WO2017206478A1 - 一种凝血酶缓释医疗器械及制备方法 - Google Patents

一种凝血酶缓释医疗器械及制备方法 Download PDF

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WO2017206478A1
WO2017206478A1 PCT/CN2016/109991 CN2016109991W WO2017206478A1 WO 2017206478 A1 WO2017206478 A1 WO 2017206478A1 CN 2016109991 W CN2016109991 W CN 2016109991W WO 2017206478 A1 WO2017206478 A1 WO 2017206478A1
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thrombin
medical device
preparation
drying
minutes
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高峰
高奇辉
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高峰
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L31/00—Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
    • A61L31/08—Materials for coatings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L31/00—Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
    • A61L31/08—Materials for coatings
    • A61L31/082—Inorganic materials
    • A61L31/086—Phosphorus-containing materials, e.g. apatite
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L31/00—Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
    • A61L31/08—Materials for coatings
    • A61L31/10—Macromolecular materials
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L31/00—Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
    • A61L31/14—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A61L31/16—Biologically active materials, e.g. therapeutic substances
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/20—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials
    • A61L2300/252—Polypeptides, proteins, e.g. glycoproteins, lipoproteins, cytokines
    • A61L2300/254—Enzymes, proenzymes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/40—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
    • A61L2300/418—Agents promoting blood coagulation, blood-clotting agents, embolising agents

Definitions

  • the invention relates to the field of medical instruments and preparation thereof, in particular to a thrombin sustained release medical device and a preparation method thereof.
  • Titanium clip is one of the important instruments for endoscopic treatment. At present, it only takes physical closure treatment for local injury. Many scholars have attempted to target sustained-release thrombin on medical metal substrates. Currently, all sustained-release studies are based on the surface of bio-inert materials, which not only have poor biological activity, but also cause toxicity of heavy metal ions to cause thrombin failure. Hydroxyapatite (abbreviated as HA or HAP, with the formula Ca 10 (OH) 2 (PO 4 ) 6 ) is the main (inorganic) component of teeth and bones, and is also the best component of artificial bone in cutting-edge medicine.
  • HA or HAP Hydroxyapatite
  • Hydroxyapatite is a natural ceramic material that stimulates or induces bone tissue growth and can form a bone bond with bone tissue. Its biocompatibility and biological activity are superior to that of tricalcium phosphate and other phosphorus-calcium ceramic materials.
  • the use of hydroxyapatite contributes to the adhesion, proliferation and function of cells. It is not only an excellent bone tissue engineering carrier material but also an additional material based on bone or tooth replacement or defect repair materials.
  • a carrier material for a medical functional material or drug is not only an excellent bone tissue engineering carrier material but also an additional material based on bone or tooth replacement or defect repair materials.
  • the object of the present invention is to solve some of the problems existing in the prior art thrombin sustained release medical device technology and to provide (the name of the invention).
  • a method for preparing a thrombin sustained release medical device comprises the following steps:
  • the metal plate is ultrasonically washed and dried with deionized water, and then immersed in a hydroxyethylidene diphosphonic acid solution, taken out for secondary drying and then washed;
  • the metal plate is a metal matrix medical device.
  • the drying in the step (1) and the step (2) is drying in a dry box at 50-70 ° C for 5-10 minutes or 20-30 ° C in air for 1-2 hours.
  • the soaking temperature in the step (1) is 20-30 ° C for 1-2 minutes.
  • the concentration of the hydroxyethylidene diphosphonic acid solution in the step (1) is 3-5 mol/l.
  • the concentration of the hydroxyethylidene diphosphonic acid solution in the step (1) is 4-5 mol/l.
  • the secondary drying in the step (1) is dried in a dry box at 50-70 ° C for 10-20 minutes or at 20-30 ° C for 2-4 hours in air.
  • the washing time in the step (1) is 2-3 minutes.
  • the concentration of the hydroxyapatite suspension in the step (2) is 5-50 g/l, the soaking temperature is 20-30 ° C, and the time is 5-10 minutes.
  • the concentration of the hydroxyapatite suspension in the step (2) is 20-30 g/l.
