The interaction of meso-tetrakis(p-sulfonatophenyl)porphyrin (TSPP) sodium salt to human serum albumin and beta-lactoglobulin was studied by steady-state and dynamic fluorescence at different pH of aqueous solutions. The formation of TSPP J-aggregates and a noncovalent TSPP-protein complex was monitored by fluorescence titrations, which depend on pH and on the protein nature and concentration. The complex between TSPP and protein displays a heterogeneous equilibrium with large changes in the binding strength versus pH. The large reduction of the effective binding constant from pH 2 to 7 suggests that electrostatic interactions are a major contribution to the binding of TSPP to the aforementioned proteins. TSPP aggregates and TSPP-protein complex exhibit circular dichroism induced by the presence of the protein. Circular dichroism spectra in the ultraviolet region show that the secondary structure of both proteins is not extensively affected by the TSPP presence. Protein-TSPP interaction was also examined by following the intrinsic fluorescence of the tryptophan residues of the proteins. Fluorescence quenching by acrylamide and TSPP itself also point to small changes on the protein tertiary structure and a critical distance R(0) approximately 56 A, between tryptophan and bound porphyrin, was estimated using the long distance Förster-type energy transfer formalism.

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The photophysical properties of Piroxicam, a nonsteroidal anti-inflammatory drug (NSAID), were investigated at different pHext values in reversed micelles of Aerosol-OT (AOT) in iso-octane, using both steady-state and picosecond time-resolved fluorescence spectroscopy. In contrast with the very complex data obtained in aqueous media, where several prototropic species are in equilibrium, the reversed micellar system essentially favors two species. The absorption spectra shows only one isosbestic point at lambda = 348 nm. Excited-state intramolecular proton transfer (ESIPT), also detected in water, is promoted at low water pool contents measured by omega 0 = [H2O]/[AOT]. A strongly shifted (lambda em = 470 nm) tautomeric emission is found. Upon the gradual increase of omega 0, striking differences with pHext are found. At pHext = 4, the drug preferentially locates itself in the interfacial region partitioning between a hydrophobic and a hydrophilic domain. Global analysis was applied to the decay data and the results were interpreted by the "two-state excited-state" formalism. At pHext = 7, the anionic species is prevalent and the probe locates itself deeper inside the water core of the reversed micelles. Thus, a strong dependence on water content is detected, approaching a behavior similar to that observed in free aqueous solutions.

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Free base porphyrin (PPhe), derivatized with aminosulfonyl groups linked to the aromatic amino acid phenylalanine at the meso-positions, was mixed with DMPC vesicles. The resulting interaction was studied by absorption, steady-state and transient state fluorescence, at different pHs. At pH=2 to pH=9, the aforementioned porphyrin predominates as an aggregated species, with a co-facial arrangement of the molecules taking into account the blue shift of the Soret band (414 nm for the monomer and 401 nm for the aggregate). Upon interaction with DMPC vesicles, the competing hydrophobic interactions with the bilayer destabilize the aggregated species in favor of monomer incorporation. Fluorescence lifetimes also show that the long component assigned to the monomer contributes only 30% to the overall decay in solution (e.g. pH=7.0) whereas in DMPC vesicles this contribution increases up to 85% independent of the solution pH, which confirms a location of the probe in an environment "protected" from free water. The picture changes completely in the case of TSPP, an anionic porphyrin which does not incorporate in DMPC vesicles. Remarkably, at pH=2.5 all the experimental findings point to the self-assembling of the porphyrin units in J-aggregates induced at the surface of the DMPC vesicle. In fact, upon removal of the aqueous solvent, we could define by fluorescence lifetime imaging microscopy (FLIM) regions where the fluorescence lifetime is that characteristic of the J-aggregate ( 0.11 ns).