Physical Chemistry Communications, Volume 3 Issue 1, April 2016 www.bacpl.org/j/pcc
Renewable Graphite Pencil Electrode as a Sensor for the Direct Determination of PerazineDimaleate Jayant I.gowda*a, Sharanappa T. Nandibewoorb P.G. Department of Chemistry, P.C. Jabin Science College, Hubballi, India‐580031.
a
P.G. Department of Studies in Chemistry, Karnatak University, Dharwad, India‐580003.
b
jayantgowda4@gmail.com; bstnandibewoor@yahoo.com
*a
Abstract Direct electrochemistry of perazinedimaleate (PDM) was performed at a graphite pencil electrode using cyclic and differential pulsevoltammetryover a wide range of pH. The oxidation of PDM is an irreversible process, pH dependent, and involves the charge transfer of twoelectrons. The results enabled themeasurement of the oxidation peak current to be used as the basis for a simple, accurate and rapid method for determining the investigated compounds, within a concentration range of 1.5 × 10‐7 M to 13.0 × 10‐5 M . The limits of detection (LOD) and quantification (LOQ) calculated from the results obtained at this pH are 8.53 × 10−9 M and 2.84 × 10−8 M, respectively. Promising results were obtained for PDM determination in real samples, without separation from the matrix. Keywords Electro‐Oxidation, Perazinedimaleate, Detection Limit, Real Sample, Pencil Graphite Electrode
Introduction Analytical chemistry plays a major role in the development of a compound from its synthesis stage to its marketing stage as a part of a drug formulation and analysis. In pharmaceutical laboratory, four basic instrumental methods spectrophotometric,2 electrochemical, and radiometric have been used: chromatography,1 3,4 analysis. Electroanalytical chemistry along with the use of oxidation–reduction reactions and other charge‐ transfer phenomena had its origins eight decades ago. It is one of the fundamental sub‐disciplines of analytical chemistry. The development of a simple, sensitive, rapid and reliable method for the determination of drugs is of great importance. The graphite pencil electrode (GPE) has been successfully used as a biosensor in modern electroanalytical field due to its high electrochemical reactivity, good mechanical rigidity, low cost, low technology and ease of modification, renewal and low background current.5,6 The GPE has good application in the analysis of neurotransmitter and detection of traces of metal ions and drugs. Recently many electro‐analytical techniques were employed for the study of different drugs using modified and unmodified pencil graphite electrode.7‐ 12Perazinedimalonate(Chemical structure as shown in Fig. 1) is a phenothiazine with general properties similar to those of chlorpromazine and is used for the treatment of psychotic conditions. It has a piperazine side‐chain. It is given by mouth as the dimalonate although doses are expressed in terms of the base; perazinedimalonate 40.3 mg is equivalent to about 25 mg of perazine. Usual doses are the equivalent of 50 to 600 mg of the base daily; up to 1000 mg daily has been given in resistant cases. It may also be given intramuscularly. There comes out a report of 5 patients receiving perazinedimalonate who developed acute axonal neuropathies of superficial nerve fibres after exposure to sunlight.13 Till now no analytical works have been reported for the determination of PDM. In view of the pharmaceutical importance and biological significance of the drug and to develop a simple, low‐cost direct current voltammetric method for the determination of PDM present work was carried out. We optimized all the experimental parameters for the determination of PDM and developed an electro analytical method for its determination. This method has the advantages such as fast response, easy repair, good reproducibility and low detection limit. The proposed method was applied to the determination of PDM in the urine samples.
