技术领域technical field
本发明属于纳米纤维管状材料的制备领域，特别涉及一种聚氨酯-角蛋白复合纳米纤维管状材料的制备方法。The invention belongs to the field of preparation of nanofiber tubular materials, in particular to a preparation method of polyurethane-keratin composite nanofiber tubular materials.
背景技术Background technique
角蛋白是一种不溶性的纤维状动物蛋白质，是外胚层细胞的结构蛋白，广泛存在于动物皮肤及皮肤附属物中，如毛发、蹄、壳、爪、角、鳞片等。近几年的大量研究表明，角蛋白是一种生物相容性好且不被机体免疫排斥的优质生物医用材料，具有广阔的应用前景。最为突出的是，经过对羊毛等来源的角蛋白进行氨基酸序列测定发现，其含有同Arg-Gly-Asp(RGD)三肽序列。此三肽序列被公认为是细胞外基质中实现细胞结合的有效结合位点，有促进细胞吸附的功能。因此，国内外已开展大量关于角蛋白基生物材料的基础研究及动物实验研究，并在创伤敷料[Wound Repair and Regeneration,2012,20:236-242]、人造骨[Journal of Bioactive and Compatible Polymers,2013,28:141-153]以及神经修复[Biomaterials34(2013)5907-5914]等方面都取得了良好效果，已有部分产品应用于临床。Keratin is an insoluble fibrous animal protein, a structural protein of ectodermal cells, widely present in animal skin and skin appendages, such as hair, hooves, shells, claws, horns, scales, etc. A large number of studies in recent years have shown that keratin is a high-quality biomedical material with good biocompatibility and not rejected by the body's immune system, and has broad application prospects. Most prominently, the amino acid sequence of keratin derived from wool and other sources was found to contain the same Arg-Gly-Asp (RGD) tripeptide sequence. This tripeptide sequence is recognized as an effective binding site for cell binding in the extracellular matrix, and has the function of promoting cell adsorption. Therefore, a large number of basic research and animal experiment research on keratin-based biomaterials have been carried out at home and abroad, and have been widely used in wound dressings [Wound Repair and Regeneration, 2012, 20: 236-242], artificial bone [Journal of Bioactive and Compatible Polymers, 2013,28:141-153] and nerve repair [Biomaterials34 (2013) 5907-5914] have achieved good results, and some products have been used clinically.
但现有研究证明，由于角蛋白的分子量较低，单一的角蛋白材料制成的膜材料通常较脆，且力学强度不高，使其应用性受到限制。因此，目前，大多数的角蛋白基生物材料往往采用角蛋白与天然高分子或人工高分子复合，这样既可以保持以改善单一角蛋白力学性能较差的不足，如丝素蛋白[Biomacromolecules,2008,9,1299–1305]、PVA[Advances inMaterials Science and Engineering,2014,Article ID163678]、PLGA[Journal of Bioactive andCompatible Polymers,2013,28:141-153]、PLLA[Biomed.Mater.2013,8:1-9]等。医用级聚氨酯是一种商业化的合成高分子材料，具有柔韧性好、力学强度高、透气防水等优点，目前已被广泛应用于伤口护理、心脏科、整形外科、血管科等领域。然而，聚氨酯材料本身无生物活性，缺少细胞识别信号位点，不利于内皮细胞等组织细胞的粘附和生长。However, existing studies have shown that due to the low molecular weight of keratin, the membrane material made of a single keratin material is usually brittle and has low mechanical strength, which limits its application. Therefore, at present, most keratin-based biomaterials often use keratin and natural polymers or artificial polymers, which can maintain and improve the poor mechanical properties of single keratin, such as silk fibroin [Biomacromolecules, 2008 ,9,1299–1305], PVA[Advances in Materials Science and Engineering,2014,Article ID163678], PLGA[Journal of Bioactive and Compatible Polymers,2013,28:141-153], PLLA[Biomed.Mater.2013,8:1 -9] etc. Medical grade polyurethane is a commercial synthetic polymer material with good flexibility, high mechanical strength, breathable and waterproof, etc. It has been widely used in wound care, cardiology, plastic surgery, vascular and other fields. However, the polyurethane material itself has no biological activity and lacks cell recognition signal sites, which is not conducive to the adhesion and growth of tissue cells such as endothelial cells.
静电纺丝技术是指利用高压电场环境使聚合物纺丝液形成带电的喷射流，该喷射流在电场作用下被拉长，溶剂挥发，最后在接收装置上形成一定形态的纳米纤维。近十几年来，该技术已成为制备纳米纤维材料的有效途径之一。Electrospinning technology refers to the use of a high-voltage electric field environment to form a charged jet flow from the polymer spinning solution. The jet flow is elongated under the action of the electric field, the solvent is volatilized, and finally nanofibers of a certain shape are formed on the receiving device. In the past ten years, this technology has become one of the effective ways to prepare nanofiber materials.
发明内容Contents of the invention
本发明所要解决的技术问题是提供一种聚氨酯-角蛋白复合纳米纤维管状材料的制备方法，本发明的优势在于通过静电纺丝的方法，将天然生物大分子材料角蛋白与聚氨酯复合制成管状纳米纤维材料，使该材料在具备高孔隙率的同时，显示出良好的力学及生物学功能。The technical problem to be solved by the present invention is to provide a method for preparing a polyurethane-keratin composite nanofiber tubular material. The advantage of the present invention is that the natural biological macromolecular material keratin and polyurethane are composited into a tubular shape by electrospinning. The nanofibrous material enables the material to exhibit good mechanical and biological functions while possessing high porosity.