  • the thrombin concentration in the thrombin solution in the step (3) is 4000-8000 U/l, prepared from thrombin lyophilized powder and 0.9% physiological saline.
  • the thrombin concentration in the thrombin solution in the step (3) is 4500-5500 U/l.
  • the soaking temperature in the step (3) is 20-30 ° C for 1-2 hours.
  • a thrombin sustained release medical device is prepared by any of the methods described above.
  • the invention realizes the preparation of a thrombin sustained-release coating on the bioactive surface of a metal-based medical device, and can realize the upgrading from a simple "naked” device to a "functional" drug composite device.
  • the invention can realize targeted sustained release of a drug which can only be applied locally, such as thrombin, and the "functional" drug composite device which can be used for preparation can be directly placed on a human target organ and a surgical target site.
  • the invention can realize the stable and sustained release of drugs such as thrombin on the metal-based coating system, and the medicine can realize sustained release at a designated site to continuously increase the concentration of surrounding tissues, and solve the problem that the locally sprayed drugs are rapidly diluted by the tissue fluid.
  • the problem of efficacy can realize the stable and sustained release of drugs such as thrombin on the metal-based coating system, and the medicine can realize sustained release at a designated site to continuously increase the concentration of surrounding tissues, and solve the problem that the locally sprayed drugs are rapidly diluted by the tissue fluid.
  • Figure 1 is a graphical representation of a product of a thrombin sustained release metal medical device of the present application
  • Figure 2 is a prepared stabilized hydroxyapatite nano/submicron coating
  • Figure 3 is a thrombin layer on the surface of a hydroxyapatite nano/submicron coating
  • Figure 4 is a photoelectron spectroscopy (XPS) curve of a hydroxyapatite layer
  • Figure 5 is a photoelectron spectroscopy (XPS) curve of a thrombin layer
  • Figure 6 is a graph showing the average concentration of thrombin at different times
  • Figure 7 is a bar graph of the dynamic thrombin release concentration at different time periods.
  • SUS304 austenitic stainless steel sample was ultrasonically washed with deionized water for 2 minutes to remove oil or rust on the surface. After the surface is clean, it is placed in a dry box and dried at 70 ° C for 5 minutes. The residual water droplets are acceptable.
  • step 5 Take out the product prepared in step 4 and put it in a dry box and dry at 70 ° C for 10 minutes, after the surface has no residual water drops.
  • the thrombin lyophilized powder and 0.9% physiological saline were prepared to prepare a 5000 U/l thrombin solution, and immersed at 25 ° C for 2 hours.
  • the sample was dried in air at 30 ° C for 2 hours, and characterized by photoelectron spectroscopy (XPS) and atomic force microscopy (AFM).
  • XPS photoelectron spectroscopy
  • AFM atomic force microscopy
  • the results of atomic force microscopy characterization test are shown in Figure 2-3. It can be seen from the AFM comparison chart that the surface changes before and after preparation, and the light color region is thrombin molecule.
  • the photoelectron spectroscopy test results are shown in Figure 4-5. It can be seen from XPS comparison that (1) thrombin molecules are successfully coated on the surface of hydroxyapatite with a certain thickness; (2) decomposed from high resolution XPS In view of this, there is no new chemical bond and production, and the sustained release product is thrombin.
  • the above results indicate that the composition and morphology of the thrombin coating changed significantly, forming a stable coating, the coating is a thrombin layer; no new chemical bond is produced
  • the SUS304 austenitic stainless steel sample was immersed in the surface cleaning solution for 2 minutes to remove the oil on the surface, and then washed twice with deionized water for 2 minutes each time. After the surface to be visually cleaned, it was placed in air at a relative humidity of 55% and 20 ° C for 2 hours, and no significant residual water droplets on the surface were qualified.
  • step 2 The sample of step 1 was taken out and immersed in a 5 mol/l solution of hydroxyethylidene diphosphonic acid at 20 ° C for 5 minutes to prepare a surface chemical exchange film of hydroxyethylidene diphosphonic acid.
