Application of the Screen Printed Planar Electrode Modified with Ruthenium Hexacyanoferrate to Glucose Biosensor Kuo-Hsiang Liao( 廖國翔 ), Chung-Min Lien( 連崇閔 ), Hau Lin( 林浩 ) Department of Chemical and Materials Engineering, Southern Taiwan University The glucose and oxygen can be catalyzed by the glucose oxidase to produce the gluconic acid and hydrogen peroxide. Because the ruthenium hexacyanoferrate( Ⅱ ) possesses the excellent conductivity and catalytic characteristic, it can be used to elevate the responding current for detection of reduction of hydrogen peroxide. A study was conducted to use the ruthenium hexacyanoferrate( Ⅱ ) to modify the screen printed planar electrode which was used as the working electrode to detect the responding current of reduction of hydrogen peroxide in the PBS buffer solution( pH = 7.4 )and the sensitivity of detection of hydrogen peroxide was determined from the responding current and consequently, the concentration of the glucose could be determined. The results showed that the responding current for the carbon paste electrode modified with the ruthenium hexacyanoferrate( Ⅱ ) was elevated significantly. At 30 ℃, -0.2V operating potential, and in 0.05 M PBS buffer solution( pH = 7.4 ), when the screen printed planar electrode was modified with the ruthenium hexacyanoferrate( Ⅱ ) [ruthenium hexacyanoferrate( Ⅱ ) : graphite carbon powders = 3 : 7 ( weight ratio )], the detection limit was 0.02 mM H 2 O 2, the linear range was 0.02 ～ 0.96 mM H 2 O 2, R 2 =0.917, and the sensitivity was 28.77μA/cm 2 ּmM H 2 O 2. For the screen printed planar glucose sensor, the detection limit was 0.02 mM C 6 H 12 O 6, the linear range was 0.02~2.24 mM C 6 H 12 O 6 (R 2 =0.9983), and the sensitivity was µA/cm 2 ． mM C 6 H 12 O 6. Introduction : At present, there is no effective method and medicine to cure the diabetes. The concentration of the blood sugar for diabetic can only be controlled by insulin. The traditional routine examination must be performed by the medical examiners in the hospitals and the examination usually requires using the expensive instruments. Also this kind of examination requires high expense and professional manipulation. Therefore, developing a rapid, convenient, and economical glucose biosensor for detecting the glucose is an important research subject. Abstract ： CONCLUSIONS The results showed that the responding current for the carbon paste electrode modified with the ruthenium hexacyanoferrate( Ⅱ ) was elevated significantly. The optimum weight ratio for ruthenium hexacyanoferrate ( Ⅱ ) : graphite carbon powders was 3 : 7. At 30 ℃, -0.2V operating potential, and in 0.05 M PBS buffer solution( pH = 7.4 ), when the screen printed planar electrode was modified with the ruthenium hexacyanoferrate( Ⅱ ) [ruthenium hexacyanoferrate( Ⅱ ) : graphite carbon powders = 3 : 7 ( weight ratio )], the detection limit was 0.02 mM H 2 O 2, the linear range was 0.02 ～ 0.96 mM H 2 O 2, R 2 =0.917, and the sensitivity was μA/cm 2 ּmM H 2 O 2. For the glucose screen printed planar sensor, the detection limit was 0.02 mM C 6 H 12 O 6 ; the linear range was 0.02 ~ 2.24 mM C 6 H 12 O 6 ; R 2 = and the sensitivity was μA/cm 2 mM C 6 H 12 O 6. REFERENCES 1. S. J. Updike and G. P. Hicks, “The Enzyme Electrode,” Nature, 214, 986, (1967). 