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S ciljem procjene potencijala primjene derivata kaliksarena 5,11,17,23-tetra-tert-butil-25,27- dimetoksi-26,28-bis-((fenantridin-6-il)metoksi)kaliks[4]aren (D) u teranostici, u ovom su radu istražena njegova kompleksacijska svojstva s bojama rodaminom B, auraminom O i fluoresceinom te lijekom timokinonom pri čemu je zbog slabe topljivosti liganda korišten sustav otapala acetonitril/diklormetan. Konstante stabilnosti određene su na temelju provedenih spektrofotometrijskih titracija. Za kompleks [D- rodaminB] lg(K/dm 3 mol -1 ) = 4,55 + 0,01 te lg(K/dm 3 mol -1 ) = 5,61 + 0,05 za kompleks [D-auraminO]. Za kompleks [D-fluorescein] nije bilo moguće odrediti vrijednost konstante stabilnosti iako su eksperimentalni podaci upućivali na to da do stvaranja kompleksa dolazi. Upravo suprotno, s timokinonom kompleksiranje nije opaženo, vjerojatno zbog njegove strukturne nekomplementarnosti s veznim mjestom liganda. Proučavani derivat kaliksarena D dobar je receptor za istraživane boje, pri čemu je najveći afinitet pokazao prema auraminu O. Iako je ustanovljeno da ovaj derivat kaliksarena ima jako dobra receptorska svojstva, on nije topljiv u vodi, što je nužno za primjenu u teranostici, ali se očekuje da bi njegov vodotopivi analog koji bi iskoristio potencijal fenantridinskih skupina u veznom mjestu, mogao pokazati velik potencijal u teranostičkoj primjeni. With the aim of evaluating the application potential of calixarene derivatives 5,11,17,23-tetra-tert- butyl-25,27-dimethoxy-26,28-bis-((phenanthridin-6-yl)methoxy)calix[4]arene (D), in this thesis, its complexation properties with the dyes rhodamine B, auramine O and fluorescein and the drug thymoquinone were investigated. The stability constants were determined based on spectrophotometric titrations and the obtained values were lg(K/dm 3 mol -1 ) = 4.55 + 0.01 for [D-rhodamineB], and for complex [D-auramineO] lg(K/dm 3 mol -1 ) = 5.61 + 0.05. The value of the stability constant for [D-fluorescein] could not be determined, only estimated based on experimental data, while the formation of the complex with thymoquinone was not observed. The results indicate good binding of the studied dyes by the studied calixarene derivative. At the same time, the highest affinity of ligand D towards the formation of a complex was shown for auramine O. Although it was shown that this calixarene derivative has very good receptor properties, it is not soluble in water, which is necessary for its application in theranostics, but it is espected that its water-soluble analogue, that would exploit the potential of phenantridine groups at the binding site, could have great potential in theranostic applications.

S ciljem da se ispita kompleksacijski afinitet nesimetrično supstrituiranog amidnog difenilantracenskog derivata kaliksarena 5,11,17,23-tetra-tert-butil-25-(9,10-difenilantracen-2-iloksietoksi)- 26,27,28-tris(N,N-dietil-karbamoilmetoksi)kaliks[4]arena, L, prema organskim bojama - rodaminu B, auraminu O i fluoresceinu te aktivnoj tvari – timokinonu, proveden je niz spektrofotometrijskih titracija u acetonitrilu. Određene su i konstante stabilnosti odgovarajuć ih kompleksa koje ukazuju na nastanak relativno stabilnih kompleksa derivata kaliks[4]arena L s istraživanim bojama, pri čemu je spoj L pokazao najveći afinitet prema auraminu O. S druge strane, s timokinonom nije došlo do vezanja zbog njegove strukturne nekomplementarnosti s veznim mjestom liganda. Iako kompleksu [L- fluorescein] nije bilo moguće odrediti konstantu stabilnosti, iz eksperimentalnih je podataka bio vidljiv nastanak kompleksa, te je njena vrijednost procijenjena kao niža u odnosu na odgovarajuće konstante za rodamin B i auramin O. Ispitani derivat kaliksarena demonstrirao je dobre receptorske sposobnosti veznog mjesta koje se sastoji od tercijarnih amidnih skupina i difenilantracenske skupine, pa je za očekivati da bi njegov vodotopivi analog imao velik potencijal u teranostici. In order to investigate the complexation affinity of the asymmetrically substituted amide diphenylanthracene derivative of calixarene 5,11,17,23-tetra-tert-butyl-25-(9,10-diphenylanthracene-2- yloxyethoxy)-26,27,28-tris(N,N-diethyl-carbamoylmethoxy) calix[4]arene, L, towards the