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Polysaccharide films containing chitosan, methylcellulose, and a mixture of these polysaccharides in various ratios were prepared and modified with meso-tetrakis(4-sulfonatophenyl)porphyrin in an aqueous medium at pH 7. The modified films were compared with the initial films using spectroscopic methods and microscopic imaging. Electronic (UV-vis absorption, electronic circular dichroism (ECD)) and vibrational (FTIR and Raman) spectra showed that the porphyrin macrocycles had a strong affinity toward chitosan and did not interact with the methylcellulose. The total porphyrin uptake depended on the chitosan: methylcellulose ratio and pure methylcellulose films did not retain porphyrin macrocycles. ECD measurements detected the presence of optically active porphyrin species bound to the films. SEM and AFM images confirmed that the porphyrin macrocycles caused structural changes on the film surface and within the film layer.

Solid meso-tetrakis(4-sulfonatophenyl)porphyrin (TPPS(4))-chitosan supramolecular complexes were prepared by addition of porphyrin to an aqueous solution of chitosan at pH values. The precipitates obtained were assigned as 1 (pH 6.8) and 2 (pH 2.5) and characterized by spectroscopic, thermal, and microscopic methods. Spectroscopic investigation confirmed the presence of TPPS(4) and chitosan in both products and that the porphyrin is highly self-associated. H-type (stacked) of TPPS(4) aggregation was proposed for 1 and J-type (tilted) for 2. Thermal analysis revealed different pyrolysis routes of the complexes depending on their structural diversity. Light microscopic analysis indicated fibrous and lamellar microstructures, respectively, for 1 and 2. SEM and AFM analysis showed that both complexes consist of compact nanostructures; their size and interconnection is different for 1 and 2. Based on structural inferences, self-assembling hierarchy models were proposed for both of the TPPS(4)-chitosan supramolecular complexes.

Interaction of meso-tetrakis(4-sulphonatophenyl)porphine (TPPS4) with chitosan (Mr approximately 400 kDa, N-acetyls approximately 20 mol.%) was studied in aqueous solutions. UV-vis absorption and circular dichroism (CD) spectroscopic titration of 10 micromol l-1 TPPS4 with chitosan demonstrated that an addition of the polysaccharide at appropriate concentrations and pH values induce and support self-aggregation of the macrocycles. The mode of aggregation was strongly dependent on pH: stacking (H-type) aggregates predominated at weak acidic conditions (pH 4.8-6.8) and tilted (J-type) aggregates at pH 2.5. At the intermediate pH value (3.6) both types of TPPS4 aggregates were detected. High amount of chitosan (>0.05 mmol l-1 of GlcN) disrupts H-aggregates forming monomeric porphyrin-chitosan complexes (pH 3.6-6.8), while J-aggregates (pH 2.5) are stable even at very high chitosan concentrations. CD titration experiments confirmed the formation of optically active species of TPPS4 in the presence of chitosan. The complex nature of CD bands assigned to both types of porphyrin aggregates indicated the occurrence of several chiral macrocyclic species dependently on pH value and chitosan concentration.

In the present study we investigated the photosensitizing properties of two novel mono- and bis-cyclodextrin tetrakis (pentafluorophenyl) porphyrin derivatives in several tumor cell lines and in BALB/c mice bearing subcutaneously transplanted syngeneic mouse mammary carcinoma 4T1. Both studied sensitizers were localized mainly in lysosomes and were found to induce cell death by triggering apoptosis in human leukemic cells HL-60. In 4T1 and other cell lines both apoptotic and necrotic modes of cell death occurred depending on drug and light doses. Mono-cyclodextrin porphyrin derivative P(beta-CD)1 exhibited stronger in vitro phototoxic effect than bis-cyclodextrin derivative P(beta-CD)2. However, in vivo P(beta-CD)2 displayed faster tumor uptake with maximal accumulation 6 h after application, leading to complete and prolonged elimination of subcutaneous tumors within 3 days after irradiation (100 J cm(-2)). In contrast, P(beta-CD)1 uptake was slower (48 h) and the reduction of tumor mass was only transient, reaching the maximum at the 12 h interval when a favorable tumor-to-skin ratio appeared. Thus, P(beta-CD)2 represents a new photosensitizing drug displaying fast and selective tumor uptake, strong antitumor activity and fast elimination from the body.

Biolocalisation and photochemical properties of novel macrocyclic photosensitisers, guanidiniocarbonyl-substituted tetraphenylporphyrin (1) and sugar-substituted sapphyrin (2) were investigated by spectroscopic methods. Both photosensitisers absorb in far visible region and showed good tumour localisation. Photosensitiser 2 demonstrated significantly larger absolute and relative to normal tissue (T/N) amount in tumour (330 microg g(-1) wet tissue, T/N=19.0) than photosensitiser 1 did (13 microg g(-1) wet tissue, T/N=2.1). According to iodometric and uric acid assays, compound 1 produced large amount of 1O2 (phidelta=0.60-0.68), while compound 2 showed non-significant 1O2 production (phidelta=0.04). The electronic spectroscopic study confirms that only photosensitiser 1 is able to mediate photooxidation of model compounds (BSA, poly(Trp), Tyr, Trp, and GMP) after light irradiation. Pour photochemical activity of compound 2 was explained by its self-aggregation. Raman spectroscopic study indicated that monomerised photosensitiser 2 effectively damaged BSA and calf thymus DNA after light excitation at the conditions of high excess of these macromolecules.

Six water-soluble macrocyclic photosensitizers, the members of two groups of expanded porphyrins (metallotexaphyrins and free-base sapphyrins) containing hydrophilic substituents and meso-tetra(4-sulfonatophenyl)-porphyrin, were tested by UV-Vis absorption and resonance Raman spectroscopy in the in vitro binding experiments with intact mitochondria isolated from swine liver. Studied macrocycles showed markedly different affinity to mitochondria. The highest uptake was observed for sapphyrin-sugar conjugate and metallotexaphyrins. Sapphyrin-polyamine conjugates exhibit something less affinity to mitochondria, while the porphyrin of anionic character showed very low mitochondrial uptake. Obtained spectroscopic results confirm that the binding process altered the self-aggregation degree of expanded porphyrins.

The synthesis, characterization, and biolocalization properties of new, fully water-soluble sapphyrins is reported. These systems, which display light absorbing features that make them of potential interest as photodynamic therapy (PDT) sensitizers, were found to localize selectively in pancreatic carcinoma tissue in a xenographic murine model. In this model, the concentration ratios for malignant tissue to surrounding muscle tissue were found to be as high as 280 in the most favorable case. The concentrations of the water-soluble sapphyrins reported here, as well as a porphyrin control system, were determined as a function of time and tissue type.