WO2025188632A1

WO2025188632A1 - Molded body and manufacturing method for the same

Info

Publication number: WO2025188632A1
Application number: PCT/US2025/018152
Authority: WO
Inventors: Kota YAGI
Original assignee: Canon Virginia Inc
Current assignee: Canon Virginia Inc
Priority date: 2024-03-04
Filing date: 2025-03-03
Publication date: 2025-09-12
Anticipated expiration: 2026-09-04
Also published as: WO2025188632A8

Abstract

Provided are a molded body containing fibroin and method for manufacture thereof. The molded body includes a first region located at a surface of the molded body, and a second region located at least 0.1 mm beneath the surface of the molded body, with a degree of crystallization of the molded body at the first region being at least 60%, and the degree of crystallization of the molded body at the second region being between 35% and 50%.

Description

MOLDED BODY AND MANUFACTURING METHOD FOR THE SAME

CROSS REFERENCE TO RELATED APPLICATIONS

[001] This application claims priority to U.S Provisional Patent Application No. 63/560,988, filed with the U.S. Patent and Trademark Office on March 4, 2024, the entire disclosure of which is incorporated herein by reference.

FIELD OF THE DISCLOSURE

[002] The present disclosure relates to a molded body composed of fibroin and a method for manufacturing a molded fibroin body.

BACKGROUND OF THE DISCLOSURE

[003] These days, efforts are being made to process highly biocompatible fibroin into various products. For example, U.S. Pub. 2021/0381129 Al to Kaplan et al. discusses obtaining a molded fibroin body having a desired degree of crystallization and desired material properties by applying at least one high temperature and high pressure to the fibroin material when manufacturing the molded fibroin body.

[004] The molded fibroin body of conventional methods may provide a high degree of crystallization on an outermost surface. However, the degree of crystallization on the inside of the molded body, that is not in direct contact with the molding die, is low. Therefore, the strength of the molded part, especially impact strength, may be reduced.

SUMMARY

[005] To address such exemplary needs and to overcome shortcomings of conventional devices, systems and methods, the presently disclosure provides a molded body having a high degree of internal crystallization and a high impact strength, using fibroin. The present disclosure also provide a method of manufacture of a molded body having a high degree of internal crystallization and a high impact strength, using fibroin.

[006] An aspect of the present disclosure provides a molded body containing fibroin, with the molded body including a first region located at a surface of the molded body, and a second region located at least 0.1 mm beneath the surface of the molded body, with a degree of crystallization of the molded body at the first region of at least 60%, and with the degree of crystallization of the molded body at the second region between 35% and 50%.

[007] Another aspect of the present disclosure provides a method of manufacturing a molded fibroin body that includes loading fibroin in a mold and maintaining a temperature of the mold between 40°C and 50°C.

[008] A further aspect of the present disclosure provides a method of manufacturing a molded fibroin body that includes loading fibroin in a mold, maintaining a temperature of the mold between 40°C and 50°C for between five and thirty minutes, raising the temperature of the mold to between 125°C and 180°C, and applying, to the fibroin, a pressure of at least lOMPa for at least five minutes.

[009] These and other objects, features, and advantages of the present disclosure will become apparent upon reading the following detailed description of exemplary embodiments of the present disclosure, when taken in conjunction with the appended drawings, and provided paragraphs.

BRIEF DESCRIPTION OF THE DRAWINGS

[0010] The accompanying drawings, which are incorporated herein and form part of the specification, illustrate various embodiments, objects, features, and advantages of the present disclosure.

[0011] FIG. 1 illustrates a crystallization process of fibroin in a conventional molding process.

[0012] FIG. 2 illustrates a crystallization process of fibroin in the molding process according to the present disclosure.

[0013] FIG. 3 is a schematic diagram of a mold.

[0014] FIG. 4 is a flowchart of a molding process of a molded fibroin body according to the present disclosure. [0015] FIG. 5 illustrates degree of crystallization distribution and degree of crystallization of a molded body according to the present disclosure.

[0016] FIG. 6 shows average degree of crystallization distribution and degree of crystallization of a molded body according to the present disclosure.

[0017] FIG. 7 summarizes aspects of various processes for various embodiments.

[0018] FIG. 8 illustrates steps of molding silk fibroin.

[0019] FIG. 9A summarizes steps of a conventional process.

[0020] FIG. 9B is a bar chart summarizing results of bending tests performed on an output of the conventional process.

[0021] FIG. 9C is a bar chart summarizing results of impact tests performed on an output of the conventional process.

[0022] FIG. 10 is a graph of crystallization degree according to conventional method.

[0023] FIG. 11 illustrates internal water evaporation according to conventional method.

[0024] FIG. 12 illustrates internal water evaporation according to the present disclosure.

[0025] FIG. 13 is a graph comparing crystallization degree by depth obtained using conventional method and using the method of the present disclosure.

[0026] FIG. 14 provides bar charts of comparative impact strengths.

