Preparation Of Biomimetic Chitosan Composite Materials And Preliminary Biological Evaluation

Bone defect tend to result in bone nonunion,which is always difficult in the treatment of orthopaedics.To find an effective replacement material of bone transplantation to repair bone defect is the direction of the researches in bone defect therapy at present.Due to a series of complications induced by transplantation of autogenous and allograft bone,the biomimetic materials will be the main direction.Chitosan,the N-deacetylated derivative of chitin,has been recommended as suitable materials because of its biocompatibility, biodegradability,reliability and wide sources.However,the melt temperature of chitosan is higher than its disassociation temperature due to the intensive hydrogen bond among the molecules.It is very difficult to prepare materials with strong intensity and good biocompatibility to be used to therapy the bone fracture and defect.The aim of this study was to prepare CaCO₃/chitosan three-dimentinal composite material and chitosan/hydroxyapatite composite material with in-situ precipitation method and evaluate their biocompatibility and repair ability for the radial bone defect in rabbits.METHODS:CaCO₃/chitosan three-dimentinal composite material was prepared by the method of in-situ precipitation.In this method the calcium ions were added into the solution of chitosan firstly and were dispersed homogeneously to attain the uniform solution.Then the uniform solution as the CaCO₃/chitosan precursory solution was poured into a cylindrical glass mold,on the internal surface of which a chitosan membrane had been precipitated. Afterward the mold was soaked in the mixture of NaOH and Na₂CO₃ aqueous solutions. Finally through the following proceedings,the three-dimensional composite in which the particles of calcium carbonate were dispersed and achieved in the experiment.The chitosan/hydroxyapatite composite material was prepared with alternation in-situ deposition method,which packed hydroxyapatite coating on the chitosan base membrane.The physicochemical properties of these materials were determined with electronmicroscope, X-ray diffraction analysis,and etc.Then,the 3T3 cells were used to evaluate the compatibility with MTT chromatometry and agar covering methods,respectively.In addition,the effects of these materials to repair the radial bone defect were observed in rabbits.RESULTS:1.It could be seen that the particles of CaCO₃ were distributed orbicularly on the cross section and its size was about 5-10μm.The mechanical properties of the composite were evaluated by three-point bending test.The bending strength enlarged firstly and then dropped with the increase of calcium carbonate's mass fraction and the modulus decreased continuously.As the mass fraction was 10%,the bending strength of the material reached its peak(about 113MPa) with 2.6MPa bending module.2.Both the coating and the upper tier particle of the hydroxyapatite in the chitosan/hydroxyapatite composite material were with porosity honeycomb appearance. The content of hydroxyapatite increased with the number of deposition cycle increasing and reached saturation(8.8%) at the end of the 4th cycle.The tensile strength of the material showed the opposite change with the content of hydroxyapatite.3.The growth of 3T3 cells had no significant change after incubated with leachate of these two chitosan composite materials(10-100%),compared with control group.No cytolysis was observed also.4.There was a little osteotylus growth directing the materials at 2nd week after the operation;abundant new periosteum parceling migrating into the materials at 4th week; newly formed bone separating and surrounding the materials at 8th week;and most of the defect zone substituted by the newly formed ripe lamellar bone and medullary cavity emerging at the 12th week after the operation.CONCLUSION:1.CaCO₃/chitosan composite material was successfully prepared with in-situ precipitation method.Hydroxyapatite coating could constructed above the chitosan base membrane through biomineralizatic altermating deposition.These structures were profit for the cell adhesion and growth.2.The content of hydroxyapatite,shape and mechanics intensity of the chitosan/hydroxyapatite composite material could be controlled through the cycles of precipitation to satisfy the need of histo-reorganization. 3.These two materials had no obvious cytotoxicity.4.Both composite materials could induce the growth of new bone and promote the repair of the radial bone defect in rabbits.