CN108236730A - 复合生物纤维膜及其制造方法 - Google Patents
复合生物纤维膜及其制造方法 Download PDFInfo
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- CN108236730A CN108236730A CN201710169757.7A CN201710169757A CN108236730A CN 108236730 A CN108236730 A CN 108236730A CN 201710169757 A CN201710169757 A CN 201710169757A CN 108236730 A CN108236730 A CN 108236730A
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- cultured
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- compound bio
- fermented
- fiber film
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Abstract
本发明提供一种复合生物纤维膜及其制造方法，包括：提供菌种；进行二阶段发酵培养；进行纯化及清洗程序，形成生物纤维素，然后打碎生物纤维素；同时，提供葡萄糖胺与多糖体，将其以第一特定比例均匀混合，形成混合溶液；然后，将上述打碎之生物纤维素以第二特定比例均匀分散于混合溶液中；进行冻干程序；最后形成复合生物纤维膜敷料。其中，上述二阶段发酵培养，包括：第一阶段发酵培养；以及第二阶段发酵培养，第二阶段发酵培养的温度较第一阶段发酵培养的温度为高。本发明之复合生物纤维膜具有极佳之止血性与抗菌性。
Description
技术领域
本发明有关于一种复合生物纤维膜及其制造方法，特别是一种含葡萄糖胺等多糖体之生物纤维膜及其制造方法，其具有极佳之止血性(hemostatic)与抗菌性(antibacterial)。
背景技术
传统上，失血是许多场合，例如战场上，死亡最重要的原因之一，一般分为体内出血和体外出血，体内出血大部分是因为脏器破裂，体外出血基本是由于动静脉损伤。如果能及时有效地止血，对挽救伤员生命，稳定伤情，为后续治疗创造条件十分重要。而敷料作为止血材料，是指盖在伤口上、有保护作用的覆盖物，可以协助控制出血，防止感染并吸收分泌物，止血敷料对于及时止血有着重要的意义。
一般而言，创伤敷料具备以下之基本物理及生物机能性：止血性、抗菌性、防止水分或体液之丧失、具有正常皮肤之柔软性及伸缩性、适度之水蒸气或气体之透过性、渗出液之吸收性、湿润强度、可消毒或灭菌、不产生因生体内劣化而产生有害物质、创面之密着性、无毒性、上皮化之促进性、镇痛作用、无刺激性等等。其中，又以止血性与抗菌性为最主要的特性要求，近年来，市面上种种形式及材料之创伤敷材不断被开发出来。
多糖体(Polysaccharide)系由多个单糖分子脱水聚合，以糖苷键连接而成，可形成直链或者有分支的长链，水解后得到相应的单糖和寡糖，例如：用来储存能量的淀粉和肝糖，以及用来组成生物结构的纤维素和甲壳素。多糖体(Polysaccharide)常常由略带修饰的重复单元构成。由于结构不同，多糖高分子和构成它的单糖分子性质迥异，可能无定形，甚至不溶于水。一般而言，自然界中存在的糖类(例如葡萄糖、果糖和甘油醛)一般为单糖，通式为(CH2O)n，其中n≧3。而多糖的通式则为(CxH2O)y，其中x通常在200到2500之间。