CN105749340B - 一种纳微米纤维组织工程三维复合支架及其制备方法 - Google Patents
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Abstract
本发明涉及一种纳微米纤维组织工程三维复合支架及其制备方法，它采用在一定条件下分别静电纺丝纳米和微米壳聚糖海藻酸钠复合纤维膜，复合纤维膜经碱洗风干后按一定配比将纳米和微米壳聚糖海藻酸钠复合纤维膜在水中高速剪切成短纤维，分样后冷冻干燥得到纳微米纤维壳聚糖海藻酸钠三维复合支架。壳聚糖和海藻酸钠复合克服两者的缺点，该制备方法工艺简单，复合支架良好的生物学性能和力学性能。
Description
技术领域
本发明涉及一种纳微米纤维组织工程三维复合支架及其制备方法，属于生物医用材料技术领域。
背景技术
壳聚糖作为一种阳离子碱性多糖，结构类似于人体内的硫酸软骨素和透明质酸等多糖类物质，在体内可被降解吸收，并具有止痛、止血、抑菌、促进伤口愈合、减小疤痕等优异特性，目前在生物医用材料领域有重要的应用。Sundararajan V等人采用相分离技术首次制备了壳聚糖支架，其结构为数十到数百微米厚度的片状蜂窝结构，具有较好的生物学性能（Biomaterials,1999;20:1133-1142），但壳聚糖制成的支架易于溶胀，结构稳定性较差。海藻酸钠可从天然褐藻中提取，是一种由β-D-甘露糖醛酸和α-L-古洛糖醛酸连结而成的阴离子多糖，具有良好的生物相容性。海藻酸钠具有良好的亲水性，可增强蛋白质的吸附，但过强的亲水性使其与细胞的结合力较差，且其二价盐在环境中易被一价盐离子取代而溶解。Zhang Miqin课题组利用相分离法制备了壳聚糖-海藻酸钠聚电解质三维支架，该复合支架具有良好的结构稳定性和生物学性能，可用于骨组织的修复（Kievit FM 等，Biomaterials, 2010, 31:5903）。马小军等公开了一种壳聚糖/海藻酸钠组织工程支架的制备方法，该支架具有较好的力学强度和吸水性。
近年来的研究表明纳米纤维支架有较大的比表面积，可提供细胞粘附和分化的更好条件，但单纯的纳米纤维力学强度较低，无法承受支架植入过程的力学载荷。相比而言，微米纤维支架具有较好的力学性能。因此，具有微纳米纤维的三维支架具有良好的生物相容性和力学性能。目前研究者多采用相分离的方法制备三维支架，但控制其纤维直径则比较困难，或采用静电纺丝的方法获得二维的纳米和微米纤维薄膜，但如何制备具有微纳米纤维的三维壳聚糖海藻酸钠复合支架制备方法尚未见报导。
发明内容
本发明的主要设想是采用静电纺丝的方法分别制备纳米和微米壳聚糖海藻酸钠复合纤维薄膜，按一定配比将纳米和微米复合纤维膜混合，并将复合薄膜高速剪切成短纤维，分样后，经冷冻干燥得到纳微米纤维壳聚糖海藻酸钠复合支架三维支架。
本发明中纳微米纤维壳聚糖三维支架的制备方法具体如下所述。
(1) 静电纺丝纳米壳聚糖纤维膜：配制壳聚糖5.0%的溶液，溶剂为三氟乙酸和二噁烷的混合物，体积比为70:30；配制海藻酸钠3.0%的溶液，溶剂为水。将壳聚糖和海藻酸钠溶液分别加入两个注射器中，用二合一针头将两个注射器连接，通以20～25kV的电压进行纺丝。
(2) 静电纺丝微米壳聚糖纤维膜：配制壳聚糖7.5%的溶液，溶剂为三氟乙酸和二噁烷的混合物，体积比为90:10；配制海藻酸钠6.0%的溶液，溶剂为水。将壳聚糖和海藻酸钠溶液分别加入两个注射器中，用二合一针头将两个注射器连接，通以12～15kV的电压进行纺丝。
(3) 将静电纺丝得到的纳米和微米壳聚糖海藻酸钠复合纤维膜浸泡在0.05mol/L的氢氧化钙溶液中去除未挥发的溶剂以及交联海藻酸钠，水洗风干后待用。
(4) 分别称取定量的纳米和微米壳聚糖海藻酸钠复合纤维膜，加入到定量的水中，在水中高速剪切成短纤维。
(5) 分样至称量瓶中，在-18℃下冷冻成型。冻干后可获得纳微米纤维壳聚糖海藻酸钠复合支架。
