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Novel nanoclay-based electrochemical sensor for highly efficient electrochemical sensing nimesulide
Journal of Physics and Chemistry of Solids 137 (2020) 109210
Contents lists available at ScienceDirect
Journal of Physics and Chemistry of Solids journal homepage: http://www.elsevier.com/locate/jpcs
Novel nanoclay-based electrochemical sensor for highly efficient electrochemical sensing nimesulide Nagaraj P. Shetti a, **, Shweta J. Malode a, Deepti S. Nayak a, Shikandar D. Bukkitgar a, Gangadhar B. Bagihalli a, Raviraj M. Kulkarni b, Kakarla Raghava Reddy c, * a
Electrochemistry and Materials Group, Department of Chemistry, K.L.E. Institute of Technology, Hubballi, 580030, Affiliated to Visvesvaraya Technological University, Karnataka, India Department of Chemistry, K.L.S. Gogte Institute of Technology (Autonomous), Affiliated to Visvesvaraya Technological University Belagavi, 590008, Karnataka, India c School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, NSW, 2006, Australia b
A R T I C L E I N F O
A B S T R A C T
Keywords: Electrochemical sensors Nanoclay Electrochemistry Nimesulide Voltammetry Heterogeneous rate constant
Quantification of bioactive molecules through voltammetry techniques is significant over the past decades, due to its adaptability and precision of assessment at the trace level. The present study involves a sensitive vol tammetric process for the detection of nimesulide (NIM) was developed at nanoclay (NC) blended carbon paste electrode (CPE) applying voltammetric techniques such as cyclic voltammetry (CV) and square wave voltam metry (SWV) in a media of pH 7.0 phosphate buffer (PB) solution. The proposed nanosensor increased peak current by a less negative shift in the potential due to electro-catalytic properties of nanoclay particles. The persuade of physicochemical constraint on the voltammetric behavior of NIM were studied. Based on the ob tained results, number of protons and electrons participated in the electrooxidation reaction, heterogeneous rate constant value, detection and quantification limits were also calculated. The proposed electrode showed lower detection limit value of 1.01 nM and hence, can be used to determine NIM in tablet and urine samples with good recovery values.
1. Introduction Nimesulide (NIM) is an anti-inflammatory drug, which functions by stopping the prostaglandin production and by this means mitigating the pain caused. NIM also has antipyretic and analgesic assets, which uti lized in the treatment of rheumatoid arthritis, osteoarthritis [1,2]. Including spectrometry, chromatography’s like expensive and tedious techniques were adopted to detect NIM [3–6]. In comparison with other techniques, the electrochemical method exhibits high precision and less time consuming for bioactive molecules estimation [7–10]. Different electrodes were used for the determination of bioactive compounds [11–16]. However, few techniques are available for the estimation of NIM [17–20]. Determination of NIM using High-performance liquid chromatography with ultraviolet technique, thin layer chromatography, spectrophotometry, and capillary zone electrophoresis [21–25]. How ever, the above-mentioned techniques were time-consuming and included complex stages. The electrochemical methods are best known due to their versatility, sensitivity, analysis of samples at the nano level
and less time consumption. Few electrochemical techniques for the detection of NIM were proposed using fabricated glassy carbon electrode (GCE) using multiwalled carbon nanotubes (MWCNTs) with detection limit of 1.6 � 10 7 M [26] and cysteic acid-carbon nanotubes incorpo rated L-cysteine modified GCE with limit of detection 5.0 � 10 8 M [27]. To best of our knowledge, there are no reports on the development of a sensor using nanoclay particles as modifier blended with carbon paste. Clay nanoparticles are employed in various fields due to their large aspect ratio, stability, and mechanical strength. In this work, montmo rillonite (MMT) nanoclay was used as sensor modifier incorporated in carbon paste electrode (CPE). The clay particles structure consists of layers of two types i.e., octahedral and tetrahedral. By interlayer cations, Vander Waals force, electrostatic force, or by hydrogen bonding, the two sheets together can form a layer, and several layers may be joined in a clay crystallite. The MMT belonging to the smectite group with chemical and physical properties has a dioctahedral structure with sheet linkage of 2:1 [28–30]. Clay minerals are porous materials with particle size ranging from 0.002 to 0.001 mm in diameter results in higher surface
* Corresponding author. ** Corresponding author. E-mail addresses: [email protected] (N.P. Shetti), [email protected] (K.R. Reddy). https://doi.org/10.1016/j.jpcs.2019.109210 Received 24 July 2019; Received in revised form 30 August 2019; Accepted 18 September 2019 Available online 19 September 2019 0022-3697/© 2019 Elsevier Ltd. All rights reserved.
