Study On Preparation And Characterization Of Double-walled Microspheres As Protein Carrier

In recent years, protein drugs delivery systems have been developed fast, especially for long-acting injectable microsphere made with biodegradable biomaterials. Because of their sustained release property, those microspheres preparation has been a hot field in pharmaceutics. Now, the activity of protein is ready to denature in the preparation process, so how to choose the proper biomaterials and preparation method based on the characterization of the protein drugs and how to prepare a safe, steady and effective preparation is an important challenge.Aim at upper questions, a chitosan- hyaluronate(CHS-HA) double-walled microsphere was fabricated in this work. The detailed process was followed:CHS-HA double-walled microspheres were prepared by emulsification-coacervation method. Tripolyphosphate (TPP) was the ion crosslinker. The individual effect of pH, volume, concentration of outer aqueous phase, andsurfactant on microspheres morphology and fabrication were tested by Zeta (ζ)potential, Scanning electron microscopy (SEM) and Fourier-transform infrared spectrometry (FT-IR). TPP concentration and pH of outer aqueous phase were crucial to microspheres fabrication, when TPP concentration was higher than 8%, spherical and smooth microspheres could be formed. The optimal pH for microspheres formation ranged from 6.0 to 7.0. Microspheres containing both span80 and tween80 in process were less shapely compared with those containing span80 only. In addition, volume and HA concentration of outer aqueous phase also affected the microspheres morphology.Insulin as a model protein drug was encapsulated in CHS-HA double-walled microphers and the encapsulation efficiency and loading efficiency were measured. Further an orthogonal design was achieved based on those results and the most perfect conditions were chose. Through the orthogonal design, the chosen factors were: the volume of the outer aqueous phase was 40mL; the pH of the outer aqueous phase was 6.0, the concentration of HA was 0.025%, and the stirring time was 3h; The release profile in vitro was detected and mimicked with mathematics equation. The mechanism of release was expounded. Meantime, Insulin amount was measured by radio-immunity assay (RIA), and detect the activity of Ins in the process of microsphere preparation, storage and release. The release profile had two phases, the fast release phase and the slow release phase and was according with the first kinetics equation. RIA results showed that Insulin bioactivity has no obvious loss in the process of microsphere preparation, storage and release.