Razvoj magnetnih kovinsko-organskih ogrodij za sočasno imobilizacijo in delovanje več encimov : magistrsko delo

Senekovič, Tajda

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Title:	Razvoj magnetnih kovinsko-organskih ogrodij za sočasno imobilizacijo in delovanje več encimov : magistrsko delo
Authors:	ID Senekovič, Tajda (Author) ID Vasić, Katja (Mentor) More about this mentor... ID Primožič, Mateja (Comentor)
Files:	MAG_Senekovic_Tajda_2025.pdf (2,89 MB) MD5: DAE355E74263F555DAF66010D3521FED
Language:	Slovenian
Work type:	Master's thesis/paper
Typology:	2.09 - Master's Thesis
Organization:	FKKT - Faculty of Chemistry and Chemical Engineering
Abstract:	Onesnaženje vodnih virov z ostanki farmacevtskih spojin, zlasti antibiotikov, predstavlja resen okoljski problem, saj že nizke koncentracije negativno vplivajo na vodno okolje in spodbujajo razvoj antimikrobne odpornosti. Ker konvencionalni postopki čiščenja teh spojin pogosto ne odstranijo v celoti, postajajo biokatalitski pristopi vse bolj zanimivi. Encimi, kot sta β-laktamaza in lakaza, omogočajo selektivno razgradnjo onesnaževal, njihovo učinkovitost pa je mogoče bistveno povečati z imobilizacijo na magnetna kovinsko-organska ogrodja (mMOFs), ki izboljšajo stabilnost, ponovno uporabnost in omogočajo enostavno ločevanje katalizatorjev iz reakcijskega medija. V magistrskem delu smo razvili mMOFs na osnovi ZIF-8 za sočasno imobilizacijo β-laktamaze in lakaze ter ovrednotili njihovo učinkovitost pri razgradnji penicilina G (PEN). Sintetizirali smo magnetne nanodelce, prevlečene z arabinogalaktanom (AG-MNPs), pripravili ZIF-8 in izdelali magnetne kompozite mZIF-8. Kot optimalne pogoje imobilizacije smo uporabili 7,5 % (v/v) glutaraldehida (GA) in HEPES pufer s pH 7,3. Preostala aktivnost imobiliziranih encimov je znašala 53,85 % za β-laktamazo in 94,21 % za lakazo, pri sočasni imobilizaciji pa 39,73 % za β-laktamazo oziroma 53,79 % za lakazo. Visoka učinkovitost imobilizacije (85–96 %) potrjuje ustreznost izbranega nosilca. Razgradnjo PEN smo preučevali pri koncentracijah 0,1, 1 in 10 mg/mL. Pri 0,1 mg/mL PEN so imobilizirani sistemi odstranili PEN hitreje od prostih encimov: β-laktamaza@mZIF-8 v 30 minutah, lakaza@mZIF-8 v 40 minutah, β-laktamaza/lakaza@mZIF-8 pa že v 10 minutah. Pri 1 mg/mL PEN je prosta β-laktamaza v 90 min razgradila le 8 % PEN, medtem ko je sistem β-laktamaza/lakaza@mZIF-8 v enakem času razgradil celotni PEN. Največje razlike med sistemi so se pokazale pri 10 mg/mL PEN. Pri mZIF-8 je bil dosežen adsorpcijski plato pri približno 30–32 %, kar potrjuje nasičenje adsorpcijskih mest. Prosta β-laktamaza je za skoraj popolno razgradnjo potrebovala 192 ur, medtem ko je pri β-laktamazi@mZIF-8 ostalo le okoli 5 % PEN. Najbolj učinkovit je bil soimobiliziran sistem, ki je delež PEN znižal na približno 3 %. Povišanje koncentracije obeh encimov pri sočasni imobilizaciji je dodatno pospešilo razgradnjo PEN. Pri najnižjih koncentracijah encimov (0,1 mg/mL β-laktamaze in 1 mg/mL lakaze) se je po 90 min razgradilo 20 % PEN, pri srednjih koncentracijah (0,2 mg/mL β-laktamaze in 2 mg/mL lakaze) 38 %, pri najvišjih koncentracijah (0,4 mg/mL β-laktamaze in 4 mg/mL lakaze) pa 45 %. Rezultati jasno potrjujejo neposredno povezavo med koncentracijo imobiliziranih encimov in katalitsko učinkovitostjo sistema.
Keywords:	imobilizacija encimov, magnetna kovinsko-organska ogrodja (mMOFs), lakaza, β-laktamaza, HPLC, čiščenje odpadnih vod
Place of publishing:	Maribor
Place of performance:	Maribor
Publisher:	[T. Senekovič]
Year of publishing:	2025
Number of pages:	1 spletni vir (1 datoteka PDF (XI, 57 str.))
PID:	20.500.12556/DKUM-96312
UDC:	628.35:577.15(043.2)
COBISS.SI-ID:	270124035
Publication date in DKUM:	02.03.2026
Views:	173
Downloads:	24
Metadata:
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:	17.12.2025