本发明的一种聚氨酯-角蛋白复合纳米纤维管状材料的制备方法，包括：A kind of preparation method of polyurethane-keratin composite nanofiber tubular material of the present invention comprises:
(1)将聚氨酯溶于溶剂中，搅拌溶解，得到聚氨酯溶液；(1) Polyurethane is dissolved in a solvent, stirred and dissolved to obtain a polyurethane solution;
(2)将角蛋白溶于溶剂中，搅拌溶解，得到角蛋白溶液；(2) dissolving keratin in a solvent, stirring and dissolving to obtain a keratin solution;
(3)将聚氨酯溶液和角蛋白溶液按质量比为95:5-5:95混合，搅拌混匀，得到聚氨酯/角蛋白纺丝液，然后进行静电纺丝，并以圆柱形收集辊为接收器，得到聚氨酯-角蛋白复合纳米纤维管状材料。(3) Mix the polyurethane solution and the keratin solution at a mass ratio of 95:5-5:95, stir and mix to obtain a polyurethane/keratin spinning solution, and then perform electrospinning, and use a cylindrical collection roller as a receiving device to obtain polyurethane-keratin composite nanofiber tubular material.
所述步骤(1)中聚氨酯为Tecothane TM、carbomers、Techphilic TM、Corthane TM、80A、90A、Chronoflex AR、Chronoflex CL、Elast Eon、Cardiomat、Avcothane中的一种或几种。In described step (1), polyurethane is Tecothane™ , carbomers, TechphilicTM , CorthaneTM , 80A, 90A, Chronoflex AR, Chronoflex CL, One or more of Elast Eon, Cardiomat, and Avcothane.
所述步骤(1)中聚氨酯溶液的浓度为5-20mg/ml。The concentration of the polyurethane solution in the step (1) is 5-20 mg/ml.
所述步骤(2)中角蛋白的分子量为3-300kDa。The molecular weight of keratin in the step (2) is 3-300kDa.
所述的角蛋白可以是采用目前已公开报道的各种方法提取制备的角蛋白，包括还原法，氧化法以及水解法等，也可以是上述角蛋白的衍生物，如羧甲基角蛋白。The keratin can be keratin extracted and prepared by various methods reported so far, including reduction method, oxidation method and hydrolysis method, etc., and can also be a derivative of the above-mentioned keratin, such as carboxymethyl keratin.
所述的角蛋白可以从人发、羊毛、家禽羽毛、牛毛等人或动物毛发提取而得，也可以是来源于其他已报道的动物体。The keratin can be extracted from human or animal hairs such as human hair, wool, poultry feathers, cow hair, or other reported animal bodies.
所述步骤(2)中角蛋白溶液的浓度为10-20mg/ml。The concentration of the keratin solution in the step (2) is 10-20 mg/ml.
所述步骤(1)、(2)中溶剂为六氟异丙醇、N，N-二甲基甲酰胺、N，N-二甲基乙酰胺、N-甲基吡咯烷酮、二氯甲烷、二氯乙烷、氯仿、四氢呋喃中的一种或几种。The solvent in the step (1), (2) is hexafluoroisopropanol, N, N-dimethylformamide, N, N-dimethylacetamide, N-methylpyrrolidone, dichloromethane, dichloromethane, One or more of ethyl chloride, chloroform, tetrahydrofuran.
所述溶剂为六氟异丙醇。The solvent is hexafluoroisopropanol.
所述步骤(3)中聚氨酯/角蛋白纺丝液加入药物、无机抗菌剂、有机抗菌剂、抗凝血剂中的一种或几种。In the step (3), one or more of drugs, inorganic antibacterial agents, organic antibacterial agents and anticoagulants are added to the polyurethane/keratin spinning solution.
所述步骤(3)中静电纺丝工艺参数为：电压13-35kV，接收距离8-22cm，纺丝速率为0.3-1.5ml/h，收集辊转动速率60-130rpm，喷丝孔内径为0.7-0.9mm，纺丝温度20-30℃，纺丝湿度45-65％。The electrospinning process parameters in the step (3) are: voltage 13-35kV, receiving distance 8-22cm, spinning rate 0.3-1.5ml/h, collection roller rotation rate 60-130rpm, spinneret hole inner diameter 0.7 -0.9mm, spinning temperature 20-30℃, spinning humidity 45-65%.
所述步骤(3)中圆柱形收集辊直径尺寸可以是0.6-2厘米，或依据所需管状材料的内径尺寸需求进行调节；收集辊的轴垂直于纺丝喷嘴开口方向，并做匀速转动。In the step (3), the diameter of the cylindrical collection roller can be 0.6-2 cm, or adjusted according to the inner diameter of the required tubular material; the axis of the collection roller is perpendicular to the opening direction of the spinning nozzle and rotates at a constant speed.
