技术领域Technical Field
本发明涉及骨粘接剂制备技术领域，具体涉及一种湿环境长期有效的骨粘接剂的制备方法与应用。The invention relates to the technical field of bone adhesive preparation, and in particular to a preparation method and application of a bone adhesive effective in a wet environment for a long time.
背景技术Background Art
临床的多种场景下需要进行颌面部骨的固定(如颌面部骨折、创伤后或种植治疗前期骨移植等等)。颌骨骨折的处理不当可造成患者咬合错乱、张口受限、咀嚼功能障碍。而骨移植的固定不佳可能影响后续的赝复体治疗或种植治疗。为了达到这一目的，临床操作中往往需要引入人工材料，如钛钉、钛板等进行骨的固定。Maxillofacial bone fixation is required in many clinical scenarios (such as maxillofacial fractures, bone transplantation after trauma or before implant treatment, etc.). Improper treatment of jaw fractures can cause malocclusion, limited mouth opening, and chewing dysfunction in patients. Poor fixation of bone transplantation may affect subsequent prosthesis treatment or implant treatment. In order to achieve this goal, artificial materials such as titanium nails and titanium plates are often introduced in clinical operations for bone fixation.
传统的固定方法虽能达到良好的效果，但存在着材料滞留在体内无法降解、应力阻挡作用、金属微电解反应和需要二次手术取出等问题。因此，开发具有生物安全性、可降解性、良好的粘接力的生物骨粘接剂成为目前研究的热点。Although traditional fixation methods can achieve good results, there are problems such as material retention in the body and inability to degrade, stress barrier effect, metal microelectrolysis reaction and the need for secondary surgery to remove it. Therefore, the development of biological bone adhesives with biosafety, degradability and good adhesion has become a hot topic in current research.
目前，现有技术中，广义的骨粘接材料包括磷酸盐骨胶(CPC)、骨水泥(PMMA基聚合物)和生物粘接剂(人工合成、仿生、天然基)等在内，涵盖了固体颗粒、粉剂、液体、凝胶等多种形式。在临床报道的产品中，CPC的成分与天然骨组织最为相似。但其自身的稳定性较差，在潮湿环境中易松散塌陷，导致不利的骨破坏。PMMA作为一种大分子高聚物，其粘附性不佳，降解产物有较强的毒性。而现有的生物粘接剂中，使用方便且粘结性好的氰基丙烯酸酯类(CA)生物相容性差，而生物相容性好的仿生粘接剂(如贻贝类粘接材料)或天然基粘接剂等又多限于软组织粘接，如皮肤创口的愈合、肠瘘的修补等等。此外，骨粘接材料还应具有在湿环境中的长期粘接能力，这是该类材料开发的难点。At present, in the prior art, the broad sense of bone adhesive materials includes phosphate bone glue (CPC), bone cement (PMMA-based polymer) and bioadhesives (synthetic, biomimetic, natural-based), etc., covering various forms such as solid particles, powders, liquids, gels, etc. Among the products reported clinically, the composition of CPC is most similar to natural bone tissue. However, its own stability is poor, and it is easy to loosen and collapse in a humid environment, resulting in adverse bone damage. PMMA, as a macromolecular polymer, has poor adhesion and its degradation products are highly toxic. Among the existing bioadhesives, cyanoacrylates (CA), which are easy to use and have good adhesion, have poor biocompatibility, while biomimetic adhesives (such as mussel adhesives) or natural-based adhesives with good biocompatibility are mostly limited to soft tissue bonding, such as healing of skin wounds, repair of intestinal fistulas, etc. In addition, bone adhesives should also have long-term bonding ability in a wet environment, which is a difficulty in the development of such materials.
因此，本申请旨在提供一种具有良好的长时间湿环境粘接能力、生物相容性、生物可降解性的操作便捷的骨粘接材料及制备方法与应用，在多种颌面骨固定场景下提供稳定的粘接力和长久的固位作用，以解决上述问题。Therefore, the present application aims to provide a bone bonding material and preparation method and application that are easy to operate and have good long-term wet environment bonding ability, biocompatibility, and biodegradability, so as to provide stable bonding force and long-term retention effect in various maxillofacial bone fixation scenarios to solve the above problems.
