Voltammetric Determination of Gabapentin by a Carbon Ceramic Electrode Modified with Multiwalled Carbon Nanotubes and Nickel-Catechol Complex

A técnica sol-gel foi usada na fabricação de um eletrodo cerâmico de carbono (CCE) renovável com complexo níquel-catecol (Ni-CA). O complexo foi depositado na superfície de nanotubos de carbono de parede múltipla-CCE por potencial cíclico na faixa de 0,0-0,8 V (vs. eletrodo saturado de calomelano, SCE). Ni-CA mostrou ter atividade catalítica sobre a reação redox de gabapentina. Observou-se uma relação linear da corrente de pico anódico com a concentração de gabapentina na faixa de 1,25-63,23 μmol L com um limite de detecção de 0,5 μmol L. A resposta do eletrodo com respeito à gabapentina era bem reprodutível e observou-se estabilidade a longo prazo do eletrodo (acima de 2 meses). O eletrodo foi aplicado com sucesso à determinação de gabapentina em preparações farmacêuticas.


Introduction
Gabapentin (GP), 1-(aminomethyl)cyclohexane acetic acid, is an antiepileptic drug which is also effective in the prevention of frequent migraine headaches, neuropathic pain, and nystagmus.It is a cyclic analogue of gammaaminobutyric acid (GABA) which may alter GABA transmission in the central nervous system. 1,2The lipophilic cyclohexane ring in its structure allows GP, unlike GABA, to cross the blood-brain barrier.GP increases the rate of synthesis and accumulation of GABA, decreases the release of monoamines (dopamine, norepinephrine, and serotonin), and weakly inhibits GABA-transaminase, an enzyme which degrades GABA into other amino acids. 3ue to the absence of chromophores and fluorophores in GP, derivatization is a part of the electrophoretic and chromatographic [20] assays for the drug analysis.For example, in electrophoretic methods, derivatization of GP with fluorescamine, 4 and 6-carboxyfluorescein succinimidyl ester 5 was used to provide chromophores for UV absorption and fluorescence detection.In chromatographic methods, reagents such as 9-fluorenylmethyl chloroformate, 6 4-fluoro-7-nitrobenzofurazan, 7 and 1-fluoro-2,4-dinitrobenzene 8 were used for the determination of GP with fluorescence detectors.
Electrochemical methods, on the other hand, have been proven to be very sensitive to the determination of drugs and related molecules.The advance in electrochemical techniques in the field of analysis of drugs is due to the simplicity, low cost and relatively short analysis time when compared to electrophoretic and chromatographic techniques.Moreover, the use of chemically modified electrodes in electrochemical methods is widely reported for sensitive and selective determination of various pharmaceuticals. 9,10ol-gel electrochemistry has rapidly developed over the last years and has become an area of active research. 11he ability to form inorganic-organic hybrids and tailoring thin films make it attractive for the field of chemically modified electrodes. 12,13Carbon-ceramic composite electrodes (CCEs) are comprised of a dispersion of carbon powder that is held together by a ceramic binder.The ceramic material is produced from the low temperature sol-gel process which involves the hydrolysis of an alkoxide precursor, followed by condensation of the hydroxylated monomer. 11,14he composite electrode benefits from the mechanical robustness of the silicate backbone, resistance to air oxidation and to hydrolysis under extreme pH conditions.Electron conductivity through the interconnected carbon grains and the ability to manipulate the physicochemical characteristics of the matrix by incorporation of suitable monomer precursors or sol-gel dopants are other advantages of CCEs.
Gavalas et al. reported the fabrication of carbon nanotube (CNT) composite electrodes based on the sol-gel technique. 15The composite carries with it the advantages of both CNTs 16,17 and CCEs.4][25] Several modified CCEs were recently used for the determination of pharmaceutical compounds. 18,21,26P is not electroactive at the surface of most unmodified electrodes; the only report is on gold electrode. 27Chemically modified electrodes such as nanotubes of nickel oxidemodified carbon paste electrode 28 and silver nanoparticle modified multiwalled carbon nanotubes (MWCNTs) paste electrode 29 were used for the voltammetric determination of GP, through an electrocatalytic mechanism.
In this work, Ni-catechol complex (Ni-CA) was deposited on the surface of a MWCNT-CCE electrode and used in the electrocatalytic determination of GP.Carbon paste electrode that has been used in the determination of GP 28 is nonporous, and therefore only its outermost surface is wetted by the electrolyte and accessible to dissolved substrates.In contrast, CCE has a porous structure, which is not clogged by the mechanical polishing step, due to the brittleness of the inorganic matrix, and is highly resistant to plastic deformations.Thus, highly reproducible responses were observed by using CCEs instead of carbon paste electrodes. 30oreover, the resistance of CCE to extreme pH conditions (in this work, pH 13), and the advantages of MWCNTs, such as improvement in the rate of electron transfer and enlargement of surface area were considered in the present work.