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Journal of Physics and Chemistry of Solids 137 (2020) 109210
area per unit volume. As a result, a large number of cations can be adsorbed, which convey a considerable electrical conductivity in clay minerals. Nowadays, the use of montmorillonite clay minerals has been appreciably amplified. These clay particles can be employed as modi fiers in physical and chemical systems. The NC as modifiers in drug determination exhibits excellent sensitivity, selectivity, and stability [29–31]. Carbon paste has been extensively functional in electrochemistry since it can be exploited at both positive and negative potentials ranging from 1.4 to þ1.3 V versus Ag|AgCl reference electrode. The modifi cation improves analytical activity by increasing its stability, sensitivity, and selectivity in the reactions. The electrocatalytic activity of the nanosensor is based on the catalytic chemical reactions taking place at the active surface of the electrode with an immobilized modifier. The modification helps to shift the potential to a lower value, separation of overlapping of voltammetric signals, and enhancement in the peak response, etc [32–35]. The literature provides that the attractive prop erties like great potential range, voltammetric scan reproducibility, less expenditure, elementary fabrication method, superior permanence, secure discard of utilized paste were encouraged the researchers to pick CPEs for an extensive series of bioactive drugs determination [30–36]. In this paper, for the first time, we report nanoclay (NC) particles for nanosensor applications in the determination of Nimesulide. The elec trochemical sensing characteristics were investigated using cyclic vol tammetry (CV), linear sweep voltammetry (LSV), and square wave voltammetry (SWV) techniques, and results showed that the NC sensor has excellent efficiency in recovery and low detection limit, demon strating that this efficient nanosensor is useful in the clinical trials.
3. Results and discussion 3.1. Surface area of the nanosensor The surface area of the nanosensor was determined using the Randles-Sevcik equation. The cyclic voltammograms at different scan rates were obtained using KCl solution (0.1 M) as supporting electrolyte and K3[Fe(CN)6] (1.0 mM) as a test solution. Using the current and po tential values and substituting in equation (1), we found an area of 0.042 cm2 for pure CPE and 0.086 cm2 area for NC-CPE [38]. 1/2 C0* Ip ¼ (2.69 � 105) n3/2 A D1/2 0 ν
(1)
3.2. Pre-concentration time and amount of modifier The effect of pre-concentration time on NIM electrochemical behavior was studied in a range of 0–120 s. At 30 s, an enhancement in the peak current was recorded (Fig. 1) results in higher adsorption of analyte on the nanosensor. Hence, 30 s was set as accumulation time for further experimental analysis. The effect of modifier amount is important in knowing the electro catalytic activity of modifier and analyte molecules. The fabrication of pure and modified nanosensor is already described in section 2.2 and from the observation, 0.05 g of NC particles were found be most favor able to develop modified CPE. 3.3. Voltammetric behavior of NIM
2. Experimental
Fig. 2 indicates CVs of CPE and NC-CPE, in the presence and absence
2.1. Instrumentation and reagents The electrochemical analyzer of CHI Company, model D630, USA was utilized for voltammetric estimations. The system was incorporated with three-electrode glass cell consisting of the main working electrode as NC modified carbon paste electrode (NC-CPE), auxiliary electrode as platinum wire, and reference electrode as Ag|AgCl (3.0 M KCl) corre spondingly. The pH of the solutions was measured using pH meter, Elico Ltd., India. The essential chemicals were procured from Sigma Aldrich (USA). In this entire experimentation water used was distilled double times. The NIM solution of 1.0 mM was prepared using ethanol. Phosphate buffer (0.2 M ionic strength) of pH ranging from 3.0 to 11.2 was used as sup porting media [37]. 2.2. Nanosensor preparation The pure paste electrode was organized by blending of graphite powder and paraffin oil (7:3) and filled in hollow polytetrafluoro ethylene tube (PTFE). The inserted paste was changed with new paste, after each measurement. Likewise, the NC-CPE was organized using clay nanoparticles in a pertinent amount to the pure CPE and used for the experiment. 2.3. Pharmaceutical and biological fluid sample analysis NIM containing pills sample solution was prepared along with filtration and sonication. Clear supernatant liquid of aliquots was diluted with pH 7.0 PB solution. For NIM detection as well as recovery SWV technique was applied and recovery values were calculated using the standard addition method. From healthy candidates, collected urine samples and centrifuged for 5 min at 25 � 0.2 � C. The resultant urine sample was diluted using pH 7.0 PB solution, and the sample was analyzed by SWV technique by adding a known amount of NIM (1.0 mM) to the filtrate.
