This master thesis presents the unique properties of silk in the platform for the design and manufacture of controlled release medicines. A great deal of interest in the biopolymer fibroin derives from a rare combination of its properties, such as the possibility of isolation and aqueous-based purification, processing in the absence of chemical cross-linkers, sterilisation by various methods, etc. Nevertheless, critical points on the path to bringing fibroin-based products to the market, while ensuring proper quality, efficacy and safety remain to be considered. Within our experimental research, we developed a robust and scalable process of isolating fibroin from the cocoons of silkworm Bombyx mori and established control steps to optimise the identity and purity of the product. The characterisation of the prepared fibroin solution was our starting point in a systematic approach to formulation development, including the selection of estradiol as a drug candidate and determination of critical parameters for the preparation of fibroin delivery systems with controlled release. We prepared various fibroin delivery forms (films, hydrogels and porous scaffolds) with the addition of ethanol, which served to dissolve estradiol and accelerate the gelation of the fibroin solution. We determined the potential of estradiol delivery systems by testing the incorporation, in vitro release in the phosphate buffer without and with the surfactant and analysis of estradiol concentrations with high-performance liquid chromatography. The properties of our delivery systems and their influence on fibroin were determined with different analytical methods: scanning electron microscopy, Fourier-transform infrared spectroscopy and X-ray powder diffraction.
Thin films were dissolved in the buffer within five minutes with initial “burst-release” of estradiol, as the fibroin in them was predominantly amorphous and water-soluble. Chemical and conformational properties of fibroin in thin films did not change, despite the incorporation of estradiol in ethanol solution. Thick films were inherently insoluble due to the gelation of fibroin with ethanol and consequently had a higher content of crystalline domains or β-sheets. For the preparation of thick films, we dried the intermediate hydrogel forms. By testing the release of estradiol in vitro, we found a prolonged release for up to 129 days, which confirms the promising use of the thick films with estradiol. Porous scaffolds prepared by lyophilisation are less suitable as delivery systems since estradiol was present only on the surface in the form of crystals and was not incorporated into the fibroin matrix.
|
