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Full Paper

Electrochemistry and Adsorptive Stripping Voltammetric

Determination of Amoxicillin on a Multiwalled Carbon Nanotubes
Modified Glassy Carbon Electrode
Behzad Rezaei,* Sajjad Damiri
Department of Chemistry, Isfahan University of Technology, Isfahan, 84156-83111 I.R. Iran
*e-mail: rezaei@cc.iut.ac.ir

Received: November 22, 2008

Accepted: March 15, 2009

Abstract
The study of electrochemical behavior of amoxicillin (AMX), a b-lactam antibiotic, is described on a multiwalled
carbon nanotubes (MWCNTs) modified electrode by electrochemical impedance spectroscopy (EIS) and adsorptive
stripping voltammetry for sensitive determination of AMX in pharmaceutical and human urine samples within a wide
pH range from 2.0 to 10.0. Also, studies by Fe2O3 nanoparticles modified carbon paste electrode show that iron oxide
impurities in the MWCNTs are not active sites for sensing of amoxicillin. Under optimized conditions, the oxidation
peak has two linear dynamic ranges of 0.6 – 8.0 and 10.0 – 80.0 mM with a detection limit of 0.2 mM and a precision of
< 4%.

Keywords: Multiwalled carbon nanotubes, Amoxicillin, Adsorptive stripping voltammetry, Fe2O3 nanoparticles,
Antibiotics, Nanotubes

DOI: 10.1002/elan.200804571

1. Introduction [11 – 23], and electrochemical methods [1 – 2, 24 – 28].

Antibiotics of the b-lactam group are important antimicro-
bial agents that are widely used to treat infectious human
and animal diseases and to enhance growth and yield in
agriculture. Amoxicillin (AMX), (D-a-amino-p-hydroxy-
benzylpenicillin trihydrate, Scheme 1), is the only phenolic
penicillin and a moderate-spectrum b-lactam antibiotic used
in humans and food-producing animals to treat several
diseases. b-lactam antibiotics presents a structure based on a
b-lactam ring responsible for the antibacterial activity and
variable side chains that account for the major differences in
their chemical and pharmacological properties. AMX
actives against a wide range of Gram-positive, and a limited
range of Gram-negative organisms. It is usually the drug of
choice within the class because it is better absorbed,
following oral administration, than other b-lactam anti-
biotics. Amoxicillin is susceptible to degradation by b-
lactamase-producing bacteria, and so may be given with
clavulanic acid to decrease its susceptibility. In human urine,
amoxicillin is excreted in a ratio of 86  8%, its urine
concentration being 1.7  7 g L1 for the first 2 h after a
single oral dose of 500 mg [1 – 4].
The therapeutic importance of AMX required the devel-
opment of sensitive and rapid method for industrial quality
control and clinical monitoring. Various analytical methods
have been reported for the separation and determination of
amoxicillin based on spectroscopy [3 – 8], capillary electro-
phoresis [9, 10], high-performance liquid chromatography
HPLC has been the most widely used technique for
determination of this group of drugs, but presents some
disadvantages such as necessity to the large amount of high
purity organic solvents, long equilibration and derivatiza-
tion treatment. Moreover, the low solubility of amoxicillin
in the organic solvents becomes difficult to develop
procedures based on extraction and pre-concentration
steps. Although, the electrochemical behavior of penicillins
has been extensively studied, papers about electrochemical
oxidation of amoxicillin describe very ill defined waves and
high positive potential.
Carbon nanotubes (CNTs) continue to receive remark-
able attention in electrochemistry [29 – 35]. Since their
discovery by Iijima [36] in 1991 using transmission electron
microscopy, CNTs have been the subject of numerous
investigations in chemical, physical and materials areas due
to their novel structural, mechanical, electronic, and chem-
Scheme 1. Chemical structure of amoxicillin.

