title: An integrated bienzyme glucose oxidase-fructose dehydrogenase-tetrathiafulvalene-3-mercaptopropionic acid-gold electrode for the simultaneous determination of glucose and fructose.
abstract: A bienzyme biosensor for the simultaneous determination of glucose and fructose was developed by coimmobilising glucose oxidase (GOD), fructose dehydrogenase (FDH), and the mediator, tetrathiafulvalene (TTF), by cross-linking with glutaraldehyde atop a 3-mercaptopropionic acid (MPA) self-assembled monolayer (SAM) on a gold disk electrode (AuE). The performance of this bienzyme electrode under batch and flow injection (FI) conditions, as well as an amperometric detection in high-performance liquid chromatography (HPLC), are reported. The order of enzyme immobilisation atop the MPA-SAM affected the biosensor amperometric response in terms of sensitivity, with the immobilisation order GOD, FDH, TTF being selected. Similar analytical characteristics to those obtained with single GOD or FDH SAM-based biosensors for glucose and fructose were achieved with the bienzyme electrode, indicating that no noticeable changes in the biosensor responses to the analytes occurred as a consequence of the coimmobilisation of both enzymes on the same MPA-AuE. The suitability of the bienzyme biosensor for the analysis of real samples under flow injection conditions was tested by determining glucose in two certified serum samples. The simultaneous determination of glucose and fructose in the same sample cannot be performed without a separation step because at the detection potential used (+0.10 V), both sugars show amperometric response. Consequently, HPLC with amperometric detection at the TTF-FDH-GOD-MPA-AuE was accomplished. Glucose and fructose were simultaneously determined in honey, cola softdrink, and commercial apple juice, and the results were compared with those obtained by using other reference methods.
authors.affiliation: Departamento de Química Analítica, Facultad de CC Químicas, Universidad Complutense de Madrid, Madrid E-28040, Spain.
authors.forename: Susana
authors.initials: S
authors.lastname: Campuzano
authors.forename: Oscar A
authors.initials: OA
authors.lastname: Loaiza
authors.forename: María
authors.initials: M
authors.lastname: Pedrero
authors.forename: F Javier Manuel
authors.initials: FJ
authors.lastname: de Villena
authors.forename: José M
authors.initials: JM
authors.lastname: Pingarrón
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D002237:Carbohydrate Dehydrogenases / Q000737:chemistry*; D020099:Coated Materials, Biocompatible / Q000737:chemistry; D045424:Complex Mixtures / Q000032:analysis / Q000737:chemistry; D004563:Electrochemistry / Q000379:methods*; D004800:Enzymes, Immobilized / Q000737:chemistry; D005632:Fructose / Q000032:analysis* / Q000737:chemistry; D005947:Glucose / Q000032:analysis* / Q000737:chemistry; D005949:Glucose Oxidase / Q000737:chemistry*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D018511:Systems Integration
chemical_list: D020099:Coated Materials, Biocompatible; D045424:Complex Mixtures; D004800:Enzymes, Immobilized; D005632:Fructose; D002237:Carbohydrate Dehydrogenases; D005949:Glucose Oxidase; C028761:D-fructose 5-dehydrogenase; D005947:Glucose