Fig. 1. Effect of pre-concentration time on the NIM peak current. 2
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Journal of Physics and Chemistry of Solids 137 (2020) 109210
Fig. 2. Electrochemical behavior of 1.0 mM NIM in pH 7.0 PB solution at ʋ of 50 mVs 1; Acc. time of 30s: CVs of, (a) blank CPE; (b) 1.0 mM NIM at CPE; (c) blank NC-CPE; (d) 1.0 mM NIM at NC-CPE. Inset: Peak current and potential at both the electrodes.
of 1.0 mM NIM at a scan rate of 50 mVs 1 in pH 7.0 PB media. In presence of 1.0 mM NIM, an abridged peak was observed at potential 0.8917 V and peak current of 3.738 μA for pure CPE with an irreversible nature. While at NC-CPE, a similar performance through an accentuated anodic peak at potential 0.8837 V and peak current of 35.35 μA.
Fig. 3. Effect of supporting electrolyte on 1.0 mM NIM at NC-CPE; ʋ of 50 mVs 1; Acc. time of 30s; Influence of, (A) pH vs Ep/V; (B) pH vs Ip/μA (No. of experimental relative to error bar is five).
involved were calculated. Here, Ep/2 is the potential, where current is half of the peak value. Using equations, obtained ‘α’ value as 0.55, ‘k0’ as 3.74 � 103 s 1, ‘n’ to be 1.8 � 2. The chemical structure of Nimesulide was shown in Scheme 1. The possible oxidation mechanism of NIM involving two protons and two electrons is shown in Scheme 2 [44].
3.4. Phosphate buffer solution The changes in peak potential and current of NIM was noticed with variation in supporting electrolyte pH (Fig. 3). The studied pH range was 3.0–11.2 with a sweep rate of 50 mVs 1. It was observed that with an increase in PB solution pH, the peak potential decreased from pH 3.0 to 7.0 and later remained pH-independent up to pH 11.2. From Fig. 3A, we can observe the two linear regions, one region between pH 3.0 to 7.0, i. e., pH < pKa and another between pH 8.0 to 11.2, i.e., pH > pKa. The linear relationship for the plot of Ep vs pH from pH 3.0 to 7.0 is Ep ¼ 0.058 pH þ 1.308; R2 ¼ 0.992 and from pH 8.0 to 11.2 is Ep ¼ 0.003 pH þ 0.928; R2 ¼ 0.903. The increase in the slope value between pH 3.0 to 7.0 indicates the presence of an antecedent acid-base equilibrium with pKa of about 6.8 which corresponds to the pKa of NIM. An enhanced peak was observed in pH 7.0, thus the same pH was main tained throughout the study (Fig. 3B). The slope of 58.0 mV/pH from Ep vs pH plot suggests the involvement of an equal number of protons and electrons in NIM oxidation [39].
4. Analytical applications 4.1. NIM concentration variation Square wave voltammograms (SWVs) for varying concentrations of NIM (0.01 μM–0.35 μM) were recorded at NC-CPE in pH 7.0 media (Fig. 5). The linear equation, Ip ¼ 29.47C þ 0.287; R2 ¼ 0.991 for the calibration plot was obtained. The limit of detection (LOD) and quan tification (LOQ) values of 1.01 nM and 3.37 nM were calculated using 3S/M and 10S/M equations (S ¼ standard deviation, M ¼ slope), respectively [45]. The lower value of detection and quantification by current method suggest the sensitivity of the method (Table 1).