[0027] Throughout the figures, the same reference numerals and characters, unless otherwise stated, are used to denote like features, elements, components or portions of the illustrated embodiments. Moreover, while the subject disclosure will now be described in detail with reference to the figures, it is done so in connection with the illustrative exemplary embodiments. It is intended that changes and modifications can be made to the described exemplary embodiments without departing from the true scope and spirit of the subject disclosure as defined by the appended claims.

DETAILED DESCRIPTION

[0028] The present disclosure has several embodiments and relies on patents, patent applications and other references for details known to those of the ai t. Therefore, when a patent, patent application, or other reference is cited or repeated herein, it should be understood that it is incorporated by reference in its entirety for all purposes as well as for the proposition that is recited.

[0029] The molded fibroin body of the present embodiment possesses a first region having a degree of crystallization of 60% or more on the surface of the molded body, and possesses a second region having a degree of crystallization of 35% to 50% in the part deeper than 0.1 mm from the surface of the molded body. In addition, it is possible to obtain the molded fibroin body of the present embodiment by heat die compression molding fibroin material.

[0030] Hereinafter, embodiments are described for fibroin material, heat die compression molding, and manufacture of a molded body.

[0031] Fibroin material can be obtained by converting degummed fibroin from which sericin is removed into an aqueous solution and drying the aqueous solution.

[0032] Fibroin is a component contained in silkworm cocoons, spider silk, hornet silk (a substance produced by wasp larvae), etc., and 70% or more is biological polymeric material composed of the amino acids: glycine and alanine. Fibroin is considered to be superior in terms of biocompatibility and biodegradability. In the case of natural silkworm cocoons (raw silk), generally 20% to 30% is sericin and 70% to 80% is fibroin. In addition, not only animal-derived fibroin, but also plant-derived fibroin and fibroin obtained by DNA modification of these can be used.

[0033] (Aqueous Solution)

[0034] An aqueous solution of fibroin can be obtained by, for example, the method of producing the aqueous solution of silkworm silk discussed in WO 2006/101223 or by the method of producing the aqueous solution of spider silk discussed in U.S. Pat. 5,245,012. When using these, an aqueous solution in which hydrophilic fibroin is uniformly dissolved can be obtained.

[0035] (Drying of Aqueous Solution)

[0036] The fibroin material in this embodiment is a powdered fibroin molecule in an amorphous state. Amorphous fibroin can be manufactured using a process of dissolving degummed fibroin in an aqueous solution and then removing the water from the aqueous solution. Although the process for removing water from an aqueous solution is not limited to a specific process, since dewatering by simple heating can accelerate the crystallization of fibroin and damage its meltability during molding, and methods may be used such as lyophilization, which allows dewatering while cooling, and spraydrying, for which the time of applying heat to the aqueous solution is extremely short.

[0037] When using the lyophilization method, the freezing temperature is not particularly limited, as long as the aqueous solution freezes, but because the freezing point of an aqueous solution in which a solute is dissolved is lower than the freezing point of water, it is preferably -10°C or lower, and more preferably -20°C or lower. When drying under reduced pressure after freezing, the temperature is preferably -20°C or higher, and more preferably -15°C or higher, because the time required for dewatering becomes longer the lower the drying temperature is. In a sufficiently frozen sample, the pressure may be approximately 600 Pa or lower, which is the pressure at the triple point of water, but 100 Pa or lower is preferable, and 50 Pa or lower is more preferable. This reduces foaming which can occur if the sample is partially liquefied. After the water has been removed from the sample, the low-pressure state is preferably released after the temperature reaches 10°C or higher. This is because if the temperature is lower than this, condensation may occur inside the sample. This is not desirable because when condensation occurs, the fibroin may gel due to adsorption of water into the dry fibroin body.

[0038] When using spray drying, the concentration of fibroin in the aqueous solution may be adjusted in advance to facilitate spraying of the sample. It is preferable to shorten the time that the sprayed sample is exposed to heat by adjusting the air pressure or flow rate of the spray. To avoid heating and crystallizing of the fibroin as a result of the equipment itself heating during spraying, spraying may be conducted while the periphery of the equipment or the sample recovery area are cooled with a cooling device or the like.

[0039] To turn it into a powder, a method may be used of pulverizing by, for example using a jet mill, a hammer mill, a ball mill, a pin mill, or the like, after lyophilization, or a method of using the powder obtained by spray drying can be used. The powdered fibroin material preferably contains 5% to 20% water content to improve flowability during fibroin molding and promote crystallization.

[0040] In addition, amorphous in this embodiment is a degree of fibroin crystallization of 20% or less. The degree of crystallization is calculated by the following method.