Reagents and solutions
Methyltriethoxysilane (MTEOS) and Ni(NO 3 ) 2 .6H 2 O were from Merck.MWCNTs (20-50 nm o.d., purity > 95%) and graphite powder were purchased from Aldrich.GP powder was from Bakhtar Bioshimi Pharmaceutical Company (Kermanshah, Iran).All other chemicals were of analytical grade and used without further purification.All solutions were freshly prepared using doubly distilled water.

Apparatus
Electrochemical measurements were performed with μ-Autolab type III potentiostat/galvanostat instrument.Data acquisition was performed using software NOVA 1.8.A three-electrode system consisting of a saturated calomel electrode (SCE) as the reference, a platinum wire as the counter electrode, and a modified MWCNT-CCE as working electrode was employed for the electrochemical measurements.All experiments were conducted at room temperature.A pHmeter (Jenway, Model 140) with a combined glass electrode was used to check the pH levels of the solutions.

Preparation of Ni-CA/MWCNT-CCE
MWCNT-CCE was prepared according to the procedure described by Lev and co-workers. 14Briefly, a mixture of MTEOS (0.2 mL), methanol (0.6 mL) and hydrochloric acid (20 μL, 11 mol L -1 ) was magnetically stirred to ensure uniform mixing, after which graphite powder (1.0 g) and MWCNTs (0.01 g) were added and the mixture was shaken for 5 min.The mixture was packed into a Teflon tube (2 mm i.d.), and dried at room temperature for 48 h.The prepared MWCNT-CCE was polished with polishing paper to remove any extra composite material.

Determination of GP in tablets
A homogenized powder was prepared from ten accurately weighed tablets (100 mg tablet -1 ).A proper portion of the powder equivalent to 0.021 g GP was added to HCl solution (2 mL, 0.01 mol L -1 ).Dissolution of the drug was assisted by means of ultrasonication.After filtration on a filter paper, the solution was made up to the mark with distilled water in a 50 mL volumetric flask.A 10 mL portion of this solution was transferred into a 25 mL volumetric flask and diluted with distilled water.The GP content was determined by linear sweep voltammetry (LSV) using the modified electrode.

Results and Discussion
Electrochemical behavior of Ni-CA/MWCNT-CCE Cyclic voltammograms of separate solutions of Ni, CA, and Ni-CA in NaOH (0.1 mol L -1 ) on MWCNT-CCE are compared in Figure 1.In the case of Ni(NO 3 ) 2 solution (curve a), well-defined anodic and cathodic peak currents of Ni(II)/Ni(III) were observed on a large background current.The corresponding half-reaction is: The formal potential, Eº', was calculated (= (Epa + Epc)/2) as about 0.65 V.The peak separation (ΔEp = Epa -Epc) of 0.16 V shows the quasi-reversible nature of the electron transfer.
CA showed no distinguishable peak (curve b) because, at highly alkaline media, CA undergoes hydrolysis and converts to quinone in solution.In a mixture of Ni(II) and CA (curve c), however, a well-defined pair of anodic and cathodic peaks were observed at lower potentials (Eº' = 0.41 V) with very low background current.Indeed, Ni-CA complex is formed between the oxidized form of CA (o-quinone) and nickel ions (Figure 2), which is more readily adsorbed at the electrode surface, 34 and a large negative shift in oxidation potential results.The presence of MWCNTs in CCE provided a larger surface area for Ni-CA to be deposited, and accelerated electron transfer rate between Ni-CA and the substrate electrode. 19nsecutive potential cycling on MWCNT-CCE (16 cycles) showed increasing anodic and cathodic currents in a 2:1 mixture of CA and Ni(NO 3 ) 2 in alkaline solution (0.1 mol L -1 NaOH).Figure 3 shows the result of repeated cyclic voltammograms in the range of 0.0-0.8V (vs.SCE) with a scan rate of 50 mV s -1 .Increasing of anodic and cathodic currents with the number of potential cycles confirms the electrodeposition of the redox species (Ni-CA) on MWCNT-CCE.The ratio of cathodic peak current (Ipc) to anodic one (Ipa) was about unity, which showed the stability of the oxidation product at the electrode surface.

