Sinteza in vrednotenje fluorescenčnih sond na osnovi kumarina in lantanidnih kompleksov za označevanje lipofilnih struktur

Smrtnik, Martina (avtor), Pajk, Stane (mentor)

Detekcija s fluorescenco predstavlja zelo občutljivo metodo, ki se vedno bolj uporablja na področju bioznanosti in raziskovanja. Gre za pojav, kjer se s svetlobo določene valovne dolžine, vzbuja elektrone fluorofora, da preidejo v višje vzbujeno singletno energijsko stanje. Po nekaj nanosekundah se elektroni spontano vrnejo v osnovno energijsko stanje, ob tem pa oddajo odvečno energijo v obliki fluorescence, ki jo mi detektiramo. Za preučevanje s fluorescenco si želimo fotostabilne fluorofore, s čim večjim kvantnim izkoristkom in molarnim ekstinkcijskim koeficientom. Obenem morajo imeti dovoljšnjo specifičnost za tarčne molekule, dobro pa je če imajo tudi velik Stokesov premik. Ker je take fluorofore običajno težko dobiti, je smiselno nadaljnjo raziskovanje in odkrivanje novih fluoroforov. Naša magistrska naloga se osredotoča na sintezo fluorescenčnih sond za označevanje lipofilnih struktur. V prvem delu se posvečamo sintezi kumarinskih derivatov, ki so zanimivi predvsem za označevanje lipidnih kapljic. Lipidne kapljice so zaenkrat še dokaj neraziskani celični organeli, ki jih najdemo v vseh evkariontskih celicah. Številne raziskave ugotavljajo, da je kopičenje lipidnih kapljic v telesu povezano s pojavom določenih infekcijskih bolezni, rakavimi boleznimi, obolenji srca in boleznimi centralnega živčnega sistema (Alzheimerjeva bolezen, Parkinsonova bolezen), zato so orodja za raziskovanje, kot so fluorofori, zelo zaželena. V sklopu magistrske naloge nam je uspelo sintetizirati dve zelo obetavni fluorescenčni sondi za označevanje lipidnih kapljic in lipofilnih struktur v okolici jedra celice (produkt 8 in 13). Spojina 13 je zaradi velikega Stokesovega premika bolj uporabna kot spojina 8, bi pa bilo pri obeh, potrebno ugotoviti še morebitno citotoksičnost. V drugem delu magistrske naloge smo sintetizirali lipofilne ligande, ki smo jih nato pretvorili v komplekse z evropijem, terbijem in/ali disprozijem. Lantanidni kompleksi so zelo dobri fluorofori, predvsem zaradi velikih Stokesovih premikov in zelo ozkih emisijskih spektrov. Pri delu nam je uspelo sintetizirati ker nekaj različnih fluoroforov z lantanidi, vendar smo pri vseh imeli enako težavo. Spojine pri analizi niso bile topne v DMSO in tako tudi niso mogle označevati struktur v celicah. Fluorofori imajo sicer visok kvantni izkoristek, vendar je zanje potrebno najti ustrezno topilo in preučiti njihove lastnosti. Na Institutu Jožef Stefan bodo, v nadaljevanju, komplekse poskušali raztopiti še v absolutnem etanolu in ugotavljali uporabnost naših spojin.

• Jezik: Slovenski jezik
• Ključne besede: fluorescenca, fluorofori, kumarini, lantanidi, lipidne kapljice
• Vrsta gradiva: Magistrsko delo/naloga
• Organizacija: FFA - Fakulteta za farmacijo
• Leto izida: 2022
• PID: 20.500.12556/RUL-143548
• Datum objave v RUL: 24.12.2022

Sekundarni jezik

• Jezik: Angleški jezik
• Naslov: Synthesis and evaluation of fluorescent probes based on coumarin scaffold and lanthanide complexes

Izvleček

Fluorescence detection is a highly sensitive method that is increasingly used in life sciences and research. It is a phenomenon in which the electrons of a fluorophore are excited by light of a specific wavelength to go to a higher excited singlet energy state. After a few nanoseconds, the electrons spontaneously return to their ground state and release excess energy in the form of fluorescence, which we can detect. For fluorescence imaging, we need photostable fluorophores with high quantum yields and high molar extinction coefficients. At the same time, fluorophores must have sufficient specificity for target molecules and large Stokes shifts. Since such fluorophores are rare, further research and discovery of new fluorophores is very important. In our master thesis we focus on the synthesis of fluorescent probes for labeling lipophilic structures. In the first part, we focus on the synthesis of coumarin derivatives, which are of particular interest for the labeling of lipid droplets. Lipid droplets are cellular organelles found in all eukaryotic cells, and much remains to be understood about them. Numerous studies have shown that the accumulation of lipid droplets in the body is related to the occurrence of certain diseases, making research tools such as fluorophores highly desirable. As part of our master's thesis, we were able to synthesize two promising fluorescent probes for labeling lipid droplets (products 8 and 13). Compound 13 is more interesting than compound 8 due to the large Stokes shift, but the potential cytotoxicity of the compounds should be investigated. In the second part of the master's thesis, we synthesized lipophilic ligands, which were then combined with europium, terbium, and/or dysprosium to form complexes. Lanthanide complexes are very good fluorophores, mainly due to their large Stokes shift and very narrow emission spectra. As part of the master's thesis, we were able to synthesize a few different fluorophores with lanthanides, but they all had the same problem. When analyzed, the compounds were not soluble in DMSO and therefore could not label cellular structures. Fluorophores glow well, but it is necessary to find a suitable solvent and study their properties. The Jožef Stefan Institute staff will continue to try to dissolve the complexes in absolute ethanol and analyze the usefulness of our compounds.

• Ključne besede: fluorescence, fluorophores, coumarins, lanthanides, lipid droplets