1. INTRODUCTION:

Use of nanomaterials for analytical applications has increased enormously in the recent years. In particular nanomaterials play a vital role in the field of chemically modified electrodes (CMEs) for the enhanced behaviour of these electrodes both with respect to sensitivity and selectivity [1]. GNP demonstrate four exclusive compensation over macro electrodes while worn for electro analysis: they are augmentation of, high surface area, catalysis, mass transport and control over electrode microenvironment [2,3].

With these advantages one can use gold nanoparticles as a chemical platform to anchor redox mediators to improve the electron transport behaviour as well as sensitivity of electrode.

Dopamine (3, 4-dihydroxyphenyl ethylamine) is an important neurotransmitter in the mammalian central nervous system (CNS), and it could be detected electrochemically because of its electroactivity. The electrochemical oxidation of dopamine has been studied mostly on carbon base electrodes [4]. Parkinson’s disease (PD) is a neurodegenerative disease which causes progressive disorders in extrapyramidal system that regulates the communication between the neurons in the brain and muscles in the human body. PD involves the breakdown of nerve cells that results in shortage of dopamine which in turn lends to interruption in the blood–brain barrier. Be short of dopamine deliver will mislay manage over the vital faction patterns such as, writing, reaching, walking for objects and other crucial programs cannot function accurately [5, 6].

We have developed a chemically modified electrode by a new approach using gold nanoparticles and nickel hexacyanoferrates for the determination of dopamine. GNP- CdHCF modified electrode was developed by anchoring CdHCF onto the surface of gold nanoparticles using 3-Mercapto-2-aminopropionic acid (L-cysteine) as cross linker, to the metal ion Ni²⁺which was attached with –NH₂group of L-cysteine, this even though COOH group is also present. The metal ion is attached effectively with –NH₂ group which was conformed by FTIR. The gold nano particles synthesized using sodium citrate method was characterized using UV-Vis and cyclic voltammetry. High surface area and size dependent behaviour of gold nanoparticles showed an enhanced electron transport behaviour and high sensitivity for the determination of dopamine. The CdHCF film was effectively used for the amperometric determination of dopamine as it was found to have excellent catalytic activity over the oxidation of dopamine. Differential pulse voltammetry (DPV) and flow injection techniques were effectively used for the determination of the analyte from environmental water samples.

2. EXPERIMENTAL:

2.1 Reagents and chemicals:

All reagents were of analytical grade; Graphite powder was from Aldrich (1-2mm), (Aldrich, Steinheim, Germany). Dopamine from Alfa Aesar (Alfa Aesar chemicals, Kolkata, India). CdCl₂, Potassium ferrocyanide were obtained from Merck (Mumbai, India). Double distilled water was used for all electrochemical experiments. Studies on effect of pH were carried out using 0.1M HCl and 0.1M NaOH solutions. pH 7.0 was maintained using 0.1M KNo_3,0.1 M PBS), L-cysteine solution (20mM) was prepared using double distilled water and cadmium solution (0.01M) was prepared by dissolving the salt in ethanol. Potassium ferrocyanide (0.02 M) in KNO₃ (0.1M) solution was used to derivatize the coordinated nickel ion. All measurements were done after carefully degassing the solutions with pure nitrogen for 10-15 min.

2.2 Apparatus and material:

Electrochemical measurements were carried out using Electrochemical workstation CH Instruments 660B, Tx, USA controlled by an IBM personal computer with standard three-electrode configuration. The surface modified CdHCF graphite paraffin wax composite electrode was used as the working electrode, a platinum wire as the counter electrode and a standard calomel electrode as the reference.

2.3 Sample Preparation:

In order to evaluate the applicability of the proposed method for the determination of DA in real samples, the utility of the developed method was tested by analysis of this compound in mixed synthetic and in real samples using standard addition methods. The results are summarized in Table I. The good recoveries of the sample indicate the successful application of the proposed method for the determination of DA. For further investigation, the recovery of DA was determined for dopamine injection. The dopamine injection solution (specified content of DA was 40.0 mg mL−1) was diluted to 100 mL with water, then a different amount of the diluted solution was transferred into each of a series of 10-mL volumetric flasks and diluted to the mark with phosphate buffer. Then, 10 mL aliquot of this test solution was placed in the electrochemical cell and the DA content was measured by the proposed method. This procedure was repeated five times and the relative standard deviation was found as 1.6 %. Different standard concentrations of DA were added to the diluted DA injection solution, with recoveries between 96.5 and 103.2 % for five measurements.

