Deposition of new thia-containing Schiff-base iron (III) complexes onto carbon nanotube-modified glassy carbon electrodes as a biosensor for electrooxidation and determination of amino acids

Article abstract of DOI:10.1016/j.electacta.2010.10.031

Multiwall carbon nanotubes (MWCNTs) were used as an immobilization matrix to incorporate an Fe (III)-Schiff base complex as an electron-transfer mediator onto a glassy carbon electrode surface. First, the preheated glassy carbon was subjected to abrasive immobilization of MWCNTs by gently rubbing the electrode surface on filter paper supporting the carbon nanotubes. Second, the electrode surface was modified by casting 100 μL of an Fe (III)-complex solution (0.01 M in ACN). The cyclic voltammograms of the modified electrode in an aqueous solution displayed a pair of well-defined, stable and nearly reversible reductive oxidation redox systems with surface confined characteristics. Combinations of unique electronic and electrocatalytic properties of MWCNTs and Fe (III)-Schiff base complexes resulted in a remarkable synergistic augmentation of the response. The electrochemical behavior and stability of the modified electrode in aqueous solutions at pH 1-9 were characterized by cyclic voltammetry. The apparent electron transfer rate constant (K_s) and transfer coefficient (a) were determined by cyclic voltammetry and were approximately 7 s^-1 and 0.55, respectively. The modified electrodes showed excellent catalytic activity towards the oxidation of amino acids at an unusually positive potential in acidic solution. They also displayed inherent stability at a wide pH range, fast response time, high sensitivity, low detection limit and had a remarkably positive potential oxidation of amino acids that decreased the effect of interferences in analysis. The linear concentration range, limits of detection (LOD), limits of quantization (LOQ) and relative standard deviation of the proposed sensor for the amino acid detection were 1-55,000, 1.10-13.70, 2.79-27.14 and 1.30-5.11, respectively.

Full text of DOI:10.1016/j.electacta.2010.10.031

Electrochimica Acta 56 (2011) 1051–1061
Contents lists available at ScienceDirect
Electrochimica Acta
journal homepage: www.elsevier.com/locate/electacta
Deposition of new thia-containing Schiff-base iron (III) complexes onto carbon
nanotube-modified glassy carbon electrodes as a biosensor for electrooxidation
and determination of amino acids
Lotfali Saghatforousha, Mohammad Hasanzadeha,b,∗, Nasrin Shadjoua,b, Balal Khalilzadehc
a Department of Chemistry, Faculty of Science, Payame Noor University (PNU), P.O. Box 58168-45164, Khoy, Iran
b Department of Chemistry, Faculty of Science, K.N. Toosi University of Technology (KNTU), Tehran, Iran
c Department of Analytical Chemistry, Faculty of Science, Arak University, Arak, Iran
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 6 August 2010
Received in revised form 4 October 2010
Accepted 10 October 2010
Available online 16 October 2010
Multiwall carbon nanotubes (MWCNTs) were used as an immobilization matrix to incorporate an Fe
(III)–Schiff base complex as an electron-transfer mediator onto a glassy carbon electrode surface. First,
the preheated glassy carbon was subjected to abrasive immobilization of MWCNTs by gently rubbing
the electrode surface on filter paper supporting the carbon nanotubes. Second, the electrode surface was
modified by casting 100 ␮L of an Fe (III)-complex solution (0.01 M in ACN). The cyclic voltammograms
of the modified electrode in an aqueous solution displayed a pair of well-defined, stable and nearly
reversible reductive oxidation redox systems with surface confined characteristics. Combinations of
unique electronic and electrocatalytic properties of MWCNTs and Fe (III)–Schiff base complexes resulted
in a remarkable synergistic augmentation of the response. The electrochemical behavior and stability
of the modified electrode in aqueous solutions at pH 1–9 were characterized by cyclic voltammetry.
