Trends in Analytical Chemistry, Vol. 24, No.

1, 2005 Trends

Electrochemical impedance
spectroscopy studies of polymer
degradation: application to
biosensor development
C

esar Fern
andez-S

anchez, Calum J. McNeil , Keith Rawson

Electrochemical impedance spectroscopy (EIS) has emerged as a useful or thin-film technology, or monitoring
analytical tool for the development of sensor devices in a wide variety of corrosion [2].
configurations. We focus this review on the particular application of EIS to Using EIS, a vast range of coatings have
the study of degradation phenomena taking place at polymer-coated sub- been widely tested as effective barriers
strates, which have primarily been of great interest in the study of corrosion against corrosion of metal surfaces in the
protection; more recently, it has led to the successful construction of last few decades. Paints and other organic
versatile polymer-coated transducers for sensor development. Impedance and inorganic emulsions deposited on the
analysis of breakdown processes of polymer coatings on electrochemical metal surface break down creating pin-
transducers through the direct or indirect action of biomolecules constitutes holes, craters and other defects when
a feasible detection protocol for the fabrication of generic integrated bio- corroded. In other words, water and any
sensors. We give a detailed description of such applications, and present a other existing free ions are allowed to
particular view on sensor devices reported so far and ideas that bring penetrate into the polymer. This so-called
significant improvements to this promising technology. ionic attack alters the insulating structure
ª 2004 Elsevier Ltd. All rights reserved. of the polymer, which modifies the
impedance characteristics of the overall
Keywords: Degradation; Enzyme sensor; Immuno sensor; Impedance spectroscopy; metal/polymer element. Microbial colonies
Polymer-coated electrodes
and marine water have been reported as
common corrosion agents, their effect on
different polymers being assessed with EIS
1. Introduction at set frequencies.
C
esar Fern

andez-S
anchez, EIS has also been increasingly success-
Calum J. McNeil*
School of Clinical and
Electrochemical impedance spectroscopy ful in the design and the development of
Laboratory Sciences, (EIS) appears to be an excellent technique sensor systems. We need to bear in mind
The Medical School, University for the investigation of bulk and interfacial the vast range of materials that may be
of Newcastle upon Tyne, electrical properties of any kind of solid or used in their fabrication, as these may
Framlington Place, Newcastle confer specific electric properties to the
liquid material connected to or being part
upon Tyne NE2 4HH, UK
of an appropriate electrochemical trans- resulting device. Also, a great number of
Keith Rawson ducer [1]. Any intrinsic property of a methodologies can be performed to
Cambridge Life Sciences, material or a specific process that could integrate the appropriate recognition
Cambridge Business Park, affect the conductivity of an electro- element with the transducer, which, in
Angel Drove, Ely, turn, gives rise to the generation of
chemical system can potentially be studied
Cambridgeshire CB7 4DT, UK
by EIS. It is a non-invasive technique that different interfaces with specific electric
does not require complex or expensive properties. In this context, the versatility
instrumentation and is easy to operate, of EIS allows its application to the control
thus allowing its current applications not and monitoring of the different stages
*Corresponding author.
only in research laboratories but also as a necessary for the fabrication of the sensor
Fax: +44-191-2227991;
E-mail: tool for the control of processes such as the and its eventual characterisation. Also,
performance of batteries, semiconductors, EIS has been used successfully as an

0165-9936/$ - see front matter ª 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.trac.2004.08.010 37
Trends Trends in Analytical Chemistry, Vol. 24, No. 1, 2005

analytical tool for the measurement of electric property
changes of the sensor system in the presence of
increasing concentrations of the corresponding analyte.
Based on the applications outlined above, the imped-
ance analysis of deterioration processes taking place on
polymer-coated transducers under controlled experi-
mental conditions appears to be an excellent transduc-
tion protocol in the development of electrochemical
sensors. To date, degradation mechanisms going from
pH changes to hydrolysis of the side chains of the poly-
mer to temperature gradients have all been monitored by
EIS and applied to the development of sensor systems for
a few specific applications. Nevertheless, those resulting
devices can be envisaged as generic platforms for the
measurement of an extensive range of analytes of
medical and environmental interest.
such circuits, a resistance ideally describes a conductive
path, such as that generated by the bulk conductivity of
the system or the charge-transfer step due to an
electrode reaction, whereas a capacitance generally
describes space-charge-polarisation regions within the
system as well as modification of an electrode surface
due to adsorption processes or polymer-layer deposition.
The Randles circuit is the simplest equivalent circuit
that describes an electrochemical cell where a single-step
Faradaic process in the presence of diffusion may occur
(Fig. 1(a)). It combines three components, namely the
electrolyte resistance between working and reference
electrodes ðRe Þ, the double-layer capacitance ðCdl Þ and
the Faradaic impedance due to the charge-transfer
process ðZf Þ at the working electrode-electrolyte inter-
face. Zf is normally subdivided into a charge-transfer