3.5. Scan rate
4.2. Estimation of NIM in pharmaceutical and urine samples
The effect of scan rate on NIM voltammetry performance, LSV technique was applied using NC-CPE in 7.0 pH PB solution (Fig. 4). The potentials of the NIM peak shifted to positive potential values as the scan rate was increased (Fig. 4A). Using the slope value of plot of log Ip vs log υ, the process was found to be diffusion-controlled i.e., 0.669; which is nearer to 0.5 theoretically (Fig. 4B) [40,41]. The corresponding equation is log Ip (μA) ¼ 0.669 log υ þ 2.23; R2 ¼ 0.999. The relationship between Epvs log υ was also found to be linear Ep (V) ¼ 0.059 log υ þ 1.221; R2 ¼ 0.991(Fig. 4C). By Laviron’s theory, ʋ and Ep relationship can be stated as [42], Using the Bard and Faulkner [43] equation, the transfer coefficient (α), the heterogeneous rate constant, and the number of electrons
The preparation of pharmaceutical and urine samples for the inves tigation was prepared as discussed in subsection 2.3. The recovery studies were carried out using SWV technique (Table 2) and the results are satisfying for NIM quantification. 4.3. Effect of excipients and metal salts The effect of excipients on NIM activity was studied to check the accuracy of the proposed method. From Fig. 6A, it can be observed that 1.0 mM excipients namely Uric acid, Urea, Tartaric acid, Sucrose, Starch, Lactose, Gum acacia, Glycine, Dextrose, Citric acid, and Ascorbic acid shifted the potential value but did not exceed �5% signal response 3
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Journal of Physics and Chemistry of Solids 137 (2020) 109210
Fig. 4. Linear sweep voltammograms of 1.0 mM NIM at scan rate of: (1) 0.01 to (13) 0.20 Vs-1. Accumulation time of 30s; pH 7.0 at NC-CPE. Plots of(A) Ip/μA vs υ/Vs 1; (B) log Ip/μA vs log υ/Vs 1; (C) Ep/V vs log υ/Vs 1. Table 2 Estimation of NIM in pharmaceutical and spiked human urine samples. Pharmaceutical Samples
Spiked (10
1 2 3
1.0 5.0 8.0
Urine Samples
Spiked (10
1 2 3
0.1 0.5 0.8
5
6
M)
Detected (10
6
M)
0.96 4.80 7.89 M)
Detected (10
Recovery 96.0 96.0 98.6
5
M)
0.097 0.468 0.789
RSD
% RSD
0.020 0.213 0.020
2.05 2.13 2.02
*Average five readings.
of 1.0 � 10 5 M NIM, which suggest that excipients did not interfere with the NIM electroactivity. Similarly, when effect of some common salts of 1.0 mM concentration such as Ammonium sulphate, Ammonium chloride, Sodium chloride, Copper acetate, Potassium chloride, Copper sulphate, Barium chloride, and Potassium nitrate shifted the 1.0 � 10 5 M NIM potential but did not exceed �5% signal response, which implies that the metal salts did not interfere with the NIM elec troactivity (Fig. 6B). Thus, the present nanosensor can be competently used for NIM finding.
Fig. 5. SWVs for NIM concentrations in pH 7.0 at NC-CPE: (1) Buffer to (17) 0.6 μM. (A) Plot of concentration vs peak current. Table 1 LOD of NIM using an array of working electrodes. Sensors utilized
pH utilized
Linearity range (M)
LOD (nM)
Reference
Barium-doped zinc oxide nanoparticles modified glassy carbon electrode Titanium dioxide nanoparticles modified glassy carbon electrode Multiwalled carbon nanotubes modified glassy carbon electrode Cysteic acid and carbon nanotubes modified glassy carbon electrode Nanoclay modified carbon paste electrode
0.2 M pH 7.0 PBS 0.2 M pH 2.0 PBS 0.2 M pH 6.6 PBS 0.05 M H2SO4 0.2 M pH 7.0 PBS
1.0 � 10 5–1.0 � 10 7 40 � 10 6 - 100 � 10 6 3.2 � 10 7–6.5 � 10 5 1.0 � 10 7–1.0 � 10 5 0.01 � 10 6–0.35 � 10 6
1.79 3.37 160.0 50.0 1.01
[15] [20] [26] [27] [Present work]
4
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Journal of Physics and Chemistry of Solids 137 (2020) 109210
Fig. 6. Effect of excipients and metal salts on NIM peak response.
the long-standing reliability of NC-CPE. The reproducibility was checked for intra-day at constant tempera ture keeping a steady concentration of the NIM. The % RSD (relative standard deviation) was 2.5% for five tiresome measurements which exhibit good reproducibility for NIM detection. 5. Conclusions In the present study, the nanoclay particles incorporated CPE were successfully used as a nanosensor for NIM detection. The remarkable increase in the peak response was due to the novel NC-CPE in pH 7.0 PB solution. From the voltammetry study observed an irreversible, diffusion-controlled process, two protons, and two electrons involve ment in the voltammetry of NIM. The LOD and LOQ values of 1.01 nM and 3.37 nM were obtained. The excipients did not interfere with the NIM electroactivity. The recovery values for pharmaceutical and urine samples were satisfying for NIM quantification. High sensitivity, selec tivity and low value of LOD suggest the applicability of the method for
Scheme 1. Chemical structure of Nimesulide (NIM).
4.4. Repeatability and reproducibility The stability of the nanosensor was made sure for 20 days, by conserving the nanosensor in an airtight container. The sensor sustained 98% of its peak response for a concentration of 1.0 mM NIM, suggests
Scheme 2. Oxidation mechanism of NIM. 5
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Journal of Physics and Chemistry of Solids 137 (2020) 109210
the detection and estimation of NIM in pharmaceutical and spiked human urine samples.