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1578
ical properties [37, 38]. Depending on their atomic structure,
CNTs behave electrically as a metal or as a semiconductor
[39, 40]. The subtle electronic properties suggest that CNTs
have the ability to promote charge-transfer reactions when
used as an electrode [41 – 44]. The modification of electrode
substrates with multiwalled carbon nanotubes (MWCNTs)
for use in analytical sensing has been documented to result
in low detection limits, high sensitivities, reduction of
overpotentials and resistance to surface fouling. MWCNTs
have been introduced as electrocatalysts [30 – 32, 45 – 46]
and CNTs modified electrodes have been reported to give
super performance in the study of a number of biological
species, including cytochrome c [47], uric acid [48], NADH
[49] and quercetin [50]. Recently, however, these reported
electrocatalytic properties of MWCNTs have been dispelled
[51, 52]. Compton and Banks group has demonstrated for
most electroactive species, via the comparison of MWCNT
modified electrodes with edge plane pyrolytic graphite
electrodes, that usually the electroactive sites of the
MWCNTs are edge plane-like sites/defects which can occur
at the ends of the nanotubes or along the nanotube where
tube compartments terminate [52, 53]. Accordingly
MWCNT modified electrodes are no more electrocatalytic
than edge plane pyrolytic graphite electrodes except in a few
cases where the observed response of the MWCNT
modified electrodes could not be mirrored with edge plane
pyrolytic graphite electrodes [54]. Specifically for the
electrochemical oxidation of hydrazine and the electro-
chemical reduction of hydrogen peroxide, they have shown
that the electrocatalysis is due to iron oxide impurities
rather than edge-plane-like sites/defects [54, 55] and the
reduction of halothane is due to the presence of copper
oxide nanoparticles occluded within the carbon nanotubes
during their production [56]. These metal impurities have
been found to never be completely removed using the
common technique used to clean CNTs by stirring in strong
acid, even for excessively long exposure times and are
trapped in-between graphite sheets. In summary, the
electroactivity of MWCNTs is a result of either edge-
plane-like sites/defects or metal impurities [45, 52, 54 – 56].
On the other hand, above group has reported a new high-
purity metal-catalyst-free class of MWCNTs which are
fabricated via a solid phase growth mechanism. This new
MWCNTs class will allow experimentalists to avoid any
potential experimental misinterpretation, not only in elec-
trochemistry and electrocatalysis, but also in related appli-
cations [57].
Electrochemical properties of iron oxides have been
widely studied [58 – 60] and they are of significant impor-
tance in both technical and natural processes. a-Fe2O3
(hematite), which is the most stable iron oxide, with n-type
semiconducting properties under ambient conditions, has
been widely investigated in many technological fields for
applications such as anode materials for lithium-ion cells
[61 – 64], gas sensors [65 – 69], catalysts [70 – 71], solar
energy conversion [72], electrochromism [73], and magnetic
materials [74, 75], owing to its low cost and high resistance to
corrosion. It is well known that the particle sizes and shapes
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B. Rezaei, S. Damiri
of nanoscale hematite affect their properties and potential
applications [75, 76]. In the electrochemical sensors, up to
now, it has been reported that mesoporous Fe2O3 thin film
electrodes catalyze glucose oxidation [77], tubelike a-Fe2O3
shows unique sensing properties for nitric oxide [78], and
mitochondrial cytochrome c (Mcc) sorbs strongly to hem-
atite electrode from aqueous solution in a narrow pH range
due to opposite charge on Mcc and hematite [79].
The objective of the current work is to develop a sensitive
and useful method for the determination of amoxicillin,
based on the unusual properties of carbon nanotubes such as
strong adsorptive ability, huge specific area, subtle elec-
tronic properties as well as excellent electrocatalytic activ-
ity. The electrochemical behavior of AMX on the multiwall
carbon nanotube-coated glassy carbon electrode (GCE)
was investigated by voltammetry and electrochemical
impedance spectroscopy techniques to better be determined
the interaction of AMX and the electrodes. The results
showed that MWCNTs strongly enhanced the electron
transfer rate of amoxicillin oxidation and the determination
sensitivity of amoxicillin was significantly improved. At the
MWCNTs coated GCE, the remarkable peak current
enhancement and negative shift of peak potential occurred
for AMX oxidation, compared with that of a bare GCE.
Consequently, an ultrasensitive adsorptive stripping vol-
tammetric method based on the carbon nanotube-modified
electrode was first developed for the determination of
amoxicillin in pharmaceutical and urine samples. This newly
proposed method possesses following advantages such as
high sensitivity, rapid response, low cost and simplicity. Also,
in the other section of this work, the activity of iron oxide
nanoparticles which are left in the nanotubes from produc-
tion process was studied on the a-Fe2O3 modified carbon
paste electrode to be recognized its capability in electro-
catalytic oxidation of AMX.
2. Experimental
2.1. Apparatus and Reagents
Electrochemical measurements were carried out in a con-
ventional three-electrode cell, powered by an electrochem-
ical system comprising the Autolab system with PGSTAT 12
and FRA2 boards (Eco Chemie B.V., Utrecht, The Nether-
lands). The system was run on a PC using GPES and FRA
4.9 software. For impedance measurements, a frequency
range of 100 kHz to 10 mHz was employed. The AC voltage
amplitude used was 5 mV, and the equilibrium time was
20 minutes. The MWCNTs modified GCE, a graphite
electrode and a saturated Ag/AgCl reference electrode
was employed as a working, auxiliary and reference
electrode, respectively. The MWCNTs were bought from
Irans Research Institute of Petroleum Industry and synthe-
sized by chemical vapor deposition (CVD) with a diameter
of 8 – 15 nm, a length 50 mM and the purity of 95%. The
modified electrodes with carbon nanotube layers were
