Novel Poly(pyrrole-co-3-acetyl pyrrole)-WO3 nanocomposites modified gold electrode as electrocatalytic oxidation and reduction of H2O2

Hydrogen peroxide have strong oxidizing property hence widely used as oxidizing agent in various food production, organic compound synthesis, pulp & paper bleaching, sterilization, clinical application, pharmaceutical and environmental analysis [1-4]. H2O2 is by product of the most of the oxidative biological reactions which is most important factor of diseases like cancer, asthma, neurodegenerative disorder heart diseases [5-8] etc. Hence the detection and quantification of H2O2 was based on a simple credible, precise, fast & economical. Several methods have been developed for qualitative and quantitative detection of H2O2 such as titration [9], spectrophotometry [10], chemilumination [11], fluorometric [12] and chromatographic [13] techniques. But most of the methods have disadvantage like high cost, time consuming, and complexity while electrochemical methods have preferably low cost, high efficiency, sensitivity, selectivity [14] and reproducibility of electrode in operation. Recent studies on various polymer nanocomposites were used as electrochemical biosensor [15], drugs sensor [16,17], an environmental pollutant sensor [18], water and soil sample analysis [19], pharmaceutical and human fluids [20]. Several nanocomposites based modified electrode such as Pt Nanoparticle-Decorated rGO–CNT Nanocomposite [21], poly(azureA)-platinum nanoparticles [22], Co-embedded N-doped hierarchical carbon [23], AgAu / RGO / TiO2 nanocomposite [24], Co3O4 nanowall [25], novel metals [26-28] and conducting polymer nanocomposites [29,30] have been used for direct oxidation, reduction and detection of H2O2. In the present study chemically synthesized novel PPAP-WO3 nanocomposites (NCPs) for the electrochemical detection of H2O2 has newer approach. In the present observation we have adapted electrochemical detection of H2O2 by cyclic voltammetry (CV), differential pulse voltammetry (DPV) and square wave voltammetry (SWV) techniques. The fabrication of an electrochemical sensor based on novel poly(pyrrole-co-3acetyl pyrrole)-WO3 nanocomposites modified gold electrode (PPAP-WO3-AuE) and its electrocatalytic oxidation and reduction of hydrogen peroxide is described here. The PPAP-WO3 nanocomposites were synthesized by chemical method and characterized by different techniques. The WO3 nanoparticles incorporated with PPAP were confirmed by x-ray diffraction pattern, scanning electron microscopy and transmission electron microscope micrograph. The electrochemical behaviour of PPAP-WO3-AuE towards the electro catalytic oxidation and reduction of hydrogen peroxide was investigated by cyclic voltammetry, differential pulse voltammetry and square wave voltammetry. The observed DPVs and SWVs response depend linearly on concentration of hydrogen peroxide in the range of 1-10 mM and with limit of detection (LOD) is 1×10 M. The correlation coefficients were found as 0.991, 0.930 and sensitivity observed was 47.64 A/mM.cm and 8.31A/mM.cm. These results indicate the PPAP-WO3-AuE exhibited good platform and could be used for electrochemical determination of hydrogen peroxide.


