CN113527895A

CN113527895A - A kind of degradable superfibroin-based material and its preparation method and application

Info

Publication number: CN113527895A
Application number: CN202110891655.2A
Authority: CN
Inventors: 郭成辰; 吕浩
Original assignee: Westlake University
Current assignee: Westlake University
Priority date: 2021-08-04
Filing date: 2021-08-04
Publication date: 2021-10-22
Anticipated expiration: 2041-08-04
Also published as: CN113527895B

Abstract

本发明涉及一种可降解超韧丝蛋白基材料及其制备方法及应用。本发明的可降解超韧丝蛋白基材料包含增塑剂、盐及丝蛋白，所述塑化剂为选自甘油、山梨醇、聚乙二醇中的一种或多种，以及所述盐为选自氯化钙、氯化锂、溴化锂中的一种或多种。通过调节超韧丝蛋白基材料中增塑剂与盐的比例，本发明的超韧丝蛋白基材料具有可调的力学性能、降解性能及水溶性，取决于具体应用，可被开发成各种生物工程材料的原料。The present invention relates to a degradable superfibroin-based material and a preparation method and application thereof. The degradable superfibroin-based material of the present invention comprises a plasticizer, a salt and silk protein, the plasticizer is one or more selected from glycerol, sorbitol, polyethylene glycol, and the salt is one or more selected from calcium chloride, lithium chloride and lithium bromide. By adjusting the ratio of plasticizer to salt in the super fibroin-based material, the super fibroin-based material of the present invention has adjustable mechanical properties, degradation properties and water solubility, and can be developed into various Raw materials for bioengineering materials.

Description

Degradable super-tough silk protein-based material and preparation method and application thereof

Technical Field

The invention relates to a silk fibroin-based material, and particularly relates to a degradable super-tough silk fibroin material, and a preparation method and application thereof.

Background

Silk protein is a protein-based biopolymer extracted from natural silkworm cocoons, and has excellent mechanical properties, biocompatibility and biodegradability. In recent years, silk protein-based materials have been widely used in biomedical fields such as tissue engineering, bioelectronics, and regenerative medicine. For example, a silk protein film can be used as a substrate for preparing a biosensor; the silk protein sponge can be used as a tissue engineering scaffold; the silk fibroin nanospheres can be used as carriers for drug delivery and release; the fibroin blocks can be processed into implantable bone pegs and the like. Generally, silk protein can be extracted from natural silkworm cocoons by a dissolving and regenerating method, and the preparation method is green and environment-friendly and has good practical application value.

The silk protein-based material prepared based on silk protein is widely applied to the biomedical fields of epidermal wound repair, cornea regeneration, tympanic membrane repair, tissue engineering scaffolds, flexible bioelectronic devices and the like, and has good application prospect. Generally, silk protein-based materials are prepared by solution processing methods. In order to control the physicochemical properties, in particular the mechanical properties and the biodegradability, of the fibroin-based materials, several post-treatment methods (such as water annealing, alcohol treatment, etc.) are employed. In addition, in order to increase the mechanical properties (e.g. stretchability and toughness) of the silk protein based material, some small molecules (e.g. glycerol, etc.) may be added as plasticizers into the silk protein solution (S.Lu et al. biomacromolecules 2010,11, 143-. Taking glycerol as an example, the addition of the glycerol increases the fluidity of the silk fibroin molecular chain, and improves the flexibility of the silk fibroin film. Meanwhile, the addition of glycerol induces fibroin molecules to form a beta-folded structure, and further forms a crystalline region, so that the degradability of the fibroin-based material is reduced, and the research and application of the fibroin-based material in the field of degradable flexible devices are limited.

In addition to the plasticizer, some metal cations (e.g., potassium, calcium, etc.) can interact with the fibroin molecules to form ion pairs or coordination, thereby affecting the molecular structure of the fibroin and the chain aggregation state during material fabrication and molding. For example, relevant studies have shown that calcium ions can interact with negatively charged amino acid residues in silk proteins (e.g., glutamate and aspartate), block ubiquitous hydrophobic interactions and modulate the self-assembly process of silk proteins (p. dubey et al. biomacromolecules 2015,16, 3936-3944).

