CHITOSAN PARTICLES FOR THE CONTROLLED RELEASE OF PROTEINS

Back Ground Chitosan as a natural polymer has been fabulated into a number of formulations such as films, hydrogels and particles based on its excellent properties such as biodegradable, biocompotable, bioadhesive, permeartion-enhancement, antibiotic, antitumor etc. properties. Among them, chitosan microparticles found a lot of applications in pharmaceutics such as vaccine delivery, mucosal delivery and gene delivery, etc. Down to the nanoscale, chitosan nanoparticles have more attractive properties more than that of chitosan microparticles, which further widen the applications of chitosan particles in biomedicine and biopharmaceutics. Objective of this Thesis This thesis is aimed to prepare chitosan micro or nanoparticles to delivery proteins. What are the questions this thesis attempted to solve? 1) What are the proper preparation conditions of chitosan micro or nanoparticles? 2) How does the pH value affect the formation and protein encapsulation of chitosan nanoparticles? 3) How to overcome the burst release of chitosan nanoparticles? 4) How to overcome the aggregation disadvantage in the contritional preparation process of TPP-gelated chitosan microparticles? 5) How to construct a composite particles system to realize the sustainable release of proteins? What are the methods used in this thesis? 1) ionotropic gelation method to prepare chitosan nanoparticles 2) Emulsification-coacervation (NaOH) method to prepare chitosan microparticles 3) a polyelectrolytes coacervation method to prepare chitosan-BSA complexes 4) a microemulsion involved emulsification-coalescence method to prepare TPP-gelated chitosan microparticles 5) a nanoparticles encapsulation method to prepare composite particles. Results and Conclutions 1) the effect of preparation parameters on the properties of chitosan nanoparticles: 1a) the concentration of chitosan has no siganificant effect on particles yield, positively associated with particles size, size distribution, positively associated with BSA encapsulation efficiency in a specific concentration range; 1b) the mass matio of chitosan to TPP negatively associated with particles yield, positively associated with particle size and size distribution, negatively associated with BSA encapsulation efficiency; 1c) the concentration of BSA has no significant effect on particles yield, particle size or size distribution, negatively associated with BSA encapsulation efficiency in a specific concentration range. 2) a chitosan polymer chain conformation related mechanism is proposed through the study of the effect of pH value on the formation and BSA encapsulation of chitosan nanoparticles. 3) the most homogeneous and smooth chitosan particles could be obtained at the parameters of: 2% (m/v) chitosan solution, 2/10 w/o volume ratio, 4% Span 85 as susfactant, 5 Krpm homogenization speed and 3 times addition of NaOH solution as a coacervation agent. The obtained particles have a mean diameter of 9.4±1.9 m. The BSA loading test found that dispersed particles only could be obtained below the BSA concentration of 0.5% under above mentioned parameters. 4) a noval polyelectrolytes complex formed by TPP and BSA is obtained attempted to solve the burst release effect of chitosan nanoparticles. 5) a microemulsion involved emulsification-coalescence method is used to overcome the aggregation problem of TPP-gelated chitosan microparticles. 6) a chitosan nanoparticles encapsulated PLA composite particles are successfully constructed. What is new in this thesis? 1) to study the formation mechanism and protein encapsulation of chitosan nanoparticles through the study of the effect of pH value on their properties and propose the role of chitosan polymer chain conformation during this process, 2) a noval TPP-BSA polyelectrolytes complex is obtained base on the purpose to overcome the burst release effect of chitosan nanoparticles, 3) apply emulsification-coalescence method in which a microemulsion of cross-linking agent-TPP is used to solve the aggregation problem of TPP-gelated chitosan microparticles, 4) propose a chitosan nanoparticles encapsulated PLA composite particles to control the release of proteins. Where is the study of this thesis in the field? 1) Chitosan nanoparticles have been extensively investigated in the past few years and the factors which can affect the properties of chitosan nanoparticles have been well documented as well. This thesis provides the evidence from a new side to understand the formation and protein encapsulation mechanisms of chitosan nanoparticles. 2) New and highly effective methods have been proposed by others to prepare protein loaded chitosan microparticles such as sieving and microfluidic methods which can reproduceably scale up the production of monodispersed chitosan microparticles. This thesis just solved a technique problem in the conventional preparation process of TPP-gelated chitosan microparticles. 3) The proposed TPP-BSA polyelectrolytes complex could be an alternative route to overcome the burst release effect of chitosan nanoparticles. 4) The proposed chitosan nanoparticles encapsulated PLA composite particles is one of the solutions among other composite particles proposed by others.