Introduction
Hydrogen peroxide have strong oxidizing property hence widely used as oxidizing agent in various food production, organic compound synthesis, pulp & paper bleaching, sterilization, clinical application, pharmaceutical and environmental analysis [1][2][3][4]. H2O2 is by product of the most of the oxidative biological reactions which is most important factor of diseases like cancer, asthma, neurodegenerative disorder heart diseases [5][6][7][8] etc. Hence the detection and quantification of H2O2 was based on a simple credible, precise, fast & economical. Several methods have been developed for qualitative and quantitative detection of H2O2 such as titration [9], spectrophotometry [10], chemilumination [11], fluorometric [12] and chromatographic [13] techniques. But most of the methods have disadvantage like high cost, time consuming, and complexity while electrochemical methods have preferably low cost, high efficiency, sensitivity, selectivity [14] and reproducibility of electrode in operation.
The fabrication of an electrochemical sensor based on novel poly(pyrrole-co-3acetyl pyrrole)-WO3 nanocomposites modified gold electrode (PPAP-WO3-AuE) and its electrocatalytic oxidation and reduction of hydrogen peroxide is described here. The PPAP-WO3 nanocomposites were synthesized by chemical method and characterized by different techniques. The WO3 nanoparticles incorporated with PPAP were confirmed by x-ray diffraction pattern, scanning electron microscopy and transmission electron microscope micrograph. The electrochemical behaviour of PPAP-WO3-AuE towards the electro catalytic oxidation and reduction of hydrogen peroxide was investigated by cyclic voltammetry, differential pulse voltammetry and square wave voltammetry. The observed DPVs and SWVs response depend linearly on concentration of hydrogen peroxide in the range of 1-10 mM and with limit of detection (LOD) is 1×10 -4 M. The correlation coefficients were found as 0.991, 0.930 and sensitivity observed was 47.64 A/mM.cm 2 and 8.31A/mM.cm 2 . These results indicate the PPAP-WO3-AuE exhibited good platform and could be used for electrochemical determination of hydrogen peroxide.

Synthesis of novel Poly(Pyrrole-co-3 acetyl Pyrrole) (PPAP)-WO3 Nanocomposites
The monomer pyrrole (3 mmol) and 3-acetyl pyrrole (3 mmol) were dissolved separately in 5 ml chloroform mixed in beaker and stirred for 30 min at room temperature. The doping of pre synthesized WO3 NPs in monomer mixture solution was done by addition of 500 mg WO3 nanoparticles in mixture of pre-stirred pyrrole and 3-acetyl pyrrole solution. The monomer mixture solution was stirred for 30 min at room temperature. Then tri chloro acetic acid (TCA) (13 mmol) in 2 ml chloroform was added drop wise to the monomer solution at room temperature to start polymerization. Trichloro acetic acid was added to the monomer and nanoparticles mixture solution, the colour changed immediately from yellowish to dark green. The polymerization was carried out for 4 hrs at room temperature after which precipitation was formed in the reaction flask. After polymerization resultant residue was washed well with water and acetone. The precipitates were stirred with 3 W% NaOH in aqueous solution for 30 min in order to remove acid catalyst. The solution was stirred with 2N HCl solution for 30 min to decrease the pH and make them acidic. The obtained green precipitate of novel PPAP-WO3 nanocomposites was dried for 24 hrs at room temperature.

Preparation of modified electrode
The gold (Au) electrode (2 mm diameter) was carefully polished using a polishing cloth with alumina slurry and then rinsed thoroughly with glass distilled water. The Au electrode was placed in ultrasound cleaner for 5 min, rinsed again with glass distilled water and allowed to dry at RT. To prepare the Au electrode modified with PPAP-WO3 nanocomposites, an alcoholic solution of 0.1 % Nafion dispersion of PPAP-WO3 NCPs (1mg mL -1 ) was prepared and the suspension (5L) was cast on to the surface of pretreated Au electrode. The solvent was allowed to evaporate at RT which resulted in immobilized PPAP-WO3 NCPs material on the Au electrode surface.

Characterization
The prepared materials of PPAP-WO3 NCPs were characterized by Fourier Transfer Infrared spectroscopy, x-ray diffraction, Scanning Electron Microscopy and Transmission Electron Microscopy techniques. The Infrared spectrum was recorded on Fourier Transfer Infrared spectrophotometer [JASCO, FT-IR/4100] Japan using dry KBr as standard reference in the range of 400-4000 cm -1 . The X-ray powder diffraction patterns of PPAP-WO3 NCPs were recorded on Bruker 8D advance X-ray diffractometer using Cu Kα radiation of wavelength = 1.54056 Å. To study the morphology PPAP-WO3 NCPs were examined using Scanning Electron Microscope. The Scanning Electron Microscopy analysis was carried out with JEOL; JSM-6330 LA operated at 20.0kV and 1.0000nA. Shape, size & morphology was calculated by Transmission Electron Microscopy analysis carried out on Philips model CM200 operated at 200kV.