有益效果Beneficial effect
(1)本发明提出将角蛋白与聚氨酯复合，并采用静电纺丝的方法制备纳米纤维型管状材料，以期改善单一角蛋白纤维材料力学性能较低的不足；(1) The present invention proposes to compound keratin and polyurethane, and adopts the method for electrospinning to prepare nanofiber tubular material, in order to improve the deficiency that the mechanical property of single keratin fiber material is lower;
(2)本发明得到的管状材料即保持了角蛋白良好的生物活性，又具备聚氨酯良好的韧性和力学强度，同时具有内径尺寸可调、制备方法简单等优点，有望用于人工血管等领域。(2) The tubular material obtained by the present invention not only maintains good biological activity of keratin, but also has good toughness and mechanical strength of polyurethane, and has the advantages of adjustable inner diameter and simple preparation method, and is expected to be used in artificial blood vessels and other fields.
具体实施方式Detailed ways
下面结合具体实施例，进一步阐述本发明。应理解，这些实施例仅用于说明本发明而不用于限制本发明的范围。此外应理解，在阅读了本发明讲授的内容之后，本领域技术人员可以对本发明作各种改动或修改，这些等价形式同样落于本申请所附权利要求书所限定的范围。Below in conjunction with specific embodiment, further illustrate the present invention. It should be understood that these examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention. In addition, it should be understood that after reading the teachings of the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms also fall within the scope defined by the appended claims of the present application.
实施例1Example 1
以羊毛为原料采用还原法提取角蛋白，具体方法为：称取5g羊毛，以石油醚为溶剂采用索氏抽提法去除羊毛表面油脂，然后用乙醇清洗羊毛，风干。将上述羊毛加入100ml、1mol/l的巯基乙醇中，用NaOH调节pH约10.0，然后在40℃条件反应12h。反应结束后，将反应液收集(记为滤液1)，同时将反应剩余的羊毛固体过滤收集，继续用100ml、0.1mol/l的Tris碱液在40℃条件下处理2h，然后过滤除去固体，收集滤液(记为滤液2)。.将滤液1和2合并，于6000rpm离心5分钟，然后用截留分子量40，000的透析袋透析36h，最后将透析液加入液氮进行速冻处理，然后置入冷冻干燥机干燥得到干态的角蛋白粉末。The keratin was extracted from wool by reduction method. The specific method was: weigh 5g of wool, use petroleum ether as solvent to remove the oil on the surface of wool by Soxhlet extraction, then wash the wool with ethanol and air-dry it. Add the above-mentioned wool into 100ml, 1mol/l mercaptoethanol, adjust the pH to about 10.0 with NaOH, and then react at 40°C for 12h. After the reaction was finished, the reaction solution was collected (referred to as filtrate 1), and the wool solid remaining in the reaction was collected by filtration, and continued to be treated with 100ml, 0.1mol/l Tris lye at 40°C for 2h, and then filtered to remove the solid. The filtrate was collected (designated as filtrate 2). .The filtrates 1 and 2 were combined, centrifuged at 6000rpm for 5 minutes, and then dialyzed with a dialysis bag with a molecular weight cut-off of 40,000 for 36 hours. Finally, the dialysate was added to liquid nitrogen for quick freezing, and then dried in a freeze dryer to obtain dry horn protein powder.
聚氨酯-角蛋白复合纳米纤维膜的制备：将质量为0.15g的聚氨酯溶于10ml六氟异丙醇，室温下进行磁力搅拌至完全溶解，得到浓度为15mg/ml的聚氨酯溶液。将0.2g角蛋白固体溶于10ml六氟异丙醇，室温下进行磁力搅拌至完全溶解，得到浓度为20mg/ml的角蛋白溶液。将上述聚氨酯与角蛋白溶液按照质量比50：50的比例进行混合，磁力搅拌至混合均匀。将上述所得聚氨酯/角蛋白溶液进行静电纺丝，静电纺丝参数为电压15kV，接收距离12cm，纺丝速率为0.5ml/h，喷丝孔内径为0.7mm，纺丝温度25℃，纺丝湿度50％。以直径1.0cm的圆柱形收集辊为接收器接收喷射产生的纳米丝，收集辊的转动速率为100rpm，经过4小时的收集，得到聚氨酯-角蛋白复合纳米纤维管状材料。Preparation of polyurethane-keratin composite nanofiber membrane: 0.15 g of polyurethane was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a polyurethane solution with a concentration of 15 mg/ml. 0.2 g of solid keratin was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a keratin solution with a concentration of 20 mg/ml. Mix the above polyurethane and keratin solution according to the mass ratio of 50:50, and magnetically stir until the mixture is uniform. The polyurethane/keratin solution obtained above is subjected to electrospinning, the electrospinning parameters are voltage 15kV, receiving distance 12cm, spinning rate 0.5ml/h, spinneret inner diameter 0.7mm, spinning temperature 25°C, spinning Humidity 50%. A cylindrical collection roller with a diameter of 1.0 cm was used as a receiver to receive the nanofilaments generated by the jet, and the rotation speed of the collection roller was 100 rpm. After collecting for 4 hours, a polyurethane-keratin composite nanofiber tubular material was obtained.
