10–2

M

10 s 60 days Xue-Cai

Tan et al.,

(2005) [72]

GOx and the electrocatalyst

cobalt phthalocyanine were

mixed with the carbon ink.

1170 nA

mM−1

- 0.025

mM

0.025–2

mM

- - Eric

Crouch

(2005) [73]

26

Poly(3,4-

ethylenedioxythiophene)fil

m

12.42 μA

mM−1

cm-2

With Ascorbic

acid current

increases 9.7%

and with Uric

Acid 39.1%.

130.0

µM

0.1-10 mM 4-10 s 20%

decrease

after 18

days

Nien et al.

(2006) [74]

Aminated silica

nanoparticles

5.11 μA

mM−1

cm-2

- 9.0 µM Upto 8

mM

4 s 90%

decrease

in 45

days.

Sun et al.

(2006) [75]

Electrochemical

entrapment of glucose

oxidase (GOD) into porous

poly(acrylonitrile-co-

acrylic acid)

6.82 mA

M−1

cm−2

3-7% by

scorbic acid

(0.1 mM),

reduced

glutathione (2

mM), L-

cysteine

(0.02 mM),

and p-

acetaminophen

ol (0.05 mM).

41% by

uric acid

(0.5 mM)

0.5 μM 5 × 10−6

to

3 × 10−3

M

<30 s Atleast 5

weeks

Dan Shan

et al.,

(2006) [76]

Immobilization of GOx on

to the electrochemically

platinized carbon paste.

- No

interference

2 × 10−5

M

6 × 10−5

to

1.2 × 10−2

M

20–30 s 45 days Liang Ming

et al.,

(2006) [77]

GOx was immob. directly

onto an oxidized boron-

doped diamond (BDD)

electrode.

- - 2.31×10−

5 mol/L

6.67×10−5

to 2×10−3

mol/L

<5 s 1 month

(61%)

Jing Wu et

al., (2006)

[50]

GOx/colloidal Au NP on

GCE by Nafion film

0.4 μA

mM−1

cm−2

- 0.37 mM 20 mM 1.5 min - Sylvain

Thibault et

27

al., (2008)

[42]

GOx is immobilized on

CNT surface by assembling

polydiallyldimethylammon

ium chloride (PDDA) layer

- No

interference

7mM 15 μM to 6

mM

- - Guodong

Liu et al.,

(2006) [78]

Polypyrrole (PPy),

functionalized cMWNT,

and GOx

95 nA

mM−1

- - 4 mM - 8 s Yu-Chen

Tsai et al.,

(2006) [79]

Au nanowires-Chitosan

was immob. on GCE

- - 5×10−6

M

10−5

-2×10-

2M

< 8 s 1 month

(85%)

Yashuang

Lu et al,

(2007) [41]

GOx

was immob.on

Pt/sulfonated-

MWCNT/GCE

0.56

μA/mM

- - 6.4

mM

- - H.J. Wang

et al.,

(2007) [80]

PAA, MWCNTs,

cysteamine and GNp,

respectively, followed by

the adsorption of GOD on

Pt electrode

(GOD/GNp/MWCNTs/Pt

electrode)

2.527

μA/mM

No

interference

6.7 μM 0.1–10 mM

glucose

<7 s 7 S Bao-Yan

Wu et al.,

(2007) [81]

Calcium carbonate

nanoparticles was used for

immob. of GOx.

58.1 mA

cm−2

M−1

3-7% by

scorbic acid

(0.1 mM),

reduced

glutathione (2

mM), L-

cysteine

(0.02 mM),

and p-

acetaminophen

ol (0.05 mM).

0.1 μM 0.001–12

mM

6 s 120 days

(86%)

Dan Shan

et al.,

(2007) [82]

28

39% by

uric acid

(0.5 mM)

―Unprotected‖ Pt

nanoclusters mixed with

the nanoscale SiO2

particles

3.85 μA

mM−1

- 1.5 μM 0.27 to 4.08

mM

- - Haipeng

Yang

(2007) [83]

Micro-patterned Prussian

blue (PB) and ferrocene

modified glucose oxidase

covered by a thin Nafion

membrane

- Slight

interference by

Ascorbic acid

on anodic

detection

75 µM 0.1 to 50

mM

1-6 min 1 week

(60%)

Na Zhang

et al.,

(2007) [84]

Layered double hydroxides

(LDHs)

60 mA M−1

cm−2

Negligible 3 μM 6.7 × 10−6

-

3.86 ×10−4

M

5 s - Dan Shan

et al.(2007)

[85]

Exfoliated Graphite

Nanoplatelets Nafion

membrane

14.17 µA

cm-2

mM-1

56.8% for

0.1mM

Ascorbic acid

and 125% for

0.2 mM uric

acid

interference

10 µM upto 6 mM 5

seconds

Stability

for 7

days.