characterized by scanning electron microscopy (SEM) and
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Determination of Amoxicillin
energy dispersive X-ray (EDX) analysis. Alpha-Fe2O3
nanoparticles (Particle size: 40 – 60 nm) purchased from
Malek-Ashtar University of Technology. Spectrally pure
graphite powder (particle size < 50 mm) from Merck and
high viscosity paraffin oil (density ¼ 0.88 g cm3) from Fluka
were used for preparation of carbon paste electrode (CPE).
Amoxicillin was purchased from Fluka. Capsules of amox-
icillin were prepared from Cosar Company (Tehran, Iran).
Each amoxicillin capsule contains amoxicillin trihydrate
equivalent to amoxicillin anhydrous 500 mg. Other non-
medicinal ingredients of capsule matrix are cellulose micro-
crystalline, stearic acid and talc (magnesium silicate).
0.002 M solutions of amoxicillin were freshly prepared in
methanol-water (1 : 1, v : v) solution and were used as the
stock solution.
Universal buffer (boric acid, phosphoric acid, acetic acid
and sodium hydroxide, 0.1 M) solutions with different pH
values were used for the study of the pH influence. All
aqueous solutions were prepared with deionized water of
resistivity not less than 18.0 MW at 25 8C. Prior to the
adsorptive stripping voltammetry experiments, solutions
were stirred by a magnet for 5 minutes.
2.2. Preparation of MWCNT Suspension and Modified
GCE
To eliminate metal oxide catalysts within the nanotubes,
multiwalled carbon nanotubes were refluxed in the 2.0 M
HNO3 for 15 hours, and then washed with twice-distilled
water and dried at room temperature. The purified
MWCNTs were dispersed in acetonitrile (0.1 mg MWCNTs
per 10 cc) by using ultrasonic agitation to obtain a relative
stable suspension. The GCE was carefully polished with
0.05 mm alumina slurry on a polishing cloth, and then
washed ultrasonically in methanol and water, respectively.
The cleaned GCE was coated by casting 300 mL of the black
suspension of MWCNTs and dried in an oven at 60 8C. The
microscopic areas of the MWCNT-modified GCE and the
bare GCE were obtained by CV using 1 mM K3Fe(CN)6 as a
probe at different scan rates. For a reversible process, the
Randles – Sevcik formula has been used:
Ipa ¼ 2.69  (A s mol1 V1/2)n3/2AC0DR1/2v1/2 (1)
where Ipa (A) refers to the anodic peak current, n is the
electron transfer number, A (cm2) is the surface area of the
electrode, DR (cm2 s1) is diffusion coefficient, C0 (mol cm3)
is the concentration of K3Fe(CN)6, v (V s1) is the scan rate.
For 1 mM K3Fe(CN)6 in the 0.1 M KCl electrolyte: n ¼ 1 and
DR ¼ 7.6  106 cm2 s1, then from the slope of the ipa  v1/2
relation, the microscopic areas can be calculated. On the
bare GCE, the electrode surface area was found 0.0314 cm2
and for MWCNT-modified GCE the microscopic area was
nearly 2.9 – 3.0 times greater.
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2.3. Preparation of Iron Oxide Nanoparticles Modified
Carbon Paste Electrode
The graphite powder and iron oxide nanoparticles mixture
was homogenized by ultrasonication in a beaker containing
100 cc acetone. The modified carbon paste electrode was
prepared by mixing 0.1 g of iron oxide nanoparticles with
2.0 g of graphite powder and subsequently adding 0.250 g of
mineral oil. The final paste was obtained on evaporation of
the solvent. The modified carbon paste was packed into an
electrode body, consisting of a plastic cylindrical tube (o.d.
5 mm, i.d. 3 mm) equipped with a copper wire serving as an
external electric contact. Appropriate packing was achieved
by pressing the electrode surface against a filter paper.
2.4. Preparation of Real Samples
Content of ten amoxicillin capsules labeled with amount of
500 mg per capsule, were completely homogenized. 20 mg
of powders was accurately weighted and dissolved with
ultrasonication in 100 cm3 of methanol-water (2 : 5, v : v)
solution. The resultant solution 10 times was diluted and
2 cm3 of the solution in 10 cm3 of 0.1 M buffer was used for
determination. The urine samples used for measurements
were centrifuged and diluted 100 times with water without
any further pretreatment. The standard addition method
was used for the determination of amoxicillin in real
samples.
3. Results and Discussion
3.1. Characterization of MWCNT Modified Electrode
The purity and dispersing state of the MWCNTs and iron
oxide nanoparticles were examined by SEM and energy
dispersive X-ray (EDX) analysis. SEM micrographs showed
most of MWCNTs are well dispersed in acetonitrile and no
longer entwine together. EDX spectrum (not shown) similar
to other reports [54], indicated the presence of significant
amount of iron and sulfur after 15 hours purification in hot
nitric acid. The composition of the unwashed MWCNTs was
99.26%, C; 0.62%, Fe; 0.09%, S and 0.03%, Cl. By using
energy dispersive X-ray (EDX) analysis, the purified nano-
tubes showed near to three times lower content for iron and
sulfur. Banks and Comptons group has also shown the
electroactive metal impurities on/within the carbon nano-
tubes are not further removed by super washing and they
may cause the observed “electrocatalysis” associated with
carbon-nanotube-modified electrodes [54 – 56].
3.2. Electrochemical Behavior of Amoxicillin on GCE
and MWCNT Modified Electrode
The voltammetric responses of 20 mM AMX on the bare and
MWCNTs modified electrode and in pH 7.5 and 3.0 have
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1580
Fig. 1. A) Stripping voltammograms of AMX at the GCE electrode, and B) background-subtracted stripping voltammogram at the
MWCNTs modified GCE. Other conditions: 20 mM AMX; scan rate of 100 mV s1; pH 7.5; and accumulation time of 6 minutes. Inset
shows corresponding background-subtracted stripping voltammograms in pH 3.0.
been compared at Figure 1. As can be seen, the oxidation of
amoxicillin on the bare glassy carbon electrode and in acidic
conditions shows an ill defined wave attributed to oxidation
of the phenolic substituent to relevant carbonyl group. In the
basic buffer solution, AMX does not show any electro-
oxidation signal on the GCE. The mentioned results, have
been performed by adsorptive stripping measurements to be
enhanced the output signals. Under the similar conditions,
amoxicillin yields a well-defined oxidation peak on the
MWCNTs modified electrode at near to 1.0 and 0.6 V for