实施例2Example 2
以人发为原料采用还原法提取角蛋白，具体方法为：称取5g人发，以石油醚为溶剂采用索氏抽提法去除人发表面油脂，然后用乙醇清洗人发，风干。将上述人发加入200ml水与5g焦亚硫酸钠混合的溶液中，用NaOH调节pH约10.0，然后在60℃条件反应5h。反应结束后，将反应液收集(记为滤液1)，同时将反应剩余的人发固体过滤收集，继续用100ml、0.1mol/l的Tris碱液在40℃条件下处理2h，然后过滤除去固体，收集滤液(记为滤液2)。.将滤液1和2合并，于6000rpm离心5分钟，然后用截留分子量8，000的透析袋透析36h，最后将透析液加入液氮进行速冻处理，然后置入冷冻干燥机干燥得到干态的角蛋白粉末。Using human hair as raw material to extract keratin by reduction method, the specific method is: weigh 5g of human hair, use petroleum ether as solvent to remove human hair surface oil by Soxhlet extraction, then wash human hair with ethanol and air-dry. Add the above-mentioned human hair into a mixed solution of 200ml of water and 5g of sodium metabisulfite, adjust the pH to about 10.0 with NaOH, and then react at 60°C for 5h. After the reaction, collect the reaction solution (referred to as filtrate 1), and collect the remaining human hair solid by filtration at the same time, continue to treat it with 100ml, 0.1mol/l Tris lye at 40°C for 2h, and then filter to remove the solid , collect the filtrate (referred to as filtrate 2). .The filtrates 1 and 2 were combined, centrifuged at 6000rpm for 5 minutes, and then dialyzed with a dialysis bag with a molecular weight cut-off of 8,000 for 36 hours. Finally, the dialysate was added to liquid nitrogen for quick freezing, and then dried in a freeze dryer to obtain dry horn protein powder.
聚氨酯-角蛋白复合纳米纤维膜的制备：将质量为0.2g的聚氨酯溶于10ml六氟异丙醇中，室温下进行磁力搅拌至完全溶解，得到浓度为20mg/ml的聚氨酯溶液。将0.15g角蛋白固体溶于六氟异丙醇中，室温下进行磁力搅拌至完全溶解，得到浓度为15mg/ml的角蛋白溶液。将上述聚氨酯与角蛋白溶液按照质量比20：80的比例进行混合，磁力搅拌至混合均匀。将上述所得聚氨酯/角蛋白溶液进行静电纺丝，静电纺丝参数为电压13kV，接收距离10cm，纺丝速率为0.8ml/h，喷丝孔内径为0.7mm，纺丝温度30℃，纺丝湿度65％。以直径0.6cm的圆柱形收集辊为接收器接收喷射产生的纳米丝，收集辊的转动速率为60rpm，经过4小时的收集，得到聚氨酯-角蛋白复合纳米纤维管状材料。Preparation of polyurethane-keratin composite nanofiber membrane: 0.2 g of polyurethane was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a polyurethane solution with a concentration of 20 mg/ml. 0.15 g of solid keratin was dissolved in hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a keratin solution with a concentration of 15 mg/ml. Mix the above-mentioned polyurethane and keratin solution according to the mass ratio of 20:80, and magnetically stir until the mixture is uniform. The polyurethane/keratin solution obtained above is subjected to electrospinning, the electrospinning parameters are voltage 13kV, receiving distance 10cm, spinning rate 0.8ml/h, spinneret inner diameter 0.7mm, spinning temperature 30°C, spinning Humidity 65%. A cylindrical collection roller with a diameter of 0.6 cm was used as a receiver to receive the nanofilaments generated by the jet. The rotation speed of the collection roller was 60 rpm. After 4 hours of collection, a polyurethane-keratin composite nanofiber tubular material was obtained.
实施例3Example 3
以羊毛为原料采用氧化法提取角蛋白，具体方法为：称取5g羊毛，以石油醚为溶剂采用索氏抽提法去除羊毛表面油脂，然后用乙醇清洗羊毛，风干。向上述羊毛中加入100ml去离子水和5ml、30％的双氧水，然后在100℃条件反应2h。反应结束后，将反应液收集(记为滤液1)，同时将反应剩余的羊毛固体过滤收集，继续用100ml、0.1mol/l的Tris碱液在40℃条件下处理2h，然后过滤除去固体，收集滤液(记为滤液2)。.将滤液1和2合并，于6000rpm离心5分钟，然后用截留分子量8，000的透析袋透析36h，最后将透析液加入液氮进行速冻处理，然后置入冷冻干燥机干燥得到干态的角蛋白粉末。The keratin is extracted from wool by oxidation method. The specific method is: weigh 5g of wool, use petroleum ether as solvent to remove the oil on the surface of wool by Soxhlet extraction, then wash the wool with ethanol and air-dry it. 100ml of deionized water and 5ml of 30% hydrogen peroxide were added to the wool, and then reacted at 100°C for 2h. After the reaction was finished, the reaction solution was collected (referred to as filtrate 1), and the wool solid remaining in the reaction was collected by filtration, and continued to be treated with 100ml, 0.1mol/l Tris lye at 40°C for 2h, and then filtered to remove the solid. The filtrate was collected (designated as filtrate 2). .The filtrates 1 and 2 were combined, centrifuged at 6000rpm for 5 minutes, and then dialyzed with a dialysis bag with a molecular weight cut-off of 8,000 for 36 hours. Finally, the dialysate was added to liquid nitrogen for quick freezing, and then dried in a freeze dryer to obtain dry horn protein powder.