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Appendix A. Supplementary data Supplementary data to this article can be found online at https://doi. org/10.1016/j.jpcs.2019.109210. References [1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] [14] [15] [16] [17] [18] [19] [20]
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6
Contents lists available at ScienceDirect
Journal of Physics and Chemistry of Solids journal homepage: http://www.elsevier.com/locate/jpcs
Novel nanoclay-based electrochemical sensor for highly efficient electrochemical sensing nimesulide Nagaraj P. Shetti a, **, Shweta J. Malode a, Deepti S. Nayak a, Shikandar D. Bukkitgar a, Gangadhar B. Bagihalli a, Raviraj M. Kulkarni b, Kakarla Raghava Reddy c, * a
Electrochemistry and Materials Group, Department of Chemistry, K.L.E. Institute of Technology, Hubballi, 580030, Affiliated to Visvesvaraya Technological University, Karnataka, India Department of Chemistry, K.L.S. Gogte Institute of Technology (Autonomous), Affiliated to Visvesvaraya Technological University Belagavi, 590008, Karnataka, India c School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, NSW, 2006, Australia b
A R T I C L E I N F O
A B S T R A C T
Keywords: Electrochemical sensors Nanoclay Electrochemistry Nimesulide Voltammetry Heterogeneous rate constant
Quantification of bioactive molecules through voltammetry techniques is significant over the past decades, due to its adaptability and precision of assessment at the trace level. The present study involves a sensitive vol tammetric process for the detection of nimesulide (NIM) was developed at nanoclay (NC) blended carbon paste electrode (CPE) applying voltammetric techniques such as cyclic voltammetry (CV) and square wave voltam metry (SWV) in a media of pH 7.0 phosphate buffer (PB) solution. The proposed nanosensor increased peak current by a less negative shift in the potential due to electro-catalytic properties of nanoclay particles. The persuade of physicochemical constraint on the voltammetric behavior of NIM were studied. Based on the ob tained results, number of protons and electrons participated in the electrooxidation reaction, heterogeneous rate constant value, detection and quantification limits were also calculated. The proposed electrode showed lower detection limit value of 1.01 nM and hence, can be used to determine NIM in tablet and urine samples with good recovery values.
1. Introduction Nimesulide (NIM) is an anti-inflammatory drug, which functions by stopping the prostaglandin production and by this means mitigating the pain caused. NIM also has antipyretic and analgesic assets, which uti lized in the treatment of rheumatoid arthritis, osteoarthritis [1,2]. Including spectrometry, chromatography’s like expensive and tedious techniques were adopted to detect NIM [3–6]. In comparison with other techniques, the electrochemical method exhibits high precision and less time consuming for bioactive molecules estimation [7–10]. Different electrodes were used for the determination of bioactive compounds [11–16]. However, few techniques are available for the estimation of NIM [17–20]. Determination of NIM using High-performance liquid chromatography with ultraviolet technique, thin layer chromatography, spectrophotometry, and capillary zone electrophoresis [21–25]. How ever, the above-mentioned techniques were time-consuming and included complex stages. The electrochemical methods are best known due to their versatility, sensitivity, analysis of samples at the nano level
and less time consumption. Few electrochemical techniques for the detection of NIM were proposed using fabricated glassy carbon electrode (GCE) using multiwalled carbon nanotubes (MWCNTs) with detection limit of 1.6 � 10 7 M [26] and cysteic acid-carbon nanotubes incorpo rated L-cysteine modified GCE with limit of detection 5.0 � 10 8 M [27]. To best of our knowledge, there are no reports on the development of a sensor using nanoclay particles as modifier blended with carbon paste. Clay nanoparticles are employed in various fields due to their large aspect ratio, stability, and mechanical strength. In this work, montmo rillonite (MMT) nanoclay was used as sensor modifier incorporated in carbon paste electrode (CPE). The clay particles structure consists of layers of two types i.e., octahedral and tetrahedral. By interlayer cations, Vander Waals force, electrostatic force, or by hydrogen bonding, the two sheets together can form a layer, and several layers may be joined in a clay crystallite. The MMT belonging to the smectite group with chemical and physical properties has a dioctahedral structure with sheet linkage of 2:1 [28–30]. Clay minerals are porous materials with particle size ranging from 0.002 to 0.001 mm in diameter results in higher surface
* Corresponding author. ** Corresponding author. E-mail addresses: [email protected] (N.P. Shetti), [email protected] (K.R. Reddy). https://doi.org/10.1016/j.jpcs.2019.109210 Received 24 July 2019; Received in revised form 30 August 2019; Accepted 18 September 2019 Available online 19 September 2019 0022-3697/© 2019 Elsevier Ltd. All rights reserved.
