Chemosensor properties of 7-hydroxycoumarin substituted cyclotriphosphazenes * Dedicated to our supervisor Prof. Dr. Adem Kılıç on his retirement.

The newly synthesized cyclotriphosphazene cored coumarin chemosensors 5, 6, and 7 were successfully characterized by 1 H NMR, 31 P NMR, and MALDI-TOF mass spectrometry. Additionally, the photophysical and metal sensing properties of the targeted compounds were determined by fluorescence spectroscopy in the presence of various metals (Li + , Na + , K + , Cs + , Mg 2+ , Ca 2+ , Ba 2+ , Cr 3+ , Mn 2+ , Fe 3+ , Co 2+ , Al 3+ , Hg + , Cu 2+ , Zn 2+ , Ag + , and Cd 2+ ) . The fluorescence titration results showed that compounds 5, 6, and 7 could be employed as fluorescent chemosensors for Fe 3+ ions with high sensitivity. The complex stoichiometry between final cyclotriphosphazene chemosensors and Fe 3+ ions was also determined by Job’s plots.


Introduction
Fluorescence chemosensor design is the key topic in the determination of ions in biological processes. It is important to examine the fluorescence properties of the compounds to be synthesized as new sensors, their location in living organisms, and their usability for metal ion detection as fluorescent sensors [1][2][3]. Fe 3+ , an indispensable element for life, both provides oxygen-forming capacity and serves as a cofactor in many enzymatic reactions in the mitochondrial respiratory chain, and both deficiency and excess amounts of Fe 3+ can cause various diseases [4]. Deficiency of Fe 3+ ions causes anemia, diabetes, liver damage, Parkinson disease, and cancer. On the other hand, excessive intake of iron in humans is equally harmful and may cause dysfunctions of certain organs, such as the heart, pancreas, and liver [5]. Furthermore, high levels of Fe 3+ ions may be toxic, promoting the oxidation of lipids, proteins, and other cellular components. For this reason, the detection of iron ions in biological media is very important [6]. Coumarin-based fluorescent chemosensors show less toxicity and are easily altered. Furthermore, the carbonyl group of coumarin can partake in coordination with metal ions if necessary. This is an ideal model for the design of chemosensors because the response will be fast and efficient when guests, e.g., protons and metal ions, are bound to the host probes. Recently, coumarins being are used as fluorescent chemosensors due to their high optical activities, high light stability, high quantum yield, wide Stokes shift, and low toxicity properties. These compounds are an important part of organic heterocyclic compounds used in sensor technology [4]. Coumarin compounds demonstrate perfect photophysical properties.
Phosphazenes are composed of repeating units of [-P=N-] in their structure. They also have a cyclic structure connecting two inorganic or organic side groups (R) to each phosphorus atom. Hexachlorocyclotriphosphazene is one of the most important members of the phosphazene family because different products are obtained as a result of various reactions on the six chlorine groups in the structure. Depending on the attached groups, they are suitable for the synthesis of different compounds for various applications [19,20]. During the past two decades, nucleophilic substitution reactions at phosphorus atoms of hexachlorocyclotriphosphazene have been extensively explored, leading to an enormous variety of materials with interesting properties such as anticancer [21] and antimicrobial agents [22] or fluorescence sensors [23]. Although there are many studies about coumarin, there are few studies about cyclotriphosphazenes bearing coumarin groups. Some of these were synthesis and characterization studies [24], as well as fluorescent chemosensor studies [25,26]. In the current study, new coumarin-based cyclotriphosphazenes (5-7) (Scheme), which are fluorescent chemosensors for Fe 3+ ions, have been successfully designed and prepared. The photophysical properties and metal-sensing behaviors of the new compounds (5-7) were investigated by UV-Vis electronic absorption and florescence spectroscopies. Scheme. 7-Hydroxycoumarin substituted cyclotriphosphazenes (5-7).

Experimental
Materials and methods are given in the Supporting information.

Synthesis of compound 5
Hexachlorocyclotriphosphazene (4) (0.5 g, 1.43 mmol) was dissolved with tetrahydrofuran in a 100-mL threenecked round-bottomed flask. After that, NaH (0.35 g, 8.58 mmol) dissolved with 5 mL of THF was added to the reaction mixture. Then 7-hydroxy-3-(4-methoxyphenyl)coumarin (1) (2.29 g, 8.58 mmol) dissolved with 10 mL of THF was added drop-wise for about 20 min and the reaction was heated with stirring at 80 • C under an argon atmosphere for 4 days. After this time, the reaction mixture was filtered off with a G4 filter for removing formed NaCl salts. The solvent was completely removed by evaporation. Finally, the reaction mixture was purified by column chromatography using silica gel as column material and an n−hexane-THF (1:5) solvent mixture as an eluent. Yellowish pure product 5 was obtained (0.62 g, 0.35 mmol, 25%). Anal. Calc. for