The apparent electron transfer rate constant (Ks) and transfer coefficient (a) were determined by cyclic
voltammetry and were approximately 7 s−1 and 0.55, respectively. The modified electrodes showed excel-
lent catalytic activity towards the oxidation of amino acids at an unusually positive potential in acidic
solution. They also displayed inherent stability at a wide pH range, fast response time, high sensitivity,
low detection limit and had a remarkably positive potential oxidation of amino acids that decreased
the effect of interferences in analysis. The linear concentration range, limits of detection (LOD), limits of
quantization (LOQ) and relative standard deviation of the proposed sensor for the amino acid detection
were 1–55,000, 1.10–13.70, 2.79–27.14 and 1.30–5.11, respectively.
Keywords:
Carbon nanotubes
Amino acids
Modified electrode
Schiff base
© 2010 Elsevier Ltd. All rights reserved.
1. Introduction
the overpotential required for either oxidation or reduction of com-
pounds. Many mixed valent compounds, such as oxides, complexes,
The electrochemical modification of electrodes with a suit-
able reagent has been widely used for analytical applications. The
resulting electrodes were designed to provide desired selective
sites towards the analytes. Electrochemically modified electrodes
have played an important role in studying electrocatalysis [1–4],
electron transfer kinetics [5,6], membrane barriers [7,8] and elec-
troorganic syntheses [9]. One of the most important electrode
modification techniques involved the formation of an electrocat-
alytic system where redox species were capable of undergoing a
rapid and reversible electrode reaction. These electrodes reduced
or alloys of copper [10–12] cobalt [13–15] and ruthenium [16–18],
have shown electrocatalytic properties.
Carbon nanotubes are new, novel and interesting membranes
comprised of nanostructure materials; they are promising for use
in immobilization substances because of their significant mechan-
ical strength, high electrical conductivity, high surface area and
good chemical stability. The electrocatalytic response of carbon
nanotubes (CNTs) has received considerable attention [19–23].
The surface of CNTs can be chemically modified to impart a spe-
cific property; this modification can be done either covalently or
by physical adsorption [24–27]. Immobilization of molecules and
biomolecules on CNTs has been pursued previously, motivated by
the prospects of using nanotubes as new types of sensors and
biosensors. Electronic properties of nanotubes coupled with the
specific recognition properties of the immobilized systems would
provide ideal miniaturized sensors. A prerequisite for research in
∗
Corresponding author at: Department of Chemistry, Faculty of Science, Payame
Noor University (PNU), P.O. Box 58168-45164, Khoy, Iran. Tel.: +98 461 2344384;
fax: +98 461 2332556.
E-mail address: mhmmd hasanzadeh@yahoo.com (M. Hasanzadeh).
0013-4686/$ – see front matter © 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.electacta.2010.10.031

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L. Saghatforoush et al. / Electrochimica Acta 56 (2011) 1051–1061
OH
EtOH
+
FeCl3
+
Reflux
N
S
NH2
Br
CHO
S
N
Fe
Cl2
O
Br
N
Scheme 1. General procedure for synthesis of the complex.
this area is the development of a method for the immobilization of
new molecules and biomolecules on the carbon nanotubes reliably.
The compatibility and electrochemical applications, including the
immobilization of a variety of species on the external and inter-
nal surfaces of MWCNTs of carbon nanotubes, have been reported
[26–32]. Recently, cobalt hydroxide nanoparticles on modified
electrodes were used for the identification of biological molecules
[33–35].
The use of bare, unmodified electrodes as electrochemical sen-
sors for morphine and its derivatives had a number of limitations.