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www.bacpl.org/j/pcc Physical Chemistry Communications, Volume 3 Issue 1, April 2016
FIG. 1. CHEMICAL STRUCTURE OF PERAZINE
Experimental Section Reagents and Chemicals The pencil‐lead rods (HB 0.5 mm in diameter and 6 cm length) were purchased from local bookstore. PDM was purchased from Sigma Aldrich. Stock solutions of PDM (1.0 mM) were prepared in millipore water. The phosphate buffering from pH 3.0 to 11.2 were prepared according to the method of Christian and Purdy.14Other reagents used were of analytical or chemical grade. All solutions were prepared with millipore water. Instrumentation and Analytical Procedure Electrochemical measurements were carried out on a CHI 630D electrochemical analyzer (CH Instruments Inc., USA). The voltammetric measurements were carried out in a 10 mL single compartment three‐electrode glass cell with Ag/AgCl as a reference electrode, a platinum wire as counter electrode and a pre‐treated graphite pencil electrode as working electrode. All the potentials are given against the Ag/AgCl (3.0 M KCl). pH measurements were performed with an Elico LI 120 pH meter (ElicoLtd.,India ). All experiments were carried out at an ambient temperature of 25 ± 0.1 0C. Measurement Procedure Stock solution of 1 mM of PDM was prepared by dissolving the desired amount in double distilled water. Required amount of the stock solution was added to electrolytic cell containing phosphate buffer solution. Voltammograms were then recorded using voltammetric analyzer under optimized parameters. The parameters for DPV were initial potential: 0.4 V; final potential: 0.8 V; increase potential: 0.004 V; amplitude: 0.05 V; quiet time: 2 s ; sensitivity: 1 × 10‐6 A/ V. Preparation of the Working Electrode The pencil lead was tightly coated with Teflon band and its tip polished on a weighing paper to a smoothed finish. Electrical contact with the lead was achieved by soldering a copper wire to the metallic holder of the working electrode. Electrochemical Pretreatment of the GPE The electrochemical pretreatment of polished PGE surface carried out potentiodynamically by scanning the potential between ‐1.5 and 2.0 V/SCE with a scan rate of 50 mV s−1 for 50 cycles in 0.2 M phosphate buffer solution (pH 11.2). Results and Discussion Electrochemical Behavior of PDM by Cyclic Voltammetry Cyclic voltammetry is the most widely used technique as it provides considerable information on the thermodynamics of redox processes, the kinetics of heterogeneous electron‐transfer reactions, coupled chemical reactions and adsorption processes; hence, initial studies were carried out using this technique.Cyclic
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Physical Chemistry Communications, Volume 3 Issue 1, April 2016 www.bacpl.org/j/pcc
voltammogramswere recorded for 0.1 mM PDM at GPE at pH 11.2 using a sweep rate of 50 mVs−1. PDM is irreversibly oxidizedgiving rise to an oxidation peak at 0.590 mVas shown in Fig. 2.
FIG. 2. CV CURVES IN 0.2 M PHOSPHATE BUFFER SOLUTION CONTAINING 1.0 × 10‐4 M PDM AT GCE(B), GPE(C). THE ACCUMULATION TIME AND SCAN RATE WERE 2S AND 50 MVS‐1 RESPECTIVELY.
Effect of pH The effect of varying pH of buffer solution on the electrochemical behavior of PDM at GPE was performed using CV in 0.2 M PBS. Fig. 3(a) depicts the response of peak current and potential of PDM to pH. The peak potential was shifted negatively when the solution pH was increased and a good linear relationship was observed between the Ep and pH values (in the range of 3–11.2, Fig. 3(b) with the following equation: Epa(V)= ‐0.053 pH+1.022, (R2 = 0.971) (1) A value of about ‐53 mV/pH unit clearly indicates that equal numbers of electrons and protons are involved in the electro‐oxidation of PDM on the surface of the electrode.15 On the other hand, the anodic peak current of PDM was maximum for pH 11.2, and then decreased with higher pH value (Fig. 3(c)). Therefore, phosphate buffer with pH 11.2 was used as supporting electrolyte in all voltammetric determinations.
(a)
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1 0.9
Ep(V)
0.8 0.7 0.6 0.5 y = -0.053x + 1.022 R² = 0.971
0.4 0.3 0
5
10
pH
15
(b)
Peak current (Ip)
10 9 8 7 6 5 4 3 2 1 0 0
1
2
3
4
5
6 pH
7
8
9
10 11 12
(c) FIG. 3. (A) CVS OF 1.0 × 10‐4 M PDM AT THE GPE IN VARIOUS PHS (FROM 3 TO 11.2: (1) 11.2, (2) 10.4, (3) 9.2, (4) 8.0, (5) 7.4, (6) 6.0, (7) 5.0, (8) 4.0, (9) 3.0) OF BUFFER SOLUTION, DEPENDENCE OF (B) THE OXIDATION PEAK POTENTIAL (EP) AND (C) THE OXIDATION PEAK CURRENT (IP) WITH PH SOLUTION; SCAN RATE 50 MV S−1.
