Trametes robiniophila mycelium culture method

Abstract

The invention relates to the field of mycelium culture, and discloses a trametes robiniophila mycelium culture method which is characterized in that a traditional culture medium is improved, and a flaky polylactic acid nano-material, chitosan nano-particles and porous silicon dioxide nano-particles are added into the traditional culture medium to form a three-dimensional porous network structure, so that physical support is provided for mycelia; hyphae can be better attached and grow, and growth and reproduction of the hyphae are effectively promoted. Compared with a traditional culture medium, the problems that the physical environment for mycelial growth is poor, and the utilization efficiency of nutrient substances is low are solved.

Description

Technical Field

[0001] The invention relates to the field of mycelium culture, and in particular to a method for culturing locust ear mycelium. Background Art

[0002] Sophora japonica is a traditional Chinese medicine with important medicinal value. Its main active ingredients include polysaccharides, triterpenoids, etc., which have multiple effects such as hemostasis, antidysentery, and anti-cancer. However, due to the sparse distribution of host tree species of Sophora japonica and the scarcity of wild resources, it is difficult to meet the huge market demand for Sophora japonica. At present, the technology of artificial cultivation of Sophora japonica mainly includes solid fermentation and liquid culture. Although the solid fermentation method can simulate the natural growth environment, the growth cycle is long, the biological effect is low, and the waste of resources is serious. The liquid culture method has the advantages of short culture cycle, high biomass, and easy industrial production, but there are still many deficiencies in the optimization of liquid culture medium in the prior art.

[0003] Traditional liquid culture media are mainly composed of ingredients such as corn flour and soybean cake powder. Although they can support the growth of mycelium, there is still room for improvement in mycelial biomass, extracellular polysaccharide production and quality. In addition, traditional culture media lack optimized design for the mycelial growth environment, resulting in poor physical environment for mycelial growth and low efficiency in nutrient utilization, making mycelium prone to nutritional deficiencies in the later stages of growth.

[0004] Therefore, in response to the above problems, a new solution needs to be proposed to optimize the growth environment of Sophora japonica mycelium. Summary of the Invention

[0005] The invention overcomes the shortcomings of the prior art and provides a method for culturing locust ear mycelium.

[0006] To achieve the above object, the present invention provides a technical solution: a culture medium for Sophora japonica mycelium, comprising the following components:

[0007] Soluble starch 2.00%–3.00%;

[0008] 0.03%–0.10% of sheet-like polylactic acid nanomaterials;

[0009] Peptone 1.00%–1.80%;

[0010] Chitosan nanoparticles 0.01%–0.05%;

[0011] Potassium dihydrogen phosphate 0.15%–0.30%;

[0012] Magnesium sulfate heptahydrate 0.08%–0.15%;

[0013] Zinc sulfate 0.002%–0.010%;

[0014] Sodium L-lactate 0.03%–0.10%;

[0015] Vitamin B1 0.0005%–0.001%;

[0016] Porous silica nanoparticles 0.005%–0.020%;

[0017] The balance is water.

[0018] In a preferred embodiment of the present invention, the specific surface area of the sheet-like polylactic acid nanomaterial is 150-250m 2 / g, flake thickness 10-50nm.

[0019] In a preferred embodiment of the present invention, the soluble starch is corn starch or tapioca starch with an amylopectin content ≥50%.

[0020] In a preferred embodiment of the present invention, the particle size of the chitosan nanoparticles is 50-150 nm.

[0021] In a preferred embodiment of the present invention, the porosity of the porous SiO2 nanoparticles is ≥80%, and the particle size is 100-300 nm.

[0022] In a preferred embodiment of the present invention, the surface of the sheet-like polylactic acid nanomaterial is carboxylated to enhance its hydrogen bond adsorption capacity with starch molecules.

[0023] On the other hand, the present invention also provides a method for culturing Sophora auricularia mycelium, wherein the liquid culture medium is used for fermentation, and the fermentation process includes the following parameters:

[0024] pH value: 6.5-7.0;

[0025] Speed: 180r / min;

[0026] Temperature: 30℃;

[0027] Fermentation time: 8-10 days.

[0028] In a preferred embodiment of the present invention, the pH value is dynamically adjusted by KH2PO4 buffer

[0029] In a preferred embodiment of the present invention, the culture medium is sterilized by filtration through a 0.2-0.25 μm filter membrane before use to ensure a sterile environment.