Low sensitivity and reproducibility, slow electron transfer reac-
tions, low stability over a wide range of solution compositions and
high over potential at which the electron transfer process occurs
were the limitations identified. Typically, the CNTs were immobi-
lized on to the electrode surfaces by using a CNT solution as the
dispersing medium. As carbon nanotubes are insoluble in most
solvents, and dissolution processes require a long period time, a
simple and reliable method to fabricate electrode surfaces modi-
fied with CNTs needed to be developed. One of the best materials
capable of modification by carbon nanotubes was a glassy carbon
(GC) electrode that was typically polished first. Unlike a basal plane
pyrolytic graphite electrode, direct adhesion of carbon nanotubes
on the surface of a GC electrode is difficult and the surface should
be pretreated for adhesion of the carbon nanotube particles. The
Schiff base derived from salicylaldehyde (salens), which is used as
a polydentate ligand, is known to form very stable complexes with
transition metal ions. An important area of research involves the
syntheses of transition metal complexes containing thiolate coor-
dinations, which have implications in bioinorganic chemistry and
medical chemistry. Transition metals complex with Schiff bases to
become ligands and have been used as catalysts in diverse processes
such as oxygen- and atom-transfer, enantioselective epoxidation,
aziridation and mediation of organic redox reactions. They have
also been used as mediators in other oxidation processes. Although
extensive scientific reports exist on the synthesis, characterization
and crystalline structure of the transition metal–salen complexes
in the solid state, information about the resulting complex species
and their formation equilibrium in solution is limited. In this paper,
we describe the first synthesis of thia-containing salens and the use
of newly synthesized salens (Scheme 1) as suitable biosensors for
the selective determination of various amino acids. In the current
study, a simple two-step procedure was used for the modification
of glassy carbon with MWCNTs and an Fe (III)–Schiff base com-
plex. Continuations of our research included the preparation of
modified electrodes and their applications as a detector of biologi-
cal molecules [36–40]. Additionally, Fe (III)–Schiff base-MWCNTs
modified onto glassy carbon electrodes were evaluated for the
detection of various amino acids.
Scheme 2. FTIR spectrum of an Fe (III)–Schiff base complex.

L. Saghatforoush et al. / Electrochimica Acta 56 (2011) 1051–1061
1053
cedure in Scheme 1 [41]. The structure of the product was
confirmed by CHN analysis and by IR (Scheme 2). Mp, 210 ◦C
(decomposed); Fw, 527.066 g mol−1, elemental analysis: Calcd. for
C16H20BrCl2FeN2SO3: C, 36.46%; H, 3.82%; N, 5.31%. Found: C,
36.68%; H, 3.77%; N, 5.52%. IR (KBr): 3450 cm−1 n(O–H), 3030 and
3080 cm−1 n(C–H) aromatic, 2840 and 2950 cm−1 n(C–H) aliphatic,
1618 cm−1 n(C N) imin.
3.2. Preparation of modified Fe (III)–Schiff base–MWCNTs-GC
and Fe (III)–Schiff base-GC electrodes
The glassy carbon electrode was carefully polished with alu-
mina and a polishing cloth, and the electrode was placed in ethanol
and subjected to vibration to remove adsorbed particles. After pol-
ishing, the electrode was heated for 5 min at 50 ◦C in an oven.
Two milligrams of purified MWCNT was placed on a piece of filter
paper (Whatman. No. 1001 110). Next, the MWCNTs were abra-
sively immobilized onto the electrode. Immediately after removing
the preheated GC electrode from the oven, the electrode surfaces
were gently rubbed against fine qualitative filter paper containing
carbon nanotubes for 1 min. The electrode was rinsed with ethanol
and distilled water. After 20 cycle scans, which were carried out in
the potential range from −1.5 to 1.5 V with a reference electrode
in a solution of 1 M H2SO4, 50 ␮L of freshly prepared Fe (III)–Schiff
base complex (0.1 M in ACN solution) was doped onto the glassy
carbon electrode fabricated with MWCNTs. The modified electrode
was dried for 1 h in room temperature. The modified electrode was
rinsed with distilled water before use.
The adsorption of the Fe (III)-complex onto the surface of the
GC electrode was carried out in two steps: (a) the GC electrode was
held under a constant potential of 1.8 V for 20 min in 1 M sulfuric
acid solution, and (b) the pre-anodized GC electrode was immersed
in 0.1 M Fe (III)-complex solution in ACN for 30 min.
Fig. 1. (A) SEM image of the GC electrode modified with MWCNTs; scale bar is
200 nm. Inset is the SEM image with higher magnification for the same sample. (B)
Same as MWCNTS modified with Fe (III)–Schiff base complexes.
3.3. Characterization of Fe (III)–Schiff base complexes
Fig. 1A shows the scanning electron microscope (SEM) images
of MWCNTs and Fe (III)–Schiff base complexes with modified
MWCNTs. It is evident that the MWCNTs are very long and
present as a highly entangled network structure; it is attributed
2. Experimental details
All chemicals used were of analytical grade from Merck (Darm-
stadt, Germany) and were used without further purification.