发明内容Summary of the invention
本发明的目的是为了解决现有技术中的骨粘接剂不能在体内湿环境中有效进行骨粘接的问题，提供一种湿环境长期有效的骨粘接剂的制备方法与应用。The purpose of the present invention is to solve the problem that the bone adhesive in the prior art cannot effectively perform bone bonding in a wet environment in vivo, and to provide a preparation method and application of a bone adhesive that is effective in a wet environment for a long time.
为了达到上述目的，本发明的技术方案如下：In order to achieve the above object, the technical solution of the present invention is as follows:
本发明提供了一种湿环境长期有效的骨粘接剂，所述骨粘接剂由组分1和组分2构成，所述组分1和组分2分别为聚-[(N-三羟甲基甲基丙烯酰胺)-氨基丙基甲基丙烯酰胺](ATMA)和醛化甲基纤维素(OMC)；所述聚-[(N-三羟甲基甲基丙烯酰胺)-氨基丙基甲基丙烯酰胺]与醛化甲基纤维素的物质的量之比为0.6:1,1:1，1.4:1。The invention provides a bone adhesive which is effective in a wet environment for a long time. The bone adhesive consists of a component 1 and a component 2. The component 1 and the component 2 are poly-[(N-trihydroxymethylmethylacrylamide)-aminopropylmethacrylamide] (ATMA) and hydroformyl methylcellulose (OMC), respectively. The molar ratio of the poly-[(N-trihydroxymethylmethylacrylamide)-aminopropylmethacrylamide] to the hydroformyl methylcellulose is 0.6:1, 1:1, or 1.4:1.
本发明还提供了上述一种湿环境长期有效的骨粘接剂的制备方法，包括以下步骤：The present invention also provides a method for preparing the above-mentioned bone adhesive which is effective in a wet environment for a long time, comprising the following steps:
S1、聚-[(N-三羟甲基甲基丙烯酰胺)-氨基丙基甲基丙烯酰胺]的制备：S1. Preparation of poly-[(N-trimethylol methacrylamide)-aminopropyl methacrylamide]:
采用自由基聚合法，将物质的量之比为0.25：0.8：1的原料氨基丙基甲基丙烯酰胺、N-三羟基甲基丙烯酰胺和过硫酸铵采用去离子水溶解混合，于惰性气体环境，持续搅拌，并于60℃加热一小时后，升温至70℃加热12小时，反应完毕后，用Mw＝3500的透析袋透析2天，之后冻干，得组分1；Using a free radical polymerization method, raw materials aminopropyl methacrylamide, N-trihydroxy methacrylamide and ammonium persulfate in a molar ratio of 0.25:0.8:1 were dissolved and mixed in deionized water, and continuously stirred in an inert gas environment, and heated at 60°C for one hour, and then heated to 70°C for 12 hours. After the reaction was completed, dialyzed with a dialysis bag with Mw=3500 for 2 days, and then freeze-dried to obtain component 1;
S2、醛化甲基纤维素的制备：S2. Preparation of hydroformylation methyl cellulose:
采用高碘酸钠氧化法，将甲基纤维素、0.3mo l/L高碘酸钠溶液于反应容器中采用去离子水溶解混合；然后，于常温黑暗条件下持续搅拌24小时；然后加入反应终止剂乙二醇，并于半小时后反应终止，用Mw＝3500的透析袋透析3天，之后冻干，得组分2；Using sodium periodate oxidation method, methyl cellulose and 0.3 mol/L sodium periodate solution were dissolved and mixed in a reaction container with deionized water; then, stirring was continued for 24 hours at room temperature in the dark; then, reaction terminator ethylene glycol was added, and the reaction was terminated after half an hour, and dialyzed with a dialysis bag with Mw=3500 for 3 days, and then freeze-dried to obtain component 2;
S3、骨粘接剂的制备：S3. Preparation of bone adhesive:
取冻干后的组分1和组分2，其中，组分1与组分2的物质的量之比为0.6:1、1.0:1、1.4:1，然后将其分别以去离子水溶解，得到质量体积比为15％的两组分，然后将两组分搅拌混匀，根据席夫碱反应，于5分钟内迅速形成水凝胶，即制得水凝胶骨粘接剂。Take freeze-dried component 1 and component 2, wherein the molar ratio of component 1 to component 2 is 0.6:1, 1.0:1, and 1.4:1, and then dissolve them in deionized water to obtain two components with a mass volume ratio of 15%, and then stir and mix the two components. According to the Schiff base reaction, a hydrogel is quickly formed within 5 minutes to obtain a hydrogel bone adhesive.