  • Step 2 After the sample was taken out, it was placed in air at a relative humidity of 55% and a temperature of 20 ° C for 2 hours, and the prepared film was dried.
  • step 3 After the sample of step 3 has no obvious liquid or colloid, it is placed in a suspension of hydroxyapatite (HA) which is continuously stirred at 20 g/l, and immersed at 25 ° C for 10 minutes. The excess hydroxyethylidene diphosphonic acid crystal residue is dissolved in water while self-assembly of the hydroxyapatite coating on the hydroxyethylidene diphosphonic acid surface chemical exchange film is completed.
  • HA hydroxyapatite
  • step 4 Take out step 4 and place the sample in air at a relative humidity of 55% and 20 ° C for 2 hours, after the surface has no obvious residual water droplets.
  • the thrombin lyophilized powder and 0.9% physiological saline were prepared to prepare a 5000 U/l thrombin solution, and immersed at 25 ° C for 1 hour.
  • step 5 Take out step 5 and dry the sample in air at 30 °C for 2 hours, take 6 samples in each group, measure the absorbance of different concentration standards with the microplate reader at 450nm wavelength, and draw the standard according to different standard concentration and corresponding absorbance. curve. On the basis of this, the average thrombin concentration calculated at different times is shown in Fig. 6. It can be seen from Fig. 6 that thrombin is released rapidly in 0-60 minutes, and thrombin release is very slow from 60-180 minutes. Can maintain a certain concentration.
  • the thrombin-coated metal plates (6 pieces in total) were respectively immersed in a beaker containing 200 ml of physiological saline, and 100 ul of the solution was sampled at 0h, 1h, 2h, 3h, 4h, 5h, and placed in the labeled EP tube. After each sampling, the original physiological saline was discarded, and the absorbance of the six groups of solutions at different times was measured by a microplate reader and the