The preparation of the degradable super-tough silk protein-based material has important significance for promoting the application of silk protein materials in more fields, particularly the biomedical field, and is also a research focus. Although some progress has been made by previous related studies, until now, there is a lack of a very efficient and easy-to-implement process for preparation. Therefore, the current urgent need exists for exploring the regulation and control method of structure and aggregation state of silk protein in the processing process and developing a preparation method of novel degradable super-tough silk protein-based material.

Disclosure of Invention

The invention aims to provide a degradable super-tough silk egg-based material and a preparation method thereof, which can realize the accurate regulation and control of the water solubility, the mechanical property and the degradability of the silk protein-based material by changing the mixing proportion of raw materials.

In one aspect, the present invention provides a super-tough silk protein-based material, which comprises a plasticizer, a salt and silk protein, wherein the plasticizer is one or more selected from glycerol, sorbitol and polyethylene glycol, and the salt is one or more selected from calcium chloride, lithium chloride and lithium bromide.

In some embodiments, the weight ratio of silk fibroin to plasticizer to salt in the super-tough silk fibroin-based material can be 1 (0.15-0.3) to (0.05-0.15), such as 1 (0.25-0.28) to (0.05-0.07), or 1 (0.27-0.29) to (0.13-0.15).

The properties and performances of the prepared material can be regulated and controlled by adjusting the mass ratio of the fibroin, the plasticizer and the salt. For example, when the salt ratio is 10% or more, the prepared material has excellent water solubility and degradability. When the plasticizer ratio is 20% or more, the prepared material has excellent toughness and stretchability. According to the invention, the toughness, water solubility, mechanical property and degradability of the prepared silk protein-based material can be adjusted by adjusting the proportion of silk protein, plasticizer and salt in the mixed solution, so that a simple, convenient and reliable method is provided for obtaining the silk protein-based material with expected performance.

In the super-tough silk fibroin-based material according to the present invention, the plasticizer, the salt, and the silk fibroin are uniformly mixed, preferably at a molecular level.

According to the requirement, the super-tough silk protein-based material can also comprise functional molecules or materials, such as drugs, antibodies, growth factors, inorganic nano materials, dyes, organic molecules and the like. Generally, these ingredients may be present in an amount of 0-50 wt%, based on the total weight of the super-tough silk fibroin-based material, for example 20% carbon nanotubes may be added.

In some embodiments, the super-tough silk fibroin-based material consists of a plasticizer, a salt, and silk fibroin.

In some embodiments, the plasticizer is glycerin and the salt is calcium chloride. In some embodiments, the mass fraction of glycerin is 15-20%, such as 15.5%, 16%, 16.5%, 17%, 17.5%, 18%, 18.5%, 19%, 19.5%, etc., and the mass fraction of calcium chloride is 5-10%, such as 5.5%, 6%, 6.5%, 7%, 7.5%, 8%, 8.5%, 9%, 9.5%, etc., based on 100% by weight of the dry weight of the super-tough silk fibroin-based material. In some embodiments, the mass fraction of glycerin is 20% and the mass fraction of calcium chloride is 10% based on 100% by weight of the dry weight of the super-tough silk fibroin-based material. In other embodiments, the mass fraction of glycerin is 20% and the mass fraction of calcium chloride is 5% based on 100% by weight of the dry weight of the super-tough silk fibroin-based material.

In some embodiments, the super-tough silk protein-based material may be in the form of a film, a sheet, a sponge, a powder, a fiber, a gel, and the like, but is not limited thereto.

The method for preparing the super-tough silk fibroin-based material according to the present invention is not particularly limited as long as it can achieve uniform mixing of the respective components, preferably molecular-level uniform mixing, and for example, it can be prepared by solution mixing, grinding in the presence of a solvent, melt mixing, kneading, and the like.

In another aspect, the present invention provides a method for preparing the above-mentioned super-tough silk protein-based material, comprising the steps of:

(1) preparing a silk protein aqueous solution;

(2) preparing a plasticizer aqueous solution;

(3) preparing an aqueous salt solution;

(4) mixing the silk protein water solution, the plasticizer water solution and the salt water solution to obtain a uniformly mixed solution;

(5) and drying the obtained solution to obtain the super-tough silk protein-based material.

In some embodiments, the super-tough silk fibroin-based material is in the form of a thin film. For example, the film may be formed by a solution casting method, but is not limited thereto. For example, the mixed solution obtained in the step (4) is transferred to a prefabricated mould and dried to obtain the modified silk protein film, wherein the drying temperature is 0-90 ℃ and the drying time is 3-48 hours. Drying may be carried out under atmospheric pressure or under vacuum.