[0041] (Method for Measuring the Degree of Crystallization)

[0042] The degree of crystallization of the molded part may be determined as follows. Measure the infrared absorption spectrum of the surface of the molded body every 1 cm'¹ in the range from 1580 cm"¹ to 1720 cm"¹. Calculate the linear function passing through the value at 1580 cm"¹ and the value at 1720 cm"¹ of the obtained spectrum, make it the baseline, and subtract it from the measured values. Regard the obtained spectrum as the sum of four spectra derived from random coil, [3- sheet I, P-sheet II, and P-turn, and refer to it as the effective spectrum.

[0043] A spectrum derived from a random coil is set as a Gaussian distribution of which the peak center is any value between 1645 cm"¹ and 1655 cm"¹. Spectra derived from P- sheet I, P-sheet II, and P-turn are set as Gaussian functions of which the peak centers possess the fixed values of 1620, 1698, and 1685 cm"¹, respectively. In the four Gaussian functions, peak intensities and deviations of the Gaussian functions are set as variables. The sum of these four Gaussian functions is set as a synthetic spectrum having nine variables.

[0044] In the range of 1580 cm"¹ to 1720 cm"¹, determine the absolute value of the difference between the effective spectrum and the synthesized spectrum every 1 cm"¹, calculate the sum thereof, and set it as the spectral error. Set the spectral error with respect to the integral value of the effective spectrum as the error ratio, and fit the nine variables of the synthetic spectrum so the error ratio becomes small. When the error ratio after fitting becomes 3% or less, set it as a converged ratio, and regal'd the four spectra constituting the synthetic spectrum as the actual spectra of each component. Finally, calculate the sum of the three spectral integral values derived from P-sheet I, P- sheet II, and P-tum with respect to the integral value of the effective spectrum in the range from 1580 cm"¹ to 1720 cm"¹ as the degree of crystallization in the present embodiment. Fitting can be executed using the software accompanying the infrared absorption spectrum measuring device, the graph analysis software, the solver add-in in Microsoft Excel software, or the like.

[0045] (Bound Water)

[0046] U.S. Pub. 2021/0381129 Al to Kaplan et al. discusses that the structural change of silk fibroin requires the presence of two types of bound water. The first type is weakly bound water which starts to evaporate around 35 °C and reaches its maximum evaporation speed at 95°C. The other type is strongly bound water which starts to evaporate around 55°C and reaches its maximum evaporation speed at 125°C.

[0047] In the present disclosure, evaporation of these two types of bound water is utilized as the fibroin material changes its structure and crystallizes. However, evaporation of the bound water inside the molded body is inhibited by the change in the degree of surface crystallization of the molded body.

[0048] FIG. 1 illustrates a crystallization process of fibroin in a conventional heat die compression molding process.

[0049] In Step 1 of FIG. 1, the fibroin material is heated to a normal temperature of 25°C in a mold whose temperature has been adjusted to 145°C. Step 2 of FIG. 1 illustrates a state in which the temperature of the fibroin material is raised from the surface, the surface temperature reaches 125 °C, while a temperature distribution is generated from the surface to the interior, and the bound water on the surface evaporates actively. Step 3 of FIG. 1 illustrates a state in which crystallization only takes place near the surface where the bound water has evaporated. Step 4 of FIG. 1 illustrates a state in which the evaporation of internal bound water is inhibited by crystallized fibroin material. Step 5 of FIG. 1 illustrates a state in which only the surface is highly crystallized, and the internal crystallization rate remains low. Thus, in conventional heat die compression molding process of fibroin material, the evaporation of bound water inside the molded body is inhibited by crystallized material on the surface, and crystallization inside the molded body does not proceed, resulting in low impact strength. [0050] FIG. 2 illustrates a crystallization process of fibroin in the molding process according to the present disclosure.

[0051] As illustrated in step 1 the crystallization process of fibroin material of FIG. 2, heating stalls of normal temperature fibroin material of 25°C in a mold whose temperature has been adjusted to 50°C, which is a temperature at which strongly bonded water does not start to evaporate. Step 2 of FIG. 2 illustrates a state in which the overall temperature of the fibroin material has become 50°C. In step 3 of FIG. 2, heating of the fibroin material, whose temperature is 50°C overall, starts in a mold whose temperature has been adjusted to 145°C. Step 4 of FIG. 2 illustrates a state in which the temperature of the fibroin material is raised from the surface. Because, compared to a conventional process, the difference in time required to reach 125°C on the surface of the molding and on the inside of the molding is smaller, not only the bound water on the surface, but also water bound within the molded body actively evaporates. Step 5 of FIG. 2 illustrates the inside of the molded body being highly crystallized.

[0052] Thus, in the present disclosure, adding a process of temperature control prior to the start of heat die compression, increases the amount of evaporation of bound water inside the molded body, improving the degree of crystallization further inside the molded body. As described herein, the impact strength of the molded body is increased.

[0053] < Heat die compression >

[0054] The molded fibroin body, in the present embodiment, can be obtained by heat die compression molding, in which the powdered fibroin material is fed into a molding die, the temperature of the powdered fibroin material is raised to a certain temperature by initial heating, and heat and pressure is applied to the mold for the molding while the initial temperature is maintained.

