The biosensor based on the pyruvate oxidase modified conducting polymer for phosphate ions determinations.

An enzymatic biosensor was fabricated by the covalent immobilization of pyruvate oxidase (PyO) onto the nano-particle comprised poly-5,2':5',2''-terthiophene-3'-carboxylic acid, poly-TTCA (nano-CP) layers on a glassy carbon electrode (GCE) for the amperometric detection of the phosphate ions. The direct electron transfer reaction of the immobilized PyO onto the nano-CP layers was investigated and the electron transfer rate constant was determined to be 0.65 s(-1). The electrochemically prepared nano-CP lowered the oxidation potential (+0.40 V versus Ag/AgCl) of an enzymatically generated H(2)O(2) by PyO in a phosphate solution. Experimental parameters affecting the sensitivity of the biosensors, such as amounts of the cofactors, the pH, the applied potential, and the temperature were optimized. A linear response for the detection of the phosphate ion was observed between 1.0 microM and 100 microM and the detection limit was determined to be about 0.3 microM. The response time of the biosensors was about 6s. The biosensor showed good selectivity towards other interfering anions. The long-term storage stability of the phosphate biosensor was studied and the sensor was applied in a human serum sample for the phosphate ions detection.

[1]  Y. Shim,et al.  The potential use of hydrazine as an alternative to peroxidase in a biosensor: comparison between hydrazine and HRP-based glucose sensors. , 2005, Biosensors & bioelectronics.

[2]  H. D. Stensel,et al.  Design and retrofit of wastewater treatment plants for biological nutrient removal , 1992 .

[3]  Y. Shim,et al.  Characterization of an EDTA bonded conducting polymer modified electrode: its application for the simultaneous determination of heavy metal ions. , 2003, Analytical chemistry.

[4]  Yoon-Bo Shim,et al.  Direct electrochemistry of horseradish peroxidase bonded on a conducting polymer modified glassy carbon electrode. , 2003, Biosensors & bioelectronics.

[5]  B. T. G. Tan,et al.  Novel heteroarylene polyazomethines: their syntheses and characterizations , 1998 .

[6]  Y. Shim,et al.  Simple preparation of terthiophene-3′-carboxylic acid and characterization of its polymer , 2002 .

[7]  J. Luong,et al.  An FIA biosensor system for the determination of phosphate. , 1991, Biosensors & bioelectronics.

[8]  Xiao Dong Chen,et al.  Electrochemical Immobilization of Enzymes on Graphite Electrode , 1998 .

[9]  N. Chaniotakis,et al.  Phosphate biosensor based on polyelectrolyte-stabilized pyruvate oxidase , 2001 .

[10]  J. Ogawa,et al.  Determination of inorganic phosphate by flow injection method with immobilized enzymes and chemiluminescence detection. , 1989, Analytical biochemistry.

[11]  H. Endo,et al.  Determination of phosphate ions with an enzyme sensor system. , 1987, Biosensors.

[12]  Izumi Kubo.  Potentiometric phosphate-sensing system utilizing phosphate-binding protein , 2002, Analytical and bioanalytical chemistry.

[13]  Serge Cosnier,et al.  A Bienzyme Electrode (Alkaline Phosphatase−Polyphenol Oxidase) for the Amperometric Determination of Phosphate , 1998 .

[14]  Itamar Willner,et al.  Biomaterials for Sensors, Fuel Cells, and Circuitry , 2002, Science.

[15]  Philip N. Bartlett,et al.  Electrochemical immobilisation of enzymes: Part I. Theory , 1987 .

[16]  S. Daunert,et al.  Phosphate binding protein as the biorecognition element in a biosensor for phosphate. , 2004, Sensors and actuators. B, Chemical.

[17]  Christopher R. Lowe,et al.  Enzyme entrapment in electrically conducting polymers. Immobilisation of glucose oxidase in polypyrrole and its application in amperometric glucose sensors , 1986 .

[18]  I Karube,et al.  A chemiluminescent FIA biosensor for phosphate ion monitoring using pyruvate oxidase. , 1997, Biosensors & bioelectronics.

[19]  I. Karube,et al.  A novel biosensor system for the determination of phosphate , 1996 .

[20]  W. Schuhmann,et al.  Amperometric biosensors produced by immobilization of redox enzymes at polythiophene‐modified electrode surfaces , 1996 .

[21]  R. L. Elsenbaumer,et al.  Handbook of conducting polymers , 1986 .

[22]  C. H. Fiske,et al.  THE COLORIMETRIC DETERMINATION OF PHOSPHORUS , 1925 .

[23]  W. Schuhmann,et al.  Covalent binding of glucose oxidase to functionalized polyazulenes. The first application of polyazulenes in amperometric biosensors , 1993 .

[24]  S O Engblom,et al.  The phosphate sensor. , 1998, Biosensors & bioelectronics.

[25]  Gordon G. Wallace,et al.  Conducting electroactive polymer-based biosensors , 1999 .

[26]  M. Umaña,et al.  Protein-modified electrodes. The glucose oxidase/polypyrrole system , 1986 .

[27]  L. Gorton,et al.  Direct electron transfer between heme-containing enzymes and electrodes as basis for third generation biosensors , 1999 .

[28]  G. Guilbault,et al.  Amperometric determination of phosphate by use of a nucleoside phosphorylase-xanthine oxidase enzyme sensor based on a Clark-type hydrogen peroxide or oxygen electrode. , 1990, Analytical biochemistry.

[29]  J. D. Burton,et al.  Problems in the analysis of phosphorus compounds , 1973 .

[30]  A P Turner,et al.  Biosensors--Sense and Sensitivity , 2000, Science.

[31]  J. V. Bannister,et al.  Amperometric dertermination of phosphate ions by biosensor , 1992 .

[32]  R. Murray,et al.  CHEMICALLY MODIFIED ELECTRODES , 1977 .

[33]  R. Renneberg,et al.  Biosensor for rapid phosphate monitoring in a sequencing batch reactor (SBR) system. , 2003, Biosensors & bioelectronics.

[34]  Y. Shim,et al.  A performance comparison of choline biosensors: anodic or cathodic detections of H2O2 generated by enzyme immobilized on a conducting polymer. , 2004, Biosensors & bioelectronics.

[35]  E. Laviron.  General expression of the linear potential sweep voltammogram in the case of diffusionless electrochemical systems , 1979 .

[36]  M. Mascini,et al.  AP-GOD Biosensor Based on a Modified Poly(PHENOL) Film Electrode and Its Application in the Determination of Low Levels of Phosphate , 1995 .

[37]  A. Yaropolov,et al.  Direct electron transfer effect biosensors , 1996 .