阅读说明：本技术 一种用于牙齿修复的引导组织再生膜的制备方法 (Preparation method of tissue regeneration guiding membrane for tooth restoration ) 是由 奚桢浩 韩宇晴 宋超波 张嘉鹏 岑莲 陆尔奕 崔立宇 于 2019-09-12 设计创作，主要内容包括：本发明公开了一种用于牙齿修复的引导组织再生膜的制备方法,包括如下步骤：(1)将合成高分子材料溶解于有机溶剂中,分散均匀,将溶液浇注于培养皿内,然后进行冷冻；得到第一层结构；(2)将天然高分子材料溶解于有机溶剂中,加入无机生物活性颗粒,分散均匀后,将上述混合溶液浇铸于所述第一层结构上,然后进行冷冻；得到复合层结构；(3)将步骤(2)得到的复合层结构进行冷冻干燥。本发明采用合成高分子和天然高分子复合材料,提高了复合膜机械性能的同时,提高其生物相容性；配合采用梯度温度冷冻的方式,调控复合膜纵向结构上孔结构的分布,形成了具有梯度结构的孔结构,更有利于细胞的粘附和生长,取得了显著的效果。(the invention discloses a preparation method of a tissue regeneration guiding membrane for tooth restoration, which comprises the following steps: (1) dissolving a synthetic high polymer material in an organic solvent, uniformly dispersing, pouring the solution into a culture dish, and freezing; obtaining a first layer structure; (2) dissolving a natural high polymer material in an organic solvent, adding inorganic bioactive particles, uniformly dispersing, casting the mixed solution on the first layer structure, and freezing; obtaining a composite layer structure; (3) and (3) freeze-drying the composite layer structure obtained in the step (2). The invention adopts the composite material of synthetic polymer and natural polymer, improves the mechanical property of the composite membrane and improves the biocompatibility of the composite membrane; the mode of gradient temperature refrigeration is adopted in a matching manner, the distribution of the pore structure on the longitudinal structure of the composite membrane is regulated and controlled, the pore structure with the gradient structure is formed, the adhesion and the growth of cells are facilitated, and the remarkable effect is obtained.)

1. A method for preparing a guided tissue regeneration membrane for dental restoration, comprising the steps of:

(1) Dissolving a synthetic high polymer material in an organic solvent, uniformly dispersing, pouring the solution into a culture dish, and freezing; obtaining a first layer structure;

the freezing step is carried out at a gradient temperature, the solution is placed in freezing chambers with different freezing temperatures from top to bottom, the temperature of a top-bottom contact freezing plate is controlled to be-200-0 ℃, and the temperature difference of the top-bottom contact freezing plate and the bottom-bottom contact freezing plate is at least 40 ℃; the temperature of the upper contact freeze plate is lower than that of the lower contact freeze plate;

(2) dissolving a natural high polymer material in an organic solvent, adding inorganic bioactive particles, uniformly dispersing, casting the mixed solution on the first layer structure, and freezing; obtaining a composite layer structure;

The natural polymer material is selected from one or more of collagen, silk fibroin, agar and pectin;

The inorganic bioactive particles are hydroxyapatite, bioglass, tricalcium phosphate, silicon dioxide or calcium carbonate;

The freezing step is carried out at a gradient temperature, the freezing chamber is placed in a freezing chamber with different upper and lower freezing temperatures, the temperature of an upper contact freezing plate and a lower contact freezing plate is controlled to be-200-0 ℃, and the temperature difference of the upper contact freezing plate and the lower contact freezing plate is at least 40 ℃; the temperature of the upper contact freeze plate is lower than that of the lower contact freeze plate;

(3) And (3) freeze-drying the composite layer structure obtained in the step (2) to obtain the guided tissue regeneration membrane for tooth restoration.

2. The method of claim 1, wherein: the synthetic polymer material in the step (1) is selected from one or more of polylactic acid, polylactic acid derivatives, polyglycolide derivatives, polylactide glycolide derivatives, polycaprolactone derivatives and polybutylene succinate.