In some embodiments, the super-tough silk protein-based material is in the form of a sponge. For example, the sponge is made by freeze-drying, but not limited thereto. Preferably, the temperature of freeze-drying is-50 to-110 ℃.

In some embodiments, the super-tough silk fibroin-based material is in a powder form. For example, the sponge is made by means of spray drying, but is not limited thereto.

In some embodiments, the super-tough silk protein-based material is in the form of a gel. For example, the gel may be prepared by concentrating the solution and then physically or chemically crosslinking the solution. For example, the concentration temperature is 5 to 60 ℃ and the concentration time is 1 to 60 hours, the physical crosslinking includes adjusting the pH to less than 5, applying electricity, adding an organic solvent such as alcohol, and the like, and the chemical crosslinking includes, but is not limited to, enzymatic crosslinking.

In some embodiments, the super-tough silk fibroin-based material is in the form of a sheet or block. For example, the solution is made into a sheet or block form by drying and curing, preferably, the drying temperature is 5 to 150 ℃.

In some embodiments, the super-tough silk fibroin-based material is in the form of a fiber. For example, the fiber may be prepared by concentrating the solution and then spinning the resulting concentrated solution by a spinning method such as dry spinning, wet spinning, electrostatic spinning, etc., but is not limited thereto.

In some embodiments, step (1) comprises: adding the crushed silkworm cocoons into a boiling sodium carbonate solution, keeping the solution degummed, rinsing the degummed silk and drying. And then adding the dried degummed silk into a lithium bromide aqueous solution for dissolving. And (4) after the silk fiber is completely dissolved, dialyzing and purifying the solution to obtain silk protein water solution, and storing the silk protein water solution in a refrigerator. The concentration of the silk fibroin can be measured by taking a certain amount of silk fibroin aqueous solution and drying.

In some embodiments, in step (1), the concentration of the sodium carbonate solution is 0.01 to 0.05mol/L and the degumming treatment time is 5 to 180 minutes.

In some embodiments, in step (1), the aqueous lithium bromide solution has a concentration of 9.3mol/L and is dissolved at 60 ℃ for 4 hours.

In some embodiments, in step (2), the concentration of the plasticizer solution is 10-1000mg/mL, such as 100 mg/mL.

In some embodiments, in step (3), the salt solution has a concentration of 0.01 to 5mol/L, for example 1 mol/L.

In some embodiments, the plasticizer may comprise from 10 to 30%, for example from 15 to 20%, by mass of all materials in the system except water in the final prepared material.

In some embodiments, the salt may comprise from 2 to 20% by mass of all materials in the system other than water, for example from 5 to 10% by mass of the finally prepared material.

In another aspect, the present invention provides an application of the above super-tough silk protein-based material as a bioengineering material, wherein the bioengineering material can be selected from epidermal wound repair material, cornea regeneration material, tympanic membrane repair material, tissue regeneration scaffold material, packaging material, flexible bioelectronic device, and the like.

Advantageous effects

The preparation method of the super-tough silk protein-based material has the following advantages:

(1) the silk protein solution required in the preparation method is directly prepared from natural silkworm cocoons, the raw materials are wide in source, cheap and easy to obtain, and the yield is controllable; the added reagents, plasticizers and salts are common industrial raw materials and are low in price. The regulation and control method is simple and easy to operate, does not influence the biocompatibility of the silk protein-based material, and provides an effective way for expanding the application of the silk protein-based material in the biomedical field.

(2) The mechanical property and degradability of the silk protein film obtained by the regulation and control method can be accurately regulated and controlled through the addition ratio of the plasticizer and the salt. According to molecular analysis, the plasticizer is added to increase the chain motion of silk protein molecules, regulate the glass transition temperature of the silk protein molecules and regulate the crystallization degree of the silk protein molecules, so that the stretchability and the toughness of the silk protein material are macroscopically enhanced. Ions, especially metal ions, in the salt can form ion pairs with negatively charged groups in the silk protein molecules, so that the chain aggregation state of the silk protein molecules is regulated, and the crystallization behavior of the silk protein molecules is influenced, so that the water solubility and the degradability of the material are regulated macroscopically. By combining plasticizer regulation and salt regulation, the degradable super-tough silk protein-based material can be prepared, and the precise regulation and control of the mechanical property and the degradation property of the material can be realized.