2. R. Garjonyte and A. Malinauskas, “Glucose Biosensor Based on Glucose Oxidase Immobilized in Electropolymerized Polypyrrole and Poly(o-phenylenediamine) Films on a Prussian Blue-Modified Electrode,” Sensors and Actuators B, 63, 122, (2000). Fig. 2 TB graphs of the carbon paste electrodes with different ratios of ruthenium hexacyanoferrate( Ⅱ ) to graphite carbon powders; the ruthenium hexacyanoferrate( Ⅱ ) : graphite carbon powders were 〔 (A) 0:10(unmodified carbon paste electrode) (B) 1:9 (C) 2:8 (D) 3:7 (E) 4:6 (F) 5:5 〕 Results and Discussion ： Fig. 3 The TB graphs of screen printed planar electrodes for detection of H 2 O 2 (ruthenium hexacyanoferrate( Ⅱ ): 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 =600 rpm; 1μL, 2μL, 4μL, 8μL and 16μL of 100mM H 2 O 2 are injected at 100 seconds, 200 seconds, 300 seconds, 400 seconds and 500 seconds respectively Fig. 5 The TB graphs of screen printed planar electrode for detection of glucose (ruthenium hexacyanoferrate( Ⅱ ) : 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 =600 rpm; 1μL, 2μL, 4μL, 8μL and 16μL of 100mM glucose are injected at 100 seconds, 200 seconds, 300 seconds, 400 seconds,and 500 seconds respectively Fig. 4 The TB graphs of screen printed planar electrodes for detection of H 2 O 2 (ruthenium hexacyanoferrate( Ⅱ ) : 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 =600 rpm; 16μL of 100mM H 2 O 2 is injected per 100 seconds Fig. 6 The TB graphs of screen printed planar electrode for detection of glucose (ruthenium hexacyanoferrate( Ⅱ ) : 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=600 rpm; 16μL of 100mM glucose is injected per 100 seconds 7 cm 0.5 cm 0.05 cm Chemicals and Reagents Ruthenium （Ⅲ） Chloride Hydrate(RuCl 3 ); Potassium Hexacyanoferrate(II)(K 4 [Fe(CN) 6 ]. 3H 2 O) ;Hydrogen Peroxide (H 2 O 2 ); Glucose Oxidase(EC , Type X-S: From Aspergillus Niger, units/mg); Graphite Carbon Powder; Carbon Paste; Cyclohexanone(C 6 H 10 O); Nafion; Potassium Dihydrogenphosphate(KH 2 PO 4 ); Potassium Chloride (KCl); D(+)-Glucose Monohydrate(C 6 H 12 O 6 ). Equipment Electrochemical Analyzer (CHI 614A, CH Instruments, Inc) was used to measure the activity of electrode by Cyclic Voltammetry ( CV ) and Time Base ( TB ) mode ; pH meter (Metrohm 731); Constant Temperature Thermal Bath (Wisdom BC-2DT 10L); Oven (DENG YNG) ; Electric Stirrer (Fargo); Carbon Paste Electrode and Screen Printed Planar Electrode were used as the working electrodes, Coiled Platinum Wire was used as the counter electrode and Ag / AgCl was used as the reference electrode. 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 ruthenium hexacyanoferrate( Ⅱ ) powders, graphite carbon powders and carbon paste were mixed with the appropriate ratio (ruthenium hexacyanoferrate( Ⅱ ) : 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. Fig 1. CV graphs for (A) carbon paste electrode modified with ruthenium hexacyanoferrate( Ⅱ ) ( the range of scanning potential: -0.8 ～ +0.8 V) (B) unmodified carbon paste electrode( the range of scanning potential: -0.6 ～ +0.6 V) 1 2μL glucose oxidase solution was put onto the electrode 2 5μL of 1% Nafion solution was dropped onto the electrode evenly ( ) Preparation of the Screen Printed Planar Electrode : The above mentioned mixture with the appropriate ratio (ruthenium hexacyanoferrate( Ⅱ ) : graphite carbon powders : carbon paste = 0.3 : 0.7 : 1) was evenly coated on the front side of screen plate and then the PE paper was placed under the screen plate and the plastic plate was used to print the mixture on the PE paper evenly. The electrode was dried in the oven and then we obtained the screen printed planar hydrogen peroxide electrode. After the screen printed planar hydrogen peroxide electrode was dried, the 2μL glucose oxidase solution( 3mg of glucose oxidase was dissolved in 500μL PBS buffer solution ) was put onto the screen printed planar hydrogen peroxide electrode and the electrode was dried at room temperature. Then 5μL of 1% Nafion solution(in 95% alcohol) was dropped onto the electrode evenly and after the electrode was dried at room temperature, we obtained the screen printed planar glucose electrode.