进一步地，步骤S1中每1mol N-三羟基甲基丙烯酰胺采用250mL去离子水。Furthermore, in step S1, 250 mL of deionized water is used for every 1 mol of N-trihydroxymethylacrylamide.
进一步地，步骤S2中反应终止剂乙二醇与去离子水总体积比为0.00152：1；步骤S2中每1g甲基纤维素需100mL去离子水。Furthermore, in step S2, the total volume ratio of the reaction terminator ethylene glycol to deionized water is 0.00152:1; and in step S2, 100 mL of deionized water is required for every 1 g of methyl cellulose.
本发明还提供了上述一种湿环境长期有效的骨粘接剂的制备方法的应用，所述方法制得的水凝胶骨粘接剂用于可在干湿环境中放置或注射至需粘接的骨表面的骨粘接剂的制备。The present invention also provides an application of the method for preparing the above-mentioned bone adhesive that is effective in a long-term wet environment. The hydrogel bone adhesive prepared by the method is used for preparing a bone adhesive that can be placed in a dry or wet environment or injected onto a bone surface to be bonded.
在本方案中，制备好的骨粘接剂具有可注射性，且操作时间长，在形成后4个小时内均可注射。可在干湿环境中放置或注射至需粘接的骨表面，固定20秒即可达到强力有效的粘接。聚-[(3-羟甲基甲基丙烯酰胺)-氨基丙基甲基丙烯酰胺]与醛化甲基纤维素的摩尔质量比为1:1的水凝胶骨粘接剂在大鼠体内应用时，可达到嵌入式骨块的粘接和Onlay式骨块的粘接，与商用氰基丙烯酸酯粘接剂对比，其生物安全性更佳。In this scheme, the prepared bone adhesive is injectable and has a long operation time. It can be injected within 4 hours after formation. It can be placed in a dry or wet environment or injected onto the bone surface to be bonded, and fixed for 20 seconds to achieve strong and effective bonding. When the hydrogel bone adhesive with a molar mass ratio of poly-[(3-hydroxymethyl methacrylamide)-aminopropyl methacrylamide] and aldehyde methyl cellulose of 1:1 is used in rats, it can achieve the bonding of embedded bone blocks and Onlay bone blocks. Compared with commercial cyanoacrylate adhesives, it has better biosafety.
与现有技术相比，本方案的有益效果：Compared with the prior art, this solution has the following beneficial effects:
1、可操作性强，操作时间长且可注射；2、湿环境骨粘接力，对湿骨的粘接力可达2MPa，在湿环境中能保持长达14天的骨粘接；3、生物相容性好，在体外、体内实验中已证实具有良好的生物相容性、生物可降解性；4、体内有效骨粘接，在体内模型中已证实可以完成嵌入式骨粘接和Onlay式骨粘接。1. Strong operability, long operation time and injectable; 2. Bone bonding in wet environment, the bonding force to wet bone can reach 2MPa, and bone bonding can be maintained for up to 14 days in a wet environment; 3. Good biocompatibility, it has been confirmed in in vitro and in vivo experiments to have good biocompatibility and biodegradability; 4. Effective bone bonding in vivo, it has been confirmed in in vivo models that embedded bone bonding and Onlay bone bonding can be completed.