N.P. Shetti et al.
Journal of Physics and Chemistry of Solids 137 (2020) 109210
area per unit volume. As a result, a large number of cations can be adsorbed, which convey a considerable electrical conductivity in clay minerals. Nowadays, the use of montmorillonite clay minerals has been appreciably amplified. These clay particles can be employed as modi fiers in physical and chemical systems. The NC as modifiers in drug determination exhibits excellent sensitivity, selectivity, and stability [29–31]. Carbon paste has been extensively functional in electrochemistry since it can be exploited at both positive and negative potentials ranging from 1.4 to þ1.3 V versus Ag|AgCl reference electrode. The modifi cation improves analytical activity by increasing its stability, sensitivity, and selectivity in the reactions. The electrocatalytic activity of the nanosensor is based on the catalytic chemical reactions taking place at the active surface of the electrode with an immobilized modifier. The modification helps to shift the potential to a lower value, separation of overlapping of voltammetric signals, and enhancement in the peak response, etc [32–35]. The literature provides that the attractive prop erties like great potential range, voltammetric scan reproducibility, less expenditure, elementary fabrication method, superior permanence, secure discard of utilized paste were encouraged the researchers to pick CPEs for an extensive series of bioactive drugs determination [30–36]. In this paper, for the first time, we report nanoclay (NC) particles for nanosensor applications in the determination of Nimesulide. The elec trochemical sensing characteristics were investigated using cyclic vol tammetry (CV), linear sweep voltammetry (LSV), and square wave voltammetry (SWV) techniques, and results showed that the NC sensor has excellent efficiency in recovery and low detection limit, demon strating that this efficient nanosensor is useful in the clinical trials.
3. Results and discussion 3.1. Surface area of the nanosensor The surface area of the nanosensor was determined using the Randles-Sevcik equation. The cyclic voltammograms at different scan rates were obtained using KCl solution (0.1 M) as supporting electrolyte and K3[Fe(CN)6] (1.0 mM) as a test solution. Using the current and po tential values and substituting in equation (1), we found an area of 0.042 cm2 for pure CPE and 0.086 cm2 area for NC-CPE [38]. 1/2 C0* Ip ¼ (2.69 � 105) n3/2 A D1/2 0 ν
(1)
3.2. Pre-concentration time and amount of modifier The effect of pre-concentration time on NIM electrochemical behavior was studied in a range of 0–120 s. At 30 s, an enhancement in the peak current was recorded (Fig. 1) results in higher adsorption of analyte on the nanosensor. Hence, 30 s was set as accumulation time for further experimental analysis. The effect of modifier amount is important in knowing the electro catalytic activity of modifier and analyte molecules. The fabrication of pure and modified nanosensor is already described in section 2.2 and from the observation, 0.05 g of NC particles were found be most favor able to develop modified CPE. 3.3. Voltammetric behavior of NIM
2. Experimental
Fig. 2 indicates CVs of CPE and NC-CPE, in the presence and absence
2.1. Instrumentation and reagents The electrochemical analyzer of CHI Company, model D630, USA was utilized for voltammetric estimations. The system was incorporated with three-electrode glass cell consisting of the main working electrode as NC modified carbon paste electrode (NC-CPE), auxiliary electrode as platinum wire, and reference electrode as Ag|AgCl (3.0 M KCl) corre spondingly. The pH of the solutions was measured using pH meter, Elico Ltd., India. The essential chemicals were procured from Sigma Aldrich (USA). In this entire experimentation water used was distilled double times. The NIM solution of 1.0 mM was prepared using ethanol. Phosphate buffer (0.2 M ionic strength) of pH ranging from 3.0 to 11.2 was used as sup porting media [37]. 2.2. Nanosensor preparation The pure paste electrode was organized by blending of graphite powder and paraffin oil (7:3) and filled in hollow polytetrafluoro ethylene tube (PTFE). The inserted paste was changed with new paste, after each measurement. Likewise, the NC-CPE was organized using clay nanoparticles in a pertinent amount to the pure CPE and used for the experiment. 2.3. Pharmaceutical and biological fluid sample analysis NIM containing pills sample solution was prepared along with filtration and sonication. Clear supernatant liquid of aliquots was diluted with pH 7.0 PB solution. For NIM detection as well as recovery SWV technique was applied and recovery values were calculated using the standard addition method. From healthy candidates, collected urine samples and centrifuged for 5 min at 25 � 0.2 � C. The resultant urine sample was diluted using pH 7.0 PB solution, and the sample was analyzed by SWV technique by adding a known amount of NIM (1.0 mM) to the filtrate.
