壳聚糖对肝素微胶囊性能的影响(英文)

Effects of chitosan on the performance of heparin microcapsule

背景:壳聚糖、海藻酸钠是良好的天然的微胶囊制备材料,在组织工程中应用广泛。本课题组以往制备的抗凝血材料主要是运用材料的惰性以及模仿血管内壁的液晶态,而本实验是在此基础上,利用低分子肝素的生物抗凝活性及其他特异性能,对肝素进行微胶囊化,以期使肝素在体内的释放达到一种缓释的效果。目的:以天然的壳聚糖、海藻酸钠为微胶囊的包囊材料,对低分子肝素进行微胶囊化,以保证肝素在体内的稳定性,分析壳聚糖含量对肝素微胶囊性能的影响。设计:开放性实验。单位:广州暨南大学材料科学与工程系。材料:实验于2004-10/2005-06在暨南大学材料科学与工程系生物材料实验室完成。肝素(山东福瑞达生物化工有限公司,分子量<5000);壳聚糖(脱乙酰度≥90%,黏度<100cps,上海伯奥生物科技有限公司);海藻酸钠(青岛明月海藻工业有限公司);乳化剂为Span80,CaCl2均为国产化学纯。方法:①肝素-壳聚糖微胶囊的制备:取一定量肝素水溶液乳化于石蜡油中,充分搅拌使反应体系呈乳液状,然后使整个体系升温到50℃并维持20min。然后缓慢滴加20g/L壳聚糖水溶液,使体系升温到60℃,再滴加戊二醛,并使反应体系于80℃保持1h。离心分离,过滤洗涤,除去残留有机物,烘干。②肝素-海藻酸钠-壳聚糖微胶囊的制备:取一定量的海藻酸钠和肝素水溶液乳化于石蜡油中,充分搅拌使反应体系呈乳状液,维持20min。然后缓慢滴加含有不同浓度壳聚糖的CaCl2水溶液,保持30min。离心分离,过滤洗涤,除去残留有机物,烘干。③测定药物含量与包封率,确定肝素标准曲线,测定肝素微胶囊体外缓释情况。主要观察指标:①壳聚糖溶液浓度对肝素-壳聚糖微胶囊制备的影响。②戊二醛用量对肝素-壳聚糖微胶囊制备的影响。③海藻酸钠溶液浓度对肝素-海藻酸钠的影响。④壳聚糖浓度对肝素-海藻酸钠-壳聚糖微胶囊制备的影响。⑤不同材料包裹的肝素微胶囊的体外释放情况。⑥肝素含量与包封率的测定结果。⑦肝素微胶囊扫描电镜观察结果。结果:①随着壳聚糖溶液初始的增大,产物颜色加深,颗粒度增大,但颗粒的均匀性和成球性提高。②戊二醛用量增大,产物颜色加深,且使产物相互黏着严重。未与壳聚糖作用的戊二醛也可以自身固化而呈不规则颗粒物。③随着海藻酸钠浓度的改变,产物的成球特性没有明显的变化。④随着壳聚糖浓度的增加,成球性好,但当浓度达到一定程度时,微球之间有相互黏结的现象。将壳聚糖的浓度控制在质量比为2%较好。⑤随着壳聚糖含量的增大,肝素的释放速度变慢。⑥当微囊中的壳聚糖含量质量比达到20%时,肝素的包封率可以达到58%。单纯利用壳聚糖包埋肝素,肝素的包封率可以达到79.9%。⑦含有壳聚糖的微胶囊表面比较致密,同时随着戊二醛含量的增大,微胶囊会粘连在一起。结论:壳聚糖在一定浓度下对微胶囊的均匀性和成球性存在影响,壳聚糖的用量可以改变肝素的包封率,且随着壳聚糖用量的增加肝素的包封率随之提高,同时肝素的缓释速率相应降低。