3. the method of claim 1, wherein: the temperature difference of the upper and lower contact freezing plates in the step (1) is 40-60 ℃.

4. The method of claim 1, wherein: the temperature difference of the upper and lower contact freezing plates in the step (2) is 30-50 ℃.

5. The method of claim 1, wherein: the organic solvents in the steps (1) and (2) are the same or different and are respectively selected from one or more of chloroform, styrene, perchloroethylene, trichloroethylene, ethylene glycol ether, triethanolamine, acetic acid, ethyl acetate and hydrochloric acid.

6. The method of claim 1, wherein: and (3) dispersing in the steps (1) and (2) by adopting ultrasonic dispersion, wherein the dispersion time is 10-120 min.

7. The method of claim 1, wherein: the guided tissue regeneration membrane comprises an upper layer structure and a lower layer structure, wherein the pore size distribution of one layer corresponding to the first layer structure in the step (1) is 20-300 micrometers, and the connectivity is more than or equal to 90%; the pore size distribution of the other layer is 1-20 microns, and the connectivity is more than or equal to 50%.

8. The method of claim 1, wherein: the number average molecular weight of the high polymer material in the step (1) is 5-30 ten thousand daltons.

9. A guided tissue regeneration membrane for dental restoration, produced by the production method according to claim 1.

10. the guided tissue regeneration membrane for dental restoration according to claim 9, characterized in that: the thickness of the guided tissue regeneration membrane is 20-500 micrometers; the composite material comprises an upper layer and a lower layer, wherein the pore size distribution of the first layer is 20-300 microns, and the connectivity is more than or equal to 90%; the pore size distribution of the other layer is 1-20 microns, and the connectivity is more than or equal to 50%.

Technical Field

the invention relates to a preparation method of a guided tissue regeneration membrane for tooth restoration.

Background

periodontal disease is one of the common oral diseases of human beings, and the structure of periodontal supporting tissue is destroyed at the early stage of the disease, and as the disease becomes worse, the periodontal structure is gradually lost, and the teeth are dropped. For this disease, guided tissue regeneration has emerged. Guided Tissue Regeneration (GTR) is a technique of placing a physical barrier by a surgical method to selectively separate different periodontal tissues, preventing the gingival epithelium and the gingival connective Tissue from growing towards the root surface, creating a space, inducing the periodontal ligament cells with the Regeneration potential of the periodontal tissues to move coronally and grow and differentiate, realizing the Regeneration of periodontal ligament, alveolar bone and cementum, and forming new attachment of periodontal. Applying the Guided Tissue Regeneration (GTR) principle to the physiological regeneration of periodontal neo-attachments from Nyman S, more and more scholars are working on applying the GTR principle to clinical practice. However, due to the choice of cases, the method of surgery, and the differences in the properties of the barrier membranes used, various studies have reported wound healing and some differences in the degree of tissue regeneration. Cortellini P indicated by a summary of the study including 17 GTR clinical study reports of 651 vertical bone defects: among 651 cases of vertical bone defects, the case of adhesion loss occurred one year after GTR surgery was about 2.7%, 11% for those with clinical adhesion acquisition less than 2mm, 24.8% for those with adhesion acquisition between 2-3mm, and 21.2% for those with adhesion acquisition over 6 mm; some reports show that the same difference also exists in the degree of tissue regeneration of root bifurcation lesion one year after GTR operation. At present, studies show that the factors influencing the clinical effect mainly comprise several aspects such as selection of indications, determination of surgical methods, selection and use of membranes and the like. Among them, the guided tissue regeneration membrane (membrane material) is a high-tech product playing a key role in guided regeneration technology, and is also a core material of GTR, and is increasingly important in the field of national health and medical care.

disclosure of Invention

The invention aims to provide a preparation method of a guided tissue regeneration membrane for tooth restoration.