鉴于多糖通常由六碳糖构成，多糖的通式也可写作(C6H10O5)n，其中40≦n≦3000。多糖是一种重要的生物高分子，纤维素和甲壳素是两种常见组成生物结构的多糖。纤维素构成植物的细胞壁，可谓地球上数量最多的有机分子。纤维素应用广泛，不仅在造纸业和纺织业中举足轻重，而且是生产人造丝、醋酸纤维素、赛璐珞、硝化纤维等的原料。甲壳素结构和纤维素类似，但支链中含有氮，所以强度更高。其存在于节肢动物的外骨骼和真菌的细胞壁中。甲壳素也有很多作用，比如可用作手术缝合线。
葡萄糖胺(glucose-amine)的化学式为C6H13NO5或(1,4)-2-amino-2-deoxy-β-D-glucan，又称葡糖胺或胺基葡萄糖，是自然界含量最丰富的单糖之一。工业上通常采用水解甲壳类动物外骨骼以制取葡萄糖胺。葡萄糖胺衍生物N-乙酰胺基葡萄糖是甲壳素(Chitin)的单体，甲壳素(Chitin)广泛存在于节肢动物的外骨骼以及真菌的细胞壁之中，是一种化学构造式和纤维素类似之天然高分子/天然聚合物，性质通常是半透明、易弯、有弹性、和十分坚韧的。几丁聚糖(Chitosan)则是一种由葡萄糖胺所聚合而成的多糖体，为几丁质或甲壳素经脱乙酰化反应后所得到之高分子物质。
由葡萄糖胺所衍生或聚合之高分子物质(例如几丁聚糖)，在生医材料上的相关应用研究非常多，然而，由其直接制成或加工成的医用敷材，却不易与人体或动物的皮肤接触，在使用时容易有脱落、无法完全与皮肤密合、以及不利于被人体或动物的皮肤吸收等问题。
发明内容
有鉴于此，本发明提供一种复合生物纤维膜，其系一种含葡萄糖胺与多糖体之生物纤维膜，具有极佳的止血性(hemostatic)与抗菌性(antibacterial)，不仅具有快速止血作用、纤维之增殖阻害及促进组织再生之效果，同时亦具有处理伤口处细菌等之功能，进而可达到防止创伤面化脓之效果。
就其中一个观点，本发明提供一种复合生物纤维膜之制造方法，包括：提供菌种；进行二阶段发酵培养；进行纯化清洗程序，形成生物纤维素，并打碎生物纤维素；同时提供葡萄糖胺与多糖体，将葡萄糖胺与多糖体以第一特定比例均匀混合，形成混合溶液；然后，将打碎之生物纤维素以第二特定比例均匀分散于复合溶液中；以及，进行冻干程序，形成复合生物纤维膜敷料。其中，上述二阶段发酵培养，包括：一第一阶段发酵培养；以及一第二阶段发酵培养，第二阶段发酵培养的温度较第一阶段发酵培养的温度为高。
一实施例中，上述菌种包括醋酸菌属，较佳的是木质醋酸菌(Gluconacetobacterxylinum)。
一实施例中，上述第一阶段的发酵培养温度在第二阶段的发酵培养温度在
一实施例中，上述葡萄糖胺加入去离子水中，利用高速均质机进行高速搅拌，以形成糊状之凝胶。
一实施例中，上述多糖体，例如几丁聚糖，以第一特定比例溶于该混合溶液中，其重量百分比系介于之间。
一实施例中，上述第二特定比例，亦即该生物纤维素于该复合生物纤维膜中之重量百分比，系介于之间。
就其中一个观点，本发明提供一种复合生物纤维膜，采用申请专利范围第1项所述之制造方法，包括：一葡萄糖胺及/或一多糖体，该葡萄糖胺与该多糖体以一特定比例混合；以及，一生物纤维素，该生物纤维素重量百分比系介于之间，其中，该生物纤维素系经过二阶段发酵培养，包括一第一阶段发酵培养以及一第二阶段发酵培养，该第二阶段发酵培养的温度较该第一阶段发酵培养的温度为高。
一实施例中，上述特定比例系介于之间。
藉由底下实施例之详细说明，更容易了解本发明之具体功效。
附图说明
图1绘示系根据本发明之一实施例，一种复合生物纤维膜之制造方法流程示意图。