Fig. 1. Effect of pre-concentration time on the NIM peak current. 2
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Journal of Physics and Chemistry of Solids 137 (2020) 109210
Fig. 2. Electrochemical behavior of 1.0 mM NIM in pH 7.0 PB solution at ʋ of 50 mVs 1; Acc. time of 30s: CVs of, (a) blank CPE; (b) 1.0 mM NIM at CPE; (c) blank NC-CPE; (d) 1.0 mM NIM at NC-CPE. Inset: Peak current and potential at both the electrodes.
of 1.0 mM NIM at a scan rate of 50 mVs 1 in pH 7.0 PB media. In presence of 1.0 mM NIM, an abridged peak was observed at potential 0.8917 V and peak current of 3.738 μA for pure CPE with an irreversible nature. While at NC-CPE, a similar performance through an accentuated anodic peak at potential 0.8837 V and peak current of 35.35 μA.
Fig. 3. Effect of supporting electrolyte on 1.0 mM NIM at NC-CPE; ʋ of 50 mVs 1; Acc. time of 30s; Influence of, (A) pH vs Ep/V; (B) pH vs Ip/μA (No. of experimental relative to error bar is five).
involved were calculated. Here, Ep/2 is the potential, where current is half of the peak value. Using equations, obtained ‘α’ value as 0.55, ‘k0’ as 3.74 � 103 s 1, ‘n’ to be 1.8 � 2. The chemical structure of Nimesulide was shown in Scheme 1. The possible oxidation mechanism of NIM involving two protons and two electrons is shown in Scheme 2 [44].
3.4. Phosphate buffer solution The changes in peak potential and current of NIM was noticed with variation in supporting electrolyte pH (Fig. 3). The studied pH range was 3.0–11.2 with a sweep rate of 50 mVs 1. It was observed that with an increase in PB solution pH, the peak potential decreased from pH 3.0 to 7.0 and later remained pH-independent up to pH 11.2. From Fig. 3A, we can observe the two linear regions, one region between pH 3.0 to 7.0, i. e., pH < pKa and another between pH 8.0 to 11.2, i.e., pH > pKa. The linear relationship for the plot of Ep vs pH from pH 3.0 to 7.0 is Ep ¼ 0.058 pH þ 1.308; R2 ¼ 0.992 and from pH 8.0 to 11.2 is Ep ¼ 0.003 pH þ 0.928; R2 ¼ 0.903. The increase in the slope value between pH 3.0 to 7.0 indicates the presence of an antecedent acid-base equilibrium with pKa of about 6.8 which corresponds to the pKa of NIM. An enhanced peak was observed in pH 7.0, thus the same pH was main tained throughout the study (Fig. 3B). The slope of 58.0 mV/pH from Ep vs pH plot suggests the involvement of an equal number of protons and electrons in NIM oxidation [39].
4. Analytical applications 4.1. NIM concentration variation Square wave voltammograms (SWVs) for varying concentrations of NIM (0.01 μM–0.35 μM) were recorded at NC-CPE in pH 7.0 media (Fig. 5). The linear equation, Ip ¼ 29.47C þ 0.287; R2 ¼ 0.991 for the calibration plot was obtained. The limit of detection (LOD) and quan tification (LOQ) values of 1.01 nM and 3.37 nM were calculated using 3S/M and 10S/M equations (S ¼ standard deviation, M ¼ slope), respectively [45]. The lower value of detection and quantification by current method suggest the sensitivity of the method (Table 1).