2.4 Fabrication of GNP-CdHCF paraffin wax composite electrode:

Synthesized GNP solution was added to 1g of graphite powder and its stirred for two hours in a normal temperature. The stirred mixture was centrifuged at 1450 rpm for 30 minutes and the residue was kept overnight for drying. The electrode was modify with 4:1 ratio (Graphite: Paraffin wax) Gold nanoparticles graphite wax mixture was tightly packed in a small glass tube of 3mm diameter. The electrode was removed gently from the tube after it turns hard. The electrode showed an admirable resistance and conductivity. The electrode was first dipped in L- cysteine solution (20 mM ) for 2 hours. Then, this L-cysteine modified GNP composite electrode was dipped in 0.01 M ethanolic solution of CdCl₂ for 2 minutes. The amino group of cysteine functionalized with gold nanoparticles has a greater affinity towards transition metal ion like Cd²⁺. The Cd²⁺ ions coordinated to amine group was then derivatized using 0.02 M potassium ferrocyanide solution dissolved in 0.1 M KNO₃by cycling the potential in the range -0.2 V to 1.0 V at the scan rate of 50 mV s^–1[7]

3. RESULTS AND DISCUSSION:

3.1 SEM characterization of the surface:

The SEM images of electrodes are performed in Fig. 1. shows the surface of the electrode prepared from GNP – graphite mixture and the presence of GNP / CdHCF particle on the electrode surface which has a size of 60-70 nm.

Fig. 1. The SEM images of presence of GNP on graphite matrix with CdHCF particles on the modified electrode.

3.3. Effect of supporting electrolytes and scan rate:

The CdHCF modified electrode was characterized by cyclic voltammetry and the effect of supporting electrolyte was studied. It was found that among the cations studied, (Na⁺, K⁺, Ba²⁺, Ca²⁺, NH₄⁺ at 0.1M concen tration) K⁺ gave a well-resolved and sharp peak. So 0.1M KNO₃was chosen as the background electrolyte that is shown in fig.2.

Fig. 2 Cyclic voltammograms of the GNP-CdHCF modified composite electrode in the presence of different supporting electrolytes 0.1 M (a) KNO_3,(b) BaNO₃ (d) LiNO₃ (g)NaNO₃; scan rate: 20 mVs^-1.

The effect of scan rate on the modified electrode was also studied at different scan rates in the range of 10-150mV/s. in fig.3. The ratio of i_pa/i_pcwas found to be almost close to unity in the range studied. The CdHCF modified electrode showed a ∆E_pof 0.54 V for a potential scan rate of 20mV/s. This slight deviation from ideal behavior arising even at low scan could be attributed to the limitations associated with charge transfer in the film. Wider splitting was observed at higher scan rates (> 200mVs^-1) indicating the limitations arising from the charge transfer kinetics. The anodic and cathodic peak currents were linearly proportional to the square root of the scan rate (v^1/2) with a correlation coefficient of 0.999, which is expected for a diffusion-controlled process (Fig. 3b). The electron transfer coefficient () was calculated from the slope of the plot of Ep Vs log v and was found to be 0.42 and the heterogeneous electron transfer rate constant (K_s) of the CdHCF modified graphite electrode was estimated to be 0.766 s^-1.[8].

Fig. 3.cyclic voltammogram of the CdHCF modified electrode at different scan rates in 0.1M KNO₃(pH 7.0).

The scan rates from inside to outer are 10-100 mVs^-1with increments of 10 from 10 mV^-1, (3a) dependence of peak current I_pa and I_pcon square root of scan rate(υ), (3b) variation of peak potential vs. logarithm of scan rates (logυ).

The performance of the modified electrode was studied under different pH conditions in the range of 2-9. The pH of the background electrolytes was varied using HCl and NaOH. It was found that the peak current did not vary much in the pH range 2-6 but a maximum response in current was obtained at pH 7 (Fig 4). The current response again decreases at pH 8 and above. The poor response at very basic pH could be due to the hydroxylation of the mediator [9]. The hydroxylated moiety formed is electrochemically inactive and gets dissolved into the background electrolyte which results in a decrease in current response. Hence a neutral pH was chosen for subsequent experiments.

Fig.4 Plot of pH Vs Current for modified electrode.

It was found that the peak current did not vary much in the pH range 2-6 but a maximum response in current was obtained at pH 7 .

3.5 Electrocatalytic oxidation of dopamine at the modified electrode:

The electrocatalytic property exhibited by the CdHCF-GNP graphite wax composite electrode for the oxidation of dopamine in 0.1M KNO₃is also shown in Fig.5. It is seen that on bare graphite wax composite electrode, the oxidation of dopamine occurs at a higher potential around 0.74V (curve c) whereas on the CdHCF-GNP graphite wax composite electrode, the oxidation of dopamine occurs at 0.55 V(curve d).Also the catalytic current at the modified electrode was nearly 14 times higher than that obtained at the bare electrode for dopamine oxidation. The modified electrode showed linear response for the catalytic oxidation of dopamine. The linear range for the determination of dopamine is from 2.6X10^-6 to 7.2X10^-3M with a correlation coefficient of 0.999.(Fig.5a) and the detection limit was 1X10^-7M.

