{Analysis of sulfamethoxazole by square wave voltammetry using new carbon paste electrode}
doi:10.5599/jese.583 281
J. Electrochem. Sci. Eng. 8(4) (2018) 281-289; DOI: http://dx.doi.org/10.5599/jese.583
Open Access : : ISSN 1847-9286
www.jESE-online.org
Original scientific paper
Analysis of sulfamethoxazole by square wave voltammetry
using new carbon paste electrode
Izabel C. Eleotério1, Marco A. Balbino1, José F. de Andrade1, Bruno Ferreira1,
Adelir A. Saczk2, Leonardo L. Okumura3, Antonio Carlos F. Batista4,
Marcelo F. de Oliveira1,
1Departamento de Química, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, USP,
14040-901, Ribeirão Preto, SP, Brazil
2Departamento de Química, Universidade Federal de Lavras, 37200–000, Lavras, MG, Brasil
3Departamento de Química, Universidade Federal de Viçosa, 36570–000, Viçosa, MG, Brasil
4Universidade Federal de Uberlândia - Campus do Pontal, 38304-402, Ituiutaba - MG, Brazil
Corresponding author - E-mail: marcelex@ffclrp.usp.br; Tel. +55-16-3315-9150
Received: February 17, 2018; Revised: April 20, 2018; Accepted: April 20, 2018
Abstract
In this work a new model of carbon paste electrode was employed to determine
sulfamethoxazole (SMX), an antibiotic used to treat infections in human and veterinary
medicine, by the square wave voltammetric modality (SWV). More specifically, the elec-
trochemical behavior of SMX was investigated by cyclic voltammetry (CV), and the
quantitative analysis of SMX was provided by SWV. The analytical curve was obtained
with a linear correlation coefficient (r) of 0.985 and standard deviation (SD) of 0.005 μA.
Limits of detection and quantification were found as 2.3×10-6 and 7.7×10-6 mol L-1,
respectively. According to the obtained results, the new carbon paste prototype electrode
can successfully be employed in this kind of electroanalytical applications.
Keywords
Carbon paste electrode; Electroanalysis of pharmaceutical compounds; electrochemical sensors;
voltammetric analysis
Introduction
Concerning a drug analysis, electroanalytical methods offer several advantages: they are
versatile, fast, sensitive, inexpensive, and environmentally friendly due to the limited use of chemi-
cals [1]. Recently developed electrochemical devices efficiently monitor pollutants through direct or
indirect reactions between the contaminant and the electrode surface, what makes them poten-
tially applicable in situ [2-4]. The large-scale use of this technology relies on the scientific knowledge
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J. Electrochem. Sci. Eng. 8(4) (2018) 281-289 ANALYSIS OF SULFAMETHOXAZOLE
282
and nowadays, scientists have investigated many electrodes for this purpose [5]. Gold electrode,
glassy carbon electrode, platinum disc electrode, graphite electrode, chemically modified
electrodes, carbon nanotube and carbon paste electrodes (CPEs) are some of the examples already
reported in the literature [1,5].
Ralph Norman Adams introduced CPE in 1958 [6-9]. This electrode consists of a mixture of carbon
powder and a non-electroactive liquid binder and offers a broad potential window, low residual
current (background), unique surface characteristics, low cost, and versatile preparation [6-16]. For
this reason, CPE has been widely employed to determine the sulfamethoxazole (SMX) by the
voltammetric analysis. SMX is chemically known as 4-amino-N-(5-methylisoxazol-3-yl)-benzene
sulfonamide and its chemical structure is shown in Figure 1. SMX constitutes a sulfonamide that helps
to treat infections in human and veterinarian medicine [17,18]. Obviously large consumption of this
antibiotic agent, however, can lead to environmental and public health problems.
Figure 1. Chemical structure of sulfamethoxazole.
Several methods for sulfamethoxazole determination have already been described in the
literature, including chromatographic methods coupled with different detectors 18,22,24, capillary
electrophoresis [25,26], spectrophotometry [27,28], and electroanalytical methods [29-39]. It is
possible to observe in the literature, however, that non-renewable surface devices for CPE application
were applied. Conventional CPEs must usually be refilled with the carbon paste, increasing thus the
consumption of material and time analysis. In this context, we developed a new model of CPE and
applied it for the sulfamethoxazole analysis using the square-wave voltammetry.
