Preparation Of Calcium Alginate/Chitosan Microsphere For Protein Oral Delivery And Its Properties Study

Parenteral administration is the only route for protein drugs now, but multiple injections and high cost are its shortcomings. It is necessary to develop the oral formulation which is convenient and safe. There are two obstacles of oral protein delivery, one is susceptibility to enzymatic degradation in the gastrointestinal tract, the other is its low permeability across the intestinal epithelium. Microsphere technique seems to be the most hopeful way to solve the problems in the developing strategies which had been proposed, because microspheres could protect the activity of the encapsulated protein and protein- loaded microspheres with 10μm diameter or smaller could be absorbed into the blood stream through peyer's patch.The preparation and properties of microspheres for protein oral administration were studied in this paper under the base of the mild and efficient impulsive electrostatic technique. Owning to their good biologic characteristics (compatibility, degradation and adhesive ability), processbile speciality and economic trait, chitosan and sodium alginate were used as materials and insulin was chosen as drug model.1. The disaggregating method of insulin-loaded calcium alginate microspheres wasestablishedIn order to quantitatively estimate the effect of the impulsive electrostatic preparation process on insulin activity, a disaggregating method of calcium alginate microsphere was established. Using citric-citrate buffer solution as medium, the interrelated factors influencing the disaggregating rate and insulin relative activity were studied through orthogonal experiments, and finally the statistic analysis of the relative activity of insulin loaded in microspheres after disaggregating was conducted. It was found that the pH value of the buffer solution and the citrate ionic strength M evidently influenced the disaggregating rate of microspheres. When pH>5.6, the disaggregatingrate significantly increased whereas the activity decreased obviously. With the increase of M, the disaggregating rate was increased obviously whereas the activity hardly changed. After insulin-loaded microspheres were disaggregated by the buffer solution composed of pH=5.6 and M=0.153mol/L, the statistic analysis showed that when confidence level was a = 0.05 and a =0.1, confidence interval of the relative activity of insulin loaded in microspheres could be (89.13, 91.63) and (89.35, 91.41). Therefore, this method could rapidly and sufficiently disaggregate calcium alginate/chitosan microspheres and meanwhile effectively keep insulin activity.2. The effect of the impulsive electrostatic technology on the activity of protein loaded in the microspheresThe effect of indicatory voltage, frequency and pulse width on the falling percentage of insulin relative activity was analyzed by the estimation of blood sugar in mice after the determination of encapsulation efficiency of insulin by CBBG The results showed that the falling percentage of insulin relative activity in the preparation process was very low (<8%). When the voltage rose from 242V to 385V, it increased from 0.8% to 7.54%, but the frequency and pulse width had no effect on it. For its slight effect on the drug activity, impulsive electrostatic technique could be used to prepare microsphere for protein oral delivery.3. The stability of the calcium alginate/chitosan microsphere membrane in the related solution of oral deliveryThe swollen rate was used to examine the stability of microsphere membrane in simulative gastric juice and simulative intestine juice which are related solution of oral delivery. It was found that the reacting amount of chitosan and calcium alginate was important to sustain the stability of the microsphere membrane. The optimal preparation conditions were determined to ensure microspheres stably getting across the gastrointestinal tract with the activity of the drug in it.4. The sustained release property of insulin-loaded microsphere in the related solution of oral deliveyBy using HPSEC to mensurate the insulin content, the sustained release property ofinsulin-loaded microsphere in the related solution of oral delivery was performed, it was found that the release profile was mainly affected by chitosan molecular weight, and large molecular weight can slow down the release speed. Considering the stability of microsphere membrane, 10 thousand chitosan seemed to be the most appropriate. The increased concentrations of the alginate and chitosan were in favor of making compact membrane, consequently retarded the drug release. In order to decrease the diffusion amount of protein drug into chitosan and its activity losing, the membrane formation time should not exceed 15 minutes.