pH 3.0 and 7.5, respectively, and the oxidation peak current
significantly increases as compared with that at the bare
GCE. The increase in the peak current and the lowering of
oxidation potential are clear evidences of the catalytic effect
of the MWCNTs toward the oxidation of amoxicillin. The
generated product of AMX oxidation is retained on the
electrode surface, which it can be identified by its reduction
at þ 0.25 V at pH 3.0 in the reverse scan.
Investigation of amoxicillin oxidation on the iron oxide
nanoparticles modified carbon paste electrode, according to
Figure 2, shows this material does not have any electro-
catalytic effect on the oxidation of amoxicillin. Therefore
iron oxide nanoparticles which are left in the nanotubes
from production process can not be electroactive sites for
oxidation of AMX and edge plane-like sites/defects of
nanotubes, along with adsorption phenomenon, are respon-
sible for amplifying of the amoxicillin oxidation signal.
3.3. Electrochemical Impedance Spectroscopy Studies
Electrochemical impedance spectroscopy was also em-
ployed to investigate the oxidation of amoxicillin on both
GC and MWCNTs electrodes. Figure 3 shows the bode plots
and the Nyquist plots of the impedance surface densities (W/
cm2) and admittance surface densities (S/cm2), on the GC
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B. Rezaei, S. Damiri
(A) and MWCNTs (B) electrodes recorded at 0.6 V as DC-
offset for 0.1 mM AMX in pH 7.5. Because impedance and
admittance quantities depends on the microscopic areas of
the electrodes and the surface area of the bare GC and
modified electrodes were different, therefore the impe-
dance elements were normalized with respect to the
electrode surface area during data analysis after the experi-
ments. The Nyquist diagram of MWCNTs electrode repre-
sent two overlapped, slightly depressed capacitive semi-
circles. The high-frequencies semicircle is related to the
electron-transfer process and so the combination of charge-
transfer resistance and double-layer capacitance. The low-
frequencies semicircle can be related to an adsorption
process, which is the adsorption of the reactions intermedi-
ate(s) on the electrode surface. Also, bode and admittance
plots on Figure 3, clearly show the presence of one charge
transfer step at the high frequencies and one adsorption
phenomenon at the low frequencies [80].
However, the GC electrode represents a linear tail with a
slope of near-unity at low frequencies and the charge
transfer resistance of electrochemical oxidation process on
this electrode is very high. The linear tail was attributed to
the semi-infinite diffusion of the electroreactant species
[81]. The electron-transfer resistance, Rct, for the bare GC
and MWCNTs electrode, equals to 410.0 and 40.9 W,
respectively. Therefore, the charge transfer rate for oxida-
tion of amoxicillin on the MWCNTs electrode surface, due
to their electrocatalytic effects, is significantly higher than
that of the GC electrode. It is interesting that admittance
diagram of MWCNTs electrode, shows a large diameter
semicircle in high frequencies and a small admittance
semicircle, related to adsorption of electrochemical reaction
intermediate(s), at low frequencies. Moreover, electroox-
idation of amoxicillin on the GCE shows only a small
admittance semicircle, indicating the low charge transfer
rate on the electrode surface.
 2009 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Determination of Amoxicillin
Fig. 2. Electrochemical behavior of amoxicillin on the a) bare carbon paste electrode (CPE) without AMX; b) as (a) with 20 mM AMX;
c) a-Fe2O3 nanoparticles modified CPE without AMX; d) as (c) in the presence of 20 mM AMX. Other conditions scan rate of 100 mV
s1: pH 7.5 and accumulation time of 6 min. at OCP.
The equivalent circuits compatible with the Nyquist
diagrams recorded for the GC and MWCNTs electrodes
are depicted in Schemes 2A and B, respectively. The model
equivalent circuits contain the solution resistance (Rs), the
charge-transfer resistance of amoxicillin oxidation (Rct) and
a constant phase element corresponding to the double-layer
capacitance (CPEdl). In addition, CPEads and Cads signify a
constant phase element and a capacitance representing the
adsorption of the reaction intermediate(s) on the MWCNTs
and GC electrodes, respectively. W, which is included in the
circuit, is the Warburg element related to the semi-infinite
diffusion process on the GC electrode and T (hyperbolic
tangent element or bounded Warburg) shows the finite
Warburg diffusion impedance of amoxicillin in the thin film
of MWCNTs. Impedance of CPE, W and T elements can be
expressed as [82]:
Scheme 2. The equivalent circuits compatible with the Nyquist
diagrams represented in Fig. 3.
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1581
ZCPE ¼ (Y0jw)n (2)
1/2
ZW ¼ Y 1
0 (jw) (3)
1/2
ZT ¼ Y 1
0 (jw) coth[B (jw)1/2] (4)
where Y0 (the admittance parameter, S cm2 sn) and n
(dimensionless exponent) are two parameters independent
of frequency; j ¼ ( 1)1/2, w ¼ angular frequency ¼ 2pf, and
B is the diffusion factor. ZCPE corresponds to the constant
phase angle element (CPE) impedance. Y0 ¼ Cdl only when
n ¼ 1, and n is related to a (phase angle) by a ¼ (1  n) 90.
So, n ¼ 1 and a ¼ 0 stand for a perfect capacitor, and lower n
values directly reflect the roughness of the electrode surface.
When n ¼ 0.5, it is equal to a Warburg impedance. When n ¼
0, CPE is reduced to a resistor.
In the mentioned circuits, the charge-transfer resistance
of the electrode reaction is the only circuit element that has a
simple physical meaning describing how fast the rate of
charge transfer during electrooxidation of amoxicillin
changes with the electrode potential. The most widely
accepted explanation for the presence of constant phase
elements and the appearance of depressed semicircles in the
Nyquist plots is the microscopic roughness, causing an
inhomogeneous distribution in the solution resistance as
well as in the double-layer capacitance [83].
In this study, we used the above process on the MWCNTs
modified electrode for determination of amoxicillin in
pharmaceutical and urine samples and optimized the
various parameters that can affect on the amount of that
electrooxidation signal.
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1582 B. Rezaei, S. Damiri

