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Namen naloge je bila imobilizacija encima glukoze oksidaze na trdni nosilec in razviti kinetični model oksidacije glukoze do glukono laktona. Z imobiliziranim encimom smo želeli doseči čim višjo presnovo glukoze ter njegovo večkratno uporabo. Najprej smo določali optimalne pogoje reakcije oksidacije glukoze s prostim encimom glukoza oksidaza. Najvišjo presnovo glukoze (100 %) po 180 min, smo dosegli pri T = 35 °C, qv,zrak = 20 L/h, f = 300 min -1 in pH = 7. Za imobilizacijo encima na trdni nosilec obstajajo različni postopki. Izbrali smo optimalnega in določili najboljše pogoje za njegovo učinkovito vezavo. Po uspešnem postopku imobilizacije je sledila optimizacija reakcije oksidacije glukoze z imobiliziranim encimom glukoza oksidaza. Po 3 h reakcije smo dosegli 95 % presnovo, procesni pogoji so bili: T = 35 °C, qv,zrak = 20 L/h, f = 300 min -1 in pH = 6. Z imobiliziranim encimom glukozo oksidazo nam je uspelo reakcijo izvesti 3- krat, vendar je bila zadnja presnova le še 35 %. Izvedli smo tudi kinetične izračune za reakcijo oksidacije glukoze z imobiliziranim encimom glukoza oksidaza. Potrdili smo Michaelis – Menten mehanizem reakcije in določili glavne parametre tega modela. Rezultati so primerljivi z literaturnimi vrednostmi. The aim of this work was focused mainly to the immobilization of the enzyme glucose oxidase on a solid carrier (support) and to the development of a kinetic model of the oxidation of glucose into glucone lactone. With the immobilized enzyme we wanted to achieve the highest possible glucose conversion and at the same time the multiple use of the immobilized encyme glucose oxidase. In the first step there were determined the optimal conditions for the reaction of glucose oxidation with the free enzyme of glucose oxidase. The highest glucose conversion (100 %) was achieved after 180 min at T = 35 °C, qv,air = 20 L/h, f = 300 min -1 in pH = 7. The immobilization of the enzyme on a solid carrier can be carried out with different procedures. We have selected the optimum and determined the best conditions for its efficient binding. A successful process of immobilization was followed by optimization of the oxidation reaction of glucose with immobilized enzyme glucose oxidase. The reaction lasted 3 h and at the given conditions, T = 35 °C, qv,air = 20 L/h, f = 300 min -1 in pH = 6, it achieved 95% conversion of glucose. The same immobilized enzyme glucose oxidase was successfuly used in three reactions, but in the last reaction the conversion reached only 35%. We have also performed kinetic calculations for the oxidation of glucose with immobilized enzyme glucose oxidase. We confirmed Michaelis - Menten reaction mechanism and determined the main parameters of this model. The results were comparable with the literature values. |