聚氨酯-角蛋白复合纳米纤维膜的制备：将质量为0.05g的聚氨酯溶于10ml六氟异丙醇中，室温下进行磁力搅拌至完全溶解，得到浓度为5mg/ml的聚氨酯溶液。将0.1g角蛋白固体溶于10ml六氟异丙醇中，室温下进行磁力搅拌至完全溶解，得到浓度为10mg/ml的角蛋白溶液。将上述聚氨酯与角蛋白溶液按照质量比95：5的比例进行混合，磁力搅拌至混合均匀。将上述所得聚氨酯/角蛋白溶液进行静电纺丝，静电纺丝参数为电压13kV，接收距离8cm，纺丝速率为1.5ml/h，喷丝孔内径为0.9mm，纺丝温度20℃，纺丝湿度45％。以直径1.5cm的圆柱形收集辊为接收器接收喷射产生的纳米丝，收集辊的转动速率为110rpm，经过3小时的收集，得到聚氨酯-角蛋白复合纳米纤维管状材料。Preparation of polyurethane-keratin composite nanofiber membrane: 0.05 g of polyurethane was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a polyurethane solution with a concentration of 5 mg/ml. 0.1 g of solid keratin was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a keratin solution with a concentration of 10 mg/ml. Mix the above-mentioned polyurethane and keratin solution according to the mass ratio of 95:5, and magnetically stir until the mixture is uniform. The polyurethane/keratin solution obtained above is subjected to electrospinning, the electrospinning parameters are voltage 13kV, receiving distance 8cm, spinning rate 1.5ml/h, spinneret inner diameter 0.9mm, spinning temperature 20°C, spinning Humidity 45%. A cylindrical collection roller with a diameter of 1.5 cm was used as a receiver to receive the nanofilaments generated by the jet. The rotation speed of the collection roller was 110 rpm. After 3 hours of collection, the polyurethane-keratin composite nanofiber tubular material was obtained.
实施例4Example 4
以羊毛为原料采用氧化法提取角蛋白，具体方法为：称取5g羊毛，以石油醚为溶剂采用索氏抽提法去除羊毛表面油脂，然后用乙醇清洗羊毛，风干。向上述羊毛中加入100ml、4％的过氧乙酸，然后在60℃条件反应5h。反应结束后，将反应液收集(记为滤液1)，同时将反应剩余的羊毛固体过滤收集，继续用100ml、0.1mol/l的Tris碱液在40℃条件下处理2h，然后过滤除去固体，收集滤液(记为滤液2)。.将滤液1和2合并，于6000rpm离心5分钟，然后用截留分子量8，000的透析袋透析36h，最后将透析液加入液氮进行速冻处理，然后置入冷冻干燥机干燥得到干态的角蛋白粉末。The keratin is extracted from wool by oxidation method. The specific method is: weigh 5g of wool, use petroleum ether as solvent to remove the oil on the surface of wool by Soxhlet extraction, then wash the wool with ethanol and air-dry it. Add 100 ml of 4% peroxyacetic acid to the above wool, and then react at 60° C. for 5 h. After the reaction was finished, the reaction solution was collected (referred to as filtrate 1), and the wool solid remaining in the reaction was collected by filtration, and continued to be treated with 100ml, 0.1mol/l Tris lye at 40°C for 2h, and then filtered to remove the solid. The filtrate was collected (designated as filtrate 2). .The filtrates 1 and 2 were combined, centrifuged at 6000rpm for 5 minutes, and then dialyzed with a dialysis bag with a molecular weight cut-off of 8,000 for 36 hours. Finally, the dialysate was added to liquid nitrogen for quick freezing, and then dried in a freeze dryer to obtain dry horn protein powder.
聚氨酯-角蛋白复合纳米纤维膜的制备：将质量为0.15g的聚氨酯溶于10ml六氟异丙醇中，室温下进行磁力搅拌至完全溶解，得到浓度为15mg/ml的聚氨酯溶液。将0.2g角蛋白固体溶于10ml六氟异丙醇中，室温下进行磁力搅拌至完全溶解，得到浓度为20mg/ml的角蛋白溶液。将上述聚氨酯与角蛋白溶液按照质量比5：95的比例进行混合，磁力搅拌至混合均匀。将上述所得聚氨酯/角蛋白溶液进行静电纺丝，静电纺丝参数为电压35kV，接收距离22cm，纺丝速率为1.5ml/h，喷丝孔内径为0.9mm，纺丝温度20℃，纺丝湿度55％。以直径1.0cm的圆柱形收集辊为接收器接收喷射产生的纳米丝，收集辊的转动速率为130rpm，经过3小时的收集，得到聚氨酯-角蛋白复合纳米纤维管状材料。Preparation of polyurethane-keratin composite nanofiber membrane: 0.15 g of polyurethane was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a polyurethane solution with a concentration of 15 mg/ml. 0.2 g of solid keratin was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a keratin solution with a concentration of 20 mg/ml. Mix the above-mentioned polyurethane and keratin solution according to the mass ratio of 5:95, and magnetically stir until the mixture is uniform. The polyurethane/keratin solution obtained above is subjected to electrospinning, the electrospinning parameters are voltage 35kV, receiving distance 22cm, spinning rate 1.5ml/h, spinneret inner diameter 0.9mm, spinning temperature 20°C, spinning Humidity 55%. A cylindrical collection roller with a diameter of 1.0 cm was used as a receiver to receive the nanofilaments produced by the jet. The rotation speed of the collection roller was 130 rpm. After 3 hours of collection, the polyurethane-keratin composite nanofiber tubular material was obtained.
