Ionic interactions of anionic thiacalix[4]arene with cationic porphyrins #

A facile synthesis of 25,26,27,28-tetrakis(alkyloxy) - 2,8,14,20-tetrathiacalix[4]arene-5,11,17,23-tetrasulfonate via ipso substitution of p-tert -butylthiacalix[4]arene tetraalkyl ether with sulfuric acid is described. Ionic interaction of anionic thiacalix[4]arene and cationic porphyrins were quantitatively studied by UV-vis, fluorescence, 1 H NMR and ESI-MS spectrometry. Binding constants were in the range of 10 8 M -1 . Binding with axial ligands was investigated and formation of ternary complexes was recognized with pyridine, 4-methylpyridine and N - methylimidazole. The factors affecting the interaction process including pH, solvents and salts were also examined in detail. The results indicated that the neutral medium (pH = 7) is most favorable for electrostatic interactions.


Introduction
Supramolecular capsules present an important class of architectures that can reversibly accommodate smaller molecules in their cavities. 1The synthesis of molecular capsules, 2 based on ionic interactions between oppositely charged calix [4]arenes (C [4]A) is useful for biochemical applications such as drug encapsulation, transport through cell membrane, drug delivery and so far their use has been explored in the stabilization of reactive intermediates for organic transformations and for catalysis. 3These capsules consist of two or more building blocks that have similar size, complementary functional groups and associate via multiple non-covalent interactions such as H-bonds, 4 metal ligands 5 and ionic interactions.hetero capsules based on functionalized calixarenes (CA), resorcinarenes and other building blocks have been reported. 7Water soluble cationic porphyrins are currently of vast interest because of their possible wide applications such as self-assembled nanostructures with welldefined shapes and dimensions, in photochemical cleavage of DNA, in water splitting reactions and as mimic of energy transfer systems. 8Cationic meso-5,10,15,20-tetrakis(N-methyl-4pyridyl)porphyrin has affinity to bind G-quadruplex DNA, has been the focus of a new avenue for the treatment of cancer. 9he CA are third generation host molecules supplementing the crown ethers 10 and cyclodextrins. 11The presence of four sulfur atoms imposes many new properties on the Thiacalix [4]arene (TC [4]A) skeleton as compared to classical C [4]A.Several reactions, 12 unknown in C [4]A chemistry have been described due to the intrinsic properties of sulfur such as coordination ability to metal ions, oxidizability to sulfoxide and sulfone, the larger size, reactivity and increased fluxionality than those of the -CH2 bridged C [4]A skeleton.The covalently linked TC [4]A with porphyrins leads to novel conjugates, which exhibit complexation abilities towards anions, 13 cations 14 or neutral molecules. 15Interaction of C [4]A with porphyrins via covalent bonding, 16 H-bonding 17 and ionic interactions 18 have been well explored but to the best of our knowledge there is no report of ionic interaction between anionic TC [4]A and cationic porphyrins.Thiacalixarenes are essentially insoluble in water, and therefore the introduction of sulfonate functions at the upper rim would enable the extension of their chemistry into aqueous solutions, a requirement for potential biological applications.Although, ipsosulfonation of tetrahydroxythiacalix [4]arene with conc.H2SO4 at 90°C is well known, 19 there is no report for the ipso sulfonation of tetraether derivative of TC [4]A.

SO 3 Na
As can be seen from Figure 2, the aromatic protons showed non-bonded interaction with only tert-butyl protons.If the compound exists in 1,3-alternate form, there would have been some type of interaction between aromatic and butyl chain protons.Thus, keeping this fact, it was confirmed that compound 3a exists only in cone conformation and not in the 1,3-alternate form.Similar observations were made with 3b.

