Synthesis and bactericidal properties of porphyrins immobilized in a polyacrylamide support: influence of metal complexation on photoactivity.

Spectroscopic and photodynamic properties of three novel polymeric hydrogels bearing porphyrins have been studied in vitro on the recombinant bioluminescent Gram-negative Escherichia coli DH5α to assess their ability to inactivate bacterial strains in solution. The three different hydrogels were formed by polymerization of 5-[4-2-(2-(2-acrylamidoethoxy)ethoxy)ethyl]carboxyphenyl-10,15,20-tris(4-N-methylpyridyl)porphyrin trichloride (5) and its complexes with Pd(ii) (6) and Cu(ii) (7) respectively, to form three optically transparent polyacrylamide hydrogels. All of the porphyrins are tricationic and they bear at the meso positions three N-methylpyridyl rings and one terminal acryloyl group connected through a flexible hydrophilic linker, particularly suitable for the later polymerization and incorporation into a hydrogel. The hydrogels were characterized by IR and scanning electron microscopy and incorporation of the dye was confirmed by UV-visible spectroscopy. All the hydrogels are characterized by a non-ordered microporous structure. The E. coli exhibited a decrease of 1.87 log after 25 min irradiation when the porphyrin hydrogel 9 was evaluated. When the Pd(ii) and Cu(ii)porphyrin hydrogels were tested (10, 11), they showed a 2.93 log decrease and 1.26 log decrease in the survival of the E. coli after 25 min irradiation, respectively. Similar results were obtained when the porphyrins were tested in solution. Of the three hydrogels, the Pd(ii)porphyrin hydrogel (10) proved to be the one with the highest photokilling ability under illumination, and also exhibited the lowest toxicity in the absence of light. Hydrogels 9 and 10 were found to be active for five cycles, suggesting the possibility of reuse.

Synthesis, characterization and biological evaluation of a new photoactive hydrogel against Gram-positive and Gram-negative bacteria.

In 2013, the World Health Organization reported that 884 million people lack access to clean potable water. Photodynamic antimicrobial chemotherapy (PACT) is a very promising alternative to conventional antibiotics for the efficient inactivation of pathogenic microorganisms. We report the synthesis, characterization and antibacterial activity of a polyacrylamide-based hydrogel (7), with a new photoactive phenothiazinium compound (6) immobilized on it, to be used as a novel water-sterilizing device. The hydrogel was characterized by IR and scanning electron microscopy and incorporation of the dye confirmed by UV-visible spectroscopy. Antibacterial tests using the recombinant bioluminescent Gram-positive Staphylococcus aureus RN4220 and Gram-negative Escherichia coli DH5α were performed to assess the ability of the hydrogel to inactivate bacterial strains in solution. The hydrogel is characterized by a non-ordered microporous structure and is able to generate reactive oxygen species. The hydrogel is able to inactivate planktonic cells of the S. aureus and E. coli (3.3 log and 2.3 log killing, respectively) after 25 min of irradiation with white light at 14.5 mW cm-2. The contact surface does not influence the kill rates while the killing rate increased by increasing the total amount of the hydrogel (0.27 log drop to 1.65 log drop with 0.5 mg cm-3 to 2.5 mg cm-3 of total amount of dye). The hydrogel was found to be active for four cycles, suggesting the possibility of reuse and it was shown to be active against both Gram-positive and Gram-negative species with no leaching of the active molecule.

Immobilized photosensitizers for antimicrobial applications.

Photodynamic antimicrobial chemotherapy (PACT) is a very promising alternative to conventional antibiotics for the efficient inactivation of pathogenic microorganisms; this is due to the fact that it is virtually impossible for resistant strains to develop due to the mode of action employed. PACT employs a photosensitizer, which preferentially associates with the microorganism, and is then activated with non-thermal visible light of appropriate wavelength(s) to generate high localized concentrations of reactive oxygen species (ROS), inactivating the microorganism. The concept of using photosensitizers immobilized on a surface for this purpose is intended to address a range of economic, ecological and public health issues. Photosensitising molecules that have been immobilized on solid support for PACT applications are described herein. Different supports have been analyzed as well as the target microorganism and the effectiveness of particular combinations of support and photosensitizer.

Towards matched pairs of porphyrin-Re(I) /(99m) Tc(I) conjugates that combine photodynamic activity with fluorescence and radio imaging.

