Application of ZnO Nanoparticle/Ion Liquid Modified Carbon Paste Electrode for Determination of Isoproterenol in Pharmaceutical and Biological Samples

Isoproterenol was purchased from Merck. Stock solutions were prepared daily by dissolving appropriate amount distilled water to prepare 1 × 10⁻³ molL⁻¹ and were stored in 4°C until use. Other standard solutions were prepared by dilution of the stock solution with buffer solution. Graphite fine powder (particle size <50 μm) and high viscosity paraffin oil (density = 0.88 g cm⁻³) were purchased from Merck. All other chemicals were purchased in the analytical grade from Merck and Fluka.

Phosphate buffer (sodium dihydrogen phosphate and disodium monohydrogen phosphate plus sodium hydroxide, 0.1 mol L⁻¹) solutions (PBS) with different pH values were used.

1-Methyl-3-butylimidazolium bromide (ion liquid) was synthesized according to the previous paper.²⁶

All Voltammetric measurements were carried out using μ-Autolab modular electrochemical system (Eco Chem. Utrecht, The Netherlands) equipped with PSTA 20 model and driven by NOVA software (Eco Chem.). All the electrochemical studies were performed at 25 ± 1°C. A conventional three electrode cell assembly was employed for the experiments in a 50 mL glass cell containing an Ag/AgCl electrode as reference electrode, a platinum wire as counter electrode and ZnO/ILCPE as working electrode. All of the potentials were measured and reported vs. Ag/AgCl reference electrode. The pH of the solutions was controlled with a Metrohm pH meter (model 827). X-ray powder diffraction studies were carried out using a STOE diffractometer with CuKα radiation (λ = 1.54 Å).

The 100 ml of 0.25 molL⁻¹ zinc nitrate solution was added drop wise to 100 ml of 0.5 molL⁻¹ sodium hydroxide solution. The mixture was stirred by magnetic stirring apparatus (1,500 rpm) at room temperature. Then, the light-green precipitate was separated from solution using buchner funnel. The precipitate was washed with deionized water and ethanol several times. It was dried at 110°C for 15 h, and then calcined at 300°C for 2 h.

To obtain the best conditions in preparation of the modified electrode, the ratios of ZnO/IL/graphite was optimized. Under optimum conditions, the constituents of the paste electrode contain 0.9 g graphite powder, 0.1 g ZnO nanoparticles, 0.2 g ion liquid and 0.8 g paraffin. The mixture was hand mixed in mortar and pestle for 10 min until uniformly wetted paste obtained. The prepared paste was inserted in to a glass tube (internal radius 2.2 mm) with a copper wire placed in to the glass tube for electrical connection. New surface by pushing an excess the paste and polishing it on a weighing paper was obtained. The unmodified carbon paste electrode was prepared in the same way without adding nanoparticles and ion liquid to the mixture.

Figure 1 shows X-ray diffraction pattern of ZnO nanoparticles. Average particles size were obtained 49.5 nm using Scherrer equation D = Kλ/(β cos θ) where K is a constant (0.9), λ is the source wavelength (λ = 1.5418 Å), β is the full width at the half-maximum of the ZnO line (101) and θ is the diffraction angle. In XRD pattern the main peaks were observed at the (100), (002), (101), (102), (110), (103), (200), (112), (201), (004), and (202) planes which very close to ZnO reference patterns. The TEM image of synthesized ZnO, shows dark spots that confirm formation of nanoparticles in our synthesis condition (Figure 2).

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Oxidation of isoproterenol is associated with production of H⁺, so the pH of solution has effect on the oxidation process. The effect of pH solution on the oxidation of IP at ZnO/ILCPE was investigated using cyclic voltammetry. The maximum peak current was appeared in pH 7.0 (PBS), so this value was selected as optimum pH for further experiments (Figure 3). Also, the results show that the peak potential of the redox couple was pH dependent, and the peak potential was shifted to lower potential with increasing the pH.

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Cyclic voltammograms of 400 μmol L⁻¹ of IP at different electrodes were shown in Figure 4. In the presence of ZnO nanoparticles and ion liquid, maximum oxidation current was obtained around 520 mV vs. the Ag/AgCl/KCl_sat (Figure 4a). At ZnO/CPE, ILCPE and unmodified CPE lower oxidation currents were observed (Figures 4b,4c,4d). Figure 4 (inset) shows the current density derived from the cyclic voltammograms of 400 μmol L⁻¹ IP (pH 7.0) at the surface of different electrodes. In addition, at ZnO/ILCPE oxidation of IP was occurred at lower potential. In contrast, modification of CPE with ZnO nanoparticles and ion liquid causes increasing oxidation current and decreasing the oxidation potential (overpotential) that indicated catalytic ability of ZnO/ILCPE.

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In order to study the effect of potential scan rate (ν) on the oxidation current, cyclic voltammograms of IP at various scan rates were recorded at pH 7.0 (Figure 5). Linear relation between peak current and square root of scan rate was obtained where indicates a diffusion controlled oxidation reaction occurs at the surface of ZnO/ILCPE.

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To obtain transfer coefficient (α), linear relationship between peak potential (E_pa) and ln(ν) was used according to equation 1.³¹

Where m = RT/ [(1 – α) n_αF]. The value of α = 0.78 was obtained from this equation.

Chronoamperometric measurements of IP at ZnO/ILCPE were carried out by setting the working electrode potential at 700 mV vs. Ag/AgCl/KCl_sat for the various concentrations of IP in buffer solutions (pH 7.0) (Figure 6). Under diffusion control, a plot of I vs. t^−1/2 will be linear, and the value of D can be obtained from the slope. The slopes of the straight lines for different concentrations of IP were plotted vs. IP concentration and the value of the D was found to be 2.9 × 10⁻⁶ cm² s⁻¹ using the Cottrell equation.³²

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Square wave voltammetry was used for determination of linear dynamic range of the method. The plot of oxidation current vs. concentration was linear in the range of 0.08 to 800.0 μmol L⁻¹ IP (I (μA) = 0.0639 C_IP + 3.3722, R² = 0.992) (Figure 7). The lower detection limit of the method was determined using the equation C_m = 3S_b/m, where S_b is the standard deviation of the blank response (μA) and m is the slope of the calibration curve. Using the data from calibration curve and measurements of blank, the lower detection limit was obtained to be 0.04 μmol L⁻¹ IP. In order to examine the reproducibility of preparation of modified electrode, three electrodes were prepared according to proposed method. Then, they were used for determination of IP in the solution containing 50 μmol L⁻¹ IP. The RSD values were found to be 1.1% and 1.5% for E_pa and I, respectively.

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To study the effect of potential interferences, the influence of various foreign species on the signal of 10 μmol L⁻¹ IP were investigated at ZnO/ILCPE. The tolerance limit was taken as the maximum concentration of foreign substances which caused no more than ±5% relative error in the determination of IP. The results showed that 1000 fold of glucose, fructose, lactose, sucrose, Na⁺, K⁺, CO₃²⁻, SCN⁻, F⁻, Br⁻, Ca²⁺, SO₄²⁻, 800 fold aniline, phenylalanine, methionine, valine, glycine, histidine and tryptophan, and saturated starch solution did not effect on IP oxidation current.

With the aim of evaluation of proposed method for analysis of real samples, the concentration of IP was measurement in a pharmaceutical sample) Injection solution (and two biological samples. The results presented in Table I showed that ZnO/ILCPE has retained its efficiency for the determination of IP in real samples with satisfactory results.