Recognition of C60 by tetra- and tri-quinoxaline cavitands

Abstract Molecular recognition of C60 fullerene by two cavitands, bearing four and three quinoxaline walls, is reported here. Fluorescence titrations show the recognition ability of the receptors with high binding constant values. The formation of a stable 1:1 supramolecular complex was also confirmed by DOSY, EI-MS and X-ray analysis. The crystal structure analysis of tetraquinoxaline cavitand and C60 shows a crystal packing, where C60 molecules are intercalated between bilayers of cavitand molecules, on the ideal fourfold symmetry axis of the receptor, and are arranged in a zigzag motif.


Introduction
Since the discovery of fullerenes (1), the interest in their chemistry, and in particular in their supramolecular complexes, has been increasing due to the potential applications in biology and in materials science (2). Supramolecular nanohybrid structures based on fullerenes (as electron acceptor) and porphyrins (3)(4)(5), tetrathiafulvalenes (TTFs) (6,7), phthalocyanines (8,9), ferrocenes or oligomers (as electron donors) (10)(11)(12) are used in electron-and energy-transfer devices (photovoltaic cells) able to mimic the photosynthetic system (13). In this context, the search of molecular receptors able to form stable supramolecular complexes with fullerenes in solution and in solid phase is a very active research field (14). Nowadays, the target is to develop supramolecular complexes in order to have an efficient electronic communication between the two components (14). This requires a careful design of the two supramolecular donor and acceptor structures, which also need to show appropriate physical properties. A wide range of synthetic 'hosts' has been explored: calix [n] arenes (15,16), cyclotriveratrylenes (17,18), corannulenes (19)(20)(21), cyclic paraphenyleneacetylenes (22), π-extended with a notably high binding constant of up to 10 5 M −1 . Fluorescence titration data, NMr diffusion and EI-MS analysis clearly indicate a strong affinity of the cavitands towards C 60 fullerene. Noteworthy, crystals suitable for X-ray single-crystal diffraction of tetraquinoxaline cavitand and C 60 confirmed the ability of this host to efficiently form 1:1 supramolecular complexes with C 60 fullerene.

