A Study of the Operating Conditions for the Sensitivity of Detection of Hydrogen Peroxide for the Carbon Paste Electrode Modified with Ferrocene
Pao-Tsai Kuo(郭寶財), Chung-Min Lien(連崇閔), Hau Lin(林浩)
Department of Chemical and Materials Engineering, Southern Taiwan University
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Abstract
Hydrogen peroxide is widely used in the industry and food preservation, and therefore developing a hydrogen peroxide sensor which can detect the hydrogen peroxide rapidly and conveniently is an important research subject. In recent years, the diabetes has become one of the top ten causes of death for the people in our country. Therefore, developing a glucose biosensor which can detect the glucose rapidly and conveniently is also an important research subject. The glucose and oxygen can be catalyzed by the glucose oxidase to produce the gluconic acid and hydrogen peroxide. A study was conducted to use the ferrocene to modify the carbon paste electrode. Because the ferrocene(Fe(C5H5)2) possesses the excellent catalytic characteristic, it can be used with the graphite carbon powders which possesses the excellent conductivity to make the carbon paste electrode and to elevate the responding current of hydrogen peroxide. The responding current of hydrogen peroxide is detected in the phosphate buffer solution(PBS) and then the concentration of the hydrogen peroxide can be obtained and consequently, the concentration of the glucose can be determined. The TB (Time Base) graphs for different operating potentials, stirring rates, and pH values were plotted to determine the optimum operating conditions. At 30℃, – 0.2V operating potential, 500 rpm stirring rate and in 0.05M PBS buffer solution ( pH = 7.4 ) , when the carbon paste electrode was modified with ferrocene [ferrocene : graphite carbon powder = 3 : 7 ( weight ratio )] , the detection limit was 0.02 mM H2O2 , the linear range was 0.02～1.2 mM H2O2 , R2 = 0.9998 and the sensitivity was 161.43µA/cm2．mM H2O2.
Keywords: Ferrocene, Carbon Paste Electrode, Reaction Parameters, Sensitivity, Hydrogen Peroxide
1 Introduction
Due to an occasional use of hydrogen peroxide in the food industry for the purpose of preservation nowadays, a rapid and convenient sensor for detecting the hydrogen peroxide is an important research subject. In recent years, diabetes has become one of the top ten causes of death for the people in our country. Therefore developing a rapid and convenient glucose biosensor also has become an important research subject. The glucose and oxygen can be catalyzed by the glucose oxidase and the glucose is oxidized to gluconic acid and the oxygen is reduced to hydrogen peroxide. The electrode releases the electrons at the reductive potential and converts the mediator to the reductive state. Then the mediator at the reductive state releases the electrons to reduce the hydrogen peroxide to water and consequently the mediator at the reductive state is converted to the oxidizing state. The responding current for detecting the hydrogen peroxide is used to measure the amount of hydrogen peroxide and consequently determine the concentration of the glucose. Therefore, the detecting technique for the hydrogen peroxide is an important research subject. Because the ferrocene(Fe(C5H5)2) possesses the excellent catalytic characteristic, it can be used with the graphite carbon powders which possesses the excellent conductivity to make the carbon paste electrode and to elevate the responding current of hydrogen peroxide. The responding current of hydrogen peroxide is detected in the phosphate buffer solution(PBS) and then the concentration of the hydrogen peroxide can be obtained and consequently, the concentration of the glucose can be determined.
2 Experimental
2.1 Chemicals and Reagents
Ferrocene ; Hydrochloric Acid (HCl); Sodium Hydroxide (NaOH) ; Hydrogen Peroxide (H2O2); Graphite Carbon Powder( C ); Carbon Paste; Cyclohexanone(C6H10O); Potassium Dihydrogenphosphate (KH2PO4); Potassium Chloride (KCl).
2.2 Equipment
Electrochemical Analyzer (CHI 401A, CH Instruments, Inc) was used to measure the activity of electrode by Cyclic Voltammetry ( CV ) and Time Base ( TB ) mode ; Electric Stirrer(Fargo) ; pH meter (Metrohm 731); Constant Temperature Thermal Bath (Wisdom BC-2DT 10L); Oven (DENG YNG) ; Carbon Paste Electrode was used as the working electrodes, Coiled Platinum Wire was used as the counter electrode and Ag / AgCl was used as the reference electrode.
2.3 Preparation of the carbon paste electrode
Take one section of 7 cm electric wire with 0.05 cm inside diameter. After depriving the coating 0.5 cm length from both ends, the nake-ended wire was washed, dried and ready for use. Then the ferrocene powders, graphite carbon powders and carbon paste were mixed with the appropriate ratio (ferrocene : graphite carbon powders : carbon paste = 0.3 : 0.7 : 1). After the mixing was complete, the mixture was evenly coated on the nake-ended electric wire and dried in the oven and then we obtained the carbon paste electrode.
2.4 Amperometric measurement for the carbon paste electrode
The Carbon Paste Electrode was used as the working electrodes, the Coiled Platinum Wire was used as the counter electrode and the Ag / AgCl was used as the reference electrode. The three electrode system was placed in the 0.05 M PBS buffer solution. The electrochemical analyzer was used to plot the CV diagrams and to detect the responding current of hydrogen peroxide for the Time Base ( TB ) mode.