average value was calculated. The absorbance measured by discarding the soaking solution at different times was analyzed by t test. The P values of the two groups were greater than 0.05, and the difference was not statistically significant. There was no significant difference in the absorbance of the solution measured before the solution was discarded every 1 hour. We can assume that the drug-coated metal plate can stably release thrombin in the same period of time. The specific results are shown in Figure 7.
  • thrombin achieves a stable sustained release value at 1 hour, and a stable sustained release of thrombin in a continuously changing solution.
  • the "thrombin-hydroxyapatite-oxide-free layer-metal" steel plate can stably and stably release thrombin in a physiological saline solution, thereby ensuring the concentration of thrombin in the solution and having a sustained release effect of thrombin. That is, the thrombin sustained-release metal matrix medical device thus prepared also has a good thrombin sustained release effect.

Abstract

一种凝血酶缓释医疗器械的制备方法，包括如下步骤：(1)将医疗器械用去离子水超声洗涤干燥后浸泡于羟基乙叉二膦酸溶液中，取出进行二次干燥后洗涤；(2)将步骤(1)处理后的医疗器械浸泡于羟基磷灰石悬浊液中，取出干燥；(3)将步骤(2)处理后的医疗器械浸泡于凝血酶溶液中取出得到凝血酶缓释医疗器械。所述方法可实现凝血酶等仅能局部应用的药物的靶向缓释，可用于制备的"功能性"药物复合器械并直接放置于人体靶器官以及手术靶部位。

Description

一种凝血酶缓释医疗器械及制备方法 技术领域
本发明涉及医疗器械及其制备领域，特别是涉及一种凝血酶缓释医疗器械及其制备方法。
背景技术
目前，随着医疗技术的飞速发展与创新，各种内镜技术、微创技术等诊疗合一的医学手段逐步成为主流医疗技术不断出现，但内镜与微创器械也带来一。上消化道出血是常见的消化道疾病，同时也是内镜下治疗中常见的手术并发症。对于上消化道出血的传统内科治疗手段主要有局部喷洒或注射止血、静脉应用止血药物等，目前内镜下止血广泛应用靶向药物凝血酶局部作用于病灶表面，血液很快形成稳定的凝血块。但靶向喷洒后存在凝血酶被组织液快速稀释的问题，使药效降低。后随着内镜下器械的不断发展与改良，用于局部止血及封闭穿孔的各种吻合夹开始出现。钛夹作为内镜下治疗的重要器械之一，目前只对局部损伤承担物理封闭治疗作用。很多学者尝试在医用金属基材上靶向缓释凝血酶，目前所有的缓释研究都基于生物惰性材料表面，不但生物活性差，而且重金属离子毒性会造成凝血酶失效。羟基磷灰石(hydroxyapatite，简写为HA或HAP，分子式为Ca10(OH)2(PO4)6)是牙齿和骨骼的主要(无机)成分，在前沿医学上也作为人造骨的最佳成分。羟基磷灰石生物兼容性好，是能刺激或者诱导骨组织生长并能与骨组织形成骨性结合的天然陶瓷材料，生物兼容性及生物活性均优于磷酸三钙及其它磷钙陶瓷材料。羟基磷灰石的使用，有助于细胞的粘附、增殖及功能发挥，在直接作为骨、牙等替代或缺损修复材料的基础上，不但是优异的骨组织工程载体材料，也可作为其它医学功能材料或药物的载体材料。
发明内容
