Autores
Jara Cornejo, E. (UNIVERSIDAD NACIONAL DE INGENIERÍA) ; Khan, S. (UNESP) ; Wong, A. (UNESP) ; Taboada Sotomayor, M.D.P. (UNESP) ; Picasso Escobar, G. (UNIVERSIDAD NACIONAL DE INGENIERÍA)
Resumo
This work aims to quantify methotrexate using electrochemical sensors based on
polypyrrole-based molecularly imprinted polymers (MIP) electrodeposited by cyclic
voltammetry on a glassy carbon electrode (GCE) modified with multi-walled carbon
nanotubes (MWCNT). The electrodeposited polymeric films were characterized by
infrared spectrometry (FTIR), scanning electron microscopy (SEM), and cyclic
voltammetry (CV). Results evaluated by differential pulse voltammetry (DPV) showed
a detection limit of 2.7×10-9 molL-1 for methotrexate for a linear range of 0.01 –
25 molL-1 and amperometric sensitivity of 0.226 μALmol-1.
Palavras chaves
MIP sensor; Polypyrrole; Electropolymerization
Introdução
The drug methotrexate is an antimetabolite of the folate group normally used in
oncology hospitals, and after being used in some treatment, the residues are
usually disposed of in hospital effluents (Arnold et al. 2014). Due to its
antimetabolitic properties of the drug it could bring harmful damage to the
human being, could inhibit metabolic processes of the organism, and can be
considered as a possible emerging contaminant (Jureczko and Kalka 2020).
Molecular imprinted polymers consist of a polymerization process of a functional
monomer in the presence of a template molecule, it is very important that the
functional monomer and the analyte have a good affinity between them (Martín-
Esteban 2016) since polymerization is expected to grow around the analyte. In
such a way that after an analyte removal process, free cavities are generated
within the polymer and these cavities are specific for the analyte. However,
MIP-based sensors generally have low electrical conductivity (Li et al. 2017).
So by modifying the surface of the electrode with some conductive substrate, it
is possible to improve that electrical deficiency, such as multi-walled carbon
nanotube (MWCNT), since this material is capable of effectively accelerating
electron transfer and improving sensitivity (Cosio et al. 2017). In addition
polypyrrole (PPy) is a conductive polymer and in several works it has been
efficiently applied in the MIP sensor process (Beluomini 2018). The synthesis of
polypyrrole by electrochemical methods has been of interest, since
electropolymerization is a simple process and allows the quantification of
molecules of small and large molecular mass of the analyte. The use of PPy and
MWCNT generates a synergistic effect in electrochemical detection favoring the
quantification of analytes.
Material e métodos
Reagents
Methotrexate (98%), pyrrole (99%), lithium perchlorate (98%), MWCNT (<5%
modified with COOH), hexacyanoferrate (II) potassium trihydrate (98%) and
potassium ferricyanide (98%) were purchased from Sigma-Aldrich. Monobasic sodium
phosphate (98%), bibasic sodium phosphate (98%) and Potassium chloride (98%)
they were bought from Merck.
Modification of GCE with MWCNT
For the increase the analytical signal of the sensor, a suspension of 1mg/ml of
MWCNT dispersed in N, N dimethylformanide was prepared. This suspension was
first sonicated in an ultrasound bath for 20 min, then deposited on the
electrode surface and using an IR lamp fr the complete solvent was evaporation
untill dry.
Preparation of molecularly imprinted polymers (MIPs) and molecularly non-
imprinted polymers (NIPs) on MWCNT/GCE
The modified electrode with MWCNT are immersed in a solution containing pyrrole
20 mmol L-1, lithium perchlorate 0.1 mol L-1 and methotrexate 100 µmol L-1 to
obtain MIP/MWCNT/GCE. In parallel, for comparison reasons, NIP/MWCNT/GCE was
prepared using the same solution as mentioned above, but without methotrexate.
The electropolymerization parameters were: Conditioning potential of 0.4V for 30
s, potential range of -0.4 to 1 V, sweep speed 50 mVs-1 and 10
electropolymerization cycles. Once the electrodeposited polymer is obtained,
then in the next step the analyte removal stage is continued, in this stage the
electrode is immersed in a sodium phosphate buffer solution at pH 10 and cyclic
voltammetry from 0.3 to 1 V was applied to completely remove methotrexate from
the printed polymer cavities and the obtained film was nominated as OPPy/MWCNT.
Resultado e discussão
The printed FTIR spectrum of polypyrrole (PPy) is shown in Figure 1A and
presents characteristic peaks of polypyrrole according to the literature
(Shamsipur, Moradi, and Pashabadi 2018) confirming the presence of PPy. SEM
image shows that the morphology of the MWCNT was slightly modified with PPy
(Figure 1B).
The prepared MIP/MWCNT/GCE sensor, as a preliminary test of its efficacy, was
evaluated in the quantification of methotrexate. For this, a standard solution
of 100 umol L-1 was prepared and the differential pulse voltammetry technique
was applied. As shown in Figure 2A, the signal obtained by the MIP/MWCNT/GCE
electrode was 3 times higher than the corresponding to NIP/MWCNT/GCE, confirming
the specificity of the former towards the analyte and the calculations made by
computational simulation. Figure 2B shows a change in the sensitivity of the
sensor. This is due to the fact that in a solution with a low concentration of
the analyte (0.01 – 25 umol L-1) the saturation of the sensor cavity is not
reached, while at higher concentration values (25 – 125 umol L-1) a decrease in
detection is observed by the saturation of the analyte in the cavities.
The application of the sensor in real samples was proposed by testing river
water (Table 1) and pharmaceutical samples (Table 2). For the pharmaceutical
samples, 2.5 mg methotrexate tablets were used (Table 1). To check the response
of the sensor in the selectivity test, it was tested on samples of river water
adding standard methotrexate at 2.5 and 25 mol L-1. The results showed
excellent recovery values for river and pharmaceutical water of 102% and 105%,
respectively. The results obtained by the sensor prepared in this work are
comparable with those tested by the analytical technique of HPLC.
Conclusões
In this work, a MIP-sensor was prepared to take advantage of the methotrexate
imprinted cavities provided by the electropolymerization of pyrrole on an
electrode modified with multi-walled carbon nanotubes (MWCNT).The
electropolymerization factors were optimized. The MIP/MWCNT/GCE sensor presents a
good repeatability in the analysis of 5 successive measurements of methotrexate
with a relative standard deviation (RSD) value of 2.5%. The reproducibility of the
MIP/MWCNT/GCE sensor was evaluated in 5 independent measurements with an RSD
result equal to 1.72%.
Agradecimentos
The authors wish to express their gratitude to PROCIENCIA (project No. 023-
FONDECYT-BM-INC.INV-2019) and OGI of the National University of Engineering of
Lima-Peru (project nº FC-PF-12-2021) for the financial support.
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