A mediated polyphenol oxidase biosensor immobilized by electropolymerization of 1,2-diamino benzene.
A biosensor based on a partially purified polyphenol oxidase (PPO) enzyme was developed by using electropolymerization of [(2,2'-bipyridine)(chloro)(p-cymene)rutenium(II)]chloride] mediator complex and 1,2-diamino benzene (DAB) on a screen printing Pt electrode (1mm diameter). The electropolymerization was carried out at +0.7V for 45min in phosphate buffer (50mM, pH 7.0) which contained 14.0U/10mL polyphenole oxidase, 200mM DAB and 2.5mM Ru-mediator complex solutions. Measurement is based on the detection of the oxidation current of the Ru-mediator complex that related to the enzymatic reaction catalyzed by PPO at +0.65V. The phosphate buffer (50mM, pH 7.0 containing 0.1M KCl) and 30 degrees C were established as being the optimum working conditions. Under the optimum experimental conditions a linear calibration curve was obtained between 5 and 100microM catechol concentration. The detection limit of the biosensor is 2.385microM. In the characterization studies of the biosensor some parameters such as effect of Ru-mediator types on the biosensor response, substrate specificity, reproducibility and storage stability were studied. From the experiments, the average value (x), standard deviation (SD) and coefficient of variation (CV%) were found to be 48.75microM,+/-1.56microM, and 3.2% respectively for 50microM catechol standard.
Department of Biochemistry, Faculty of Science, Ege University, 35100 Bornova-Izmir, Turkey. erol.akyilmaz@ege.edu.tr
Erol
E
Akyilmaz
Ozge
O
Kozgus
Hayati
H
Türkmen
Bekir
B
Cetinkaya
D015374:Biosensing Techniques / Q000379:methods*; D004156:Catechol Oxidase / Q000737:chemistry* / Q000378:metabolism*; D004563:Electrochemistry; D004566:Electrodes; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D016014:Linear Models; D009942:Organometallic Compounds / Q000737:chemistry; D010655:Phenylenediamines / Q000737:chemistry*; D010984:Platinum / Q000737:chemistry; D011108:Polymers / Q000737:chemistry*; D011327:Printing; D015203:Reproducibility of Results; D011198:Solanum tuberosum / Q000201:enzymology; D013379:Substrate Specificity
D004800:Enzymes, Immobilized; D009942:Organometallic Compounds; D010655:Phenylenediamines; D011108:Polymers; D010984:Platinum; C034193:1,2-diaminobenzene; D004156:Catechol Oxidase