Entwicklung eines amperometrischen Biosensors zur Erfassung von Polyphenolen

In der durchgeführten Arbeit wurden neue Immobilisierungsstrategien zur Etablierung eines amperometrischen Biosensors, welcher Polyphenole erfasst, untersucht. Ziel war es eine „Handheld-Unit“ unter Verwendung einer Dickschichtelektrode (Thick-Film Technology, TFT) zu entwickeln. Zur Erfassung von...

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Bibliographic Details
Main Author: Schmidt, Sascha Gabriel
Contributors: Keusgen, Michael (Prof. Dr.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2011
Online Access:PDF Full Text
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The aim of this dissertation was to establish a new immobilization method for an amperometric biosensor, which detects polyphenols. The use of a handheld unit using a thick-film electrode (thick-film technology, TFT) for the detection of polyphenols was a further aim of this work. For detection of polyphenols the enzyme Laccase was selected, which catalyzes the oxidation of phenols to quinones. In this work the Laccases from Rhus vernicifera, Agaricus bisporus and Trametes versicolor were tested for the possible use as the biological component of the biosensor. For the tested immobilization methods the Laccase from Trametes versicolor was suitable. As a new approach for the immobilization of laccase for the detection of polyphenols, the immobilization reagent polylactic acid (PLA) was selected. Different polylactic acids were analyzed. The conductivity of the polymer could be achieved with activated carbon as well as with carbon nanotubes (CNT). The use of CNTs shows a more stable signal. Furthermore the immobilization of enzymes with photolinkers on TFT sensors was analyzed. As another part of this work the photolinkers N-hydroxysulfosuccinimidyl-4-azidobenoate (Sulfo-HSAB), 4-fluoro-3-nitrophenyl azide (FNPA) and the combination of 3-aminopropyltriethoxysilane/glutaraldehyde (APTES/GDA) were tested. Best sensitivity and robustness of the sensor were reached, when the enzyme Laccase was immobilized by FNPA on the TFT-sensor. Real samples (green tea (Camellia sinensis), aronia juice and cranberry juice) have been measured using the TFT biosensor. These results were compared with the results of the reference method Folin-Ciocalteu. The biosensor results are very suitable for screening. For the immobilization method with PLA and for the immobilization method with the photolinker FNPA the storage stability of coated biosensors was checked. Both coating strategies have a long shelf life (PLA coating: at least 83 days, FNPA coating at least 28 days).