organic dyes – rhodamine B, auramine O and fluorescein and the active compound – thymoquinone, a series of spectrophotometric titrations in acetonitrile were carried out. The stability constants of the corresponding complexes were determined and their values indicated that relatively stable complexes of calix[4]arene derivatives L with the studied dyes were formed, with compound L showing the highest affinity for the binding of auramine O. On the other hand, the complex with thymoquinone did not form under the experimental conditions used, due to its structural incompatibility with the ligand's binding site. Although the value of the stability constant for [L-fluorescein] complex could not be determined, there was experimental evidence of the complex formation and its value was estimated to be lower than the corresponding constants for rhodamine B and auramine O. Since the studied calixarene derivative demonstrated good receptor properties of the binding site composed of tertiary amide groups and a diphenylanthracene group, it can be expected that its water-soluble analogue should have great potential for applications in theranostics.

U ovom radu dan je teoretski uvid u supramolekulsku kemiju, teranostiku i kaliksarene, te njihovu primjenu. Eksperimentalno je spektrofotometrijskim titracijama istraživano veže li tetrasupstituirani fenantridinski derivat kaliks[4]arena: 5,11,17,23-tetra-tert-butil-25,26,27,28-tetrakis-((fenantridin-6- il)metoksi)kaliks[4]aren (L), odabrane boje: rodamin B, auramin O i fluorescein, s ciljem primjene dobivenih kompleksa za molekulsko označavanje u teranostici i kemoterapeutike: timokinon i paklitaksel, također za primjenu u teranostici. Pokazano je da L kompleksira navedene boje i farmakološki aktivnu tvar timokinon, međutim za paklitaksel nije opažen nastanak kompleksa. Na temelju eksperimentalnih podataka, izračunate su konstante stabilnosti odgovarajućih kompleksa. Prema dobivenim vrijednostima, u korištenim eksperimentalnim uvjetima, afinitet kaliksarenskog derivata L bio je najveći prema vezanju auramina O. Nadalje, vezanje istraživanih boja bilo je efikasnije od vezanja istraživanih lijekova, što je i očekivano s obzirom na njihove strukturne karakteristike i komplementarnost sa strukturom samog liganda Iz svega navedenog zaključeno je da se ovaj kaliksaren, unatoč dobrim receptorskim svojstvima njegovog veznog mjesta kojeg čine fenantridinske podjedinice na donjem obodu, ne može koristiti u teranostici jer nije topljiv u vodi. Međutim, njegov bi analog, s mogućnošću otapanja u vodi, mogao imati izniman potencijal primjene u teranostici. This paper provides a theoretical insight into supermolecural chemistry, theranostics and calixarene and their application. It was experimentally investigated, by means of spectrophotometric titrations, whether the tetrasubstituted phenanthridine derivative of calix[4]arene: 5,11,17,23-tetra-tert- butyl-25,26,27,28-tetrakis-((phenanthridin-6-yl)methoxy)calix[4]arene (L) binds selected dyes: rhodamine B, auramine O and fluorescein, with the aim of applying the resulting complexes for molecular labeling in theranostics and chemotherapeutic drugs: thymoquinone and paclitaxel, also for use in theranostics. It was shown that L forms complexes with these dyes and the pharmacologically active substance thymoquinone, however, the formation of complexwith paclitaxel was not observed. Based on experimental data, the stability constants of the corresponding complexes were calculated. According to the obtained values, under the experimental conditions applied, the affinity of the calixarene derivative L was the highest for the binding of auramine O. Furthermore, the binding of the three studied dyes were generally more efficient than the binding of the investigated drugs, which was expected considering their structural characteristics and complementarity with the structure of ligand itself. Keeping all of the above in mind, it is concluded that this calixarene derivative, despite of good receptor properties of its binding site formed by phenantridin substituents at the lower rim, cannot be used in theranostics because of its insolubility in water. Nevertheless, its water-soluble analog could have exeptional potential of application in theranostics.