实施例5Example 5
将以羊毛为原料采用还原法提取角蛋白，并对角蛋白进行衍生化改性。具体方法为：称取5g羊毛，以石油醚为溶剂采用索氏抽提法去除羊毛表面油脂，然后用乙醇清洗羊毛，风干。将上述羊毛加入100ml、1mol/l的巯基乙醇中，用NaOH调节pH约10.0，然后在40℃条件反应12h。然后，用醋酸将上述反应液pH调整到7.5，向其中加入100ml、0.33mol/l的NaBrO3溶液，在室温条件下继续搅拌24h。反应结束后，过滤除去固体物，将滤液收集并用截留分子量8，000的透析袋透析36h，最后将透析液加入液氮进行速冻处理，然后置入冷冻干燥机干燥得到干态的羧甲基角蛋白粉末。The wool is used as raw material to extract keratin by reduction method, and the keratin is derivatized and modified. The specific method is as follows: take 5 g of wool, use petroleum ether as a solvent to remove the oil on the surface of the wool by Soxhlet extraction, then wash the wool with ethanol and air-dry it. Add the above-mentioned wool into 100ml, 1mol/l mercaptoethanol, adjust the pH to about 10.0 with NaOH, and then react at 40°C for 12h. Then, the pH of the above reaction solution was adjusted to 7.5 with acetic acid, 100 ml of 0.33 mol/l NaBrO3 solution was added thereto, and stirring was continued for 24 h at room temperature. After the reaction, remove the solid matter by filtration, collect the filtrate and dialyze with a dialysis bag with a molecular weight cut-off of 8,000 for 36 hours, and finally add the dialysate to liquid nitrogen for quick-freezing treatment, and then put it in a freeze dryer to dry to obtain dry carboxymethyl angle protein powder.
聚氨酯-角蛋白复合纳米纤维膜的制备：将质量为0.15g的聚氨酯溶于10ml六氟异丙醇中，室温下进行磁力搅拌至完全溶解，得到浓度为15mg/ml的聚氨酯溶液。将0.1g羧甲基角蛋白固体溶于10ml六氟异丙醇中，室温下进行磁力搅拌至完全溶解，得到浓度为10mg/ml的羧甲基角蛋白溶液。将上述聚氨酯与角蛋白溶液按照质量比5：95的比例进行混合，磁力搅拌至混合均匀。将上述所得聚氨酯/角蛋白溶液进行静电纺丝，静电纺丝参数为电压15kV，接收距离12cm，纺丝速率为1.0ml/h，喷丝孔内径为0.7mm，纺丝温度15℃，纺丝湿度55％。以直径1.0cm的圆柱形收集辊为接收器接收喷射产生的纳米丝，收集辊的转动速率为100rpm，经过4小时的收集，得到聚氨酯-角蛋白复合纳米纤维管状材料。Preparation of polyurethane-keratin composite nanofiber membrane: 0.15 g of polyurethane was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a polyurethane solution with a concentration of 15 mg/ml. 0.1 g of solid carboxymethyl keratin was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a carboxymethyl keratin solution with a concentration of 10 mg/ml. Mix the above-mentioned polyurethane and keratin solution according to the mass ratio of 5:95, and magnetically stir until the mixture is uniform. The polyurethane/keratin solution obtained above is subjected to electrospinning, the electrospinning parameters are voltage 15kV, receiving distance 12cm, spinning rate 1.0ml/h, spinneret inner diameter 0.7mm, spinning temperature 15°C, spinning Humidity 55%. A cylindrical collection roller with a diameter of 1.0 cm was used as a receiver to receive the nanofilaments generated by the jet, and the rotation speed of the collection roller was 100 rpm. After collecting for 4 hours, a polyurethane-keratin composite nanofiber tubular material was obtained.
实施例6Example 6
以人发为原料采用还原法提取角蛋白，并对角蛋白进行衍生化改性。具体方法为：称取5g人发，以石油醚为溶剂采用索氏抽提法去除人发表面油脂，然后用乙醇清洗人发，风干。将上述人发加入200ml水与5g焦亚硫酸钠混合的溶液中，用NaOH调节pH约10.0，然后在60℃条件反应5h。然后，用醋酸将上述反应液pH调整到8.5，向其中加入8.0g碘乙酸，在室温条件下继续搅拌6h。反应结束后，过滤除去固体物，将滤液收集并用截留分子量8，000的透析袋透析36h，最后将透析液加入液氮进行速冻处理，然后置入冷冻干燥机干燥得到干态的羧甲基角蛋白粉末。Human hair is used as raw material to extract keratin by reduction method, and the keratin is derivatized and modified. The specific method is as follows: 5g of human hair is weighed, oil on the surface of the human hair is removed by Soxhlet extraction with petroleum ether, then the human hair is washed with ethanol and air-dried. Add the above-mentioned human hair into a mixed solution of 200ml of water and 5g of sodium metabisulfite, adjust the pH to about 10.0 with NaOH, and then react at 60°C for 5h. Then, the pH of the above reaction solution was adjusted to 8.5 with acetic acid, 8.0 g of iodoacetic acid was added thereto, and stirring was continued for 6 h at room temperature. After the reaction, remove the solid matter by filtration, collect the filtrate and dialyze with a dialysis bag with a molecular weight cut-off of 8,000 for 36 hours, and finally add the dialysate to liquid nitrogen for quick-freezing treatment, and then put it in a freeze dryer to dry to obtain dry carboxymethyl angle protein powder.