Results and discussion
Cavitands 1 and 2 were synthesised starting from resorcinol, which in presence of HCl leads to Octol macrocycle in high yield (see Scheme 1) (39).
reaction of Octol with an excess of 2,3-dichloroquinoxaline, in the presence of K 2 CO 3 , affords tetraquinoxaline cavitand 1 (39). The triquinoxaline cavitand 2 was obtained with the high yield of 61% through selective excision of one quinoxaline unit by following the procedure reported tetrathiafulvalene derivatives and porphyrins (23)(24)(25). Cavitands represent an intriguing alternative as host materials. They are robust molecules with a bowl shape consisting of four resorcinol rings linked by four methylene bridges (26)(27)(28)(29)(30)(31)(32)(33)(34). An important possibility is the ad hoc tailoring of the cavity wall size, which for instance can be enlarged by the insertion of quinoxaline units, making the cavity deeper and at the same time more rigid and lipophilic. By this means, the cavity can provide π-electron-rich regions able to develop CH-π and π-π stacking, thus facilitating the recognition of complementary organic molecules (35)(36)(37).
To the best of our knowledge, only one example of cavitand derivative able to recognise efficiently fullerene C 60 has been reported by rebek et al. with a K b of 900 M −1 (38).
by Tunstad et al. in the presence of catechol and caesium fluoride (39). The compounds were characterised by NMr and ESI-MS (see Supplementary Material, S2-S4).
Quinoxaline cavitands are particularly interesting due to their ability in solution to undergo a reversible thermal or pH-induced switch from the vase to the kite conformation. This equilibrium is essential to guarantee the aptitude to include a wide range of guests inside the cavity (40,41). As previously reported by rebek et al., tetraquinoxaline cavitand bearing four undecyl-aliphatic chains in the lower rim adopts the 'vase conformation' in toluene-d 8 solution, as indicated by the chemical shift of methine hydrogens in the 1 H NMr spectrum (6.05 ppm) (38). In our case, in both cavitand receptors, the chemical shift of methine protons is near to 5.6 ppm, indicating a 'vase-like' conformation, probably due to the shorter aliphatic chains.
Fluorescence titration of cavitand 1 with C 60 in toluene leads to a progressive quenching of the fluorescence with a gradual red-shift of the maximum of emission ( Figure  1). By fitting the quenching profiles using HypSpec (42), a software designed to extract equilibrium constants from potentiometric and/or spectrophotometric titration data, a binding constant value of log K b = 5.24 M −1 was determined for the 1:1 stoichiometry (see Supplementary  Material, S9).
Similarly, the addition of C 60 leads to a quenching of the fluorescence also in the case of the cavitand 2, from which we derived a binding constant of log K b = 4.95 M −1 ( Figure 2).
In both cases, the strong quenching of fluorescence emission by the addition of C 60 is due to an energy-transfer process between quinoxaline cavity and fullerene (30). In fact, a FrET process is excluded by the absence of the characteristic emission of fullerene. It means that, in solution, a stable supramolecular complex between quinoxaline cavitands and C 60 occurs.
The formation of supramolecular complex between cavitand 1 and C 60 was also investigated by Diffusion-ordered spectroscopy (DOSY) experiments (see Supplementary Material, S6-S7). DOSY is a powerful NMr technique which distinguishes NMr signals according to the diffusion constant (43). recently, Morris and co-workers developed a new method to correlate the diffusion coefficient obtained by DOSY experiments and the molecular weight of the analyte, in order to reveal the formation of monomers, dimers or supramolecular complexes (44). In our case, in toluene-d 8 solution, receptor 1 gave rise to a diffusion coefficient of 5.01 × 10 −10 m 2 s −1 , corresponding to an experimental molecular weight of 1412, attributable to the receptor with one molecule of toluene (theoretical weight 1420). When 1 equivalent of C 60 was added, the diffusion coefficient decreased to 4.30 × 10 −10 m 2 s −1 , with an experimental molecular weight of 2013, indicating the presence of the supramolecular complex between 1 and fullerene (theoretical weight 2048).  The C 60 establishes π⋯π stacking interactions with two other cavitand molecules and a benzene molecule, as shown in Figure 4(a). Thus, the resulting crystal packing shows that the C 60 molecules are intercalated between bilayers of cavitand molecules, and are arranged in a zigzag motif (Figure 4(b)). At solid phase, the inclusion of C 60 into the quinoxaline cavity was not observed due to the presence of benzene molecules inside the host (45). The limited size of the quinoxaline cavity, compared to the fullerene diameter, precludes the inclusion into the aromatic pocket. However, the possibility to establish many C−H⋯π interactions between aliphatic chains and C 60 is the main driving force in the recognition event. In this system, quinoxaline walls act as 'antenna' moiety for the energy transfer process through cavitand and C 60 , as previously demonstrated in the covalent assembly (30).

Conclusions
recognition properties of fullerene C 60 by two quinoxaline cavitands, bearing four and three aromatic walls, were here reported. Fluorescence titrations, DOSY and EI-MS measurements indicate the formation of a stable 1:1 supramolecular complex, confirmed also by X-ray analysis.
Furthermore, crystal structure analysis of tetra-quinoxaline cavitand and C 60 shows a crystal packing where fullerene molecules are intercalated between bilayers of cavitand molecules, arranged in a zigzag motif. At solid phase, C 60 is stabilized via CH-π interactions by the Furthermore, also EI-MS analysis of a solution containing cavitand 1 and C 60 in equimolar ratio indicated the formation of a stable supramolecular complex, showing a peak at m/z 2048 (see Supplementary Material, S8).

Crystal structure description of 1 with fullerene C 60
To lead a characterisation of the supramolecular assembly also in the solid state, X-ray studies were undertaken. Crystals suitable for the X-ray analysis were obtained by slow evaporation of a benzene solution containing a 1:1 mixture of receptor 1 and C 60 . The asymmetric unit of 1 consists of a cavitand molecule hosting a benzene molecule (solvent of crystallisation) in the cavity, a C 60 molecule and three other benzene solvent molecules (Figure 3(a)). Notably, fullerene is not recognised by the quinoxaline cavity, but the supramolecular complex is due to the stabilisation of C 60 by the aliphatic chains of the host. In fact, the C 60 is placed at the lower rim of the cavitand and it interacts with the methylene groups of the aliphatic chains through C−H⋯π interactions (distance range 3.5-3.7 Å, Figure 3(a)). As shown in Figure 3(b), the C 60 is located on the ideal fourfold symmetry axis of the receptor, at a distance of about 4 Å from the mean plane described by the methine carbon atoms of the cavitand. Thus, the formation of 1:1 supramolecular complex between tetraquinoxaline cavitand and C 60 , suggested by DOSY and EI-MS, was also confirmed by X-ray analysis. detection probe (ID-PFG). EI mass spectra were acquired on a ES-MS Thermo-Finnigan lCQ-DECA using MeOH (positive ion mode).