Sustavno je istraženo kompleksiranje kationa zemnoalkalijskih metala s derivatom kaliks[4]arena koji na donjem obodu posjeduje dvije fluorescentne fenantridinske podjedinice i dva supstituenta s tercijarnim amidnim skupinama, koji zajedno čine efikasno vezno mjesto za katione. U svrhu detaljnijeg proučavanja utjecaja otapala na ravnoteže procesa kompleksiranja, provedena su istraživanja reakcija u acetonitrilu, metanolu i etanolu. Kompleksiranje je istraženo spektrofotometrijskim, fluorimetrijskim te izotermnim mikrokalorimetrijskim izravnim i kompeticijskim titracijama. Određene su konstante stabilnosti nastalih kompleksa (i iz njih izračunane odgovarajuće standardne reakcijske Gibbsove energije) te standardne reakcijske entalpije i entropije. Proučavani kaliksaren pokazuje iznimno velik afinitet prema zemnoalkalijskim kationima u acetonitrilu, daleko manji u etanolu te još manji u metanolu. U acetonitrilu termodinamički je najstabilniji kompleks s Ca2+, dok u metanolu i etanolu najveća konstanta stabilnosti odgovara kompleksu sa Sr2+. Razlike u afinitetima za vezanje kationa uglavnom su posljedica različite solvatacije vrsta prisutnih u otopini i kompatibilnosti veličine kationa i veznog mjesta liganda. U acetonitrilu su u svim slučajevima i entalpijski i entropijski doprinos standardnoj reakcijskoj Gibbsovoj energiji povoljni, dok su u metanolu i etanolu reakcije kompleksiranja endotermne i entropijski kontrolirane. Istraživani ligand pokazao se vrlo efikasnim i osjetljivim fluorescentnim receptorom za Ca2+ i Sr2+ u acetonitrilu. The complexation of alkaline-earth metal cations with tertiary-amide lower-rim fluorescent calix[4]arene derivative comprising two phenanthridine moieties was studied by means of spectrophotometric, fluorimetric, and microcalorimetric direct and competitive titrations. In order to investigate the solvent effect on cation-binding processes, the complexation reactions were studied in acetonitrile, methanol, and ethanol, whereby the complex stability constants (and derived standard reaction Gibbs energies) as well as standard reaction enthalpies and entropies of complexation reactions were determined. The investigated calixarene exhibited high affinity towards alkaline-earth metal cations in acetonitrile, with low selectivity and peak affinity corresponding to Ca2+, whereas the complex stabilities were significantly lower in ethanol and methanol, in which the affinity for Sr2+ was highest. The differences in cation-binding abilities were a consequence of the differences in solvation of the species present in solution as well in compatibility of the cation and binding-site sizes. The extensive solvent effect could be, at least partially, rationalized by much more favourable cation solvation in methanol and ethanol, as compared to acetonitrile. The binding reactions were enthalpically controlled in acetonitrile, as opposed to methanol and ethanol in which the reactions were endothermic and entropically driven. The investigated ligand was proven to be very effective and rather sensitive fluorescent receptor for Ca2+ and Sr2+ in acetonitrile.