附图说明BRIEF DESCRIPTION OF THE DRAWINGS
图1是本发明实施例中ATMA和OMC的合成反应、交联反应原理图；FIG1 is a schematic diagram of the synthesis reaction and cross-linking reaction of ATMA and OMC in an embodiment of the present invention;
图2是本发明实施例中ATMA的氢谱(400MHz)；FIG2 is a hydrogen spectrum (400 MHz) of ATMA in an embodiment of the present invention;
图3是本发明实施例中OMC的傅里叶红外光谱图(FTIR)；FIG3 is a Fourier transform infrared spectrum (FTIR) of OMC in an embodiment of the present invention;
图4是本发明实施例中聚-[(N-三羟甲基甲基丙烯酰胺)-氨基丙基甲基丙烯酰胺]，醛化甲基纤维素制得的水凝胶骨粘接剂的傅里叶红外光谱图(FTIR)；4 is a Fourier transform infrared spectrum (FTIR) of a hydrogel bone adhesive prepared from poly-[(N-trihydroxymethyl methacrylamide)-aminopropyl methacrylamide] and hydroformylation of methyl cellulose in an embodiment of the present invention;
图5是本发明实施例中水凝胶骨粘接剂的扫描电镜图；FIG5 is a scanning electron microscope image of a hydrogel bone adhesive in an embodiment of the present invention;
图6是本发明实施例中骨粘接剂具有可注射性和较长的操作时间；FIG6 is an embodiment of the present invention showing that the bone adhesive has injectability and a longer operation time;
图7是本发明实施例中骨粘接剂具有自愈合性；FIG7 is a diagram showing that the bone adhesive has self-healing properties in an embodiment of the present invention;
图8是本发明实施例中骨粘接剂具有快速湿粘接特性；FIG8 is a diagram showing that the bone adhesive has rapid wet bonding properties in an embodiment of the present invention;
图9是本发明实施例中骨粘接剂成型后具有稳定性；FIG9 is a diagram showing the stability of the bone adhesive after molding in an embodiment of the present invention;
图10是本发明实施例中骨粘接剂具有流变剪切稀释特性；FIG10 is a diagram showing a bone adhesive having rheological shear thinning properties in an embodiment of the present invention;
图11是本发明实施例中骨粘接剂的湿环境粘接力检测；11 is a wet environment bonding force test of the bone adhesive in an embodiment of the present invention;
图12是本发明实施例中骨粘接剂的体外溶胀和降解性能；FIG12 is a diagram showing the in vitro swelling and degradation properties of the bone adhesive in an embodiment of the present invention;
图13是本发明实施例中骨粘接剂的体外生物安全性；FIG13 is a diagram showing the in vitro biological safety of the bone adhesive in an embodiment of the present invention;
图14是本发明实施例中骨粘接剂的体内生物安全性；FIG14 is a diagram showing the in vivo biological safety of the bone adhesive in an embodiment of the present invention;
图15是本发明实施例中骨粘接剂在大鼠体内的嵌入式粘接；FIG15 is an embedded bonding of a bone adhesive in a rat according to an embodiment of the present invention;
图16是本发明实施例中骨粘接剂在大鼠体内的Onlay式粘接。FIG. 16 is an Onlay bonding of the bone adhesive in a rat according to an embodiment of the present invention.
具体实施方式DETAILED DESCRIPTION
为了使本技术领域的人员更好地理解本发明方案，下面将结合本发明的实施例及附图，对本发明的技术方案进行进一步详细地描述，显然，所描述的实施例仅仅是本发明一部分的实施例，而不是全部的实施例。基于本发明中的实施例，本领域普通技术人员在没有做出创造性劳动前提下所获得的所有其他实施例，都应当属于本发明保护的范围。In order to enable those skilled in the art to better understand the solution of the present invention, the technical solution of the present invention will be further described in detail below in conjunction with the embodiments of the present invention and the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work should fall within the scope of protection of the present invention.