Effect of Potential Scan Rate Useful information involving electrochemical mechanism usually can be gained from the investigation of CVs in the different potential sweep rates. Therefore, the CV investigations for 0.1 mM PDM were performed on the surface of GPE in buffered solution of pH 11.2 at different potential sweep rates. Fig. 4(a) illustrates the influence of scan rate on the cyclic voltammograms of PDM in the range of 25–300 mV s−1. A slope of 0.45 was revealed for the linear relation between the log Ipa and the log v (Fig. 4(b)) indicating a diffusion controlled process16on the surface of the electrode. The regression equation for this relationship is logIpa(10‐5 A)= 0.450 log v (Vs‐1) + 0.965(R2 = 0.970) (2) The relationship between the oxidation peak potential and logarithm of scan rate is shown in Fig. 4(c). It can be seen that the oxidation peak potential shifts positively with increasing scan rate. There is a linear relationship between Epa and the logarithm of the scan rate, Eq.(3). Such a behavior reveals the irreversible nature of the electrochemical process for PDM. Furthermore, only an oxidation peak was observed even at low scan rates, suggesting that the electrode reaction of PDM under these conditions were totally irreversible: Epa(V) = 0.049 log v (Vs‐1) + 0.663 (R2 = 0.979) (3)
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Physical Chemistry Communications, Volume 3 Issue 1, April 2016 www.bacpl.org/j/pcc
As for an irreversible electrode process, according to Laviron,17Ep is defined by the following equation
RTk 0 2.303RT 2.303RT E pa =E 0' + + log log ν αnF αnF αnF
where α is the transfer coefficient, k0 the standard heterogeneous rate constant of the reaction, n the number of electrons transferred, v the scan rate and E0’ is the formal standard redox potential. Other symbols have their usual meaning. Thus,the value of αn can be easily calculated from the slope of Epa vs. log v plot. In this system, the slope is 0.049, taking T = 298 K, R = 8.314 JK‐1mol‐1, and F = 96480 C mol‐1, the αn value was calculated to be 1.205. Taking α=0.5 number of electrons (n) transferred in the electro‐oxidation of PDM was calculated to be 2.41~ 2. The value of k0 can be determined from the intercept of the previous plot, if the value of E0’ is known. The value of E0’ in eqn. (4) can be obtained from the intercept of Epavs. v curve by extrapolating to the vertical axis at v = 0.18In our system, the intercept for Epa vs. log v plot was 0.663 and E0’ was obtained to be 0.608 and the k0 was calculated to be 150.28 s‐1.
(a)
1.2
y = 0.450x + 0.965 R² = 0.970
0.67
1
0.4
0.64 0.63 0.62
Ep(V)
0.6
0.65
log Ip(10-5 A)
0.8
0.66
y = 0.049x + 0.663 R² = 0.979
0.61
0.2
0.6 0 -2
-1.5
-1 log v (V/s)
-0.5
0
0.59 -2
-1.5
-1 log v (V/s)
-0.5
0
(b) (c) FIG. 4. (A) CVS OF 1.0 × 10‐4 M PDM AT GPE AT DIFFERENT SCAN RATES IN 0.2 M PBS (PH 11.2), (B) THE PLOT OF LOG IPA(10‐5A) VS. LOG V (VS‐1) AND (C) VARIATION OF PEAK POTENTIAL (EPA) WITH LOG V (VS‐1).
Analytical Application The DPV method with high current sensitivity and low charging contribution to the background current was applied to estimate the lower limit of detection in the electrochemical determination of the PDM. The DPV obtained with increasing of PDM concentration shows that the peak current increased linearly with increasing of PDM concentration (Fig. 5(a)). The calibration curve for the DPV peak current in PDM oxidation vs. its concentration (Fig. 5(b)) shows a linear calibration curve in the concentration range of 1.5 × 10‐7M to 13.0 × 10‐5 M and the linear equation is Ipa(10‐6A) = 0.089C(μM) + 3.132, (R2 = 0.991) (5)
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The limit of detection (LOD) is obtained as 8.53× 10−9 M and consequently the limit of quantification (LOQ) is calculated as 2.84 × 10−8M, corresponding to following equations; LOD = 3s/m; LOQ = 10s/m Whereas s is the standard deviation of the currents of the blank solution, and m is the slope of the calibration curve.19
(a)
16 y = 0.008x + 3.217 R² = 0.992
14
Current (µA)
12 10 8 6 4 2 0 0
500 1000 Concentration(10-7 M)
1500
(b) FIG. 5. A. DPVS FOR THE ADDITION OF DIFFERENT AMOUNT OF PDM IN PBS (PH 11.2) AT GPE; SCAN RATE 50 MV S−1, B. THE PLOT OF PEAK CURRENT VS. ADDED CONCENTRATION OF PDM.