本发明的目的是解决现有凝血酶缓释医疗器械技术中存在的部分问题，提供(发明名称)。
本发明的目的是通过以下技术方案实现的：
一种凝血酶缓释医疗器械的制备方法，包括如下步骤：
(1)将金属板用去离子水超声洗涤干燥后浸泡于羟基乙叉二膦酸溶液中，取出进行二次干燥后洗涤；
(2)将步骤(1)处理后的金属板浸泡于羟基磷灰石悬浊液中，取出干燥；
(3)将步骤(2)处理后的金属板浸泡于凝血酶溶液中取出即得；
所述金属板为金属基质的医疗器械。
优选地，所述步骤(1)和步骤(2)中的干燥是在干燥箱中50-70℃干燥5-10分钟或20-30℃空气中干燥1-2小时。
优选地，所述步骤(1)中的浸泡温度为20-30℃，时间为1-2分钟。
优选地，所述步骤(1)中的羟基乙叉二膦酸溶液浓度为3-5mol/l。
优选地，所述步骤(1)中的羟基乙叉二膦酸溶液浓度为4-5mol/l。
优选地，所述步骤(1)中的二次干燥为干燥箱中50-70℃干燥10-20分钟或20-30℃空气中干燥2-4小时。
优选地，所述步骤(1)中的洗涤时间为2-3分钟。
优选地，所述步骤(2)中的羟基磷灰石悬浊液浓度为5-50g/l，浸泡温度为20-30℃，时间为5-10分钟。
优选地，所述步骤(2)中的羟基磷灰石悬浊液浓度为20-30g/l。
优选地，所述步骤(3)中的凝血酶溶液中凝血酶浓度为4000-8000U/l，由凝血酶冻干粉与0.9％生理盐水制备。
优选地，所述步骤(3)中的凝血酶溶液中凝血酶浓度为4500-5500U/l。
优选地，所述步骤(3)中的浸泡温度为20-30℃，时间为1-2小时。
本发明的另一方面，一种凝血酶缓释医疗器械，由上述任一所述方法制备而得。
本发明的有益效果：
1.本发明实现了医疗器械金属基的生物活性表面的凝血酶缓释涂层制备，可实现从单纯“裸”器械到“功能性”药物复合器械的升级。
2.本发明可实现凝血酶等仅能局部应用的药物的靶向缓释，可用于制备的“功能性”药物复合器械可直接放置于人体靶器官以及手术靶部位。
3.本发明可实现凝血酶等药物在金属基涂层体系上的稳定缓释，药物可实现在指定部位缓释从而可持续增加周围组织浓度，解决了局部喷洒的药物被组织液快速稀释而影响药效的问题。
附图说明
图1是本申请凝血酶缓释金属医疗器械的产品图示；
图2是制备的稳定羟基磷灰石纳米/亚微米涂层；
图3是羟基磷灰石纳米/亚微米涂层表面的凝血酶层；
图4是羟基磷灰石层的光电子能谱(XPS)曲线；
图5是凝血酶层的光电子能谱(XPS)曲线；
图6是不同时间下凝血酶平均浓度曲线图；
图7是不同时间段动态凝血酶缓释浓度的柱图。
具体实施方式
为了更好地说明本发明，下面结合本发明实施例中的附图，对本发明实施例中的技术方案进行清楚、完整地描述。
具体实施例1：实验室制备凝血酶缓释涂层
1.SUS304奥氏体不锈钢样品用去离子水超声洗涤2分钟，清除表面的油污或锈蚀。待表面干净后，放置于干燥箱中70℃干燥5分钟，以表面无 残留水滴为合格。
2.浸泡于25℃、5mol/l的羟基乙叉二膦酸溶液中2分钟，进行羟基乙叉二膦酸表面化学装换薄膜的制备。
3.取出后待多余溶液流下后，放置于干燥箱中70℃干燥10分钟，使制备的薄膜干燥。
4.待样品表面无明显液体或胶体，用去离子水冲洗样品表面3分钟，冲洗掉多余的羟基乙叉二膦酸结晶残留，剩下不易被冲洗掉的羟基乙叉二膦酸金属表面化学转换膜。放置于20g/l不断进行搅拌的羟基磷灰石(HA)悬浊液中，25℃浸泡5分钟。完成羟基乙叉二膦酸表面化学转换薄膜上的羟基磷灰石涂层的自组装制备。保留一部分样品进行羟基磷灰石涂层的光电子能谱(XPS)与原子力显微镜(AFM)表征，见附图2-5。
5.取出步骤4制备好的产品放入干燥箱中70℃干燥10分钟，待表面无残留水滴后。放入凝血酶冻干粉与0.9％生理盐水制备5000U/l凝血酶溶液中，25℃浸泡2小时。
6.取出在30℃空气中干燥2小时，进行光电子能谱(XPS)与原子力显微镜(AFM)表征。原子力显微镜表征测试结果见附图2-3,从AFM对比图可以看出，制备前后表面出现明显变化，浅色区域为凝血酶分子。光电子能谱表征测试结果见附图4-5，经过XPS对比可以看出，(1)凝血酶分子成功涂覆在羟基磷灰石表面，且有一定厚度；(2)从高分辨XPS解谱来看，没有新的化学键和产生，缓释产物是凝血酶。上述结果说明凝血酶涂层前后成分与形貌产生了明显的变化，形成了稳定的涂层，涂层为凝血酶层；没有新的化学键结合产生，说明凝血酶无其它污染物产生。
具体实施例2：工厂车间制备凝血酶缓释涂层