需要说明的是，在不冲突的情况下，本发明中的实施例及实施例中的特征可以相互组合。下面将结合实施例来详细说明本发明。It should be noted that, in the absence of conflict, the embodiments and features in the embodiments of the present invention can be combined with each other. The present invention will be described in detail below in conjunction with the embodiments.
实施例：Example:
本发明实施例提供的方案如上述发明内容所述，提供了一种湿环境长期有效的骨粘接剂的制备方法与应用。The solution provided by the embodiment of the present invention is as described in the above invention content, and provides a preparation method and application of a bone adhesive that is effective in a wet environment for a long time.
具体实施例过程为：The specific implementation process is:
1、本水凝胶骨粘接剂由两个组分构成，分别为：①聚-[(N-三羟甲基甲基丙烯酰胺)-氨基丙基甲基丙烯酰胺](ATMA)，②醛化甲基纤维素(OMC)。1. The hydrogel bone adhesive is composed of two components: ① poly-[(N-trihydroxymethyl methacrylamide)-aminopropyl methacrylamide] (ATMA) and ② aldehyde methyl cellulose (OMC).
组分1ATMA的制备原理为自由基聚合法。具体制备方法如下。The preparation principle of component 1ATMA is free radical polymerization. The specific preparation method is as follows.
①条件：惰性气体环境，持续搅拌；①Conditions: inert gas environment, continuous stirring;
②温度：60℃加热一小时后，升温至70℃加热12小时；② Temperature: After heating at 60℃ for one hour, heat to 70℃ for 12 hours;
③原料：氨基丙基甲基丙烯酰胺，N-三羟基甲基丙烯酰胺和过硫酸铵，摩尔比为：0.25：0.8：1。③ Raw materials: aminopropyl methacrylamide, N-trihydroxymethyl acrylamide and ammonium persulfate, the molar ratio is 0.25:0.8:1.
④溶剂：去离子水(每1mol N-三羟基甲基丙烯酰胺需250ml去离子水)。反应完毕后，用Mw＝3500的透析袋透析2天，之后冻干。④Solvent: deionized water (250 ml of deionized water is required for every 1 mol of N-trihydroxymethyl acrylamide). After the reaction is completed, dialyze for 2 days using a dialysis bag with Mw=3500, and then freeze-dry.
组分2OMC的制备原理为高碘酸钠氧化法。具体制备方法如下。The preparation principle of component 2OMC is sodium periodate oxidation method. The specific preparation method is as follows.
①条件：黑暗条件，持续搅拌24小时；①Conditions: dark conditions, continuous stirring for 24 hours;
②温度：常温；②Temperature: normal temperature;
③原料：甲基纤维素，0.3mol/L高碘酸钠溶液(体积为去离子水的1/10)；③ Raw materials: methyl cellulose, 0.3 mol/L sodium periodate solution (the volume is 1/10 of deionized water);
④溶剂：去离子水(每1g甲基纤维素需100ml去离子水)；④Solvent: deionized water (100 ml of deionized water is required for every 1 g of methyl cellulose);
⑤反应终止剂：乙二醇(与去离子水总体积比为0.00152：1)；⑤ Reaction terminator: ethylene glycol (total volume ratio to deionized water is 0.00152:1);
加入反应终止剂后半小时，反应终止，用Mw＝3500的透析袋透析3天，之后冻干。The reaction was terminated half an hour after the addition of the reaction terminator, and the product was dialyzed for 3 days using a dialysis bag with Mw=3500, followed by freeze-drying.
