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Bachelor's Thesis
Author of thesis: Bc. Richard Struhár
Acad. year: 2024/2025
Supervisor: doc. Ing. Filip Mravec, Ph.D.
Reviewer: Ing. Ivana Málková
Temperature-sensitive liposomes (TSLs) represent an attractive platform for the delivery of controlled-release biologically active agents, whereby upon a slight increase in temperature, their lipid bilayer undergoes a phase transition and thus becomes more permeable to the encapsulated content. This work focuses on the preparation, characterization, and functional testing of TSLs based on phospholipid dipalmitoylphosphatidylcholine (DPPC) with the addition of different lysolipid monopalmitoylphosphatidylcholine (MPPC) concentrations. Compositions both with and without sodium dilauroylglycerophosphate (PA) were chosen. In the first stage, vesicles were prepared by the thin lipid film hydration method (for different compositions of DPPC:PA:MPPC) at a total lipid concentration of 5 mmol/dm3. Subsequently, they were unilamellarized by 15-fold extrusion through a 100 nm membrane. Size and PDI (polydispersity index) were determined by DLS (dynamic light scattering), zeta potential by ELS (electrophoretic light scattering). The phase transitions of the membranes were monitored by fluorescence spectrometry, which measured anisotropy with two probes DPH (diphenylhexatriene) and its derivative TMA-DPH (trimethylammonium diphenylhexatriene) at temperatures between 10 and 70 °C. The kinetics of the encapsulated hydrophilic calcein was tested on the most promising 90:10 mol. % composition (DPPC:Lyso PC). Samples with 10 mol. % Lyso-PC and 90 mol. % DPPC were monodisperse, Z-average was around 125 nm, PDI < 0.07, and surface charge was neutral (ζ potential = 3 mV). Anisotropy measurements showed an inflection point of Tm = 37.5 ± 0.5 °C. The width of the phase transition (x0 ± 2dx) was 34-41 °C corresponding to the range of mild hyperthermia. However, the expected rapid release of the encapsulated calcein was not achieved when heated to 45 °C, after 24 h, < 5 % of the calcein had escaped. The results of this work suggest that transient pores are formed only during the phase transition and disappear after the transition to the liquid-crystalline phase. In this work, a composition of 90:10 mol. % (DPPC:MPPC) was selected as a suitable base for low-temperature TSL. Further investigation of the membrane properties during the phase transition will be required to reproduce the rapid release of the contents when a certain temperature is reached.
DPPC, temperature-sensitive liposomes, lysolipid, membrane phase transition, fluorescence anisotropy, membrane fluidity, dynamic light scattering (DLS), release kinetics
Date of defence
12.06.2025
Result of the defence
Defended (thesis was successfully defended)
Grading
A
Process of defence
Obhajoba bakalářské práce proběhla podle následujícího schématu: prezentace studenta-vyjádření vedoucí/ho-oponentský posudek-reakce na posudek-diskuse s komisí. Student přednesl výborný výtah výsledků své práce, řádně zodpověděl všechny dotazy oponentské i členů komise, pohotově reagoval na připomínky. V diskusi tak student prokázal výbornou schopnost orientace v teoretických i praktických základech problematiky práce. Komise zhodnotila jeho práci celkově jako výbornou. Klučáková: Jak jste určil počáteční koncentraci kalceinu při uvolňování? Dzik: Prosím upřesněte metodiku stanovení uvolněného kalceinu. Krčma: Jaké jsou nejistoty uvedených výsledků?
Language of thesis
Slovak
Faculty
Fakulta chemická
Department
Institute of Physical and Applied Chemistry
Study programme
Chemistry and Chemical Technologies (BPCP_CHCHTE)
Composition of Committee
prof. Ing. Martina Klučáková, Ph.D. (předseda) doc. Ing. Vojtěch Enev, Ph.D. (člen) prof. RNDr. František Krčma, Ph.D. (člen) doc. Ing. Zdenka Kozáková, Ph.D. (člen) doc. Ing. Petr Dzik, Ph.D. (člen) doc. Ing. Filip Mravec, Ph.D. (místopředseda)
Supervisor’s reportdoc. Ing. Filip Mravec, Ph.D.
Grade proposed by supervisor: A
Reviewer’s reportIng. Ivana Málková
Grade proposed by reviewer: A
Responsibility: Mgr. et Mgr. Hana Odstrčilová