Electrochemical oxidation of provitamin B5, d-panthenol and its analysis in spiked human urine
© Nayak and Shetti. 2016
Received: 5 November 2015
Accepted: 3 May 2016
Published: 11 May 2016
The behavior of biomolecules and the advancements in the electrochemical techniques play a tremendous role in the development of voltammetric sensors. Redox reactions of biologically active molecules can be studied by using different voltammetry techniques which guide us to understand the metabolic fact of the targeted drug. In the present work, we describe the electrochemical oxidation of d-panthenol (DP) by using a versatile glassy carbon electrode (GCE).
Experimental section was carried out by using cyclic voltammetry and square wave voltammetry.
Under the optimized conditions (pH 4.2), the square wave voltammetric peak current of d-panthenol increased linearly with its concentration. The detection limit was found to be 5.0 × 10−7 M. The number of protons and electrons involved in the oxidation process were calculated. The heterogeneous rate constant was found to be (3.67 × 103 s−1).
The method detects the trace level of the analyte with low detection limit which imparts the development of a sensor for selective and sensitive determination of d-panthenol. This sensor was successfully applied to determine the d-panthenol in spiked urine samples.
Voltammetric methods gathered much attention of researchers owing to their rapidity of analysis, no requirement of sample pretreatment, fairly high sensitivity, and inexpensive instrumentation. In addition, application of electroanalytical techniques includes the determination of electrode mechanisms [Malode et al. 2012; Bukkitgar et al. 2015]. The performance of the voltammetric techniques is purely affected by the characteristic feature of the working electrode material such as chemical and physical properties. An overview of the improvement in electroanalytical chemistry demonstrates that solid metal electrodes represent the most rapidly rising class of electrodes [Shetti et al. 2009; Jorge et al. 2010; Wudarska et al. 2013]. Among all those electrodes, the utilization of glassy carbon electrode for the electrochemical measurements has increased in recent years as they provide good sensitivity, negligible porosity, and superior mechanical rigidity [Nayak and Shetti 2016]. Oxidation and reduction property of drugs can give insight into its metabolic providence or their in vivo redox process or pharmaceutical activity [Kumar et al. 2008; Diculescu et al. 2006].
Panthenol has been determined by chromatographic methods [Bui-Nguyen 1984; Kulikov and Zinchenko 2007; Prosser and Shreppad 1969; Nagamallika and Arunadevi 2013]: thin-layer chromatographic determination of panthenol with spectro-fluorimetric detection [Shehat Mostafa et al. 2004], fluorimetry, and colorimetric methods for the detection of panthenol [Shehat Mostafa et al. 2002].
A review of the literature exposes that, till date, there is only one report on the electrochemical behavior of DP. In earlier work [Wang and Tseng 2001], authors used carbon paste electrode (CPE), cobalt oxide-modified carbon paste electrode (CoO/CPE) and cadmium oxide-modified carbon paste electrode (CdO/CPE). Looking at the oxidation mechanism and the linearity range, we have undertaken this work at glassy carbon electrode. The pharmaceutical and cosmological importance of d-panthenol necessitates a sensitive method to be developed. The plan of the present study is to establish the suitable experimental conditions to explore the oxidation mechanism of d-panthenol and its determination in spiked human urine samples using cyclic, square wave voltammetric techniques.
Apparatus and chemicals
Electrochemical analyzer (CHI Company, D630, USA) was used to study the electrochemical activities of the drug under investigation at an ambient temperature of 25 ± 0.1 °C. A three-electrode system consisting of GCE as working electrode, platinum wire as counter electrode, and Ag/AgCl (3 M KCl) as reference electrode were used in a 10-ml single compartment. d-panthenol (Sigma-Aldrich, USA) was used to prepare 1.0 mM stock solution in double-distilled water. The phosphate buffer solutions ranging 3.0–11.2 pH with ionic strength 0.2 M were prepared according to literature [Christian and Purdy 1962; Bukkitgar and Shetti 2015], and pH of the solutions was measured by pH meter (Elico Ltd., LI120, India). Double-distilled water, analytical-grade chemicals, and reagents without further purification were used throughout the experiments.
Pretreatment of electrode
Prior to use, the GCE was carefully polished using a 0.3-μm Al2O3 slurry on a polishing cloth before each experiment. The GCE was first activated in phosphate buffer (pH 4.2) by cyclic voltammetric sweeps between −2.0 and 3.0 V until stable cyclic voltammograms were obtained. Then, electrodes were transferred into another 10 ml of phosphate buffer (pH 4.2) containing proper amount of DP.