Synthesis of compound 6
Hexachlorocyclotriphosphazene (4) (0.5 g, 1.43 mmol) was dissolved with tetrahydrofuran in a 100-mL threenecked round-bottomed flask. After that, NaH (0.35 g, 8.58 mmol) dissolved with 5 mL of THF was added to the reaction mixture. Then 7-hydroxy-3-(3,4-dimethoxyphenyl)coumarin (2) (2.55 g, 8.58 mmol) dissolved with 10 mL of THF was added drop-wise for about 20 min and the reaction was heated with stirring at 80 • C under an argon atmosphere for 4 days. After this time, the reaction mixture was filtered off with a G4 filter for removing formed NaCl salts. The solvent was completely removed by evaporation. Finally, the reaction mixture was purified by column chromatography using silica gel as column material and an n−hexane-THF

Synthesis of compound 7
Hexachlorocyclotriphosphazene (4) (0.5 g, 1.43 mmol) was dissolved with tetrahydrofuran in a 100-mL threenecked round-bottomed flask. After that, NaH (0.35 g, 8.58 mmol) dissolved with 5 mL of THF was added to the reaction mixture. Then 7-hydroxy-3-(3,4,5-trimethoxyphenyl)coumarin (3) (2.81 g, 8.58 mmol) dissolved with 10 mL of THF was added drop-wise for about 20 min and the reaction was heated with stirring at 80 • C under an argon atmosphere for 4 days. After this time, the reaction mixture was filtered off with a G4 filter for removing formed NaCl salts. The solvent was completely removed by evaporation. Finally, the reaction mixture was purified by column chromatography using silica gel as column material and an n−hexane-THF (1:5) solvent mixture as an eluent. Then yellowish pure product 7 was obtained (0.63 g, 0.30 mmol, 21%). Anal. Calc. for

Syntheses and characterizations
7-Hydroxycoumarin derivatives substituted cyclotriphosphazenes (5-7) were synthesized and their fluorescence chemosensor properties were also reported. 7-Hydroxy-3-(4-methoxyphenyl)coumarin (1), 7-hydroxy-3-(3,4-dimethoxyphenyl)coumarin (2), and 7-hydroxy-3-(3,4,5-trimethoxyphenyl)coumarin (3) were synthesized according to the procedures given in the literature [27][28][29]. Compounds 1-3 and hexachlorocyclotriphosphazene (4) were separately reacted in a 6:1 molar ratio in the presence of NaH as a base in THF under an argon atmosphere at room temperature (Scheme). All the products (5-7) were purified by column chromatography and characterized by MALDI-TOF MS, FT-IR, 1 H NMR, 31 P NMR, and fluorescence spectroscopy. The MS, 1 H NMR, and 31 P NMR results for each new compound were compatible with the data given in Section 2. All of the proton decoupled 31 P NMR spectra of compounds 5-7 showed a sharp single peak due to the equivalent chemical environment of all the phosphorus nuclei, suggesting an A 3 spin system. The 31 P NMR spectrum of compound 5 is depicted as an example in Figure 1b.
The 1 H NMR data also confirmed the structures of compounds 5-7. The aromatic protons for all compounds were observed between 8.06 and 6.73 ppm and some of them were distinguishable from each other. The NMR signals of the -OCH 3 groups were observed as a singlet or two singlets between 3.76 and 3.89 ppm.

Electronic absorption and fluorescence behavior
Absorption and emission properties of coumarin-based cyclotriphosphazene compounds were investigated in different solvents at room temperature ( Figures S7-S9). The absorption spectra of the studied compounds (5-7) did not show any changes in the different studied solvents. The absorption bands observed at around 340 nm were attributed to the substituted coumarin groups because cyclotriphosphazenes are optically inert in the UV-Vis region [30]. In addition, the ground state absorption spectra of the compounds in THF/H 2 O (20:1) were measured at different concentrations. From these spectra, the molar extinction coefficients of the compounds (5-7) were calculated according to the Beer-Lambert law (Figures S10-S12). The solvent effects on the fluorescence emission behavior of the studied compounds (5-7) were determined in different organic solvents as well as THF/H 2 O (20:1) solution ( Figures S13-S15). All studied coumarin substituted cyclotriphosphazenes   The fluorescence lifetimes ( τ F ) of coumarin substituted cyclotriphosphazenes (5-7) were also measured using the time-correlated single-photon counting (TCSPC) technique (Figures 4, S18, and S19). The lifetimes were found to be biexponential and the τ F values are given in the Table. Changes in the fluorescence lifetime values indicated the binding of Fe 3+ ions to the studied compounds (5-7).

Chemosensor studies
Spectroscopic changes upon addition of metal salts to the ligand were recorded using a fluorescence spectropho-