Supercritical assisted processes for the production of biopolymeric micro and nanocarriers

Abstract

This work provides an innovative point of view on obtaining nanoparticles by supercritical fluids. Supercritical fluids have already been exploited in the production and processing of micro or sub-micro particles, but the production of nanoparticles is still more ambitious, requiring a deep understanding of the thermodynamic of multiphase systems involved and of the fluid dynamics and mass transfer of the process. This thesis focused on the development of supercritical assisted process for the production of biopolymeric micro and nanocarrier for pharmaceutical and biomedical applications. After a wide study of the state of the, two different processes have been focused on the production of different kind of devices: 1) Supercrititical Emulsion Extraction process (SEE) for the production of multifunctional nanodevices; 2) Supercritical Assisted Injection in Liquid Antisolvent process (SAILA) for the production of stabilized nanoparticle water suspensions. Supercritical Emulsion Extraction (SEE) has been recently proposed in the literature for the production of drug/polymer microspheres with controlled size and distribution, starting from oil-in-water (o-w) and water-in-oil-in-water (w-o-w) emulsions. This process uses supercritical carbon dioxide (SC-CO) to extract the “oil” phase of emulsions, leading to near solvent-free microparticles. SEE offers the advantage of being a one-step process and is superior to other conventional techniques for the better particle size control, higher product purity and shorter processing times; the SEE process has been proposed in the continuous layout, using a high pressure packed tower. However, monodisperse sub-micro, nanoparticles of biopolymer suitable for pharmaceutical formulations have not been produced until now by SEE. Therefore, this part of the work was performed on the production of monodisperse biopolymer nanoparticles. Particularly, emulsion formulation parameters have been tested, such as, different surfactant concentrations and biopolymer percentages in the oily phase, and several emulsification techniques (ultrasound or high speed emulsification) and their interactions with SEE processing have been tested to obtain small droplets dimensions from the micro size to the nano size range. Poly-lactic-co-glycolic acid (PLGA), poly-lactic acid (PLA) and poly-caprolactone (PCL) were the polymers selected to produce micro and nano devices, relative results are shown in Chapter 7. Then, after the optimization of the process conditions for the production of monodisperse sub-micro and nanoparticles encapsulation of drugs, proteins, peptides and metals has been carried out. For the encapsulation study the SEE technique starting from single or double emulsions was used for the production of nanospheres. The SEE technology has a great potential regarding the field of protein encapsulation thanks to the mild extraction condition and the short process time, and mainly thanks to the possibility to use multiple emulsions. For this reason, the SEE has been applied to the production of biopolymeric micro and nanoparticles of PLA and PLGA encapsulating peptides and proteins starting from double emulsion. Results are shown in Chapter 8. Bovine serum albumin (BSA) has been selected as protecting protein for growth factor, such as Vascular Endothelial Growth Factor (VEGF) and Bone Morphogenetic Proteins (BMP). Both micro and nanoparticles were produced; the effect of size of particles on protein loading and release has been evaluated. PLGA microspheres loaded with GFs were charged inside a bioactive alginate scaffold to monitor the effect of the local release of these biosignals on cells differentiation. Human Mesenchimal Steam Cells (h-MSC), where used in view of the fact that they are a promising cell source for bone tissue engineering. Good cells differentiation indicates that the SEE process was successful in the encapsulation of proteins and peptides, preserving the functional structure of the proteins, thanks to the mild operating conditions used. Another important challenge that has been managed is also the production of nanocomposite biopolymeric particles encapsulating metal nanoparticles for the production of light sensitive drug delivery devices. The encapsulation of TiO2 and Au nanoparticles has been performed for different biomedical applications (Chapter 9 and 10). In this work PLA/nano-TiO2 microparticles have been produced using SEE for photodynamic therapy of cells and bacteria. Anatase type nano-TiO2 ethanol stabilized suspension has been synthesized by precipitation from solutions of titanium alcoxides and directly used as the water internal phase a of double emulsion water in oil in water and also TiO2 fine powder was produced and used for encapsulation experiments using a solid in oil in water emulsion. Both micro and sub-microparticles have been produced. [edited by Author]