2. Theoretical background
Cdl
(a)
EIS measures the impedance of a system (Z) as a function
of frequency of an applied perturbation. When working
with electrochemical systems, this perturbation is
Re
normally an AC voltage of small amplitude (typically
5–10 mV peak-to-peak) and the response is a current
Zf
that differs in amplitude and phase (phase difference, /)
with the applied voltage.
The ratio of applied voltage to measured current is the Rct Zw
impedance of the system ðZ ¼ E=IÞ, which is easily
calculated over a wide frequency range, thus giving a (b) - Z’’
spectrum where processes with different kinetics that Charge transfer-limited process Diffusion-limited
process
may occur at the system under study are dominant at
different frequency regions. In this context, dipolar ωmax = 1/ Rct Cdl
properties will be reflected at high frequencies, whereas
bulk and surface properties will be evident at inter- ω
mediate and low frequencies, respectively. π/4
From a physical point of view, impedance is just a
totally complex resistance (measured in Ohms, X) that
appears when an AC current flows through a circuit Z’
made of resistors, capacitors, inductors or any combi- Re Rct
nation of these. This magnitude shows a complex nota- Re+Rct-2σ2Cdl
tion, with a resistive or real part attributable to resistors (c)
(in phase with the applied voltage) and a reactive or Zw 90
imaginary part attributable to the contribution of Rct
log Z φ
capacitors (out of phase with the applied voltage by
þp=2) and/or inductors (out of phase with the applied Cdl 45
voltage by
p=2):
pffiffiffiffiffiffiffi Re
Z ¼ Z 0 þ jZ 00 ¼ R
jX ; X ¼ 1=xC; j ¼
1;
0
where R is the resistance (measured in X), X the
reactance, C the capacitance (measured in Farads, F), log f
and x the applied angular frequency (measured in rad/s;
x ¼ 2pf , f is the frequency measured in Hz). Figure 1. (a) Randles equivalent circuit, (b) Nyquist Plot, and (c)
Experimental impedance data of an electrochemical Bode Plot that describe impedance behaviour of a simple electro-
chemical cell involving a single Faradaic process. Each dot of the
cell can be easily fitted to the impedance of an equivalent
Nyquist Plot represents the impedance at a given frequency.
circuit mainly comprising resistors and capacitors. In

38 http://www.elsevier.com/locate/trac
Trends in Analytical Chemistry, Vol. 24, No. 1, 2005 Trends

resistance ðRct Þ and the so-called Warburg Impedance

ðZw Þ, which reflects the influence of the mass transport of
the electroactive species on the total impedance of the
electrochemical cell. Thus, for those diffusion-limited
processes, Zw becomes dominant, whereas for those
charge-transfer-controlled processes, Zf is just Rct . All
these components can be identified by studying the
variation in impedance of an electrochemical cell over a
wide frequency range (normally 100 kHz–0.1 Hz). A
common way of showing the resulting data is the
complex plane or Nyquist Plot (Fig. 1(b)), in which
the real ðZ 0 Þ versus the imaginary ðZ 00 Þ components of
the impedance are plotted. In this plot, two separate
processes are very well differentiated, that is, a semicircle
relating to a charge-transfer-controlled process, the
intercept of which with the X axis gives Re and Rct
values, and a straight line with a slope of 1 due to Zw ,
whose extrapolation to the X axis allows calculation of
the Warburg coefficient ðrÞ, from which the diffusion
coefficients of the electroactive species can be estimated.
From the frequency at the top of the semicircle, where Z 00
is maximum, the time-relaxation constant ðsÞ for the
Faradaic process can also be calculated.
Another way of presenting impedance data is the Bode
Plot, in which the logarithm of the absolute value of Z
and the phase ð/Þ are plotted against the logarithm of
frequency ðf Þ. Fig. 1(c) shows the Bode Plots for the
Randles circuit. Unlike the Nyquist Plots, these data
presentations give direct information about f and / that
help ascertain the different constituent phases of the
system more easily. Thus, in those frequency regions
where a resistive behaviour is dominant, a horizontal
line is observed for the log Z– log f representation and a
/ close to 0
is measured. Also, capacitive behaviour
within a frequency region is described by a straight line
with a slope of )1 in the log Z– log f plots and a /
around 90
, whereas diffusion-controlled phenomena
(Warburg Impedance) would give a straight line with a
slope of )1/2 and a / of 45
. A detailed mathematical
description of all the above parameters can be found
elsewhere [3,4].
Impedance behaviour described by Nyquist Plots with
more than one semicircle or Bode Plots exhibiting dif-
ferent capacitive/resistive regions may describe electro-
chemical systems with several phases and/or more
complicated Faradaic processes (i.e., coating formation/
deterioration or adsorption phenomena on the trans-
ducer as well as coupled chemical reactions). Indeed, EIS
has proved to be an excellent technique to verify the
properties of polymer coatings as insulating materials
and follow their deterioration processes under specific
experimental conditions [5]. Fig. 2 shows the impedance
spectrum of an insulated polymer-coated electrode in
contact with an electrolyte solution. It can be modelled
by a resistance, the electrolyte resistance ðRe Þ, in series
with a capacitance, the geometric capacitance ðCg Þ. The
(a) Rb
Re Cg
(b) - Z’’
ω
Z’
Rb
Re
(c) Rb
90
log Z φ
Cg
45
Re
0
log f
Figure 2. (a) Equivalent circuit that best describes the impedance
behaviour of an insulating polymer-coated transducer and, also, of
a partially degraded system following a delamination process
(dotted-line circuit element describes possible imperfections on the
surface of the polymer coating). (b) Nyquist Plot (black dots indi-
cate intact polymer; white dots partially degraded polymer).
(c) Bode Plot (solid lines indicate intact polymer; dashed lines
partially degraded polymer). No contribution of Zw has been
considered.
polymer-coated electrode itself theoretically resembles a
pure capacitor, the capacitance of which ðCg Þ can be
calculated with the following equation:
Cg ¼ ee0 A=d;
where e is the dielectric constant of the polymer, e0 is the
permittivity of the free space, A is the electrode area and
d is the thickness of the polymer layer.
In practice, a bulk resistance ðRb Þ in parallel with
Cg normally appears and accounts for initial conduc-
tive pathways across the polymer ascribed to small
defects in the coating. This resistance is very high,
so the resulting RC parallel combination is not
reflected in the Nyquist Plot as a semicircle but as a
bend of the straight line, which is characteristic of
pure capacitive behaviour. By contrast, it is easily
identified in the Bode Plot by a horizontal line at low
frequency values.