聚氨酯-角蛋白复合纳米纤维膜的制备：将质量为0.15g的聚氨酯溶于10ml六氟异丙醇，室温下进行磁力搅拌至完全溶解，得到浓度为15mg/ml的聚氨酯溶液。将0.2g羧甲基角蛋白固体溶于10ml六氟异丙醇，室温下进行磁力搅拌至完全溶解，得到浓度为20mg/ml的羧甲基角蛋白溶液。将上述聚氨酯与角蛋白溶液按照质量比50：50的比例进行混合，磁力搅拌至混合均匀。将上述所得聚氨酯/角蛋白溶液进行静电纺丝，静电纺丝参数为电压15kV，接收距离12cm，纺丝速率为1.2ml/h，喷丝孔内径为0.7mm，纺丝温度15℃，纺丝湿度55％。以直径1.0cm的圆柱形收集辊为接收器接收喷射产生的纳米丝，收集辊的转动速率为100rpm，经过4小时的收集，得到聚氨酯-角蛋白复合纳米纤维管状材料。Preparation of polyurethane-keratin composite nanofiber membrane: 0.15 g of polyurethane was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a polyurethane solution with a concentration of 15 mg/ml. 0.2 g of carboxymethyl keratin solid was dissolved in 10 ml of hexafluoroisopropanol, and magnetically stirred at room temperature until completely dissolved to obtain a carboxymethyl keratin solution with a concentration of 20 mg/ml. Mix the above polyurethane and keratin solution according to the mass ratio of 50:50, and magnetically stir until the mixture is uniform. The polyurethane/keratin solution obtained above is subjected to electrospinning, the electrospinning parameters are voltage 15kV, receiving distance 12cm, spinning rate 1.2ml/h, spinneret inner diameter 0.7mm, spinning temperature 15°C, spinning Humidity 55%. A cylindrical collection roller with a diameter of 1.0 cm was used as a receiver to receive the nanofilaments generated by the jet, and the rotation speed of the collection roller was 100 rpm. After collecting for 4 hours, a polyurethane-keratin composite nanofiber tubular material was obtained.
实施例7Example 7
按照实施例1中所述的制备方法，以Carbothane TMPC-3585A聚氨酯材料与羊毛角蛋白混合制备的聚氨酯/角蛋白复合纳米纤维材料，经分析测定，复合纳米纤维材料在干燥状态下的断裂强度为55MPa，断裂伸长率为225％。以血管内皮细胞为细胞模型，采用扫描电子显微镜观察细胞在材料上的黏附形态，并通过MTT法对细胞的增殖行为进行评价，结果表明，内皮细胞在聚氨酯/角蛋白复合纳米纤维材料上表现出良好的黏附形态和增殖行为，与对照组盖玻片具有显著性差异。按照实施例1中所述的制备方法，以单一的角蛋白制备的膜材料在干燥状态下脆性强、易破碎。按照实施例1中所述的制备方法，以单一的Carbothane TMPC-3585A聚氨酯制备的纳米纤维材料经过细胞评价表明，内皮细胞在单一聚氨酯膜材料上表现出与对照组盖玻片相近的增殖行为，无显著性差异。According to the preparation method described in Example 1, the polyurethane/keratin composite nanofiber material prepared by mixing Carbothane™ PC-3585A polyurethane material with wool keratin, after analysis and determination, the breaking strength of the composite nanofiber material in dry state It is 55MPa, and the elongation at break is 225%. Taking vascular endothelial cells as cell models, the adhesion morphology of cells on the material was observed by scanning electron microscopy, and the proliferation behavior of cells was evaluated by MTT method. The results showed that endothelial cells exhibited Good adhesion morphology and proliferation behavior, which are significantly different from the control coverslip. According to the preparation method described in Example 1, the membrane material prepared from a single keratin is highly brittle and easily broken in a dry state. According to the preparation method described in Example 1, the nanofibrous material prepared with a single Carbothane™ PC-3585A polyurethane was evaluated by cells, and the endothelial cells showed similar proliferation behavior to the control cover glass on the single polyurethane membrane material , no significant difference.