Experimental
Materials and reagents
A stock solution of 0.005 mol L-1 SMX (Sigma Aldrich) was prepared in the methanol (Merck). The
following electrolyte solutions were used in this research: 0.1 mol L-1 potassium chloride (Synth),
0.1 mol L-1 sodium nitrate containing 0.01 mol L-1 potassium ferricyanide, 0.1 mol L-1 sodium
perchlorate; 0.05 mol L-1 sulfuric acid (Merck)/methanol in 70:30 % (v/v) ratio and 0.04 mol L-1 Britton–
Robinson (BR) buffer, pH 2.18, prepared by mixing 0.04 mol L-1 boric acid (Carlo Erba), acetic acid and
orthophosforic acid (Merck). Mixtures of graphite powder (particle size = 19.2-168.5 µm, Analítica)
and mineral oil (Nujol, União Química) were used to prepare the carbon paste [14,16].
Carbon paste electrode construction
The carbon paste electrode was prepared by mixing 2.25 g of graphite powder with 0.75 g of
mineral oil. This mixture was homogenized by magnetic stirring in a 25 mL beaker containing
10 mL of chloroform (Merck). The paste was obtained after evaporation of the solvent. The carbon
paste was packed into a versatile electrode body fabricated in our laboratory. It consisted of a glass
cylindrical tube in the form of a syringe (o.d. 7 mm, i.d. 3 mm) and contained a platinum rod to
establish the electric contact (Figure 2).
NH2
S
O O
NH
N O
CH3
I. C. Eleotério et al. J. Electrochem. Sci. Eng. 8(4) (2018) 281-289
doi:10.5599/jese.583 283
1 - Electric contact (platinum) 2 - poston (Teflon)
3 - cylindrical glass tube 4 - carbon paste
5 - electric contact (stainless steel)
Figure 2. Carbon paste electrode developed in our research group.
Instrumentation
The voltammetric experiments were conducted using a potentiostat model μAUTOLAB III (Eco
Chemie) connected to a personal computer. The experiments were carried out in a three-electrode
system consisting of a carbon paste working electrode, platinum spiral wire auxiliary electrode and
Ag/AgCl, 3.0 mol L-1 KCl, reference electrode, all arranged in a 5-mL electrochemical cell (Figure 3).
The solutions were deoxygenated with nitrogen for 15 min prior measurements.
Figure 3. Electrode arrangement in electrochemical cell.
Voltammetric analysis
The voltammetric behavior of the carbon paste electrode was initially investigated by CV
measurements performed in different solutions, such as 0.1 mol L-1 KCl, 0.1 mol L-1 NaNO3 containing
0.01 mol L-1 K3Fe(CN)6, 0.04 mol L-1 BRbuffer (pH 2.18) and 0.1 mol L-1 NaClO4. CV analysis was carried
out at potentials ranging from 0.1 V up to 1.8 V and scan rates from 10 to 100 mV s-1. Subsequently,
the quantitative analysis of SMX was conducted by SWV with previously optimized experimental
parameters (frequency and pulse amplitude) and a constant step potential of 50 mV. Potential
window ranged from 0.1 to 1.8 V vs. Ag/AgCl and 0.04 mol L-1 BRbuffer (pH 2.18) was used as the
supporting electrolyte. The voltammetric parameters investigated for the SMX assay were:
frequencies of 8, 12, 18, 20, and 24 Hz and pulse amplitude ranging from 10 to 100 mV. SMX was
J. Electrochem. Sci. Eng. 8(4) (2018) 281-289 ANALYSIS OF SULFAMETHOXAZOLE
284
analyzed at different concentrations by the standard addition method, i.e. the analytical curve was
constructed by adding aliquots of the SMX stock solution to the electrochemical cell. A linear curve
was achieved for SMX concentrations ranging from 7.9 to 24.0 µmol L-1.