3 Results and Discussion
3.1 CV diagram of carbon paste electrode
The CV diagrams were plotted for the carbon paste electrode modified with ferrocene (ferrocene : graphite carbon powders : carbon paste = 0.3 : 0.7 : 1) and the unmodified carbon paste electrode. The above mentioned electrode was placed in 0.1 M KCl of 5 mL 0.05 M PBS buffer solution (pH =7.4) and after deoxygenating by purging nitrogen gas, the system was scanned with 50 mV/s scanning rate. Figure 1 shows the CV graphs of (A) carbon paste electrode modified with ferrocene (the range of scanning potential: -0.8～+0.8 V) and (B) unmodified carbon paste electrode(the range of scanning potential: -0.8～+0.8 V). Figure1 shows that the carbon paste electrode modified with ferrocene can have the higher responding current. In order to reduce the interference of oxygen and avoid the inteferring substances ( Ascorbic acid, Uric acid, and Acetaminophen etc.) in human body, the operating potential at –0.2 V was used in this research.
3.2 Determination of the optimum operating potential by TB graphs
Figure 2 shows the TB graphs of carbon paste electrodes for detection of H2O2 at different operating potentials (ferrocene: graphite carbon powders = 3 : 7). Figure 2 shows that when the operating potential = –0.25V, the responding current is the highest. But because at –0.25V operating potential, it might cause the interference of the interfering substances in the human blood, –0.2V operating potential was used in this research.
3.3 Determination of the optimum stirring rate by TB graphs
Figure 3 shows the TB graphs of carbon paste electrodes for detection of H2O2 at different stirring rates (ferrocene : graphite carbon powders = 3 : 7). Figure 3 shows that when the stirring rate = 500 rpm, the responding current is the highest. When the stirring rate was higher than 500 rpm, it might cause the electrode to be unstable and therefore 500 rpm stirring rate was used in this research.
3.4 Determination of the optimum pH value of PBS buffer solution by TB graphs
Figure 4 shows the TB graphs of carbon paste electrodes for detection of H2O2 at different pH values of PBS buffer solution (ferrocene : graphite carbon powders = 3 : 7). Figure 4 shows that when the pH = 8.0, the responding current is the highest. Because the pH of human blood is about 7.4, pH = 7.4 PBS buffer solution was used in this research.
3.5 The study of the TB graphs of carbon paste electrodes for detection limit and linear range of hydrogen peroxide
Figure 5 shows the TB graphs of carbon paste electrodes for detection of the detection limit of H2O2 for pH = 7.4 PBS buffer solution. Figure 6 shows the TB graphs of carbon paste electrodes for detection of the linear range of H2O2 for pH = 7.4 PBS buffer solution . The results showed that the detection limit was 0.02 mM H2O2 , the linear range was 0.02～1.2 mM H2O2, R2=0.9998, and the sensitivity was 161.43 μA/cm2ּmM H2O2 .
4 Conclusions
The results showed that the responding current for the carbon paste electrode modified with ferrocene was elevated significantly. The TB (Time Base) graphs for different operating potentials, stirring rates, and pH values were plotted to determine the optimum operating conditions. The results showed that at –0.2 V operating potential, 500 rpm stirring rate and in 0.05M PBS buffer solution(pH=7.4), the detection limit was 0.02 mM H2O2, the linear range was 0.02~1.2 mM H2O2, R2=0.9998 and the sensitivity was 161.43 μA/cm2ּmMH2O2. This research can be further applied to the glucose biosensor in the future.
Figure 1: CV graphs for (A) carbon paste electrode modified with ferrocene ( the range of scanning potential: -0.8～+0.8 V) (B) unmodified carbon paste electrode( the range of scanning potential: -0.8～+0.8 V); after deoxygenating by purging nitrogen gas for 20 minutes; in 0.1 M KCl of 5 mL 0.05 M PBS buffer solution (pH =7.4) ; scanning rate = 50 mV/s.
Figure 2: The TB graphs of carbon paste electrodes for detection of H2O2 at different operating potentials(ferrocene : graphite carbon powders = 3 : 7); the operating potentials are [ (A) 0V (B) –0.1V (C) –0.15V (D) –0.2V (E) –0.25 V ] ; At 30 ℃, after deoxygenating by purging nitrogen gas for 20 minutes; in 0.1 M KCl of 5 mL 0.05M PBS buffer solution( pH= 7.4 ) ; stirring rate =500 rpm; 10 μL of 100mM H2O2 is injected per 100 seconds.
Figure 3: The TB graphs of carbon paste electrodes for detection of H2O2 at different stirring rates(ferrocene : graphite carbon powders = 3 : 7); the stirring rates are [ (A) 100 rpm (B) 200 rpm (C) 300 rpm (D) 400 rpm (E) 500 rpm ] .
Figure 4: The TB graphs of carbon paste electrodes for detection of H2O2 at different pH values of PBS buffer solution (ferrocene : graphite carbon powders = 3 : 7); the pH values are [ (A) pH = 4.0 (B) pH = 5.0 (C) pH = 6.0 (D) pH = 7.4 (E) pH = 8.0 ] .
Figure 5: The TB graphs of carbon paste electrode electrodes for determining the detection limit of H2O2 (ferrocene: graphite carbon powders = 3 : 7).
Figure 6: The TB graphs of carbon paste electrodes for determining the linear range of H2O2 (ferrocene : graphite carbon powders = 3 : 7); At 30 ℃, the operating potential = –0.2 V; in 0.1 M KCl of 5 mL 0.05 M PBS buffer solution( pH= 7.4 ) ; stirring rate =500 rpm ; 10μL of 100mM H2O2 is injected per 100 seconds.
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