Collagen yarn, method for producing same, and biomaterial

Abstract

This collagen yarn contains collagen nanofibrils, and has a diameter of 10-50 μm and a total length of 2 × 105 times or more greater than the diameter, wherein the collagen nanofibrils exhibit high alignment properties along the longitudinal direction of the collagen yarn, the high alignment properties are indicated by a fact that the refractive index difference determined by a birefringence measurement performed on a hydrated collagen yarn is 4.3 × 10-4 or more, and the hydrated collagen yarn is obtained by immersing the collagen yarn in a neutral phosphate buffer solution.

Description

Collagen thread, its manufacturing method, and biomaterial

[0001] The present invention relates to a collagen thread, a method for producing the same, and a biomaterial.

[0002] As disclosed in JP 2019-085373 A (Patent Document 1) and elsewhere, progress is being made in the development of artificial collagen threads that mimic collagen fibers found in vivo. Artificial tendons made by bundling the above-mentioned artificial collagen threads, membrane materials made by weaving the above-mentioned artificial collagen threads, and other materials may be useful as cell scaffold materials, and the application of the above-mentioned artificial tendons and membrane materials as medical materials is expected.

[0003] Japanese Patent Application Laid-Open No. 2019-085373

[0004] Haynl C et al. NANO LETTERS 2016: 16: 5917-5922Paten J A et al. ACS Nano 2016: 10: 5027-5040

[0005] By using an artificial collagen thread that has at least the following four characteristics, it may be possible to artificially reproduce collagen fibers found in the body: 1) It is safe for the body; 2) It has a thinness similar to that of collagen fibers found in the body (diameter: approximately 20 μm); 3) The collagen nanofibrils that make up the artificial collagen thread are aligned inside the artificial collagen thread, preferably all the way to the core; and 4) It has a length sufficient for producing artificial tendons and membrane materials.

[0006] The highly oriented collagen fiber bundles disclosed in Patent Document 1 have fibers aligned throughout the artificial collagen yarn. However, even the smallest diameter is approximately 100 μm, and further improvement is required to reproduce collagen fibers found in vivo. The method disclosed in Haynl C et al. NANO LETTERS 2016: 16: 5917-5922 (Non-Patent Document 1) combines traditional wet spinning with microchannels, enabling continuous production of artificial collagen yarns with a diameter of approximately 4 μm. However, wet spinning is known to align collagen fibers on the yarn surface but not the interior. The flow-induced crystallized collagen disclosed in Paten J A et al. ACS Nano 2016: 10: 5027-5040 (Non-Patent Document 2) is capable of aligning collagen nanofibrils throughout the interior and core of the artificial collagen yarn, but it is difficult to achieve sufficient length for the fabrication of artificial tendon and membrane materials. Therefore, an artificial collagen thread that artificially reproduces collagen fibers found in vivo has not yet been obtained, and its development is eagerly awaited.

[0007] In view of the above, an object of the present invention is to provide a collagen thread that can artificially reproduce collagen fibers found in vivo, a method for producing the same, and a biomaterial.

[0008] The present inventors have conducted extensive research to solve the above problems and have completed the present invention. The present inventors have discovered a method for producing a collagen fiber by passing a raw material consisting of a neutral collagen solution with a high concentration (for example, 3.0% by mass or more and 5.0% by mass or less) through a narrow-diameter (for example, an inner diameter of 100 μm or more and 300 μm or less) through a flow path with a high shear rate (for example, 100 s -1 The present inventors have come up with the idea of ​​producing collagen threads by discharging the collagen nanofibrils into a coagulation bath filled with a high-concentration (70% by volume or more) ethanol solution at a concentration of 70% by volume or more and 95% by volume or less. The collagen threads obtained by the above production method have a diameter of 50 μm or less, and the collagen nanofibrils that make up the collagen threads are aligned inside the collagen thread, preferably to the core, and have a diameter of 2×10 5 This finding led to the present invention.

[0009] The present invention has the following characteristics: [1] A collagen thread according to the present invention is a collagen thread containing collagen nanofibrils, the collagen thread having a diameter of 10 μm or more and 50 μm or less, and a total length of the collagen thread that is 2×10 times longer than the diameter. 5 The collagen nanofibrils have a length of 4.3×10 or more times that of the hydrated collagen yarn, and the collagen nanofibrils exhibit high alignment along the longitudinal direction of the collagen yarn, and the high alignment is such that the refractive index difference determined from birefringence measurements of the hydrated collagen yarn is 4.3×10 -4 The hydrated collagen thread can be obtained by immersing the collagen thread in a neutral phosphate buffer solution. [2] The collagen nanofibrils are preferably composed of atelocollagen. [3] The total length is 1 × 10 relative to the diameter. 6 [4] A method for producing a collagen thread according to the present invention includes the steps of preparing a neutral collagen solution containing collagen at a concentration of 3.0% by mass or more and 5.0% by mass or less, and obtaining a collagen thread by discharging the neutral collagen solution into an ethanol solution having a concentration of 70% by volume or more and 95% by volume or less, wherein the collagen is atelocollagen, and in the step of obtaining the collagen thread, the neutral collagen solution is passed through a flow path having an inner diameter of 100 μm or more and 300 μm or less for 100 seconds. -1 More than 1500s -1 The collagen thread is extruded into the ethanol solution at a shear rate of 300 s or less. [5] The collagen thread manufacturing method preferably further comprises a step of stretching the collagen thread by winding the collagen thread at a winding speed of 1.5 to 4.0 times the linear extrusion velocity. [6] The neutral collagen solution preferably contains the collagen at a concentration of 4.5 to 5.0 mass %. [7] The shear rate is 300 s or less. -1 More than 1000s -1 [8] The biomaterial according to the present invention includes the collagen thread. [9] The biomaterial is preferably an artificial tendon or membrane material.

[0010] According to the present invention, it is possible to provide a collagen thread that can artificially reproduce collagen fibers found in vivo, a method for producing the same, and a biomaterial.

[0011] FIG. 1 is a scanning electron microscope image of a cross section taken along the axial direction of a collagen thread according to the present embodiment. FIG. 2 is an explanatory diagram illustrating birefringence measurement performed by line analysis on a hydrated collagen thread in order to evaluate the high alignment of collagen nanofibrils that make up the collagen thread according to the present embodiment. FIG. 3 is an image taken with a digital camera of the appearance of a biomaterial (artificial tendon) obtained from the collagen thread according to the present embodiment. FIG. 4 is a flowchart showing an example of a method for manufacturing a collagen thread according to the present embodiment. FIG. 5 is a schematic diagram of a manufacturing apparatus for producing a collagen thread according to the present embodiment. FIG. 6 is another schematic diagram of a manufacturing apparatus for producing a collagen thread according to the present embodiment.

[0012] Hereinafter, an embodiment of the present invention (hereinafter also referred to as "the present embodiment") will be described in more detail. In this specification, the notation "A to B" means the upper and lower limits of a range (i.e., greater than or equal to A and less than or equal to B), and when no unit is specified for A and only a unit is specified for B, the units of A and B are the same. The term "alignment" means that the axes of collagen nanofibrils formed by the regular arrangement of multiple collagen molecules are aligned. In some prior art documents, the term "orientation," which refers to the alignment of not only the axes but also the orientation of the collagen nanofibrils, is used synonymously with the above-mentioned "alignment," but the two terms are used separately in this specification.

[0013] As used herein, the "interior" of a collagen thread refers to the entire portion excluding the surface of the collagen thread. The "core" of a collagen thread refers to the central portion that appears on a cross section obtained by cutting the collagen thread along a plane perpendicular to its axial direction, and particularly refers to the portion where the central portion is continuous along the axial direction.

[0014] [Collagen thread] The collagen thread according to this embodiment is a collagen thread containing collagen nanofibrils. The diameter of the collagen thread is 10 μm or more and 50 μm or less. The total length of the collagen thread is 2×10 times the diameter. 5 The collagen nanofibrils exhibit high alignment along the longitudinal direction of the collagen thread. The high alignment is demonstrated when the refractive index difference determined from birefringence measurements of the hydrated collagen thread is 4.3 × 10 -4 This is demonstrated by the above. The hydrated collagen thread can be obtained by immersing the collagen thread in a neutral phosphate buffer solution. A collagen thread with these characteristics can artificially reproduce collagen fibers found in living organisms. Therefore, the collagen thread may be useful as a biomaterial such as an artificial tendon or membrane material.

[0015] <Collagen Nanofibrils> FIG. 1 is a scanning electron microscope image of a cross section of a collagen yarn according to this embodiment taken along the axial direction. As shown in FIG. 1, the collagen yarn 1 includes collagen nanofibrils 11. Collagen nanofibrils 11 are collagen fibers formed by the spontaneous assembly and alignment of countless collagen molecules in a neutral environment upon humidification, resulting in a thread-like appearance with an overall thickness of approximately 100 nm. Collagen nanofibrils 11 can be obtained from collagen with fibril-forming ability (fibril-forming collagen). Preferred examples of the fibril-forming collagen include type I collagen, which is a collagen constituting bone, skin, tendons, and ligaments; type II collagen, which is a collagen constituting cartilage; and type III collagen, which is contained in biological tissues composed of type I collagen. The fibril-forming collagen can be obtained by extraction and purification from biological tissues using standard methods, or by purchasing a commercially available product (e.g., product name: "Collagen BM" manufactured by Nitta Gelatin Co., Ltd.). The fibril-forming collagen can consist of each type of collagen alone or a mixture of multiple types of collagen. The animal species from which the fibril-forming collagen is derived is not particularly limited, but from the viewpoint of application as a biomaterial, collagen derived from mammals such as pigs and cows is preferred.

[0016] The collagen nanofibrils 11 are preferably composed of atelocollagen. This makes it possible to provide a collagen thread with low antigenicity and high biocompatibility. "Atelocollagen" refers to telopeptide-removed collagen prepared by enzymatically decomposing and removing the telopeptides at the N- and C-termini of collagen molecules using proteolytic enzymes such as pepsin, chymosin, cathepsin D, and renin. In particular, as the atelocollagen constituting the collagen nanofibrils, among the above telopeptide-removed collagens, mammal-derived telopeptide-removed collagens approved as raw materials for medical devices are preferred, with telopeptide-removed collagen derived from pigskin being more preferred.

[0017] <Diameter> The diameter of the collagen thread 1 is 10 μm or more and 50 μm or less. From the viewpoint of application to biomaterials such as artificial tendons or membrane materials, the diameter of the collagen thread 1 is preferably 10 μm or more and 40 μm or less, and more preferably 20 μm or more and 30 μm or less. If the diameter of the collagen thread 1 is less than 10 μm, it may be too thin and may not have sufficient strength, and may have poor windability, such as being broken during winding. If the diameter of the collagen thread 1 exceeds 50 μm, it may be too thick and may be difficult to reproduce the function of a collagen fiber in a living body (for example, cell scaffold function). The diameter of the collagen thread 1 can be determined by determining the cross-sectional area through image analysis of an image obtained by scanning electron microscope observation of the cross section of the collagen thread 1, and regarding the diameter of a circle having the same area as that cross-sectional area (circle-equivalent diameter) as the diameter of the collagen thread 1.

[0018] <Total Length> The total length of the collagen thread 1 is 2×10 5 This allows a single collagen thread 1 to have a length sufficient for producing an artificial tendon and membrane material as a biomaterial. From the viewpoint of producing the artificial tendon and membrane material with a single collagen thread 1, the total length is 1×10 times the diameter. 6 It is preferable that the collagen thread 1 has a length of at least 100 times the length of the collagen thread 1. Specifically, the total length of the collagen thread 1 is preferably 10 m or more, more preferably 100 m or more, and most preferably 200 m or more. There is no particular upper limit to the total length of the collagen thread 1. For example, from the viewpoint of producing the above-mentioned artificial tendon and membrane material, the ideal total length of the collagen thread 1 is 1000 m. The total length of the collagen thread 1 can be determined by measuring the length of the collagen thread obtained by the collagen thread manufacturing method described below that can be wound up without breaking.

[0019] <High Alignment> The collagen nanofibrils 11 exhibit high alignment along the longitudinal direction of the collagen thread 1. The high alignment is achieved when the refractive index difference determined from birefringence measurements of a hydrated collagen thread is 4.3 × 10 -4This is indicated by the above. The hydrated collagen thread is obtained by immersing the collagen thread 1 in a neutral phosphate buffer. As used herein, "high alignment" means that the collagen nanofibrils 11 constituting the collagen thread 1 are aligned at least inside the collagen thread 1, preferably all the way to the core. The neutral phosphate buffer is not particularly limited in terms of pH, ion concentration, amount of sodium chloride added, etc., as long as the conditions are such that collagen fibers can exist stably. However, from the viewpoint of minimizing salt precipitation and hardening the hydrated collagen thread to make it easier to handle, a low-concentration (e.g., 10 mM) sodium hydrogen phosphate solution containing no sodium chloride and adjusted to pH 7 is preferably used as the neutral phosphate buffer.