In order to achieve the purpose, the invention adopts a technical scheme that: a preparation method of a guided tissue regeneration membrane for tooth restoration comprises the following steps:

(1) Dissolving a synthetic high polymer material in an organic solvent, uniformly dispersing, pouring the solution into a culture dish, and freezing; obtaining a first layer structure;

The freezing step is carried out at a gradient temperature, the solution is placed in freezing chambers with different freezing temperatures from top to bottom, the temperature of a top-bottom contact freezing plate is controlled to be-200-0 ℃, and the temperature difference of the top-bottom contact freezing plate and the bottom-bottom contact freezing plate is at least 40 ℃; the temperature of the upper contact freeze plate is lower than that of the lower contact freeze plate;

(2) Dissolving a natural high polymer material in an organic solvent, adding inorganic bioactive particles, uniformly dispersing, casting the mixed solution on the first layer structure, and freezing; obtaining a composite layer structure;

the natural polymer material is selected from one or more of collagen, silk fibroin, agar and pectin;

The inorganic bioactive particles are hydroxyapatite, bioglass, tricalcium phosphate, silicon dioxide or calcium carbonate;

the freezing step is carried out at a gradient temperature, the freezing chamber is placed in a freezing chamber with different upper and lower freezing temperatures, the temperature of an upper contact freezing plate and a lower contact freezing plate is controlled to be-200-0 ℃, and the temperature difference of the upper contact freezing plate and the lower contact freezing plate is at least 40 ℃; the temperature of the upper contact freeze plate is lower than that of the lower contact freeze plate;

(3) And (3) freeze-drying the composite layer structure obtained in the step (2) to obtain the guided tissue regeneration membrane for tooth restoration.

as hereinbefore, the inorganic bioactive particles are preferably nanoscale particles. The mass fraction of the added inorganic bioactive particles is preferably 0.1-10% calculated by taking the natural polymer material as 100 parts by mass; more preferably 1% to 6%, and still more preferably 3% to 5%.

Above, the natural polymer material is preferably collagen and/or silk fibroin.

The freezing temperatures in the steps (1) and (2) are the same or different and are respectively selected from any section of temperature in the range of-200 to 0 ℃, the temperature in the section is not specifically limited in the invention, and typically but not limited to-200 to-40 ℃, 60 to-20 ℃, 120 to 0 ℃, 150 to-30 ℃, 200 to-30 ℃ and the like.

In the above technical scheme, the synthetic polymer material in step (1) is selected from one or more of polylactic acid, polylactic acid derivatives, polyglycolide derivatives, polylactide glycolide derivatives, polycaprolactone derivatives, and polybutylene succinate.

In the step (1), the synthetic polymer material is preferably polylactide glycolide or a polylactide glycolide derivative; and the ratio of the polylactide to the polyglycolide monomer is 1: 9-9: 1, preferably 7: 3-5: 5.

in the technical scheme, the temperature difference of the upper and lower contact freezing plates in the step (1) is 40-60 ℃. Preferably 45 to 55 ℃. More preferably 50 to 52 ℃.

In the technical scheme, the temperature difference of the upper and lower contact freezing plates in the step (2) is 30-50 ℃. Preferably 40 to 45 ℃. More preferably 46 to 48 ℃.

In the above technical scheme, the organic solvents in the steps (1) and (2) are the same or different and are respectively selected from one or more of chloroform, styrene, perchloroethylene, trichloroethylene, ethylene glycol ether, triethanolamine, acetic acid, ethyl acetate and hydrochloric acid.

In the technical scheme, ultrasonic dispersion is adopted for dispersion in the steps (1) and (2), and the dispersion time is 10-120 min. The ultrasonic dispersion time is not particularly limited, and may be, typically but not limited to, 10min, 20min, 30min, 40min, 50min, 60min, 70min, 80min, 90min, 100min, and the like.