本发明制得的纳微米纤维壳聚糖海藻酸钠复合支架外观为白色圆盘，其形状和高度可在分样中采用不同的容器和分样体积调节。壳聚糖和海藻酸钠的复合可克服两者的缺点，支架具有更好的综合性能。制备方法中主要通过静电纺丝获得纳微米尺度的纤维，制备中也可方便的调整纺丝条件得到不同尺度的纳米和微米壳聚糖海藻酸钠复合纤维。该支架具有微纳米纤维，赋予支架良好的生物相容性和力学强度。该制备方法工艺简单，易于得到不同直径的壳聚糖海藻酸钠复合纤维，相比于单纯的静电纺丝法，该法还可容易地制备三维结构的支架。
具体实施方式
下面结合具体实施例，对本发明内容作进一步的说明，但本发明的实现方式并不局限于此。
实施例1：分别取7ml的三氟乙酸和3ml的二噁烷，混合均匀，称取0.5g壳聚糖，搅拌溶解，静置12小时。取10ml的水，称取0.3g海藻酸钠，搅拌溶解，静置12小时。将壳聚糖和海藻酸钠溶液分别加入到两个注射器中，用二合一针头将两个注射器连接，通以20kV的电压纺丝，收集在接地的铝箔上，可获得壳聚糖海藻酸钠纳米复合纤维薄膜，纤维直径在50-500nm之间。分别取9ml的三氟乙酸和1ml的二噁烷，混合均匀，称取0.75g壳聚糖，搅拌溶解，静置12小时。取10ml的水，称取0.3g海藻酸钠，静置12小时。将壳聚糖和海藻酸钠溶液分别加入到两个注射器中，用二合一针头将两个注射器连接，通以15kV的电压纺丝，收集在接地的铝箔上，可获得壳聚糖海藻酸钠微米复合纤维薄膜，纤维直径在1-10μm之间。将静电纺丝得到的纳米和微米纤维膜浸泡在0.05mol/L的氢氧化钙溶液中去除未挥发的溶剂以及交联海藻酸钠，水洗风干。分别称取0.2g和0.8g纳米和微米壳聚糖海藻酸钠复合纤维膜，在100ml水中高速剪切成短纤维，分样至称量瓶中，在-18℃下冷冻成型。冻干后可获得纳微米纤维壳聚糖海藻酸钠复合支架。
实施例2：分别取14ml的三氟乙酸和6ml的二噁烷，混合均匀，称取1.0g壳聚糖，搅拌溶解，静置12小时。取20ml的水，称取0.6g海藻酸钠，搅拌溶解，静置12小时。将壳聚糖和海藻酸钠溶液分别加入到两个注射器中，用二合一针头将两个注射器连接，通以25kV的电压纺丝，收集在接地的铝箔上，可获得壳聚糖海藻酸钠纳米复合纤维薄膜，纤维直径在50-500nm之间。分别取9ml的三氟乙酸和1ml的二噁烷，混合均匀，称取0.75g壳聚糖，搅拌溶解，静置12小时。取10ml的水，称取0.6g海藻酸钠，搅拌溶解，静置12小时。将壳聚糖和海藻酸钠溶液分别加入到两个注射器中，用二合一针头将两个注射器连接，通以12kV的电压纺丝，收集在接地的铝箔上，可获得壳聚糖海藻酸钠微米复合纤维薄膜，纤维直径在1-10μm之间。将静电纺丝得到的纳米和微米纤维膜浸泡在0.05mol/L的氢氧化钙溶液中去除未挥发的溶剂以及交联海藻酸钠，水洗风干。分别称取1.2g和0.3g纳米和微米壳聚糖海藻酸钠复合纤维膜，在100ml水中高速剪切成短纤维，分样至称量瓶中，在-18℃下冷冻成型。冻干后可获得纳微米纤维壳聚糖海藻酸钠复合支架。
Claims (4)
1.一种纳微米纤维组织工程三维复合支架，其特征在于在一定条件下分别静电纺丝纳米和微米壳聚糖海藻酸钠复合纤维膜，在0.05mol/L的氢氧化钙溶液中去除未挥发的溶剂以及交联海藻酸钠，并水洗风干后按一定配比将纳米和微米壳聚糖海藻酸钠复合纤维膜在水中高速剪切成短纤维，分样后，经冷冻干燥得到纳微米纤维壳聚糖海藻酸钠复合三维支架。
2.根据权利要求1所述的纳微米纤维组织工程三维复合支架，其特征在于静电纺丝纳米纤维膜时，壳聚糖浓度为5.0%，溶剂为三氟乙酸和二噁烷的混合物，体积比为70:30，海藻酸钠浓度为3%，溶剂为水，纺丝电压为20～25kV。
3.根据权利要求1所述的纳微米纤维组织工程三维复合支架，其特征在于静电纺丝微米壳聚糖纤维膜时，壳聚糖浓度为7.5%，溶剂为三氟乙酸和二噁烷的混合物，体积比为90:10，海藻酸钠浓度为6.0%，溶剂为水，纺丝电压为12～15kV。
4.根据权利要求1所述的纳微米纤维组织工程三维复合支架，其特征在于纳米壳聚糖海藻酸钠复合纤维和微米壳聚糖海藻酸钠复合纤维的质量比为20:80或80:20，壳聚糖海藻酸钠纳米微米纤维在水中的总浓度为1%(w/v)或1.5%(w/v)。
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