All solutions were prepared with doubly distilled water. Elec-
trochemical measurements were carried out in a conventional
three-electrode cell powered by an electrochemical system com-
prising an AUTOLAB system with PGSTAT12 boards (Eco Chemie,
Utrecht, Netherlands). The system operated on a PC using GPES
4.9 software. A saturated calomel electrode (SCE) was used as a
reference electrode. All potentials were measured with respect
to the SCE, which was positioned as close to the working elec-
trode as possible by means of a Luggin capillary. Buffer solutions
(0.1 M) were prepared from sulfuric acid (H2SO4), phosphoric acid
(H3PO4) and disodium hydrogen phosphate (Na2HPO4). Hydrogen
chloride (HCl) and sodium hydroxide (NaOH) were used for pH
adjustment. The solutions were degassed by bubbling high purity
(99.99%) argon gas through them prior to the experiments. Elemen-
tal analyses for CHN were performed using a Heraeus CHN-O-RAPID
elemental analyzer. IR spectra were recorded on a Shimadzu model
FTIR prestige 21 spectrophotometer using KBr discs. The surface
morphology of the modified electrodes was evaluated with a Vega-
Tescan electron microscope (SEM).
3. Results and discussion
3.1. Synthesis of Fe (III)-complexes
Fig. 2. (a) Cyclic voltammetry response for MWCNTs modified GC electrode, (b) as
The target Schiff-base metal-complex, Fe (III)–Schiff Base,
was synthesized in one-step reaction according to the pro-
(a) for the Fe (III)-complex-MWCNTs modified GC electrode, buffer solution (pH 2)
and scan rate 100 mV s−1
.

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40
35
30
25
20
15
10
5
0
-5
-10
0.2
0.3
0.4
0.5
0.6
0.7
0.8
E(V)
300
200
100
0
50
40
30
20
10
0
y = 9.321x - 56.28
R2 = 0.991
a
y = 0.526x - 1.165
R2 = 0.99
b
-100
-200
-300
-10
-20
-30
y = -11.518x + 10.45
y = -0.2898x + 1.6724
2
2
R = 0.9945
R
= 0.9971
30
0
60
90
8
13 18
23
28
33
v/(mV s-1
)
v0.5/ (mV s-10.5
)
0.8
0.7
c
0.6
0.5
0.4
-3
-2
-1
log v/(mV s-1
0
1
)
Fig. 3. Cyclic voltammetry response of the Fe (III)-complex-MWCNTs modified GC electrode in the buffer solution (pH 2) at scan rates (inner to outer) 10, 20, 40, 50, 70 and
100 mV s−1 (main panel). (a and b) The plots of peak currents vs. square root of scan rates and scan rate. (c) The plot of peak potential separation vs. log v.
to low solubility of the CNTs in most solvents. Furthermore, the
images reveal that the CNTs randomly distributed across the elec-
trode surface. The inset of this figure shows the SEM image of
the same sample with a higher magnification (the scale bar is
approximately 200 nm). The diameter of MWCNTs is approxi-
mately 15–20 nm. Fig. 1B displays the SEM images of MWCNTs
modified with the adsorption of Fe (III)–Schiff base complexes.
The inset of this figure clearly shows that the diameters of the
MWCNTs are about 40 nm (two times the diameter of the MWC-
NTs without Fe (III)–Schiff base complexes). This finding indicates
that a high quantity of the Fe (III)–Schiff base complex has
adsorbed onto the surface of the MWCNTs after immersing a
MWCNT-modified electrode in the Fe (III)–Schiff base complex
solution.
3.4. Electrochemical properties and the stability of glassy carbon
electrodes modified with Fe (III)–Schiff base complexes and
carbon nanotubes
Fig. 2 illustrates the cyclic voltammograms of glassy carbon
electrodes modified with the immobilization of CNTs and a glassy
carbon electrode modified with CNTs and Fe (III)–Schiff base com-
plexes in buffer solution (pH 2) at scan rate of 100 mV s−1. A
well-defined single redox couple with oxidation/reduction poten-
tials of 0.415 and 0.530 mV were discovered for glassy carbon
electrodes modified with Fe (III)-complexes by CNTs (Fe (III)–Schiff
base complex modified MWCNTs). However, in the glassy carbon
electrode modified with MWCNTs, no peak was observed at the
potential range from 0.0 to 1.0 V.