Synthesis of Octol
According to a known procedure from the literature (39), Octol was synthesised as follows: in a round bottom flask, 16.5 g (0.150 mol) of resorcinol were dissolved in 20 ml of ethanol and 20 ml of HCl conc. The mixture was cooled at 5 °C and 17.1 ml (0.150 mol) of heptanal were added in one hour. The solution was refluxed for 8 h and a solid precipitate was obtained. After addition of water, the precipitate was filtered and washed with water. Octol (15.8 g, 51%) was obtained by crystallization from methanol. 1

Synthesis of 1
According to a known procedure from the literature (39), cavitand 1 was synthesised as follows: 1.40 g (1.65 mmol) of Octol, 2.00 g (10.0 mmol) of 2,3-dichloroquinoxaline and 1.50 g (10.0 mmol) of anhydrous K 2 CO 3 were suspended in 30 ml of dry DMF. The mixture was stirred at room temperature for 8 h, and at 50 °C for 18 h. The reaction was quenched by addition of 60 ml of water, and filtered to obtain a crude solid. Compound 1 (1.10 g, 50%) was purified by chromatography (CHCl 3 :EtOAc 95:5). 1 H NMr (CDCl 3 , 500 MHz) δ 0.92 (t, J = 6.5 Hz, 12H, CH 3 ), 1.29 aliphatic chains, while the inclusion of fullerene into the quinoxaline cavity was not observed. Due to the strong energy-transfer process showed in solution, this mode of supramolecular assembly based on quinoxaline cavitands and C 60 can lead to the realisation of supramolecular nanohybrid structures with potential applications in electronand energy-transfer devices.

General
The NMr experiments were carried out at 27 °C on a Varian uNITY Inova 500 MHz spectrometer ( 1 H at 499.88 MHz, 13 C NMr at 125.7 MHz) equipped with pulse field gradient module (Z axis) and a tuneable 5-mm Varian inverse  in Paratone, as cryo-protectant, was mounted in a loop and flash frozen to 100 K with liquid nitrogen. unfortunately since the crystals of this compound were very small, the intensities of diffraction were weak and of poor quality. Diffraction data of compound 1 were indexed and integrated using MOSFlM (46). Scaling was carried out with SCAlA (47,48). The structure was solved by direct methods using SIr2011 (49) in the triclinic P-1 space group. During the refinement cycle two alkylic chains at the lower rim of the cavitand were found disordered over two positions and refined at equal occupancy. Non-hydrogen atoms with population higher than 0.5 were anisotropically refined (H atoms at the calculated positions) by full-matrix leastsquares methods on F 2 using SHElXl-13 (50). The presence of two angles close to 90° suggested the possibility of a non-merohedral twinning.

UV-vis and fluorescence titrations
Emission spectra were recorded with a Fluoromax 3 (Horiba) spectro-fluorometer. uV-vis absorption spectra were recorded with lambda 2 (PerkinElmer) or Cary5000 (Varian). During the fluorescence titrations, the concentration of the host was kept constant and guest was added in small portions. Data analysis was carried out using HypSpec (version 1.1.33), a software designed to extract equilibrium constants from potentiometric and/ or spectrophotometric titration data. HypSpec starts with an assumed complex formation scheme and uses a least-squares approach to derive the spectra of the complexes and the stability constants. χ 2 test (chi-square) was applied, where the residuals follow a normal distribution (for a distribution approximately normal, the χ 2 test value is around 12 or less). In all of the cases, χ 2 ≤ 10 were found, as obtained by 3 independent measurements sets.

Crystal structure determination
Crystals of compound 1 were of small dimensions and data collections was carried out using synchrotron radiation at the X-ray diffraction beam-line of the Elettra Synchrotron, (Trieste, Italy) employing the rotating-crystal method with the cryo-cooling technique. routinely, the crystal dipped