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| CN201410264033.7ACN104018247B (en) | 2014-06-13 | 2014-06-13 | The preparation method of a kind of urethane-Keratin sulfate composite nano fiber tubular material |
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| CN201410264033.7ACN104018247B (en) | 2014-06-13 | 2014-06-13 | The preparation method of a kind of urethane-Keratin sulfate composite nano fiber tubular material |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104784758A (en)* | 2015-05-09 | 2015-07-22 | 南京师范大学 | Preparation method of polymer/keratin composite anticoagulation vascular tissue engineering scaffold |
| CN105803676A (en)* | 2016-03-23 | 2016-07-27 | 天津工业大学 | Preparation method of keratin/PEO nanofiber membrane with high keratin content |
| CN107190349A (en)* | 2017-05-10 | 2017-09-22 | 天津中智科技发展有限公司 | A kind of method that utilization yak hair prepares nanofiber |
| CN110528174A (en)* | 2019-09-09 | 2019-12-03 | 仲恺农业工程学院 | Preparation method of dopamine modified keratin composite membrane |
| WO2019238610A1 (en) | 2018-06-12 | 2019-12-19 | Kerline S.R.L. | Keratin nanofibers as delivery vehicles of active ingredients, methods for the production and uses thereof |
| CN110670168A (en)* | 2019-10-06 | 2020-01-10 | 武汉纺织大学 | Highly spinnable wool keratin/silicon dioxide composite fiber and preparation method thereof |
| CN111778635A (en)* | 2020-07-14 | 2020-10-16 | 河南工业大学 | A kind of preparation method of peanut protein-polyurethane nanofiber membrane |
| CN114045574A (en)* | 2021-11-15 | 2022-02-15 | 华峰化学股份有限公司 | Preparation method of skin-friendly polyurethane elastic fiber |
| CN115516024A (en)* | 2020-05-01 | 2022-12-23 | 现代牧场股份有限公司 | Protein polyurethane alloy and layered material comprising same |
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101078134A (en)* | 2007-06-27 | 2007-11-28 | 东华大学 | Preparation of natural material/polymer material coaxial electrostatic spinning nano fibre |
| CN101100766A (en)* | 2007-07-13 | 2008-01-09 | 东华大学 | Preparation of Nanofibrous Carriers Loaded and Slowly Released Drugs and Bioactive Factors |
| CN101130902A (en)* | 2007-08-07 | 2008-02-27 | 东华大学 | Preparation and application of fibers containing heparin and bioactive molecules and their fabrics |
| CN101785875A (en)* | 2010-02-25 | 2010-07-28 | 王深明 | Preparation method of nano superfine fiber vascular prosthesis |
| DE102011105369A1 (en)* | 2011-06-20 | 2012-12-20 | Carl Freudenberg Kg | Stable, aqueous protein solution or protein dispersion, process for the preparation of the stable, aqueous solution or dispersion and process for the production of moldings, fabrics, impregnations or coatings from the stable, aqueous protein solution or protein dispersion and their use |
| CN103276472A (en)* | 2013-06-03 | 2013-09-04 | 郑州大学 | Collagen/polyvinyl alcohol composite microspheres as well as preparation method and application thereof |
| CN103394131A (en)* | 2013-07-26 | 2013-11-20 | 宁夏医科大学 | Novel double-layered composite transmitting tissue regeneration membrane and preparation method thereof |
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101078134A (en)* | 2007-06-27 | 2007-11-28 | 东华大学 | Preparation of natural material/polymer material coaxial electrostatic spinning nano fibre |
| CN101100766A (en)* | 2007-07-13 | 2008-01-09 | 东华大学 | Preparation of Nanofibrous Carriers Loaded and Slowly Released Drugs and Bioactive Factors |
| CN101130902A (en)* | 2007-08-07 | 2008-02-27 | 东华大学 | Preparation and application of fibers containing heparin and bioactive molecules and their fabrics |
| CN101785875A (en)* | 2010-02-25 | 2010-07-28 | 王深明 | Preparation method of nano superfine fiber vascular prosthesis |
| DE102011105369A1 (en)* | 2011-06-20 | 2012-12-20 | Carl Freudenberg Kg | Stable, aqueous protein solution or protein dispersion, process for the preparation of the stable, aqueous solution or dispersion and process for the production of moldings, fabrics, impregnations or coatings from the stable, aqueous protein solution or protein dispersion and their use |
| CN103276472A (en)* | 2013-06-03 | 2013-09-04 | 郑州大学 | Collagen/polyvinyl alcohol composite microspheres as well as preparation method and application thereof |
| CN103394131A (en)* | 2013-07-26 | 2013-11-20 | 宁夏医科大学 | Novel double-layered composite transmitting tissue regeneration membrane and preparation method thereof |
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104784758A (en)* | 2015-05-09 | 2015-07-22 | 南京师范大学 | Preparation method of polymer/keratin composite anticoagulation vascular tissue engineering scaffold |
| CN104784758B (en)* | 2015-05-09 | 2017-05-10 | 南京师范大学 | Preparation method of polymer/keratin composite anticoagulation vascular tissue engineering scaffold |
| CN105803676A (en)* | 2016-03-23 | 2016-07-27 | 天津工业大学 | Preparation method of keratin/PEO nanofiber membrane with high keratin content |
| CN107190349B (en)* | 2017-05-10 | 2020-01-07 | 天津中智科技发展有限公司 | Method for preparing nanofiber by using yak hair |
| CN107190349A (en)* | 2017-05-10 | 2017-09-22 | 天津中智科技发展有限公司 | A kind of method that utilization yak hair prepares nanofiber |
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| Publication number | Publication date |
|---|---|
| CN104018247B (en) | 2016-06-08 |
| Publication | Publication Date | Title |
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| CF01 | Termination of patent right due to non-payment of annual fee | Granted publication date:20160608 Termination date:20190613 | |
| CF01 | Termination of patent right due to non-payment of annual fee |