2、骨粘接剂的制备2. Preparation of bone adhesive
取冻干后的组分1(以下又称A)和组分2(以下又称O)，A与O的摩尔比为0.6、1.0、1.4(后称为A-O-0.6,A-O-1.0,A-O-1.4)，分别以去离子水溶解，得到质量体积比为15％的两组分，然后将两组分搅拌混匀，由于席夫碱反应的存在，可于5分钟内迅速形成水凝胶。Take freeze-dried component 1 (hereinafter referred to as A) and component 2 (hereinafter referred to as O), the molar ratio of A to O is 0.6, 1.0, and 1.4 (hereinafter referred to as A-O-0.6, A-O-1.0, and A-O-1.4), respectively, and dissolve them in deionized water to obtain two components with a mass volume ratio of 15%. Then, the two components are stirred and mixed. Due to the presence of the Schiff base reaction, a hydrogel can be quickly formed within 5 minutes.
3、骨粘接剂的应用3. Application of bone adhesive
制备好的骨粘接剂具有可注射性，且操作时间长，在形成后4个小时内均可注射。可在干湿环境中放置或注射至需粘接的骨表面，固定20秒即可达到强力有效的粘接。A-O-1.0水凝胶骨粘接剂在大鼠体内应用时，可达到嵌入式骨块的粘接和Onlay式骨块的粘接，与商用氰基丙烯酸酯粘接剂对比，其生物安全性更佳。The prepared bone adhesive is injectable and has a long operation time. It can be injected within 4 hours after formation. It can be placed in a dry or wet environment or injected onto the bone surface to be bonded. Fixing for 20 seconds can achieve strong and effective bonding. When A-O-1.0 hydrogel bone adhesive is used in rats, it can achieve the bonding of embedded bone blocks and Onlay bone blocks. Compared with commercial cyanoacrylate adhesives, it has better biosafety.
在本实施例中，图1为ATMA和OMC的合成反应、交联反应，ATMA上含有胺基，OMC上含有醛基，二者可通过席夫碱反应交联；如图2所示，其为ATMA的氢谱(400MHz)，图2中的峰的分布与走向表明成功合成了ATMA。如图3所示，其为OMC的傅里叶红外光谱图(FTIR)，图3中，1740cm-1左右的特征峰证明在MC中引入了醛基，成为OMC。图4为A-O水凝胶骨粘接剂的傅里叶红外光谱图(FTIR)，图4中可看出，水凝胶具有两组分的特征峰，在1638cm-1处的峰变化证明发生了席夫碱反应；图5为水凝胶骨粘接剂的扫描电镜图，显示其具有均匀的多孔形态。图6为骨粘接剂具有可注射性和较长的操作时间，刚成型的水凝胶具有可注射性，放置4个小时后，仍可注射。图7为骨粘接剂具有自愈合性，图7中分别将两块染成不同颜色的水凝胶切成两半，重新组合后经过5分钟，水凝胶恢复为完整的圆形。图8为骨粘接剂具有快速湿粘接特性，图8中将水凝胶注射到骨块表面，与浸泡在PBS中的骨块按压20秒后即可形成牢固粘接。图9为骨粘接剂成型后具有稳定性，如图9所示，在1％应变，25℃加载下，随着时间的增加，水凝胶的储存模量(G”)和损耗模量(G’)下降，提示其具有剪切变稀性能。图10为骨粘接剂具有流变剪切稀释特性，如图10所示，在1％应变，25℃下，随着剪切速率的增加，水凝胶黏度下降，提示其具有剪切变稀性能。图11为骨粘接剂的湿环境粘接力检测，如图11的结果所示，将粘接好的骨块浸泡于PBS中进行后续的拉伸测试。发现在0.5小时后粘接力达到峰值，并能保持长达14天的湿环境粘接能力。图12为骨粘接剂的体外溶胀和降解性能，图12中，将不同组水凝胶浸泡在PBS中，24小时后测试其重量变化，计算出溶胀率。A-O-1.0组水凝胶具有较为适宜的溶胀率。将不同组水凝胶浸泡在PBS中，观察不同时间其降解率。发现6周后水凝胶大部分降解。图13为骨粘接剂的体外生物安全性，图13中，通过CCK8测试研究三组水凝胶对MC-3T3-E1(小鼠胚胎成骨细胞)活性的影响，可以看出三组水凝胶具有良好的生物相容性，其中A-O-1.0水凝胶最佳。图14为骨粘接剂的体内生物安全性，图14中，通过大鼠背部皮下埋置，观察到随时间增加，水凝胶逐渐降解。对7天和28天的大鼠血样进行分析，发现红细胞、白细胞和血小板的值均与对照组相比无明显变化。对埋置处皮肤进行取样染色，HE、Masson染色均未发现炎性变化。同时，进行器官毒性研究，发现水凝胶对心、肝、脾、肺、肾无组织学方面的影响。图15为骨粘接剂在大鼠体内的嵌入式粘接，图15中，通过制造大鼠颅骨缺损，再将直径5mm的骨片利用水凝胶原位粘接，对空白、对照、骨粘接剂组的1、2、4周骨片固位力进行检测。发现水凝胶固定的骨片固位力最佳。图16为骨粘接剂在大鼠体内的Onlay式粘接，图16中，通过制取直径5mm的大鼠颅骨骨片，再将其粘接到大鼠皮质骨上，完成Onlay式粘接。发现4周后骨片在位，且与皮质骨发生紧密的结合，证明骨粘接剂对于Onlay粘接有效。In the present embodiment, FIG. 1 is a synthesis reaction and a cross-linking reaction of ATMA and OMC. ATMA contains an amine group and OMC contains an aldehyde group. The two can be cross-linked by a Schiff base reaction; as shown in FIG. 2, it is a hydrogen spectrum (400MHz) of ATMA. The distribution and trend of the peaks in FIG. 2 indicate that ATMA has been successfully synthesized. As shown in FIG. 3, it is a Fourier infrared spectrum (FTIR) of OMC. In FIG. 3, the characteristic peaks around 1740cm-1 prove that an aldehyde group is introduced into MC to become