1.SUS304奥氏体不锈钢样品用表面清洗液浸泡2分钟，清除表面的油污，随后用去离子水清洗两次，每次2分钟。待目测表面干净后，放置于相对湿度55％温度20℃空气中干燥2小时，以表面无明显残留水滴为合格。
2.取出步骤1样品浸泡于20℃、5mol/l的羟基乙叉二膦酸溶液中5分钟，进行羟基乙叉二膦酸表面化学装换薄膜的制备。
3.步骤2样品取出后放置于相对湿度55％温度20℃空气中干燥2小时，让制备的薄膜干燥。
4.待步骤3样品表面无明显液体或胶体，放置于20g/l不断进行搅拌的羟基磷灰石(HA)悬浊液中，25℃浸泡10分钟。多余的羟基乙叉二膦酸结晶残留会在水中溶解，同时在完成羟基乙叉二膦酸表面化学装换薄膜上的羟基磷灰石涂层的自组装制备。
5.取出步骤4样品放置于相对湿度55％温度20℃空气中干燥2小时，待表面无明显残留水滴后。放入凝血酶冻干粉与0.9％生理盐水制备5000U/l凝血酶溶液中，25℃浸泡1小时。
6.取出步骤5样品在30℃空气中干燥2小时，每组取6块样品，利用酶标仪在450nm波长下分别测定不同浓度标准品的吸光度，根据不同标准品浓度及对应的吸光度绘制标准曲线。在此基础上不同时间下计算的出的平均凝血酶浓度如附图6所示，从附图6可知在0-60分钟内凝血酶释放迅速，从60-180分钟凝血酶释放十分缓慢但仍可维持一定的浓度。
将镀有凝血酶的金属板(共6块)分别浸泡于盛有200ml生理盐水的烧杯中，分别于0h、1h、2h、3h、4h、5h取样本液100ul置于标记好的EP管中，每次取样后弃去原有生理盐水，利用酶标仪测定不同时间下六组溶液的吸光度并计算平均值。不同时间下弃去浸泡溶液测得的吸光度应用t检验两两组间比较P值均大于0.05，差异无统计学意义，每隔1小时弃去浸泡溶液前测得的溶液的吸光度无显著差异，我们可以认为相同时间段内药物涂层金属板可稳定持续释放凝血酶。具体结果如附图7所示。
上述结果说明凝血酶在1小时候达到稳定缓释值，而在持续变化的溶液中可以稳定的缓释凝血酶。
通过上述方法制备的凝血酶缓释金属板，所述金属板为金属基质的医疗器械时，该凝血酶缓释医疗器械如图1所示，包括医疗器械的金属基底12、 金属表面15、磷酸盐层13、磷酸化学转换膜14、羟基磷灰石层和16和凝血酶层17。
上述试验研究表明“凝血酶-羟基磷灰石-无氧化物层-金属”钢板于生理盐水溶液中可持续稳定地释放凝血酶，从而保证溶液中凝血酶的浓度，具有凝血酶缓释效果，即，如此制备的凝血酶缓释金属基质医疗器械也具有良好的凝血酶缓释效果。
以上所述，仅为本发明较佳的具体实施方式，但本发明的保护范围并不局限于此，任何熟悉本技术领域的技术人员在本发明披露的技术范围内，可轻易想到的变化或替换，都应该涵盖在本发明的保护范围之内。因此，本发明的保护范围应该以权利要求书的保护范围为准。

Claims (10)

  1. 一种凝血酶缓释医疗器械的制备方法，其特征在于，包括如下步骤：
    (1)将金属板用去离子水超声洗涤干燥后浸泡于羟基乙叉二膦酸溶液中，取出进行二次干燥后洗涤；
    (2)将步骤(1)处理后的金属板浸泡于羟基磷灰石悬浊液中，取出干燥；
    (3)将步骤(2)处理后的金属板浸泡于凝血酶溶液中取出，得到凝血酶缓释医疗器械。
  2. 根据权利要求1所述的制备方法，其特征在于，所述步骤(1)和步骤(2)中的干燥是在干燥箱中50-70℃干燥5-10分钟或20-30℃空气中干燥1-2小时。
  3. 根据权利要求1所述的制备方法，其特征在于，所述步骤(1)中的浸泡温度为20-30℃，时间为1-2分钟。
  4. 根据权利要求1所述的制备方法，其特征在于，所述步骤(1)中的羟基乙叉二膦酸溶液浓度为3-5mol/l。
  5. 根据权利要求1所述的制备方法，其特征在于，所述步骤(1)中的二次干燥为干燥箱中50-70℃干燥10-20分钟或20-30℃空气中干燥2-4小时。
  6. 根据权利要求1所述的制备方法，其特征在于，所述步骤(1)中的洗涤时间为2-3分钟。
  7. 根据权利要求1所述的制备方法，其特征在于，所述步骤(2)中的羟基磷灰石悬浊液浓度为5-50g/l，浸泡温度为20-30℃，时间为5-10分钟。
  8. 根据权利要求1所述的制备方法，其特征在于，所述步骤(3)中的凝血酶溶液中凝血酶浓度为4000U/l-8000U/l，由凝血酶冻干粉与0.9％生理盐水制备。
  9. 根据权利要求1所述的制备方法，其特征在于，所述步骤(3)中的 浸泡温度为20-30℃，时间为1-2小时。
  10. 一种凝血酶缓释医疗器械，其特征在于，由权利要求1-9任一所述方法制备而得。
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