CN108926741B - 一种羊膜生物补片的制备方法 - Google Patents

一种羊膜生物补片的制备方法 Download PDF

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CN108926741B
CN108926741B CN201810636306.4A CN201810636306A CN108926741B CN 108926741 B CN108926741 B CN 108926741B CN 201810636306 A CN201810636306 A CN 201810636306A CN 108926741 B CN108926741 B CN 108926741B
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陈俊
陈世益
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Huashan Hospital of Fudan University
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Abstract

本发明提供了一种羊膜生物补片的制备方法，其包括如下步骤：S1、将羊膜经紫外光照射后进行清洗；S2、将清洗后的羊膜浸泡于银离子溶液中，在超声条件下处理后，用紫外光照射，使羊膜表面修饰出纳米银涂层；S3、将修饰有纳米银涂层的羊膜浸泡于无菌生理盐水中，真空包装后在4℃下保存备用。与现有技术相比，本发明具有如下的有益效果：1、纳米银表面修饰可改善羊膜的组织诱导，并使其具有一定的预防感染、炎症的作用，促进羊膜生物补片与软组织的整合；2、该制备方法可在医用羊膜表面形成较为均匀的涂层，涂层结合力牢靠、稳定性良好；3、本制备过程简单、条件温和、节能环保、成本低廉，具有良好的市场应用前景。

