Preparation method of micron and nano short fibers

Abstract

The invention discloses a preparation method of micron and nano short fibers. The preparation method comprises the following steps: preparing an embedding medium composed of a high-molecular water-soluble polymer and an oligosaccharide solution; embedding the prepared fiber membrane with an embedding agent, and freezing to obtain a fiber embedding block; cutting the embedding block by using a slicing machine according to a direction perpendicular to the fiber membrane to obtain a sheet-shaped cut object containing short fibers; melting the cut sheet, stirring to form a uniformly dispersed short fiber fusion solution, then freezing and cutting again, and repeating for 1-5 times; and repeatedly washing with deionized water, filtering the short fibers, removing the embedding agent, and freeze-drying to obtain the short fibers with the length of 50-1000 microns. The invention provides a preparation method of micron and nano short fibers. The micron and nano superfine short fibers which are controllable in length, good in uniformity and good in biocompatibility can be efficiently prepared. The method has the advantages that the cost is low, the method is suitable for large-scale production, the whole process is green and pollution-free, and the use of toxic chemical reagents is avoided.

Description

Technical Field

[0001] This invention relates to the field of micron and nanofiber preparation, and more particularly to a method for preparing micron and nano short fibers. Background Technology

[0002] Microfibers and nanofibers refer to ultrafine fiber materials with diameters in the micrometer and nanometer range. Their manufacturing technologies include micro-extrusion bioprinting, melt spinning, electrospinning, and air-jet spinning. By adjusting the preparation parameters and material ratios, the fiber diameter can be effectively controlled. To enable more flexible applications in different research fields, these materials often require specific morphological designs. For example, in tissue engineering and regenerative medicine, developing injectable microcarriers requires processing microfibers and nanofibers into short fibers with lengths ranging from micrometers to millimeters to fully utilize the unique injectable properties of short fibers.

[0003] Existing technologies for preparing micron and nano short fibers include homogenization, ultrasonic disruption, and cryo-slicing. However, these methods often introduce or generate factors detrimental to material properties and biocompatibility during the preparation of short fibers. Homogenization uses the mechanical shearing force generated by the high-speed rotation of the homogenizer blade to shear and disperse the fibers. This method is simple to operate and low in cost, but its core drawback is that it cannot produce short fibers with uniform size. To improve dispersion efficiency, organic solvents (such as toluene and xylene) or strong alkaline solutions are often added before or during homogenization to pre-hydrolyze and degrade the fibers. The introduction of these chemical additives may not only change the original chemical structure of the fibers and affect their mechanical strength, but also affect biocompatibility due to residual toxic substances. Ultrasonic disruption utilizes the cavitation effect of ultrasound to break down fibers. This process requires the addition of ultrasonic media such as ethanol and isopropanol, and generates intense local high temperatures, which can lead to thermal degradation of the fiber material, destroying its original physicochemical properties and also being detrimental to maintaining the intrinsic properties and biocompatibility of the material. The cryosection method requires embedding a fiber membrane in a solution containing a commercially available OCT embedding agent, followed by embedding and cutting to prepare short fibers of uniform size. However, these commercial OCT embedding agents typically contain potentially cytotoxic components (such as polyvinylpyrrolidone and polyethylene glycol), and their residues can severely affect the biocompatibility of the short fibers. In summary, current mainstream technologies (cryosection, homogenization, and ultrasonic disruption) generally suffer from reliance on toxic chemical additives, resulting in destructive thermal effects or damage to fiber structure / strength when preparing micron and nanofibers. Summary of the Invention

[0004] In view of the aforementioned deficiencies in existing technologies, the technical problem to be solved by this invention is that existing mainstream technologies (freeze-slicing method, homogenization method, and ultrasonic disruption method) generally rely on toxic chemical additives in the preparation of micron and nano short fibers, resulting in destructive thermal effects or damage to fiber structure / strength. This invention provides a method for preparing micron and nano short fibers using an embedding agent composed of oligosaccharides and water-soluble polymers. By embedding the fiber membrane in this embedding agent and then slicing it, micron and nano-scale ultrafine short fibers with controllable length, good uniformity, and excellent biocompatibility can be efficiently prepared. This method has advantages such as low cost, suitability for large-scale production, and being green and pollution-free throughout the entire process, avoiding the use of toxic chemical reagents.

[0005] To achieve the above objectives, the present invention provides a method for preparing micron and nano short fibers, comprising the following methods:

[0006] Step (1): Prepare an encapsulation agent consisting of a water-soluble polymer and an oligosaccharide solution;

[0007] Step (2): The prepared polymer fiber membrane is embedded with the embedding agent and frozen in an ultra-low temperature freezer at -20°C to -80°C to obtain a fiber embedded block;

[0008] Step (3): Cut the embedded block with a slicer in a direction perpendicular to the fiber membrane to obtain a sheet-like cut containing short fibers;

[0009] Step (4): Melt the sheet-like cut material, stir to form a uniformly dispersed short fiber fusion liquid, then freeze and cut the uniformly dispersed short fiber fusion liquid again, repeating 1-5 times;

[0010] Step (5): repeatedly rinse and filter the short fibers with deionized water to remove the embedding agent. After washing the short fibers with deionized water 1-3 times, freeze-dry them to obtain short fibers with a length of 50-1000μm.

[0011] Furthermore, an encapsulating agent composed of a water-soluble polymer and an oligosaccharide solution is prepared, specifically comprising a mixture of 0.1-30 wt% of a water-soluble polymer and 0.1-20 wt% of an oligosaccharide solution in a certain mass ratio.

[0012] Furthermore, the high molecular weight water-soluble polymer includes one or more of polyvinyl alcohol (PVA), polyethylene glycol (PEG), polymaleic anhydride (PMA), polyacrylamide (PAM), and polyvinylpyrrolidone (PVP).

[0013] Furthermore, the oligosaccharide solution includes one or more of glucose, maltose, sucrose, lactose, fructose, and glyceraldehyde.

[0014] Further, in step (2), the polymer is one or more of the following: lactic acid and caprolactone copolymer (PLCL), polyurethane (PU), polycaprolactone (PCL), polyglycerol sebate (PGS), polyoctyl citrate (POC), polylactic acid (PLA), polymethyl methacrylate (PMMA), copolymer of 3-hydroxybutyrate and 3-hydroxyvalerate (PHBV), and copolymer of lactic acid and glycolic acid (PLGA).

[0015] Furthermore, the methods for preparing the fiber membrane include one or more of the following: micro-extrusion bioprinting, melt spinning, electrospinning, and air-jet spinning.

[0016] Furthermore, the prepared fiber membrane is embedded in the embedding agent by laying the fiber membrane layer by layer in the embedding agent, laying 1-5 layers.

[0017] Furthermore, when removing the encapsulating agent using short fibers, a nylon filter screen with a mesh size of 200-500 is used for filtration.

[0018] Technical effect

[0019] This invention provides a method for preparing micron- and nano-sized short fibers. By using an embedding agent formed from oligosaccharides and water-soluble polymers, combined with an embedding and slicing method, ultrafine short fibers with uniform and controllable length and fineness at the micron and nanometer scale can be successfully prepared. The embedding agent used is a biocompatible material and does not involve organic bases or other reagents, resulting in short fibers with good biocompatibility. This method is characterized by low cost, environmental friendliness, and high efficiency. The preparation process does not contain strong alkalis or organic solvents. The ultrafine short fibers prepared by this invention can serve as excellent carriers for biomedical research and clinical applications. This method has advantages such as low cost, suitability for large-scale production, and a completely green and pollution-free process, avoiding the use of toxic chemical reagents.

[0020] The following will further explain the concept, specific structure, and technical effects of the present invention in conjunction with the accompanying drawings, so as to fully understand the purpose, features, and effects of the present invention. Attached Figure Description

[0021] Figure 1 Ordinary optical microscope images of the micron and nano-scale ultrafine short fibers of PLCL prepared in Example 2 and Comparative Example 1.

[0022] Figure 2 The image shows the statistical distribution of fiber lengths of the micron and nano-scale ultrafine short fibers of PLCL prepared in Example 2 and Comparative Example 1.

[0023] Figure 3 These are ordinary optical microscope images of the PLGA micron and nano-scale ultrafine short fibers prepared in Example 3 and Comparative Example 2.

[0024] Figure 4 These are ordinary optical microscope images of the PLGA micron and nano-scale ultrafine short fibers prepared in Example 3 and Comparative Example 2.

[0025] Figure 5 Ordinary optical microscope images of the PLLA micron and nano-scale ultrafine short fibers prepared in Example 4 and Comparative Example 3.

[0026] Figure 6 Ordinary optical microscope images of the PLLA micron and nano-scale ultrafine short fibers prepared in Comparative Examples 4 and 3.

[0027] Figure 7 The images show live / dead staining of L929 cells after culturing the micron and nano-sized ultrafine short fiber extract prepared in Example 5 for 24 hours.

[0028] Figure 8 The live / death rate of L929 cells after culturing with the micron and nano-scale ultrafine short fiber extract prepared in Example 5 for 24 hours is statistically analyzed. Detailed Implementation

[0029] To make the technical problems to be solved, the technical solutions, and the beneficial effects of the present invention clearer, the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the present invention and are not intended to limit the present invention.

[0030] In the following description, specific details, such as particular internal procedures and techniques, are set forth for illustrative purposes and not for limitation, in order to provide a thorough understanding of the embodiments of the invention. However, those skilled in the art will appreciate that the invention may be practiced in other embodiments without these specific details. In other instances, detailed descriptions of well-known systems, apparatuses, circuits, and methods have been omitted so as not to obscure the description of the invention with unnecessary detail.

[0031] Preparation of fiber membranes:

[0032] At room temperature, 0.5 g of lactide-caprolactone copolymer (PLCL, LA:CL = 75:25 mol:mol) (purchased from Jinan Daigang Biotechnology Co., Ltd., China, Mw = 100,000 Da) and 0.026 g of polyethylene oxide (PEO) were dissolved in 5 mL of hexafluoroisopropanol and stirred overnight to obtain a spinning solution. The prepared solution was subjected to stable jet electrospinning under the following conditions: voltage 5-7 kV, receiving distance 20 cm, roller speed 1000 r / min (diameter 10 cm), ambient temperature 20-25℃, and humidity 25-30%. The resulting fibers were dried in a vacuum drying oven for more than one week for subsequent experiments.

[0033] Using the above spinning system, the PLCL in the system was replaced with polylactic acid-glycolic acid copolymer (PLGA, GA:LA = 75:25 mol:mol) (purchased from Jinan Daigang Biotechnology Co., Ltd., China, Mw = 340,000 Da) and polylactic acid (PLLA) (purchased from Jinan Daigang Biotechnology Co., Ltd., China, Mw = 100,000 Da) to prepare electrospun fiber membranes.

[0034] Example 1

[0035] This embodiment describes the preparation method of the embedding agent:

[0036] Polyvinyl alcohol and sucrose were dissolved separately in deionized water and stirred overnight to prepare 5 wt% polyvinyl alcohol solution and 20 wt% sucrose solution. The above solutions were mixed 1:1 to prepare a homogeneous solution for subsequent experiments.

[0037] Example 2

[0038] (1) Prepare the embedding agent in advance according to Example 1;

[0039] (2) The PLCL fiber membrane prepared by electrospinning was placed in parallel in the embedding agent and frozen in an ultra-low temperature freezer at -20℃ to -80℃.

[0040] (3) The embedded block is cut with a slicer in a direction perpendicular to the fiber membrane; in this embodiment, the width of the cutting blade is fixed at 1 mm, and the width of the cut embedded block is 1 mm. A sheet-like cut containing short fibers is obtained by cutting, and after melting it at room temperature, the short fiber-embedding agent mixture after one cut is collected and recorded as C1;

[0041] (4) The C1 fiber mixture obtained in step (3) is frozen again, and the cutting and melting steps in step (3) are repeated 3 times and 5 times respectively to collect C3 and C5 short fibers.

[0042] (5) Filter each group of short fibers with a 200-mesh filter screen and wash them with deionized water three times to obtain short fibers with a length of 50-1000μm.

[0043] The short fiber samples obtained above were prepared into slides and observed under an optical microscope. ImageJ was used to statistically analyze the fiber lengths. Images of the short fibers in each group under the optical microscope are shown below. Figure 1 As shown, its length is statistically analyzed, such as... Figure 2 As shown, the average lengths of short fibers in groups C1, C3, and C5 are 592.5±323.5μm, 484.4±263.1μm, and 398.0±219.6μm, respectively.

[0044] The short fibers prepared by cutting are relatively uniform in length and have the characteristics of a wider length range and better dispersibility.

[0045] Comparative Example 1

[0046] (1) Take the C1, C3 and C5 short fibers of PLCL collected in Example 2, homogenize them in 200 mL of deionized water at 25,000 rpm for 5 min using a homogenizer, filter and wash them through a 200 mesh screen, and record them as C1H1, C3H1 and C5H1.

[0047] (2) The PLCL short fiber samples obtained above were prepared into slides and observed under an optical microscope. The fiber length was then statistically analyzed using ImageJ. Images of the short fibers in each group under the optical microscope are shown below. Figure 1 As shown, its length is statistically analyzed, such as... Figure 2 As shown, the average lengths of short fibers of C1H1, C3H1, and C5H1 are 545.26±271.3μm, 456.3±298.9μm, and 331.0±188.9μm, respectively.

[0048] The combined use of cutting and homogenization methods can improve fiber dispersibility, and homogenization has a synergistic effect on the length and dispersibility of short fibers.

[0049] Example 3

[0050] According to Example 2, the PLCL fiber membrane in Example 2 was replaced with a PLGA fiber membrane, while all other aspects remained the same as in Example 2, resulting in C1, C3, and C5 short fibers. The photographs of the short fibers in each group under an optical microscope are shown below. Figure 3 As shown, its length is statistically analyzed, such as... Figure 4 As shown, the average lengths of short fibers in C1, C3, and C5 are 457.2±224.1μm, 408.2±196.1μm, and 337.0±168.9μm, respectively.

[0051] Similar to PLCL, the length of PLGA short fibers decreases after multiple cuts, and the embedding-cutting method is also suitable for preparing PLGA short fibers.

[0052] Comparative Example 2

[0053] According to Comparative Example 1, the C1, C3, and C5 short fibers of PLGA obtained in Example 3 were homogenized to prepare C1H1, C3H1, and C5H1 short fibers. Photographs of each group of short fibers under an optical microscope are shown below. Figure 3 As shown, its length is statistically analyzed, such as... Figure 4As shown, the average lengths of the short fibers of PLGA C1H1, C3H1, and C5H1 are 437.4±189.9μm, 389.6±184.0μm, and 313.5±180.7μm, respectively.

[0054] Similar to PLCL, the combined use of embedding and cutting methods and homogenization methods can improve the dispersibility of PLGA short fibers and also have a certain effect on reducing their length.

[0055] Example 4

[0056] According to Example 2, the PLCL fiber membrane in Example 2 was replaced with a PLLA fiber membrane, while all other aspects remained the same as in Example 1, resulting in C1, C3, and C5 short fibers. Photographs of the short fibers in each group under an optical microscope are shown below. Figure 5 As shown, its length is statistically analyzed, such as... Figure 6 As shown, the average lengths of C1, C3, and C5 short fibers are 635.5±434.7μm, 458.5±331.3μm, and 265.2±250.5μm, respectively.

[0057] Repeated embedding and cutting can effectively reduce the length of PLLA short fibers and promote the dispersion of PLLA short fibers.

[0058] Comparative Example 3

[0059] According to Comparative Example 1, the C1, C3, and C5 short fibers obtained in Example 4 were homogenized to prepare C1H1, C3H1, and C5H1 short fibers. Photographs of the short fibers under an optical microscope are shown below. Figure 5 As shown, its length is statistically analyzed, such as... Figure 6 As shown, the average lengths of the short fibers of C1H1, C3H1, and C5H1 are 412.8±360.1μm, 320.9±282.9μm, and 253.5±239.6μm, respectively.

[0060] The combined use of embedding and cutting methods and homogenization methods can more significantly shorten the length of short fibers in rigid polymers such as PLLA.

[0061] Example 5

[0062] The C5 short fibers obtained in Example 2 were washed with deionized water and freeze-dried. 0.1 g of the short fibers were then soaked in 1 mL of DMEM medium containing 10% FBS for 24 h. The extract was collected and used to culture L929 fibroblasts. The cell viability / deadness staining after 24 h of culture is shown below. Figure 7 As shown, the survival / mortality statistics are as follows: Figure 8 As shown, the cell viability reached 98.53±0.57%, indicating that the prepared short fibers had no cytotoxicity and good biocompatibility.

[0063] The preferred embodiments of the present invention have been described in detail above. It should be understood that those skilled in the art can make numerous modifications and variations based on the concept of the present invention without creative effort. Therefore, all technical solutions that can be obtained by those skilled in the art based on the concept of the present invention through logical analysis, reasoning, or limited experimentation on the basis of existing technology should be within the scope of protection defined by the claims.

Claims

1. A method for preparing micron and nano short fibers, characterized in that, Including the following methods: Step (1): Prepare an encapsulation agent consisting of a water-soluble polymer and an oligosaccharide solution; Step (2): The prepared polymer fiber membrane is embedded with the embedding agent and frozen in an ultra-low temperature freezer at -20°C to -80°C to obtain a fiber embedded block; Step (3): The embedded block is cut with a slicer in a direction perpendicular to the fiber membrane to obtain a sheet-like cut containing short fibers; Step (4): Melt the sheet-like cut material, stir to form a uniformly dispersed short fiber fusion liquid, then freeze and cut the uniformly dispersed short fiber fusion liquid again, repeating 1-5 times; Step (5): repeatedly rinse and filter the short fibers with deionized water to remove the embedding agent. After washing the short fibers with deionized water 1-3 times, freeze-dry them to obtain short fibers with a length of 50-1000μm.

2. The method for preparing micron and nano short fibers as described in claim 1, characterized in that, The encapsulation agent is composed of a water-soluble polymer and an oligosaccharide solution. Specifically, it is prepared by mixing 0.1-30 wt% of a water-soluble polymer and 0.1-20 wt% of an oligosaccharide solution in a certain mass ratio.

3. The method for preparing micron and nano short fibers as described in claim 2, characterized in that, The water-soluble polymer includes one or more of polyvinyl alcohol (PVA), polyethylene glycol (PEG), polymaleic anhydride (PMA), polyacrylamide (PAM), and polyvinylpyrrolidone (PVP).

4. The method for preparing micron and nano short fibers as described in claim 2, characterized in that, The oligosaccharide solution includes one or more of glucose, maltose, sucrose, lactose, fructose, and glyceraldehyde.

5. The method for preparing micron and nano short fibers as described in claim 1, characterized in that, The polymer of the polymer fiber membrane in step (2) includes one or more of the following: lactic acid and caprolactone copolymer (PLCL), polyurethane (PU), polycaprolactone (PCL), polyglycerol sebate (PGS), polyoctyl citrate (POC), polylactic acid (PLA), polymethyl methacrylate (PMMA), copolymer of 3-hydroxybutyrate and 3-hydroxyvalerate (PHBV), and copolymer of lactic acid and glycolic acid (PLGA).

6. The method for preparing micron and nano short fibers as described in claim 1, characterized in that, The fiber membrane is prepared by one or more of the following methods: micro-extrusion bioprinting, melt spinning, electrospinning, and air-jet spinning.

7. The method for preparing micron and nano short fibers as described in claim 6, characterized in that, The prepared fiber membrane is embedded in the embedding agent, specifically by laying the fiber membrane layer by layer in the embedding agent, laying 1-5 layers.

8. The method for preparing micron and nano short fibers as described in claim 1, characterized in that, When removing the encapsulating agent by filtering short fibers, use a nylon filter screen with a mesh size of 200-500.

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