Fig. 3. The Nyquist plots of the impedance surface densities (Z’’/cm2 vs. Z’/cm2) acquired for 0.1 mM AMX in pH 7.5 on the GC
electrode (A) and MWCNTs electrode (B). Bias in both diagrams was 0.6 V with 5 mV ac voltage amplitude and frequency range of
0.01 Hz to 100 kHz. Insets show their related a) bode plots and b) the admittance surface densities (Y’’/cm2 vs. Y’/cm2) plots. Points show
the experimental data and the full line is calculated from the optimized parameters.

3.4. Effect of Solution pH on the Peak Potentials and
Peak Currents
The peak potential and the peak current closely depend on
the pH of buffer solution. Experimental results of 20 mM
AMX in 0.1 M buffer solution at different pHs from 2.0 to
10.0 are shown in Figure 4. Direct oxidation of AMX
presents voltammetric signal on all pH range. When the pH
is increased it is observed that the peak potential shifts to
less positive values and shows some breaks in the slope at
pHs 2.5 and 8.0 – 9.0. AMX presents three pKa values at
2.41; 7.19 and 9.38. In acidic medium, amino group of
amoxicillin is protonated to give a cationic species. When
the pH is increased, in a first stage, the deprotonation of
carboxylic group (pKa1 ¼ 2.41) take places, yielding the
zwitter ion species, existent in the pH range 4 – 6. In the next
stages, the deprotonation of the amino group NH þ3 (pKa2 ¼
7.19) and phenolate group (pKa3 ¼ 9.38) are performed [84].
By treatment of MWCNTs in nitric acid, carboxylic group
can be created on the surface of nanotubes. Therefore in the
acidic solutions, protonated AMX can not properly inter-
acted with working electrode surface and its oxidation
current amount is relatively low. In the conditions that AMX
is deprotonated, there is better interaction with nanotubes