Ionic interaction studies
UV-vis spectroscopy.The influence of anion-cation interactions on the structure and electronic absorption spectra of cationic porphyrins and anionic thiacalix [4]arenes were investigated by absorption spectroscopy.Solutions of porphyrins (1a-1g, 2.0 × 10 -7 M) in water (2.5 mL) were titrated with increasing volumes of anionic thiacalix [4]arenes 4. Stepwise addition of compound 4b to porphyrins 1a-1g resulted in pronounced UV-vis spectral changes.A typical example is shown in Figure 3. Thus, the intensities of the Soret band at 428 nm (corresponding to the porphyrin B transition) and visible bands (corresponding to the porphyrin Q transitions) of the porphyrins gradually decreased upon increasing the concentration of the thiacalix [4]arene 4b and new bathochromically shifted (by 3-8 nm) B-and Q-bands appeared.At the same time two clear isobestic points were observed at 390 nm and 438 nm in the Soret region.The 1:1 stoichiometry was observed for the porphyrin:thiacalix [4]arene assemblies by method of continuous variation.
The absorption spectroscopic titrations gave the apparent binding constants (Table 1).From Table 1, it is clear that metal free porphyrins strongly bind with TC [4]A.This is presumably due to the greater flexibility of the metal free porphyrins vs the metallated porphyrins.Metal centers were available for ligand binding as well as for pH effects.Porphyrin 1g strongly binds with 4b relative to porphyrins 1a and 1c because in 1a and 1c, the charges are in the rings while in 1g, the positive charge on external nitrogen increases the size of the porphyrin and easily available for negative charges of anionic TC [4]A.Porphyrin 1b somewhat strongly binds with 4b than 1d due to the close proximity of 4-pyridyl cationic nitrogen with the anionic part of TC [4]A.Porphyrin 1c exists in four different atropisomeric forms (αααα, αααβ, ααββ, αβαβ) due to the non-symmetric substitution at the meso aromatic rings.Ionic interactions induced by anionic TC [4]A forces all molecules to convert only to the αααα form.On the contrary, the spectral changes of 5,15-bis(N-methyl-4-pyridinium)-10,20-diphenylporphyrin 1e and mono-(4-pyridyl)-triphenylporphyrin 1f with 4b were not sufficient to determine reasonable binding constants.The results indicate that a change of the alkyl chain has almost no effect on the strength of the ion pair complex (entry 1 and 2).In order to exclude the possibility that the observed spectral changes are due to nonspecific aggregation of porphyrins 1a-1g, the UV-vis spectra of porphyrins 1a-1g were measured at various concentrations (between 10 -7 and 10 -6 M in H2O).Aggregation of the porphyrins was not observed at this concentration range.

Ligand binding
The coordination of axial ligands L to the zinc center of assembly, forming ternary complexes was also studied.Zinc porphyrins are known to bind only one axial ligand resulting in a fivecoordinated zinc atom. 20In order to demonstrate the binding ability of the complex, binding studies were performed in H2O with pyridine, 4-methylpyridine and N-methylimidazole ligands.In all the cases, the binding of ligands L with Zn-porphyrins was accompanied by bathochromic shifts and with noticeable decreases in the chromophoric basic absorption bands (Figure 4).Presumably, this was due to the increase of electron density at the zinc cation and at the porphyrin nitrogen atoms.The growth of a fractional negative charge at N atoms establishes the destabilization of the a2u molecular orbital at a1u constant level.The bathochromic shift, probably, was caused by the increase in the energy of a2u -type MO.Upon addition of increasing amount of ligand to the solution of porphyrins 1b and 1d in H2O (2×10 -7 M), characteristic Soret peak at 430 nm of porphyrin changed with 5-7 nm bathochromic shifting.
Porphyrin 1b showed augmented affinity towards N-methylimidazole and 4-methylpyridine than pyridine itself.Moreover, UV-vis titrations of a 1:1 complex of porphyrin and TC [4]A showed the formation of a 1:1:1 complex with axial ligands as confirmed by Job's method.
Association constants for ligand complexation are tabulated in Table 2.In particular, binding of 4-methylpyridine and N-methylimidazole was very strong with complexes.The binding constants for the axial ligand L to porphyrin 1b were determined in a separate experiment and K1b•4b is the formation constant for assembly 1b•4b under the same experimental conditions.The ratio K1b•4b•L/K1b•4b represents the ligand affinity displayed by assembly 1b•4b.The results (Table 2) clearly demonstrate that in general K1b•4b•L / K1b•4b ≤ K1b•L, which indicates that no cavity effect is operating.The binding affinity of ligands towards porphyrin is; Nmethylimidazole > 4-methylpyridine > pyridine.In the case of covalently linked capped porphyrin, 18 there is an attractive interaction between axial ligand and the -wall of calixarene that is why the binding of ligands with covalently linked capped porphyrins was much stronger than with the free porphyrins, but the binding affinity of ligands with porphyrin calix complexes 1b.4b and 1d.4 is  free porphyrins indicate minimal or no cavity effect is there.1.In MeOH, the aggregates hold together very strongly with binding constant of 10 8 M -1 and thus MeOH was the best solvent for the assembly formation, possibly due to a reasonable strong template effect.In polar aprotic solvents like DMSO, the association of 1b and 4b was less relative to other polar solvents.This indicated a different solvation shell around the porphyrin on formation of the complex in this solvent.Complex formation in the presence of salts like NaCl and NaClO4 were also examined (Table 1).On titrations with 4b (9.96 × 10 -7 M to 9.61 × 10 -6 M), in the presence of NaCl and NaClO4 (100 µl, 3 × 10 -4 M, each) in 2.5 mL solution of 1b (2 × 10 -7 M) a red shift of 7-9 nm was observed.Here, the data suggests that at lower salt concentration (resulting in a more ordered solvent shell around the ionic groups), the importance of desolvation in driving the assembly process becomes greater.
pH Effect Cationic porphyrins self assembled in acidic or basis media to form nanowires whereas metal free porphyrins form diacid in the presence of HCl.Thus metal free porphyrins were not suitable for pH effect studies.At different pH (4.0, 7.0, 9.0), titrations of metallated porphyrins 1b and 1f (2.5 mL, 4.2 × 10 -7 M) with 4b (4 × 10 -7 M) were examined.Titration of 1b (2.5 mL, 4 × 10 -7 M) with increasing amount of 4b showed a blue shift of 4 nm at pH 4 (phosphate buffer) whereas a shift of 6 nm was observed at pH 9. Furthermore, at neutral pH, the maximum red shift was 7.5 nm under similar conditions.Results from Table 1 indicate that neutral conditions were most suitable for ionic interaction of porphyrins and 4b.Consistent and reproducible results were obtained with 1d.