Jue Lu et

al.(2007)

[86]

Amino functionalized

Multi-wall carbon

nanotubes (MWNTs)

7.46 μA

mM−1

cm-2

Minimum

interference.

8.0 µM - <10 s - Sun et al.

(2008) [87]

Single Walled Carbon

Nanohorns

15.0 μA

mM−1

cm-2

Can avoid the

commonly

coexisted

interference.

6.0 µM 0-6 mM - - Liu et al.

(2008) [88]

Layer-by-layer (LBL) self-

assembling of chitosan and

glucose oxidase (GOD) on

a Prussian blue film was

developed

- Low

interference

3.1 μM 6 μM to 1.6

mM

10 s 4 weeks

(73%)

Bing Yin et

al., (2008)

[89]

29

Gold nanoelectrode array

was fabricated by template-

assisted electrodeposition

on general electrodes

1.52 mA

mM-1

cm-2

- 5 x 10-6

M

1 x 10-6

to 1

x 10-2

M

8 s 3

months

(80%)

Yanyan Liu

et al.,

(2009) [90]

Composite material based

on layered double

hydroxides (LDHs) and

chitosan (CHT)

62.6 mA

M−1

cm−2

Weak

interference

0.1 μM 1 × 10−6

to

3 × 10−3

M

5 s 60 days

(70%)

Qiaofang

Shi et al.,

(2008) [91]

Immobilizing glucose

oxidase (GOD) in a titania

sol-gel film, which was

prepared by a vapor

deposition method, on a

Prussian Blue (PB)-

modified electrode

12.74 μA

cm-2

mM-1

For ascorbic

acid, current

decreased

about 3%, &

for Uric Acid

and cysteine

the current

increased 0.2

and

1.2%,

respectively

5 μM 0.02 to 15

mM

< 10 s 3

months

(91%)

Ruping

Liang et

al., (2008)

[92]

Adsorption of

GOx on an AuNPs–

AgCL@polyaniline

(PANI) core-shell

nanocomposites on GCE

- 3, 2.4 and

1.8% for

ascorbic acid,

uric acid &

cysteine.

4 pM 4–34 pM - 2 weeks

(80%)

Wei Yan et

al., (2008)

[93]

Graphite nanoplatelets

(xGnPs) decorated with Pt

and Pd nanoparticles was

used

61.5 ± 0.6

μA mM-1

cm-2

Interference by

ascorbic acid

and uric acid

1 μM 20 mM 2 s 1 week Jue Lu et

al., (2008)

[94]

Encapsulating GOx in the

Nafion– single-walled

carbon

nanohorns(SWCNHs)

composite film

1.06

μA/mM

No

interference of

L-lactate,

glutathione, L-

cysteine, and

p-

aminophenol,

6 μM 0 to 6.0

mM

- - Xiaoqing

Liu et al.,

(2008) [88]

30

L-ascorbate

acid

GOx with

chitosan-AuNP on gold-

Prussian Blue (Au-PB)

nanoparticles (GCE).

9.5 µA M-1

cm-2

- - 3 mM 10 s - Juozas

Kulys et

al.(2008)

[95]

Poly (3,4-

ethylenedioxythiophene) /

Prussian blue bilayer and

multi-walled carbon

nanotubes

2.67 μA

mM−1

cm-2

- - 1–10 mM - 18%

decrease

in 30

days

Chiu et al.

(2009) [96]

Gold nanorods/cellulose

acetate composite film

8.4 μA

mM−1

cm-2

- 20 µM 0.03-

2.2 mM

- 20%

decrease

in 30

days.

Ren et al.

(2009) [97]

Entrapping GOx onto the

inner wall of highly

ordered polyaniline

nanotubes (nanoPANi)

97.18 ±

4.62 μA

mM−1

cm−2

No

interference

0.3 ± 0.1

μM

0.01−5.5

mM

3 s 2 weeks

(91%)

Ziyi Wang

et al.,

(2009) [98]

Alginate (Alg)/layered

double hydroxides (LDHs)

organic-inorganic

composite film

68.9

µA/mM/cm

0- 3.6% by

Ascorbic acid

(0.1 mM), Uric

acid (0.5 mM),

Glutathone

reduced (2

mM), L-

cysteine (0.02

mM), p-

acetaminophen

ol (0.05 mM)

4 × 10−5

M

1.6 × 10−5

−

2 × 10−3

M

10 s 28 days

(87%)

Shou-Nian

Ding et al.,

(2009) [99]

Graphene

- - - 2-14 mM - - Shan et al.

(2009)

[100]

31

Silver nanoparticles/carbon

nanotubes/chitosan film

135.9

µA mM-1

20µM for uric

acid and 8µM

for ascorbic

acid

0.1µM 0.5-50 µM - 9%

decrease

in 10

days and

20% in

about 40

days

Jiehua lin

et al.

(2009)

[101]

Poly (3,4-

ethylenedioxythiophene) /

Prussian blue bilayer and

multi-walled carbon

nanotubes

2.67 μA

cm−2

mM−1

.

- - 1–10 mM - 18%

decrease

in 30

days

Jing-Yang

Chiu et al.

(2009) [96]

GOX is immobilized onto

the CNT/Pt nanosphere.

70

μA/mM/cm

2

- 380 nM 1 μM to

0.75 mM

8 s - Jonathan C.

Claussen

(2010)

[102]

GOx was immob. thin

films of chitosan

containing nanocomposites

of graphene and gold

nanoparticles (AuNPs) at a

gold electrode

0.55μAmM

−1

- 180 μM 2 to 10 mM

and from 2

to 14 mM

- 15 days

(4.6%

reductio

n)

Changshen

g Shan et

al., (2010)

[103]

Utilizes CNTs

electrochemically

decorated with platinum

(Pt) nanospheres to sense

glucose

70

µA/mM/cm

- 380 nM 1 µM to

0.75 mM

8 s - Jonathan C.

Claussen et

al., (2010)

[102]

Silicon dioxide coated

magnetic nanoparticle

decorated multiwalled

carbon nanotubes

(Fe3O4@SiO2/MWNTs)

on a glassy carbon

electrode (GCE)

58.9μA/m

M cm2

Weak

interference by

ascorbic acid

and uric acid

800 nM 1 μM to 30

mM

- - Tessy

Theres

Baby et al.,

(2010) [11]

32

Immobilization of GOx on

nanoflake-like SnS2

modified GCE

7.6 ± 0.5

mA M⁻¹

cm⁻2

None by Uric

acid, and

Ascorbic acid

1.0 ×

10⁻⁵ M

2.5 × 10⁻⁵

M to 1.1 ×

10⁻³ M

8 s 20 days

(8%

reductio

n)

Zhanjun

Yang et al.,

(2011)

[104]

Electrodepositing chitosan

(CS)-glucose

oxidase(GOD)

biocomposite onto the

stainless steel needle

electrode (SSN electrode)

modified by Pt–Pb

nanoparticles (Pt–Pb/SSN

electrode)

0.4485

μA/mM

- - 0.03 to 9

mM

15 s - Meiqing

Guo et al.,

(2011)

[105]

p-tert-

butylthiacalix[4]arene

tetra-amine (TC4TA) is

used for immob. of GOx.

10.2 mA

M−1

cm−2

Weak

interference

20 μM 0.08–10

mM

5 s 20%

decrease

in 30

days

Ming Chen

et al.,

(2011)

[106]

AuNPs and MWCNT

nanocomposite materials

were constructed by

alternate self assembly of

thiol functionalized

MWCNTs and AuNPs,

19.27 μA

mM−1

cm−2

Weak for

ascorbic acid,

uric acid and

acetaminophen

2.3 μM 20 μM to

10 mM

3 s 1 week

(95.4%)

Peng Si et

al., (2011)

[9]

GOx was immob. on

polyaniline-

polyvinylsulphonate

(Pani-Pvs) via the

entrapment technique

- 15% and 25%

for ascorbic

acid & uric

acid.

1.0 ×

10−7

M

1.0 ×

10−7–1.0 ×

10−5 M

200 s 40 days

(80.6%)

Fatma

Arslan

et al.,

(2011)

[107]

Pt nanoparticle

homogeneously decorated

on polyaniline (Pani)-

wrapped boron nitride

nanotubes (BNNTs),

19.02 mA

M–1

cm–2

Negligible for

Ascorbic acid

and Uric acid

0.18 μM 0.01 to 5.5

mM

3 s 40 days

(95%)

Jianmin

Wu et al.,

(2011)

[108]

Palladium 31.2 µA Significant 0.2 µM 0.001-1.0 - 20% Zeng et

33

nanoparticle/chitosan-

grafted graphene

nanocomposites

cm-2

mM-1

interference mM decrease

in 21

days.

al.(2011)

[109]

Immobilizing the PtPd-

MWCNTs catalysts in a

Nafion film on a glassy

carbon electrode.

112 μA

mM−1

cm−2

Negligible

interference

0.031

mM

0.062–

14.07 mM

5 s 28 days

(85%)

Kuan-Jung

Chen et al.,

(2012) [14]

Pt nanoparticles-chitosan

composite film (PtNPs-CS)

- Insignificant

interference by

ascorbic acid,

threonine,

L-cysteine,

uric acid

0.4 μM 1.2 μM to

4.0 mM

<5 s 1 month

(89.6%)

Jingjing Li

et al.,

(2012)

[110]