Description

一种羊膜生物补片的制备方法
技术领域
本发明涉及一种羊膜生物补片的制备方法，属于医疗耗材技术领域。
背景技术
软组织例如韧带或者肌腱损伤是临床最为常见的疾病之一。目前临床常常采取手术将移植材料替代和修补受损的软组织。当前常用的移植材料包括自体、异体和人工移植物三大类。其中自体或者异体移植组织由于具有良好的生物活性，因此，它们占据主要市场份额。临床主要利用自体或者移植组织主要进行两类手术。1、修复增强，当软组织例如韧带或者肌腱受损撕裂时，通过手术将自体或者异体组织例如阔筋膜等对受损的软组织进行缝合，利用自体或者异体组织的天然生物活性，诱导自身组织再生，实现恢复受损组织的原来功能。2、替代手术，当软组织例如韧带或者肌腱断裂无法进行缝合修补的时候，通过取自体异体的肌腱进行重建手术，例如前交叉韧带断裂损失断裂时，通过取自体四股腘绳肌腱最为移植材料替代原来的前交叉韧带。但是不论是修复增强还是替代手术，因为受损的韧带或肌腱组织是维持身体运动的主要的力学结果，因此对于移植物的生物力学和生物活性都有着很高的要求，导致符合客观临床需求的自体或者异体移植组织不多。同时，随着临床的需要越来越多，同时自体或者异体组织来源受限，寻找新的移植材料是软组织修复领域具有非常重要的临床和经济意义。
羊膜是胎盘的最内层的主要由上皮细胞组成的薄膜，其表面光滑，无血管、神经及淋巴，具有良好的生物力学性能，厚约0.02～0.5mm，在电镜下，其分为五层：上皮层、基底膜、致密层、纤维母细胞层和海绵层。最新的研究发现，由于具有良好的生物活性以及力学性能，更为关键的是其无免疫原性，羊膜非常适合作为组织修复材料。目前，羊膜作为组织修复材料已经在临床上被广泛应用于眼科，泌尿外科和烧伤外科等。同时最新的动物研究也证实羊膜也可以作为肌腱的损伤修复材料。但是目前羊膜进行通过酶处理等灭菌或者冷冻干燥处理过程，会导致羊膜的生物活性和力学性能降低，这样会到作为软组织例如韧带或者肌腱这样的受力组织修复时效果。因此，如何最大限度保留羊膜的原有生物特性是羊膜生物制品临床应用前处理的关键技术。
银具有良好的抗菌消炎效果，同时又具有生物安全性，银相关的抗菌医用产品在临床广泛应用。值得注意最新的研究发现纳米银不仅保留银的抗菌消炎效果，同时具有良好的组织诱导性，在再生组织上前景十分广阔。目前，纳米银的相关制备方法非常多，但是对于如何在不影响羊膜力学性能和生物活性的情况下，将纳米银负载于羊膜表面，目前尚无的相关文献和专利报道。本发明专利通过利用紫外的天然杀菌消毒效果，以及在羊膜表面原位还原得到纳米银，对羊膜进行快速消毒处理，避免了额外的化学试剂实验以及额外的表面修饰反应，最大限度保留了羊膜的天然活性，并提高羊膜的抗菌消炎性和生物诱导性，促进软组织如肌腱细胞粘附、增殖，从而促进羊膜移植物与宿主组的整合，并在术后有效降低滑膜炎、感染等并发症的发现。
发明内容
本发明的目的在于针对上述现有技术存在的缺陷，提供一种纳米银修饰羊膜生物补片的制备方法；主要解决羊膜生物补片在前处理中生物活性下降以及临床应用中可能导致的感染、炎症问题。本发明利用紫外和纳米银材料能够机械破坏细胞膜并氧化胞内代谢物质，且不易产生耐药的优势，对于羊膜生物补片进行高效前处理，表面进行纳米银修饰，制备得到生物力学性能佳，组织诱导性良好，具有预防感染、炎症作用的新型羊膜生物补片，以减少在软组织修复手术后可能发生的感染、炎症等并发症，加强羊膜与宿主组织的整合。
本发明是通过以下技术方案实现的：
本发明提供了一种羊膜生物补片的制备方法，其包括如下步骤：
S1、将羊膜经紫外光照射后进行清洗；
S2、将清洗后的羊膜浸泡于银离子溶液中，在超声条件下处理后，用紫外光照射，使羊膜表面修饰出纳米银涂层；
S3、将修饰有纳米银涂层的羊膜浸泡于无菌生理盐水中，真空包装后在4℃下保存备用。
作为优选方案，所述紫外光的功率为50～100W、波长为120～380nm。
作为优选方案，所述银离子溶液的制备方法为：
将2.5mg的硝酸银粉末溶解于5mL去离子水中，在40℃下超声振荡10min，得到浓度为0.5mg/mL的银离子溶液。
作为优选方案，步骤S2中所述的超声处理时间为0.5～15min。
作为优选方案，步骤S3中所述的羊膜在无菌缓冲液中的浸泡深度为1～10mm。
作为优选方案，所述羊膜的来源为人、猪、牛、羊或狗。
作为优选方案，所需的银离子溶液是所需的银离子溶液是硝酸银，硫酸银，醋酸银的一种或者几种。
与现有技术相比，本发明具有如下的有益效果：
1、纳米银表面修饰可改善羊膜的组织诱导，并使其具有一定的预防感染、炎症的作用，促进羊膜生物补片与软组织的整合；
2、该制备方法可在医用羊膜表面形成较为均匀的涂层，涂层结合力牢靠、稳定性良好；
3、本制备过程简单、条件温和、节能环保、成本低廉，具有良好的市场应用前景。
附图说明
通过阅读参照以下附图对非限制性实施例所作的详细描述，本发明的其它特征、目的和优点将会变得更明显：
图1为本发明的羊膜生物补片的制备方法流程示意图。
具体实施方式
下面结合具体实施例对本发明进行详细说明。以下实施例将有助于本领域的技术人员进一步理解本发明，但不以任何形式限制本发明。应当指出的是，对本领域的普通技术人员来说，在不脱离本发明构思的前提下，还可以做出若干变形和改进。这些都属于本发明的保护范围。
本发明公开一种软组织修复的新型羊膜生物补片的制备方法。本发明方法操作简单，条件温和，成本低且对环境污染小；纳米银涂层修饰后的羊膜生物补片的生物力学性能佳、组织诱导性良好、具有抗菌抗炎的性能，是一种对目前羊膜生物补片产品前处理以及临床应用中所存在的缺陷进行改良的简单方法，拥有巨大的市场应用价值。
具体如以下各实施例：
实施例1
如图1所示，将羊来源的羊膜进行简单的裁剪，得到长约5cm的正方形小片，将此羊膜片置于功率100W，波长185nm的紫外灯下密闭空间内照射下，3min内用去离子水冲洗2遍，使羊膜生物补片表面的污垢被清除。
在室温条件下，以去离子水为溶剂，将2.5mg的硝酸银粉末溶解于5mL去离子水中，置于超声震荡仪中40℃震荡10min，得到浓度为0.5mg/mL的银离子溶液。配制好银离子溶液的1min内，将上述清洁后的羊膜浸泡于新鲜制备的0.5mg/mL的银离子溶液中，在超声仪内超声处理1min，使体系中的银离子能均匀分布在羊膜表面，然后迅速转移到紫外灯照射下的反应皿，使羊膜表面与去离子水的页面高度在于1cm左右，在紫外灯照射下，充分让银离子与羊膜片充分反应，静置反应1min后，用镊子轻轻提拉翻面，再照射反应1min后取出。将反应完毕的羊膜生物补片用去离子水重新两遍，然后将所制备的纳米银修饰的羊膜补片浸泡于无菌生理盐水进行真空包装，4℃下保存待用。
所制备的羊膜生物补片可以单层也可以是多层叠加使用，其单层拉伸强度大于10MPa，所负载的纳米银颗粒与羊膜的结合力大于5MPa，植入体内4周内拉伸强度不小于原有拉伸强度，同时羊膜生物补片与宿主组织开始融合，有新生组织形成且无免疫排斥反应，1年后产品在体内完全降解。
实施例2
将人来源的羊膜片裁剪成长约10cm，宽约3cm的片，在功率300W，波长250nm的紫外灯下密闭空间内照射下，然后将此羊膜片5min内生理盐水，冲洗3遍使羊膜生物补片表面无明显污垢。在7℃条件下，以去离子水为溶剂，将5mg的醋酸银粉末溶解于5mL去离子水中，置于超声震荡仪中7℃震荡4min，得到浓度为1mg/mL的银离子溶液。迅速将表面清洁好的羊膜浸泡此1mg/mL的银离子溶液中，使体系中的银离子能够充分分布在羊膜表面，功率200W，波长360nm的紫外灯照射下，将表面覆盖银离子溶液的羊膜生物补片充分反应，静置3min后，用镊子轻轻提拉翻面，再照射3min后取出。将反应完毕的羊膜生物补片用去离子水重新两遍，然后将所制备的纳米银修饰的羊膜补片浸泡于无菌生理盐水进行真空包装，4℃下保存待用。
所制备的羊膜生物补片的拉伸强度大于10MPa，所负载的纳米银颗粒粒径在50～100nm之间。
实施例3
将牛来源的羊膜片裁剪成长约10cm，宽约8cm的片，置于功率100W，波长380nm的紫外灯下，用去离子水冲洗5遍。在4℃条件下，以去离子水为溶剂，将5mg的硫酸银粉末溶解于50mL去离子水中，置于超声震荡仪中4℃震荡3min，得到浓度为0.1mg/mL的银离子溶液。迅速将表面清洁好的的羊膜浸泡于0.1mg/mL的银离子溶液中，超声振荡15min，使体系中的银离子能够充分分布在羊膜表面。然后功率100W，波长380nm的紫外灯照射下，将此羊膜片静置于生理盐水中，液面高度为10mm，紫外照射3min，然后用镊子翻面再照射3min后取出。将反应完毕的羊膜生物补片用生理盐水水重新两遍，然后将所制备的纳米银修饰的羊膜补片浸泡于无菌生理盐水进行真空包装，4℃下保存待用。
所制备的羊膜生物补片的拉伸强度大于10MPa，所负载的纳米银颗粒粒径在10～100nm之间。
实施例4
将猪来源的羊膜片裁剪成长约2cm，宽约2cm的片，置于功率50W，波长120nm的紫外灯下，在2min内用去离子水冲洗2遍。在室温条件下，以去离子水为溶剂，将100mg的硫酸银粉末溶解于10mL去离子水中，置于超声震荡仪中室温条件震荡15min，得到浓度为10mg/mL的银离子溶液。迅速将表面清洁好的的羊膜浸泡于10mg/mL的银离子溶液中，超声振荡1min，使体系中的银离子能够充分分布在羊膜表面。然后功率50W，波长120nm的紫外灯照射下，室温条件下将此羊膜片静置于生理盐水中，液面高度为1mm，紫外照射1min，然后用镊子翻面再照射1min后取出。将反应完毕的羊膜生物补片用生理盐水水重新两遍，然后将所制备的纳米银修饰的羊膜补片浸泡于无菌生理盐水进行真空包装，4℃下保存待用。
所制备的羊膜生物补片的拉伸强度大于10MPa，所负载的纳米银颗粒粒径在5～50nm之间。
以上对本发明的具体实施例进行了描述。需要理解的是，本发明并不局限于上述特定实施方式，本领域技术人员可以在权利要求的范围内做出各种变形或修改，这并不影响本发明的实质内容。

