Development of in vitro method for characterization of release and diffusion behavior of small-molecular drugs and biomolecules of subcutaneous administration. Investigation of protein interactions with the drug model temoporfin using the dispersion releaser technology.

Thesis title: "Study of technologies for nano and microencapsulation of active agents with different physicochemical characteristics"Abstract: Diseases such as cancer, diabetes and other that depress the immune system are quite common and can cause serious limitations. In the treatment of these diseases, many types of drugs can be used, and the nano and microparticles are one of the most investigated systems for drug delivery. Properties such as solubility, stability and metabolism of the drug to be encapsulated are important because they determine the method and materials to be used. This study aimed to evaluate techniques more industrially viable and the use of different materials, natural or synthetic, to obtain nano and microparticulate delivery systems containing drugs with different physicochemical properties. Thus, formulations consisting of PLGA-PEG nanoparticles were obtained by the nanoprecipitation method assisted by microreactor for encapsulation of the photosensitizer mTHPC to use in photodynamic therapy. Moreover, L-alanyl-L-glutamine (AGP) was encapsulated by spray drying using pectin and Surelease® to the prevention of insulin-induced hypoglycemia. Further, by the same technique, AGP was microencapsulated using pectin and extract of propolis byproduct (EPB) as a system for stimulation of the immune system. The formulations were physicochemically characterized and evaluated in vitro and/or in vivo. Nanoparticles with mTHPC were able to reduce its toxicity while maintaining efficacy, and possibly improved the in vivo biodistribution. The microparticles of pectin and Surelease® containing AGP did not show a satisfactory in vitro release profile, but in vivo showed better potential in the maintenance of blood glucose than the non-encapsulated AGP. The system prepared with the EPB demonstrated immunostimulant action. Thus, nano and microparticulate systems have shown promising results and the chosen methods would facilitate a future upscale.

Dissertation title: Preparation and characterization of microparticles of ethylcellulose containing L-alanyl-L-glutamineAbstract: The glycemic control is essential for diabetic patients. However, some studies have shown that the strict glycemic control increases the incidence of insulin-induced hypoglycemia (IIH), particularly of nocturnal hypoglycemia. The administration of L-alanyl-L-glutamine (AGP) is effective on glycemia recovery in long-term IIH cases and it is suggested that its oral administration prevents prolonged episodes of IIH. However, AGP is rapidly metabolized. Therefore, it would be interesting the development of an oral modified release system to prolong the drug release, particularly during the sleep, in which diabetic patients have a decreased perception sense of hypoglycemia. Thus, this study aimed to obtain and characterize microparticles (MP) containing AGP. The MP were obtained by the emulsification and solvent evaporation method, either by simple and double emulsification, using ethyl cellulose. The MP showed spherical shape, smooth surface and mean diameter between 9.0 and 14 μm. It was proved by TG and DSC that AGP has high thermal stability which was even increased after encapsulation. The MP prepared by double emulsification presented higher encapsulation efficiencies than those prepared by simple emulsification. The microencapsulation technology has enabled the prolonged release of the drug and the MP prepared by simple emulsification demonstrated a quicker and more extensive release of AGP compared to the MP prepared by double emulsification.

The development and manufacture of novel nanocarriers for drug delivery has proved challenging with regards to scale-up and pharmaceutical quality. Polymeric nanocarriers composed of poly(lactic-coglycolic acid)-b-poly(ethylene glycol) (PLGA–PEG) were prepared and the photosensitizer meso-tetrakis (3-hydroxyphenyl) chlorin (mTHPC) was effectively encapsulated. Furthermore, the interplay of various process and formulation parameters and their impact on the most important product specifications were investigated by using a factorial design and a central composite design in a microfluidic manufacturing process. These nanoparticles for intravenous administration with a size of 97 +- 0.13 nm, narrow size distribution, and an encapsulation efficiency of more than 80% were produced at high throughput. In vitro stability and in vitro drug release testing were applied for quality control purposes. Finally, the toxicity of the photosensitizer was tested in vitro. The cytotoxicity was successfully reduced while the efficacy of the formulation was maintained. First observations using in vivo imaging suggest effective distribution of the nanocarrier system after injection into rodents. Thus, further in vivo testing of the beneficial effects of nanoencapsulation into the matrix system and its formulation will be considered for the delivery of mTHPC to tumor tissues during photodynamic therapy.