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Determination of Amoxicillin
Fig. 4. Effect of pH on the peak current and peak potential at
the modified electrode. Other conditions, scan rate of 100 mV s1:
20 mM AMX and accumulation time of 6 min at OCP.
film and electrooxidation current is higher. The peak
current reaches maximum at pH 7.5, which was selected as
the optimized pH value for determination of amoxicillin.
The calibration curve of E versus pH, in the pH range of
3.0 – 8.0, has a slope of  44.4 mV pH1. This slope
approximately suggests that the number of electron transfer
is equal with that of hydrogen ions taking part in the
electrode reaction.
3.5. Effect of Scan Rate on the Peak Currents and Peak
Potentials
The effects of scan rate on the peak current and peak
potential at the modified GCE in a 0.1 M universal buffer
were investigated by cyclic voltammetry in the presence of
20 mM AMX at pH 7.5. The anodic peak current linearly
increases as the scan rate increases, and the following
equation was obtained: I(mA) ¼ 1133.90  v (V s1) þ 23.43,
R2 ¼ 0.9961, indicating an adsorption controlled oxidation
process occurring at the modified GCE. The scan rate of
100 mV s1 was selected as the optimum for amoxicillin
determination. The linear relationship of peak potential (E)
and logarithmic scan rate (v) of an irreversible process obeys
the following equation [85];
  1=2   stripping cyclic voltammetry under the optimum conditions
RT D were selected as: pH 7.5 (universal buffer) with a scan rate of
EðVÞ ¼ E ’ðVÞ þ ðVÞ 0:780 þ ln =ðs1=2 Þ
ð1  aÞnF k0
  