[0055] (Initial Heating)

[0056] Initial heating is a process in which the temperature of the material is raised to a specified temperature prior to heat die compression molding.

[0057] FIG. 3 is a schematic diagram of a mold. In particular, FIG. 3 illustrates a mold that can be used for the initial heating of a fibroin material. The mold is composed of an upper punch 1, a lower punch 2, and a frame form 3, and after feeding the fibroin material 4 into the frame form 3, insert the upper punch 1 and the lower punch 2 through the opening of the frame form 3, and the fibroin material 4 can be compressed. A hot plate is brought into contact with the upper punch 1 and the lower punch 2, and the temperature adjustment of this hot plate makes it possible to heat the fibroin material 4 to the appropriate temperature. In addition to the hot plate, a heater directly grounded to the mold or a heating furnace, or the like, can also be used as a means for heating. The means of heating is not particularly limited as long as it is a temperature raising means capable of appropriate temperature adjustment. The temperature of the mold in the initial heating process is preferably from 40°C to 50°C, and 50°C is most preferable. Below 40°C, the effect of introducing initial heating may not be sufficiently obtained, and sufficient strength may not be obtained. On the other hand, at temperatures higher than 50°C, evaporation of the strongly bound water bound to the fibroin may begin. Therefore, due to a lack of moisture required for crystallization during heat die compression molding, the degree of crystallization may be low and sufficient strength may not be obtained. It is preferable that the pressure at the time of pressurization in the initial heating process is conducted from the atmospheric pressure or more to 1 MPa. At 1 MPa or more, crystallization of the fibroin material will start due to the pressurization, but sufficient strength may not be obtained because the degree of crystallization inside the molded body is not high enough. For example, when the pressure in the initial heating process was set to 10 MPa, it was confirmed that the impact strength was reduced by approximately 55% compared to when it was set to 1 MPa. In addition, the time for maintaining the specified temperature in the initial heating process is preferably 5 to 30 minutes, and most preferably 10 to 20 minutes. In the case of 5 minutes or shorter, the temperature of the fibroin material overall may not change completely, and sufficient strength may not be obtained. On the other hand, when heating it for 30 minutes or longer, due to the effect that the amount of evaporation of weakly bound water which evaporates starting at 35°C increases, the progress of crystallization during heat die compression (this molding) may not be enough, and sufficient strength may not be obtained. The above parameters such as temperature, pressure, and time only exemplify typical numerical values that are particularly preferable, and the present disclosure is not limited to these preferable conditions.

[0058] (Heat Die Compression Molding)

[0059] With the initially heated fibroin material 4 placed in the mold used in the initial heating is further heated or pressurized, a molded body having a high crystallinity may be obtained.

[0060] The temperature of the mold in the heating and pressurizing process is preferably from 125°C to 180°C, and from 145°C to 170°C is even more preferable. At 125°C or lower, crystallization does not progress sufficiently, due to the fact that the evaporation of strongly bound water does not take place actively, and sufficient strength may not be obtained. On the other hand, at temperatures higher than 180°C, decomposition of fibroin starts, and sufficient strength may not be obtained.

[0061] The pressure when pressurizing is preferably 10 MPa or more. Below 10 MPa, the fibroin may not integrate sufficiently.

[0062] In addition, after reaching the specified pressure, the time for maintaining said pressure is preferably 5 minutes or longer. If it is shorter than 5 minutes, the time for crystallization of the fibroin to proceed is short, and sufficient strength may not be obtained. The above parameters such as temperature, pressure, and time only exemplify typical numerical values that are particularly preferable, and the present disclosure is not limited to these preferable conditions.

[0063] < Molded Body >

[0064] The molded body in this embodiment has a distribution in the degree of crystallization. The degree of crystallization in the region near the surface (the first region) is 60% or more. In addition, the degree of crystallization of the region (the second region) having a depth equal to or greater than a specified depth from the vicinity of the surface, specifically the region having a depth of 0.1 mm or more from the outermost surface of the molded body, is 35% to 50%. In other words, the molded body of the present embodiment has a region where the degree of crystallization is 35% to 50% in the region having a depth deeper than 0.1 mm from the outermost surface, which is referred to as the second region. [0065] Thus, since the molded body of the present embodiment has a distribution of the degree of crystallization from the surface to the inside of the molded body, and also has a second region having a certain degree of crystallization or more even at a certain depth or more, the impact strength can be improved.

[0066] To maintain a higher impact strength, it is preferable that the second region exists possessing a certain thickness. Specifically, it is preferable that the second region exists possessing a thickness of 0.25 mm or more. An upper limit is not particularly set for the thickness of the second region, but a thickness of about 5.0 mm, for example, is deemed to be sufficient.

[0067] (Measurement of Internal Degree of Crystallization)

[0068] The measurement of the degree of crystallization of the inside of the molded body may be performed by any method capable of measuring the degree of crystallization of the inside of the molded body, such as a method in which a cross section of the molded body is taken out and a selected point of the cross section is measured using micro IR, or a method in which the surface of the molded body is gradually shaved using a processing machine such as a microtome and the infrared absorption spectrum of the shaved cross section and the shaved flake is measured using U-ATR every time the surface of the molded body is shaved for a certain distance.

[0069] (Mechanical Properties)

[0070] Confirmation of mechanical properties of the molded body was obtained using a Charpy impact strength test in accordance with ISO 179-1 and using a flexural- properties test in accordance with ISO178. In the present embodiment, the strength of the molded body is preferably an impact strength of 1 kJ/m ² or more and a maximum flexural property of 110 MPa or more, and more preferably an impact strength of 1.5 kJ/m ² or more and a maximum flexural property of 130 MPa or more.

[0071] [Examples]

[0072] FIG. 4 is a flowchart of a molding process of a molded fibroin body according to the present disclosure.

[0073] [Fibroin powder is preparation] [0074] In step 401 of FIG. 4, the fibroin powder is prepared. After washing with water, the silkworm cocoons were boiled in a 0.02 mol/L sodium carbonate aqueous solution for 30 minutes to conduct degumming. The degummed cocoons were put into a 9.3 mol/L LiBr aqueous solution and dissolved by conducting 4 hours of agitation at 60°C. Cellulose tubes 30/32 (fractionated molecular weight of 12000-14000) manufactured by Sekisui Chemical Co., Ltd. were used for desalination. After further dilution with pure water, it was confirmed that the aqueous solution is homogeneous without precipitate in a visual inspection.

[0075] After the aqueous fibroin solution obtained as described above was spread on a tray so the thickness became 5 mm, the tray was placed on a shelf in a freeze dryer (Model No. FD-550P, manufactured by Tokyo Electric Machine) and cooled to -30°C. After 3 hours, and after freezing was confirmed in a visual inspection, decompression was started at -6°C and 30 Pa, and dewatering was started. After 24 hours, decompression was released after letting the sample temperature rise to 15 °C, and the sheet- shaped dried body was taken out.

[0076] After that, it was pulverized by grinding it for 30 seconds in a mill apparatus (Crash Milser IFM-C 20G, manufactured by Iwatani Industries).

[0077] When the infrared absorption spectrum of the powder was measured with the ATR method using an FT-IR/NIR spectrometer Frontier, the crystallization rate was determined to be 10%, and it was confirmed to be an amorphous body. This was used as a powdered fibroin material 4.

[0078] [Initial Heating Process]

[0079] In step 402 of FIG. 4, a frame form 3 in which a hole of 10 mm in the shorter side direction, 85 mm in the longer side direction, and 50 mm in depth was machined, was prepared, and 4.8 g of powdered fibroin material 4 was poured into the frame form 3 in a state in which a lower punch 2 slidably fitted to the frame form 3 was inserted. After pouring completion, an upper punch 1 was inserted into the opening of the frame form 3. With the upper punch 1 and the lower punch 2 inserted into the frame form 3, it was moved to a hand press. The temperature of a hot plate installed in the hand press was adjusted to 50°C, and the fibroin material 4 was compressed to become a molding pressure of 1 MPa. At this time, the temperature of the hot plate transmits to the abutting upper punch 1 and lower punch 2, and as a result the fibroin material 4 is initially heated. The compression time was set to 10 minutes from the start of compression. After 10 minutes had passed, the pressure of the press was released, the temperature adjusted mold was immediately moved to the press for molding, and the molding process of step 3 (the heat press or the molding process) was conducted.

[0080] [Molding Process]

[0081] For the molding process of step 403 of FIG. 4, a hot plate was also installed in the press. The hot plate temperature was temperature adjusted to be 145°C. The mold described above which was moved immediately to the press for molding after the completion of step 2 compressed the fibroin material 4 so the molding pressure became 100 MPa. In this process, the fibroin material 4 was heated up to 145°C. The compression time was set to 15 minutes from the start of compression, the pressure of the press was released after 15 minutes had passed, and the mold was taken out of the press for molding and air-cooled. When the temperature of the mold had cooled to 30°C or lower, the mold was disassembled and the molded body was taken out. The size of the molded body was 10 mm in the shorter side direction, 85 mm in the longer side direction, approximately 4 mm thick, and it was orange with a translucent appearance. Three molded bodies of the same condition were obtained by the same procedure.

[0082] [Step A. Degree of Crystallization Measurement Process]

[0083] The degree of crystallization of the outermost surface of the molded body obtained in step 403 was measured. When the infrared absorption spectrum was measured with the ATR method using the FT-IR/NIR spectrometer Frontier for the measurement, the crystallization rate of the outermost surface, i.e., the first region, was determined to be 81 .5%.

[0084] Next, the molded body was machined to measure the degree of crystallization inside the molded body. A sample of 5 mm x 5 mm x 4 mm was prepared by cutting out the center 5 mm of the 10 mm of the shorter side direction and the center 5 mm of the 85 mm of the longer side direction of the molded body. [0085] Using a manual rotary microtome manufactured by Leica Microsystems, flakes of 10 m were cut from the outermost surface. By collecting the flakes at every 5 cuts, flake groups of 50 pm each were obtained for a total of 40 areas until the thickness center reached 2 mm. At this time, the flakes of the outermost surface were removed.

[0086] FIG. 5 illustrates degree of crystallization distribution and degree of crystallization of a molded body according to the present disclosure.

[0087] The results of FIG. 5 are for measuring the infrared absorption spectrum of the flake samples for the above-mentioned 40 areas worth with the ATR method. As a trend of the degrees of crystallization, the outermost surface is the highest with 81.5%, the degree of crystallization rapidly decreases to 43.7% just by going 50 pm to the inside, and after that it maintains a degree of crystallization of 35% to 50% from 50 pm to 850 pm. That is, the first region of this example is thinner than 0.05 mm. Further, there is a region having a degree of crystallization of 35% or more and 50% or less in a region deeper than 0.1 mm from the outermost surface of the molded body of this example. Next, the degree of crystallization begins to decrease again from 900 pm, and it reduces to the center at 2000 pm, with values ranging between 20% and 40%.

[0088] FIG. 6 shows average degree of crystallization distribution and degree of crystallization of a molded body according to the present disclosure.

[0089] To plot the overall trend in order to reduce the variation of the results of the above-mentioned distribution of the degree of crystallization, the depth was divided into 8 areas of 250 pm each, and the plotted results of the average values of 5 depths belonging to each area are shown in FIG. 6. By such averaging processing, it was clearly observed that the degree of crystallization of the molded body gradually reduced with increasing depth from the surface.

[0090] [Step B. Evaluation Process of Mechanical Properties]

[0091] A notching machine (Model No. 189-PN, manufactured by Yasuda Seiki Seisakusho, Ltd.) was used on the second molded body, and a notch was made in the molded body to a depth of 2 mm. After that, the molded body was set in a Charpy impact tester (Model No. 258 D, manufactured by Yasuda Seiki Seisakusho, Ltd.), and when the impact test was performed, the impact strength was determined to be 1.59 kJ/m².

[0092] The flexural strength of the third molded body was measured using a universal material tester (Model 5582, manufactured by Instron Japan Co., Ltd.), at a pressing speed of 2 mm/min. The maximum bending stress was determined to be 132.1 MPa.

[0093] [EXAMPLE 2]

[0094] FIG. 7 summarizes aspects of various processes for various embodiments.

[0095] The heating and compression time in step 402 of Example 1 was set to 5 minutes. The other steps were performed in the same manner as in Example 1 . The degree of crystallization of the obtained molded body was as shown in FIG. 7, resulting in an impact strength of 1.08 kJ/m² and a maximum bending stress of 122.3 MPa.

[0096] [EXAMPLE 3]

[0097] The heating and compression time in step 402 of Example 1 was set to 30 minutes. The other steps were performed in the same manner as in Example 1. The degree of crystallization and mechanical properties of the obtained molded body were as shown in FIG. 7, resulting in an impact strength of 1.42 kJ/m^2, and a maximum bending stress of 128.4 MPa.

[0098] [EXAMPLE 4]

[0099] The heating temperature in step 402 of Example 1 was set to 40°C. The other steps were performed in the same manner as in Example 1. The degree of crystallization and mechanical properties of the obtained molded body were as shown in FIG. 7, resulting in an impact strength of 1.19 kJ/m² and a maximum bending stress of 125.3 MPa.

[00100] [COMPARATIVE EXAMPLE 1 ]

[00101] The initial heating process of step 402 of Example 1 was omitted. The other steps were performed in the same manner as in Example 1. The degree of crystallization and mechanical properties of the obtained molded body were as shown in FIG. 7, resulting in an impact strength of 0.85 kJ/m² and a maximum bending stress of 121.2 MPa. [00102] [COMPARATIVE EXAMPLE 2]

[00103] The heating and compression times in step 402 of Example 1 were set to 60 minutes. The other steps were performed in the same manner as in Example 1 . The degree of crystallization and mechanical properties of the obtained molded body were as shown in FIG. 7, resulting in an impact strength of 0.94 kJ/m² and a maximum bending stress of 120.6 MPa.

[00104] [COMPARATIVE EXAMPLE 3]

[00105] The heating temperature in step 402 of Example 1 was set to 60°C. The other steps were performed in the same manner as in Example 1. The degree of crystallization and mechanical properties of the obtained molded body were as shown in FIG. 7, resulting in an impact strength of 0.76 kJ/m² and a maximum bending stress of 38.1 MPa.

