Preparation Of PLGA Microspheres / P(NIPAAm-co-AAm) Hydrogels Combination System And Their Drug Delivery Application

With the fast growth of biomedical and biotechnology, protein drugs have been being extensively investigated because of its high bioactivity, incredible selectivity, low toxicity, fewer side effects, and great potential to clinical application. But the clinic applications through oral delivery systems still face many obstacles, duo to their low stability, short half-life and easily degraded by photolytic enzymes in the gastrointestinal tract. Therefore, controlled-release drug delivery was developed by encapsulating proteins into polymeric materials for sustained release of protein drugs in expected time.Both degradable polymeric microspheres and thermoresponsive hydrogels are the most important carrier devices. Microspheres prepared with biodegradable polymers such as poly(D,L-Lactide-co-glycolide) (PLGA) show much prolonged drug release for an extended time period. However, the high initial bursts release and denaturing of proteins has been reported as a drawback for the application of PLGA microspheres. The denaturing of proteins is caused by exposure to organic solvents and high sheering force during microspheres fabrication processes. Poly(N-isopropylamide) (PNIPAAm) hydrogels have been intensely investigated as protein drug vehicles for recent decades because the release rate of loaded drugs could be modulated by external environmental changes due to their temperature sensitivity property. Furthermore, a highlight of PNIPAAm hydrogel is that the pore size of network can be changed by volume phase transition, which made proteins could be loaded into PNIPAAm hydrogel by post-fabrication encapsulation with avoiding denaturing of proteins. But the hydrogel system is not able to control drug release in an ideal manner because of its limited pore size. Thus, a combination system of microspheres and hydrogels reciprocally compensate for individual disadvantages to produce a novel drug delivery system for protein. Firstly, PLGA microspheres were prepared by emulsion-solvent evaporation, and investigated the influences of manufacturing parameters on the properties of microspheres. Then drug-loaded microspheres were introduced into the thermosensitive hydrogel implant for sustained delivery of protein. In vitro release of drugs and their release mechanism of combination systems were studied. The main works and conclusions are included as follows:PLGA was chosen as the carrier material. Bovine serum albumin (BSA) was employed as the model drug. Drug-loaded PLGA microspheres were prepared by W/O/W emulsion-solvent evaporation method. The influences of manufacturing parameters on the properties of microspheres such as particle size, entrapment efficiency, drug loading efficiency, and drug release in vitro were investigated. Microspheres with high encapsulation efficiency and well sustained drug release were prepared.Proteins were loaded into PNIPAAm hydrogel by post-fabrication encapsulation technique because the pore size of hydrogel network can be changed by volume phase transition. This technique can avoid denaturing the bioactivity of proteins from the potentially harsh environments during the loading process. The conventional thermoresponsive hydrogels for protein delivery, however, suffer from certain limitations duo to low entrapment rates, loading efficiency and incomplete release of proteins. To improve the loading efficiency and to increase the cumulative release of protein from the hydrogels, porous thermoresponsive P(NIPAAm-co-AAm) hydrogels were synthesized by two-step freezing polymerization method. The influence of the reaction time of the first step polymerization and the amount of deionized water on the porous structures of P(NIPAAm-co-AAm) hydrogel was investigated using a scanning electron micro scope (SEM ) and Auto surface area analyzer. Scanning electron microscopy (SEM) studies revealed that the hydrogel frozen after 3 min and 2.5 ml of deionized water in the reaction system exhibited the most highly interconnected porous structures and the largest pore size. These open pore structures were very useful to increase the drug loading efficiency and the cumulative released amount of the hydrogels. In vitro BSA release from the hydrogels exhibited that the drug release rate was mainly affected by the porous structure of the hydrogels. So creating an interconnecting pore structure within the hydrogel matrices is an effective strategy to improve the drug loading efficiency and the cumulative released amount of the hydrogels.To overcome the above limitations, Drug-loaded PLGA microspheres were dispersed into porous P(NIPAAm-co-AAm) hydrogels for prolonging sustained delivery of proteins with the auto adjustable function to external temperature changes.Various amounts PLGA-based microspheres were incorporated physically into P(NIPAAm-co-AAm) hydrogel to form the combination drug delivery system. FTIR and SEM showed that PLGA microspheres the originally chemical properties of PLGA microspheres and P(NIPAAm-co-AAm) hydrogel were not changed, but the porous structure of hydrogels was destroyed. With the PLGA microspheres addition to the hydrogels, the number of porous structure of the gel was reduced. The in vitro BSA release behaviors showed that a novel drug delivery system was successfully constructed for long-term sustained release of proteins. The mechanisms of drug release from the microspheres, hydrogels and combination systems were deduced by Peppas'formula. The results show that the mixture formulation exhibits sustained release of BSA for over 30 days without any burst effect. The drug release was controlled by both drug diffusion and material erosion. Cytotoxicity study proved that PLGA microspheres/P (NIPAAm-co-AAm) hydrogels have good biocompatibility. Such results demonstrated that the combination drug delivery system may have great potential application for delivering proteins.