We recently prepared two novel water soluble porphyrins bearing a single peripheral chelator, either diethylenetriamine (1) or bipyridyl (2), tethered to one meso position. The preparation of their conjugates with a fac-{(99m) Tc(CO)3 }(+) fragment and the potential of these resulting conjugates as fluorescence and radio imaging tools were also described. In this work, we focused on the corresponding non-radioactive analogues that bear the fac-{Re(CO)3 }(+) fragment (diethylenetriamine 3 and bipyridyl 4). We report on the uptake, in vitro PDT activity, and cellular localization of Re(I) conjugates 3 and 4 in comparison to the parent porphyrins 1 and 2. Compounds 1-4 have modest or negligible cytotoxicity in the dark against HeLa human cervical cancer cells but become remarkably cytotoxic after exposure to moderate doses of red visible light (590-700 nm). This phototoxicity was found to be directly proportional to the total light dose. Although the four compounds show distinct uptake patterns, they have comparable PDT activity. Confocal fluorescence measurements showed that porphyrin 1 and its Re(I) conjugate 3 have different cellular localization patterns in HeLa cells.

Novel water-soluble (99m)Tc(I)/Re(I)-porphyrin conjugates as potential multimodal agents for molecular imaging.

The synthesis and characterization of two novel water soluble porphyrins with three meso pyridyl rings and one peripheral chelator - either a diethylenetriamine unit (4) or a bipyridyl fragment (8) - for binding to the {(99m)Tc(CO)3}(+) moiety is reported. In 8, despite the presence of a flexible and hydrophilic PEG-like linker that connects the bpy unit to the macrocycle, good water solubility was only obtained by methylation of the pyridyl N atoms that provided three extra positive charges. Furthermore, the water-soluble conjugates of 4 and 8 with either one fac-{Re(CO)3}(+) (9 and 10, respectively) or one fac-{(99m)Tc(CO)3}(+) fragment (9a and 10a, respectively) are described.

Ruthenium-porphyrin conjugates with cytotoxic and phototoxic antitumor activity.

We report here two novel "extended-arms" porphyrins, TetbpyPP and TedabpyPP, in which four peripheral bpy fragments are connected to the meso positions of the macrocycle through flexible linkers of different length and hydrophilicity. We describe also the new, water-soluble, tetracationic conjugate [TedabpyPP{Ru([9]aneS3)Cl}(4)][Cl](4) (6). Compound 6 belongs to the series of cationic Ru-porphyrin conjugates 1-5, each bearing four peripheral Ru(II) half-sandwich coordination compounds, that we recently prepared as potential photosensitizing chemotherapeutic agents. The in vitro cell growth inhibition of conjugates 1-6 toward MDA-MB-231 human breast cancer cells and HBL-100 human nontumorigenic epithelial cells are reported, together with the phototoxic effects of 1, 4, and 6 on MDA-MB-231 cells. All conjugates have IC(50) values in the low micromolar range that decrease by 1 order of magnitude upon irradiation of cell cultures with visible light. The most promising compounds 1 and 6 are phototoxic at low light and drug doses.

Synthetic strategies towards ruthenium-porphyrin conjugates for anticancer activity.

The conjugation of porphyrins to metal fragments is a strategy for making new compounds that are expected to combine the phototoxicity and the tumour-localization properties of the porphyrin chromophore with the cytotoxicity of the metal fragment for additive antitumour effect. We report here the preparation of new classes of porphyrin-ruthenium conjugates with potential bio-medical applications. Ruthenium was chosen because several Ru compounds have shown promising anticancer activity. The conjugation with the porphyrin moiety was accomplished either through peripheral pyridyl rings (e.g.meso-4'-tetrapyridylporphyrin, 4'TPyP) or through bpy units (e.g.meso-(p-bpy-phenyl)porphyrins, bpy(n)-PPs, n = 1-4). The number of Ru fragments attached to the porphyrins ranges from 1 to 4 and the total charge of the conjugates from -4 to +8. Different types of peripheral fragments, both Ru(III) and Ru(II), have been used: in some cases they are structurally similar to established anticancer compounds. Examples are [Na](4)[4'TPyP{trans-RuCl(4)(dmso-S)}(4)] (2), that bears four NAMI-type Ru(III) fragments, or [4'TPyP{Ru([9]aneS3)(en)}(4)][CF(3)SO(3)](8) (3) and [bpy(4)-PP{Ru([9]aneS3)(dmso-S)}(4)][CF(3)SO(3)](8) (9) (en = ethane-1,2-diamine, [9]aneS3 = 1,4,7-trithiacyclononane) that have four half-sandwich Ru(II) compounds. The Ru fragments may either contain one or more labile ligands, such as in 2 or in 9, or be coordinatively saturated and substitutionally inert, such as in 3 or in [bpy(4)-PP{Ru([12]aneS4)}(4)][CF(3)SO(3)](8) (11) ([12]aneS4 = 1,4,7,10-tetrathiacyclododecane). Most of the ruthenium-porphyrin conjugates described in this work are soluble--at least moderately--in aqueous solution and are thus suitable for biological investigations, in particular for cytotoxicity and photo-cytotoxicity tests.