图2绘示系应用本发明之复合生物纤维膜，进行人血止血性评估之示意图。
符号说明：
S1：提供菌种
S2：两阶段发酵培养
S3：纯化清洗，形成生物纤维素
S4：打碎生物纤维素
S5：提供葡萄糖胺和多糖体
S6：特定比例均匀混合，形成混合溶液
S7：将生物纤维素均匀分散于混合溶液中
S8：进行冻干程序
S9：形成复合生物纤维膜敷料
具体实施方式
本发明的特色之一是利用生物纤维素与葡萄糖胺/多糖体的有效结合，包括透过调整生物纤维素与水的比例，再经由机械力作用打碎生物纤维素，再将生物纤维素均匀分散于特定比例之葡萄糖胺/多糖体混合溶液中，最后经冻干程序，将复合材料制造成不同型式之生物纤维膜敷料或海绵敷料。
本发明上述将生物纤维素(又称细菌纤维素)打碎的原因，是因为若是以含浸之方式将葡萄糖胺/多糖体材料导入生物纤维素中，容易受限于葡萄糖胺/多糖体之浓度与分子量影响，且吸血之菜单现有限。因此，本发明是将生物纤维素打碎分散，可均匀混合葡萄糖胺/多糖体混合溶液中，以作为止血敷料之强度补强。此外，多糖体于弱酸条件下会带正电，此带电之特性有助于出血时协助止血机制作用，故可提升细菌纤维素的止血表现。
此外，本发明另一特色是引进一种生物纤维微晶技术，以单菌培养方式生成生物纤维素，因为培养过程中全程在受控制的无麈室环境下进行，以及培养后续制程也受到严格管理，因此，能有效控制其纯净度。利用单一菌株、单片培养的方式，形成天然生物纤维素，系属于奈米级生物科技产品，其可以被环境自然分解，且其奈米多孔结构，可以阻隔外界微生物，达到保护皮肤的效果。本发明培养出的此种生物纤维素还具备材质柔软、高亲肤性、超饱和含水性等特质，使得葡萄糖胺和多糖体，或是甲壳素或几丁聚糖，可以完全地与人体或动物的皮肤接触密合，不容易脱落，且利于被人体或动物的皮肤吸收。
本发明的培养方式，为一种两阶段的发酵培养，其目的系用以控制发酵前后期间的生物纤维膜的形成速度。此种两阶段的培养包含两阶段培养温度的静置培养，包括第一阶段在室温的温度下进行培养，以及第二阶段在高于室温的温度下进行培养，以下将有进一步详细之叙述。
请参照第1图，其绘示系根据本发明之一实施例，一种复合生物纤维膜之制造方法流程示意图。首先，进行S1步骤，提供一菌种，较佳的是木质醋酸菌(Gluconacetobacterxylinum)。本发明所述之生物纤维膜可由微生物的培养而获得，此处之微生物包括可生产纤维素的细菌或真菌等，更具体地为醋酸菌属。
然后，进行S2步骤，进行二阶段发酵培养，包括：进行第一阶段发酵培养；以及进行第二阶段发酵培养，其中，第二阶段发酵培养的温度较第一阶段发酵培养的温度为高。本发明利用木质醋酸菌在装有液体培养基的培养盘中，进行单片培养，经过发酵程序(fermentation)产生的纤维素所构成。上述纤维素在菌体外形成横断面直径为约30～100nm的微生物纤维。复数个微生物纤维以非织造(nonwoven)的形态在培养基的表面形成网状结构(reticular structure)，形成半透明的生物纤维膜。
本发明之生物纤维膜的形成，主要是利用接菌量及培养条件的变化来形成较紧致的生物纤维膜结构。由于静置培养时，培养液的液面处是培养液与空气直接接触，溶氧量较高，可促进微生物的生长及生物纤维的生成，故生物纤维素的形成多发生在培养液的液面处。所以在培养初期，如果能将醋酸菌浓度降低及/或是降低微生物纤维素的生产速度，即可形成较柔软的微生物纤维素的网状结构；随后因培养时间增加，微生物的快速增生，使得微生物浓度相对增加；及/或是提高生产微生物纤维素的速度，微生物纤维素的产量也相对增加，以形成较为紧密的微生物纤维素网状结构。
