IMMOBILIZATION OF LIPASE ONTO MAGHEMITE NANOPARTICLES, MODIFIED WITH AMINOSILANE

Namen diplomske naloge je sinteza maghemitnih nanodelcev, prevlečenih z aminosilanom, ter imobilizacija encima lipaze na pripravljen nosilec. Maghemitne nanodelce smo pripravili z obarjalno reakcijo ali koprecipitacijo železovih II in železovih III ionov s 25 % amonijakom in jih prevlekli s funkcionalno plastjo silike, nato pa še s plastjo aminosilana. Optimirali smo pogoje za dosego najvišje učinkovitosti imobilizacije ter preostale aktivnosti imobiliziranega encima, kot so: koncentracija encima, vrsta mrežnega povezovalca, čas imobilizacije ter koncentracija mrežnega povezovalca glutaraldehida. Proučevali smo stabilnost imobiliziranega encima, vpliv časa in temperature izpostavitve na ohranitev aktivnosti imobiliziranega ter prostega encima. Najvišjo učinkovitost imobilizacije smo dosegli po aktivaciji magnetnih nanodelcev z 1 % glutaraldehidom (GA), s koncentracijo encima 7,5 µL/mL ter časom imobilizacije 24 ur pri hitrosti stresanja 410 rpm. Kljub visoki učinkovitosti imobilizacije smo v večini primerov dobili zelo nizke ohranjene aktivnosti imobiliziranega encima, imobiliziran encim pa je pokazal boljšo stabilnost pri povišani temperaturi v primerjavi s prostim encimom. The purpose of the diploma thesis is the synthesis of magnetic maghemite nanoparticles coated with aminosilane and immobilization of lipase on a ready carrier. Maghemite nanoparticles were synthesized by the coprecipitation technique of ferrous II and ferric III ions with 25 % ammonia and coated with silica and afterwards with aminosilane. Process conditions for achieving the maximum efficiency and the residual activity of the immobilized enzyme were optimized, by changing the concentration of the enzyme, the type of a cross-linker, the time of immobilization and the concentration of a cross-linker glutaraldehyde. Our subject of interest was also the stability of the enzyme and how the remaining activity of the immobilized enzyme was affected by exposed time and temperature. Maximum efficiency was achieved after activating maghemite nanoparticles with 1 % glutaraldehyde (GA), with the enzyme concentration of 7,5 µL/mL after 24 hours at 410 rpm. Despite a good binding efficiency of the immobilization, very low residual activity was obtained in most cases. However, the immobilized enzyme showed a better stability at increased temperatures as compared to the wild enzyme.