Fluorescence spectroscopy
Fluorescence decays of cationic porphyrins in the presence of anionic TC [4]A were obtained upon excitation and emission at 425 and 650 nm, respectively and were best analyzed with a sum of two exponentials.Solutions of porphyrins (1a-1f, 2.0 × 10 -7 M) in water (2.5 mL) were titrated with an increasing volume of 4b (Figure 5).On titrating 1a with increasing volumes of 4b, the peak at 651 nm continuously decreased with arrival of new peak at 613 nm.A clear isobestic point was observed at 640 nm.Similarly, in the case of 1b and 1d, characteristic peak at 600 nm gradually decreased upon titrations of these metalloporphyrins with 4b.It is worth mentioning here that heavy atom effect of iodide as in 5,10,15,20-tetrakis(N-methyl-4pyridyl)porphyrin tetraiodide quenches the fluorescence of porphyrins, reflecting the importance of counter anion in fluorescence spectroscopy.For this reason porphyrins with tosylate rather than iodide counterions were most suitable for fluorescence studies.The emission spectra and intensity of trans-meso-bis(4-pyridyl)-diphenylporphyrin 1e and mono(4-pyridyl)triphenylporphyrin 1f remain unchanged upon addition of 4b, indicating no interaction between 1e, 1f and 4b.Binding constants calculated by fluorescence spectroscopy were in good agreement with those calculated by UV-vis spectroscopy (Table 1, entry 1 to 7).

Conclusions
We have introduced a novel class of supramolecular self-assembly driven by ionic interactions between tetracationic porphyrins and tetraanionic thiacalix [4]arene derivatives in polar solvents.
A detailed spectroscopic analysis revealed the formation of 1:1 ionic complex between two investigated derivatives.The fashioned, 1:1 complexes were further utilized in the formation of 1:1:1 ternary complex by axial coordination of ligands.Spectroscopic findings indicate that anionic TC [4]As strongly bind with cationic porphyrins under different conditions than classical C [4]As which had previously proved successful for ionic interactions.Higher flexibility of metal free porphyrins promote them to strongly bind with anionic thiacalix [4]arene than metallated porphyrins.
We believe that present studies based on ionic interactions will be a welcome addition to TC [4]A chemistry and can be further extended to more complicated supramolecular systems for encapsulation of different guest molecules or in the formation of nanocapsules under physiological conditions.

Figure 6 .
Figure 6.Selected region of 1 H NMR spectra of 1b (2  10 -4 M) in D2O upon addition of 4b at 298 K.The ortho pyridyl proton, meta pyridyl protons and aromatic protons of TCA are designated as blue, magenta and green colors.

Table 1 .
Binding constants (Kass) calculated for ionic interaction between cationic porphyrin 1a-1g and anionic thiacalix[4]arenes 4a and 4b at 25C a Determined by absorption spectroscopic titrations; b determined by fluorescence spectroscopic titrations.Errors are less than  18%; c nd = not determined; d all pH solutions were prepared in phosphate buffer. Ionic interaction studies between classical C[4]A and porphyrins gave lower value of Kass (10 7 M) in MeOH: see ref. 18b.

Table 2 .
Binding constants, calculated for complexation of ligands with porphyrin 1b and assembly 1b.4b