Fig.5. Cyclic voltammograms in 0.1 M KNO₃ (pH 7.0)at a scan rate of 20 mVs^-1 (a) bare electrode in the absence of dopamine (b) bare electrode in the presence of dopamine l (c) modified electrode in the absence of dopamine (d) CdHCF modified electrode in the presence of 7.6x10^-5M dopamine .Fig.5acalibration graph for dopamine measurement.

Hydrodynamic voltammetric studies on the bare and the CdHCF –GNP graphite wax composite electrode were carried out in the potential range between 0 to 1.0V in 0.1M KNO₃in order to check the applicability of the modified electrode in flow system. The bare electrode showed a poor response to dopamine whereas there was a considerable increase in response with the modified electrode around 0.54V (Fig.6). Hence 0.55V was chosen as the operating potential for amperometric studies.

Fig.6. Hydrodynamic voltammograms obtained with (a, b) CdHCF film modified and bare composite electrode in the presence of 9.196×10⁻⁵M DA. 0.1 M KNO₃ stirring rate: 300 rpm.

3.6. Flow injection analysis:

The flow- rate dependence of the current response at a constant concentration of dopamine was examined by recording the peak currents at different flow rates. The current response was found to be decrease with increasing flow rate. At a flow rate of 0.5 min^-1, the determination of dopamine can be performed in 2 min including the sampling and washing.Fig.7 displayed the flow injection response of the CdHCF-GNP- graphite wax composite electrode for dopamine solution of increasing concentration from 8.9, 19.2 23.3 µM. Well-defined and sharp peaks were observed at a detection potential of 0.54V (versus Ag/Agcl). The flow injection peak currents were proportional to the dopamine concentration. The resulting calibration plot had a slope of 18.9 µA mM^-1for CdHCF modified electrode, and the correlation coefficient of 0.997. A detection limit of 1X10^-7M, can be estimated on the signal-to-noise ratio (S/N=3). Fig.7a shows the modified electrode response to same concentration of dopamine (spiked) in environmental water sample. Both the results appear almost in agreement with each other in sensitivity. The results indicated that the detection limits are appreciably low and are more sensitive. In all cases the response was rapid and reproducible. After an initial loss activity, the electrodes exhibited a very stable response during hours of continuous flow injection.

Fig. 7. Amperometric response of the CdHCF-NP modified graphite- wax composite electrode for the successive addition of 0.1 ml of .001 M dopamine in 0.1 M KNO₃, Working potential: 0.55V

3.7. Interferences:

In order to emphasize the selectivity of the modified electrode, the effect of possible interferents such as ascorbic acid, uric acid, citric acid, glutathione, H₂O₂, glucose and fructose on the determination of dopamine was examined. Except ascorbic acid, all the other compounds were not found to alter the response for the dopamine determination even if they are found in 100-fold excess. However, ascorbic acid found to interfere severely in the determination of dopamine when found in 1:1 ratio or in excess and hence it has to be removed prior to analysis

3.8 Determination of dopamine from urine samples:

Urine samples were analyzed directly after diluting 25 times with buffer solution (pH 5.0) without any further pretreatment. Then, 5 mL of this test solution was transferred into the electrochemical cell and the DA content was determined. Table 1.

Table 1 Real Sample analysis of dopamine

Samples

Concentration of Dopamine (mM)

Recovery (%)

Added

Found^a

Sample I

19.20± 0.70

39.81± 0.49

98.4

99.5

Sample II

19.9 ± 0.50

39.68 ± 0.48

99.6

99.2

Sample III

19.76 ± 0.48

39.16 ± 0.47

99.2

97.9

Sample IV

19.82 ± 0.49

38.99 ± 0.45

96.06

97.47

4. CONCLUSION:

A GNP graphite paraffin wax composite electrode was prepared by the oxidative electroderivatization of CdHCF in 0.1 M KNO₃, and the resulting modified electrode exhibited a good electrocatalytic activity and good stability for dopamine. In a flow injection analysis, the detection limit of dopamine was estimated to be of the order of 1X10^-7M.

5. CONFLICT OF INTEREST:

Nill.

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Received on 28.04.2018 Accepted on 11.05.2018

Asian J. Pharm. Tech. 2018; 8 (2):83-87 .

DOI: 10.5958/2231-5713.2018.00013.2