BACKGROUND： Chitosan and sodium alginate are the good natural materials for microcapsule, and also used widely in tissue engineering. Our research teams have made thorough work at anticoagulant materials, but these materials are inert or simulate the liquid crystal form of blood vessel wall. While in this experiment, on the base of our previous study, we microencapsulated heparin with biotic anticoagulation activity and other specific performances in order to enable microcapsule to have a long time releasing effect of medicine. OBJECTIVE： To microencapsulate the low molecular heparin so as to ensure the stability of heparin in vivo and analyze the effect of content of chitosan on the performance of heparin microcapsules basing on the natural chitosan and sodium alginate as the enwrapped materials of microcapsules. DESIGN： Open experiment. SETTING： Department of Material Science and Engineering, Jinan University. MATERIALS： The experiment was performed at the laboratory of Department of Material Science and Engineering, Jinan University from October 2004 to June 2005. Heparin, with relative molecular mass〈 5 000, was provided by Shandong Freda Biochem Co., Ltd.,; Chitosan was provided by Shanghai Bio Life Science ＆ Technology Co., Ltd, DD≥90%, η〈 100 cps; Sodium alginate was provided by Qingdao Bright Moon Seaweed Industrial Co., Ltd. Emulsions were Span80, and CaCl2, which were both made in China. METHODS： ① Preparation of heparin/chitosan microcapsules （HCM）： Some heparin aqueous solution was emulsified in liquid paraffin. The reaction system was stirred fully and presented emulsion. Then the whole reaction system was wanned to be at 50 ℃ and maintained for 20 minutes. Afterwards, 20 g/L chitosan solution was added slowly, subsequently with raising the temperature to be at 60 ℃ and then glutaraldehyde was dropwised keeping the reaction system at 80 ℃ for lhour. Centrifugation, filtration and washing followed by washing with kerosene fully, remain organic was extracted by dehydrated alcohol with extractor were performed. Drying and xeransis in＇ vacuum were done at last. ② Preparation of heparin-sodium alginate-chitosan microcapsules （HSCM） ：Heparin aqueous solution and sodium alginate were emulsified in paraffin, and the reaction system was stirred into emulsion at room temperature for 20 minutes, then 3% CaCl2 solution containing different concentrations of chitosan was added slowly. 30 minutes later, Microcapsules were separated, washed and dried as the treatments as before.③ Drug content and envelope efficiency were measured, heparin standard curve was determined and in vitro releasing effect of heparin microcapsules was also measured. MAIN OUTCOME MEASURES： ①Effect of chitosan solution concentration on preparation of heparin-chitosan microcapsules; ② Effect of glutaraldehyde dosage on preparation of heparin-chitosan microcapsules; ③ Effect of sodium alginate concentration on hepatin-sodium alginate; ④Effect of chitosan concentration on hepatin-sodium alginate-chitosan microcapsules. ⑤ In vitro release of heparin microcapsules enwrapped by different materials. ⑥Measurement of heparin content and envelope efficiency. ⑦ Observation of heparin microcapsule under scanning electron microscope. RESULTS：①With the increasing concentration of chitosan, the color of production changed from yellow to dark, and microcapsules were increscent, but the microcapsules uniformity and property of bailing were increased. ②The increasing content of glutaraldehyde led darker production. Increase of glutaraldehyde content made production bond each other severely. The glutaraldehyde, which did not react with chitosan, can solidify itself and presented anomalous microcapsules forming. ③There was not obvious bailing property of the production with the change of concentration of sodium alginate. ④The bailing property of microsphere was good with increasing concentration of chitosan. However, microcapsules conglutinated with each other. 2% chitosan would be better. ⑤With the increase of chitgsan content, the releasing speed of heparin became slow. ⑥The envelope efficiency was about 58% when microcapsule contained 20%（wt） of chitosan, and used chitosan only the envelope efficiency could approach to 79.9%. ⑦ The surface of microcapsules with chitosan was very compact, and with increasing of content of glutaraldehyde, microcapsules would bond each other. CONCLUSION： Chitosan at certain concentration will affect the uniformity and bailing property of microcapsules. Chitosan dosage can alter the envelope efficiency of hepafin. Envelope efficiency of heparin is increased and releasing speed of hepafin is decreased with the increase of content of chitosan.