ð1  aÞnFn 1=2 1=2
þ ln =ðs Þ ð5Þ
RT
Here, E8’ is the formal potential, a the transfer coefficient, n
the number of electrons involved in an electrode reaction, F
the Faraday constant, DR the diffusion coefficient, k0 the
standard heterogeneous rate constant, R the gas constant, T
the absolute temperature (300 K), and in the parentheses
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1583
are the units of the arguments. The relationship between the
peak potential and the scan rate (40 – 120 mV s1) was
studied. Based on the slope of the fitted line for the relevant
figure (not shown), with the equation: E (V) ¼ 0.0272  ln [v
(V s1)] þ 0.6384, R2 ¼ 0.9982, RT/2(1  a)nF ¼ 0.0272 V,
the value of n(1  a) was calculated to be 0.525. Also, the
slopes of tofel plots (E vs. log Ip) in low scan rates (10 –
50 mV s1) confirm this value of n(1  a). For n ¼ 1, a ¼
0.525, which indicates that the activation free energy curve
is approximately symmetrical in such an irreversible oxida-
tion process. On the other hand, according to the Equation 6
for totally irreversible process [85], a value in the optimized
conditions was found 0.58.
j Ep  Ep/2 j¼ 1.85 RT/aF ¼ (47.7/a) mV (6)
From the mentioned results, the overall reaction mechanism
can be attributed to the contribution of one electron and one
proton for oxidation of the phenolic group of amoxicillin to
generate the related quinone.
3.6. Effects of Accumulation Potential and Time
The oxidation peak currents of 20 mM AMX were measured
by cyclic voltammetry after 6 min of accumulation at
different potential from  0.5 to þ 0.3 V in pH 7.5. It was
observed the adsorptive stripping AMX response increases
rapidly upon increasing the accumulation potential. We also
observed the accumulation of AMX under open circuit
conditions, with a higher signal than that obtained at the
mentioned accumulation potentials. Further experiments
thus employed at open circuit potential.
In addition, the accumulation time affects significantly on
the adsorptive stripping voltammograms. By increasing the
time, the oxidation peak current amplifies greatly at the first,
and then reaches a steady amount after 6 minutes due to
adsorption saturation.
3.7. Performance of the System for Amoxicillin
Measurements
The calibration plot of amoxicillin determination is linear in
two concentration region of 0.6 – 8.0 and 10.0 – 80.0 mM with
detection limit of 0.2 mM (see Fig. 5). Using adsorptive
100 mV s1, and the stripping time of 6 min at open circuit
potential. Equations of linear least square calibration curves
over this ranges are:
I(mA) ¼ 14.67  CAMX  0.82 (R2 ¼ 0.9976) and I(mA) ¼
2.75  CAMX þ 127.96 (R2 ¼ 0.9964). Relative standard devi-
ation (RSD) of < 4% for 20 mM amoxicillin (for 8 analysis)
showed excellent reproducibility. No obvious changes in the
anodic peak current were found for the same sample
concentration when the modified electrode was kept under
ambient conditions with the room temperature of about
Electroanalysis 2009, 21, No. 14, 1577 – 1586
1584 B. Rezaei, S. Damiri