本发明所述之发酵培养，除习知的发酵条件以外，更包括一种两阶段的发酵培养，来控制发酵前后期间的生物纤维膜的形成速度。该两阶段的培养包含两阶段培养温度的静置培养，包括第一阶段在室温的温度下进行培养，以及第二阶段在高于室温的温度下进行培养。更具体地说，第一阶段的培养温度在静置培养小时；第二阶段的培养温度在静置培养小时，但前述条件仅为例示不限于此。本发明所述之两阶段发酵培养可根据所欲形成的生物纤维膜之膜厚、含水量等因素，适当调整培养条件。
接着，进行S3步骤，进行纯化清洗程序，形成一生物纤维素。经二阶段发酵培养后，取出初步之培养成品，以0.1％～5％的NaOH清洗、除菌，然后静置一段时间。再重复以清水清洗次，再以柠檬酸中和后，得到初步的生物纤维素，以供后续使用。
以及，进行S4步骤，将上述生物纤维素打碎。例如系利用高速分散器将步骤S3形成之生物纤维素打碎。
然后，与S1步骤同时地或先行地，进行S5步骤，提供葡萄糖胺与多糖体等材料，将葡萄糖胺和多糖体以特定比例均匀混合，以形成一混合溶液。此处葡萄糖胺加入去离子水中。然后，利用高速均质机进行高速搅拌，例如为12,000rpm以上的速度，以形成糊状之凝胶。同时地或者可先行地，利用一酸性溶液溶解多糖体，其中多糖体例如可以是薄膜状，而酸性溶液例如可以是2％之醋酸溶液。
接着，进行S6步骤，将上述葡萄糖胺与上述溶解的多糖体均匀混合，特别地是以一特定比例均匀混合，该比例系介于(多糖体于混合溶液中之重量百分比)系介于之间，以形成一复合溶液。多糖体之重量百分比若大于20％，作为创伤覆材的医疗用时，对于体液之吸收性差，同时其柔软性差，对于人体或动物体皮肤之创伤处易造成不适之感觉，增加患者之疼痛。多糖体之重量百分比若小于0.2％，其抗拉强度差容易破损，特别是对于伤口渗出液体多之创伤更加容易破损，易造成手术上作业性之不便。因此，多糖体之重量百分比最好在之间，最佳为本发明主要利用具有促进表皮再生之葡萄糖胺及具预防化脓效果之薄膜状多糖体复合成形体，以开发出具有加强多孔性成型体之干、湿强度以减少其覆材应用上容易破损之缺点，同时具有防止伤口化脓加速伤口愈合之创伤用保护材。
然后，进行S7步骤，将已打碎之生物纤维素均匀分散于上述混合溶液中，其中生物纤维素之重量百分比较佳的是介于之间。
接着，进行S8步骤，进行冻干程序，最后，再加工制成一复合生物纤维膜敷料(S9步骤)，生物纤维膜厚度例如是在0.1mm～0.5mm之间。
一实施例中，本发明之复合生物纤维膜因经过两阶段发酵培养，具有较柔软的性质，若是平铺于人体或动物体的皮肤上，会产生优异的皮肤密合度。此外，藉由微生物纤维分子上大量的羟基(-OH)，容易与人体或动物体皮肤产生氢键，而与皮肤形成良好的密合效果，进而让生物纤维膜中的葡萄糖胺和多糖体能够易于快速被人体或动物的皮肤吸收。
请参照第2图，其绘示系应用本发明之复合生物纤维膜，进行人血止血性评估之实验数据示意图。可以看到的是，本发明之复合生物纤维膜，当应用在人体表面出血处时，在极短的时间内，例如约30秒左右，吸光值就达到了一稳定值，亦即人体表面已不再出血，成功达到了止血的功能。而市售的一般止血敷材，当应用在人体表面出血处时，即使到了约40秒左右，其吸光值仍一直不断下降，并未达到稳定值，虽达到了初步止血的功能但须时较长。