OMC. FIG. 4 is a Fourier infrared spectrum (FTIR) of AO hydrogel bone adhesive. It can be seen from FIG. 4 that the hydrogel has characteristic peaks of two components, and the peak change at 1638cm-1 proves that a Schiff base reaction has occurred; FIG. 5 is a scanning electron microscope image of a hydrogel bone adhesive, showing that it has a uniform porous morphology. FIG. 6 shows that the bone adhesive has injectability and a long operation time. The newly formed hydrogel has injectability and can still be injected after being placed for 4 hours. Figure 7 shows that the bone adhesive has self-healing properties. In Figure 7, two hydrogels dyed in different colors were cut into two halves. After 5 minutes of reassembly, the hydrogels returned to a complete circle. Figure 8 shows that the bone adhesive has rapid wet bonding properties. In Figure 8, the hydrogel was injected into the surface of the bone block and pressed against the bone block soaked in PBS for 20 seconds to form a strong bond. Figure 9 shows that the bone adhesive has stability after molding. As shown in Figure 9, under 1% strain and 25°C loading, the storage modulus (G”) and loss modulus (G’) of the hydrogel decreased with time, indicating that it has shear-thinning properties. Figure 10 shows that the bone adhesive has rheological shear-thinning properties. As shown in Figure 10, under 1% strain and 25°C, the viscosity of the hydrogel decreased with the increase of shear rate, indicating that it has shear-thinning properties. Figure 11 shows the wet environment bonding test of the bone adhesive. As shown in the results of Figure 11, the bonded bone blocks were immersed in PBS for subsequent tensile tests. It was found that after 0.5 hours The bonding force reaches its peak value after that and can maintain the bonding ability in a wet environment for up to 14 days. Figure 12 shows the in vitro swelling and degradation properties of the bone adhesive. In Figure 12, different groups of hydrogels were immersed in PBS, and their weight changes were tested after 24 hours, and the swelling rate was calculated. The AO-1.0 group of hydrogels has a more suitable swelling rate. Different groups of hydrogels were immersed in PBS, and their degradation rates were observed at different times. It was found that most of the hydrogels were degraded after 6 weeks. Figure 13 shows the in vitro biosafety of the bone adhesive. In Figure 13, the CCK8 test was used to study the effects of the three groups of hydrogels on the activity of MC-3T3-E1 (mouse embryonic osteoblasts) It can be seen that the three groups of hydrogels have good biocompatibility, among which AO-1.0 hydrogel is the best. Figure 14 shows the in vivo biosafety of the bone adhesive. In Figure 14, the hydrogel was gradually degraded over time by subcutaneous implantation in the back of rats. Blood samples from rats at 7 and 28 days were analyzed, and it was found that the values of red blood cells, white blood cells, and platelets were no significantly changed compared with those of the control group. The skin at the implantation site was sampled and stained, and no inflammatory changes were found