In Eq. (1), I p refers to the anodic peak current and n is the number of electrons transferred during the electrode reaction = 1. A 0 is the surface area of the electrode, D R is the diffusion coefficient, i.e., 7.6 × 10−6 cm2 s−1 [Adams 1969; Gosser 1994], υ is the scan rate, and C 0 is the concentration of K3Fe (CN)6. From the slope of the plot of I p versus υ 1/2, the area of the electrode surface was calculated to be 0.04 cm2.
Preparation of spiked human urine sample
Human urine was obtained from four healthy volunteers of similar sex and age. Aliquots were centrifuged at 7000 rpm for 5 min at room temperature (25 ± 0.1 °C). These urine samples were analyzed immediately or they were stored at low temperature until analysis.
Results and discussion
Voltammetric behavior of d-panthenol
Effect of solution pH
E P = 0.062 pH + 0.657; R 2 = 0.891 (pH = 3.0–5.0)
E P = 0.020 pH + 0.474; R 2 = 0.843 (pH = 5.0–8.0)
E P = 0.049 pH + 0.686; R 2 = 0.990 (pH = 7.0–9.2)
The slope of E P versus pH being close to the theoretical value 0.059 suggested that the number of electrons transferred is equal to that of hydrogen ions taking part in the electrode reaction [Bukkitgar et al. 2016]. From the plot of I P versus pH (Fig. 2b), it is clear that the best result with respect to sensitivity and sharper response was obtained at pH = 4.2, hence it was selected for further work. The peak current depends on the deprotonation and protonation form of the electro active species in electrochemical cell. Further, the magnitude of current is directly proportional to rate of the electrochemical reaction. Hence, it is apparently concluded that the oxidation of DP is very high at pH 4.2.
Scan rate variation
I P = 50.06 υ + 1.238; R 2 = 0.980
From the plot of logarithm of anodic peak current versus logarithm of scan rate, a straight line was obtained with a slope of 0.54 (Fig. 3b) which is closer to the theoretical value of 0.5 for a purely diffusion-controlled process [Gosser 1994; Yadav et al. 2014]. The dependence log I P versus log υ is used as criterion to determine whether the reaction is reversible or irreversible, based on the shift in the peak. In our study, the forward peak was shifted, indicating an irreversible electrode process. Gosser also quoted that in an irreversible system, both the electrode kinetics and chemical kinetics are slow [Demir et al. 2014, 1997].
log I P = 0.545 log υ + 1.310; R 2 = 0.986
Characteristics of d-panthenol calibration plot using square wave voltammetry at glassy carbon electrode
Analytical parameters Values
Linearity range (M)
1.0 × 10−3–8.0 × 10−6
Slope of the calibration plot (μA M−1)
Correlation coefficient (r)
RSD of slope (%)
RSD of intercept (%)
Number of data points
5.0 × 10−7
17.0 × 10−7
Repeatability (RSD %)
Reproducibility (RSD %)
Comparison of linearity range and detection limits of d-panthenol by different reported methods
Linearity range (μg/mL)
Reversed phase HPLC method
Kulikov and Zinchenko 2007
Shehat Mostafa et al. 2004
Shehat Mostafa et al. 2002
Differential pulse voltammetry
Wang and Tseng 2001
Square wave voltammetry
Effect of interfering substances
Influence of potential interferents on the voltammetric response of 1.0 × 10−4 M d-panthenol
Signal change (%)
Detection of DP in spiked urine samples
Application of square wave voltammetry for the determination of d-panthenol in spiked human urine
Urine sample 1
0.1 × 10−4
0.099 × 10−4
Urine sample 2
0.2 × 10−4
0.197 × 10−4
Urine sample 3
0.5 × 10−4
0.485 × 10−4
The developed voltammetric method provides a simple and quick tool for the direct determination of provitamin B5 using glassy carbon electrode. The investigation clearly reveals that the electrochemical oxidation of d-panthenol was found to be irreversible and diffusion-controlled, and it involves a two-electron two-proton electrode mechanism. From the results obtained, a probable electrochemical mechanism was proposed. A square wave voltammetric technique was developed for the determination of d-panthenol in human urine samples quantitatively. As compared to other methods, the proposed method offered an improvement in simplicity and accuracy.
One of the authors, Deepti S. Nayak, expresses thanks to the Department of Science and Technology, Government of India, New Delhi, for the award of Inspire Fellowship in Science and Technology
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