Preparation Of Chitosan - Polylactic Acid Graft Copolymer, Drug Release And Biocompatibility

Chitosan (CS) and polylactide (PLA) are polymers with excellent biocompatibility and biodegradability. CS is bioactive, but the intermolecular hydrogen-bonding interactions restrict the solubility of CS with high molecular weight (Mn) in water and in many organic solvents. Low molecular weight chitosan oligosaccharide exhibits excellent hydrophilicity. PLA presents outstanding mechanical strength, but the high hydrophobicity greatly constrains its biomedical applications. CS-graft-PLA copolymer (CS-g-PLA), prepared by attaching PLA to CS backbone, could combine the hydrophobicity of PLA with the hydrophilicity of CS, and therefore presents great potential as controlled drug release carriers and tissue engineering scaffolds. CS-g-PLA was synthesized by coupling reaction of the end carboxyl of PLA-COOH and the amine group of CS, using EDC·HCl and NHS as catalysts.The structure, properties and controlled drug release behavior were characterized to evaluate the potential biomedical applications of CS-g-PLA.Firstly, monodisperse PLA-COOH was synthesized using low toxic zinc lactate as catalyst. PLA-COOH was then activated and coupled onto the CS backbone to yield CS-g-PLA by using EDC·HCl and NHS. Experimental parameters, such as the Mn of CS and PLA, EDC·HCl to NHS ratio, mixed solvent system and reacting time etc, were found to influence on the structure of CS-g-PLA, especially the degree of substitution (DS) of PLA. CS-g-PLA prepared from CS with Mn of8000, PLA with Mn of3200and catalyst ratio of5presents the highest DS of PLA-25. At the early stage of synthesis, prolonging reaction time helps to enhance the DS of PLA; the DS reaches the highest when the reaction time is8h; while after then, the DS decreases. The size of CS-g-PLA micelles examined by DLS is in the range of50-200nm, and micelles remain stable in one week.Self-assembled micelles were prepared by direct dissolution and solvent evaporation. The drug release behavior of the CS-g-PLA micelles was investigated using paclitaxel (PTX) as the model drug. Release profiles are characterized by an initial burst followed by a slower release.Various parameters such as preparation method, micelle composition, preparation duration, drug load and stereocomplexation influence the release behavior of PTX from CS-g-PLA micelles. Micelles prepared by solvent evaporation or from CS-g-PLA with higher DS of PLA enhance the release rate of PTX. Longer preparation time leads to lower EE and higher release rate,while lower initial feed ratio of drug to copolymer leads to lower release rate. In addition, the release rates of mixed micelles appear slightly slower than that of single ones, indicating the existence of stereocomplexation between CS-g-PLLA and CS-g-PDLA.The coagulation properties of CS-g-PLA were evaluated from hemolysis, plasma recalcification profile, dynamic blood clotting. The cell viability was assessed by MTT, agar diffusion and LDH experiments. The samples actually have no effect on the viability to cells. Inflammatory cytokine analysis using sandwich ELISAs revealed that CS-g-PLA could lower the presence of inflammatory cytokine markers.