by HE and Masson staining. At the same time, organ toxicity studies were conducted, and it was found that the hydrogel had no histological effect on the heart, liver, spleen, lungs, and kidneys. Figure 15 is Embedded bonding of bone adhesive in rats. In Figure 15, a rat skull defect was created, and then a bone piece with a diameter of 5 mm was bonded in situ using hydrogel. The retention force of the bone piece in the blank, control, and bone adhesive groups was tested at 1, 2, and 4 weeks. It was found that the bone piece fixed with hydrogel had the best retention force. Figure 16 shows the Onlay bonding of bone adhesive in rats. In Figure 16, a rat skull bone piece with a diameter of 5 mm was prepared and then bonded to the rat cortical bone to complete the Onlay bonding. It was found that the bone piece was in place after 4 weeks and was tightly bonded to the cortical bone, proving that bone adhesive is effective for Onlay bonding.
通过本发明的上述实施例，本发明设计了一种双组分水凝胶骨粘接剂，具有持续时间长的湿环境骨粘接力，该水凝胶骨粘接剂可在体内实现多种类型的骨粘接，尤其报道较少的Onlay式骨粘接。本发明方法制得的该骨粘接剂材料操作时间长，具有可注射性，对湿骨的粘接力可达2MPa，在湿环境中能保持长达14天的骨粘接，并具有良好的生物相容性、生物可降解性，在体内模型中已证实可以完成嵌入式骨粘接和Onlay式骨粘接。Through the above embodiments of the present invention, the present invention designs a two-component hydrogel bone adhesive with a long-lasting bone bonding force in a wet environment. The hydrogel bone adhesive can achieve various types of bone bonding in vivo, especially the less reported Onlay bone bonding. The bone adhesive material prepared by the method of the present invention has a long operation time, is injectable, has an adhesion force of up to 2MPa to wet bones, can maintain bone bonding for up to 14 days in a wet environment, and has good biocompatibility and biodegradability. It has been confirmed in an in vivo model that embedded bone bonding and Onlay bone bonding can be completed.
以上具体实施例仅仅是对本发明的解释，其并不是对本发明的限制，本领域技术人员在阅读完本说明书后可以根据需要对本实施例做出没有创造性贡献的修改，但只要在本发明的权利要求范围内都受到专利法的保护。The above specific embodiments are merely explanations of the present invention, and are not limitations of the present invention. After reading this specification, those skilled in the art may make modifications to the embodiments without any creative contribution as needed. However, such modifications are protected by the patent law as long as they are within the scope of the claims of the present invention.
| Application Number | Priority Date | Filing Date | Title |
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| CN202310367952.6ACN116440313B (en) | 2023-04-07 | 2023-04-07 | Preparation method and application of bone adhesive capable of being used in wet environment for long term |
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| CN202310367952.6ACN116440313B (en) | 2023-04-07 | 2023-04-07 | Preparation method and application of bone adhesive capable of being used in wet environment for long term |
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| CN202310367952.6AActiveCN116440313B (en) | 2023-04-07 | 2023-04-07 | Preparation method and application of bone adhesive capable of being used in wet environment for long term |
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