CN103272268A - 一种抗菌角膜修复材料及其制备方法 - Google Patents
一种抗菌角膜修复材料及其制备方法 Download PDFInfo
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Abstract
本发明公开了一种抗菌角膜修复材料的制备方法,包括以下步骤:(1)将从牛筋中提取的Ι型胶原纯化,用乙酸或盐酸溶液配制浓度为6.0~10.0mg/ml的胶原溶液;(2)将胶原溶液浇铸到角膜修复材料成型模具中,在室温条件下自然风干成膜;(3)将所得的胶原膜浸泡到抗生素溶液中,搅拌使膜与溶液充分接触;然后向上述溶液中加入交联剂和催化剂,搅拌发生交联反应;(4)将步骤(3)所得的交联膜材料取出,用去离子水冲洗3~5次,然后在室温条件下自然风干,即得到抗菌角膜修复材料。该材料具有较好的力学性能、光学性能和生物相容性,且具有良好的抗菌效果。可用于医疗领域中受损角膜组织的修复和替代。
Description
技术领域
本发明涉及一种抗菌角膜修复材料的制备方法,具体涉及到一种能够在修复受损角膜组织的同时还能降低发生角膜炎症反应的风险的生物材料的制备方法。本发明所得的材料可应用于受损角膜组织的修复和替代。
背景技术
角膜疾病是一种十分常见的眼科病,世界上约有三分之一的失明患者是由于角膜疾病而导致的。角膜移植是治疗角膜盲的一种有效手段,而同种异体的捐赠角膜是目前临床上唯一可行的角膜修复材料。但是由于健康的捐赠角膜的数量远少于角膜移植的需求量,因此人工角膜的研发就显得十分必要。
除了角膜移植材料的匮乏之外,如何避免角膜移植手术后的炎症反应是角膜移植能否成功所面临的另外一个难题。目前常见的方法就是在术后频繁地点药,这种方法不但浪费了大量的药物,同时很难保证药物的用量和加药时间,对患者本身来说又是一件极其麻烦的事情。因此很多组织工程修复材料会通过对药物的包裹或吸附等物理作用来实现药物负载,而针对角膜独特的光学透明性和韧性要求,这类修复材料往往很难保证材料在具有药物缓释效果的同时还具有较好的光学性能和力学性能。因此在保证角膜修复材料所需理化性能和生物学性能的基础上,使之在术后伤口恢复的一段时间里具有抗菌的效果,那将极大地降低角膜移植手术后发生炎症反应的风险。
发明内容
本发明的目的在于克服现有技术的缺点,提供一种新的抗菌角膜修复材料的制备方法。本发明采用高纯度Ι型胶原和眼科手术后常用的抗生素小分子为原料,按照一定的比例,经过模具成膜,抗生素溶液浸泡,搅拌交联,洗涤和风干等工艺,得到一种具有较好的力学性能,光学性能和生物相容性的角膜修复材料。同时,本发明提出的抗菌角膜修复材料还能在一定时间内对导致眼科手术后炎症反应的诸如金黄色葡萄球菌等细菌具有良好的抗菌效果。本材料可用于医疗领域中受损角膜组织的修复和替代,同时还能降低移植手术后细菌感染的风险。
本发明的目的通过以下技术方案实现:
一种抗菌角膜修复材料的制备方法,包括以下步骤:
(1)将从牛筋中提取的Ι型胶原纯化,用乙酸或盐酸溶液配制浓度为6.0~10.0mg/ml的胶原溶液;
(2)将胶原溶液浇铸到角膜修复材料成型模具中,在室温条件下自然风干成膜;
(3)将步骤(2)所得的胶原膜浸泡到浓度为5.0~25.0mg/ml的抗生素溶液中,搅拌使膜与溶液充分接触;然后向上述溶液中加入交联剂和催化剂,搅拌发生交联反应;所述交联剂与催化剂的质量比为4:1,胶原和抗生素的总质量与交联剂的质量比为(5~7):1;
(4)将步骤(3)所得的交联膜材料取出,用去离子水冲洗3~5次,然后在室温条件下自然风干,即得到抗菌角膜修复材料。
优选地,步骤(2)中所述角膜修复材料成型模具具有与角膜组织相似的几何外形。