3.5. Scan rate
4.2. Estimation of NIM in pharmaceutical and urine samples
The effect of scan rate on NIM voltammetry performance, LSV technique was applied using NC-CPE in 7.0 pH PB solution (Fig. 4). The potentials of the NIM peak shifted to positive potential values as the scan rate was increased (Fig. 4A). Using the slope value of plot of log Ip vs log υ, the process was found to be diffusion-controlled i.e., 0.669; which is nearer to 0.5 theoretically (Fig. 4B) [40,41]. The corresponding equation is log Ip (μA) ¼ 0.669 log υ þ 2.23; R2 ¼ 0.999. The relationship between Epvs log υ was also found to be linear Ep (V) ¼ 0.059 log υ þ 1.221; R2 ¼ 0.991(Fig. 4C). By Laviron’s theory, ʋ and Ep relationship can be stated as [42], Using the Bard and Faulkner [43] equation, the transfer coefficient (α), the heterogeneous rate constant, and the number of electrons
The preparation of pharmaceutical and urine samples for the inves tigation was prepared as discussed in subsection 2.3. The recovery studies were carried out using SWV technique (Table 2) and the results are satisfying for NIM quantification. 4.3. Effect of excipients and metal salts The effect of excipients on NIM activity was studied to check the accuracy of the proposed method. From Fig. 6A, it can be observed that 1.0 mM excipients namely Uric acid, Urea, Tartaric acid, Sucrose, Starch, Lactose, Gum acacia, Glycine, Dextrose, Citric acid, and Ascorbic acid shifted the potential value but did not exceed �5% signal response 3
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Journal of Physics and Chemistry of Solids 137 (2020) 109210
Fig. 4. Linear sweep voltammograms of 1.0 mM NIM at scan rate of: (1) 0.01 to (13) 0.20 Vs-1. Accumulation time of 30s; pH 7.0 at NC-CPE. Plots of(A) Ip/μA vs υ/Vs 1; (B) log Ip/μA vs log υ/Vs 1; (C) Ep/V vs log υ/Vs 1. Table 2 Estimation of NIM in pharmaceutical and spiked human urine samples. Pharmaceutical Samples
Spiked (10
1 2 3
1.0 5.0 8.0
Urine Samples
Spiked (10
1 2 3
0.1 0.5 0.8
5
6
M)
Detected (10
6
M)
0.96 4.80 7.89 M)
Detected (10
Recovery 96.0 96.0 98.6
5
M)
0.097 0.468 0.789
RSD
% RSD
0.020 0.213 0.020
2.05 2.13 2.02
*Average five readings.
of 1.0 � 10 5 M NIM, which suggest that excipients did not interfere with the NIM electroactivity. Similarly, when effect of some common salts of 1.0 mM concentration such as Ammonium sulphate, Ammonium chloride, Sodium chloride, Copper acetate, Potassium chloride, Copper sulphate, Barium chloride, and Potassium nitrate shifted the 1.0 � 10 5 M NIM potential but did not exceed �5% signal response, which implies that the metal salts did not interfere with the NIM elec troactivity (Fig. 6B). Thus, the present nanosensor can be competently used for NIM finding.
Fig. 5. SWVs for NIM concentrations in pH 7.0 at NC-CPE: (1) Buffer to (17) 0.6 μM. (A) Plot of concentration vs peak current. Table 1 LOD of NIM using an array of working electrodes. Sensors utilized
pH utilized
Linearity range (M)
LOD (nM)
Reference
Barium-doped zinc oxide nanoparticles modified glassy carbon electrode Titanium dioxide nanoparticles modified glassy carbon electrode Multiwalled carbon nanotubes modified glassy carbon electrode Cysteic acid and carbon nanotubes modified glassy carbon electrode Nanoclay modified carbon paste electrode
0.2 M pH 7.0 PBS 0.2 M pH 2.0 PBS 0.2 M pH 6.6 PBS 0.05 M H2SO4 0.2 M pH 7.0 PBS
1.0 � 10 5–1.0 � 10 7 40 � 10 6 - 100 � 10 6 3.2 � 10 7–6.5 � 10 5 1.0 � 10 7–1.0 � 10 5 0.01 � 10 6–0.35 � 10 6
1.79 3.37 160.0 50.0 1.01
[15] [20] [26] [27] [Present work]
4
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Journal of Physics and Chemistry of Solids 137 (2020) 109210
Fig. 6. Effect of excipients and metal salts on NIM peak response.
the long-standing reliability of NC-CPE. The reproducibility was checked for intra-day at constant tempera ture keeping a steady concentration of the NIM. The % RSD (relative standard deviation) was 2.5% for five tiresome measurements which exhibit good reproducibility for NIM detection. 5. Conclusions In the present study, the nanoclay particles incorporated CPE were successfully used as a nanosensor for NIM detection. The remarkable increase in the peak response was due to the novel NC-CPE in pH 7.0 PB solution. From the voltammetry study observed an irreversible, diffusion-controlled process, two protons, and two electrons involve ment in the voltammetry of NIM. The LOD and LOQ values of 1.01 nM and 3.37 nM were obtained. The excipients did not interfere with the NIM electroactivity. The recovery values for pharmaceutical and urine samples were satisfying for NIM quantification. High sensitivity, selec tivity and low value of LOD suggest the applicability of the method for
Scheme 1. Chemical structure of Nimesulide (NIM).
4.4. Repeatability and reproducibility The stability of the nanosensor was made sure for 20 days, by conserving the nanosensor in an airtight container. The sensor sustained 98% of its peak response for a concentration of 1.0 mM NIM, suggests
Scheme 2. Oxidation mechanism of NIM. 5
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the detection and estimation of NIM in pharmaceutical and spiked human urine samples.
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