Fig. 5. Calibration curves for determination of amoxicillin at the optimum conditions. Inset show the some of the raw voltammograms.

30 8C for one month or greater, thus, the modified electrode determination of AMX. 700-fold of Naþ, Kþ, NO 3 , Ca2þ,
  
is stable. Mg2þ, SO 2 4þ
4 , F , urea, 300-fold of NH , Cl , Br , CO 3 ,
2

The detection limit, linear dynamic range and sensitivity
for amoxicillin determination at this method, according to
Table 1, are significantly better than other electrochemical
methods reported by Biryol [2], Cavalheiro [27], and Zanoni
[28]. Also the analysis can be performed on the wide pH
range 2.0 – 10.0. As compared to most of the published
chromatographic and electrophoresis methods, which re-
quire lengthy and tedious extraction procedures, the pro-
posed method does not require any filtration for analysis of
drugs from undissolved excipients and is rapid, very
sensitive and simple. Although the chromatographic proce-
dures has the advantage of simultaneous quantitation of the
drug and its related substances when the chromatographic
conditions are properly selected.
3.8. Interference Studies
Under optimized experimental conditions described above,
the effects of some foreign species on the determination of
AMX at 20 mM level were evaluated in detail. The tolerance
limit was defined as the maximum concentration of the
interfering substance that caused an error less than 3% for
glucose, sucrose, lactose and fructose have almost no
influence on the current response of AMX. All these
indicated the peak current of amoxicillin is not affected by
all conventional cations, anions, and organic substances, but
the iron or manganese cations can be interfered.
3.9. Application
To evaluate the applicability of proposed method, the
recovery of AMX was determined in the capsule and urine
samples by adding the standard value of AMX to them. The
standard addition method was used for the analysis of
prepared samples. The data given in Table 2 show the
satisfactory results for analytical determination of amox-
icillin in real samples.
4. Conclusions
The results discussed above demonstrate that the electro-
chemical response of amoxicillin by adsorption stripping
voltammetry on MWCNTs film can remarkably be en-

Table 1. Analytical parameters of several modified electrodes for AMX determination. LOD: limit of detection; LDR: linear dynamic
range; CPE: carbon paste electrode; SWV: square wave voltammetry.
Electrode Method LOD (mM ) LDR (mM ) Applicable pH range References
Modified CPE Cyclic voltammetry 0.812 10 – 200 < 2.3 [2]
Modified CPE SWV 8.49 18.9 – 91.9 5.5 [27]
Modified GCE SWV 0.92 2 – 25 < 7.0 [28]
MWCNTs modified GCE Cyclic voltammetry 0.2 0.6 – 8.0 and 10.0 – 80.0 2.0 – 10.0 This work

Electroanalysis 2009, 21, No. 14, 1577 – 1586 www.electroanalysis.wiley-vch.de  2009 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Determination of Amoxicillin
Table 2. Determination of AMX in drug and urine samples in pH 7.5
No. Sample Added (mM )
Capsules
1 40
2 80
Urine
3 10
4 40
[a] Average of five replicate measurements
hanced in high pH range of 2.0 – 10.0 and iron oxide
nanoparticles in the nanotubes which are left from produc-
tion process can not be active sites for amoxicillin oxidation.
Furthermore, an adsorption process occurred during the
redox process of AMX on the electrode surface; revealed by
impedance measurements. Electrochemical studies for
identifying of AMX oxidation mechanism confirm the
contribution of one electron and one proton in the process.
Analysis by this method as compared to other reported
electrochemical methods can be performed with a higher
sensitivity and determination range with low experimental
detection limit of 0.2 mM. This modified electrode can be
properly used for determination of amoxicillin in drug and
human urine samples with satisfactory results.
5. Acknowledgements
The authors wish to thank Isfahan University of Technology
(IUT) Research Council and Center of Excellence in sensor
(IUT) for support of this work.
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