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Title:Preučevanje mehanizma oksidacije pomožnih snovi v proteinskih formulacijah
Authors:ID Matves, Sanja (Author)
ID Zalar, Matja (Mentor) More about this mentor... New window
ID Fermišek, Miha (Comentor)
Files:.pdf UN_Matves_Sanja_2026.pdf (3,94 MB)
MD5: A75644DB6AA2851E5FD38D8B0D4B66BC
 
Language:Slovenian
Work type:Bachelor thesis/paper
Typology:2.11 - Undergraduate Thesis
Organization:FKKT - Faculty of Chemistry and Chemical Engineering
Abstract:Pomožne snovi imajo v farmaciji ključno vlogo pri zagotavljanju stabilnosti, varnosti in ustrezne oblike končnega zdravila. Ena izmed pogostejših pomožnih snovi je aminokislina histidin, ki se uporablja kot pomembna puferska pomožna snov za stabilizacijo bioloških zdravil. Vendar pa lahko prisotnost sledov prehodnih kovinskih ionov v formulaciji povzroči kovinsko katalizirano oksidacijo histidina, kar vodi do degradacije pufra in posledično agregacije zdravilne učinkovine. V diplomskem delu smo s pomočjo jedrske magnetne resonance (NMR) preučevali kinetiko oksidacije histidina ter primerjali katalitični vpliv železovih, bakrovih in nikljevih ionov (Fe(II), Cu(II), Ni(II)) ter kontrolnega vzorca brez dodanih kovin v raztopini 50 mM histidina in 0,2 M vodikovega peroksida pri vrednosti pH 6. S spremljanjem časovne spremembe integriranih površin NMR signalov za protona Hε in Hδ na imidazolnem obroču histidina smo ocenili konstante reakcijskih hitrosti za posamezne kovine. Analiza NMR spektrov je razkrila izrazite razlike v kinetiki oksidacije v odvisnosti od uporabljene kovine. Prisotnost železovih ionov je pospešila oksidativno razgradnjo imidazolnega obroča, medtem ko je le-ta v prisotnosti Cu(II) potekala bistveno počasneje. Ioni Ni(II) so se izkazali za redoks neaktivne, saj v izbranem časovnem okviru oksidativna razgradnja histidina ni potekla. Ugotovili smo, da je stabilnost histidina kot pomožne snovi močno odvisna od prisotnosti prehodnih kovin, kar je ključnega pomena pri zagotavljanju varnosti in stabilnosti zdravilne učinkovine. Poznavanje kinetike razgradnje omogoča natančnejše napovedovanje roka uporabnosti proteinskih formulacij ter utemeljuje potrebo po strogi kontroli kovinskih nečistoč v proizvodnih procesih in v produktih.
Keywords:pomožne snovi, histidin, kovinsko katalizirana oksidacija, NMR spektroskopija, namizni NMR spektrometer
Place of publishing:Maribor
Year of publishing:2026
PID:20.500.12556/DKUM-99564 New window
Publication date in DKUM:09.09.2026
Views:220
Downloads:7
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:20.08.2026

Secondary language

Language:English
Title:Investigating the oxidation mechanism of excipients in protein formulations
Abstract:Excipients play a crucial role in pharmacy by ensuring the stability, safety, and appropriate dosage form of the final medicinal product. One of the more common excipients is the amino acid histidine, which is used as an important buffering excipient to stabilize biological drugs. However, the presence of trace transition metal ions in the formulation can cause metal-catalyzed oxidation of histidine, leading to buffer degradation and subsequently the aggregation of the active pharmaceutical ingredient. In this study, we investigated the kinetics of histidine oxidation using nuclear magnetic resonance (NMR) spectroscopy and compared the catalytic effect of iron, copper, and nickel ions (Fe(II), Cu(II), Ni(II)) alongside a metal-free control sample in a solution of 50 mM histidine and 0.2 M hydrogen peroxide at pH 6. By monitoring the time-dependent changes in the integrated areas of the NMR signals for the Hε and Hδ protons on the imidazole ring of histidine, we estimated the reaction rate constants for individual metals. Analysis of the NMR spectra revealed distinct differences in oxidation kinetics depending on the metal used. The presence of iron ions accelerated the oxidative degradation of the imidazole ring, whereas in the presence of Cu(II), this process proceeded significantly slower. Ni(II) ions proved to be redox-inactive, as no oxidative degradation of histidine took place within the selected timeframe. We established that the stability of histidine as an excipient depends heavily on the presence of transition metals, which is of critical importance for ensuring the safety and stability of the active ingredient. Understanding the degradation kinetics allows for more accurate shelf-life predictions of protein formulations and justifies the need for strict control of metal impurities in production processes and final products.
Keywords:excipients, histidine, metal-catalyzed oxidation, NMR spectroscopy, benchtop NMR


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