L. Saghatforoush et al. / Electrochimica Acta 56 (2011) 1051–1061
1055
Fig. 4. (A) The cyclic voltammograms of the Fe (III)-complex-MWCNTs modified GC electrode in buffer solution (pH 2) in the absence (a) and the presence (b) 5 mM Cys.
Potential sweep rate was 100 mV s−1. (B) Dependency of the anodic peak current electro-oxidation on Cys concentrations. (C) Current function vs. v0.5 in the buffer solution
(pH 2) in the presence of 5 mM Cys.
Fig. 3 displays typical cyclic voltammograms for modified glassy
carbon electrodes in a buffer solution (pH 2) at different scan rates.
Fig. 3(a and b) shows that the anodic and cathodic peak currents
are directly proportional to the scan rate of the potential at a sweep
rate from 10 to 100 mV s−1. The plot of peak current vs. the scan
rate up to 100 mV s−1 had good linearity as expected for surface
confined redox processes. The small peak-to-peak separation and
good linearity between the peak currents and scan rates at sweep
rates of 10–100 mV s−1 suggested facile charge transfer kinetics. At
higher scan rates, the peak currents vs. the scan rate plots deviated
from linearity and the peak currents became proportional to the
square root of the potential scan rate. The data indicated that the
peak current was diffusion controlled. In addition, with increas-
ing scan rates, the peak separations began to increase, indicating
that the limitation arose from charge transfer kinetics. Based on
Laviron theory [40], the electron transfer rate constant (Ks) and
charge transfer coefficient (˛) could be determined by measuring
the variation of peak potential with scan rate.
the associated anodic charge increased drastically. Additionally, the
ratio Ipa/Ipc was 79.4. In absence of Cys, the ratio was decreased to
28.7. The results indicated that Cys was oxidized by an active Fe (III)
moiety through a cyclic mediation redox process. Iron species were
immobilized onto the electrode surface, and the one with the higher
valence number oxidized to Fe (III) via a chemical reaction followed
by the generation of low-valence iron (Fe (II) species). Similarly, the
high-valence oxide is regenerated through the external electrical
circuit. Accordingly, Cys is oxidized via an ECꢀ mechanism.
Curiously, the Fe (III)-catalyzed oxidation of free amino acids is
almost completely dependent upon the presence of the bicarbonate
ion, and it is greatly stimulated by the Fe (III)–Schiff base complex.
It is assumed that the most active catalytic complex is composed
of two equivalents of HCO3, an amino acid and at least one equiv-
alent of iron; however, two forms of iron must be involved. In the
presence of Fe (III) or Cu (II), O2, and an electron donor, a num-
ber of metal-catalyzed oxidation (MCO) systems are able to oxidize
amino acids. These studies have established that Val, l-Phe, l-Arg,
Gly, Ser, Tyr, Cys and l-Trp are among the most common sites of
oxidation by MCO systems.
The peak potential values were proportional to the logarithm
of the scan rate for rates higher than 100 mV s−1 (Fig. 4). The cal-
culated values for (Ks) and (˛) were approximately 6 s−1 and 0.56,
respectively. The surface concentration of electroactive species (ꢀ )
can be calculated from the slope of the plot of Ipa vs. scan rate and
is about 2.04 × 10−9 mol cm2.
The redox transition of iron species present in the film is:
k
1
Fe(II) ꢀ Fe(III) + e
(1)
k−1
Fig. 4A presents the CVs of Fe (III)–Schiff base complex elec-
trodes, both in the absence (a) and in the presence (b) of 5 mmol L−1
Cys in buffer solution. At the Fe (III)–Schiff base complex electrode,
oxidation of Cys resulted in a typical electrocatalytic response. In
the presence of Cys, it was observed that the anodic current and
and Cys is oxidized on the modified surface via the following reac-
tion:
K
(E)
2
Fe(III)-complex + cysteine−→ intermediate + Fe(II)
(2)

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L. Saghatforoush et al. / Electrochimica Acta 56 (2011) 1051–1061
Scheme 3. Oxidation mechanism of Gly (A), Cys (B), Val (C), l-Phe (D), Ser (E), Tyr (F), l-Trp (G) and l-Arg (H).
