Comparison of oxidative state in the cells of wild type and highproducer strain of bacteria Saccharopolyspora erythraea during erythromycin production

Cilj magistrske naloge je bil primerjati oksidacijsko stanje naravnega in visokodonosnega seva bakterije Saccharopolyspora erythraea tekom produkcije eritromicina in s pomočjo tega ugotoviti povezavo med oksidativnim stresom in sekundarnim metabolizmom. Pripravili smo bioproces z naravnim in visokodonosnim sevom, kjer smo spremljali parametre, kot so pH, parcialni volumen micelija, koncentracija glukoze in eritromicina. Za ovrednotenje oksidacijskega stanja smo z uporabo barvila 2′,7′-diklorodihidrofluorescein (H2DCF) merili znotrajcelično oksidacijo. Nivo znotrajcelične oksidacije je bil v času intenzivne produkcije eritromicina povišan pri visokodonosnem sevu, medtem ko pri naravnem sevu ni bilo opaznih sprememb. V nadaljevanju smo naredili še analize aktivnosti katalaze, in sicer s spremljanjem razgradnje vodikovega peroksida preko zniževanja absorbance vzorca. Tekom bioprocesa se aktivnost katalaze ni spreminjala, tako pri visokodonosem kot pri naravnem sevu. Kljub zvišanem nivoju znotrajcelične oksidacije v času intenzivne produkcije eritromicina pri visokodonosnem sevu, je aktivnost katalaze ostala primerljiva z naravnim sevom. Naši rezultati tako nakazujejo, da bi lahko obstajala pozitivna korelacija med oksidativnim stresom in donosom eritromicina pri bakteriji Saccharopolyspora erythraea.The aim of this thesis was to compare the oxidative state in cells of wild type and the highproducer strain of bacterium Saccharopolyspora erythraea during the erythromycin production and also to establish potencial connection between oxidative stress and secondary metabolism. We set up bioprocess with wild type and the industrial high-producing strain, where parameters like pH, partial mycelium volume, glucose and erythromycin concentration were measured. To evaluate oxidative state, we used 2\u27,7\u27-dichlorodihydrofluorescein to measure intracellular oxidation. Intracellular oxidation level was higher during the intensive production of erythromycin in the industrial strain, while in the wild type strain there were no noticeable changes. Furthermore, we determined the catalase activity by measuring the decrease in absorbance of sample during hydrogen peroxide degradation. We have not observed any changes in catalase activity, independently on the strain used. Although we observed higher intracellular oxidation during intensive erythromycin production in the high-producing strain, catalase activity was still comparable with the wild type strain. Our results indicate, that there is a positive correlation between oxidative stress and erythromycin production in Saccharopolyspora erythraea