优选地,步骤(3)所述抗生素的浓度为10.0~15.0mg/ml。
优选地,步骤(3)所述抗生素为妥布霉素、庆大霉素、氧氟沙星或环丙沙星。
优选地,步骤(3)所述交联剂为1-乙基-3(3-二甲基氨丙基)碳化二亚胺(EDC),催化剂为N-羟基丁二酰亚胺(NHS)。
优选地,所述胶原和抗生素的总质量与交联剂的质量比为6:1。
优选地,步骤(3)中所述交联反应的时间为2~6小时。
与现有技术相比,本发明具有如下优点:
(1)本发明制备的角膜修复材料能够在术后伤口恢复的一段时间里具有抗菌性,能够极大地降低角膜移植手术后发生炎症反应的风险。
(2)本发明添加抗生素后制备的角膜修复材料仍然具有与天然角膜组织类似的光学性能、力学性能和生物学性能。
(3)本发明制备的角膜修复材料能够极大地方便患者的术后护理,并降低大量药物所需的费用。
(4)本发明所采用的成型工艺简单,原料成本较低,有利于规模生产。
附图说明
图1是实施例1的Tob-Col膜的样品图。
图2是实施例1的Tob-Col膜与对照组Col膜的透光率曲线图。
图3是实施例1的Tob-Col膜与对照组Col膜的抗张强度图。
图4是金黄色葡萄球菌在Col膜上的生长图。
图5是金黄色葡萄球菌在实施例1的Tob-Col膜上的生长图。
图6是实施例1的Tob-Col膜与对照组Col膜的细胞毒性测试结果图。
图7是人眼角膜上皮细胞在实施例1的Tob-Col膜材料表面的生长图。
图8是实施例1的Tob-Col膜在动物眼表的角膜移植图。
具体实施方式
为了更好的理解本发明,下面结合实施例对本发明做进一步地说明,但本发明要求保护的范围并不局限于此。
实施例1
以妥布霉素(Tobramycin)和胶原(Col)为原料,制备一种妥布霉素-胶原交联膜(Tob-Col)材料。这种Tob-Col膜材料的制备步骤如下:
(1)将从牛筋中提取的Ι型胶原纯化,用盐酸配制浓度为6mg/mL的胶原溶液;
(2)将8mL浓度为6mg/mL的胶原溶液浇铸到角膜修复材料成型模具中,在室温条件下自然风干成膜;
(3)将步骤(2)所得的胶原膜浸泡到浓度为15mg/mL的妥布霉素溶液中,搅拌使胶原膜与妥布霉素溶液充分接触;
(4)在步骤(3)的溶液中加入EDC和NHS(EDC:NHS=4:1),其中胶原和妥布霉素的总质量与EDC的质量比为6:1,搅拌发生交联反应,2小时之后将所得的交联膜材料取出,用去离子水冲洗3次,然后在室温条件下自然风干得到Tob-Col膜。
实施例2
以庆大霉素(Gentamicin)和胶原(Col)为原料,制备一种庆大霉素-胶原交联膜(Gen-Col)材料。这种Gen-Col膜材料的制备步骤如下:
(1)将从牛筋中提取的Ι型胶原纯化,用盐酸配制浓度为8mg/mL的胶原溶液;
(2)将8mL浓度为8mg/mL的胶原溶液浇铸到角膜修复材料成型模具中,在室温条件下自然风干成膜;
(3)将步骤(2)所得的胶原膜浸泡到浓度为15mg/mL的庆大霉素溶液中,搅拌使胶原膜与庆大霉素溶液充分接触;
(4)在步骤(3)的溶液中加入EDC和NHS(EDC:NHS=4:1),其中胶原和庆大霉素的总质量与EDC的质量比为6:1,搅拌3小时使抗生素小分子与胶原发生交联反应;
(5)将步骤(4)所得的交联膜材料取出,用去离子水冲洗3次,然后在室温条件下自然风干得到Gen-Col膜。
实施例3
以环丙沙星(Ciprofloxacin)和胶原(Col)为原料,制备一种环丙沙星-胶原交联膜(Cip-Col)材料。这种Cip-Col膜材料的制备步骤如下:
(1)将从牛筋中提取的Ι型胶原纯化,用乙酸配制浓度为8mg/mL的胶原溶液;
(2)将8mL浓度为8mg/mL的胶原溶液浇铸到角膜修复材料成型模具中,在室温条件下自然风干成膜;