Electrochemical analysis
Cyclic voltammetry (CV), differential pulse voltammetry (DPV) and square wave voltammetry (SWV) measurements were performed on a Metrohm Autolab PGSTAT128N (Metrohm B.V., Utrecht, Netherlands) having a single compartment with three electrode system containing, a saturated Ag/AgCl reference electrode, a platinum wire auxiliary electrode and a modified PPAP-WO3-Au electrode as working electrode. Hydrogen peroxide measurements were carried out at pH 7.4 in 0.1 M potassium phosphate buffer solution (PBS) at room temperature. The CV, DPV and SWV were done at different concentration of hydrogen peroxide in the range 1-10 mM. For CV measurements, the potential scan was taken from -0.90 to 1.10 V at the different scan rate in the range 25-150 mVs -1 .

Fourier transfer infrared spectroscopy
The PPAP-WO3 NCPs powder prepared were analysed by Fourier Transfer Infrared spectrophotometer in range 400-4000 cm -1 in Fig. 1. The peak at 3402 cm -1 was attributed to the N-H bond while the peak at 1624 cm -1 attributed to the C-N-C bond or the C=O group form during the PPAP polymerization. The strong peak at 2872 cm -1 and 2824 cm -1 were related to the aliphatic C-H vibration. On the other hand, the peaks at 1240 cm -1 and 996 cm -1 were due to the C-N and C-H groups in the composite spectrum respectively. In addition, the peak at 1520 cm -1 was characteristic of a typical PPAP ring vibration. The Fourier Transfer Infra-red spectrum of the PPAP-WO3 NCPs powder shows characteristic bipolaron bands at 1240 cm −1 and broad band at 1520 cm −1 , indicating the formation of PPAP in its doped state [31]. The peak at 1715 cm -1 to the C=O functional group presence in indicates that the copolymerization occurred in pyrrole and 3-acetyl pyrrole. The carbonyl group frequency peak were described at 1703 cm -1 in pure PPAP. After doping of WO3 NPs carbonyl group frequency shifted to 1715 cm -1 which indicate doping of WO3 NPs. In Fourier Transfer Infra-red spectrum of PPAP-WO3 NCPs sharp new peaks appears at 924 cm -1 , 8320 cm -1 , 668 cm -1 and 619 cm -1 which is the characteristic peak of WO3 [32] indicating the presence of WO3 in PPAP.

X-ray diffraction
The x-ray diffraction pattern of PPAP-WO3 NCPs is shown in Fig. 2. The x-ray diffraction analysis data of PPAP-WO3 nanocomposites is described in Table S1 Fig. 2. The intercalation of WO3 nanoparticles on PPAP leads to the formation to broadening peak and also decreases in peak intensity than pure WO3 nanoparticles which indicate the formation of PPAP-WO3 crystalline nanocomposites. The average particle size was calculated by using Debye-Scherrer formula. The PPAP-WO3 NCPs showed 2 and full width of half maximum (FWHM) which are values shown in Table S1. The average crystalline size of PPAP-WO3 NCPs was found to be 13.60 nm.

Scanning electron microscopy & energy dispersive spectroscopy
The morphology of PPAP-WO3 nanocomposites studied by Scanning Electron Microscope shows the micrograph of PPAP-WO3 nanocomposites that are having irregular shape and their distribution is not uniform showed in Fig. 3(a). WO3 nanoparticles showed agglomeration on the PPAP composites, which formed porous PPAP-WO3 nanocomposites. The elemental composition of prepared PPAP-WO3 nanocomposites was analysed by qualitatively and quantitatively by Energy Dispersive Spectroscopy. The elemental composition of PPAP-WO3 nanocomposites by atomic % (weight %) from Fig. 3(b)

Transmission electron microscope
The Transmission electron micrograph Fig. 4(a) of PPAP-WO3 nanocomposites showed that WO3 nanoparticles deposited on the polypyrrole and their size was found to be 30-50 nm. The selected area electron diffraction pattern inset of Fig. 4(a), PPAP-WO3 Nanocomposites was found to be spotty ring which indicate the crystalline nature of nanocomposite. Fig. 4(c) shows a histogram of PPAP-WO3 Nanocomposites, the average diameter (daverage) and standard deviation () was found to be 36.46 nm and 13.98 respectively.

