Nano-fiber extracorporeal membrane lung oxygenation membrane based on bacterial cellulose and preparation method of nano-fiber extracorporeal membrane lung oxygenation membrane

Abstract

The invention relates to a bacterial cellulose-based nanofiber in-vitro membrane lung oxygenation membrane and a preparation method thereof. The bacterial cellulose-based nanofiber in-vitro membrane lung oxygenation membrane is prepared by a hydrophobic modification method or a pore size increasing and mechanical enhancing method. The nanofiber extracorporeal membrane lung oxygenation membrane obtained by the invention has good mechanical properties, blood compatibility and oxygen transmission performance, and can strive for treatment time for severe lung failure patients; the operation method is simple, conditions are mild, and the method has great application potential in the field of respiratory support.

Description

Technical Field

[0001] The present invention belongs to the technical field of biocompatible material preparation, and particularly relates to a nanofiber extracorporeal membrane oxygenation membrane based on bacterial cellulose and a preparation method thereof. Background Art

[0002] As a key life support technology, extracorporeal membrane oxygenation (ECMO) plays a crucial role in the treatment of patients with cardiopulmonary failure and major cardiac surgeries. However, the high cost of ECMO equipment has become a major obstacle to its widespread application. At present, the domestic production of ECMO equipment has not been achieved, and domestic enterprises can only provide auxiliary consumables such as puncture catheters and connecting catheters. For key components such as the membrane lung, which plays a core role in the operation of the equipment, it is completely dependent on imported channels.

[0003] As an in vitro alternative device for the functions of the human native lungs, the core function of the artificial lung membrane is to achieve efficient oxygen and carbon dioxide gas exchange. Currently, materials such as polypropylene, poly(4-methyl-1-pentene), and silicone are commonly used in commercial artificial lung membrane products. However, these traditional materials generally have limited gas permeation efficiency, are prone to platelet activation and coagulation cascade reactions, leading to thrombosis and other problems. In addition, during long-term clinical use, the performance stability of these materials is insufficient and difficult to meet the growing clinical needs. Therefore, the development of new high-performance membrane lung materials is of great significance for promoting the widespread application and in-depth development of ECMO technology in clinical practice and is also a key link in the localization of ECMO equipment.

[0004] Bacterial NanoCellulose (BNC), also known as bacterial cellulose (BC), is a nanoscale cellulose material formed by bacteria gradually assembling glucan chains through a series of complex and precise enzymatic reactions. BNC has been widely used in blood-contact biomedical fields such as wound dressings, hemostatic materials, and artificial blood vessels due to its excellent blood compatibility. This material has extremely high specific surface area and porosity, so it has great application potential in the field of gas exchange. However, in the actual application scenarios of artificial lung membranes, BNC materials expose a series of problems that need to be solved. First, the high hydrophilicity of BNC makes its surface easily adsorbed to form a layer of water film. This phenomenon seriously hinders the mass transfer process of gas in the membrane material and reduces the gas exchange efficiency. Second, the bacterial culture process has a significant impact on the performance of BNC materials. Prolonging the bacterial culture time will lead to the densification of the fiber network structure, which will cause a significant decrease in the oxygen permeability of the material; on the contrary, shortening the culture cycle can increase the fiber pore size and promote oxygen transmission to a certain extent, but it will weaken the overall mechanical strength of the material and affect the durability and reliability of the material in practical applications. These problems have seriously restricted the large-scale application and promotion of bacterial cellulose nanofiber membranes in the field of ECMO.

[0005] Among the existing preparation processes for gas permeable membranes, the casting method is more commonly used. However, the gas permeable membranes prepared by this method still have room for improvement in terms of oxygen permeability and blood compatibility, and the preparation process is cumbersome, so the stability of product quality is difficult to guarantee. At present, no relevant research results and technical solutions for applying bacterial cellulose nanofiber membranes to extracorporeal membrane oxygenation membranes have been found. Summary of the invention

[0006] The technical problem to be solved by the present invention is to provide a nanofiber extracorporeal membrane oxygenation membrane based on bacterial cellulose and a preparation method thereof. The preparation method has the remarkable characteristics of simple operation and strong feasibility, and can effectively retain the nanofiber network structure unique to bacterial cellulose nanofibers. By means of hydrophobic modification or increasing the pore size and enhancing the mechanical properties, the prepared nanofiber extracorporeal membrane oxygenation membrane can further improve the oxygen transfer efficiency. In addition, the material exhibits outstanding advantages such as excellent mechanical properties, excellent blood compatibility and environmental degradation.

[0007] The invention provides a nanofiber extracorporeal membrane oxygenation membrane based on bacterial cellulose, which is prepared by a hydrophobic modification method or a pore size enlargement and mechanical enhancement method.

[0008] The present invention also provides a method for preparing a nanofiber extracorporeal membrane oxygenation membrane based on bacterial cellulose, comprising the following steps:

[0009] (1) Inoculate *Acetobacter xylinum* into a culture medium and perform static culture;

[0010] (2) Purify the obtained cellulose membrane with sodium hydroxide solution and water to obtain a purified bacterial cellulose nanofiber membrane;

[0011] (3) Perform hydrophobic modification on the bacterial cellulose nanofiber membrane by physical modification method or chemical modification method, and finally obtain a nanofiber extracorporeal membrane oxygenation membrane by vacuum drying.

[0012] Preferably, the *Acetobacter xylinum* strain in the step (1) is CGMCC 27910, ATCC 23770 or ATCC 23767.

[0013] Preferably, the static culture temperature in the step (1) is 25 - 40 °C.

[0014] Preferably, the static culture days in the step (1) are 5 - 9 days. If the static culture time is short, the nanofiber extracorporeal membrane oxygenation membrane has a loose network and high oxygen permeability. If the static culture time is long, the nanofiber extracorporeal membrane oxygenation membrane has a dense network and low oxygen permeability.

[0015] Preferably, the concentration of the sodium hydroxide solution in the step (2) is 0.5% - 2% (w / v).

[0016] Preferably, the purification temperature in the step (2) is 50 - 90 °C.

[0017] Preferably, the physical modification method in the step (3) includes physical adsorption method or physical spraying method; the chemical modification method includes silanization method, alkanoylation method, esterification method or graft polymerization method.

[0018] Preferably, the reagents used in the physical adsorption method include one or more of cationic surfactants, quaternary ammonium salts, and diblock copolymer dispersants; the reagents used in the physical spraying method include commercial waterproof sprays.

[0019] More preferably, the cationic surfactant is one or more of dodecyl trimethyl ammonium chloride, tetradecyl trimethyl ammonium chloride, hexadecyl trimethyl ammonium chloride or hexadecyl trimethyl ammonium bromide.

[0020] More preferably, the quaternary ammonium salt is one or more of stearyl trimethyl ammonium chloride, benzyl trimethyl ammonium chloride, diallyl dimethyl ammonium chloride or quaternary alkyl ammonium.

[0021] More preferably, the diblock copolymer dispersant is one or more of 2-hydroxyethyl polymethacrylate or dodecyl polymethacrylate diblock copolymer.

[0022] More preferably, the commercial waterproof spray is one or more of Gear Aid ReviveX Nubuck, 3M Scotchgard, TRG, or MOOTAA.

[0023] Preferably, the silanization method uses a silanization reagent; the acylation method uses an acylating reagent; the esterification method uses an esterifying reagent; the graft polymer method uses a polymer side chain.

[0024] More preferably, the silanization reagent is one or more of trimethylchlorosilane, trichloromethylsilane, methyltrimethoxysilane, 3-methacryloxypropyltrimethoxysilane, 1H,1H,2H,2H-perfluorooctyltriethoxysilane, or γ-aminopropyltriethoxysilane.

[0025] More preferably, the acylating reagent is one or more of acetyl chloride, butyryl chloride, or 10-undecenoyl chloride.

[0026] More preferably, the esterifying reagent is one or more of lactic acid or acetic acid.

[0027] More preferably, the polymer side chain is one or more of polycaprolactone, polymethyl methacrylate, or poly(methyl acrylate).

[0028] Preferably, the vacuum drying parameters in step (3) are: vacuum drying temperature 30 - 100 °C, vacuum drying time 6 - 24 h.

[0029] The present invention also provides a second method for preparing a nanofiber extracorporeal membrane oxygenation membrane based on bacterial cellulose, comprising the following steps:

[0030] (1) Dissolve the synthetic polymer in a solvent to prepare a spinning solution; ultrasonically remove the bubbles from the spinning solution and obtain electrospun fibers using electrospinning technology.

[0031] (2) Add a medium containing Acetobacter xylinum to a petri dish and statically culture to obtain a bacterial cellulose nanofiber membrane.

[0032] (3) Sterilize the electrospun fibers and place them on the bacterial cellulose nanofiber membrane, and supplement the medium with a medium containing Acetobacter xylinum, and statically culture.

[0033] (4) Purify the cellulose membrane obtained by culturing in step (3) with sodium hydroxide solution and water to finally obtain a nanofiber extracorporeal membrane oxygenation membrane.

[0034] Preferably, the synthetic polymer in step (1) includes one or more of cellulose acetate (CA), polyvinylidene fluoride (PVDF), polyether ketone, or polysulfone.

[0035] Preferably, the solvent in step (1) is one or more of N,N-dimethylformamide, acetone, chloroform, or hexafluoroisopropanol. When there are two solvents, the volume ratio is 1:9 - 9:1.

[0036] Preferably, the mass fraction of the spinning solution in step (1) is 8 - 16%.

[0037] Preferably, the ultrasonic frequency in step (1) is 50 - 100 Hz, and the ultrasonic time is 1 - 5 min.

[0038] Preferably, the electrospinning conditions in step (1) are 10 - 20 kV, the receiving distance is 10 - 15 cm, the roller rotation speed is 10 - 30 r / min, the injection rate is 0.5 - 2 mL / h, and the spinning time is 0.5 - 4 h.

[0039] Preferably, the Acetobacter xylinum strain in step (2) is CGMCC 27910, ATCC 23770, or ATCC 23767.

[0040] Preferably, the static culture temperature in steps (2) and (3) is 25 - 40 °C.

[0041] Preferably, the static culture days in steps (2) and (3) are 1 - 3 days. If the static culture time is short, the nanofiber extracorporeal membrane oxygenation membrane network is loose and has high oxygen permeability. If the static culture time is long, the nanofiber extracorporeal membrane oxygenation membrane network is dense and has low oxygen permeability.

[0042] Preferably, the sterilization conditions in step (3) are sterilization at 115 °C for 30 min, or sterilization at 121 °C for 20 min.

[0043] Preferably, the volume of the added liquid in step (3) is 5 - 15 mL.

[0044] Preferably, the concentration of the sodium hydroxide solution in step (4) is 0.5% - 2% (w / v).

[0045] Preferably, the purification temperature in step (4) is 50 - 90 °C.

[0046] With the help of the biosynthesis process of Acetobacter xylinum, the present invention successfully prepares a bacterial cellulose nanofiber membrane with high porosity and a unique three-dimensional network structure, and this structural characteristic makes it an ideal oxygen transport carrier. Given that the gas transmission rate in air is much higher than that in water, the present invention performs hydrophobic modification on the bacterial cellulose nanofiber membrane to reduce the resistance during gas transmission, thereby significantly improving the oxygen transmission efficiency. At the same time, considering that increasing the pore size can accelerate the oxygen transmission rate, by appropriately shortening the culture time and introducing a synthetic polymer electrospun membrane as a mechanical support structure, the oxygen transmission rate is further increased. The finally obtained nanofiber extracorporeal membrane oxygenation membrane has high oxygen permeability, excellent mechanical properties and good biocompatibility, and the whole preparation process is simple, easy to operate and highly repeatable, without damaging the fiber network structure of the membrane itself.

[0047] Beneficial effects

[0048] (1) The present invention performs hydrophobic modification on the bacterial cellulose nanofiber membrane (which may also include commercially available bacterial cellulose membranes) to reduce the resistance during gas transmission, thereby significantly improving the oxygen transmission efficiency. At the same time, considering that increasing the pore size can accelerate the oxygen transmission rate, by appropriately shortening the culture time and introducing a synthetic polymer electrospun membrane as a mechanical support structure, the oxygen transmission rate is further increased.

[0049] (2) Improving oxygen transmission rate and application potential: The nanofiber extracorporeal membrane oxygenation membrane prepared by the present invention significantly increases the oxygen transmission rate of the bacterial cellulose nanofiber membrane. This membrane shows potential application value in the treatment of heart failure, lung injury, etc., and is expected to be applied to the extracorporeal life support system, providing a new technical means for the clinical treatment of related diseases.

[0050] (3) Retaining the structural and comprehensive performance advantages: The preparation method of the present invention effectively retains the original nanofiber network structure of the bacterial cellulose nanofiber membrane, making the membrane not only have good mechanical properties, but also excellent blood compatibility and environmental degradability, which can meet the requirements of long-term use and lay a solid foundation for its wide application in the biomedical field. The preparation conditions of the present invention are simple, the reaction efficiency is high, and the operability is strong.

[0051] (4) Simple and efficient preparation conditions: The preparation process of the present invention has simple conditions, high reaction efficiency, and strong operability, and is easy to achieve large-scale preparation in actual production, which is conducive to promoting the industrial application and popularization of this technology. Description of the Drawings

[0052] Figure 1 Field emission scanning electron microscope images of the BNC membrane and the TBNC extracorporeal membrane oxygenation membrane before and after oxygen transmission.

[0053] Figure 2 Axial tensile mechanical property diagrams of BNC membranes and TBNC extracorporeal membrane oxygenation membranes.

[0054] Figure 3 Electron micrographs of platelet and red blood cell adhesion of BNC membranes and TBNC extracorporeal membrane oxygenation membranes.

[0055] Figure 4 Diagrams of oxygen transfer rates in blood of BNC membranes and TBNC extracorporeal membrane oxygenation membranes.

[0056] Figure 5 Field emission scanning electron micrographs of BNC membranes, PVDF / BNC extracorporeal membrane oxygenation membranes, and PVDF-CA / BNC extracorporeal membrane oxygenation membranes.

[0057] Figure 6 Burst pressure diagrams of BNC membranes, PVDF / BNC extracorporeal membrane oxygenation membranes, and PVDF-CA / BNC extracorporeal membrane oxygenation membranes.

[0058] Figure 7 Electron micrographs of platelet and red blood cell adhesion of BNC membranes, PVDF / BNC extracorporeal membrane oxygenation membranes, and PVDF-CA / BNC extracorporeal membrane oxygenation membranes.

[0059] Figure 8 Diagrams of oxygen transfer rates in water of BNC membranes, PVDF / BNC extracorporeal membrane oxygenation membranes, and PVDF-CA / BNC extracorporeal membrane oxygenation membranes.

[0060] Figure 9 Process flow chart for the preparation of nanofiber extracorporeal membrane oxygenation membranes based on bacterial cellulose. Specific implementation manners

[0061] The present invention will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and not to limit the scope of the present invention. In addition, it should be understood that after reading the content taught by 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 this application.

[0062] Example 1

[0063] (1) Inoculate the Acetobacter xylinum ATCC 23770 strain into a liquid medium and incubate it statically at a constant temperature of 30°C for 7 days to obtain a bacterial cellulose nanofiber membrane.

[0064] (2) At 80 °C, immerse the bacterial cellulose nanofiber membrane in a 1% (w / v) sodium hydroxide solution for 4 h, then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0065] (3) Adsorb cetyltrimethylammonium bromide on the purified bacterial cellulose nanofiber membrane for 1 h, then wash it repeatedly with ethanol, and finally dry it in vacuum at 60 °C for 24 h to obtain a bacterial cellulose nanofiber extracorporeal membrane oxygenation membrane.

[0066] Example 2

[0067] (1) Inoculate the Komagataeibacter xylinus CGMCC 27910 strain into a liquid medium and incubate it statically at a constant temperature of 30 °C for 7 days to obtain a bacterial cellulose nanofiber membrane.

[0068] (2) At 80 °C, immerse the bacterial cellulose nanofiber membrane in a 1% (w / v) sodium hydroxide solution for 4 h, then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0069] (3) Adsorb benzyltrimethylammonium chloride on the purified bacterial cellulose nanofiber membrane for 1 h, then wash it repeatedly with ethanol, and finally dry it in vacuum at 60 °C for 24 h to obtain a bacterial cellulose nanofiber extracorporeal membrane oxygenation membrane.

[0070] Example 3

[0071] (1) Inoculate the Komagataeibacter CGMCC 27910 strain into a liquid medium and incubate it statically at a constant temperature of 30 °C for 7 days to obtain a bacterial cellulose nanofiber membrane.

[0072] (2) At 80 °C, immerse the bacterial cellulose nanofiber membrane in a 1% (w / v) sodium hydroxide solution for 4 h, then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0073] (3) Adsorb 2-hydroxyethyl methacrylate on the purified bacterial cellulose nanofiber membrane for 1 h, then wash it repeatedly with ethanol, and finally dry it in vacuum at 60 °C for 24 h to obtain a bacterial cellulose nanofiber extracorporeal membrane oxygenation membrane.

[0074] Example 4

[0075] (1) Inoculate the Komagataeibacter ATCC 23770 strain into a liquid medium and incubate it statically at a constant temperature of 35 °C for 5 days to obtain a bacterial cellulose nanofiber membrane.

[0076] (2) At 90 °C, immerse the bacterial cellulose nanofiber membrane in a 2% (w / v) sodium hydroxide solution for 4 h, and then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0077] (3) Spray the commercial waterproof spray GearAid ReviveX Nubuck onto the bacterial cellulose nanofiber membrane from a distance of 15 cm, and finally dry it in a vacuum at 30 °C for 24 h to obtain the extracorporeal membrane oxygenation membrane of bacterial cellulose nanofibers.

[0078] Example 5

[0079] (1) Inoculate the Acetobacter xylinum ATCC 23770 strain into a liquid medium and incubate it statically at a constant temperature of 35 °C for 5 days to obtain a bacterial cellulose nanofiber membrane.

[0080] (2) At 90 °C, immerse the bacterial cellulose nanofiber membrane in a 2% (w / v) sodium hydroxide solution for 4 h, and then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0081] (3) Spray the commercial waterproof spray 3M Scotchgard onto the bacterial cellulose nanofiber membrane from a distance of 15 cm, and finally dry it in a vacuum at 30 °C for 24 h to obtain the extracorporeal membrane oxygenation membrane of bacterial cellulose nanofibers.

[0082] Example 6

[0083] (1) Inoculate the Acetobacter xylinum ATCC 23767 strain into a liquid medium and incubate it statically at a constant temperature of 30 °C for 5 days to obtain a bacterial cellulose nanofiber membrane.

[0084] (2) At 80 °C, immerse the bacterial cellulose nanofiber membrane in a 0.5% (w / v) sodium hydroxide solution for 4 h, and then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0085] (3) Place the glass dish containing the bacterial cellulose nanofiber membrane and the beaker containing 0.5 mL of trimethylchlorosilane in a desiccator simultaneously, and carry out chemical deposition at 30 °C for 20 min. After taking it out, dry the membrane in a vacuum at 40 °C for 24 h to obtain the extracorporeal membrane oxygenation membrane of bacterial cellulose nanofibers.

[0086] Example 7

[0087] (1) Inoculate the Acetobacter xylinum ATCC 23767 strain into a liquid medium and incubate it statically at a constant temperature of 30 °C for 5 days to obtain a bacterial cellulose nanofiber membrane.

[0088] (2) At 80 °C, immerse the bacterial cellulose nanofiber membrane in a 0.5% (w / v) sodium hydroxide solution for 4 h, then immerse it in deionized water for 4 h. Repeat the above operations until the membrane becomes white and the solution is neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0089] (3) Mix and react the bacterial cellulose nanofiber membrane and an ethanol solution of γ-aminopropyltriethoxysilane at room temperature for 1 h, then wash it repeatedly with ethanol. After taking it out, dry the membrane in a vacuum at 40 °C for 24 h to obtain a bacterial cellulose nanofiber extracorporeal membrane oxygenation membrane.

[0090] Example 8

[0091] (1) Inoculate the Acetobacter xylinum ATCC 23770 strain into a liquid medium and incubate it statically at a constant temperature of 40 °C for 5 days to obtain a bacterial cellulose nanofiber membrane;

[0092] (2) At 70 °C, immerse the bacterial cellulose nanofiber membrane in a 2% (w / v) sodium hydroxide solution for 4 h, then immerse it in deionized water for 4 h. Repeat the above operations until the membrane becomes white and the solution is neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0093] (3) Mix and react the bacterial cellulose nanofiber membrane and butyryl chloride at room temperature for 24 h, then wash it repeatedly with ethanol. After taking it out, dry the membrane in a vacuum at 30 °C for 24 h to obtain a bacterial cellulose nanofiber extracorporeal membrane oxygenation membrane.

[0094] Example 9

[0095] (1) Inoculate the Acetobacter xylinum ATCC 23770 strain into a liquid medium and incubate it statically at a constant temperature of 40 °C for 5 days to obtain a bacterial cellulose nanofiber membrane.

[0096] (2) At 70 °C, immerse the bacterial cellulose nanofiber membrane in a 2% (w / v) sodium hydroxide solution for 4 h, then immerse it in deionized water for 4 h. Repeat the above operations until the membrane becomes white and the solution is neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0097] (3) Mix and react the bacterial cellulose nanofiber membrane and acetyl chloride at room temperature for 24 h, then wash it repeatedly with ethanol. After taking it out, dry the membrane in a vacuum at 30 °C for 24 h to obtain a bacterial cellulose nanofiber extracorporeal membrane oxygenation membrane.

[0098] Example 10

[0099] (1) Inoculate the Acetobacter xylinum ATCC 23767 strain into a liquid medium and incubate it statically at a constant temperature of 30 °C for 7 days to obtain a bacterial cellulose nanofiber membrane.

[0100] (2) At 80 °C, immerse the bacterial cellulose nanofiber membrane in a 1% (w / v) sodium hydroxide solution for 4 h, then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0101] (3) Mix and react the bacterial cellulose nanofiber membrane with acetic anhydride, concentrated sulfuric acid, and glacial acetic acid at room temperature for 1 h, then wash it repeatedly with ethanol. After taking it out, dry the membrane in vacuo at 30 °C for 24 h to obtain a bacterial cellulose nanofiber extracorporeal membrane oxygenation membrane.

[0102] Example 11

[0103] (1) Inoculate the Acetobacter xylinum ATCC 23767 strain into a liquid medium and incubate it statically at a constant temperature of 30 °C for 7 days to obtain a bacterial cellulose nanofiber membrane.

[0104] (2) At 80 °C, immerse the bacterial cellulose nanofiber membrane in a 1% (w / v) sodium hydroxide solution for 4 h, then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0105] (3) Mix and react the bacterial cellulose nanofiber membrane with lactic acid at 100 °C for 24 h, then wash it repeatedly with ethanol. After taking it out, dry the membrane in vacuo at 30 °C for 24 h to obtain a bacterial cellulose nanofiber extracorporeal membrane oxygenation membrane.

[0106] Example 12

[0107] (1) Inoculate the Acetobacter xylinum ATCC 23767 strain into a liquid medium and incubate it statically at a constant temperature of 30 °C for 9 days to obtain a bacterial cellulose nanofiber membrane.

[0108] (2) At 80 °C, immerse the bacterial cellulose nanofiber membrane in a 2% (w / v) sodium hydroxide solution for 4 h, then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0109] (3) Mix and react the bacterial cellulose nanofiber membrane with polycaprolactone at 60 °C for 24 h, then wash it repeatedly with ethanol. After taking it out, dry the membrane in vacuo at 60 °C for 12 h to obtain a bacterial cellulose nanofiber extracorporeal membrane oxygenation membrane.

[0110] Example 13

[0111] (1) Inoculate the Acetobacter xylinum ATCC 23767 strain into a liquid medium and incubate it statically at a constant temperature of 30 °C for 9 days to obtain a bacterial cellulose nanofiber membrane.

[0112] (2) At 80 °C, immerse the bacterial cellulose nanofiber membrane in a 2% (w / v) sodium hydroxide solution for 4 h, and then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0113] (3) Mix the bacterial cellulose nanofiber membrane and polymethyl methacrylate at 60 °C and react for 24 h. Then wash it repeatedly with ethanol. After taking it out, dry the membrane in vacuum at 60 °C for 12 h to obtain an extracorporeal membrane oxygenation membrane of bacterial cellulose nanofiber.

[0114] Comparative Example 1

[0115] (1) Inoculate the Acetobacter xylinum CGMCC 27910 strain into a liquid medium and incubate it statically at 30 °C for 5 days to obtain a bacterial cellulose nanofiber membrane.

[0116] (2) At 80 °C, immerse the bacterial cellulose nanofiber membrane in a 0.5% (w / v) sodium hydroxide solution for 4 h, and then immerse it in deionized water for 4 h. Repeat the above operations until the membrane turns white and the solution becomes neutral to obtain a purified bacterial cellulose nanofiber membrane.

[0117] Example 14

[0118] (1) Prepare a mixed solvent of N,N-dimethylformamide and acetone (volume ratio 7:3). Dissolve 0.6 g of PVDF and 0.6 g of CA in 8.8 g of the mixed solvent to prepare a spinning solution with a mass fraction of 12% (w / w).

[0119] (3) Ultrasonic the spinning solution at 50 Hz for 1 min to remove bubbles. Then load the spinning solution into a syringe. Under the conditions of a voltage of 13 kV, a receiving distance of 15 cm, a roller rotation speed of 10 r / min, an injection rate of 1 mL / h, and a spinning time of 2 h, obtain electrospun fibers by electrospinning technology.

[0120] (4) Add a medium containing Acetobacter xylinum ATCC 23770 to a petri dish and incubate it statically at 30 °C for 1 day. Then sterilize the electrospun fibers at 115 °C for 30 min and place them on the bacterial cellulose nanofiber membrane, and supplement 10 mL of the medium containing Acetobacter xylinum ATCC 23770, and incubate it statically at 30 °C for 3 days.

[0121] (5) Wash the cultured cellulose membrane repeatedly with a 1% (w / v) sodium hydroxide solution and water at 80 °C under high temperature conditions to obtain a purified extracorporeal membrane oxygenation membrane of bacterial cellulose nanofiber.

[0122] Example 15

[0123] (1) Prepare a mixed solvent of N,N-dimethylformamide and acetone (volume ratio 7:3), and dissolve 1.2 g of PVDF in 8.8 g of the mixed solvent to prepare a spinning solution with a mass fraction of 12% (w / w).

[0124] (3) Ultrasonic the spinning solution at 50 Hz for 1 min to remove bubbles, then load the spinning solution into a syringe, and under the conditions of a voltage of 13 kV, a receiving distance of 15 cm, a roller rotation speed of 10 r / min, an injection rate of 1 mL / h, and a spinning time of 2 h, obtain electrospun fibers by electrospinning technology.

[0125] (4) Add a medium containing Gluconacetobacter xylinus ATCC 23770 to a petri dish and statically culture it at 30 °C for 1 day. Then sterilize the electrospun fibers at 115 °C for 30 min and place them on the bacterial cellulose nanofiber membrane, and supplement 10 mL of the medium containing Gluconacetobacter xylinus ATCC 23770, and statically culture it at 30 °C for 3 days.

[0126] (5) Wash the cultured cellulose membrane repeatedly with 1% (w / v) sodium hydroxide solution and water at 80 °C to obtain a purified bacterial cellulose nanofiber extracorporeal membrane oxygenation membrane.

[0127] Comparative Example 2

[0128] (1) Add a medium containing Gluconacetobacter xylinus ATCC 23770 to a petri dish and statically culture it at 30 °C for 4 days.

[0129] (2) Wash the cultured cellulose membrane repeatedly with 1% (w / v) sodium hydroxide solution and water at 80 °C to obtain a purified bacterial cellulose nanofiber membrane.

[0130] Perform tests on the morphology, mechanical strength, red blood cell and platelet adhesion, and oxygen permeability of the TBNC extracorporeal membrane oxygenation membrane prepared in Example 6 and the BNC membrane prepared in Comparative Example 1. Figure 1 It can be seen that after oxygen transmission, some of the fibers of the BNC membrane are bonded, affecting the permeability of the network pores. While the fiber structure of the TBNC extracorporeal membrane oxygenation membrane has not changed significantly and still maintains the original pores intact. Figure 2 It can be seen that the prepared TBNC extracorporeal membrane oxygenation membrane has excellent mechanical properties, with a Young's modulus of 1.09 MPa, an elongation at break of 45.15%, and a tensile strength of 0.47 MPa. Figure 3 It can be seen that the prepared TBNC extracorporeal membrane oxygenation membrane has good blood compatibility, and there is no obvious platelet activation and red blood cell deformation. Figure 4It can be seen that the prepared TBNC extracorporeal membrane oxygenation membrane has strong blood oxygen transmission performance, and the oxygen transmission rate in deoxygenated arterial blood reaches 135 mL·m -2 ·min -1 , and the oxygen transmission rate in deoxygenated venous blood reaches 78 mL·m -2 ·min -1 . The oxygen transfer performance of the TBNC extracorporeal membrane oxygenation membrane is about twice that of the BNC membrane.

[0131] The PVDF-CA / BNC extracorporeal membrane oxygenation membrane prepared in Example 14, the PVDF extracorporeal membrane oxygenation membrane prepared in Example 15, and the BNC membrane prepared in Comparative Example 2 were respectively tested for morphology, bursting pressure, red blood cell and platelet adhesion, and oxygen permeability. From Figure 5 it can be seen that after the BNC membrane is compounded with PVDF or PVDF-CA, an obvious interfacial structure of the hydrogel membrane and the spun membrane appears in the cross section, proving the successful preparation of the material. From Figure 6 it can be seen that the prepared PVDF-CA / BNC extracorporeal membrane oxygenation membrane has excellent compressive performance, and the bursting pressure is 37.60 kPa. From Figure 7 it can be seen that the prepared PVDF-CA / BNC extracorporeal membrane oxygenation membrane has good blood compatibility, and no obvious platelet activation and red blood cell deformation occur. From Figure 8 it can be seen that the oxygen transmission rates in water of the prepared BNC membrane, PVDF / BNC extracorporeal membrane oxygenation membrane, and PVDF-CA / BNC extracorporeal membrane oxygenation membrane are 14 mL·m -2 ·min -1 , 40 mL·m -2 ·min -1 and 43 mL·m -2 ·min -1 . This is because the ultrathin BNC membrane structure is easily damaged by oxygen pressure. After compounding with PVDF, the bursting pressure of the PVDF / BNC membrane is significantly enhanced and can remain stable under higher oxygen pressure, so the overall oxygen permeability rate increases. The incorporation of hydrophilic CA makes the spun membrane and the BNC membrane combine more closely, further improving the compressive performance and oxygen permeability of the material.

[0132] Table 1

[0133]

[0134] Table 1 (continued)

[0135]

[0136] As can be seen from the above results, the present invention improves the oxygen transmission performance of the bacterial cellulose nanofiber membrane. The prepared extracorporeal membrane oxygenation membrane of bacterial cellulose nanofibers has the advantages of high mechanical strength, good blood compatibility, and high oxygen transmission rate. This invention contributes to the development and application of bacterial nanocellulose in the field of artificial lung membranes.

Claims

1. An in vitro membrane lung oxygenation membrane based on bacterial cellulose nanofibers, characterized in that: It is prepared by a hydrophobic modification method or a pore size increasing and mechanical strengthening method.

2. A preparation method of a nanofiber extracorporeal membrane oxygenation membrane based on bacterial cellulose, comprising the following steps: (1) Inoculating Acetobacter xylinum into a culture medium and culturing it statically; (2) Purifying the obtained cellulose membrane with a sodium hydroxide solution and water to obtain a purified bacterial cellulose nanofiber membrane; (3) Hydrophobically modifying the bacterial cellulose nanofiber membrane by a physical modification method or a chemical modification method, and finally vacuum drying to obtain a nanofiber extracorporeal membrane oxygenation membrane.

3. The preparation method according to claim 2, wherein: The physical modification method in the step (3) includes a physical adsorption method or a physical spraying method; the chemical modification method includes a silanization method, an alkanoylation method, an esterification method or a graft polymer method.

4. The preparation method according to claim 4, characterized in that: The reagents used in the physical adsorption method include one or more of a cationic surfactant, a quaternary ammonium salt, and a diblock copolymer dispersant; the reagents used in the physical spraying method include a commercial waterproof spray.

5. The preparation method according to claim 4, characterized in that: The silanization method uses a silanization reagent; the alkanoylation method uses an alkanoylation reagent; the esterification method uses an esterification reagent; the graft polymer method uses a polymer side chain.

6. A preparation method of a nanofiber extracorporeal membrane oxygenation membrane based on bacterial cellulose, comprising the following steps: (1) Dissolving a synthetic polymer in a solvent to prepare a spinning solution; ultrasonically removing bubbles from the spinning solution, and obtaining electrospun fibers by an electrospinning technique; (2) Adding a culture medium containing Acetobacter xylinum into a petri dish and culturing it statically to obtain a bacterial cellulose nanofiber membrane; (3) Sterilizing the electrospun fibers and placing them on the bacterial cellulose nanofiber membrane, and supplementing the culture medium with a culture medium containing Acetobacter xylinum, and culturing it statically; (4) Purifying the cellulose membrane obtained by culturing in the step (3) with a sodium hydroxide solution and water, and finally obtaining a nanofiber extracorporeal membrane oxygenation membrane.

7. The preparation method according to claim 6, characterized in that: The synthetic polymer in the step (1) includes one or more of cellulose acetate, polyvinylidene fluoride, polyether ketone or polysulfone.

8. The preparation method according to claim 6, characterized in that: The solvent in the step (1) is one or more of N,N-dimethylformamide, acetone, chloroform or hexafluoroisopropanol.

9. The preparation method according to claim 6, characterized in that: The electrospinning conditions in the step (1) are 10-20 kV, the receiving distance is 10-15 cm, the roller rotation speed is 10-30 r / min, the injection rate is 0.5-2 mL / h, and the electrospinning time is 0.5-4 h.

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