Synthesis, characterization and investigation of biological properties of non-periferally tetrasubstituted novel phthalocyanines with 2-morpholinoethoxy groups

Phthalocyanines were discovered by chance in 1907, as an impurity during the synthesis of o-cyanobenzamide. In the following years, phthalocyanines used as dyes and pigments, also a lot of study were patented. Linstead and co-workers were pioneers to characterize the chemical structure of phthalocyanine molecule using the X-ray method. The core of phthalocyanine is capable of coordinating more than 70 elements. Different metal ions give unique character and change not only physical properties but also chemical properties. Phthalocyanines do not exist in the nature and are obtained by synthetic methods. They can be regarded as tetrabenzotetraazaporphyrins and as tetramerization products of four iso-indoline units and consist of a planar structure with an 18-π electron system. Phthalocyanines are synthetic macrocycles structurally similar to naturally available porphyrins such as hemoglobin, chlorophyl and vitamin B12. The carbons which located at α position in phthalocyanine structure are named as non-peripheral and others located β position according to nitrile groups are named as peripheral carbons. Almost all early phthalocyanine complexes were unsubstituted on the periphery and had a low solubility in most known solvents even in such high-boiling aromatic solvents. Introduction of substituents can dramatically increase the solubility of the target phthalocyanine in water or common organic solvents and can be improve some properties of phthalocyanines designed for many application. This aromatic macrocyclic compound shows strong absorption bands at visible region due to π-π* transitions. As a consequence, phthalocyanines have dark blue-green colour. This makes them the demanded pigments in dye industry. Phthalocyanines are used as a redox agent and catalyst because of their ease of oxidation and reduction behaviour. Phthalocyanine and related compounds have attracted attention for many decades because of their comprehensive applications. Phthalocyanines find commercial applications as a photoconductor in Xerography and in optical data storage as the laser absorption layer within recordable compact disc. In addition, phthalocyanines are widely studied for applications in non-linear optics, Langmuir-Blodgett films, gas sensors, electrochromic displays and fuel cells. Thanks to their high resistance of light, heat and chemicals, phthalocyanines are prefered for photodynamic cancer therapy. Photodynamic therapy (PDT) uses a combination of a photosensitizing drug and light in the presence of molecular oxygen to obtain a therapeutic effect that depends on selective cell injury. PDT has been developed as an alternative to conventional treatments such as radiotherapy and chemotherapy. Phthalocyanines have received particular attention due to their high molar absorption coefficient in the red part of the spectrum, which allows increased tissue penetration of the activating light. Low toxicity of phthalocyanines makes them promising for PDT application. Both lipophilic and water-soluble phthalocyanines have been considered as candidates for PDT. Therefore, the addition of carboxy, sulphoxy or quarternize amino groups in the phthalocyanine structure makes them hydrophilic. In this study, it is aimed to sythesize novel nonperipherally tetrasubstituted metal-free and metallophthaloyanines (Zn In) and their cationic derivatives bearing 2-morpholinoethoxy groups. The photophysical properties and biological activities of novel phthalocyanines will also be investigated. All of the structures of synthesized compounds are determined using UV-Vis, FT-IR, 1H NMR and mass spectroscopy methods. In the first part of the work, we synthesized 3-(2-morpholinoethoxy)phthalonitrile (1) as starting material. Compound 1 was prepared from 3-nitrophthalonitrile and 2-morpholinoethanol by the nucleophilic displacement reaction in the presence of K2CO3 at room temperature in dry DMSO under N2 atmosphere for 72 hours. After synthesizing compound 1, non-peripherally tetra substituted metal free (2), zinc (3) and indium phthalocyanine (4) derivatives were synthesized. These new phthalocyanines (2-4) are soluble in