[0030] In a preferred embodiment of the present invention, the dissolved oxygen level is maintained at 30%-50% by a dissolved oxygen controller during the fermentation process.

[0031] The present invention solves the defects existing in the background technology and has the following beneficial effects:

[0032] (1) The present invention achieves a significant improvement in the culture efficiency and product quality of Sophora japonica mycelium by adding flaky polylactic acid nanomaterials to the culture medium and optimizing the synergistic effect of its surface treatment and other components. The flaky PLA nanomaterials, chitosan nanoparticles, and porous SiO2 nanoparticles synergistically construct a three-dimensional porous network structure, which provides physical support for the mycelium, enables the mycelium to attach and grow better, and effectively promotes the growth and reproduction of the mycelium. Compared with traditional culture media, this design solves the problems of poor physical environment for mycelium growth and low nutrient utilization efficiency. At the same time, through the slow-release-adsorption dual-mode nutrient supply, it ensures that the mycelium evenly disperses nutrients throughout the growth cycle, and maintains sufficient nutrients in the bacterial community, thereby increasing the growth rate and biomass of the mycelium.

[0033] (2) The present invention adopts a multi-component collaborative optimization design, through which the surface carboxyl groups of the sheet-like PLA nanomaterials form hydrogen bond adsorption complexes with starch molecules, gradually releasing glucose to provide a continuous carbon source for the mycelium; at the same time, PLA itself degrades to produce lactic acid monomers, which together with sodium L-lactate activate the mycelial metabolic pathway; this slow-release-adsorption dual-mode nutrient supply method, combined with the "core-shell" structure formed by the electrostatic self-assembly of chitosan and PLA, not only improves the adsorption capacity of nutrients, but also optimizes the mass transfer process, so that nutrients can be more efficiently delivered to the mycelial cells.

[0034] (3) The present invention inhibits the aggregation of mycelium clusters through the three-dimensional network formed by PLA sheets and SiO2, reducing the feedback inhibition caused by local accumulation of metabolic products. This physical barrier drag reduction effect enables the mycelium to be more evenly distributed and grow, thereby improving the metabolic efficiency of the entire culture system. DETAILED DESCRIPTION

[0035] The following is a clear and complete description of the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of the present invention.

[0036] In the following description, many specific details are set forth to facilitate a full understanding of the present invention. However, the present invention may also be implemented in other ways different from those described herein. Therefore, the scope of protection of the present invention is not limited to the specific embodiments disclosed below.

[0037] The sources of the materials are shown in Tables 1 and 2:

[0038] Table 1 can directly obtain the material list

[0039]

[0040]

[0041] Table 2 Special treatment materials

[0042]

[0043] Example 1

[0044] A method for culturing Sophora japonica mycelium, comprising the following steps:

[0045] S1. Prepare the culture medium. The specific components and concentrations are as follows:

[0046] Soluble starch: 25 g / L (amylopectin content ≥ 50% amylopectin corn starch);

[0047] Flake polylactic acid (PLA) nanomaterial: 0.8g / L (specific surface area 200m 2 / g, sheet thickness 30nm, surface carboxylation treatment);

[0048] Peptone: 15 g / L;

[0049] Chitosan nanoparticles: 0.3 g / L (particle size 100 nm);

[0050] Potassium dihydrogen phosphate: 2.0 g / L;

[0051] Magnesium sulfate heptahydrate: 1.2 g / L;

[0052] Zinc sulfate: 0.05g / L;

[0053] Sodium L-lactate: 0.5 g / L;

[0054] Vitamin B1: 8 mg / L;

[0055] Porous silica (SiO2) nanoparticles: 0.1 g / L (porosity 85%, particle size 200 nm);

[0056] Dissolve the above components in deionized water, dilute to 1 L, and stir evenly.

[0057] S2. Sterilize the prepared culture medium by filtering it with a 0.22 μm filter membrane and transfer it into a 5 L fermentation tank with a liquid volume of 3 L.

[0058] S3, inoculated with Sophora japonica mycelium, inoculum size of 10% (v / v), set fermentation parameters: pH 6.8 (dynamically adjusted by KH2PO4 buffer), temperature 30 ° C, rotation speed 180 r / min, dissolved oxygen level maintained at 30%-50%. Fermentation culture for 9 days, during which periodic sampling was performed to detect mycelial biomass, exopolysaccharide production and mycelial mortality.

[0059] Example 2

[0060] A method for culturing Sophora japonica mycelium, comprising the following steps:

[0061] S1. Culture medium preparation: Prepare the culture medium. The specific components and concentrations are as follows:

[0062] Soluble starch: 25g / L (corn starch with amylopectin content ≥50%);

[0063] Flake polylactic acid (PLA) nanomaterial: 0.6g / L (specific surface area 200m 2 / g, sheet thickness 30nm, surface carboxylation treatment);

[0064] Peptone: 15 g / L;

[0065] Chitosan nanoparticles: 0.3 g / L (particle size 100 nm);

[0066] Potassium dihydrogen phosphate: 2.0 g / L;

[0067] Magnesium sulfate heptahydrate: 1.2 g / L;

[0068] Zinc sulfate: 0.05g / L;

[0069] Sodium L-lactate: 0.5 g / L;

[0070] Vitamin B1: 8 mg / L;

[0071] Porous silica (SiO2) nanoparticles: 0.1 g / L (porosity 85%, particle size 200 nm);

[0072] Dissolve the above components in deionized water, dilute to 1 L, and stir evenly.

[0073] S2. Sterilize the prepared culture medium by filtering it with a 0.22 μm filter membrane and transfer it into a 5 L fermentation tank with a liquid volume of 3 L.

[0074] S3, inoculated with Sophora japonica mycelium, inoculum size of 10% (v / v), set fermentation parameters: pH 6.8 (dynamically adjusted by KH2PO4 buffer), temperature 30 ° C, rotation speed 180 r / min, dissolved oxygen level maintained at 30%-50%. Fermentation culture for 9 days, during which periodic sampling was performed to detect mycelial biomass, exopolysaccharide production and mycelial mortality.

[0075] Example 3

[0076] A method for culturing Sophora japonica mycelium, comprising the following steps:

[0077] S1. Prepare the culture medium. The specific components and concentrations are as follows:

[0078] Soluble starch: 25g / L (corn starch with amylopectin content ≥50%);

[0079] Flake polylactic acid (PLA) nanomaterial: 1.0g / L (specific surface area 200m 2 / g, sheet thickness 30nm, surface carboxylation treatment);

[0080] Peptone: 15 g / L;

[0081] Chitosan nanoparticles: 0.3 g / L (particle size 100 nm);

[0082] Potassium dihydrogen phosphate: 2.0 g / L;

[0083] Magnesium sulfate heptahydrate: 1.2 g / L;

[0084] Zinc sulfate: 0.05g / L;

[0085] Sodium L-lactate: 0.5 g / L;

[0086] Vitamin B1: 8 mg / L;

[0087] Porous silica (SiO2) nanoparticles: 0.1 g / L (porosity 85%, particle size 200 nm);

[0088] Dissolve the above components in deionized water, dilute to 1 L, and stir evenly.

[0089] S2. Sterilize the prepared culture medium by filtering it with a 0.22 μm filter membrane and transfer it into a 5 L fermentation tank with a liquid volume of 3 L.

[0090] S3, inoculated with Sophora japonica mycelium, inoculum size of 10% (v / v), set fermentation parameters: pH 6.8 (dynamically adjusted by KH2PO4 buffer), temperature 30 ° C, rotation speed 180 r / min, dissolved oxygen level maintained at 30%-50%. Fermentation culture for 9 days, during which periodic sampling was performed to detect mycelial biomass, exopolysaccharide production and mycelial mortality.

[0091] Example 4

[0092] S1. Prepare the culture medium. The specific components and concentrations are as follows:

[0093] Soluble starch: 20 g / L (corn starch with amylopectin content ≥ 50%);

[0094] Flake polylactic acid (PLA) nanomaterial: 0.8g / L (specific surface area 200m 2 / g, sheet thickness 30nm, surface carboxylation treatment);

[0095] Peptone: 15 g / L;

[0096] Chitosan nanoparticles: 0.3 g / L (particle size 100 nm);

[0097] Potassium dihydrogen phosphate: 2.0 g / L;

[0098] Magnesium sulfate heptahydrate: 1.2 g / L;

[0099] Zinc sulfate: 0.05g / L;

[0100] Sodium L-lactate: 0.5 g / L;

[0101] Vitamin B1: 8 mg / L;

[0102] Porous silica (SiO2) nanoparticles: 0.1 g / L (porosity 85%, particle size 200 nm);

[0103] Dissolve the above components in deionized water, dilute to 1 L, and stir evenly.

[0104] S2. Sterilize the prepared culture medium by filtering it with a 0.22 μm filter membrane and transfer it into a 5 L fermentation tank with a liquid volume of 3 L.

[0105] S3, inoculated with Sophora japonica mycelium, inoculum size of 10% (v / v), set fermentation parameters: pH 6.8 (dynamically adjusted by KH2PO4 buffer), temperature 30 ° C, rotation speed 180 r / min, dissolved oxygen level maintained at 30%-50%. Fermentation culture for 9 days, during which periodic sampling was performed to detect mycelial biomass, exopolysaccharide production and mycelial mortality.

[0106] Example 5

[0107] S1. Prepare the culture medium. The specific components and concentrations are as follows:

[0108] Soluble starch: 30 g / L (corn starch with amylopectin content ≥ 50%);

[0109] Flake polylactic acid (PLA) nanomaterial: 0.8g / L (specific surface area 200m 2 / g, sheet thickness 30nm, surface carboxylation treatment);

[0110] Peptone: 15 g / L;

[0111] Chitosan nanoparticles: 0.3 g / L (particle size 100 nm);

[0112] Potassium dihydrogen phosphate: 2.0 g / L;

[0113] Magnesium sulfate heptahydrate: 1.2 g / L;

[0114] Zinc sulfate: 0.05g / L;

[0115] Sodium L-lactate: 0.5 g / L;

[0116] Vitamin B1: 8 mg / L;

[0117] Porous silica (SiO2) nanoparticles: 0.1 g / L (porosity 85%, particle size 200 nm);

[0118] Dissolve the above components in deionized water, dilute to 1 L, and stir evenly.

[0119] S2. Sterilize the prepared culture medium by filtering it with a 0.22 μm filter membrane and transfer it into a 5 L fermentation tank with a liquid volume of 3 L.

[0120] S3, inoculated with Sophora japonica mycelium, inoculum size of 10% (v / v), set fermentation parameters: pH 6.8 (dynamically adjusted by KH2PO4 buffer), temperature 30 ° C, rotation speed 180 r / min, dissolved oxygen level maintained at 30%-50%. Fermentation culture for 9 days, during which periodic sampling was performed to detect mycelial biomass, exopolysaccharide production and mycelial mortality.

[0121] Comparative Example 1

[0122] A method for culturing Sophora japonica mycelium, comprising the following steps:

[0123] S1. Medium preparation: Prepare liquid culture medium according to the traditional culture medium formula. The specific components and concentrations are as follows:

[0124] Soluble starch: 30 g / L (corn starch with amylopectin content ≥ 50%);

[0125] Maltose: 5g / L;

[0126] Peptone: 15 g / L;

[0127] Soybean cake powder hydrolyzate: 10g / L;

[0128] KH2PO4: 2.0g / L;

[0129] MgSO4·7H2O: 1.0g / L;

[0130] ZnSO4: 0.05g / L;

[0131] Na2SeO3: 0.02g / L;

[0132] Wheat bran extract: 5% (v / v);

[0133] VB1: 6mg / L.

[0134] Dissolve the above components in deionized water, dilute to 1 L, and stir evenly.

[0135] S2. Sterilize the prepared culture medium by filtering it with a 0.22 μm filter membrane and transfer it into a 5 L fermentation tank with a liquid volume of 3 L.

[0136] S3, inoculated with Sophora japonica mycelium, inoculum size of 10% (v / v), set fermentation parameters: pH 6.8 (dynamically adjusted by KH2PO4 buffer), temperature 30 ° C, rotation speed 180 r / min, dissolved oxygen level maintained at 30%-50%. Fermentation culture for 9 days, during which periodic sampling was performed to detect mycelial biomass, exopolysaccharide production and mycelial mortality.

[0137] Comparative Example 2

[0138] S1. Medium preparation: Prepare liquid culture medium according to the traditional culture medium formula. The specific components and concentrations are as follows:

[0139] Soluble starch: 30 g / L (corn starch with amylopectin content ≥ 50%);

[0140] Maltose: 5g / L;

[0141] Peptone: 15 g / L;

[0142] Soybean cake powder hydrolyzate: 10g / L;

[0143] KH2PO4: 2.0g / L;

[0144] MgSO4·7H2O: 1.0g / L;

[0145] ZnSO4: 0.05g / L;

[0146] Na2SeO3: 0.02g / L;

[0147] