CN115737909A - Preparation method and application of nanofiber material taking polycaprolactone as matrix - Google Patents

Abstract

The invention provides a nanofiber material with polycaprolactone as a matrix and a preparation method thereof, comprising the following steps: mixing and dissolving a polycaprolactone masterbatch and an organic solvent in a certain proportion, and electrospinning the obtained slurry The fiber-forming film is prepared, the surface modification and drug loading are carried out after drying, and the nanometer fiber material is obtained after drying and sterilization. The polycaprolactone-based nanofiber material provided by the invention has relatively stable performance and chemical stability in vivo, has good biocompatibility, and has excellent performance in promoting tissue repair and drug loading.

Description

A kind of preparation method and application of nanofiber material with polycaprolactone as matrix

technical field

The invention relates to the technical field of biomaterials, in particular to a nanofiber material with polycaprolactone as a matrix, a preparation method and application thereof.

Background technique

Biomaterials are a class of new high-tech materials used to diagnose, treat, repair or replace human tissues and organs or enhance their functions. They involve the health of hundreds of millions of people and are necessary to ensure human health. Its application has not only saved the lives of tens of thousands of critically ill patients, significantly reduced the mortality of cardiovascular diseases, cancer, trauma and other major diseases, but also played an important role in improving the quality of life and health of patients and reducing medical costs.

Gastrointestinal tract and pancreaticobiliary duct are important parts of the digestive system, involving food delivery, digestion, absorption, and excretion. Gastrointestinal diseases may cause strictures of the esophagus, intestinal tract, biliary tract, and pancreas, and some patients may suffer from malignant Gastrointestinal blockage caused by tumors. When these cancer patients no longer have the option of surgery and cannot tolerate chemotherapy or radiation, stent implantation has been used as an excellent modality of palliative care because it can provide patients with immediate symptom relief. In addition, temporary internal stenting may be beneficial in the treatment of several of these GI disorders and to help ensure the flow of bile, pancreatic juice, or fluid into the GI tract after endoscopic or surgical procedures. Implantation of stents has great advantages and attractiveness in relieving pain and promoting tissue repair for patients with digestive tract stenosis.

Endogenous electric fields and electrical stimulation have been widely proven to promote cell growth and wound healing. Conductive polymers such as polyaniline, polypyrrole, and polythiophene have been proven to be non-cytotoxic and can be used in tissue engineering. After adding these conductive additives, endogenous electric field or external electrical stimulation can be used to promote tissue repair. Heparan sulfate proteoglycans have a wide range of functions in mammals, and play an important role in maintaining the homeostasis of the human system. They have a wide range of regulatory functions in the digestive system, respiratory system, endocrine system, nervous system, urinary system, immune system and circulatory system of the human body. Polydopamine has excellent biocompatibility and has been widely used in the surface modification of biomaterials. The reason is that the groups of polydopamine are mainly phenolic hydroxyl groups, amine groups, and imine groups, which can further improve the surface properties of biomaterials by covalently bonding target molecules. Under the bonding action of dopamine, the prepared nanofibrous material can transport and carry the conduction aid and heparan sulfate proteoglycan.

Polycaprolactone is a hydrophobic semi-crystalline polymer whose crystallinity tends to decrease with increasing molecular weight. The good solubility, low melting point, and excellent blending compatibility of polycaprolactone have inspired extensive research on its potential applications in the biomedical field. Polycaprolactone was once popular during the upsurge of absorbable polymers in the 1970s and 1980s, but the degradation cycle of polycaprolactone for more than 1 year was quickly replaced by polyglycolide and polypropylene with shorter degradation cycles. Lactide and its copolymers. After being forgotten for nearly 20 years, the advent of tissue engineering and the beginning of a resurgence of interest in polycaprolactones, which were recognized to possess superior rheological and viscoelastic properties over many absorbable polymer counterparts, This makes it easy to fabricate and operate into a wide range of scaffolding. Additionally, the fact that many drug delivery devices manufactured using polycaprolactone are already FDA-approved and CE-mark registered provides a faster path to market.

In the field of biomaterials, polycaprolactone has good biocompatibility, excellent rheological and viscoelastic properties, relatively stable chemical properties, simple processing methods, and can be designed and prepared according to actual applications. Polycaprolactone is dissolved in an organic solvent, and nanofibers can be obtained after electrospinning, which has a large specific surface area. The surface conductive layer is obtained by in-situ oxidation polymerization, and then soaked in heparan sulfate proteoglycan solution. When the drug is used, dopamine is used to realize the function of transportation and loading. Surface modification, polycaprolactone nanofiber materials loaded with drugs can improve the adhesion of nutrients such as proteins on the surface of the material, increase biocompatibility, reduce inflammation and cytotoxicity, and have excellent performance in promoting cell growth and tissue repair .