[0020] The high alignment property is such that the refractive index difference is 4.3×10 -4 The reason for this is as follows. That is, the refractive index difference is a value obtained by dividing the optical path difference that occurs when a predetermined intrinsic polarized light passes through the inside of the hydrated collagen yarn by the thickness of the hydrated collagen yarn, as will be described later. The larger the refractive index difference, the more collagen nanofibrils 11 in the hydrated collagen yarn exist side by side with their axes aligned. And, when the refractive index difference is 4.3 × 10 -4 The above numerical values ​​are obtained only when the collagen nanofibrils 11 are aligned at least inside the collagen thread 1, preferably up to the core, as a result of comparison with the cross-sectional image of the hydrated collagen thread.

[0021] As a result, even if the surface of the collagen thread 1 is digested or decomposed by enzymes or the like in the body and the inside is exposed, the collagen thread 1 remains recognizable by cells due to the high alignment described above, and is therefore able to function as a cell scaffold material. Below, we will explain birefringence measurements of hydrated collagen threads to evaluate the high alignment of the collagen nanofibrils 11.

[0022] (Birefringence Measurement) In this embodiment, the high alignment of the collagen nanofibrils 11 is determined by measuring the refractive index difference of 4.3×10 6 obtained from the birefringence measurement of the hydrated collagen yarn as described above.-4 This is indicated by the above. Birefringence measurement refers to determining the phase difference (retardation, unit: nm) of light that occurs when circularly polarized light is incident on the hydrated collagen yarn using a known two-dimensional birefringence measurement device (for example, product name: "WPA-200" manufactured by Photonic Lattice, Inc.). The refractive index difference can be determined by dividing the phase difference by the thickness of the hydrated collagen yarn (i.e., the distance traveled by the light).

[0023] Because the cross section of a hydrated collagen thread is nearly circular, linear analysis of two-dimensional phase-contrast images can be used to calculate the refractive index difference. The phase difference obtained from linear analysis is represented by a convex shape in a graph in which the horizontal axis represents the position in the direction perpendicular to the axial direction of the hydrated collagen thread (hereinafter also referred to as the "width direction") and the vertical axis represents the phase difference. In this case, the phase-contrast data used to calculate the refractive index difference is obtained from the central 50% of the convex shape, and the thickness used for the calculation can conveniently be 0.95 times the width of the hydrated collagen thread measured separately using a known digital microscope. This is because the thickness of the hydrated collagen thread rapidly decreases at the edges of the convex shape, but the thickness in the central 50% is approximately equivalent to 0.95 times the diameter. Furthermore, the refractive index difference can be calculated by performing the above-mentioned linear analysis at any five locations on the hydrated collagen thread and averaging the five values ​​obtained. "The central 50% of the arched shape" means, for example, the central portion of the shape at a position (width) of 4±2 pixels when the arched shape is shown on a scale of 0 to 8 pixels on the horizontal axis (see Figure 2).

[0024] Fig. 2 is an explanatory diagram illustrating birefringence measurement performed by line analysis on a hydrated collagen yarn in order to evaluate the high alignment of the collagen nanofibrils that make up the collagen yarn according to this embodiment. Fig. 2 shows an image (left side) illustrating birefringence measurement performed by line analysis on a hydrated collagen yarn 10, and a graph (right side) in which the horizontal axis represents the position in the width direction of the hydrated collagen yarn 10 obtained by the birefringence measurement and the vertical axis represents the phase difference at that position. From the graph and the thickness of the hydrated collagen yarn 10, the refractive index difference of the hydrated collagen yarn 10 shown in Fig. 2 was calculated to be 5.88 x 10 -4 It can be calculated that:

[0025] Here, the hydrated collagen thread 10 is obtained by hydrating the collagen thread 1 with a neutral phosphate buffer solution and then introducing chemical crosslinks. Specifically, the hydrated collagen thread 10 is prepared by immersing the collagen thread 1 in an excess amount of neutral phosphate buffer solution (pH range of 6.5 to 7.5) until the swelling state of the collagen thread 1 or the collagen thread in an ethanol solution before drying reaches saturation, and then adding 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC) to introduce chemical crosslinks into the collagen. The EDC concentration in the phosphate buffer solution does not affect birefringence measurement, but a range of 10 mM to 200 mM is preferred from the viewpoint of stabilizing the collagen thread and providing flexibility that prevents it from becoming brittle during handling. Prior to birefringence measurement, the hydrated collagen thread 10 is preferably transferred back into an excess amount of phosphate buffer solution to remove residual crosslinkers and by-products from within the thread. EDC does not color collagen and does not intercalate between collagen molecules, making it suitable as a collagen crosslinker for optical analysis.

[0026] The refractive index difference is 5.0 × 10 -4 It is preferable that the ratio is 7.0×10 or more. -4 It is more preferable that the refractive index difference is 4.3 × 10 or more. This means that the collagen thread 1 has excellent alignment of the collagen nanofibrils 11 constituting the collagen thread 1, particularly all the way to the core, and can therefore be more effective as a cell scaffold material.-4 If it is less than this, there is a risk that the collagen nanofibrils 11 will not be aligned to the inside of the collagen thread 1. There is no particular restriction on the upper limit of the refractive index difference, and it may be, for example, 12×10 -4 In the case of collagen threads obtained by the collagen thread manufacturing method described below, the refractive index difference may be 12×10 -4 The following is realistic.

[0027] <Effects> As described above, the collagen thread according to this embodiment can achieve the above-mentioned features 1) to 4), and therefore has the potential to artificially reproduce collagen fibers found in living organisms. In other words, by using atelocollagen as its raw material, the collagen thread can be safe for living organisms. The collagen thread can have a diameter approximating 20 μm, which is the diameter of collagen fibers found in living organisms. Due to the high alignment property described above, the collagen thread can align collagen nanofibrils inside the collagen thread, preferably all the way to the core. Furthermore, the overall length of the collagen thread can be sufficient for the production of artificial tendons and membrane materials.

[0028] [Biomaterial] The biomaterial according to this embodiment includes the collagen thread described above. A biomaterial having such characteristics can function as a cell scaffold material in vivo, since the collagen thread is an artificial reproduction of collagen fibers found in vivo. In particular, since the biomaterial can function as a cell scaffold material, it is preferable that the biomaterial is an artificial tendon or membrane material. Therefore, the biomaterial is expected to be applied in the medical field, etc., as an artificial tendon or membrane material.

[0029] FIG. 3 is an image taken with a digital camera showing the appearance of a biomaterial (artificial tendon) obtained from the collagen thread according to this embodiment. As shown in FIG. 3 , the biomaterial obtained from the collagen thread can be used to form an artificial tendon 200 based on the above-mentioned characteristics 1) to 4). The artificial tendon 200 shown in FIG. 3 is an example of a 2,000-strand bundle of collagen threads (specifically, the collagen thread of Sample 6 in the Examples described below) with a diameter of 26.8±1.5 μm and a total length of 110 m, each of which has a length of 55 mm and a cross-sectional diameter of 1.7 mm. The length and cross-sectional diameter of the artificial tendon can be adjusted to a predetermined value by adjusting the diameter and number of bundles of collagen threads. Furthermore, although not shown, if the biomaterial obtained from the collagen thread is a membrane material, the membrane material can be formed, for example, by weaving the collagen threads.

[0030] [Method for Manufacturing Collagen Thread] The method for manufacturing the collagen thread is not particularly limited, as long as it can produce a thread having the above-mentioned characteristics. However, the collagen thread can be obtained with a high yield by using the following manufacturing method. That is, the collagen thread manufacturing method according to this embodiment includes the steps of preparing a neutral collagen solution containing collagen at a concentration of 3.0 mass % to 5.0 mass %, and obtaining a collagen thread by discharging the neutral collagen solution into an ethanol solution with a concentration of 70 vol % to 95 vol %. The collagen is atelocollagen. In the step of obtaining the collagen thread, the neutral collagen solution is passed through a flow path with an inner diameter of 100 μm to 300 μm for 100 s. -1 More than 1500s -1The collagen solution is discharged into the ethanol solution at a shear rate of no more than 1.5 times and no more than 4.0 times the linear discharge velocity. In addition, the collagen thread manufacturing method preferably further comprises a step of stretching the collagen thread by winding the collagen thread at a winding speed of no less than 1.5 times and no more than 4.0 times the linear discharge velocity. By winding the collagen thread at a speed exceeding the linear discharge velocity, the collagen solution immediately after being discharged into the ethanol solution is stretched and fibrillated (coagulated), the diameter of the thread becomes thinner, and the degree of fibril alignment becomes higher. A collagen thread manufacturing method having these characteristics can obtain a collagen thread that can artificially reproduce collagen fibers found in living organisms.

[0031] Specifically, the above-mentioned manufacturing method preferably includes the steps shown in the flowchart of Fig. 4. Fig. 4 is a flowchart showing an example of a collagen thread manufacturing method according to the present embodiment. According to Fig. 4, the collagen thread manufacturing method includes a step S10 of preparing a neutral collagen solution containing collagen at a concentration of 3.0 mass % or more and 5.0 mass % or less (first step: preparation step), and a step S20 of obtaining a collagen thread by discharging the neutral collagen solution into an ethanol solution having a concentration of 70 vol % or more and 95 vol % or less (second step: collagen thread obtaining step). According to Fig. 4, the collagen thread manufacturing method preferably further includes a step S30 of drawing the collagen thread by winding the collagen thread at a winding speed of 1.5 to 4.0 times the linear extrusion speed (third step: collagen thread drawing step).

[0032] As used herein, the term "neutral collagen solution" refers to a liquid in which collagen molecules are dissolved without fibrillation in a neutral solvent (e.g., a salt-containing solvent such as phosphate-buffered saline). Therefore, a neutral collagen solution is distinct from a cloudy dispersion in which collagen molecules are fibrillated and dispersed in a neutral solvent, even if the solvent is neutral. During the preparation process, some collagen may become irreversibly fibrillated, causing the solution to become slightly cloudy. However, this does not affect the present invention as long as the optical density at a wavelength of 600 nm and an optical path length of 10 mm does not exceed 0.5. Furthermore, a neutral collagen solution is also distinct from a liquid in which collagen molecules are dissolved in an acidic solvent. For example, the pH of the neutral collagen solution is preferably 6.0 or higher and 9.0 or lower.

[0033] The inventors speculate that the reason why the above-described manufacturing method can produce a collagen thread that artificially reproduces collagen fibers found in vivo is as follows. First, the neutral collagen solution contains collagen at a high concentration of 3.0% by mass to 5.0% by mass (preferably 4.5% by mass to 5.0% by mass). This "high concentration" refers to a concentration near the limit at which collagen does not fibrillate and precipitate, even if a large amount of collagen is contained in the neutral solvent, as long as the solution temperature is maintained at 10°C or below. The inventors have discovered that the concentration at which collagen can maintain a molecularly dispersed state in the neutral solvent is approximately 5.0% by mass. In other words, the neutral collagen solution is a solution that is close to saturated for collagen molecules. Therefore, when the collagen molecules in the neutral collagen solution are ejected into a coagulation bath filled with an ethanol solution in step S20 of obtaining a collagen thread, they are instantly fibrillated by the dehydration of the neutral solvent (especially water) by the ethanol. This allows the collagen molecules in the neutral collagen solution to precipitate as a collagen thread composed of collagen nanofibrils, which are bundles of fibrillated collagen molecules. In addition, the neutral collagen solution is discharged at a high shear rate through a narrow flow channel with an inner diameter of 100 μm to 300 μm, in which case the collagen molecules in the neutral collagen solution are aligned in the flow direction and discharged from the flow channel.

[0034] Specifically, with the above-described manufacturing method, collagen molecules in the neutral collagen solution are first discharged into the coagulation bath in a unidirectionally aligned state through the narrow-diameter flow channel. Furthermore, in the coagulation bath, the collagen molecules undergo fibrillation before relaxing their alignment in the flow direction, forming collagen nanofibrils consisting of bundles of collagen molecules. These collagen nanofibrils are then spun into collagen threads. In this case, the collagen nanofibrils can remain highly aligned within the collagen thread, preferably even to its core. The collagen thread can have a desired diameter by appropriately adjusting the inner diameter and shear rate of the flow channel. In principle, the above-described manufacturing method does not limit the length of the collagen thread to be obtained. Additionally, by using, for example, atelocollagen as the collagen molecules, collagen threads that are safe for living organisms can be obtained. Based on the above, it is believed that the above-described manufacturing method can produce collagen threads that artificially reproduce collagen fibers found in living organisms.

[0035] Below, with reference to Figures 4, 5, and 6, each step included in the collagen thread manufacturing method and an overview of a manufacturing apparatus for manufacturing the collagen thread used in the manufacturing method will be described. Figure 5 is a schematic diagram of a manufacturing apparatus for manufacturing a collagen thread according to this embodiment. Figure 6 is another schematic diagram of a manufacturing apparatus for manufacturing a collagen thread according to this embodiment. In the collagen thread manufacturing method, for example, step S20 of obtaining a collagen thread is carried out using the manufacturing apparatus shown in Figure 5. Alternatively, step S20 of obtaining a collagen thread and step S30 of stretching a collagen thread are carried out using the manufacturing apparatus shown in Figure 6. Below, an overview of the manufacturing apparatus will be described, and then each step included in the collagen thread manufacturing method will be described in detail.

[0036] <Manufacturing Apparatus for Producing Collagen Thread> As shown in Figure 5, a manufacturing apparatus 100 for producing collagen thread 1 includes a container 101 that contains a neutral collagen solution, a needle 102 as a flow path for discharging the neutral collagen solution from container 101 to the outside, and a drive device 103 (e.g., a syringe pump) that generates a shear rate for discharging the neutral collagen solution to the outside through needle 102. The manufacturing apparatus 100 further includes a coagulation bath 104 that contains the neutral collagen solution discharged to the outside. This coagulation bath 104 is filled with an ethanol solution with a concentration of 70% by volume or more and 95% by volume or less. As described above, the collagen in the neutral collagen solution is discharged into coagulation bath 104 through needle 102, whereby it precipitates as collagen thread 1 in the ethanol solution.

[0037] The shape of the container 101 is not particularly limited as long as it can hold the neutral collagen solution and discharge the neutral collagen solution to the outside through the needle 102. The container 101 preferably includes, for example, a syringe that holds the neutral collagen solution and a plunger that is driven by the drive device 103 to discharge the neutral collagen solution from the syringe to the outside through the needle 102. This allows the needle 102 to be easily attached to the syringe in the same manner as a syringe. The syringe and plunger can be sized appropriately depending on the amount of neutral collagen solution to be contained. The syringe and plunger may be made of resin, glass, or metal. The drive device 103 can be any pump capable of performing the above-described function by sucking up or sending liquid, gas, or the like under pressure. For example, when this manufacturing method is industrialized, a pneumatic drive device or the like may be used.

[0038] The inner diameter of the needle 102 is 100 μm or more and 300 μm or less. The inner diameter of the needle 102 is preferably 125 μm or more and 250 μm or less. This allows the shear rate of the neutral collagen solution discharged from the needle 102 to be appropriately adjusted using the driving device 103 in the step S20 of obtaining a collagen thread, thereby making it possible to obtain a collagen thread 1 having a desired diameter. The shear rate of the neutral collagen solution discharged from the needle 102 is 100 s -1 More than 1500s -1 The shear rate is set to 300 s or less from the viewpoint of easily adjusting the diameter of the collagen thread. -1 More than 1000s -1 The shear rate is preferably equal to or less than r (mm), where r is the radius of the needle 102 and Q is the flow rate at which the neutral collagen solution is discharged. ３ ・s -1 ) 4Q / πr ３ (unit: s －１ ) (π is the constant of the circumference of a circle). However, when the collagen concentration in the neutral collagen solution is high and the needle inner diameter is small, the actual Q may fall below the input value depending on the performance of the driving device. In such cases, it may be necessary to find the condition under which the winding speed of the winding roller 105 and the discharge speed of the collagen thread 1 are balanced, and to determine the actual Q.

[0039] As shown in Fig. 6, the manufacturing apparatus 100 preferably includes a take-up roller 105 for taking up the collagen thread 1 precipitated in the ethanol solution in the coagulation bath 104, and a motor 106 for driving the take-up roller 105, in order to perform the collagen thread drawing step S30. The manufacturing apparatus 100 further preferably includes a relay roller 107 that is disposed between the coagulation bath 104 and the take-up roller 105 and relays the collagen thread 1 in the ethanol solution to reach the take-up roller 105. By including the relay roller 107, the manufacturing apparatus 100 can wind the collagen thread 1 onto the take-up roller 105 without tangling during winding. Conventionally known rollers and motors can be used for the take-up roller 105, motor 106, and relay roller 107, as long as they each have the required functions.

[0040] <Steps Included in the Collagen Thread Manufacturing Method> (First Step: Preparation Step) The collagen thread manufacturing method according to this embodiment includes a step S10 of preparing a neutral collagen solution containing collagen at a concentration of 3.0% by mass or more and 5.0% by mass or less. The collagen is atelocollagen. The purpose of the preparation step S10 is to prepare the neutral collagen solution that will serve as the raw material for obtaining the collagen thread 1.

[0041] The neutral collagen solution can be prepared with an atelocollagen concentration of 3.0% by mass or more and 5.0% by mass or less by mixing an acidic atelocollagen solution with a concentration exceeding the target concentration with a neutral solvent (e.g., a salt-containing solvent such as phosphate buffer saline) using a conventionally known method. When the neutral collagen solution is discharged into a coagulation bath filled with an ethanol solution with a concentration of 70% by volume or more and 95% by volume or less in the collagen thread obtaining step S20 described below, the collagen molecules are instantaneously fibrillated in the coagulation bath, thereby obtaining collagen thread 1. As described above, the pH of the neutral collagen solution is preferably 6.0 to 9.0. The neutral collagen solution prepared in this step is contained in a container 101 (particularly a syringe) of the manufacturing apparatus 100. Air bubbles contained in the neutral collagen solution are preferably removed using a conventionally known method such as a centrifuge. Air bubbles may cause the collagen thread to break during manufacturing, resulting in a reduced yield.

[0042] If the collagen concentration in the neutral collagen solution is less than 3.0% by mass, it may be difficult for the collagen molecules to instantaneously fibrillate in the coagulation bath in step S20 of obtaining a collagen thread, and it may not be possible to obtain the desired collagen thread 1. If the collagen concentration in the neutral collagen solution exceeds 5.0% by mass, even when the liquid temperature is maintained at 10°C or below, the collagen may fibrillate in the neutral solvent, and it may not be possible to obtain the desired collagen thread 1. The neutral collagen solution preferably contains collagen (particularly atelocollagen) at a concentration of 4.5% by mass or more and 5.0% by mass or less.

[0043] (Second Step: Step of Obtaining Collagen Thread) The collagen thread manufacturing method according to this embodiment includes a step S20 of obtaining a collagen thread by discharging the neutral collagen solution into an ethanol solution having a concentration of 70% by volume or more and 95% by volume or less. In the step S20 of obtaining a collagen thread, the neutral collagen solution is passed through a flow path having an inner diameter of 100 μm or more and 300 μm or less and is then discharged for 100 s. -1 More than 1500s -1 The collagen is discharged into the ethanol solution at a shear rate of 100 s / min. The purpose of step S20 for obtaining a collagen thread is to align the collagen molecules in the neutral collagen solution in the flow direction through a small-diameter flow path, and then discharge them into the ethanol solution to instantaneously fiberize them, in order to obtain a collagen thread 1.

[0044] Specifically, the step S20 of obtaining collagen threads is carried out by the above-described manufacturing apparatus 100. In the step S20 of obtaining collagen threads, first, the neutral collagen solution contained in the container 101 of the manufacturing apparatus 100 is heated for 100 seconds. -1 More than 1500s -1 The neutral collagen solution is extruded by a plunger driven by a drive device 103 that generates a shear rate of 100 μm or more and 300 μm or less. As a result, the neutral collagen solution is ejected from the container 101 through a needle 102 with an inner diameter of 100 μm or more and 300 μm or less toward a coagulation bath 104. Furthermore, in the coagulation bath 104, the collagen (atelocollagen) in the neutral collagen solution is instantaneously fibrillated and precipitated as collagen thread 1.

[0045] If the concentration of the ethanol solution is less than 70% by volume, it may be difficult to instantaneously fibrillate the atelocollagen in the neutral collagen solution in the coagulation bath 104. If the concentration of the ethanol solution exceeds 95% by volume, the dehydration rate of the discharged neutral collagen solution may be too fast, making thread breakage more likely. If the inner diameter of the needle 102 in the manufacturing apparatus 100 is less than 100 μm, the pressure required for extrusion may be too high, making it difficult to obtain the desired shear rate, or the flow of the neutral collagen solution immediately after being discharged from the flow path may become unstable, causing thread breakage. If the inner diameter of the needle 102 exceeds 300 μm, it may be impossible to obtain a collagen thread with the desired thinness, or drying during winding may be difficult.

[0046] Furthermore, in the step S20 for obtaining collagen threads, the shear rate is 100 s -1 If the shear rate is less than 1500 s, the collagen molecules in the neutral collagen solution may not be aligned in the flow direction and ejected into the ethanol solution. -1 If the shear rate exceeds 300 s, the collagen molecules in the neutral collagen solution may not be aligned in the flow direction and may not be discharged into the ethanol solution due to the occurrence of a die swell effect in which the flow of the neutral collagen solution discharged from the flow channel expands. -1 More than 1000s -1 It is preferable that:

[0047] In the collagen thread obtaining step S20, the collagen thread 1 is finally removed from the coagulation bath 104 and wound up while drying. The collagen thread 1 removed from the coagulation bath 104 is wet with an ethanol solution, which immediately evaporates in the air. This allows the collagen thread 1 to be easily dried. The collagen thread 1 obtained by the above manufacturing method has collagen nanofibrils that are highly aligned throughout the collagen thread 1, preferably all the way to the core. Furthermore, the above manufacturing method does not, in principle, limit the length of the collagen thread 1. Additionally, by appropriately adjusting the inner diameter of the needle 102 and the shear rate, the diameter of the collagen thread 1 can be adjusted to 10 μm or more and 50 μm or less. By using atelocollagen for the collagen thread 1, safety for the living body can be ensured. As described above, a collagen thread that artificially reproduces collagen fibers found in living organisms can be obtained.

[0048] (Third Step: Step of Drawing the Collagen Thread) The collagen thread manufacturing method according to this embodiment preferably further includes a step S30 of drawing the collagen thread by winding the collagen thread at a winding speed of 1.5 to 4.0 times the linear extrusion speed. The purpose of the collagen thread drawing step S30 is three-fold: to make the collagen thread 1 thinner to the same extent as collagen fibers in vivo, to increase the degree of alignment of collagen fibrils, and to increase the production rate of the collagen thread.

[0049] Specifically, step S30 of stretching the collagen thread is performed by a manufacturing apparatus 100 as shown in FIG. 6 . That is, in the manufacturing apparatus 100, the collagen thread 1 precipitated in the coagulation bath 104 is relayed by a relay roller 107 and wound around a winding roller 105 driven by a motor 106. At this time, the winding speed at which the collagen thread 1 is wound around the winding roller 105 is preferably 1.5 to 4.0 times the linear discharge speed at which the neutral collagen solution is discharged from the container 101 into the coagulation bath 104 through the needle 102. The winding speed is more preferably 2.0 to 3.5 times the linear discharge speed. This allows the collagen thread 1 to be stretched when the collagen in the neutral collagen solution is spun while being precipitated as the collagen thread 1 in the coagulation bath 104, thereby thinning the diameter of the collagen thread 1 to, for example, 30 μm or less. Furthermore, the total length of the collagen thread 1 can be reduced by, for example, 1×10 6 The length can be more than twice as long. As a result, a thinner and longer collagen thread 1 can be obtained that artificially reproduces collagen fibers found in living organisms. The winding speed for winding the collagen thread 1 can be calculated from the rotation speed of the motor and the diameter of the roller. The linear discharge velocity when discharging the neutral collagen solution is calculated by calculating the flow rate Q (mm ３ ・s －１ ), where r (mm) is the radius of the flow path, Q / πr ２ (Unit: mm / s －１ = 3.6 m / h). However, as mentioned above, there are cases where it may be necessary to determine the actual Q.

[0050] The present invention will be described in more detail below with reference to examples, but the present invention is not limited to these examples. In these examples, the inventors produced collagen thread samples using a manufacturing apparatus such as that shown in Figure 5 or Figure 6. In the following description, Samples 1 to 6 are examples of collagen threads that fall within the scope of the present invention, and Samples 11 to 19 are comparative examples.

[0051] [Collagen Thread Production] <Sample 1> (Step 1: Preparation Step) First, an acidic atelocollagen solution (product name: "Collagen BM," manufactured by Nitta Gelatin Co., Ltd., collagen concentration: 0.571%) was prepared. Furthermore, to neutralize the acidic atelocollagen solution, a pH 7 buffer solution (12xNPB) was prepared, which was 12 times the concentration of a 50 mM sodium hydrogen phosphate buffer solution containing 140 mM sodium chloride (hereinafter referred to as 1xNPB). The acidic atelocollagen solution was concentrated in an evaporator at 29°C to a concentration of 2-8% by mass to obtain a concentrated collagen solution. The exact concentration of the concentrated collagen solution was determined from the dry weight of the acidic atelocollagen solution. The blending ratio of the concentrated collagen solution, pure water, and 12xNPB was appropriately adjusted to prepare the neutral collagen solution shown in Table 1, with a buffer concentration of 1xNPB. As a result, a neutral collagen solution containing collagen at a concentration of 4.8% by mass was prepared. Note that many neutral collagen solutions have poor fluidity and small volumes, making it difficult to measure the pH directly. Therefore, a solution (deemed solution) was prepared by replacing the concentrated collagen solution with dilute hydrochloric acid at pH 3. From this deemed solution, pure water and 12x NPB were added based on the blending ratio adjusted as described above to prepare a pH measurement solution, and the pH of this solution was measured with a pH meter. It was confirmed that the pH of the pH measurement solution was in the range of 7.1 to 7.4.

[0052] (Second step: step of obtaining collagen thread) Next, the neutral collagen solution contained in the container was ejected through a needle with an inner diameter of 182 μm toward a coagulation bath 104 filled with a 90% by volume ethanol solution (see FIG. 5). The shear rate at this time was 1000 s -1 These conditions corresponded to a linear extrusion velocity of 82 m / h. In the coagulation bath, the collagen (atelocollagen) in the neutral collagen solution was instantaneously fibrous, precipitated as collagen thread, and deposited in the coagulation bath. The collagen thread was then wound up on a winding roller. The collagen thread did not break or otherwise deteriorate during the process from extrusion to winding, and was evaluated to have good spinnability. As a result, the collagen thread of Sample 1 was obtained.

[0053] <Samples 2 to 4> The collagen solutions (neutral collagen solutions) shown in Table 1 were prepared, and the collagen threads of Samples 2 to 4 were obtained in the same manner as for producing the collagen thread of Sample 1, except that the needle inner diameter, shear rate, and extrusion linear velocity of the production equipment were set as shown in Table 1. The collagen threads of Samples 2 to 4 were evaluated to have good spinnability, with no breakage or other issues occurring in the processes from extrusion to winding.

[0054] <Sample 5> A collagen solution (neutral collagen solution) shown in Table 1 was prepared, and the needle inner diameter, shear rate, and extrusion linear velocity of the manufacturing apparatus were set as shown in Table 1. Furthermore, the collagen thread precipitated in the coagulation bath was wound up on a take-up roller driven by a motor in the manufacturing apparatus. The winding speed at which the collagen thread was wound up on the take-up roller was set to 2.0 times the extrusion linear velocity at which the neutral collagen solution was extruded from the container into the coagulation bath through the needle, and the collagen was stretched while the extruded collagen solution was being transformed into a thread (see Figure 6). The collagen thread of Sample 5 was otherwise obtained in the same manner as the collagen thread of Sample 1. The collagen thread of Sample 5 was evaluated to have good spinnability, with no breakage or other issues occurring during the process from extrusion to winding.

[0055] <Sample 6> The collagen thread of Sample 6 was obtained in the same manner as for producing the collagen thread of Sample 5, except that the winding speed at which the collagen thread was wound around the winding roller was set to 2.5 times the linear discharge speed at which the neutral collagen solution was discharged from the container into the coagulation bath through a needle. The collagen thread of Sample 6 did not break or otherwise occur in the processes from discharge to winding, and was evaluated to have good spinnability.

[0056] <Samples 11 to 12> Collagen solutions (neutral collagen solutions) shown in Table 1 were prepared, and collagen threads of Samples 11 to 12 were obtained in the same manner as for producing collagen thread of Sample 1, except that the needle inner diameter, shear rate, and extrusion linear velocity of the production equipment were set as shown in Table 1. As shown in Table 1, in the production of collagen threads of Samples 11 to 12, the shear rate was set to 2000 s -1 The collagen threads were evaluated as having good spinnability, with no breakage or other issues occurring during the process from extrusion to winding.

[0057] <Samples 13 to 15> Samples 13 to 15 were obtained by referring to the manufacturing method described in Patent Document 1. Specifically, the collagen concentration of the neutral collagen solution was set to 2.5%, a needle with a diameter of 521 μm was used, the shear rate and linear extrusion velocity were set as shown in Table 1, and the coagulation bath was changed from ethanol to a 10 mM phosphate buffer solution (heated to 37°C) with a pH of 7.0. The collagen thread of Sample 13 was obtained in the same manner as Sample 13, except that the collagen concentration of the neutral collagen solution was set to 5.0%. The collagen thread of Sample 15 was obtained in the same manner as Sample 14, except that the collagen concentration of the neutral collagen solution was set to 2.5%, and 90% ethanol was connected after the phosphate buffer solution as the coagulation bath. Samples 13 and 15 were evaluated as having good spinnability despite the low spinning speed. On the other hand, sample 14 was evaluated as having poor spinnability because the collagen thread frequently broke during winding.

[0058] <Samples 16 to 19> Collagen solutions shown in Table 1 (acidic collagen solutions with a pH of 3) were prepared, and collagen threads of Samples 16 to 19 were obtained in the same manner as for producing the collagen thread of Sample 1, except that the needle inner diameter, shear rate, and extrusion linear velocity of the production equipment were set as shown in Table 1. As shown in Table 1, in producing the collagen threads of Samples 16 and 17, the needle inner diameter was set to 1500 μm, the shear rate was set to 20 s -1In the preparation of the collagen thread of sample 16, the collagen concentration in the acidic collagen solution was set to 0.6% by mass, and in the preparation of the collagen thread of sample 17, the collagen concentration in the acidic collagen solution was set to 5.0% by mass. In the preparation of the collagen thread of sample 18, the collagen concentration in the acidic collagen solution was set to 0.6% by mass, and the shear rate was set to 20 s -1 These samples 16 to 19 were obtained as a reproduction experiment of the prior art wet spinning.

[0059] As a result, for sample 16, the collagen solution discharged into the coagulation bath fused together and precipitated, and a collagen thread could not be spun (poor spinnability). For sample 17, only the surface of the collagen thread became cloudy in the coagulation bath, suggesting that fiber formation had occurred, but the interior remained transparent for several minutes (poor spinnability). For sample 18, the collagen fused together in the coagulation bath before forming threads, forming clumps, and the thread could not be spun (poor spinnability). The collagen thread of sample 19 turned translucent, deposited at the bottom of the coagulation bath, and could then be wound up (good spinnability).

[0060] Table 1 shows a list of the preparation procedures (collagen solution, needle inner diameter, shear rate, extrusion linear velocity, spinnability, etc.) for the collagen threads of Samples 1 to 6 and Samples 11 to 19.

[0061]

[0062] [Evaluation of Collagen Threads] The diameter (average value and standard deviation), total length, and refractive index difference (average value and standard deviation) were determined for the collagen threads of Samples 1 to 6, 11 to 13, 15, and 19 using the methods described above. Furthermore, the strength of the collagen threads of Samples 1 to 6, 11 to 13, 15, and 19 was evaluated by measuring the Young's modulus (average value and standard deviation) using the following method. The results are shown in Table 2. The collagen threads of Samples 14 and 16 to 18 could not be wound up, so the above-mentioned items could not be evaluated.

[0063] Young's modulus was calculated from the initial slope of the stress-strain curve obtained from the tensile test. The collagen thread was cut to approximately 40 mm, and both ends were fixed with vinyl tape so that the sample length was 20 mm. The collagen thread was fixed to the chuck of a strength tester (product name "TA.XTplus", manufactured by Stable Micro Systems) with a chuck distance of 20 mm, and a strength test was performed at a deformation rate of 1 mm / sec to obtain a load-displacement curve. The load-displacement curve was converted to a stress-strain curve by calculating the stress by dividing the load by the cross-sectional area of ​​the collagen thread (average value of n = 5), which was measured separately using a scanning electron microscope, and by calculating the strain by dividing the displacement (mm) by the initial chuck distance (mm). Young's modulus was calculated from the slope of the linear region appearing in the strain range of 0.002 to 0.008 (n = 5). When the Young's modulus is 2.0 GPa or more, it is evaluated that the collagen thread has sufficient strength to be used as a biomaterial.

[0064]

[0065] [Discussion] The collagen threads of Samples 1 to 6 have a diameter of 10 μm or more and 50 μm or less, and the total length is 2×10 5 The refractive index difference was 4.3 × 10 -4 The collagen nanofibrils constituting each collagen thread were evaluated to be highly aligned along the longitudinal direction of the collagen thread. Furthermore, because the Young's modulus was 2.0 GPa or greater, the collagen threads were evaluated to have sufficient strength for use as biomaterials. The above value of 2.0 GPa corresponds to the elastic modulus of collagen threads obtained by classical wet spinning, in which collagen is sufficiently fibrillated in a neutral coagulation bath and then spun into threads. From the above, it was suggested that the collagen threads of Samples 1 to 6 could artificially reproduce collagen fibers found in the body and be used as biomaterials such as artificial tendons or membrane materials.

[0066] On the other hand, the collagen threads of Samples 11 and 12 have a diameter exceeding 50 μm and a refractive index difference of 4.3×10 -4The collagen thread of sample 13 had a diameter of more than 50 μm, a total length of 0.2 to 0.5 m, and a refractive index difference of 4.3 × 10 -4 The collagen thread of sample 15 was not subjected to diameter and tensile tests because the thread significantly shrunk when dried and was clearly not useful as a biomaterial. The collagen thread of sample 15 also had a refractive index difference of 4.3 × 10 -4 The collagen thread of sample 19 had a diameter of more than 50 μm and a refractive index difference of 4.3×10 -4 It was impossible to wind the collagen threads of Sample 14 and Samples 16 to 18 into threads. Therefore, none of the collagen threads of Samples 11 to 19 could be evaluated as artificially reproducing collagen fibers in vivo.

[0067] Although the embodiments and examples of the present invention have been described above, it is also intended from the beginning that the configurations of the above-described embodiments and examples may be appropriately combined.

[0068] The embodiments and examples disclosed herein should be considered to be illustrative in all respects and not restrictive. The scope of the present invention is defined by the claims, not by the above description, and is intended to include all modifications within the meaning and scope of the claims.

[0069] 1 collagen thread, 11 collagen nanofibril, 10 hydrated collagen thread, 200 artificial tendon, 100 manufacturing apparatus, 101 container, 102 needle (flow path), 103 driving device, 104 coagulation bath, 105 winding roller, 106 motor, 107 relay roller.

Claims

1. A collagen thread containing collagen nanofibrils, the diameter of the collagen thread being 10 μm or more and 50 μm or less, and the total length of the collagen thread being 2×10 5 the collagen nanofibrils have a length of 4.3×10 or more times that of the hydrated collagen yarn, the collagen nanofibrils exhibit high alignment along the longitudinal direction of the collagen yarn, and the high alignment is such that the refractive index difference determined by birefringence measurement of the hydrated collagen yarn is 4.3×10 -4 The hydrated collagen thread is obtained by immersing the collagen thread in a neutral phosphate buffer solution.

2. The collagen yarn of claim 1, wherein the collagen nanofibrils are composed of atelocollagen.

3. The total length is 1 x 10 to the diameter. 6 The collagen thread according to claim 1 or 2, having a length that is at least twice as long.

4. A process for obtaining a collagen thread by discharging a neutral collagen solution containing collagen at a concentration of 3.0% by mass or more and 5.0% by mass or less, the neutral collagen solution being discharged into an ethanol solution having a concentration of 70% by volume or more and 95% by volume or less, the collagen being atelocollagen, in the process for obtaining a collagen thread, the neutral collagen solution being passed through a flow path having an inner diameter of 100 μm or more and 300 μm or less for 100 s. -1 More than 1500s -1 The method for producing collagen thread, wherein the collagen fiber is extruded into the ethanol solution at a shear rate of:

5. The method for producing collagen thread described in claim 4, further comprising a step of stretching the collagen thread by winding the collagen thread at a winding speed of 1.5 to 4.0 times the linear extrusion speed.

6. A method for producing collagen thread as described in claim 4 or claim 5, wherein the neutral collagen solution contains the collagen at a concentration of 4.5 mass% or more and 5.0 mass% or less.

7. The shear rate is 300 s -1 More than 1000s -1 The method for producing a collagen thread according to any one of claims 4 to 6, wherein the method is as follows:

8. A biomaterial comprising the collagen thread according to any one of claims 1 to 3.

9. The biomaterial of claim 8, wherein the biomaterial is an artificial tendon or membrane material.

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