The intermediate is further oxidized to the product through a
similar electro-oxidation process:
acids, therefore, demonstrating its exhibiting capability for selec-
tive oxidation of amino acids. All mechanisms for the studied amino
acids are presented in Scheme 3.
Fig. 4B and C shows that upon increasing the Cys concentration,
irreversible oxidation develops in the region of the electrochemical
formation of Fe (III). Thus, it is likely that the electro-generated
K
(E)
3
Fe(III)-complex + intermediate−→ product + Fe(II)
(3)
The oxidation process of the Fe (III)–Schiff base complex elec-
trode exhibited similar electrocatalytic responses for other amino

L. Saghatforoush et al. / Electrochimica Acta 56 (2011) 1051–1061
1057
Scheme 3. (Continued.)
Fe (III) species is the active moiety that efficiently speeds up the
oxidation of the amino acids. Any increase in the concentration of
amino acids causes a proportional (almost linear) enhancement of
the anodic wave. It is emphasized that the anodic formation of Fe
(III) appears to be an irreversible process. Also, plotting the current
function (peak current divided by the square root of the potential
sweep rate) against the square root of the potential sweep rate
(Fig. 4C) revealed a negative slope confirming the electrocatalytic
nature of the process. Similar cyclic voltammograms were recorded
for Val, l-Phe, l-Arg, Gly, Ser, Tyr, Cys and l-Trp. The values of ˛ were

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L. Saghatforoush et al. / Electrochimica Acta 56 (2011) 1051–1061
Scheme 3. (Continued.)
obtained for the listed amino acids according to the documented
method are reported in Table 1.
surface was almost 94% of its initial value after 200 cycles (Fig. 5).
In addition, no significant decrease was observed after replacing
the electrolyte used for 200 repetitive cycles with the fresh buffer
solution. Furthermore, the storage stability of the chemically mod-
ified electrodes was very good. The electrodes reserved 97% of their
initial activity for more than 1 month when stored in air at room
temperature.
The high stability of the adsorbed Fe (III)–Schiff base complex
against desorption in an aqueous solution is related to the fol-
lowing: chemical and mechanical stability of the nanotube film,
the strong interaction of aromatic groups of the complex with
␲-staking of CNTs, and the possible interaction between the Fe
(III)–Schiff base complex and activated CNTs. Upon examination
of the reproducibility of the electrode preparation procedure, five
independent GC electrodes were modified with MWCNTs and the
Fe (III)–Schiff base complex. Cyclic voltammograms of prepared
modified electrodes in the buffer solution were recorded. The rel-
ative standard deviation values of the measured cathodic peak
currents were 4%. Due to the chemical stability, electrochemi-
cal reversibility, and high electron transfer rate constant of Fe
(III)–Schiff base complex used in this study, they can be widely
The working stability of the modified electrode was verified by
monitoring the remaining amount of active substance after succes-
sive sweeps of cyclic voltammograms. The peak height and peak
potential of the immobilized redox system during potential cycling
over the range 0.40–0.58 V remained nearly constant. The quan-
tity of the Fe (III)–Schiff base complex remaining on the electrode
Table 1
Values of the electrocatalytic reaction rate constants (k) and the diffusion coeffi-
cients (D) obtained from chronoamperometry curves.