Contents of the invention

The invention provides a nanofibrous material with polycaprolactone as a matrix and a preparation method thereof, aiming at improving the functionality of polycaprolactone and its ability to have different acid-base environments in the pancreaticobiliary tract, gastrointestinal tract, etc. Stability, after surface modification, the polycaprolactone nanofiber material loaded with drugs can improve the adhesion of nutrients such as protein on the surface of the material, increase biocompatibility, reduce inflammation and cytotoxicity, and promote cell growth and Excellent in tissue repair

In order to achieve the above object, the utility model adopts the following scheme:

A method for preparing a nanofibrous material based on polycaprolactone, characterized in that it comprises the following steps:

S1: mixing and dissolving the polycaprolactone masterbatch with an organic solvent to obtain a slurry;

S2: preparing the obtained slurry into a polycaprolactone fiber film by electrospinning;

S3: After drying the polycaprolactone fiber film, carry out surface modification and drug loading, and then dry and sterilize to obtain the nanofibrous material.

Further, the nanofiber material is composed of polycaprolactone fiber film, modified material, and drug-loaded.

Further, the polycaprolactone fiber film is obtained by dissolving the polycaprolactone masterbatch in an organic solvent according to the mass fraction (10%-30%), and then preparing by electrospinning and drying.

Further, the organic solvent is mainly composed of two solvents A and B in a mass ratio of 7:3, A is one of methylene chloride and ethylene dichloride, and B is N,N-dimethylmethane One of amides and N,N-dimethylacetamide.

Further, the surface modification is performed by soaking the prepared polycaprolactone fiber film in a dopamine solution at a temperature of 15-45°C for 10-48 hours, and placing it at a temperature of -15-5°C after soaking. In the inorganic acid solution containing oxidant and conductive polymer for 10~48h.

Further, in the dopamine solution, the solute is dopamine, the solvent is Tris-HCl buffer or phosphate buffer, and the concentration of the dopamine is 0.01-100 mg/mL.

Further, the inorganic acid solution containing an oxidant and a conductive polymer, wherein the oxidant is one of ammonium persulfate and ferric chloride, the conductive polymer is one of aniline, pyrrole, and thiophene, and the inorganic acid is One of hydrochloric acid, sulfuric acid, perchloric acid, according to the molar ratio of oxidant: conductive polymer: inorganic acid = (0.005 ~ 0.1): (0.005 ~ 0.1): (0.1 ~ 10) add deionized water to make up 1L into the inorganic acid solution.

Further, the drug loading mainly uses the soaking method, the drug used is heparan sulfate proteoglycan, ultrapure water is used as the solvent, the drug content is 1-100 mg/ml, and the fiber film after the surface modification is completed Soak in heparan sulfate proteoglycan aqueous solution at 30-40°C for 10-48 hours.

Further, the sterilization is any one of ultraviolet sterilization, dry heat sterilization and high pressure steam sterilization.

Further, the method for preparing nanofibrous materials based on polycaprolactone can be used as a culture and proliferation carrier for cells such as vascular endothelial cells (HUVEC), gastric parietal cells, gastric mucosal cells, and pancreatic islet cells in vitro, and can also be used to prepare Stents are used for tissue repair related to the digestive system such as the gastrointestinal tract and pancreaticobiliary duct.

In summary, the present invention has the beneficial effects relative to the prior art as follows:

The polycaprolactone-based nanofiber material provided by the invention has relatively stable performance and chemical stability in vivo, has good biocompatibility, and has excellent performance in promoting tissue repair and drug loading.

Description of drawings

Figure 1 is a flow chart for the preparation of polycaprolactone nanofibers.

Figure 2 is a comparison chart between the control group and the experimental group.

Detailed ways

Various exemplary embodiments of the present invention will now be described in detail. The detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features and embodiments of the present invention.

It should be understood that the terminology described in the present invention is only used to describe specific embodiments, and is not used to limit the present invention. In addition, regarding the numerical ranges in the present invention, it should be understood that each intermediate value between the upper limit and the lower limit of the range is also specifically disclosed. Any stated value or intervening value in a stated range, and each smaller range between any other stated value or intervening value in a stated range, is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included or excluded from the range.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although only the preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and/or materials in connection with which the documents are described. In case of conflict with any incorporated document, the contents of this specification control.

It will be apparent to those skilled in the art that various modifications and changes can be made in the specific embodiments of the present invention described herein without departing from the scope or spirit of the present invention. Other embodiments will be apparent to the skilled person from the description of the present invention. The description and examples of the invention are illustrative only.

As used herein, "comprising", "comprising", "having", "comprising" and so on are all open terms, meaning including but not limited to.

In the present invention, all raw materials are conventional commercially available products.

Example 1

Take 3g of polycaprolactone plastic masterbatch, add it into a mixed solution containing 18.9g of methylene chloride and 8.1g of N,N-dimethylformamide, and stir evenly at 50°C for 4h. The obtained slurry was filled into a 5ml syringe, the spinning voltage was set to 20KV, the receiving distance was 15cm, and the injection speed was 0.8ml/h. After the electrospinning is completed, put it into a vacuum drying oven at 40°C for 24 hours.

Cut the dried polycaprolactone nanofiber film into 10mm discs with a cutting machine. After ultrasonic cleaning, immerse the discs in a 2 mg/ml dopamine solution for 24 hours, and take them out with deionized water. Rinse three times and dry. Submerge the dried disc in 100ml of inorganic acid solution for 24 hours. The composition and molar ratio of the solution are: ammonium persulfate: aniline: hydrochloric acid = (0.01mol) (0.01mol) (1mol) plus deionized water to volume 1L into solution. The fiber film after surface modification was soaked in 10 mg/ml heparan sulfate proteoglycan aqueous solution at 35° C. for 24 hours. After drying, the nanofibrous material with polycaprolactone as the matrix was obtained after ultraviolet sterilization for 30 minutes.

Put the polycaprolactone disc into a forty-eight-well plate, put a prepared polycaprolactone disc into each hole, and inoculate 8000 cells in each hole, the cells are vascular endothelial cells, and put them into an incubator for cultivation 1, 3, and 7 days, and a blank polycaprolactone disc was used as a control. The cell viability was evaluated by MTT assay.

The cell viability of the control group on day 1, day 3 and day 7 was: 0.30, 0.41, 0.45.

The cell viability of the experimental group on day 1, day 3, and day 7 was: 0.70, 0.82, and 0.93.

Compared with the control group, the cell activity of the experimental group was greatly improved, and the biocompatibility was effectively improved.

Example 2

Take 3g of polycaprolactone plastic masterbatch, add it into a mixed solution containing 11.9g of methylene chloride and 5.1g of N,N-dimethylformamide, and stir evenly at 45°C for 5h. The obtained slurry was filled into a 5ml syringe, the spinning voltage was set to 20KV, the receiving distance was 15cm, and the injection speed was 0.8ml/h. After the electrospinning is completed, put it into a vacuum drying oven at 45°C for 20 hours.

Cut the dried polycaprolactone nanofiber film into 10mm discs with a cutting machine, after ultrasonic cleaning, immerse the discs in 4mg/ml dopamine solution for 18h, take out with deionized water Rinse three times and dry. Submerge the dried disc in 100ml of inorganic acid solution for 12h, the solution composition and molar ratio are: ammonium persulfate: pyrrole: perchloric acid = (0.05mol) (0.05mol) (5mol) plus deionized water to volume 1L dubbed solution. The fiber film after surface modification was soaked in 20 mg/ml heparan sulfate proteoglycan aqueous solution at 40° C. for 18 hours. After drying, the nanofibrous material with polycaprolactone as the matrix was obtained after ultraviolet sterilization for 30 minutes.

Put the polycaprolactone disk into a forty-eight-well plate, put one piece of the prepared polycaprolactone disk into each well, then add 50 μL Matrigel to each well, and then put the multiwell plate into a 37°C incubator for 4h , to cure the matrix glue into a glue. Afterwards, 8000 cells were inoculated in each well, and the cells were vascular endothelial cells, cultured in an incubator for 18 hours, stained with calcein (AM), observed vascularization with an inverted fluorescence microscope, and a blank polycaprolactone disk was used as a control.

Compared with the control group, the activity of blood vessels in the experimental group was greatly improved, which helped to promote tissue repair.

Example 3

Take 6g of polycaprolactone plastic masterbatch, add it into a mixed solution containing 9.8g of dichloroethane and 4.2g of N,N-dimethylacetamide, and stir evenly at 45°C for 4h. The obtained slurry was filled into a 5ml syringe, the spinning voltage was set to 20KV, the receiving distance was 15cm, and the injection speed was 0.8ml/h. After the electrospinning is completed, put it into a vacuum drying oven at 40°C for 24 hours.

Cut the dried polycaprolactone nanofiber film into 10mm discs with a cutting machine. After ultrasonic cleaning, immerse the discs in a 3 mg/ml dopamine solution for 24 hours, and take them out with deionized water. Rinse three times and dry. Submerge the dried disc in 100ml of inorganic acid solution for 12 hours. The composition and molar ratio of the solution are: ferric chloride: thiophene: sulfuric acid = (0.02mol) (0.02mol) (2mol) plus deionized water to make up to 1L. into solution. The fiber film after surface modification was soaked in 50 mg/ml heparan sulfate proteoglycan aqueous solution at 30° C. for 12 hours. After drying, the nanofibrous material with polycaprolactone as the matrix was obtained after ultraviolet sterilization for 30 minutes.

The prepared polycaprolactone nanofiber discs were used to measure the degradability of the material under different pH values by the mass loss method. After drying, the mass before the experiment was recorded, and the discs were soaked in pH = 8.5, =1 solution, incubate at 37°C for 21 days, change the solution every five days, dry at 40°C and weigh after constant weight. And a blank polycaprolactone disc was used as a control.

Mass loss in the control group: 20% (PH=8.5), 15% (PH=7.4), 14% (PH=1).

The mass loss of the experimental group: 11% (PH=8.5), 14% (PH=7.4), 8% (PH=1).

Compared with the control group, the stability of the experimental group was improved in the acidic or alkaline environment.

Claims (10)

1. a kind of nanofiber material preparation method taking polycaprolactone as matrix, is characterized in that, may further comprise the steps: S1: mixing and dissolving the polycaprolactone masterbatch with an organic solvent to obtain a slurry; S2: preparing the obtained slurry into a polycaprolactone fiber film by electrospinning; S3: After drying the polycaprolactone fiber film, carry out surface modification and drug loading, and then dry and sterilize to obtain the nanofibrous material. 2. A method for preparing a nanofiber material based on polycaprolactone according to claim 1, wherein the nanofiber material is composed of a polycaprolactone fiber film, a modified material, and a loaded drug. 3. a kind of nanofiber material preparation method taking polycaprolactone as matrix according to claim 1, is characterized in that, described polycaprolactone fiber film is by massfraction (10 %～30%) is dissolved in an organic solvent, then prepared by electrospinning, and obtained after drying. 4. a kind of nanofiber material preparation method taking polycaprolactone as matrix according to claim 3, is characterized in that, described organic solvent is mainly made up of A and B two kinds of solvents by the mass ratio of 7:3, A is one of dichloromethane and dichloroethane, and B is one of N,N-dimethylformamide and N,N-dimethylacetamide. 5. A kind of nanofibrous material preparation method taking polycaprolactone as matrix according to claim 1, is characterized in that, described surface modification is by making the polycaprolactone fiber film that makes at 15～45 Soak in the dopamine solution for 10 to 48 hours at a temperature of 10°C, and place it in an inorganic acid solution containing an oxidant and a conductive polymer at a temperature of -15 to 5°C for 10 to 48 hours after soaking. 6. a kind of nanofiber material preparation method taking polycaprolactone as matrix according to claim 5, is characterized in that, described dopamine solution, solute is dopamine, and solvent is Tris-HCl buffer or phosphate buffer, The concentration of the dopamine is 0.01-100 mg/mL. 7. a kind of nanofiber material preparation method taking polycaprolactone as matrix according to claim 6, is characterized in that, described inorganic acid solution containing oxidizing agent and conductive macromolecule, wherein oxidizing agent is ammonium persulfate, chlorine One of iron oxides, conductive polymer is one of aniline, pyrrole, and thiophene, and inorganic acid is one of hydrochloric acid, sulfuric acid, and perchloric acid, according to the molar ratio of oxidant: conductive polymer: inorganic acid =(0.005-0.1): (0.005-0.1): (0.1-10) add deionized water to a constant volume of 1 L to prepare the inorganic acid solution. 8. a kind of nanofiber material preparation method taking polycaprolactone as matrix according to claim 1, is characterized in that, what described drug loading mainly used is soaking method, and used medicine is heparan sulfate proteoglycan , using ultrapure water as a solvent, with a drug content of 1-100 mg/ml, soaking the surface-modified fiber film in an aqueous solution of heparan sulfate proteoglycan at 30-40°C for 10-48 hours. 9. A nanofibrous material based on polycaprolactone, characterized in that it is prepared according to the preparation method described in any one of claims 1-8. 10. a kind of application according to claim 9 taking polycaprolactone as the nanofiber material of matrix, it is characterized in that: described nanofiber material can be used as the culture proliferation carrier of cell in vitro, such as vascular endothelial cell ( HUVEC), gastric parietal cells, gastric mucosal cells, and pancreatic islets can also be prepared into scaffolds for tissue repair related to the digestive system, such as gastrointestinal tract, pancreas and gallbladder.

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