Secondary language

Language:	English
Title:	Development of magnetic metal-organic frameworks for simultaneous immobilization and function of multiple enzymes
Abstract:	Pollution of water resources with residues of pharmaceutical compounds, particularly antibiotics, represents a serious environmental problem, as even low concentrations can adversely affect aquatic ecosystems and promote the development of antimicrobial resistance. Since conventional wastewater treatment processes often fail to completely remove these compounds, biocatalytic approaches are becoming increasingly attractive. Enzymes such as β-lactamase and laccase enable the selective degradation of pollutants, and their efficiency can be significantly enhanced through immobilization on magnetic metal–organic frameworks (mMOFs), which improve stability and reusability and allow for easy separation of the catalysts from the reaction medium. In this master’s thesis, we developed ZIF-8-based mMOFs for the simultaneous immobilization of β-lactamase and laccase and evaluated their efficiency in the degradation of penicillin G (PEN). Magnetic nanoparticles coated with arabinogalactan (AG-MNPs) were synthesized, ZIF-8 was prepared, and magnetic composites (mZIF-8) were fabricated. The optimal immobilization conditions were 7.5% (v/v) glutaraldehyde (GA) and HEPES buffer at pH 7.3. The residual activity of the immobilized enzymes was 53.85% for β-lactamase (β-lactamase@mZIF-8) and 94.21% for laccase (laccase@mZIF-8), while in the case of co-immobilization (β-lactamase/laccase@mZIF-8) the residual activity was 39.73% for β-lactamase and 53.79% for laccase. The high immobilization efficiency (85–96%) confirms the suitability of the selected support. The degradation of PEN was investigated at antibiotic concentrations of 0.1, 1, and 10 mg/mL. At 0.1 mg/mL PEN, the immobilized systems removed/degraded PEN more rapidly than the free enzymes: β-lactamase@mZIF-8 within 30 minutes, laccase@mZIF-8 within 40 minutes, and the β-lactamase/laccase@mZIF-8 system within only 10 minutes. At 1 mg/mL PEN, free β-lactamase degraded only 8% of PEN within 90 minutes, whereas the β-lactamase/laccase@mZIF-8 system completely removed/degraded PEN within the same time period. The greatest differences among the systems were observed at a PEN concentration of 10 mg/mL. For mZIF-8, an adsorption plateau of approximately 30–32% was reached. Free β-lactamase required 192 hours for near-complete degradation, whereas only about 5% of PEN remained unremoved/undegraded when using β-lactamase@mZIF-8. The co-immobilized system was the most efficient, achieving almost complete removal/degradation of PEN (97%). Increasing the concentrations of both enzymes in the co-immobilized system further accelerated PEN degradation. At the lowest concentrations of immobilized enzymes (0.1 mg/mL β-lactamase and 1 mg/mL laccase), 20% of PEN was degraded after 90 minutes; at intermediate concentrations (0.2 mg/mL β-lactamase and 2 mg/mL laccase), 38% was degraded; and at the highest tested concentrations (0.4 mg/mL β-lactamase and 4 mg/mL laccase), 45% degradation was achieved. These results clearly confirm a direct relationship between the concentration of immobilized enzymes and the catalytic efficiency of the system.
Keywords:	enzyme immobilization, magnetic metal-organic frameworks (mMOFs), laccase, β-lactamase, HPLC, wastewater treatment

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