[00106] FIG. 8 illustrates steps of molding silk fibroin. As shown in FIG. 8, fibroin material may be obtained by converting degummed fibroin from which sericin is removed into an aqueous solution and drying the aqueous solution.

[00107] FIG. 9A summarizes steps of a conventional process. In step 901, spraydrying is performed to obtain powder.

[00108] In step 902, the powder is filled into molds. In step 903, the powder is pre-processed and, in step 904, the mold is changed for pressing. In step 905, heating and pressing are performed. In step 906, the mold is colling and the bar is removed in step 907.

[00109] FIG. 9B is a bar chart summarizing results of bending tests performed on an output of the conventional process.

[00110] FIG. 9C is a bar chart summarizing results of impact tests performed on an output of the conventional process.

[00111] As illustrated in FIG. 9B, silk provides superior bending results. However, as illustrated in FIG. 9C, the impact strength of silk is not sufficient.

[00112] FIG. 10 is a graph of crystallization degree according to conventional method. As illustrated in FIG. 10, low crystallization results in weak strength.

[00113] FIG. 11 illustrates internal water evaporation according to conventional method. As illustrated in FIG. 11 , water removal from fibroin is needed for crystallization. Conventional methods provide surface heating, resulting in rapid crystallization. In conventional methods, surface crystallization blocks evaporation from within. Thus, internal crystallization remains low, even after pressing is complete.

[00114] FIG. 12 illustrates internal water evaporation according to the present disclosure. In FIG. 12, pre-heating is introduced and a temperature difference between the surface and internal regions is reduced. Crystallization of inner areas occurs sooners and internal water evaporation is not blocked, improving internal crystallization.

[00115] FIG. 13 is a graph comparing crystallization degree by depth obtained using conventional method and using the method of the present disclosure. As shown in FIG. 13, the method of the present disclosure provides improved degree of crystallization.

[00116] FIG. 14 provides bar charts of comparative impact strengths.

[00117] On the left side, FIG. 14 provides a bar chart of impact strength obtained according to the conventional method. On the right side, FIG. 14 provides a bar chart of impact strength obtained according to the present disclosure. Comparison of the charts illustrates improved impact strength obtained by the present disclosure.

[00118] Accordingly, the present disclosure provides a molded body containing fibroin, with the molded body comprising a first region located at a surface of the molded body, and a second region located at least 0.1 mm beneath the surface of the molded body, wherein a degree of crystallization of the molded body at the first region is at least 60%, and wherein the degree of crystallization of the molded body at the second region is between 35% and 50%. In the molded body, the degree of crystallization decreases from the surface to a center of the molded body. In the molded body, a thickness of the second region is between 0.25mm and 5.0mm. In the molded body, a thickness of the first region may be 0.05mm or less. The molded body has a predefined region where the degree of crystallization is 35% or less. The predefined region of the molded body may be located 0.5mm beneath the surface of the molded body. A thickness of the molded body may be between 0.5mm and 10.0mm.

[001 19] The present disclosure also provides a method of manufacturing a molded fibroin body that includes loading fibroin in a mold and maintaining a temperature of the mold between 40°C and 50°C. Pressure is applied to the fibroin to maintain the temperature of the mold above 40°C. The applied pressure may be IMPa or less. The method may include raising the temperature of the mold to between 125°C and 180°C. The method may include applying, to the fibroin, a pressure of at least lOMPa for at least 5 minutes. The fibroin loaded in the mold may have a water content between 5% and 20%.

[00120] The present disclosure also provides a method of manufacturing a molded fibroin body that includes loading fibroin in a mold; maintaining a temperature of the mold between 40°C and 50°C for between five and thirty minutes; raising the temperature of the mold to between 125°C and 180°C; and applying, to the fibroin, a pressure of at least lOMPa for at least five minutes. A pressure of at least lOMPa may be applied to the fibroin to maintain the temperature of the mold between 40°C and 50°C. The fibroin may be loaded in the mold having a water content between 5% and 20%.

[00121] In referring to the description, specific details are set forth in order to provide a thorough understanding of the examples disclosed. In other instances, well- known methods, procedures, components and circuits have not been described in detail as not to unnecessarily lengthen the present disclosure.

[00122] It should be understood that if an element or part is referred herein as being "on", "against", "connected to", or "coupled to" another element or part, then it can be directly on, against, connected or coupled to the other element or part, or intervening elements or parts may be present. In contrast, if an element is referred to as being "directly on", "directly connected to", or "directly coupled to" another element or part, then there are no intervening elements or parts present. When used, term "and/or", includes any and all combinations of one or more of the associated listed items, if so provided. [00123] Spatially relative terms, such as "under" "beneath", "below", "lower", "above", "upper", "proximal", "distal", and the like, may be used herein for ease of description to describe one element or feature's relationship to another element(s) or feature(s) as illustrated in the various figures. It should be understood, however, that the spatially relative terms are intended to encompass different orientations of the device in use or operation in addition to the orientation depicted in the figures. For example, if the device in the figures is turned over, elements described as "below" or "beneath" other elements or features would then be oriented "above" the other elements or features. Thus, a relative spatial term such as "below" can encompass both an orientation of above and below. The device may be otherwise oriented (rotated 90 degrees or at other orientations) and the spatially relative descriptors used herein are to be interpreted accordingly. Similarly, the relative spatial terms "proximal" and "distal" may also be interchangeable, where applicable.

