RICERCA ITALIANA     

Research program

Nanoscale self-assembled porphyrin based complexes: properties and technological applications

University Co-ordinator

Università degli Studi di MESSINA - CHIMICA INORGANICA,CHIMICA ANALITICA E CHIMICA FISICA - ()

Research Unit Leader

Luigi Monsù Scolaro

Description

The research program of the Messina Research Unit (RU) is expected to continue and expand previous investigations on self-aggregation and supramolecular assembling of porphyrins on templating polymers. The earlier activity has been supported in the last years thanks to MIUR grants (Cofin 2002). Our previous results will constitute the starting point to search for potential applications in opto-electronics and in drug-delivery. The aim of this project will be: (i) to extend kinetic and structural investigations on porphyrin aggregates to get a control of their size, especially on the nanoscale, and of their photophysical properties in order to test their potential applicability in opto-electronics, (ii) to design and investigate non-covalent supramolecular systems as sensors, and (iii) to study the interaction of porphyrins with model proteins and vesicles, formed by amphiphilic cyclodextrins, to investigate their potential applications as delivering systems in PDT applications.
In particolar, the research can be subdivided in the following lines:
1) Structural and kinetic investigation on porphyrin nano-aggregates.
This subject is the logical prosecution of the previous supported proposal. In order to get deeper insights into the factors controlling aggregation (concentration, ionic strength, pH and temperature), detailed investigations on the self-aggregation of water solubile porphyrins and their supramolecular assembling on small biologically relevant matrices have been carried out. In this context, we focused our attention on two specific porphyrins, i.e. tetrakis-(4-sulfonatophenyl)porphyrin (TPPS4) and trans-bis(N-4-methylpyridinium)diphenylporphyrin (trans-H2Pagg). The diacid form of the anionic porphyrin TPPS4 aggregates at acidic pH and high ionic strength, leading to J-aggregates. These aggregates are characterized by a lateral arrangment of porphyrins, which form a linear array, stabilized through strong electrostatic interactions between the peripheral sulfonate groups and the protonated core in the macrocycle, apart from hydrogen bonding and van der Waals forces. The strong electronic coupling between porphyrins determines relevant effects both on absorption bands (bathochromic shift larger than 50 nm for the visibile bands) and photophysical properties. Furthermore, these aggregates exhibit “resonance light scattering” (RLS), that is responsible for a large ehnancemment of the scattered light in proximity of an absorption band.[1] The combined use of various spectroscopic techniques (UV/vis, fluorescence and light scattering) shed light onto the nature of these aggregates. The mesoscopic clusters obtained by fostering aggregation with acids can be described as fractals and their internal structures are strongly dependent on acid concentration, mixing order of reagents and ageing of stock solutions.[2] The behavior of this porphyrin on interacting with linear polyamines is rather interesting. These reagents are able to induce J-aggregation at higher pH values, with respect to the usual conditions for self-aggregation. The nature of the polyamine and in particolar the number of protonable nitrogen atoms largely influence the morphology of the final clusters. Additino of spermine leads to peculiar J-aggregates having dendritic morphology and behaving as efficient antenna systems.[3]
Chiral organized supramolecular assemblies have been obtained interacting the porphyrin trans-H2Pagg with poly-glutamate in alfa-helix conformation or on nucleic acids.[4] Assembling kinetics have been studied mainly through circular dichroism (CD), taking advantage of the presence of large induced CD signals in the visibile range of the spectra, where electronic absorption of porphyrins usually occurs. The experimental evidences show that a high degree of reproducibility and a fine control of the aggregation extent can be achieved by a careful choice of the usual medium parameters. Once again, the protocol for mixing the reagents and ageing of stock solutions play an important role in determining the kinetics of aggregation. During these investigations, nucleation phenomena have been fully addressed.
We intend to take profit of the knowledge gained with these two porphyrins to extend the methodology to their metal derivatives. The use of metallo-porphyrins allows for a wide modulation of the photophysical properties, introduces potential redox activity and can impose structural changes, due to the coordination ability of the central metal ion.
In order to get further insights into the aggregation dynamics, detailed investigations will be carried out exploiting time resolved spectroscopy (UV/vis, fluorescence emission, resonant light scattering). We will extend our attention to other anionic porphyrins using the same polymers, or to cationic porphyrins interacting with polyanionic substrates. Structural information in solution phase will be obtained through light scattering techniques (elastic and dynamic). Morphology of the resulting aggregates can be gained by high resolution microscopy, AFM (Bari RU) and confocal optical microscopy. Collaborating with the Bari RU, potential applications in opto-electronics of these aggregates will be investigated (optical limiting and two-photons absorption).
2) UV-induced deposition of porphryin nanoaggregates.