Amino acid
D (cm2 s−1
)
k (cm3 mol−1 s−1
)
˛
Cys
Gly
Tyr
Ser
10−5 × (4.61 0.03)
10−5 × (5.19 0.02)
10−5 × (6.41 0.02)
10−5 × (2.06 0.04)
10−6 × (4.94 0.03)
10−5 × (4.08 0.03)
10−5 × (2.11 0.03)
10−6 × (8.22 0.02)
10+5 × (2.09 0.03)
10+6 × (1.20 0.02)
10+5 × (3.44 0.02)
10+6 × (6.23 0.04)
10+6 × (5.33 0.03)
10+5 × (2.79 0.03)
10+6 × (3.46 0.03)
10+5 × (4.11 0.02)
0.56 0.03
0.47 0.02
0.54 0.03
0.61 0.03
0.55 0.02
0.47 0.04
0.62 0.03
0.47 0.03
Val
l-Arg
l-Phe
l-Trp

L. Saghatforoush et al. / Electrochimica Acta 56 (2011) 1051–1061
1059
40
30
20
10
0
d
a
-10
-20
-0.4
-0.2
0
0.2
0.4
0.6
0.8
1
E/V
Fig. 5. (a) The 1st, (b) 20th, (c) 50th and (d) 200th cyclic voltammograms of GC
electrode modified with MWCNTs and Fe (III)–Schiff base complexes (immersing
time for electrode modification was 30 s) in the acidic buffer solutions (pH 2). Scan
rate was 100 mV s−1
.
used in electrocatalysis as electron transfer mediators. One of the
modified electrodes and the GC electrode modified with the Fe
(III)–Schiff base complex and MWCNTs were used for the evaluation
of the catalytic ability of the modified electrode in the electrocat-
alytic process.
Fig. 6. (A) Variation of anodic peak current and (B) variation of peak potential with
pH for 1 mM Cys solution at an Fe (III)–Schiff base complex modified electrode.
Sweep rate was 100 mV s−1
.
During the oxidation of Cys, the anodic peak potential shifted
to more negative values with increasing pH. However, as seen in
Fig. 6A, with an increase of the solution pH, the anodic wave became
more broad at pH 2, revealing that the Fe (III)–Schiff base com-
plex had better electrocatalytic activity for the oxidation of Cys as
the acidity increases. Fig. 6B illustrated that Ep decreased when
increasing the pH value, and the current response for dopamine
oxidation was high at pH 2. It was clearly supported that pro-
tons are involved in the oxidation process. A similar behavior was
observed for other amino acids. Based on both high peak current
and low possible peak potential, pH 2 was selected as the work-
ing pH value to carry out the square wave voltammetry and DPV
determination of these compounds. It was concluded that the Fe
(III)–Schiff base complex catalyzed the oxidation of these amino
acids.
Chronoamperometry, as well as cyclic voltammetry
, was employed for the investigation into the processes occurring
via the ErCi mechanism [42]. Double step chronoamperograms
were recorded by selecting the working electrode potentials
to desired values and were used to measure the catalytic rate
constant on the modified surface. Fig. 7A displays double step
chronoamperograms for the modified electrode in the absence (a)
and presence (b: 1 mM, c: 4 mM, d: 6 mM, e: 8 mM, f: 10 mM and
g: 25 mM) of Cys within a concentration range of 1–25 mM. The
applied potential steps were 415 and 530 mV, respectively. The
plot of net current vs. t−0.5, which had been obtained by removing
the background current by the point-by-point subtraction method,
provided a straight line (Fig. 7B). The transient current must be
controlled by a diffusion process. The transient current was due to
the catalytic oxidation of Cys, which increased as the Cys concen-
tration was raised. No significant cathodic current was observed
when the electrolysis potential was stepped to 415 mV/SCE. When
examining the irreversible nature of the oxidation of amino acids
by using the slopes of these lines, we could obtain the diffusion
coefficients of the amino acids according to the Cottrell equation
[42, p. 163]:
I = nFAD1/2C∗ꢁ−1/2t−1/2
(4)
where D is the diffusion coefficient, and C* is the bulk concentration.
The mean value of the diffusion coefficients of Cys was determined
to be (4.61 0.03) × 10−5 cm2 s−1. These values are in agreement
with those obtained using cyclic voltammetry (Fig. 7).