Reproducibility and Stability The reproducibility for five GPE was carried out by comparing the oxidation peak current of 10 μmol L−1 PDM in a solution. The RSD was 5.4 % for PDM, indicating the good reproducibility of the modified electrode. Only a small decrease of the oxidation peak current of PDM was observed with the RSD of 3.1%, which could be attributed to the excellent repeatability of the GPE. Interference Study On the other hand, some inorganic ions and organic compounds were tested to check their levels of interference in the determination of PDM. The interfering substances used in the study are most commonly used in the pharmaceutical preparations. The tolerance limit was defined as the concentrations of foreign substances, which gave an error of less than ±5.0% at a concentration of 5.0 × 10−6mol L−1 PDM. The results suggested that 100‐fold concentration of citric acid, gum acacia, dextrose, oxalic acid, starch, tartaric has no influence on the signals of PDM in the selected potential range with deviations below 5%. Among the selected organic interferents, ascorbic acid showed significant influence on the oxidation process (Table 1).
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Physical Chemistry Communications, Volume 3 Issue 1, April 2016 www.bacpl.org/j/pcc
TABLE 1.INflUENCE OF POTENTIAL EXCIPIENTS ON THE VOLTAMMETRIC RESPONSE OF 1.0 × 10‐6 M PDM.
Excipients(1.0 mM) + Potential observed (V) Signal change (%) Drug (1.0 × 10‐6) Only PDM 0.590 0 Tartaric acid + PDM 0.598 0.33 Citric acid + PDM 0.575 ‐2.54 Glucose + PDM 0.612 3.72 Gum acacia + PDM 0.601 1.86 Lactose + PDM 0.59 0 Dextrose + PDM 0.602 2.06 Sucrose + PDM 0.589 ‐0.16 Starch + PDM 0.6 1.69 ‐13.22 Ascorbic acid+PDM 0.512 + sign indicates the interfering substance changes the potential of the drug towards the less positive ‐ Sign indicates the interfering substance changes the potential of the drug towards the more positive
Determination of PDM in Urine Samples To investigate the applicability of the proposed sensor for the determination of PDM in real samples, we selected urine samples for the analysis of PDM. The obtained results clearly demonstrate and confirm the capability of the GPE in the voltammetric determination of PDM with high selectivity, accuracy, and good reproducibility. The developed differential voltammetric method for the PDM determination was applied to urine samples. The recoveries from urine were measured by spiking drug free urine with known amounts of PDM. A quantitative analysis can be carried out by adding the standard solution of PDM into the detect system of urine samples, and the peak linearly increased in height. The calibration graph was used for the determination of spiked PDM in urine samples. The detection results of four urine samples obtained are listed in Table 2. The result shows an average recovery of 100.81% for PDM added to the urine samples. TABLE 2.APPLICATION OF DPV TO THE DETERMINATION OF PDM IN SPIKED HUMAN URINE SAMPLE.
Sample
Added (106 M)
Founda (106 M)
Recovery (%)
1 1.0 0.9955 99.55 2 3.0 3.0406 101.35 3 5.0 4.9880 99.76 4 8.0 8.2090 102.621 aAverage of five determinations + sign indicates recovery is less compare to added analyte ‐sign indicates recovery is more compare to added analyte
Average Recovery (%) 100.81
R.S.D (%) 0.437 0.455 0.119 0.613
Bias (%) 0.45 ‐1.35 0.24 ‐2.61
Conclusions In this work, it was demonstrated that renewable graphite pencil as an electrochemical sensor for electrochemical measurements. DPV technique has been developed for the determination of PDM in biological samples. The principal advantage of the proposed method is that the excipients do not interfere and the separation procedure is not necessary. Developed procedure has also been successfully used for serum and urine samples, with good recoveries obtained at the levels tested. The method is rapid, requiring below 5 minutes to run samples and involves no sample preparation other than dissolving and transferring an aliquot to the supporting electrolyte. Furthermore, the present method could possibly be adopted for pharmacokinetic studies as well as clinical and quality control laboratories. REFERENCES
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