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ORKThe research work carried out in the present PhD thesis is part of a long–standing research activity regarding the design and synthesis of polymeric materials for applications in Pharmaceutical Applications and in Regenerative Medicine. This activity is ongoing in a research group comprising chemists, biologists, pharmaceutical technologists, engineers and computer scientists at the Laboratory of Polymeric Materials for Biomedical and Environmental Applications (Biolab) of the Department of Chemistry and Industrial Chemistry of the University of Pisa.
The study has been undertaken within the framework of two research projects, the first funded by the Italian Ministry of University Research on the “Development of New Blood Substitutes” and the second funded by the European Community, entitled “Novel Approaches for the Development of Customized Skin Treatments and Services” (SKINTREAT). A six month stage at the Laboratory of Polymeric Particles and Surface Chemistry, Department of Materials and Chemistry at SINTEF company (Trondheim, Norway), was part of the activity performed within the framework of the second project during the PhD research program.
Controlled drug delivery technology represents one of the most rapidly advancing areas of science in which several disciplines, such as chemistry, pharmaceutical technology, and medicine are contributing to human health care. For many years, fundamental and applied investigations have been focused on the development of pharmaceutical formulates allowing for maximization of the therapeutic efficacy and minimization of the adverse effects of the drugs of interest.
The main goal of the first research project is to produce Hemoglobin (Hb) loaded polymeric nanoparticles as artificial blood substitutes. Medical interest in developing these systems derives from the many inconveniences related to the use of blood transfusions in order to re-establish tissue oxygen homeostasis, as well as from the inability of administering free Hb due to its short stability once in-vivo and to the administration associated to nephritic toxicity phenomena. At present, one of the most investigated systems in developing artificial oxygen carriers is represented by cellular Hb Based Oxygen Carriers (HBOC).
Since the use of polymeric biomaterials is one of the most promising approach in developing polymeric nanoparticles to be used as drug delivery systems, the use of Poly(maleic anhydride-alt-butyl vinyl ether) 5% grafted with methoxy-PEG (2000)
Alberto Dessy - PhD Thesis
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and 95% grafted with 2-methoxyethanol (VAM41) will be investigated in the preparation of polymeric nanoparticles loaded with Human Hemoglobin to develop cellular HBOC. The co-precipitation technique, which is an original straightforward procedure developed by the research group involved in the present study, will be applied in the formulation of Hb loaded nanoparticles, since it avoids the use of aggressive organic solvents and does not promote the denaturation of the loaded proteins. Nanoparticles features will be investigated in terms of size, morphology, surface charge, Hb content and protein release. Moreover the maintenance of Hb functional properties, after its loading into nanoparticles, will be detected in terms of the ability of oxygen binding reversibly, maintenance of secondary structure and of cooperative activity.
Hemoglobin oxidation must be avoided or limited during nanoparticles formulation, thus different strategies focused on this aim will be undertaken including the introduction of reducing agents (such as ascorbic acid, citric acid and leucomethilene) into the formulation system, as well as the modification of VAM41 polymeric backbone with the introduction of a conductive moiety represented by 2-methoxyaniline which can minimize oxidative phenomena.
In parallel, the use of Alginate as alternative material will be also investigated in the production of polymeric nanoparticles loaded with Hb. Particles will be prepared by means of a ionic gelation based formulation process; the attention will be focused on the optimization of the over mentioned formulation system as well as on the characterization of the so obtained particles.
The activity to be developed in the second research line will be aimed at applying VAM41 polymer in the production of Dead Sea Salts loaded polymeric nanoparticles for the development of topical applications in the treatment of skin diseases. Mini-emulsion based formulation process will be optimised in order to obtain nanoparticles possessing suitable features to be used in the mentioned application. The prepared nanoparticles will be characterized for their size, morphology, salts content and release.