http://www.elsevier.com/locate/trac 39
Trends Trends in Analytical Chemistry, Vol. 24, No. 1, 2005

Degradation processes occurring at this polymer-

(a) Cg
coated transducer are easily detected in the impedance
spectra, which also allow different processes of deterio-
ration to be differentiated. Delamination of polymer
Rct
coatings will lead to a decrease of the film thickness ðdÞ
and a consequent increase in Cg of the system, together
Re
with a decrease in the Rb that is now more easily
detected at low frequency values. This is reflected in the
Nyquist and Bode Plots, as shown in Fig. 2. The same Rb Cdl
equivalent circuit shown in Fig. 2 for an intact polymer-
coated transducer can be used to model the degradation (b) - Z’’
process.
By contrast, polymer breakdown due to the formation ω
of pores would give rise to significant differences in the
impedance spectra. Nyquist and Bode Plots at different
stages of degradation, together with the equivalent
circuit that best fits these results, are shown in Fig. 3.
Initially, the appearance of pores on the surface of the
coating gives rise to a decrease in Rb , which, in this case,
Z’
is a measurement of the resistance of the electrolyte
Re Rb Rct
filling such pores. The in-depth growth of those pores
would allow the electrolyte to penetrate the polymer and (c)
eventually reach the electrode surface and spread over it. 90
Rb
This new electrode–electrolyte interface is easily detected log Z φ
in the impedance spectra by the appearance at low
frequencies of a second semicircle (characterised by a Cg
Rct 45
second s), which is fitted to a resistor/capacitor parallel
combination in the equivalent circuit, namely a charge- Cdl Rb
transfer resistance ðRct Þ and a double-layer capacitance
Cg Re
ðCdl Þ, respectively. 0
For both degradation mechanisms described above,
the eventual dissolution of the polymer would lead to log f
impedance spectra similar to those previously described Figure 3. (a) Equivalent circuit that best describes the impedance
for a bare electrode and fitted by the Randles equivalent behaviour of a polymer-coated transducer partially degraded by
circuit. formation of pores and their corresponding, (b) Nyquist Plot (black
It should be pointed out that the impedance spectra dots indicate intact polymer; white dots and black triangles par-
tially degraded polymer), and (c) Bode Plot (solid lines indicate
and related equivalent circuits described above do not
intact polymer; dotted and dashed lines partially degraded poly-
always fit real situations. In such cases, the so-called mer). Semicircles 1 and 2 represent the partially degraded coating
distributed circuit elements are used to account for those and exposed transducer interface beneath the coating, respectively.
deviations from ideal behaviour. Those most commonly No contribution of Zw has been considered.
used are the Warburg Impedance ðZw Þ mentioned above
and the constant-phase element (CPE). Zw was first
introduced to explain diffusion-limited behaviour of
electrode processes at low frequencies by considering just impedance spectra due to the appearance of depressed
an ideal infinite-length diffusion line of electroactive semicircles in Nyquist Plots (with their centre below the
species from the bulk of the solution to the electrode real axis) and phase-angle values lower than 90
in Bode
surface. But an electrochemical cell is always finite in Plots. A thorough mathematical and physical descrip-
dimensions, so the Warburg Impedance had to be tion of distributed circuit elements is available [2,4, and
modified to fit more adequate finite-length diffusion references therein].
processes. The CPE is another generic distributed circuit
element, which stems from the fact that the electrode
surface is normally rough and/or the bulk properties of 3. Applications
the electrode material are inhomogeneous. This also
applies to polymer coatings in those real applications 3.1. Corrosion
described below. CPEs are normally added in equivalent To date, most of the research carried out on the degra-
circuits instead of ideal capacitors and are identified in dation of polymer coatings using EIS has aimed to

40 http://www.elsevier.com/locate/trac
Trends in Analytical Chemistry, Vol. 24, No. 1, 2005 Trends

address the behaviour of different polymers as protective

barriers against corrosion. The use of polymers to protect

References
different materials, such as steel and other metals/alloys,

[14–16]

[18,19]

[22,23]
against the attack of the environmental conditions to

[7,8]

[10]

[17]
[6]

[9]
which they are exposed has been very popular, with
many different polymers being designed for this purpose.