Tehnikom spektrometrije masa uz ionizaciju elektroraspršenjem analizirana su vezna svojstva sedam različitih kaliks[4]arenskih te kaliks[6]arenskih derivata s odabranim kationima lantanoida (La3+, Eu3+, Yb3+). Kompleksiranje izostaje u slučaju esterskih derivata kaliksarena bez obzira na veličinu kaliksarenske šupljine, dok amidni derivati sa svim kationima lantanoida grade stabilne 1:1 komplekse. Provedena je tandemna spektrometrija masa na odabranim ionima kaliksarena i kompleksnih spojeva, predložene su odgovarajuće sheme fragmentiranja te je procijenjena stabilnosti derivata kaliksarena i njihovih kompleksnih spojeva u plinskoj fazi. Pri fragmentiranju izoliranih iona kaliksarenskih derivata i njihovih kompleksnih spojeva uočeno je heterolitičko (induktivno) i homolitičko cijepanje veza. Disocijacija kompleksnih spojeva nije uočena. Analize su provedene na dva različita instrumenta, trostrukom kvadrupolu i ionskoj stupici te su uspoređeni rezultati dobiveni uz različite analizatore masa. Binding properties of seven different calix[4]arenes and calix[6]arenes with selected lanthanide cations (La3+, Eu3+, Yb3+) were investigated by electrospray ionization mass spectrometry. Complexation was not observed in the case of ester calixarene derivatives, regardless of the size of the calixarene cavity, while amide derivatives formed stable 1:1 complexes with all lanthanide cations. Tandem mass spectrometry analysis of the selected calixarene and complex ions was performed, corresponding fragmentation schemes and stability estimation of calixarene derivatives and their complex compounds in the gas phase were suggested. During the fragmentation of the isolated ions of the calixarenes and their complex compounds, heterolytic (inductive) bond cleavage and homolytic bond cleavage were observed. Dissociation of the complex compounds was not observed. The analyses were performed by using two different instruments, triple quadrupole and quadrupole ion trap, and comparisons of the results obtained with different mass analyzers were made.

Sintetizirano je sedam novih fluorescentnih derivata kaliks[4]arena s fenantridinskim (13) i difenilantracenskim (47) supstituentima na donjem obodu kaliksarenskog prstena. Spojevi su pripravljeni konvencionalnom organskom i mikrovalovima potpomognutom sintezom te su uspoređena reakcijska vremena i iskorištenja. Okarakterizirani su na temelju podataka dobivenih 1H i 13C NMR, FTIR spektroskopijom, elementnom analizom kao i spektrometrijom masa. Spektrofotometrijskim, fluorimetrijskim, potenciometrijskim i 1H NMR titracijama istraženo je kompleksiranje dobivenih liganada s alkalijskim kationima u smjesama otapala acetonitril/diklormetan i metanol/diklormetan ( fi= 0,5). Pri tome su određene konstante stabilnosti odgovarajućih kompleksa stehiometrije 1:1 (kation:ligand). Istraženo je i nastajanje ionskih parova u istim smjesama otapala, budući da ono također utječe na proces kompleksiranja. Kao što je očekivano s obzirom na izbor supstituenata, svi ligandi vežu alkalijske katione uz smanjenje afiniteta prema metalnim ionima s porastom njihova radiusa. Opaženo je da otapalo znatno utječe na stabilnost kompleksa, pri čemu je afinitet liganada prema kationima znatno veći u smjesi diklormetana s acetonitrilom nego onoj s metanolom. Može se zaključiti da bi pripravljeni kaliksarenski derivati mogli poslužiti kao osjetljivi fluorimetrijski senzori za alkalijske katione. In the framework of this work seven fluorescent calix[4]arene derivatives with phenanthridine (13) and diphenylanthracene (47) substituents at the lower calixarene rim were synthesized. Compounds were prepared by conventional organic synthesis as well as by microwave-assisted synthesis, and the reaction times and yields were compared. The compounds were characterized by means of FTIR, 1H and 13C NMR spectroscopies, elemental analysis as well as by mass spectrometry. The complexation of these ligands with alkali-metal cations was studied by spectrophotometric, fluorimetric, potentiometric, and NMR titrations in acetonitrile/dichloromethane and methanol/dichloromethane solvent mixtures ( phi= 0.5). The stability constants of the corresponding 1:1 (cation:ligand) complexes were determined. Ion association in these solvent mixtures was also investigated because this phenomenon can considerably affect the complexation process. The ligands were proven to bind the alkali-metal cations rather strongly with lowering of their binding abilities with the increase of the cation radius. It was found that solvent considerably affected the stability of the complexes, whereby the ligands showed significantly higher affinities for cations in acetonitrile than in methanol/dichloromethane mixture. It could be concluded that the prepared calixarene ligands could serve as sensitive fluorimetric sensors for alkali-metal cations.