Results and discussion
Electrochemical properties of CPE in different electrolyte solutions
The CPE was initially tested in various aqueous systems, in order to provide information about its
basic response and stability in different electrolytes. Figure 4(A-D) illustrates the cyclic
voltammograms of CPE measured at different scan rates in 0.1 mol L-1 KCl, 0.1 mol L-1 NaNO3
containing 0.01 mol L-1 K3Fe(CN)6, 0.01 mol L-1 NaClO4, and 0.04 mol L-1 BR–buffer (pH 2.18). Among
almost all presented CVs suggesting stability of CPE with not specific response in the specific
electrolyte and given potential region, Figure 4B presents a typical cyclic voltammogram for the
K3Fe(CN)64-/3- redox couple. This particular reaction usually served as the test for the redox activity
of an electrode. Peaks at 0.36 V (Epa) and 0.19 V (Epc) suggest reversible behavior of redox couple at
the CPE in the potential range from 0.1 V to 0.7 V vs. Ag/AgCl, with scan rate between 10 to 100
mV s-1 [12].
E / V vs. Ag/AgCl E / V vs. Ag/AgCl
E / V vs. Ag/AgCl E / V vs. Ag/AgCl
Figure 4. Cyclic voltammograms of the CPE in (A) 0.1 mol L-1 KCl (100 mV s-1), (B) 0.1 mol L-1 NaNO3 +
0.01 mol L-1 K3Fe(CN)6, (C) 0.04 mol L-1 BRbuffer (pH 2.18) (100 mV s-1) and (D) 0.1 mol L-1 NaClO4,
at denoted scan rates and potential range from 0.1 to 1.8 V vs. Ag/AgCl.
i
/
A
i
/
A
i
/
A
i
/
A
I. C. Eleotério et al. J. Electrochem. Sci. Eng. 8(4) (2018) 281-289
doi:10.5599/jese.583 285
Literature reveals that the supporting electrolyte plays an essential role in the voltammetric
signal of SMX [29,40-42]. Besides, the sulfa drugs have two dissociation constants (pKa), which in
the case of the SMX correspond to the amino functional group with pKa value of 1.8 and amide
functional group with a pKa value of 5.6 [29,33,37,40,44]. Therefore the BRbuffer solution
(0.04 mol L-1 in acetic, phosphoric and boric acids) (pH 2.18), with CV shown in Figure 4C was chosen
as the experimental medium in the voltammetric studies of SMX.
Figure 5(A-B) displays representative CVs of 0.005 mol L-1 SMX together with the corresponding
background currents recorded for the proposed CPE and a commercial glassy carbon electrode,
respectively. Figure 5A shows CV profiles of CPE in a blank solution of 0.04 mol L-1 BRbuffer
(pH 2.18) and in the same solution containing 0.005 mol L-1 SMX, whereas Figure 5B shows CV
profiles of glassy carbon in a blank solution of 0.05 mol L-1 s acid/methanol 70:3 (v/v) (pH 1.38) and
in the same solution containing 0.005 mol L-1 SMX. An irreversible two-electron oxidation
voltammetric peak appeared in both cases when 0.005 mol L-1 SMX was present in the solution
[29,40]. Also, much lower background current that was obtained for the CPE as compared with the
solid glassy carbon, suggests that the proposed CPE electrode could be more sensitive for SMX
oxidation [6-16]. The higher background current observed for the glassy carbon electrode in Figure
5B stems from the oxygen evolution [43].
E / V vs. Ag/AgCl E / V vs. Ag/AgCl
Figure 5. Cyclic voltammograms at 100 mV s-1 of 5 mmol L-1 SMX for (A) CPE in 0.04 mol L-1 BRbuffer
(pH 2.18), potential range = 0.1 V to 1.8 V vs. Ag/AgCl and (B) glassy carbon electrode in 50 mmol L-1
sulfuric acid/methanol 70:3 (v/v) (pH 1.38), potential range = 0.5 V to 1.5 V vs. Ag/AgC).
CPE electrochemical response toward SMX
The optimized experimental parameters that pointed out the best results for SMX determination
using SWV technique were obtained by variations of pulse amplitude and frequency. The pulse
amplitude was varied in the range of 10–100 mV, at the constant frequency of 12 Hz. In this case,
the optimized result was defined as the parameter value that produced increase of the peak current
without shifting the peak potential or making any significant increase in the peak width. Hence,
100 mV was chosen as the square-wave pulse amplitude value. Afterwards, the effect of frequency
was evaluated in the range 8–24 Hz, keeping constant the pulse amplitude of 100 mV. The best
result was achieved at f = 12 Hz.