In the technical scheme, the guided tissue regeneration membrane comprises an upper layer and a lower layer which are 2 layers, wherein the pore size distribution of one layer corresponding to the first layer of the structure in the step (1) is 20-300 microns, and the connectivity is more than or equal to 90%; the pore size distribution of the other layer is 1-20 microns, and the connectivity is more than or equal to 50%.

In the technical scheme, the number average molecular weight of the high polymer material in the step (1) is 5-30 ten thousand daltons. Preferably 10 to 20 kilodaltons. More preferably 12 to 18 kilodaltons. More preferably 15 to 17 kilodaltons.

The invention also discloses a guided tissue regeneration membrane for tooth restoration, which is prepared by the preparation method.

In the technical scheme, the thickness of the guided tissue regeneration membrane is 20-500 micrometers; the composite material comprises an upper layer and a lower layer, wherein the pore size distribution of the first layer is 20-300 microns, and the connectivity is more than or equal to 90%; the pore size distribution of the other layer is 1-20 microns, and the connectivity is more than or equal to 50%.

the working principle of the invention is as follows: on one hand, the composite membrane with a double-layer framework structure is adopted, the synthetic polymer material of the first layer plays a role in mechanical support, the mechanical property of the composite membrane is enhanced, and the natural polymer material and the inorganic bioactive particles of the second layer enhance the biocompatibility of the composite membrane, so that the adhesion, proliferation and differentiation of cells are facilitated; on the other hand, the invention adopts a gradient freezing mode, and by controlling the temperature of the upper and lower contact freezing plates, a solution forms smaller ice crystals during quick freezing and larger ice crystals during slow freezing, so that the smaller ice crystals are formed near the freezing plate with lower temperature and the larger ice crystals are formed near the freezing plate with higher temperature, and pore canals with different sizes are formed in the composite membrane through freeze drying, thereby realizing the distribution structure of the gradient pore structure in the material, forming a compact structure with smaller pore diameter in the synthetic polymer material layer, and forming the gradient pore structure with larger pore diameter in the natural polymer material layer; when the periodontal bone repairing material is used, one side of the compact structure is attached to one side of periodontal soft tissue, so that infiltration of connective tissue in the repairing process can be prevented, the natural polymer material layer is attached to one side of periodontal bone defect, and the gradient pore structure can promote differentiation of cells and provide a space for bone cell growth.

Due to the application of the technical scheme, compared with the prior art, the invention has the following advantages:

1) The invention adopts the composite material of synthetic polymer and natural polymer, improves the mechanical property of the composite membrane and improves the biocompatibility of the composite membrane; simultaneously, the cooperation adopts the frozen mode of gradient temperature, and the distribution of regulation and control complex film longitudinal structure goes up pore structure has formed the pore structure that has the gradient structure, and the experiment proves: compared with the existing uniform pore structure, the invention is more beneficial to the adhesion and growth of cells and obtains remarkable effect;

2) According to the invention, the membrane material with a porous structure distributed in multiple layers can be controllably prepared by adjusting the concentration of a natural high polymer material and the addition of inorganic bioactive particles, so that the medical biodegradable composite membrane with an upper and lower 2-layer structure with different pore diameters is prepared, the tissue regeneration of a defect area can be well guided, the invasion of soft tissues can be prevented, the requirement of guided regeneration can be met, and the purpose of tissue repair can be achieved;

3) the invention creatively introduces natural polymer materials and is matched with inorganic bioactive particles, which is beneficial to the adhesion and growth of cells, provides excellent conditions for postoperative recovery and has positive practical significance.

drawings

FIG. 1 is a schematic representation of a composite membrane structure of example 1 of the present invention.

FIGS. 2, 5, 6 and 7 are SEM images of the product interface of example 1 of the present invention.

FIG. 3 is an SEM image of the product interface of example 2 of the present invention.

FIG. 4 is an SEM image of the product interface of example 3 of the present invention.

Detailed Description

the invention is further described with reference to the accompanying drawings and examples: