Preparation method of a peelable mycelium sheet with in-situ enhanced mechanical properties and its application

Abstract

The present invention belongs to the field of biomaterial technology, and more particularly relates to a method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties, and its application. The cultivation method provided herein precisely controls parameters such as temperature, humidity, and light during the mycelium growth phase, combined with a double-layer degradable gel peeling layer system and the use and process design of specialized reinforcing materials, resulting in a cultivated mycelium sheet with both excellent mechanical properties and peelability. The mycelium sheets obtained using this method can be used in a variety of fields, including footwear and bags, soft furniture packaging, and automotive interior materials, providing green and efficient biomaterial solutions for related industries.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomaterials, and in particular relates to a method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties and an application thereof. Background Art

[0002] As an emerging biomaterial, mushroom mycelium sheets are biodegradable and renewable, and can be grown using agricultural waste such as corn cobs, straw, and sawdust, offering significant advantages in environmental protection and resource utilization. In the packaging sector, mushroom mycelium sheets are expected to replace traditional polystyrene cushioning packaging, addressing the problem of white pollution. In the biomedical field, mushroom mycelium sheets, due to their excellent biocompatibility, are suitable for use in tissue engineering scaffolds and wound dressings, shoe and bag fabrics, and automotive interiors.

[0003] However, the current mechanical properties of mushroom mycelium materials are poor, and there are two major problems in practical applications: first, the tear strength is not as good as cowhide when making materials such as shoes, bags, and car interiors, which limits the application scenarios; second, it is difficult to peel it off from the culture medium smoothly. When peeling it using traditional methods, the material is easily damaged, and its integrity and quality are difficult to guarantee, which greatly limits the large-scale production and application of the material.

[0004] Existing methods for improving the mechanical properties of mycelium materials and achieving exfoliation mostly focus on post-molding processing, such as adding reinforcements like carbon fiber and glass fiber. However, these reinforcements can interfere with the mycelium's growth and make it difficult to disperse evenly, resulting in uneven thickness and strength and unstable performance. Post-processing methods like high temperature, high pressure, and chemical cross-linking not only increase production steps and costs but can also leave residual chemicals that affect the biodegradability of the mycelium. Furthermore, mechanical cutting, currently used to achieve exfoliation, can damage the mycelium. Summary of the Invention

[0005] To address the above technical issues, the present invention provides a method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties, and its application. The cultivation method provided by the present invention can significantly improve both the mechanical properties and peelability of the mycelium sheet, while maintaining high sheet integrity and eliminating residual chemical residues, which does not affect the biodegradability of the mycelium sheet.

[0006] In order to achieve the above-mentioned object of the invention, the present invention adopts the following technical solutions:

[0007] A first aspect of the present invention provides a method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties, comprising the following steps:

[0008] S1. Inoculating mushroom spawn onto a solid culture medium and culturing until mycelium uniformly and densely covers the surface of the culture medium, inoculating the mycelium into a liquid culture medium and fermenting until the mycelium wet weight reaches 25% to 30%, thereby obtaining a liquid spawn; placing the mycelium culture medium into an opaque culture box for sterilization, cooling it, inoculating the liquid spawn, and placing the culture box in a light-proof culture chamber for mycelium cultivation; the top cover of the culture box having evenly distributed air holes with a diameter of 0.3 to 2 mm;

[0009] S2. When the mycelium is cultured until the aerial mycelium covers the surface of the culture medium, a chitosan-sodium alginate composite gel layer solution is uniformly coated on the surface of the mycelium culture medium to form a chitosan-sodium alginate composite gel layer with a thickness of 0.3-0.8 mm, and then a lubricating gel layer solution is sprayed on the surface of the chitosan-sodium alginate composite gel layer to form a lubricating gel layer with a thickness of 0.3-1 mm; wherein the chitosan-sodium alginate composite gel layer solution is formed by mixing a chitosan solution and a sodium alginate aqueous solution in a volume ratio of 1:(1-1.2), and contains 0.3%-0.5%wt of calcium chloride, the chitosan concentration of the chitosan solution is 1.5%-2%wt, the solvent is 1%-1.2%wt of acetic acid aqueous solution, and the sodium alginate concentration of the sodium alginate aqueous solution is 1.5%-2%; the lubricating gel layer solution is an aqueous solution containing 2.5%-3% sodium carboxymethyl cellulose and 3%-5%wt of glycerol;

[0010] S3. When the mycelium sheet is initially formed, a nanocellulose-chitin nanocrystal suspension is evenly sprayed onto the surface of the mycelium sheet, and culture is continued at 22-26° C. and a humidity of 60%-80%. During the culture process, ventilation is performed to adjust the oxygen content to 19%-21% v / v and the carbon dioxide concentration to 1%-5%. The mass ratio of nanocellulose to chitin nanocrystals in the nanocellulose-chitin nanocrystal suspension is 1:(1-1.2), and the mass concentrations of the nanocellulose and chitin nanocrystals are both 0.2%-0.3%.

[0011] S4. When the mycelium is cultured for 15 to 17 days, the mycelium sheet formed on the surface of the mycelium culture medium is lifted up.

[0012] Chitosan, sodium alginate, and sodium carboxymethyl cellulose are widely available, low in cost, and have good biocompatibility and degradability. The composite gel layer formed by the two and the lubricating gel layer together constitute a double-layer degradable gel peeling layer system. The chitosan-sodium alginate composite gel layer is the bottom layer of the double-layer degradable gel peeling layer system. It has a porous structure that allows mycelium to penetrate and grow, and can, to a certain extent, guide the mycelium to colonize and grow upward. The lubricating gel layer is the upper layer of the double-layer degradable gel peeling layer system, which can effectively reduce the friction between the mycelium sheet and the double-layer degradable gel peeling layer system, thereby achieving easy and complete peeling of the mycelium sheet from the mycelium culture medium. In addition, the composite gel layer and the lubricating gel layer can also reduce the excessive loss of surface moisture, thereby forming a microenvironment that is relatively suitable for the formation of mycelium material, further making it easier to peel off the mycelium sheet in the later stage.

[0013] During mycelial growth, nanocellulose and chitin nanocrystals play a reinforcing role. Nanocellulose, with its high specific surface area, high strength, and high modulus, can physically entangle and chemically interact with the mycelium, building a three-dimensional network structure. Chitin nanocrystals, as components of the mushroom mycelial cell wall, have good compatibility with the mycelium and promote close connection and orderly arrangement of the mycelium, making the mycelium more tightly connected.

[0014] When aerial mycelium has completely covered the surface of the culture medium, a double-layer degradable gel peeling layer system is added. Once a mycelial sheet has initially formed, a nanocellulose-chitin nanocrystal suspension is added to ensure that the early mycelium can penetrate the peeling layer. Continued cultivation after adding the nanocellulose-chitin nanocrystal suspension promotes solidification and structural densification of the mycelial sheet. After 15-17 days of culture, the mycelial sheet formed on the surface of the mycelial culture medium can be quickly and completely peeled away from the double-layer degradable gel peeling layer system, leveraging the self-forming interface of the double-layer degradable gel peeling layer system.

[0015] The culture box provides a stable culture space for mycelium growth, which is conducive to the uniform growth of mycelium and further improves the quality and performance of mycelium sheets. The air holes on the top cover ensure gas exchange between the culture medium and the outside world, providing sufficient oxygen for mycelium growth.

[0016] At the same time, this method precisely controls the temperature, humidity and carbon dioxide concentration of the culture environment to create a stable environment for mycelium growth, thereby affecting the growth metabolism and gene expression of the mycelium, prompting it to synthesize more high-strength biomacromolecules, optimizing the micro- and macro-structures of the mycelium, and further achieving in situ enhancement of the mechanical properties of mycelium sheets.

[0017] In step S2 of this preparation method, the thicknesses of the chitosan-sodium alginate composite gel layer and the lubricating gel layer refer to the thicknesses in the wet state. The time required for mycelium cultivation in step S2 to reach the point where aerial mycelia cover the surface of the culture medium is 3-5 days, and the time required for the mycelium sheet to be initially formed in step S3 is 7-9 days of mycelium cultivation.

[0018] Preferably, the mushroom species in S1 is Ganoderma lucidum or Ganoderma lucidum. Ganoderma lucidum and Ganoderma lucidum have fast growth rates, strong adaptability, and excellent mycelium mechanical properties.

[0019] Preferably, in S1, the mushroom spawn is inoculated onto a solid culture medium under a strict sterile environment.

[0020] Preferably, the solid culture medium in S1 is a slant culture medium.

[0021] Further preferably, the slant culture medium is prepared by cutting 200 g of potato into pieces, adding 1 L of water and boiling for 15 min, filtering through 4 layers of gauze, adding 20 g of glucose and 20 g of agar to the obtained filtrate, and fixing the volume to 1 L.

[0022] Preferably, the preparation method of the mycelium culture medium described in S1 is: corn cobs, straw, and sawdust are mixed in a mass ratio of 2:(1-1.2):(1-1.2), crushed to a particle size of less than 5 mm, 1%-2%wt of sucrose and 1%-1.5%wt of gypsum powder are added, mixed evenly, and water is added to adjust the moisture content to 60%-65%.

[0023] Preferably, 50-100 ml of the liquid fungus is inoculated into every 8.5 liters of the mycelium culture medium in S1.

[0024] Preferably, the mushroom strains in S1 are cultured on the solid culture medium at a temperature of 24-26° C. and a humidity of 60%-80%.

[0025] Optionally, the culture box is a rectangular culture box with a length of 40 to 60 cm, a width of 30 to 40 cm, and a height of 8 cm. This specification can not only meet the reasonable utilization of space when cultivating mushroom mycelium on a large scale, but also facilitate operation and transportation.

[0026] Optionally, the culture box is made of polypropylene.

[0027] Preferably, the preparation method of the chitosan-sodium alginate composite gel layer solution in S2 is: dissolving chitosan in an acetic acid solution with a mass fraction of 1% to prepare a chitosan solution with a mass fraction of 2%; preparing sodium alginate into a sodium alginate aqueous solution with a mass fraction of 2%; mixing the chitosan solution and the sodium alginate aqueous solution in a volume ratio of 1:1, adding 0.5%wt calcium chloride, and mixing evenly.

[0028] Preferably, the solvent of the nanocellulose-chitin nanocrystal suspension in S3 is water.

[0029] Preferably, in S3, 500-800 mL of the nanocellulose-chitin nanocrystal suspension is sprayed per square meter of the mycelium culture medium surface.

[0030] Preferably, the preparation method further comprises squeezing the mycelium sheet obtained in S4 to remove 50% to 55% wt of water, and storing the sheet at 4° C. Squeezing out some of the water helps optimize the internal structure and enhance stability and durability.

[0031] Alternatively, a double-roller water squeezer may be used to squeeze out the water.

[0032] The second aspect of the present invention provides a mycelium sheet prepared by the above preparation method.

[0033] A third aspect of the present invention provides an application of the mycelium sheet, comprising:

[0034] Application in the preparation of shoes and bags;

[0035] Application in the preparation of furniture soft packaging;

[0036] Application in the preparation of automotive interior materials.

[0037] The beneficial effects of the present invention are:

[0038] 1. The method provided by this invention achieves a significant improvement in both the mechanical properties and the peelability of mycelium sheets by precisely controlling each stage of mycelium growth, precisely regulating the growth environment, adding specially formulated reinforcing additives, and combining an innovative peeling layer design with an optimized square tray culture process. The tensile strength of the mycelium sheets can be increased by 50%-80%, meeting the stringent mechanical performance requirements for applications such as wound dressings, shoe and bag fabrics, and automotive interior materials. The double-layered degradable gel peeling layer ensures a peeling success rate exceeding 95%, and the sheets remain intact after peeling, significantly improving production efficiency and reducing costs, laying the foundation for large-scale production.

[0039] 2. The method provided by the present invention not only enhances the tensile strength of the mycelium sheet, but also uses biodegradable materials in the entire process without residual chemical synthetic substances, ensuring that the biological properties of the mycelium sheet, such as biodegradability and renewability, are not affected and meet environmental protection requirements.

[0040] 3. The materials used in the method provided by the present invention are of low cost and the process is simple. Combined with the characteristic that the culture boxes can be of uniform specifications, it facilitates large-scale, standardized and efficient production of mycelium sheets.

[0041] 4. The mycelium sheet prepared by the method provided by the present invention has excellent mechanical properties and can be used in the fields of shoes and bags, furniture soft bags, automotive interior materials, etc., which expands the application range of mycelium sheets and has extremely important practical significance for promoting the widespread application of mushroom mycelium materials in multiple fields. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] Figure 1 This is the peeling condition of the mycelium sheet in Comparative Example 1 of the present invention;

[0043] Figure 2 This is the peeling condition of the mycelium sheet in Comparative Example 2 of the present invention;

[0044] Figure 3 This is the peeling condition of the mycelium sheet in Comparative Example 3 of the present invention;

[0045] Figure 4 This is the appearance of the mycelium sheet after peeling off in Comparative Example 5 of the present invention;

[0046] Figure 5 This is the appearance of the mycelium sheet after peeling off in Comparative Example 6 of the present invention;

[0047] Figure 6 This is the peeling condition of the mycelium sheet in Example 1 of the present invention;

[0048] Figure 7 This is the appearance of the mycelium sheet after peeling off in Example 2 of the present invention. DETAILED DESCRIPTION

[0049] In order to make the purpose, technical solutions and advantages of the present invention more clear, the present invention is further described in detail below with reference to specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0050] At present, the methods to improve the mechanical properties of mycelium materials and achieve stripping are mainly to add reinforcing materials such as carbon fiber and glass fiber or adopt post-processing methods such as high temperature, high pressure, and chemical cross-linking. The reinforcing materials will change the biological properties of the mycelium material and cause the performance of the mycelium material to be unstable. The post-processing method will increase the processing cost and may leave residual chemical substances, affecting the degradability of the mycelium material.

[0051] To address the above problems, an embodiment of the present invention provides a method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties, comprising the following steps:

[0052] S1. Inoculating mushroom spawn onto a solid culture medium and culturing until mycelium uniformly and densely covers the surface of the culture medium, inoculating the mycelium into a liquid culture medium and fermenting until the mycelium wet weight reaches 25% to 30%, thereby obtaining a liquid spawn; placing the mycelium culture medium into an opaque culture box for sterilization, cooling it, inoculating the liquid spawn, and placing the culture box in a light-proof culture chamber for mycelium cultivation; the top cover of the culture box having evenly distributed air holes with a diameter of 0.3 to 2 mm;

[0053] S2. When the mycelium is cultured until the aerial mycelium covers the surface of the culture medium, a chitosan-sodium alginate composite gel layer solution is uniformly coated on the surface of the mycelium culture medium to form a chitosan-sodium alginate composite gel layer with a thickness of 0.3-0.8 mm, and then a lubricating gel layer solution is sprayed on the surface of the chitosan-sodium alginate composite gel layer to form a lubricating gel layer with a thickness of 0.3-0.8 mm; wherein the chitosan-sodium alginate composite gel layer solution is formed by mixing a chitosan solution and a sodium alginate aqueous solution in a volume ratio of 1:(1-1.2), and contains 0.3%-0.5%wt of calcium chloride, the chitosan concentration of the chitosan solution is 1.5%-2%wt, the solvent is 1%-1.2%wt of acetic acid aqueous solution, and the sodium alginate concentration of the sodium alginate aqueous solution is 1.5%-2%; the lubricating gel layer solution is an aqueous solution containing 2.5%-3% sodium carboxymethyl cellulose and 3%-5%wt of glycerol;

[0054] S3. When the mycelium sheet is initially formed, a nanocellulose-chitin nanocrystal suspension is evenly sprayed onto the surface of the mycelium sheet, and culture is continued at 22-26° C. and a humidity of 60%-80%. During the culture process, ventilation is performed to adjust the oxygen content to 19%-21% v / v and the carbon dioxide concentration to 1%-5%. The mass ratio of nanocellulose to chitin nanocrystals in the nanocellulose-chitin nanocrystal suspension is 1:(1-1.2), and the mass concentrations of the nanocellulose and chitin nanocrystals are both 0.2%-0.3%.

[0055] S4. When the mycelium is cultured for 15 to 17 days, the mycelium sheet formed on the surface of the mycelium culture medium is lifted up.

[0056] The embodiment of the present invention also provides a mycelium sheet prepared by the above preparation method.

[0057] The solutions of the present invention are described below through specific embodiments.

[0058] The slant culture medium used in the following examples was prepared as follows: 200 g of potato cubes were added to 1 L of water and boiled for 15 minutes. The mixture was filtered through four layers of gauze. To the resulting filtrate (approximately 400 mL) was added 20 g of glucose and 20 g of agar, and the volume was adjusted to 1 L.

[0059] The Ganoderma lucidum, Korean Ganoderma lucidum and American Ganoderma lucidum used in the following examples and comparative examples are all commercial strains purchased from Wuhan Zhou Yulin Edible Fungi Research Institute. Among them, Ganoderma lucidum was identified as tree Ganoderma lucidum ( Ganoderma sessile ), Korean Ganoderma lucidum and American Ganoderma lucidum were identified as Ganoderma lucidum ( Ganoderma lingzhi The big red Ganoderma lucidum and red Ganoderma lucidum used in the following comparative examples are also commercial strains purchased from Weizhiyuan Fungi Industry. The big red Ganoderma lucidum was identified as tree Ganoderma lucidum, and the red Ganoderma lucidum was identified as Ganoderma lucidum.

[0060] The experimental methods used in the following examples are conventional methods unless otherwise specified. The materials, reagents, etc. used in the following examples are all commercially available unless otherwise specified.

[0061] Example 1

[0062] This embodiment provides a method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties, the specific steps of which are as follows:

[0063] 1. Material Preparation

[0064] Select the first-level strain of sweet lily, inoculate it into the test tube slant culture medium under sterile conditions, place it in a constant temperature incubator at 25℃ and 80% humidity for 10 days, until the mycelium grows all over the slant, inoculate the mycelium into PDA liquid culture medium, incubate at a constant temperature of 120rpm and 25℃, and terminate the fermentation when the wet weight of the mycelium reaches 30% to obtain the liquid strain.

[0065] Corn cobs, straw, and sawdust were mixed in a mass ratio of 2:1:1 and ground to a particle size of less than 5 mm. 1% sucrose and 1% gypsum powder were added to the mixture and mixed thoroughly. Water was then added and stirred to a moisture content of 65%. This served as the mycelial culture medium. This mycelial culture medium was placed in an opaque culture box (60 cm long, 40 cm wide, 8 cm high, 2 mm thick, made of heat-resistant polypropylene, with evenly distributed 2 mm diameter air holes in the top cover). Sterilize the box at 121°C and 0.1 MPa for 2 hours and cool before inoculation.

[0066] 2. Liquid strain inoculation

[0067] In a clean bench, inoculate the mycelial culture medium with the liquid spawn, using a standard inoculation volume of 100 ml per box of mycelial culture medium. Place the inoculated culture box in a chamber at 25°C and 80% relative humidity until aerial mycelium covers the entire surface of the culture medium (four days from the date of inoculation).

[0068] 3. Add gel double peeling layer

[0069] Chitosan was dissolved in a 1% acetic acid solution to prepare a 2% chitosan solution. Sodium alginate was prepared as a 2% sodium alginate aqueous solution. The chitosan solution and sodium alginate aqueous solution were mixed in a 1:1 volume ratio, and 0.5% calcium chloride was added as a cross-linker. The mixture was stirred to form a chitosan-sodium alginate composite gel layer solution. Sodium carboxymethyl cellulose was prepared as a 3% aqueous solution, and 5% glycerol was added as a lubricant. The solution was stirred to form a lubricating gel layer solution. The lid of the culture box was opened, and the chitosan-sodium alginate composite gel layer solution was evenly applied to the surface of the aerial mycelium inoculated with liquid bacteria and cultured until it covered the surface of the culture medium. This formed a gel-like chitosan-sodium alginate composite gel layer with a thickness of 0.3 mm when wet. The lubricating gel layer solution was then sprayed onto the chitosan-sodium alginate composite gel layer to form a lubricating gel layer with a thickness of 0.8-1 mm. Replace the top cover of the culture box and continue culturing.

[0070] 4. Add strengthening additives

[0071] Nanocellulose and chitin nanocrystals were mixed in a 1:1 mass ratio and prepared using an ultrasonic dispersion device to create a 0.5% nanocellulose-chitin nanocrystal suspension (water as the dispersion medium). When the mycelial material had initially formed into a mycelial sheet (9 days from the date of inoculation of the liquid spawn into the mycelial culture medium), the lid of the culture box was opened and the nanocellulose-chitin nanocrystal suspension was evenly sprayed onto the surface of the mycelial culture medium using a sprayer, with 80 mL sprayed per box. The lid of the culture box was then replaced and culture continued in a culture room at 22°C and 70% relative humidity. Ventilation was performed twice daily for 30 minutes each time to ensure a fresh air environment with an oxygen content of 19%-21% v / v and a carbon dioxide concentration of 1%-5%.

[0072] 5. Mycelium culture and mycelium sheet forming

[0073] When the mycelium is cultured to a state where mycelium sheets can be harvested (15 days from the date of inoculation of the liquid spawn into the mycelium culture medium), the mycelium sheet formed on the surface of the mycelium culture medium is lifted up, placed on a flat plate, and passed through a double-roller water squeezer to remove 50% wt of the water, and then stored at 4°C.

[0074] Example 2

[0075] This embodiment provides a method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties. The specific steps are basically the same as those in Example 1, except that the fungus species in the material preparation step is the first-level American Ganoderma lucidum fungus species.

[0076] Example 3

[0077] This embodiment provides a method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties. The specific steps are basically the same as those in Example 1, except that the fungus species in the material preparation step is a first-grade Korean Ganoderma lucidum fungus species.

[0078] Example 4

[0079] This embodiment provides a method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties, the specific steps of which are as follows:

[0080] 1. Material Preparation

[0081] Select the first-level strain of sweet lily, inoculate it into the test tube slant culture medium under sterile conditions, place it in a constant temperature incubator at 24℃ and 60% humidity for 10 days, until the mycelium grows all over the slant, inoculate the mycelium into PDA liquid culture medium, incubate at a constant temperature of 120rpm and 25℃, and terminate the fermentation when the wet weight of the mycelium reaches 30% to obtain the liquid strain.

[0082] Corn cobs, straw, and sawdust were mixed in a mass ratio of 2:1:1.2 and ground to a particle size of less than 5 mm. 2% sucrose and 1.5% gypsum powder were added to the mixture and mixed thoroughly. Water was then added and stirred until the moisture content reached 65%. This served as the mycelial culture medium. This mycelial culture medium was placed in an opaque culture box (40 cm long, 30 cm wide, 8 cm high, 2 mm thick, made of heat-resistant polypropylene, with evenly distributed 0.3 mm diameter air holes in the top cover). Sterilize the box at 121°C and 0.1 MPa for 2 hours and cool before inoculation.

[0083] 2. Liquid strain inoculation

[0084] In a clean bench, inoculate the mycelial culture medium with the liquid spawn, using a standard inoculation volume of 100 ml per box of mycelial culture medium. Place the inoculated culture box in a chamber at 25°C and 80% relative humidity until aerial mycelium covers the entire surface of the culture medium (day 3 from the date of inoculation).

[0085] 3. Add gel double peeling layer

[0086] Chitosan was dissolved in a 1% acetic acid solution to prepare a 1.8% chitosan solution. Sodium alginate was prepared as a 1.8% sodium alginate aqueous solution. The chitosan solution and sodium alginate aqueous solution were mixed in a 1:1 volume ratio, and 0.4% calcium chloride was added as a cross-linker. The mixture was stirred to form a chitosan-sodium alginate composite gel layer solution. Sodium carboxymethyl cellulose was prepared as a 2.5% aqueous solution. 4% glycerol was added as a lubricant, and the mixture was stirred to form a lubricating gel layer solution. The lid of the culture box was opened, and the chitosan-sodium alginate composite gel layer solution was evenly applied to the surface of the aerial mycelium inoculated with liquid bacteria and cultured until it covered the surface of the culture medium. This formed a gel-like chitosan-sodium alginate composite gel layer with a thickness of 0.8 mm when wet. The lubricating gel layer solution was then sprayed onto the chitosan-sodium alginate composite gel layer to form a lubricating gel layer with a thickness of 0.3-0.5 mm. Replace the top cover of the culture box and continue culturing.

[0087] 4. Add strengthening additives

[0088] Nanocellulose and chitin nanocrystals were mixed in a 1:1 mass ratio and prepared using an ultrasonic dispersion device to create a 0.6% nanocellulose-chitin nanocrystal suspension (water as the dispersion medium). When the mycelial material had initially formed into a mycelial sheet (eight days from the date of inoculation of the liquid spawn into the mycelial culture medium), the lid of the culture box was opened and the nanocellulose-chitin nanocrystal suspension was evenly sprayed onto the surface of the mycelial culture medium using a sprayer, with 80 mL sprayed per box. The box was then replaced with the lid and cultured in a room maintained at 26°C and 80% relative humidity. Ventilation was maintained twice daily for 30 minutes each time to ensure fresh air, an oxygen content of 19%-21% v / v, and a carbon dioxide concentration of 1%-5%.

[0089] 5. Mycelium culture and mycelium sheet forming

[0090] When the mycelium is cultured to a state where mycelium sheets can be harvested (15 days from the date of inoculation of the liquid spawn into the mycelium culture medium), the mycelium sheet formed on the surface of the mycelium culture medium is lifted up, placed on a flat plate, and passed through a double-roller water squeezer to remove 55% wt of the water, and then stored at 4°C.

[0091] Example 5

[0092] This embodiment provides a method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties, the specific steps of which are as follows:

[0093] 1. Material Preparation

[0094] Select the first-level strain of sweet lily, inoculate it into the test tube slant culture medium under sterile conditions, place it in a constant temperature incubator at 26℃ and 70% humidity for 10 days, until the mycelium grows all over the slant, inoculate the mycelium into PDA liquid culture medium, incubate at a constant temperature of 120rpm and 25℃, and terminate the fermentation when the wet weight of the mycelium reaches 30% to obtain the liquid strain.

[0095] Corn cobs, straw, and sawdust were mixed in a mass ratio of 2:1.2:1 and ground to a particle size of less than 5 mm. 1.5% sucrose and 1.2% gypsum powder were added to the mixture and mixed thoroughly. Water was then added and stirred to a moisture content of 60%. This served as the mycelial culture medium. This mycelial culture medium was placed in an opaque culture box (60 cm long, 40 cm wide, 8 cm high, 2 mm thick, made of heat-resistant polypropylene, with evenly distributed 0.8 mm diameter air holes in the top cover). Sterilize the box at 121°C and 0.1 MPa for 2 hours and cool before inoculation.

[0096] 2. Liquid strain inoculation

[0097] In a clean bench, inoculate the mycelial culture medium with the liquid spawn, using a standard inoculation volume of 100 ml per box of mycelial culture medium. Place the inoculated culture box in a chamber at 25°C and 80% relative humidity until aerial mycelium covers the entire surface of the culture medium (day 5 from the date of inoculation).

[0098] 3. Add gel double peeling layer

[0099] Chitosan was dissolved in a 1.2% acetic acid solution to prepare a 1.5% chitosan solution. Sodium alginate was prepared into a 1.5% sodium alginate aqueous solution. The chitosan solution and sodium alginate aqueous solution were mixed in a volume ratio of 1:1.2, and 0.3% calcium chloride was added as a cross-linking agent. The mixture was stirred to form the chitosan-sodium alginate composite gel layer solution. Sodium carboxymethyl cellulose was prepared into a 2.7% aqueous solution, and 3% glycerol was added as a lubricant. The mixture was stirred to form the lubricating gel layer solution. Open the lid of the culture box and evenly apply the chitosan-sodium alginate composite gel layer solution to the surface of the aerial mycelium inoculated with liquid bacteria and cultured until it covers the surface of the culture medium, forming a gel-like chitosan-sodium alginate composite gel layer with a thickness of 0.3 mm in a moist state. Then, spray the lubricating gel layer solution on top of the chitosan-sodium alginate composite gel layer to form a lubricating gel layer with a thickness of 0.6-0.8 mm. Replace the lid of the culture box and continue culturing.

[0100] 4. Add strengthening additives

[0101] Nanocellulose and chitin nanocrystals were mixed in a mass ratio of 1:1.2 and prepared using an ultrasonic dispersion device to prepare a 0.44% nanocellulose-chitin nanocrystal suspension (water as the dispersion medium). When the mycelial material had initially formed into a mycelial sheet (7 days from the date of inoculation of the liquid spawn into the mycelial culture medium), the lid of the culture box was opened and the nanocellulose-chitin nanocrystal suspension was evenly sprayed onto the surface of the mycelial culture medium using a sprayer, with 80 mL sprayed per box. The lid of the culture box was then replaced and culture continued in a culture room at 24°C and a relative humidity of 60%. Ventilation was performed twice daily for 30 minutes each time to ensure a fresh air environment with an oxygen content of 19%-21% v / v and a carbon dioxide concentration of 1%-5%.

[0102] 5. Mycelium culture and mycelium sheet forming

[0103] When the mycelium is cultured to a state where mycelium sheets can be harvested (15 days from the date of inoculation of the liquid spawn into the mycelium culture medium), the mycelium sheet formed on the surface of the mycelium culture medium is lifted up, placed on a flat plate, and passed through a double-roller water squeezer to remove 50% wt of the water, and then stored at 4°C.

[0104] Comparative Example 1

[0105] This comparative example provides a method for preparing a mycelium sheet, and the specific steps are as follows:

[0106] 1. Material preparation: same as Example 1.

[0107] 2. Liquid spawn inoculation: In a clean bench, inoculate the liquid spawn onto the mycelial culture medium, using a standard inoculation volume of 100 ml per box of mycelial culture medium. Place the inoculated culture box in a culture room at 25°C and 80% relative humidity, ventilating twice daily for 30 minutes each time. On the ninth day, after mycelia form a mycelial sheet on the surface of the mycelial culture medium, transfer the square tray to a fruiting room at 22°C and 70% relative humidity and continue incubation until the 15th day.

[0108] Try to peel the mycelium sheet off the medium.

[0109] Comparative Example 2

[0110] This comparative example provides a method for preparing a mycelium sheet, and the specific steps are as follows:

[0111] 1. Material preparation: same as Example 1.

[0112] 2. Liquid strain inoculation: Same as Example 1.

[0113] 3. Adding a peeling layer: same as Comparative Example 1.

[0114] Ordinary nylon fiber mesh cloth (with a gap size of 50 μm) was laid only on the surface of the mycelial culture medium that had been inoculated with liquid fungus and cultured for 4 days in a culture box as a peeling layer.

[0115] 4. Mycelium culture and mycelium sheet forming

[0116] Place the square tray after laying the peeling layer in a culture room at 25°C and 80% relative humidity, and ventilate twice a day for 30 minutes each time. On the 9th day after the mycelium forms a mycelium sheet on the surface of the mycelium culture medium, transfer the square tray to a fruiting room at 22°C and 70% relative humidity and continue to culture for 15 days.

[0117] Try to peel the mycelium sheet off the medium.

[0118] Comparative Example 3

[0119] This comparative example provides a method for preparing a mycelium sheet, and the specific steps are as follows:

[0120] 1. Material preparation: same as Example 1.

[0121] 2. Liquid strain inoculation: Same as Example 1.

[0122] 3. Adding a double-layer gel peeling layer: Same as Example 1.

[0123] 4. Mycelium cultivation and mycelium sheet forming: same as in Example 1.

[0124] Try to peel the mycelium sheet off the medium.

[0125] Comparative Example 4

[0126] This comparative example provides a method for preparing a mycelium sheet. The specific steps are basically the same as those in comparative example 3, except that in the material preparation step, the mycelium sheet is crushed to a particle size of less than 3 mm.

[0127] Comparative Example 5

[0128] This comparative example provides a method for preparing a mycelium sheet. The specific steps are basically the same as those in comparative example 4, except that the fungus species in the material preparation step is Ganoderma lucidum.

[0129] Comparative Example 6

[0130] This comparative example provides a method for preparing a mycelium sheet. The specific steps are basically the same as those in comparative example 3, except that the fungus species in the material preparation step is Ganoderma lucidum.

[0131] Test Example 1

[0132] The mycelium sheets of Examples 1 to 3 and Comparative Examples 1 to 6 were compared in terms of the ease of peeling and the integrity of the mycelium sheets, and the mycelium sheets were cut into 3×8 cm2 The tensile strength of the mycelium sheet samples was tested using a 1000N tensile tester, and the thickness of the mycelium sheet samples was tested using a thickness meter. The results are shown in Table 1.

[0133] Table 1

[0134]

[0135] Note: The more “+” in “Ease of mycelial material peeling”, the easier it is to separate, and “-” means difficult to separate.

[0136] The mycelium sheet peeling conditions of Comparative Examples 1 to 3 are as follows: Figures 1 to 3 As shown in Table 1 and Figures 1 to 3 It can be seen that among the mycelium sheets of Comparative Examples 1 to 3, Comparative Example 3 has the best separation effect, followed by Comparative Example 2; the mycelium sheet of Comparative Example 2 has slightly higher strength, but lower overall thickness; the double-layer gel peeling layer can provide a new plane for mycelium growth and has no effect on strength.

[0137] The appearance or peeling conditions of the mycelium sheets after peeling of Comparative Example 5, Comparative Example 6 and Examples 1 and 2 are as follows: Figures 4 to 7 As shown in Table 1 and Figures 3 to 5 It can be seen that different strains are suitable for the gel double-layer peeling layer and the thickness of the mycelium sheet does not differ much. The gel double-layer peeling layer has a certain universality and can bring about a good separation effect.

[0138] From Table 1 and Figure 3 、 Figure 6 It can be seen that the addition of reinforcing additives has little effect on the thickness variability of the mycelium sheet, but it can significantly increase the tensile strength of the mycelium sheet by up to 78%. This shows that the gel double-layer peeling layer provides a microenvironment suitable for mycelium growth, which is conducive to the formation of mycelium sheet structure and facilitates matrix separation. On this basis, the addition of reinforcing additives can significantly improve the initial strength of the mycelium sheet.

[0139] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A method for preparing a peelable mycelium sheet with in-situ enhanced mechanical properties, characterized in that: The following steps are involved: S1. Inoculating mushroom spawn onto a solid culture medium and culturing until mycelium uniformly and densely covers the surface of the culture medium, inoculating the mycelium into a liquid culture medium and fermenting until the mycelium wet weight reaches 25% to 30%, thereby obtaining a liquid spawn; placing the mycelium culture medium into an opaque culture box for sterilization, cooling it, inoculating the liquid spawn, and placing the culture box in a light-proof culture chamber for mycelium cultivation; the top cover of the culture box having evenly distributed air holes with a diameter of 0.3 to 2 mm; S2. When the mycelium is cultured until the aerial mycelium covers the surface of the culture medium, a chitosan-sodium alginate composite gel layer solution is uniformly coated on the surface of the mycelium culture medium to form a chitosan-sodium alginate composite gel layer with a thickness of 0.3-0.8 mm, and then a lubricating gel layer solution is sprayed on the surface of the chitosan-sodium alginate composite gel layer to form a lubricating gel layer with a thickness of 0.3-0.8 mm; wherein the chitosan-sodium alginate composite gel layer solution is formed by mixing a chitosan solution and a sodium alginate aqueous solution in a volume ratio of 1:(1-1.2), and contains 0.3%-0.5%wt of calcium chloride, the chitosan concentration of the chitosan solution is 1.5%-2%wt, the solvent is 1%-1.2%wt of acetic acid aqueous solution, and the sodium alginate concentration of the sodium alginate aqueous solution is 1.5%-2%; the lubricating gel layer solution is an aqueous solution containing 2.5%-3% sodium carboxymethyl cellulose and 3%-5%wt of glycerol; S3. When the mycelium sheet is initially formed, a nanocellulose-chitin nanocrystal suspension is evenly sprayed onto the surface of the mycelium sheet, and culture is continued at 22-26° C. and a humidity of 60%-80%. During the culture process, ventilation is performed to adjust the oxygen content to 19%-21% v / v and the carbon dioxide concentration to 1%-5%. The mass ratio of nanocellulose to chitin nanocrystals in the nanocellulose-chitin nanocrystal suspension is 1:(1-1.2), and the mass concentrations of the nanocellulose and chitin nanocrystals are both 0.2%-0.3%. S4. When the mycelium is cultured for 15 to 17 days, the mycelium sheet formed on the surface of the mycelium culture medium is lifted up.

2. The method for preparing a mycelium sheet according to claim 1, wherein: The mushroom species in S1 is Ganoderma lucidum or Ganoderma lucidum; and / or The solid culture medium in S1 is a slant culture medium; and / or The mycelium culture medium described in S1 is prepared by mixing corn cobs, straw, and sawdust in a mass ratio of 2:(1-1.2):(1-1.2), crushing the mixture to a particle size of less than 5 mm, adding 1%-2%wt of sucrose and 1%-1.5%wt of gypsum powder, mixing the mixture evenly, and then adding water to adjust the moisture content to 60%-65%; and / or In S1, 50-100 ml of the liquid bacteria seed is inoculated into every 8.5 liters of the mycelium culture medium; and / or The mushroom strains in S1 are cultured on the solid culture medium at a temperature of 24-26° C. and a humidity of 60%-80%; and / or The culture box is a rectangular parallelepiped culture box with a length of 40 to 60 cm, a width of 30 to 40 cm, and a height of 8 cm.

3. The method for preparing a mycelium sheet according to claim 1, wherein: The preparation method of the chitosan-sodium alginate composite gel layer solution described in S2 is as follows: dissolving chitosan in an acetic acid solution with a mass fraction of 1% to prepare a chitosan solution with a mass fraction of 2%; preparing sodium alginate into a sodium alginate aqueous solution with a mass fraction of 2%; mixing the chitosan solution and the sodium alginate aqueous solution in a volume ratio of 1:1, adding 0.5%wt calcium chloride, and mixing uniformly.

4. The method for preparing a mycelium sheet according to claim 1, wherein: The solvent of the nanocellulose-chitin nanocrystal suspension in S3 is water; and / or In S3, 500-800 mL of the nanocellulose-chitin nanocrystal suspension is sprayed on each square meter of the mycelium culture medium surface.

5. The method for preparing a mycelium sheet according to claim 1, wherein: The preparation method further includes squeezing the mycelium sheet obtained in S4 to remove 50% to 55% wt of moisture, and storing the sheet at 4°C.

6. A mycelium sheet prepared by the method for preparing a mycelium sheet according to any one of claims 1 to 5.

7. The use of the mycelium sheet according to claim 6, characterized in that: include: Application in the preparation of shoes and bags; Application in the preparation of furniture soft packaging; Application in the preparation of automotive interior materials.

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