Transport and aerospace

Performance of primers,
Table 1 summarises several examples of corrosion

midcoats and topcoats

Architectural coatings

Domestic appliances
processes studied by EIS. This reflects the variety of

Aerospace industry

Aerospace industry

Integrated circuits
polymers tested against corrosion, the different electric
properties calculated from the impedance studies as well

Application
as the different range of industrial sectors where the set-

industry

on steel
up of a suitable procedure for monitoring corrosion

n.r.
phenomena has become very important.
Polyesters are often used as decorative architectural

Rb , Bulk (pore) resistance; Rct , Charge transfer resistance; Cg , Geometric capacitance; Cdl , Double-layer capacitance; fb , Breakpoint frequency.
coatings and, at the same time, as barriers against cor-

Water uptake and ionic diffusion

Blister formation in the coating

Moisture adsorption and pore

rosion. Because of the former role, they have usually

pores and high temperatures

Pitting on defective zones
been doped with pigments that negatively influence their

within the polymer voids

Degradation mechanism
protective properties. Mild steel panels were electrostat-
ically sprayed with clear and pigmented polyesters and
exposed to UV light for several time periods [6]. Imped-

Pore formation

Pore formation
of the coating
ance measurements were recorded as Nyquist Plots over

formation
a wide frequency range. The modelling of the system
with an appropriate equivalent circuit allowed calcula-

n.r.

n.r.
tion of bulk resistance ðRb Þ, charge-transfer resistance
ðRct Þ, coating (geometric) capacitance ðCg Þ, and interfa-
cial (double-layer) capacitance ðCdl Þ. From the values
Electric parameter

obtained, the deterioration of the polymer coatings was

Rb , Rct , Cg , Cdl

Rb , Rct , Cg , Cdl
assessed and related to an increased level of porosity of
Rct , Cg , Cdl

the films.
Similar studies were carried out on the evaluation of
Rb , Cg
Table 1. Main features of several EIS studies on polymer degradation due to corrosion

plasma-polymerised coatings using hexamethyldisilox-

Rb

Rb
fb
ane monomer on aluminium [7] and magnesium [8] Z
alloys as effective pre-treatments against corrosion. An
and high temperatures

appropriate equivalent circuit was modelled; it allowed

Artificial UV radiation

salt fog and UV light

Natural and artificial

Harrison’s solution,
Degradation agent

Harrison’s solution

evaluation of Cg and calculation of the water volume

Phosphatised mild steel Detergent solution

Harrison’s solution: 0.35% (NH4 )2 SO4 + 0.05% NaCl. n.r., Not reported.
NaCl or Na2 SO4

Microorganisms
within the film at the different stages of the degradation
(270–390 nm)

NaCl solution

process, thus suggesting initial delamination and blis-

seawater
solution

tering phenomena that would subsequently result in a

pitting process and failure of the protective coating.
Working in a similar way, lacquer- and epoxy-resin-
coated copper plates were exposed to artificial seawater
and their corrosion behaviour tested in terms of a
Al and Mg alloys

Aluminium alloy

Aluminium alloy

change in Cg , Cdl , the coating (pore) resistance ðRb Þ and

Stainless steel

Rct [9]. From this data, a three-stage degradation

Substrate

mechanism was suggested, namely water saturation of

Copper
Steel

Steel

the free volume of the coating, followed by electrolyte

penetration into the metal-coating interface that inten-
sified the corrosion process and caused a final loss of
adhesion of the film on the substrate.
poly(3-octadecyl pyrrole)

Alkyd, epoxy polyamide,

Poly(3-octyl pyrrole) and
Lacquer and epoxy resin

Acrylic electrodeposited

Cg and Rb changes were also useful in assessing the

hexamethyldisiloxane

conducting polymers
Plasma coating from

latex, polyurethane

protective behaviour against detergents of acrylic coat-

ings electrodeposited on phosphatised mild steel used in
domestic appliances [10].
Polyimide
Polyester
Polymer

Those applications shown above are described by two

coating
Sol–gel

well-differentiated processes shown in the corresponding

Nyquist Plots as two semicircles with different time-

http://www.elsevier.com/locate/trac 41
Trends
relaxation constants ðs1 =s1 > 20; 0:2 < ðR1 =R2 Þ < 5Þ,
which represent the coating and substrate (metal)
impedances at high and low frequency ranges, respec-
tively. This fact allowed a suitable description of the
polymer deterioration in terms of Cg , Cdl , Rb and Rct .
However, it is quite usual to have more complex
impedance spectra with superimposed semicircles, which
reflect processes with closer s, and/or straight lines
(slope ¼ 1) at low values of frequency, which relate to
diffusion-limiting phenomena within the system under
study (Warburg Impedance). The latter could also
be superimposed on other processes appearing in
the spectra at low-frequency ranges. All this makes the
calculation of the different parameters and the
interpretation of those processes cumbersome.
Interestingly, this drawback was easily circumvented
in the study of the degradation of an alkyd polymer
coating under saline and high humidity environments
[11]. All Nyquist Plots exhibited an apparent diffusion
tail at low frequencies, the slope of which increased as
the coating degraded, reaching a final value of 1 when
the coating detached from the substrate. Another
process appeared to be overlapped at this frequency
region and was responsible for this apparent change in
the Warburg Impedance, which was easily related to the
different stages of the corrosion process.
The study of the protective behaviour of an organo-
mineral polymer-based coating used as a topcoat in the
automotive industry introduced another useful correla-
tion between the so-called breakpoint frequency (fb ,
frequency at which / first falls to 45
when scanning the
spectrum from high to low frequencies) and the exposure
time of the coated substrate to a corrosive environment
[12]. This method enabled the quick detection of any
deterioration of the coating.
Unlike the examples described above, where an
evaluation of a specific coating was carried out to some
extent, some other reported studies made use of
impedance measurements just to compare the feasibility
of applying different coating materials and procedures
to a specific technology [13]. In this context, sol-gel
based coating systems [14–16] and electroactive-
conducting polymer coating systems [17] were envis-
aged as excellent alternatives to hazardous chromate
conversion coatings currently used in the aircraft
industry as surface treatments (primers) to prevent
corrosion on aluminium alloys just by recording and
comparing Nyquist and Bode Plots of the different
systems over long periods of time.
Following similar studies, several polymer-coating
systems on steel were tested upon exposure to natural
and artificial seawater [18,19]. It is remarkable that
these tests were carried out for more than two years at
remote test places using an experimental set-up that
allowed the collection of impedance spectra once a
week via modem. Coatings included primer, midcoat
42 http://www.elsevier.com/locate/trac
Trends in Analytical Chemistry, Vol. 24, No. 1, 2005
and topcoat layers of different materials (e.g., alkyd
solid, epoxy polyamide, latex and polyurethane). A
large amount of impedance data was collected and
interpretation of corrosion phenomena was carried out
in the first place by simply comparing the shape of
Bode Plots. A general decrease in impedance was an
indication of larger delaminated areas within the
substrate due to corrosion. Further evaluation involved
estimation of pore resistance, where the phase-angle
plot showed a minimum ðRbo Þ, and the breakpoint
frequency ðfb Þ, from which the delaminated or
corroded area was calculated. Rbo decreased steadily
with exposure time up to a certain time, at which it
increased because pores were plugged by corrosion
products and also bacteria.
In an another study from the same authors, imped-
ance spectra in the form of Bode Plots were collected to
help assess the growth of bacteria colonies on corrosion
products developed on polymer-coated steel when
exposed to marine water, independent of the coating
formulations [20].
Other related works have provided evidence of the
active role that colonies of microorganisms on the
surface of coated materials play in degradation [21].
The biosusceptibility of polyimide coatings was
assessed by measuring the decrease in Rb with time
observed in changes in the shape of Bode Plots at low-
frequency ranges [22,23]. Fungi caused an enhancement
of the transport of water and ionic species into the
polymer matrix, thus changing the dielectric properties of
the material and accelerating corrosion phenomena of
the coated substrates. This was very important in the
electronic field, where polyimides have been widely used
as insulating layers of integrated circuits.
Similar investigations were carried out with a wide
range of materials with potential uses in the aerospace
industry [24].
As described above, EIS has provided useful informa-
tion about corrosion phenomena occurring evenly on a
coated substrate. However, it is well documented that
corrosion is a localised process that tends to start at
areas of lowest ionic resistance, then spread very quickly
along the material surface. Local EIS (LEIS) is a recently
developed technique that has the potential to measure
localised degradation processes [25]. The design of dis-
placed micro-probes containing reference and counter
electrodes in a variety of arrangements has proved very
useful for recording impedance measurements locally
and creating three-dimensional surface-impedance maps
at a single frequency, which allow location of those
defects responsible for corrosion on the coated substrate.
From all the works referred to above, it is evident that
EIS plays an important role in evaluating and monitor-
ing the protective performance of polymer coatings
against corrosion. Such is the significance that it has led
to the creation of two artificial neural networks for the
Trends in Analytical Chemistry, Vol. 24, No. 1, 2005 Trends

automatic classification of polymer-coating quality on

Z , Impedance; Rb , Bulk (pore) resistance; Rct , Charge transfer resistance; Cg , Geometric capacitance; Cdl , Double-layer capacitance; IDEs, Interdigitated electrodes; hIgG, Human
steel substrates based on phase angle-log f and

References
log Z- log f data (Bode Plots) [26].

[28,29]

[32,41]

a-Chymotrypsin activity [31,34]

[35,36]
[27]

[30]

[33]
3.2. Sensors
Impedance measurements of degradation processes
occurring at polymer-coated transducers have recently

Collagenase activity
emerged as a means of detecting biological/chemical

Urea, creatinine

Oxidase activity
interactions directly or indirectly responsible for the

Inorganic salts
degradation of those coatings; thereby they have been

hIgG, urea
Analyte/s
exploited for analytical purposes. Table 2 gathers several
representative examples of these novel applications.

PSA
A few reports have dealt with a thorough description
of the impedance of polymers deposited on electrode
surfaces and their mechanism of degradation, the ulti-

Degradation mechanism

nucleophilic substitution
mate objective being to use these polymers in sensor
development. A good example was the study of different

Overoxidation and
phenomena occurring at polythiophene conducting

Pore formation

Pore formation

Delamination
Delamination
films, doped with CF3 SO3
, on platinum electrodes by EIS
[27]. This polymer was proposed as a good candidate for

Swelling
the development of electrochemical sensors for inorganic

n.r.
ions. The redox exchange behaviour of polythiophene
films was explained by studying the variation of Rb , Rct ,
Cg and Cdl with applied polarisation potential and film
thickness in aqueous solutions containing the doping Electric parameter
ion. As an example a significant increase of Rb at high Rb , Rct , Cg , Cdl
positive polarisation potentials accounted for a deacti-
vation of the polymer by overoxidative degradation and Rb , Cg

Rb , Cg
a total loss of its redox exchange properties.
Table 2. Main features of EIS studies on polymer degradation applied to sensor development

Cg

Z
Z
The thorough characterisation of two different poly-
acids, which underwent pH-dependent degradation
processes, gave insight into their potential use for the
Urease-induced increase of pH

Urease-induced increase of pH

Urease-induced increase of pH

development of biosensor devices involving enzyme-

related systems, whose corresponding reactions altered
a-Chymotrypsin enzyme
High positive potentials
(>1.2 V vs. Ag/AgClsat )

the pH of the reaction media. Methyl methacrylate/

Collagenase enzyme
methacrylic acid copolymer (Eudragit S100, R€ ohm
Degradation agent

immunglobulin G; PSA, Prostate specific antigen. n.r., Not reported.

Pharma) and methyl vinyl ether/maleic anhydride
copolymer modified by esterification with 1-octanol
(pH > 7.6)

(EMAC) degraded at pH above a threshold value because

(pH > 7)

(pH > 7)

H2 O2

of ionization of the carboxylic side groups within their

structure. The former broke down at pH > 7, while the
latter remained stable up to pH 7.6. Impedance studies
with gold screen-printed electrodes either spray or spin
coated with Eudragit S100 [28,29], and with gold or
Glycidylmethacrylate methylmethacrylate

carbon screen-printed electrodes coated with EMAC by

Methyl methacrylate-methacrylic acid

Methyl vinyl ether-maleic anhydride

either spin or screen-printing processes [29,30], were

copolymer (Eudragit S100)/Gold

copolymer (EMAC)/carbon IDEs

carried out at different degradation stages by halting the

Polythiophene/platinum wire

degradation process in buffered saline solutions of pHs

Eudragit S100/carbon IDEs

below their degradation-pH threshold value. Impedance

Poly(ester amide)/Gold

spectra in a wide frequency range were shown in the

copolymer/Gold disc
Polymer/transducer

Gelatin/Gold IDEs

form of Bode Plots. Both polymers exhibited capacitive

behaviour over a large frequency region, which is an
indication of their good insulating properties. However,
their degradation behaviour was completely different.
Bode Plots of Eudragit S100-coated gold electrodes (Fig.
4) and the calculation of derived electric parameters,

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Trends
Figure 4. Bode Plots of (a) intact and (b)–(e) partially degraded
Eudragit S100 polymer-coated transducers, and (f) bare transducer.
Spectra from (b) to (e) were obtained as the polymer film was more
extensively degraded. Degradation processes were carried out at
pH 7.8 and halted by rinsing the transducer thoroughly.
Measurements were then recorded at pH 5.2. Pore formation can
be inferred from the changes in the impedance spectra. (Reprinted
with permission from [28]. 1995 American Chemical Society.)
such as Cg , Cdl , Rb and Rct , accounted for the formation
of pores on the surface, which, at a certain stage,
reached the electrode surface and allowed the electrolyte
to penetrate and spread over it [28,29]. From these
Figure 5. Sensor arrangement for the determination of urea and human immunoglobulin G (IgG) based on the capacitance measurement of the
degradation of Eudragit S100 polymer-coated electrode.
44 http://www.elsevier.com/locate/trac
Trends in Analytical Chemistry, Vol. 24, No. 1, 2005
values, the relative porosity and the fraction of wetted
electrode area were estimated. The eventual dissolution
of the polymer layer was observed. More recently, com-
bined quartz crystal microbalance and electrochemical
impedance studies on Eudragit S100 spin-coated gold
electrodes were in agreement with this breakdown
mechanism [31].
Studies carried out with EMAC-coated electrodes
showed that this polymer did not dissolve but swelled,
remaining as a film on the electrode surface [29,30].
Swelling is due to the incorporation of water and elec-
trolyte cations that compensate the negative charges
created by deprotonation of the carboxylic moieties
within the polymer. The percentage of swelling of the
polymer could be estimated by comparing the Rb of
EMAC films before and after different breakdown stages.
The catalytic hydrolysis of urea by the enzyme urease
gives carbon dioxide and ammonia, thus causing a pH
increase in the reaction medium. This enzymatic reac-
tion was easily coupled to the breakdown process taking
place either on Eudragit S100- or EMAC-coated
electrodes at alkaline pH for the development of
electrochemical single-use biosensors for urea and
human immunoglobulin G (hIgG) [28], and urea and
creatinine [30], respectively.
Disposable Eudragit S100 spray-coated gold screen-
printed electrodes were put in close contact with acti-
vated cellulose membranes previously loaded with
urease and inserted into a Perspex well template [28].
The well was filled with a certain volume of an electro-
lyte solution at pH 6.5 containing a specific urea
concentration. All measurements were carried out in a
two-electrode arrangement using a glassy carbon
counter electrode (Fig. 5). Impedance measurements at a
Trends in Analytical Chemistry, Vol. 24, No. 1, 2005
single frequency were recorded. More precisely, the
increase of the electrode capacitance (extracted from the
imaginary component of the impedance) with time was
monitored at a set frequency of 20 kHz, where the
electrode behaviour was purely capacitive. Using
Eudragit S100 polymer layers of 1 lm thickness, their
degradation yielded a four orders of magnitude change
in capacitance, which proved the excellent sensitivity of
the device. Calibration curves for urea in a simulated
serum matrix over a concentration range of 1–100 mM
were obtained when plotting the ratio of the capacitance
value recorded at a specific time to the initial value of
capacitance for each urea concentration. The same
set-up was used for the determination of hIgG. Urease-
labelled anti-hIgG was used as a tracer for the develop-
ment of the analytical signal. Competitive and
non-competitive immunoassays were carried out with
activated cellulose membranes loaded with hIgG and
anti-hIgG, respectively. Following the several steps of the
corresponding immunoassay, the membranes were
placed over the electrode surface, inserted into the
Perspex well and measurements recorded in 100-mM
urea solutions in the same fashion as above. HIgG was
measured over a concentration range 0.0001–100 lg/
ml using both assay formats.
Because of the low pH-breakdown threshold of
Eudragit S100 (pH > 7.0), measurements of urea with
the urease device in whole blood, serum or plasma at
physiological pH cannot be carried out. When working
with immunoassays using urease as a label, all the
required steps have to be performed on the cellulose
membrane before placing it over the coated electrode,
which makes each experiment more tedious and the
Figure 6. Construction of an impedimetric sensor system for PSA based on a lateral-flow immunostrip and Eudragit S100 polymer-coated carbon
interdigitated electrodes (IDEs).
Trends
resulting device less appealing for commercial purposes.
However, the same research group has successfully
circumvented the above drawback by coupling a similar
sensitive impedimetric transducer with lateral flow
immunoassay formats developed on membrane strips
(Fig. 6) and demonstrated the approach for the
development of a fast, easy-to-use, single-use immuno-
sensor device for prostate specific antigen (PSA), the
most reliable tumour marker for the early diagnosis of
prostate cancer [32]. The non-specific degradation of the
polymer-coated transducer due to the natural pH of bi-
ological samples was easily avoided by keeping samples
in contact with the transducer for just a few seconds.
The anti-PSA antibody capture zone of a nitrocellulose
membrane strip was put in close contact with the
transducer, as depicted in Fig. 6. A certain volume of a
sample containing PSA was mixed with anti-PSA urease
conjugate and allowed to flow along the strip, then
reacting on the antibody capture zone and eventually
reaching the sink pad. The further rapid addition of a
urea solution on the opposite edge of the strip, required
for the detection of the immunocomplex formed on the
antibody capture zone, simultaneously washed away the
unbound species and the sample matrix from the strip,
thus preventing the non-specific breakdown of the
Eudragit S100 polymer layer. The overall process took
less than 2 min. Indeed, the sample reached the sink pad
in around 45 s and the washing step took just 30 s.
Capacitance measurements were started immediately
after adding the urea solution under the same experi-
mental conditions described for the capacitive sensor
approaches above. Fig. 7 provides a fine example of those
measurements recorded for increasing concentrations of
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Trends Trends in Analytical Chemistry, Vol. 24, No. 1, 2005

(Fig. 8). Thereafter, an adequate volume of the

900 [PSA] ngml-1 corresponding unaltered sample containing analyte
0 was added to the pad. Simultaneously, impedance
800
2 measurements at a single frequency were recorded
10 following the same experimental procedure as in the
700
30 devices described previously. Urea calibration curves
600
were obtained over a concentration range of 5–30
mM. In addition, this analyte was successfully deter-
500 mined in normal serum. However, this device was 5
C / C0

times less sensitive than that developed using Eudragit

400
300
200
100
0
0 200 400 600 800 1000
Time / s
Figure 7. Capacitance responses of the sensor depicted in Fig. 6
for several concentrations of PSA. C =C0 is the ratio of the recorded
capacitance (C) to initial capacitance ðC0 Þ. This ratio helps correct
the minimal variation of C0 of intact Eudragit S100 polymer-coated
transducers.
PSA, including concentration levels required for the
early detection of prostate tumours.
The higher pH at which EMAC polymer degrades
made this polymer a good feasible alternative to
Eudragit S100 for the construction of sensors for
clinical applications [30]. In order to address its per-
formance, single-use urea and creatinine sensors based
on the use of EMAC-coated-carbon screen-printed
interdigitated electrodes (IDEs) were developed. Pads of
a wicking material were loaded with either a urease
solution or a creatinine deiminase solution and stuck
over the working area of the coated IDE system
S100-coated transducers. Moreover, the calibration
range was narrower. Creatinine was measured at
concentration levels as low as 2 mM. Nevertheless, this
device was not sensitive enough for clinical analysis.
The degradation of acrylate bulk polymers by the
action of the hydrogen-peroxide product of an oxidase
enzyme has also been proposed as a feasible transduction
method for the development of biosensors based on
impedance measurements [33]. However, to our
knowledge, no true applications have yet been reported.
Some biodegradable polymers break down by the
direct action of enzymes (i.e., they act as the enzyme
substrate) and constitute another excellent approach
for the development of impedimetric biosensors. In this
context, a poly(ester amide) polymer, which is degraded
by the proteolytic enzyme a-chymotrypsin, was spun
onto a gold electrode and used as an impedimetric
transducer for the detection of the enzyme activity in
standard solutions [31,34]. Impedance spectra were
recorded over a frequency range 5 Hz–100 kHz every
30 s during the course of degradation, using a network
analyser that allowed fast collection of data. Results
were presented as log Z- log f Bode Plots (Fig. 9).
Resistive behaviour was observed at high frequencies
(electrolyte resistance, Re ), while, at lower frequencies,
the increase of impedance with decreasing frequency
indicated capacitive behaviour (geometric capacitance,
Cg ). Degradation of the film by the enzyme led to a

Figure 8. Sensor arrangement for the determination of urea and creatinine based on capacitance measurement of EMAC polymer-coated carbon
interdigitated electrodes (IDEs).

