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Title:Imobilizacija terapevtskega encima na magnetne nanodelce : magistrsko delo
Authors:ID Potisk, Patricija (Author)
ID Leitgeb, Maja (Mentor) More about this mentor... New window
ID Primožič, Mateja (Comentor)
ID Vasić, Katja (Comentor)
Files:.pdf MAG_Potisk_Patricija_2024.pdf (2,03 MB)
MD5: 958477429B05A64454A39DE2B282DE72
 
Language:Slovenian
Work type:Master's thesis/paper
Typology:2.09 - Master's Thesis
Organization:FKKT - Faculty of Chemistry and Chemical Engineering
Abstract:Namen magistrskega dela je uspešna imobilizacija encima lizocima na magnetne nanodelce. Praktična uporaba prostih encimov je pogosto omejena zaradi njihove relativno nizke stabilnosti in aktivnosti. Z namenom, da se le-ta ohrani ali poveča, se poslužujemo različnih metod imobilizacije encimov. V okviru magistrskega dela smo najprej sintetizirali magnetne nanodelce, ki so služili kot nosilci za imobilizacijo biokatalizatorja. Nanodelce smo prevlekli z različnimi polimeri: z aminosilanom, arabinogalaktanom in dekstranom. Preden smo nanje imobilizirali lizocim, smo jih funkcionalizirali z mrežnim povezovalcem glutaraldehidom oziroma epiklorohidrinom. Uporabljena koncentracija mrežnega povezovalca je pomembno vplivala na aktivnost imobiliziranega lizocima in na učinkovitost imobilizacije. Najvišje aktivnosti imobiliziranega lizocima smo pri aminosilanskih nanodelcih dosegli, ko smo uporabili mrežni povezovalec gluteraldehid s koncentracijo 1 % (v/v), pri arabinogalaktanskih nanodelcih pa s koncentracijo 15 % (v/v). V primeru dekstranskih nanodelcev je optimalno koncentracijo mrežnega povezovalca predstavljal 1 % (v/v) epiklorohidrina. Za prosti in imobilizirani lizocim smo nato določili dva kinetična parametra, maksimalno hitrost in Michaelis-Mentenovo konstanto. Raziskali smo tudi termično stabilnost lizocima. Rezultati so pokazali, da je imobilizirani lizocim ohranil aktivnost tudi po 24-urni izpostavitvi pri temperaturi 50 °C, medtem ko je prosti encim pri enakih pogojih denaturiral. Proučili smo tudi stabilnost imobiliziranega encima, ki smo ga tri tedne hranili na 4 °C. Encim, ki je bil imobiliziran na aminosilanskih in arabinogalaktanskih nanodelcih, je po dveh tednih skladiščenja ohranil 86 % oziroma 78 % začetne aktivnosti, po treh tednih skladiščenja pri 4 °C pa je njegova aktivnost upadla. Lizocim, imobiliziran na dekstranske nanodelce, se je tako pri študiji termične stabilnosti kot tudi pri študiji stabilnosti pri 4 °C izkazal za najmanj stabilnega.
Keywords:lizocim, imobilizacija, magnetni nanodelci, mrežni povezovalec, aktivnost
Place of publishing:Maribor
Place of performance:Maribor
Publisher:[P. Potisk]
Year of publishing:2024
Number of pages:1 spletni vir (1 datoteka PDF (X, 39 f.))
PID:20.500.12556/DKUM-89292 New window
UDC:577.151(043.2)
COBISS.SI-ID:212147971 New window
Publication date in DKUM:25.07.2024
Views:369
Downloads:97
Metadata:XML DC-XML DC-RDF
Categories:KTFMB - FKKT
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Licences

License:CC BY-NC-ND 4.0, Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International
Link:http://creativecommons.org/licenses/by-nc-nd/4.0/
Description:The most restrictive Creative Commons license. This only allows people to download and share the work for no commercial gain and for no other purposes.
Licensing start date:02.07.2024

Secondary language

Language:English
Title:Immobilization of therapeutic enzyme on magnetic nanoparticles
Abstract:The aim of the master's thesis is the successful immobilization of the enzyme lysozyme on magnetic nanoparticles. The practical use of free enzymes is often limited due to their relatively low stability and activity. To preserve or increase these properties, we use various methods of enzyme immobilization. As part of the master's thesis, we synthesized magnetic nanoparticles that served as carriers for the immobilization of the biocatalyst. The nanoparticles were coated with different polymers: aminosilane, arabinogalactan and dextran. Before lysozyme was immobilized on them, the nanoparticles were functionalized with cross-linkers glutaraldehyde and epichlorohydrin. The concentration of the cross-linker used significantly affected the activity of the immobilized lysozyme and the efficiency of immobilization. The highest activities of immobilized lysozyme were achieved with aminosilane nanoparticles using a glutaraldehyde concentration of 1% (v/v), and with arabinogalactan nanoparticles using a concentration of 15% (v/v). In the case of dextran nanoparticles, the optimal concentration of the cross-linker was 1% (v/v) epichlorohydrin. For both free and immobilized lysozyme, two kinetic parameters were determined: the maximum velocity and the Michaelis-Menten constant. We also investigated the thermal stability of lysozyme. The results showed that immobilized lysozyme retained activity even after 24 hours of exposure at 50 °C, while the free enzyme denatured under the same conditions. We also studied the stability of the immobilized enzyme stored at 4 °C for three weeks. The enzyme immobilized on aminosilane and arabinogalactan nanoparticles retained 86 % and 78 % of its initial activity after two weeks of storage, respectively, but its activity declined after three weeks of storage at 4 °C. Lysozyme immobilized on dextran nanoparticles proved to be the least stable in both the thermal stability study and the stability study at 4°C.
Keywords:lysozyme, immobilization, magnetic nanoparticles, cross-linker, activity


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