Claims (4)

1.一种羊膜生物补片的制备方法，其特征在于，包括如下步骤：
S1、将羊膜经紫外光照射后进行清洗；
S2、将清洗后的羊膜浸泡于银离子溶液中，在超声条件下处理后，用紫外光照射，使羊膜表面修饰出纳米银涂层；
S3、将修饰有纳米银涂层的羊膜浸泡于无菌生理盐水中，真空包装后在4℃下保存备用；
所述紫外光的功率为50～100W、波长为120～380nm；
所述银离子溶液的制备方法为：
将2.5mg的硝酸银粉末溶解于5mL去离子水中，在40℃下超声振荡10min，得到浓度为0.5mg/mL的银离子溶液。
2.如权利要求1所述的羊膜生物补片的制备方法，其特征在于，步骤S2中所述的超声处理时间为0.5～15min。
3.如权利要求1所述的羊膜生物补片的制备方法，其特征在于，步骤S3中所述的羊膜在无菌缓冲液中的浸泡深度为1～10mm。
4.如权利要求1所述的羊膜生物补片的制备方法，其特征在于，所述羊膜的来源为人、猪、牛、羊或狗。
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CN101623518A (zh) * 2009-06-30 2010-01-13 中国人民解放军第二军医大学 一种抗感染生物衍生疝和体壁修复材料及制备和应用
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