46 http://www.elsevier.com/locate/trac
Trends in Analytical Chemistry, Vol. 24, No. 1, 2005
Figure 9. Impedance spectra (Bode Plots) of poly(ester amide)
films recorded during its degradation by the enzyme a-chymo-
trypsin (electrolyte solution pH 7.3). A delamination process is
easily inferred from the shifts of the overall film impedance with
time. (Reprinted with permission from [31].  2002 American
Chemical Society.)
decrease of the impedance in the low-frequency range,
corresponding to an increase in the geometric capaci-
tance, which was related to the thinning of the polymer
film. When the polymer film was dissolved completely,
the observed capacitance in the spectrum equalled the
double-layer capacitance of the bare electrode. In this
way, a-chymotrypsin concentrations in the nanomolar
range were easily detected.
Similar experiments were carried out with a dextran
hydrogel polymer, which was degraded by the enzyme
dextranase [31,34]. However, the biodegradation pro-
cess that this polymer underwent could not be moni-
tored by EIS. Unlike the good insulating properties of
poly(ester amide) films, intact dextran hydrogels were
easily filled up with electrolyte molecules that wetted the
electrode surface. Thus, the geometric capacitance of
the system equalled the double-layer capacitance before
the degradation took place, which led to the recording
of the same impedance spectra before and after its
enzymatic dissolution. The need to use electrically
insulating polymers susceptible to undergoing degrada-
tion processes narrowed the range of materials used in
these approaches. Indeed, it has constituted one of the
major drawbacks of application of this methodology to
biosensor development.
The same problem was encountered in the develop-
ment of a biosensor for collagenase based on impedance
measurements of the enzymatic digestion of gelatin
deposited on interdigitated gold electrodes [35].
Collagenase could be detected only in solutions at very
low electrolyte concentrations (<2 mM), so as to avoid
electrolyte uptake by the polymer that would have made
the impedance of a non-degraded gelatin layer equal to
that of the electrolyte solution on a bare gold electrode.
Trends
Although not very suitable for the design of biosensors to
be used in electrolyte solutions, the same approach was
proposed for the detection of collagenase in air with
more or less success [36].
4. Summary and future trends
Overall, the easy monitoring of polymer-degradation
phenomena by EIS has been carried out extensively in
corrosion studies and, in recent years, has given rise to
the design of polymer-coated transducers for sensor
development.
Polymer-coated transducers have been seen as generic
platforms that could be coupled to biochemical interac-
tions i.e., enzyme-substrate or immunoassay systems,
directly or indirectly responsible for the deterioration of
those polymers, which can constitute or be part of the
biological recognition element for the design and the
development of biosensor devices [37,38]. Taking into
account those polymers reported so far and the wide
range of existing biochemical reactions that could po-
tentially be coupled to their breakdown processes, this
technology offers outstanding possibilities.
To date, just a few applications have turned out to be
true sensor devices. In all cases, disposable transducers
were used; that means that all the devices developed are
single-use sensors. Regarding the definition given by
IUPAC for an electrochemical biosensor i.e., a reagent-
less, self-contained, integrated device that is robust and
does not require additional separation steps [39], it is
well known that many of the biosensors reported in the
literature, especially affinity sensors based on immuno-
chemical or hybridisation reactions, do not meet all
these requirements.
Working with polymer-coated transducers and
enzyme-labelled immunoassay formats, scientists have
envisaged the design and development of integrated
single-use immunosensor systems for clinical applica-
tions that would meet the requirements for use as point-
of-care and bedside testing devices and, also, for the
analysis of environmentally relevant compounds that
could be carried out in situ by test systems in the field.
Indeed, a possible impedimetric immunosensor based on
a competitive assay format would make use of a-chymo-
trypsin as a label and enzyme-degradable poly(ester
amide) polymer-coated transducers [34]. It would
comprise a self-contained flow-through system that
would use a-chymotrypsin-analyte conjugates. The
excess of conjugate would be responsible for the break-
down of the polymer, thus giving an analytical signal
directly proportional to the concentration of analyte in
the sample.
Similarly, Eudragit S100 polymer-coated transducers,
in combination with urease-based immunoassay
systems, have proved to be excellent for the design of
http://www.elsevier.com/locate/trac 47
Trends
integrated, single-use immunosensors. Following the
work on a system described previously [32], such sensors
are based on lateral flow immunoassay formats, which
allow all the different assay steps to be integrated and
automated. In this case, the polymer is degraded by the
product of the enzymatic reaction, rather than by the
enzyme itself; that requires another step (addition of
substrate) to be integrated within the sensor system.
Small storage blisters containing urea-substrate solution
are easily inserted within the sensor in such a way that
the solution is automatically released into the system
once the immunoassay reactions have taken place [40],
thus providing an effective washing of any unbound
species from the system and initiating the specific
degradation of the polymer due to urea catalytic
hydrolysis by the bound urease label. The first one-step
immunosensor based on this set-up was proposed for the
analysis of free and total PSA in serum [41].
This is a novel measurement concept for the devel-
opment of electrochemical biosensors, which shows
significant advantages over other more commonly used
approaches. and, as such, it could evolve towards the
construction of single-use sensor devices with real
applications to the detection of a wide variety of mole-
cules of interest in those areas where it is very important
to obtain fast, reliable measurements with cost-effective,
easy-to-use devices.
Acknowledgements
C
esar Fern
andez-S

anchez is grateful to the European
Commission for the award of a Marie Curie Fellowship
(Contract number HPMF-CT-2002-01992) of the
Human Potential Programme. We thank Ana M.
Gallardo-Soto for helpful discussions during preparation
of this review.
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