Electrochemical behavior of PPAP-WO3-Au electrode
The electrochemical behavior of bare AuE and PPAP-WO3-AuE was studied by cyclic voltammetry of 0.1 mM potassium ferricyanide in 0.1 M KNO3 at the scan rate 100 mV/s. The cyclic voltammograms show that the PPAP-WO3-AuE showed higher currents in 0.1 mM ferricyanide compared to the bare AuE. The increase peak currents are largely the result of increased electroactive surface area of the PPAP-WO3-AuE. Using diffusion coefficient 6.8 × 10 -6 cm 2 /s [33] and Randeles Sevick equation it was seen that the electroactive surface area for modified PPAP-WO3-AuE was found to be 4.5 times higher than that of bare electrode (Fig. 5(a)).
The effect of scan rate on the oxidation and reduction current on PPAP-WO3-AuE was investigated by cyclic voltammetry of 0.1 mM ferricyanide in 0.1 M KNO3 at different scan rate in 25-150 mV/s in the potential range -0.2V to 0.9V (Fig. 5(b)). As shown in plot (Fig. 5(c)) from linear regression equation, the regression coefficient of ferricyanide oxidation and reduction was with R 2 = 0.983 and R 2 = 0.980 respectively.   oxidation peak current with dramatically increase in the reduction peak current increase with concentration of H2O2. As shown in plotted Fig. 6(b), from linear regression equation, the regression coefficient of oxidation and reduction was with R 2 = 0.988 and R 2 = 0.999 respectively. The sensitivity of electrode in oxidation and reduction peak was found to be 152.03 A/mMcm 2 and 374 A/mMcm 2 respectively. -0.

Electrochemical detection of hydrogen peroxide by differential pulse voltammetry
The voltammetric sensor using differential pulse voltammetry (DPV) was developed for the detection of hydrogen peroxide. Fig. 8(a) shows the obtained differential pulse voltammograms at PPAPy-WO3-AuE for different concentration of hydrogen peroxide. The concentration was varied from 1 mM to 10 mM in 0.1 PBS solutions at 7.4 pH. The oxidation peak current obtained was directly proportional to the concentration of hydrogen peroxide was plotted as shown in Fig. 8(b)

Conclusion
In the present study we successfully synthesized of novel PPAP-WO3 nanocomposites by chemical method. The PPAP-WO3 nanocomposites were crystalline, porous materials, WO3 nanoparticles incorporated with PPAP and average size 30-50nm resulted from TEM analysis. The histogram was showed average size daverage and standard deviation () was found to be 36.46 nm and 13.98 respectively. The PPAP-WO3 NCPs modified gold electrode was excellent used for detection of hydrogen peroxide. The CV, DPV and SWV curves resulted increase in peak current with increasing the concentration of hydrogen peroxide. The result obtained from CVs, the regression coefficient of oxidation and reduction was with R 2 = 0.988 and R 2 = 0.999 and sensitivity of electrode in oxidation and reduction peak was found to be 152.03 A/mMcm 2 and 374 A/mMcm 2 respectively. The observed DPVs and SWVs response depend linearly on concentration of hydrogen peroxide in the range of 1-10 mM and with limit of detection (LOD) is 1×10 -4 M.
The correlation coefficients were found as 0.991, 0.930 and sensitivity observed was 47.64 A/mM.cm 2 and 8.31A/mM.cm 2 . These results indicate the PPAP-WO3-AuE exhibited simple, effective tool and good platform could be used for electrochemical determination of hydrogen peroxide.