Effect of pH on redox behavior of Ni-CA/MWCNT-CCE
The influence of pH was examined on the voltammetric behavior of Ni-CA/MWCNT-CCE in the pH range from 10 to 13, and the results are shown in Figure 4.As is shown, by increasing the pH, enhancement of anodic and cathodic peak currents was observed and the formal potential Eº' (= (Epa + Epc) / 2) of the redox couple shifted to less positive potentials (Figure 4 inset), suggesting that OH -participated in the redox process of the modified electrode.The large slope (which was reported in literature, previously) 35,36 could not be related to the ratio of the number of H + and electrons in the electron transfer mechanism, due to the quasi-reversible behavior of Ni(II)/Ni(III) redox couple.The peak disappeared at pH ≤ 9 as was reported earlier for the redox couple. 36ltammetric behavior of Ni-CA/MWCNT-CCE in the presence of GP Due to the strong complex formation between GP and Ni(II), 37 the effect of the presence of the drug was examined on the response of Ni-CA/MWCNT-CCE.The electrocatalytic properties of nanotubes of nickel oxides (dispersed in a carbon paste electrode) on the electrooxidation of GP was previously reported. 28s is shown in Figure 5, in the presence of GP (26.7 μmol L -1 ), both anodic and cathodic peaks increased.The increase of oxidation current was attributed to the production of Ni(II) through the reaction between Ni(III) and GP: However, the cathodic current increased as well.This behavior was observed at the surface of nickel-modified electrodes for the electrocatalytic determination of glucose 38 and l-histidine. 39Because Ni(II)/Ni(III) redox couple serves as both an electronical medium (enhancing cathodic and anodic currents simultaneously) and a catalyst, the electrochemical behavior is different from typical mediated catalytic oxidation in which the cathodic peak current decreases.

Effect of scan rate on Ni-CA/MWCNT-CCE in the presence of GP
The influence of scan rate was investigated in the range of 10-100 mV s -1 on the electrochemical behavior of Ni-CA/MWCNT-CCE in the presence of GP.Both cathodic and anodic currents increased with scan rate (Figure 6a).Linear increase of peak current with scan rate (Figure 6b) indicates an adsorptive redox process, which may be due to the tendency of GP to interact with nickel ions at the electrode surface. 36The plot of log(Ipa) against log(υ) was linear (Figure 6c) with a slope of about 0.83, which shows the large contribution of adsorption of GP to the current flow; in the case of diffusion currents, the slope approaches 0.5. 40

Effect of accumulation time
Due to the importance of GP adsorption, the effect of accumulation time (t acc ) was investigated on current response of the modified electrode, as depicted in Figure 7.The peak current increased with t acc up to 10 min, beyond which it leveled off due to the saturation of the active  surface of the electrode.Therefore, the sample solutions were equilibrated with the electrode surface for 10 min before each measurement.

Linear range and limit of detection
Linear sweep voltammetry (LSV) was applied to the determination of GP on Ni-CA/MWCNT-CCE (Figure 8).In the absence of GP, the oxidation peak current is due to Ni(II)/Ni(III) redox couple at the surface of Ni-CA/MWCNT-CCE.In the presence of GP, peak current was increased linearly with GP concentration in the range of 1.25-63.23μmol L -1 , with the regression line: Ip / μA = 0.072 [GP] / μmol L -1 + 4 × 10 -6 , with R² = 0.995.The limit of detection (LOD) was calculated by using the equation: Y LOD = Y B + 3S B , 41 in which Y LOD is the peak current at LOD, Y B is the average current for blank solution (= intercept of the calibration curve), and S B is the standard deviation of replicate measurements of the blank measurements.A value of 0.5 μmol L -1 was calculated for LOD.