The rate constants of the reactions for Cys and the ensuing inter-
mediates with the redox sites of the Fe (III)–Schiff base complex
electrode can be derived from the chronoamperograms according
to Eq. (5) [42, p. 503]:
ꢀ
ꢁ
Icatal
exp(−ꢂ)
= ꢂ1/2 ꢁ1/2erf (ꢂ1/2) +
(5)
ꢂ1/2
Id
where Icatal is the catalytic current in the presence of Cys, Id the
limiting current in the absence of Cys and ꢂ = kCt (k, C and t are the
catalytic rate constant, bulk concentration of Cys and the elapsed
time, respectively) is the argument of the error function. For ꢂ > 1.5,
erf(ꢂ1/2) almost equals unity and Eq. (5) reduces to:
Icatal
= ꢂ1/2ꢁ1/2 = ꢁ1/2(kCt)1/2
(6)
Id
Using the slope of the Icatal/Id plot, the value of k at a given
concentration of Cys can be derived (Fig. 4C). The mean value of

1060
L. Saghatforoush et al. / Electrochimica Acta 56 (2011) 1051–1061
Fig. 7. (A) Double step chronoamperograms of the Fe (III)-complex-MWCNTs modified GC electrode in the absence (a) and the presence of 1 (b), 4 (c), 6 (d), 8 (e), 10 (f) and
25 mM (g) Cys in buffer solution (pH 2). Potential steps were 530 mV and 415 mV, respectively. (B) Dependency of transient current on t−0.5. (C) Dependence of Icatal/Id on t0.5
derived from the data of chronoamperograms of (a) and (g) in main panel.
k in the concentration range of 1–25 mM Cys was found to be
tions in the range of the standard series. The intra-assay RSDs of the
proposed method, that was determined on the basis of peak cur-
rent for 10 replications, was 2.97% for peaks a1, and showed good
repeatability. The analytical parameters obtained for the analyzed
samples are summarized in Table 2.
Similar DPV curves were collected for Val, l-Phe, l-Arg, Gly, Ser,
Tyr and l-Trp. The values of limits of detection (LOD), limits of
quantization (LOQ) and linear range (␮M) obtained according to
differential pulse voltammetry (DPV) technique for these amino
acids is reported in Table 3.
(2.09 0.03) × 10+5 cm3 mol−1 s−1
.
Similar chronoamperograms were collected for Val, Phe, Cys,
Arg, Gly, Ser, Tyr and Trp. The values of ˛, D and k were obtained
according to the method described and are reported in Table 1.
The calibration curve for the Cys concentration in buffer solu-
tion (pH 2) was obtained by differential pulse voltammetry (DPV).
Fig. 6A shows typical DPV curves for different concentrations of Cys
in an alkaline solution (Fig. 8).
The dependency between peak current and Cys concentra-
tion was rectilinear for peak a1 within the range of 0.01–15 mM
(with a regression equation of y = 0.1608x + 0.543, R2 = 0.9987, n = 6
(Fig. 6B). The limits of detection (LOD) and quantization (LOQ) of the
procedure were calculated according to the 3 SD/m criteria where
SD was the standard deviation of the blank and m was the slope
of the calibration curves [43]. The limits of detection and quanti-
tation were found to be 3.21 × 10−6 M and 7.04 × 10−6 M for peak
Cys. Precision and accuracy were assessed by performing replicate
analyses of Cys samples. The precision of the method was calcu-
lated as the relative standard deviation (RSD). The procedure was
repeated on the same day using the same solutions at concentra-
Table 2
Results obtained for Cys analysis in the buffer solution (pH 2).
Linear range (␮M)
1–55,000
0.1608
0.543
2.97
Slope (␮A M−1
Intercept (M)
RSD (%)a
)
LOD (␮M)
3.21
LOQ (␮M)
7.04
a
Each value was obtained from ten experiments.

L. Saghatforoush et al. / Electrochimica Acta 56 (2011) 1051–1061
1061
12
10
8
determined by an amperometric method at the surface of the mod-
ified electrode. The prepared modified electrode can be used as
a chronoamperometric detector for amino acids determination in
flow systems or in chromatographic instruments.
A
Acknowledgement
6
We gratefully acknowledge the support of this work by Payame
Noor University.
4
2
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Gly
Tyr
Ser
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25–1000
3–2500
3.21
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The Fe (III)–Schiff base complex adsorbed strongly and irre-
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with MWCNTs. Due to the chemical stability, electrochemical
reversibility and high electron transfer rate constant of the Fe
(III)–Schiff base complex, it can be used in electrocatalysis as an
electron transfer mediator to shuttle electrons between the ana-
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