(3)将步骤(2)所得的胶原膜浸泡到浓度为5mg/mL的环丙沙星溶液中,搅拌使胶原膜与环丙沙星溶液充分接触;
(4)在步骤(3)的溶液中加入EDC和NHS(EDC:NHS=4:1),其中胶原和环丙沙星的总质量与EDC的质量比为6:1,搅拌4小时使环丙沙星分子与胶原发生交联反应;
(5)将步骤(4)所得的交联膜材料取出,用去离子水冲洗3次,然后在室温条件下自然风干得到Cip-Col膜。
实施例4
以氧氟沙星(Ofloxacin)和胶原(Col)为原料,制备一种氧氟沙星-胶原交联膜(Ofl-Col)材料。这种Ofl-Col膜材料的制备步骤如下:
(1)将从牛筋中提取的Ι型胶原纯化,用盐酸配制浓度为10mg/mL的胶原溶液;
(2)将8mL浓度为10mg/mL的胶原溶液浇铸到角膜修复材料成型模具中,在室温条件下自然风干成膜;
(3)将步骤(2)所得的胶原膜浸泡到浓度为25mg/mL的氧氟沙星溶液中,搅拌使胶原膜与氧氟沙星溶液充分接触;
(4)在步骤(3)的溶液中加入EDC和NHS(EDC:NHS=4:1),其中胶原和氧氟沙星的总质量与EDC的质量比为6:1,搅拌6小时使抗生素小分子与胶原发生交联反应;
(5)将步骤(4)所得的交联膜材料取出,用去离子水冲洗3次,然后在室温条件下自然风干得到Ofl-Col膜。
通过对实施例1中的材料进行理化性能表征,图1和图2显示该Tob-Col角膜修复材料具有较好的光学性能,图3表明该Tob-Col膜具有合适的力学性能;图4和图5表明该Tob-Col膜相对Col膜而言,具有较好的抗菌性能;图6和图7分别显示了人眼角膜上皮细胞在Tob-Col膜表面的增殖以及细胞的形貌,细胞能够在材料表面稳定增殖,紧密地贴附于材料表面并且保持细胞正常的纺锤形貌。图8是将新西兰大白兔的自身角膜用环钻和刀片切除和剥离后,将Tob-Col膜进行板层移植后的术后照片,可以看到本材料能够耐受眼科手术缝线的缝合,并且能在眼表保持透明。
Claims (8)
1.一种抗菌角膜修复材料的制备方法,其特征在于,包括以下步骤:
(1)将从牛筋中提取的Ι型胶原纯化,用乙酸或盐酸溶液配制浓度为6.0~10.0mg/ml的胶原溶液;
(2)将胶原溶液浇铸到角膜修复材料成型模具中,在室温条件下自然风干成膜;
(3)将步骤(2)所得的胶原膜浸泡到浓度为5.0~25.0mg/ml的抗生素溶液中,搅拌使膜与溶液充分接触;然后向上述溶液中加入交联剂和催化剂,搅拌发生交联反应;所述交联剂与催化剂的质量比为4:1,胶原和抗生素的总质量与交联剂的质量比为(5~7):1;
(4)将步骤(3)所得的交联膜材料取出,用去离子水冲洗3~5次,然后在室温条件下自然风干,即得到抗菌角膜修复材料。
2.根据权利要求1所述的制备方法,其特征在于,步骤(2)中所述角膜修复材料成型模具具有与角膜组织相似的几何外形。
3.根据权利要求1所述的制备方法,其特征在于,步骤(3)所述抗生素的浓度为10.0~15.0mg/ml。
4.根据权利要求1所述的制备方法,其特征在于,步骤(3)所述抗生素为妥布霉素、庆大霉素、氧氟沙星或环丙沙星。
5.根据权利要求1或2或3或4所述的制备方法,其特征在于,步骤(3)所述交联剂为1-乙基-3(3-二甲基氨丙基)碳化二亚胺,催化剂为N-羟基丁二酰亚胺。
6.根据权利要求1或2或3或4所述的制备方法,其特征在于,所述胶原和抗生素的总质量与交联剂的质量比为6:1。
7.根据权利要求1或2或3或4所述的制备方法,其特征在于,步骤(3)中所述交联反应的时间为2~6小时。
8.权利要求1~7任意一项方法制备的抗菌角膜修复材料。
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