THF, DMF, DMSO, and chloroform. In the IR spectrum of compound 1, stretching vibrations of C≡N, aromatic C-H, aliphatic C-H and C-O-C appeared at 2228, 3084, 2939, 1255 cm-1 respectively. In the 1H-NMR spectrum of compound 1, aromatic protons were observed between 7.67-7.18 ppm. The OCH2 and the NCH2 protons were confirmed around 4.29-3.71 and 2.91-2.62 ppm, respectively. In the IR spectrum of 1 the intense absorption band at 2228 cm-1, corresponding to C≡N vibrations, dissappears after its conversion into the phthalocyanines (2-4). The IR spectrum of 2-4 were similar to each other. The metal-free phthalocyanine (2) showed an additional absorption band at 3283 cm−1 assigned to the NH stretching vibrations. The 1H NMR spectra of compounds 2-4 are consistent with the proposed structure. In the 1H NMR spectrum of metal-free phthalocyanine (2) in CDCl3 the aromatic protons, the OCH2 and the NCH2 protons resonated between 7.74-7.65, 4.85-3.77 and 3.48-2.97 ppm, respectively. The inner core protons of free-base phthalocyanine (2) were monitored at around -0.96 ppm. In the 1H NMR spectra of ZnPc (3) and InPc (4) in CDCl3 the aromatic protons resonated between 9.02-7.57 ppm for 3 and for 4 between 9.25-7.20 ppm. The OCH2 and the NCH2 protons resonated between 4.92-2.69 ppm respectively. The electronic absorption spectra of 2-4 exhibit a Q band absorption at 691/722, 699 and 729 nm, respectively. B bands of these phthalocyanines appear in the UV region at about 328-321 nm. In the mass spectra of compounds 2-4, the presence of the characteristic molecular ion peaks at m/z = 1031.03 [M]+ for the phthalocyanine 2, m/z = 1094.74 [M]+ for 3, and m/z = 1178.42 [M]+ for 4 confirmed the proposed structure. In the second part of this study, in order to obtain water-soluble phthalocyanine compounds (2Q-4Q), new phthalocyanine complexes (2-4) were aimed to be converted into cationic derivatives. Water-soluble metal-free (2Q), zinc (3Q) and indium (4Q) phthalocyanines were obtained from the reaction of corresponding phthalocyanines (2-4) with the excess of methyl iodide in CHCl3 at reflux temperature for 4 h. All of these new compounds (2Q-4Q) were washed successively with hot ethanol, ethyl acetate, THF, chloroform, hexane and diethyl ether. The compounds (2Q-4Q) prepared in this study show excellent solubility in DMF, DMSO and water as expected. The IR spectra of the phthalocyanines 2Q- 4Q are very similar to each other. Aromatic CH, aliphatic CH and C-O-C vibrations were observed about 3018-3007, 2947-2869 and 1235-1217 cm−1, respectively. The metal-free phthalocyanine (2Q) showed an additional absorption band at 3283 cm−1 assigned to the NH stretching vibrations. In the 1H NMR spectrum of metal-free phthalocyanine (2Q) in d6-DMSO the aromatic protons resonated between 9.22-8.04 ppm. The OCH2 and NCH2 protons were observed between 5.45-3.71 ppm. The inner core protons of free-base phthalocyanine (2Q) and the N+-CH3 protons were monitored at -0.13 and 3.67 ppm, respectively. In the 1H NMR spectra of zinc (3Q) and indium (4Q) phthalocyanines in d6-DMSO the aromatic protons of the Pc core resonated between 9.19-7.97 ppm for 3Q and for 4Q between 9.19-7.43 ppm, integrating for 12 protons for each complex The OCH2 and NCH2 protons of tetracationic phthalocyanines (3Q, 4Q) were monitored between 5.59 and 3.43 ppm. One additional signal for N+-CH3 protons was observed as a singlet at 3.70 ppm for 3Q and 3.69 ppm for 4Q respectively. A typical UV–Vis spectrum of the quaternized tetrasubstituted metal-free (2Q) and metallophthalocyanines (3Q, 4Q) in DMF showed Q bands at 697/724 nm for 2Q, 698 nm for 3Q and 717 nm for 4Q. B band region was observed around 318-325 nm. In the last part of this work, the photophysical properties of compounds 2-4 and biological properties of the cationic phthalocyanines (2Q-4Q) were investigated. The binding constants, the disappearance of quenching effect of all quaternized phthalocyanines on the fluorescence intensity of SYBR Gold-DNA complex and the changes in Tm of double helix DNA with thermal denaturation profile were investigated by UV-Vis and fluorescence spectrophotometric methods. To investigate the spontaneity of the reactions between DNA and novel quaternized Pcs (2Q-4Q) in buffer, thermodynamic parameters were employed.