[00124] The term "about," as used herein means, for example, within 10%, within 5%, or less. In some embodiments, the term "about" may mean within measurement error.

[00125] The terms first, second, third, etc. may be used herein to describe various elements, components, regions, parts and/or sections. It should be understood that these elements, components, regions, parts and/or sections should not be limited by these terms. These terms have been used only to distinguish one element, component, region, part, or section from another region, part, or section. Thus, a first element, component, region, pail, or section discussed below could be termed a second element, component, region, part, or section without departing from the teachings herein.

[00126] The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting. The use of the terms "a" and "an" and "the" and similar referents in the context of describing the disclosure (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. The terms comprising, having, includes, including, containing and the like are to be construed as open-ended terms (meaning, e.g., including, but not limited to) unless otherwise noted. Such terms, when used in the present disclosure, specify the presence of stated features, integers, steps, operations, elements, and/or components, but do not preclude the presence or addition of one or more other features, integers, steps, operations, elements, components, and/or groups thereof not explicitly stated. Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the disclosure as if it were individually recited herein. For example, if the range 10-15 is disclosed, then 11, 12, 13, and 14 are also disclosed. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., "such as") provided herein, is intended merely to better illuminate the disclosure and does not pose a limitation on the scope of the disclosure unless otherwise claimed. No language herein should be construed as indicating any non-claimed element as essential to the practice of the disclosure.

[00127] Each patent, patent publication and other publication identified herein and examples described herein are expressly incorporated herein by reference. These publications are provided solely for their disclosure prior to the filing date or applicable priority date of the present disclosure. Nothing in this regard shall be construed as an admission against antedating such publication(s). All statements as to the date or representation as to content/disclosure of such publication(s) is based on information available and does not constitute any admission as to accuracy of date of disclosure/publication of such publication(s) or content of such publication(s).

[00128] It will be appreciated that the methods and compositions of the instant disclosure can be incorporated in the form of a variety of embodiments, only a few of which are disclosed herein. Variations of those embodiments may become apparent to those of ordinary skill in the ail upon reading the foregoing description. Skilled artisans may employ such variations as appropriate, and the present disclosure may be practiced in manners other than as specifically described herein. Accordingly, this disclosure includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the disclosure unless otherwise indicated herein or otherwise clearly contradicted by context.

Claims

WHAT IS CLAIMED IS:

1. A molded body containing fibroin, the molded body comprising: a first region located at a surface of the molded body; and a second region located at least 0.1 mm beneath the surface of the molded body, wherein a degree of crystallization of the molded body at the first region is at least 60%, and wherein the degree of crystallization of the molded body at the second region is between 35% and 50%.

2. The molded body according to claim 1, wherein the degree of crystallization of the molded body decreases from the surface to a center of the molded body.

3. The molded body according to claim 1, wherein a thickness of the second region is between 0.25mm and 5.0mm.

4. The molded body according to claim 1, wherein a thickness of the first region is 0.05mm or less.

5. The molded body according to claim 1, wherein the molded body has a predefined region where the degree of crystallization is 35% or less.

6. The molded body according to claim 5, wherein the predefined region is located 0.5mm beneath the surface of the molded body.

7. The molded body according to claim 1, wherein a thickness of the molded body is between 0.5mm and 10.0mm.

8. A method of manufacturing a molded fibroin body, the method comprising: loading fibroin in a mold; and maintaining a temperature of the mold between 40°C and 50°C.

9. The method of claim 8, wherein pressure is applied to the fibroin to maintain the temperature of the mold above 40°C.

10. The method of claim 9, wherein the applied pressure is IMPa or less.

11. The method of claim 8, further comprising: raising the temperature of the mold to between 125°C and 180°C.

12. The method of claim 11, further comprising: applying, to the fibroin, a pressure of at least lOMPa for at least 5 minutes.

13. The method of claim 8, wherein the fibroin loaded in the mold has a water content between 5% and 20%.

14. A method of manufacturing a molded fibroin body, the method comprising: loading fibroin in a mold; maintaining a temperature of the mold between 40°C and 50°C for between five and thirty minutes; raising the temperature of the mold to between 125°C and 180°C; and applying, to the fibroin, a pressure of at least lOMPa for at least five minutes.

15. The method of claim 14, wherein a pressure of at least lOMPa is applied to the fibroin to maintain the temperature of the mold between 40°C and 50°C.

16. The method of claim 14, wherein the fibroin loaded in the mold has a water content between 5% and 20%.