Repeatability and stability of Ni-CA/MWCNT-CCE
For six successive determinations of GP (26.7 μmol L -1 ) under optimum conditions, the anodic current was nearly constant (relative standard deviation, RSD% = 0.67%).The excellent repeatability (intra-day) of the results is one of the advantages of CCEs, because, unlike carbon paste electrodes, the porous structure of the electrode is not clogged by the mechanical polishing step, and it is highly resistant to plastic deformations.The inter-day repeatability of the method was examined by using  Ni-CA/MWCNT-CCE in the voltammetric determination of GP (25 μmol L -1 ) in four different days.A value of RSD% = 2.94% was obtained, which shows the stability of the electrode response during using, and storage in a long time period.
The electrode showed excellent stability over a period of 2 months (the peak current for GP was changed to 90% of its initial response after this period), which is partly due to the mechanical robustness of CCEs and resistance to extreme pH conditions.The electrode was shelf-stored when it was not used.
Comparison of the analytical figures of Ni-CA/ MWCNT-CCE for GP determination with similar reports is shown in Table 1.The linear range and LOD of the proposed method is comparable with some of the previous reports.The sensitivity was improved compared to nickel-oxide nanotube carbon paste electrode, 28 which may be due to the larger amount of Ni(II)/Ni(III) redox couple at the surface of Ni-CA/MWCNT-CCE.The presence of MWCNTs and highly porous structure of CCE provide a larger surface area for Ni-CA deposition.

Interference studies
The influence of various inorganic ions and organic compounds on the determination of GP (20 μmol L -1 ) was studied at optimum conditions.The results showed that in the presence of 1000-fold of Ca 2+ , Mg 2+ , Br -, NH 4+ , SO 4 2-, citric acid, and glucose the current change was less than ± 5%.Examination of histidine, glycin, and uric acid showed their tolerance limit about 10-fold compared to GP.

Analytical applications
The applicability of Ni-CA/MWCNT-CCE to the determination of GP was examined in the recovery of certain GP amounts in solution.Two concentrations of GP (33.82 and 49.88 μmol L -1 ) were prepared and the corresponding LSVs were recorded at Ni-CA/MWCNT-CCE under optimum conditions.The recovery of GP was calculated by applying the peak currents in the calibration curve, obtaining the concentrations, and comparing them with the real amounts of GP.The recoveries for the two concentrations of GP used in this experiment were 100.06 and 98.97%, respectively.
Application of the proposed method was also tested by the analysis of GP tablets (100 mg tablet -1 ). Figure 9a shows the LSVs of Ni-CA/MWCNT-CCE in a solution (50 μmol L -1 ) prepared from GP tablets in a standard addition experiment.Figure 9b shows the standard addition

Conclusions
A carbon ceramic composite electrode was prepared in the presence of MWCNTs.Nickel-catechol complex was deposited electrochemically at the surface of the electrode.The porosity of ceramic materials, along with the improvement of electron transfer rate and large surface area provided by MWCNTs, made the electrode a proper microenvironment for the deposition of nickel complex.The prepared electrode showed good catalytic activity in the oxidation of gabapentin.The electrode had a long lifetime and shelf storage advantages, so can be used as the detector of gabapentin in flow system.The robustness of the electrode was advantageous in the polishing step, because of the resistance of CCEs to clogging and deformations, therefore, highly reproducible results were obtained.Determination of gabapentin in dosage forms by the proposed method was satisfactory with a recovery of 97.33 ± 1.73%.

Figure 2 .
Figure 2. Mechanism of the formation of the deposited Ni-CA complex.

Figure 7 .
Figure 7. Effect of accumulation time (t acc ) on anodic peak current of Ni-CA/MWCNT-CCE in the presence of GP.

Table 1 .
Comparison of the efficiency of some modified electrodes in the electrochemical determination of GP Vol. 25, No. 9, 2014 plot.The average of 4 replicate measurements of GP was 97.3 ± 1.73 mg of GP per tablet.The accuracy and precision of the analysis were satisfactory in the determination of GP in formulations.
a Ag nanoparticles.