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Production of lignin-degrading enzymes by Phanerochaete chrysosporium
Péčiová, Bianka ; Hudečková, Helena (referee) ; Kovalčík, Adriána (advisor)
This diploma thesis deals with the study of the production of ligninolytic enzymes produced by the mold P. chrysosporium and subsequently their immobilization. The theoretical part describes the selected production microorganism, principles of action of ligninolytic enzymes (lignin-peroxidase, manganese-dependent peroxidase and laccase) and methods of their immobilization. In the experimental part, optimal conditions of the production of ligninolytic enzymes by P. chrysosporium were monitored as a composition of culture medium with different glucose content and with using two different inducers: lignin and guaiacol. During the 14-day cultivation cycles, the enzyme activity of lignin-peroxidase, manganese-dependent peroxidase and laccase was monitored on each day of cultivation. Based on the optimized enzyme production procedure, the enzymes were subsequently immobilized by various methods: by method of forming enzyme aggregates (CLEA), adsorption on bacterial cellulose (BC) and a combination of both (CLEA + BC). The CLEA method proved to be the most effective, but there was problem with isolation of enzyme aggregates. Therefore, immobilization on bacterial cellulose associated with the enzymes aggregates was chosen as the most advantageous. With this method, the operational stability of the immobilized enzymes was monitored in 7 consecutive reaction cycles. Subsequently, CLEA and CLEA with BC were used to modify the model phenolic substance, where the alkali lignin was selected. For both sets of immobilizers was confirmed the effect of the enzymes on the lignin structure by measuring the molecular weight (Mw) and polydispersity index (PI).
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