(3) The invention realizes a method for accurately regulating and controlling the mechanical property and the degradation property of the silk fibroin-based material at the same time, prepares the super-tough silk fibroin-based material with excellent degradation property, and greatly promotes the application potential of the silk fibroin-based material in the biomedical field, such as the preparation of degradable tissue engineering scaffolds, implantation interventional devices, flexible electronic device substrates and the like with excellent performance.

Drawings

FIG. 1 is a photograph of a degradable super-tough silk protein film prepared in example 1 of the present invention.

FIG. 2 is an infrared absorption spectrum of the degradable super-tough silk protein thin film prepared in examples 1 to 4 of the present invention.

FIG. 3 is a Differential Scanning Calorimetry (DSC) characterization chart of the degradable super-tough fibroin films prepared in examples 1-4 of the present invention.

FIG. 4 is a graph showing the mechanical properties of the degradable super-tough fibroin films prepared in examples 1 to 4 of the present invention, wherein A is a graph showing the elongation at break and the breaking strength of each film, and B is a graph showing the Young's modulus of each film.

FIG. 5 is a water-solubility characterization diagram of the degradable super-tough silk protein thin film prepared in examples 1-4 of the present invention.

FIG. 6 is a degradation characterization diagram of the degradable super-tough silk protein thin film prepared in example 3 of the present invention under different enzyme actions.

FIG. 7 is a photograph of a concentrated silk protein/glycerol/calcium chloride solution prepared in example 5 at a high concentration.

Fig. 8 is a photograph of a fiber obtained by dry spinning using the high concentration fibroin/glycerin/calcium chloride concentrate prepared in example 5.

Fig. 9 is a graph showing the mechanical property test results of the fiber prepared in example 6.

FIG. 10 is a graph showing infrared absorption spectra of silk protein films prepared in comparative examples 1 to 5 of the present invention.

FIG. 11 is a differential scanning calorimetry characterization of the fibroin films prepared in comparative examples 1-5 of the present invention.

FIG. 12 is a graph showing the mechanical properties of the silk protein films prepared in comparative examples 1 to 5 of the present invention, wherein A is a graph showing the elongation at break and the strength at break of each film, and B is a graph showing the Young's modulus of each film.

FIG. 13 is a water solubility characterization of silk protein films prepared in comparative examples 1-5 of the present invention.

Detailed Description

The regulation and control method of the present invention is further illustrated in detail by the following examples. The examples are for illustration only and do not limit the invention in any way.

Examples 1 to 4

The degradable super-tough silk protein film is prepared according to the following method:

(1) as shown in Table 1, a fibroin solution, a glycerol solution and a calcium chloride solution were prepared, and the solutions were mixed uniformly by the volume shown in Table 1. 3.5mL of the mixed solution was poured into a petri dish with a diameter of 6mm so that the solution completely covered the surface of the petri dish.

(2) And (3) drying the culture dish poured with the mixed solution at room temperature for 12 hours in an open manner, then placing the dried culture dish in a vacuum drying oven for continuously drying at room temperature for 2-4 hours to prepare each silk protein film (15% of glycerol and 5% of calcium chloride, 15% of glycerol and 10% of calcium chloride, 20% of glycerol and 5% of calcium chloride, and 20% of glycerol and 10% of calcium chloride).

Photographs of the resulting silk protein films (20% glycerol + 10% calcium chloride) were prepared as shown in fig. 1.

TABLE 1

In table 1 above, the mass fraction is represented as the total mass of all materials except water in the system.

Comparative examples 1 to 5

In order to contrast the performance of the degradable super-tough silk protein film prepared according to the present invention, silk protein films modified without using a plasticizer and a salt and silk protein films modified with a single plasticizer or a single salt were prepared as follows:

(1) as shown in Table 2, a fibroin solution, a glycerol solution and a calcium chloride solution were prepared, and the solutions were mixed uniformly by the volume shown in Table 2. 3.5mL of each of the mixed solutions was poured into a petri dish having a diameter of 6mm so that the solution completely covered the surface of the petri dish.

(2) And (3) drying the culture dish poured with the mixed solution at room temperature for 12 hours in an open manner, then placing the dried culture dish in a vacuum drying oven for continuously drying at room temperature for 2-4 hours to prepare each silk protein film (pure silk film; 15% glycerol; 20% glycerol; 5% calcium chloride; 10% calcium chloride).

TABLE 2

In table 2 above, the mass fraction is represented as the total mass of all materials except water in the system.

Test example 1:

the modified silk protein films prepared in examples 1 to 4 and comparative examples 1 to 5 were subjected to infrared absorption spectrum characterization using an infrared spectrometer equipped with an attenuated total reflection accessory. The film samples were placed in the grooves of a diamond plate to collect the signals, and each sample was scanned 64 times with a resolution of 4cm^-1Wave number range of 400cm^-1To 4000cm^-1. The infrared absorption spectra are shown in fig. 2 and 10.

Test example 2:

the thermal properties of examples 1-4 and comparative examples 1-5 were characterized by differential scanning calorimetry. The samples were cut into small pieces and loaded into a 40. mu.L aluminum crucible, each sample weighing 3-7 mg. The differential scanning calorimetry measurement procedure consisted of two scans with a heating rate of 20 deg.C/min. The first scan was heated from 20 to 200 ℃ to remove free water from the film and the second scan was heated from-30 to 270 ℃ to measure the glass transition temperature of the different films. Differential scanning calorimetry characterization results are shown in figures 3 and 11.

Test example 3:

the mechanical properties of examples 1 to 4 and comparative examples 1 to 5 were tested by means of a universal mechanical testing machine equipped with a 50N load cell. The samples were cut into dumbbell strips according to ASTM standards and the film was then loaded onto the jaws of the machine. For each test, the stretching rate was 20mm/min for all samples, and the stretching was stopped until the samples broke, and at least 4 replicates were tested per set of films. The cross-sectional area of each sample was calculated by multiplying the film thickness by its gauge width. The tensile stress and strain are calculated based on the original cross-sectional area and length, respectively. Young's modulus, elongation at break and strength at break are determined by tensile stress and strain, plotted as cross-sectional area and length variation. The mechanical property characterization results are shown in fig. 4 and 12.

Test example 4:

the water solubility of examples 1 to 4 and comparative examples 1 to 5 was tested by uv spectrophotometry. Samples from different groups were cut into 10mm by 10mm squares and soaked in 20mL of deionized water. And after 24 hours, respectively taking the supernatant of all samples, measuring the ultraviolet absorbance of the solution at 280nm, determining the silk protein content of the samples dissolved in water according to different absorbances, and finally calculating the water solubility of different samples. The results of the water solubility test are shown in fig. 5 and 13.

Test example 5:

the degradation behaviour of the films obtained in example 3 was characterized by the degradation behaviour in different enzyme solutions (protease xiv, collagenase and a-chymotrypsin). The samples were cut into 10mm by 10mm squares and soaked in 2mL of each of the different enzyme solutions. The remaining sample mass was weighed on day 1 and day 3, respectively, and the degradation rate of the sample in different enzyme solutions was calculated. The results of the enzymatic degradation test are shown in FIG. 6.

And (4) analyzing results: from the data in fig. 2 and 3, it can be seen that as the proportion of glycerol added increases, the β -sheet content in the degradable super-tough fibroin film also increases, and the glass transition temperature of the fibroin molecules decreases.

From the data in fig. 4, it can be seen that as the ratio of glycerol and calcium chloride is increased, the elongation at break (stretchability) of the degradable super-tough silk protein film is increased.

As can be seen from the data in fig. 5, the water solubility of the degradable super-tough silk protein film increases when the calcium chloride ratio reaches or exceeds 10%.

As can be seen from the data in FIG. 6, the prepared degradable super-tough silk protein film has good enzymatic degradability.

In addition, it can be seen from a of fig. 4 and a of fig. 12 that silk protein films modified without using a plasticizer and a salt have inferior stretchability compared to the degradable super-tough silk protein films prepared in examples 1 to 4.

As can be seen from fig. 4 and 12 and fig. 5 and 13, silk protein films modified using only a plasticizer have poor water solubility and toughness, compared to the degradable super-tough silk protein films prepared in examples 1 to 4.

As can be seen from fig. 4 and 12, the silk protein film modified using only salt has inferior stretchability compared to the degradable super-tough silk protein films prepared in examples 1 to 4.

The data prove that the degradable super-tough silk protein film with adjustable mechanical property can be prepared through the synergistic regulation effect of the glycerol and the calcium chloride, and particularly when the proportion of the added glycerol is 20% and the proportion of the added calcium chloride is 10%, the novel super-tough silk protein film with excellent water solubility can be prepared, and the film has good application prospect in the fields of implantable medical devices and the like in the future.

Example 5

The low-concentration silk protein/glycerin/calcium chloride mixed aqueous solution prepared in example 3 was concentrated to prepare a high-concentration mixed silk protein aqueous solution. The concentrated silk fibroin aqueous solution can be stably stored at low temperature after being centrifugally purified, and the concentration is 40%. FIG. 7 is a graph of a high concentration silk protein/glycerol/calcium chloride concentrate.

Example 6

The high-concentration fibroin/glycerin/calcium chloride concentrate obtained in example 5 was added to a 5mL syringe for dry spinning, and the syringe port was connected to a stainless steel needle having a diameter of 0.6 mm. The solution flow rate was controlled by a syringe pump at a rate of 40. mu.L/h. The diameter of the collecting device was 12mm and the rotational speed was 140 rpm. The distance between the needle tip and the collection means was 150 mm. The temperature during the experiment was 27 ℃ and the humidity was 40%. The diameter of the finally prepared fiber is 7.0 +/-1.4 mu m. Fig. 8 is a diagram of a fiber obtained by dry spinning using a concentrated solution of high-concentration fibroin/glycerin/calcium chloride, and a mechanical property test curve of the obtained fiber is shown in fig. 9, and the result shows that the prepared fiber has strong toughness.

Claims

1. Super-tough silk protein-based material, comprising a plasticizer, a salt and silk protein, wherein the plasticizer is one or more selected from glycerol, sorbitol and polyethylene glycol, and the salt is one or more selected from calcium chloride, lithium chloride and lithium bromide.

2. The super-tough silk fibroin-based material of claim 1, wherein the weight ratio of silk fibroin to plasticizer to salt in the super-tough silk fibroin-based material is 1 (0.15-0.3): 0.05-0.15), such as 1 (0.25-0.28): 0.05-0.07, or 1 (0.27-0.29): 0.13-0.15.

3. The super-tough silk protein-based material according to claim 1, wherein the super-tough silk protein-based material further comprises functional molecules or materials, such as drugs, antibodies, growth factors, inorganic nanomaterials, dyes, organic molecules.

4. The super phloretin-based material of claim 1, wherein the plasticizer is glycerol and the salt is calcium chloride.

5. The super-tough silk fibroin-based material according to claim 4, wherein the mass fraction of glycerol is 15-20% and the mass fraction of calcium chloride is 5-10% based on 100 wt% of the dry weight of the super-tough silk fibroin-based material, preferably the mass fraction of glycerol is 20% and the mass fraction of calcium chloride is 5% based on 100 wt% of the dry weight of the super-tough silk fibroin-based material.

6. The super bast silk protein-based material of claim 1, wherein said super bast silk protein-based material is in the form of a film, a sheet, a block, a sponge, a powder, a fiber, or a gel.

7. A method of preparing the super-tough silk fibroin-based material of any one of claims 1-6, the method comprising the steps of:

(1) preparing a silk protein aqueous solution;

(2) preparing a plasticizer aqueous solution;

(3) preparing an aqueous salt solution;

8. The method of claim 7, wherein step (1) comprises: adding the crushed silkworm cocoons into a boiling sodium carbonate solution, keeping the silkworm cocoons in the boiling sodium carbonate solution for degumming, rinsing and drying the degummed silk, adding the dried degummed silk into a lithium bromide aqueous solution for dissolving, and dialyzing and purifying the solution after all silk fibers are dissolved to obtain a silk protein aqueous solution.

9. The method of claim 7, wherein,

in step (2), the concentration of the plasticizer solution is 10-1000mg/mL, for example 100 mg/mL;

in step (3), the salt solution has a concentration of 0.01 to 5mol/L, for example, 1 mol/L.

10. Use of the super-tough silk protein-based material according to any one of claims 1 to 6 as a bioengineering material, wherein the bioengineering material is selected from the group consisting of epidermal wound repair material, corneal regeneration material, tympanic membrane repair material, tissue regeneration scaffold material, packaging material and flexible bioelectronic devices.