一实施例中，本发明之复合生物纤维膜还可含有水或其他活性成份，例如为抗生素(antibiotics)、抗菌剂(antimicrobials)、抗病毒药物(antivirals)、止血剂hemostatics)、麻醉剂(anesthetics)、减缓发炎药物、类固醇、抗组织胺、促进细胞生长药剂、生长激素(growth factors)、蛋白质(proteins)、核苷酸(nucleotides)、酵素(enzymes)、保湿剂、透明质酸(hyaluroic acid)、抗坏血酸(ascorbic acid)、面酸(kojicacid)、熊果素(arbutin)等类似的化合物、或这些化合物的组合。熟知此技术领域之人士可根据市场需求，适量添加于复合生物纤维膜中。
实验数据显示，本发明复合生物纤维膜具有止血作用、纤维之增殖阻害及促进组织再生之效果，同时亦具有活化巨嗤细胞、白血球之作用，故具处理伤口处细菌等之功能，进而具有防止创伤面化脓之效果。更进一步言，由于复合生物纤维膜具有带正电荷的胺基，因此还具有良好的抗菌性。本发明复合生物纤维膜还具有极佳的伤口愈合的能力，对细胞无排斥力，具有修复细胞之功效，并能减缓过敏性肌肤等之功能。此外，由于葡萄糖胺和多糖体具有良好的生物兼容性、无生物毒性、价格低廉、容易改质、机械强度佳等优点。因此，本发明复合生物纤维膜可适合被商品化，并制成或加工成各种创伤敷材、外科用可吸收手术缝合线、伤口敷料、或人造皮肤等医用材料。
综上所述，本发明提供之复合生物纤维膜，是一种具止血性、可消毒或灭菌、可抗菌且不产生因生体内劣化而产生有害物质、创面之密着性、无毒性、上皮化之促进性、镇痛作用、无刺激性的具有防止创伤面化脓效果之创伤覆材。更由于复合生物纤维均具有类似纤维的构造，所以本发明提供一种防止水分或体液之丧失、具有正常皮广之柔软性及伸缩性、适度之水蒸气或气体之透过性、渗出液之吸收性、湿润强度治疗创伤用之创伤覆材。
Claims (10)
1.一种复合生物纤维膜之制造方法，包括：
提供一菌种；
进行二阶段发酵培养，包括：
进行一第一阶段发酵培养；以及
进行一第二阶段发酵培养，该第二阶段发酵培养的温度较该第一阶段发酵培养的温度为高；
进行纯化及清洗程序，形成一生物纤维素；
提供一葡萄糖胺和一多糖体，将该葡萄糖胺与该多糖体以一第一特定比例均匀混合，形成一混合溶液；
打碎该生物纤维素，将该打碎之生物纤维素以一第二特定比例均匀分散于该混合溶液中；以及
进行冻干程序，形成一复合生物纤维膜敷料。
2.根据权利要求1所述的复合生物纤维膜制造方法，其中该菌种包括醋酸菌属。
3.根据权利要求2所述的复合生物纤维膜制造方法，其中该菌种包括木质醋酸菌(Gluconacetobacter xylinum)。
4.根据权利要求1所述的复合生物纤维膜制造方法，其中该第一阶段的发酵培养温度在
5.根据权利要求4所述的复合生物纤维膜制造方法，其中该第二阶段的发酵培养温度在
6.根据权利要求1所述的复合生物纤维膜制造方法，其中该打碎该生物纤维素之步骤可先于形成该混合溶液之步骤。
7.根据权利要求1所述的复合生物纤维膜制造方法，其中该第一特定比例指该多糖体于该混合溶液中之重量百分比系介于之间。
8.根据权利要求1所述的复合生物纤维膜制造方法，其中该第二特定比例指该生物纤维素于该复合生物纤维膜中之重量百分比系介于间。
9.一种复合生物纤维膜，采用根据权利要求1所述的制造方法，包括：
一葡萄糖胺和一多糖体，该葡萄糖胺以一特定比例与该多糖体混合；以及
一生物纤维素，该生物纤维素以重量百分比介于20％～60％之间的比例与该葡萄糖胺/该多糖体混合，其中，该生物纤维素系经过二阶段发酵培养，包括一第一阶段发酵培养以及一第二阶段发酵培养，该第二阶段发酵培养的温度较该第一阶段发酵培养的温度为高。
10.根据权利要求9所述的复合生物纤维膜，其中该葡萄糖胺之特定比例系介于之间。
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