During previous investigations on aggregative behavior of tetra(4-pyridyl)porphyrin in chloroform or dichloromethane solutions, this RU showed that nano and microcrystals of this compound can be deposited on the surface of the cell exposed to the UV/Vis beam. The phenomenon has been explained by UV-induced photodecomposition of the halogenated solvent, formation of hydrochloric acid, protonation of the porphyrin and nucleation on the silica surface.[5] Preliminary studies have pointed out that it is possibile to extend this methodology to several porphyrins. Therefore, we intend to find out the optimal conditions (exposition time, power of the UV beam, temperature) to deposit crystals controlling the size on various substrates (quartz, silicon, metals). It is interesting to note that different halogenated solvents (brominated or iodinated) could lead to deposition of porphyrin acidic species containing counteranions different from chloride. This opportunity could allow for a better tuning of the photophysical properties of the nanocrystals, due to the dependence of these properties on the nature of counter-ions.[6] Obviously, metallo-porphyrins represent another way to change the properties of the nanodeposits. The nanocrystals will be characterized by usual spectroscopic techniques (UV/Vis, fluorescence emission and RLS). The morphological analysis will be carried out by confocal optical (in trasmission and fluorescence mode), atomic force (AFM, Bari RU) and near-field optical scanning microscopy (SNOM) available at the Dipartimento di Metodologie Fisiche Avanzate of the Messina University. Possible applications of such nanocrystals as sensors (exploiting quartz microbalance) or as active part in opto-electronic devices will be explored in collaboration with the Lecce RU.
3) Interaction of water soluble porphyrins with model proteins.
Previous results obtained on the interaction of porphyrins with simple polyamines and polypeptides will costitute the starting point for the spectroscopic, structural and kinetic study of porphyrin aggregation on proteins. This RU plans to investigate the formation of supramolecular complexes of porphyrins and porphyrin aggregates on simple proteins, such as albumin. This latter protein has been selected because it is considered as the major porphryin carrier in blood, and consequently it plays an important role when using porphyrins treating cancer in PDT. A detailed spectroscopic characterization (UV/Vis absorption, fluorescence emission, circular dichroism, IR and NMR spectroscopy, resonant light scattering) of these complex biostructures will be carried out. In particular, timeresolved techniques will be exploited to obtain kinetic data on these aggregation processes. Specific experiments will be designed to investigate either the aggregation of porphyrins on the selected protein models or the interaction of preformed aggregates with proteins. Specific effects of porphyrins on self-aggregation properties of lysozime and albumin [7] will be studied, essentially through resonant light scattering technique. The formation of more extended structural networks in solution could be investigated through light scattering techniques. A combination of elastic light scattering and small angle light scattering allows for the determination of the dimension of aggregated species, i.e. of their apparent molecular weights. Dynamic light scattering and time-resolved fluorescence can furnish further details on rotational dynamics of these species.
4) Interaction of porphryins with amphiphilyc cyclodextrins.
TPPS4 porphyrin interacts with vesicles formed by amphiphilic cyclodextrins, leading to different aggregated species depending on the relative concentrations. Upon irradiation, these supramolecular species produce singlet oxygen and they are able to vehicle porphyrins inside tumor cells, specifically targeting the nuclear compartment.[8] On considering that TPPS4 alone binds aspecifically to the cell membrane, the obtained results are very interesting for the development of new systems suitable of application in PDT. Following this line of research, we intend to study the interaction between chlorophylls (in collaboration with Bari RU) and various metal derivatives (e.g. Zn(II)) of anionic porphyrins with amphiphilic cyclodextrins, bearing amino-terminated hydrophylic groups (positively charged at physiological pH). These interactions will be modellized through theoretical methods in collaboration with the Bari UR. The characterization of such systems will be achieved through the combined use of the already described spectroscopic techniques.
Furthermore, Z-potential measurememnts (electrophoretic light scattering) will be exploited. Such technique allows for an accurate description of aggregated species in terms of size and charge. SNOM and confocal microscopy can be conveniently used to visualize vesicles in solution, together with the distribution of porphyrins inside the cells. The photogeneration of singlet oxygen will be determined through conventional chemical and photochemical methods (in collaboration with Catania RU). In vitro test on different tumor cells will be carried out in collaboration with the Dipartimento di Scienze Microbiologiche, Genetiche e Molecolari of the Messina University.

[1] R.F. Pasternack et al. Science 1995, 269, 935.
[2] M. Castriciano, et al. J. Phys. Chem. B, 2003, 107, 8765.
[3] L. Monsù Scolaro et al .Chem. Commun. 2005, 3018. N. Micali et al. Phys. Rev. E. 2005, 72, 050401.
[4] L. Monsù Scolaro, et al. J. Am. Chem. Soc. 2004, 126, 7178
[5] L. Monsù Scolaro et al. J. Am. Chem. Soc., 2003, 125, 2040.
[6] A. Rosa et al. J. Phys. Chem. A, 2003, 107, 11468. G. De Luca et al. J. Phys. Chem. B, 2005, 109, 7149. G. De Luca et al. Chem. Mat. 2006, 18, 2005.
[7] T.T. Tominaga, et al. J. Inorg. Biochem. 1997, 65, 235.
[8] A. Mazzaglia, et al. Chem., Eur. J. 2003, 9, 5762. S. Sortino et al. Biomaterials 2006 in press.