Preparation And Characterization Of Polyelectrolyte Complex Nanoparticles Based On Chitosan And The Primary Study On Their Drug-loaded Properties

Polyelectrolyte complexes (PEC) are formed by static interactions between macromolecules that carry oppositely charged polyions. In the recent twenty years, PEC were studied broadly due to their special properties, and applied in many fields such as environment, new coatings materials, template polymerization, protein separation, drug delivery and molecule self-assembly. Chitosan(CS) is a natural cationic polyelectrolyte. As a biocompatible and slowly degradable polymer, chitosan has been widely used in drug delivery systems especially in drug-controlled release, targeting, and intellectualized delivery system. The information has emphasized the importance of size, and revealed the advantages of nanoparticles over the microspheres, it has been observed that the number of nanoparticles that cross the epithelium is greater than the number of microsphere macromolecular. Meanwhile, nanoparticles drug delivery can also decrease the side-effects and enhance the drug bioavailabilty. It is a significant work to study the effect of CS molecular parameter and the introduction of a second ingredient on the properties of nanoparticles drug delivery. They also may affect the increase of their versatility in terms of the encapsulation and delivery of drug and their susceptibility to interact with biological surface.According to the research background, the research work in this thesis was focused on the preparation, characterization and primary application of the nanoparticles based on chitosan polyelectrolyte complex for drug delivery. Specially, five sections of work were carried as follows:(1)Under the condition of weakly acid solution, hydrogen peroxide was used to degrade the high molecular weights (Mws) CS. Low Mws CS was attained by regulating the amount of hydrogen peroxide, reaction temperature and reaction time, measured by viscosity-method. Alizarin red S (ARS) can interact with chitosan and the CS-ARS complexes are formed with the appearance of the new absorption peak at 425 nm. The decrease in absorbance at 425 nm is in proportion to CS concentration.Based on those above, a spectrophotometer method is proposed for the determination of CS. The conditions for measurement were determined, and the linear equations for different Mws CS were built. The quantities of CS in different compound samples have been determined in this way.(2)CS can react with ionic drug directly and forms polyelectrolyte complex to embed the drug. Using glycyrrhetic acid as a model drug, we prepared CS-GLA nanoparticles through the process of ionic gelation. The interaction involved in the complex coaceration was mainly electrostatic interaction. The remarkable advantage of this system is that all CS-GLA nanoparticles are obtained under mild conditions without any organic solvents and surfactants. Only when the weight ratio of GLA to CS was larger than 0.04, could the nanoparticles be formed. The mean diameter, size distribution and zeta potential of the nanoparticles can be controlled by some factors including the weight ratio of CS to GLA, the average molecular weight of CS and the pH value of the medium. Encapsulation efficient (EE) of GLA within the nanoparticles decreased from 88.1% to 55.5% when the weight ratio of GLA to CS in the precursor solution increased from 0.07 to 0.14, but the GLA loading capacity (LC) of the nanoparticles increased from 6.4% to 7.3%. The optimal conditions for formation CS-GLA nanoparticles were the Mw of CS 17.9kDa, pH 6.5, and a weight ratio of GLA to CS of 0.09. These conditions produced the smallest mean size 298nm. The polydispersity index of 0.04 was consistent with a relatively narrow size distribution. Electron microscopy suggested that the particles are spherical nanoparticles with a smooth surface. The experiment of GLA release in vitro and in vivo showed that these nanoparticles provided a continuous release.(3)In order to increase the loading capacity and control the release rate, we prepared chitosan- polyaspartic acid (CS-PAsp) nanoparticles. Two titration methods were applied to study the forming process of polyelectrolyte complex nanoparticles: CS solution added into PAsp solution (positive system, CS-PAsp nanoparticles) and PAsp solution added into CS solution (reverse system, PAsp-CS nanoparticles). Theresults indicated that the slow dropwise addition of chitosan into PAsp allowed to elaborate either anionic or cationic spherical nanoparticles in the size range of 80-400nm with proper CS/PAsp unit molar ratios. Variations in CS and PAsp molecular weight, solution pH, temperature, ionic strength and crosslinker concentration were examined systematically for their effects on nanoparticle size and tendency of particles aggregation so as to enable speedy fabrication of CS nanoparticles with predetermined properties.(4) CS-PAsp nanoparticles were used to embed hydrophilic drugs doxorubicin (DOX) and 5-Fluorouracil (5FU) by two methods (mixing method and absorption method). The results indicated that the nanoparticles could entrap the two drugs with high LC and EE. Drug loaded nanoparticles were stable for three months under room temperature after being crosslinked by Glutaraldehyde (GA). The in vitro experiment stated that DOX was mainly entrapped onto the surface layer of the nanoparticles due to its hydrophilic properity and larger molecular weight. These DOX molecules associated to the nanoparticles surface diffused out easily and caused the rapid release of DOX in the incubation time. In addition, the nanoparticles couldn't control the drug release. However, after being crosslinked, the 5FU drug loaded nanoparticls prepared by two methods could provide a continuous release for more than 144 hours. The rapid release was decreased greatly and the in vitro release mechanism belonged to Hixson-Crowell model. In vivo experiment of the drug-loaded nanoparticles showed that CS-PAsp nanoparticles encapsulation presented a sustained release of 5FU comparing to the 5FU solution and the half-life times were prolonged and the areas under curve (AUC) were higher. It also found that there were significant pertinence between in vitro and in vivo release, therefore, in vitro release could provide academic foundation for in vivo release.(5) 5FU loaded CS-PAsp nanoparticles by mixing method (CS-PAsp-5FU-m) was used to therapy the mice tumors. It was found that the inhibitory percentage of volume and weight arrived at 81.10% and 72.79% respectively. Bring into comparison, pure5FU solution was administrated, showing the inhibitory percentage of volume and weight 74.53% and 65.3% respectively. The studies in side-effect showed there was no significant discrepancy in physical characterization after the mice was administrated with CS-PAsp-5FU-m nanoparticles and physical brine. However, pure 5FU had great discrepancy compared to physical brine.