According to previous literature investigations, sulfonamide oxidation results in a formation of
the corresponding iminobenzoquinone intermediate, shown as peak (1) in Figure 6. The SWV
i
/
A
i
/
A
J. Electrochem. Sci. Eng. 8(4) (2018) 281-289 ANALYSIS OF SULFAMETHOXAZOLE
286
responses presented in Figure 6 showed that the oxidation current peaks increased with increase of
the frequency. Hence the frequency of 12 Hz was chosen for further analysis because of the best
resolution of the voltammetric peak (Figure 6, peak (2)).
E / V vs. Ag/AgCl
Figure 6. Effect of the frequency parameter on the SWV response of the CPE in 0.04 mol L-1 BR–buffer
(pH 2.18) containing 24.0 µmol L-1 SMX: (a) 12 Hz, (b) 18 Hz, (c) 20 Hz, (d) 24 Hz.
Potential range = 0.5 to 1.6 V vs. Ag/ACl, pulse amplitude = 100 mV, scan increment = 2 mV.
According to the literature, the SMX electrochemical oxidation occurs at the primary amino
groups (-NH2) [29,40]. Figure 7 illustrates the mechanism of SMX oxidation that as a two-electron
and pH dependent reaction, possibly takes place in an acid medium.
Figure 7. SMX oxidation at carbon paste electrode in acid medium
The electrochemical behavior of SMX in different concentrations was assessed by successive
additions of this drug in concentrations ranging from 7.9 to 24.0 µmol L-1 to the electrochemical cell.
As seen in Figure 8(A), the anodic peak current at 1.07 V vs. Ag/AgCl (irreversible oxidation peak)
increased upon rising of the SMX concentration. The analytical curve drawn in Figure 8(B) shows a
linear correlation coefficient r = 0.985 with a standard deviation SD = 0.005 μA. The corresponding
linear equation was adjusted as ipa = 0.24 μA + 6.5×103 μA /mol L-1 [SMX]. The limit of detection
calculated according to the criterion 3SD/m ratio, where m is the slope of the analytical curve, gave
2.3×10-6 mol L-1, while the limit of quantification based on the criterion of 10SD/m ratio, was
adjusted as 7.7×10-6 mol L-1.
N
+
H
H
H S
O
O
NH
O
N
CH3
-2e
-
, -2H
-
ONH + H2SO3 +
N
O
CH3
N
+
H
H
H
i
/
A
I. C. Eleotério et al. J. Electrochem. Sci. Eng. 8(4) (2018) 281-289
doi:10.5599/jese.583 287
Figure 8. (A) Influence of SMX concentration in 0.04 mol L-1 Britton–Robinson buffer (pH 2.18) solution on
the voltammetric response of CPE: (a) 24.0 µmol L-1, (b) 21.9 µmol L-1, (c) 20.1 µmol L-1, (d) 18.0 µmol L-1,
(e) 15.9 µmol L-1, (f) 14.0 µmol L-1, (g) 12.1 µmol L-1, (h) 10.0 µmol L-1, and (i) 7.9 µmol L-1.
Potential range = 0.5 to 1.6 V vs. (Ag/ACl), frequency = 12 Hz, pulse amplitude = 100 mV,
scan increment = 2 mV. (B) Analytical curve of the peak current, µA vs. SMX concentration, µmol L-).
Conclusions
The novel and efficient support for the carbon paste substrate is developed allowing
determination of sulfamethoxazole at the mol L-1 level. The developed CPE showed an excellent
stability in different electrolyte media and excellent voltammetric response for the K3Fe(CN)64-/3-
redox couple probe. Oxidation of SMX at the CPE occurring at about 1.07 V vs. Ag/AgCl was found
to be an irreversible 2-electron and pH dependent process. An electrocatalytic effect was observed
in comparison with glassy carbon electrode. Another peak occurring at about 0.49 V vs. Ag/AgCl was
observed when frequency values higher than 12 Hz were applied and ascribed to the formation of
the corresponding iminobenzoquinone intermediate. The developed CPE is an inexpensive and
versatile electrode, having high potential for application as a transducer in a device serving for
determination of sulfamethoxazole.
Acknowledgments: The authors acknowledge the financial support of FAPESP (Processes
2011/10216-5 and 2016/23825-3) and CAPES (Edital Pro-Forenses 25/2014). The authors also
acknowledge Dr. Cynthia M. C. P. Manso for editing and revising the text.
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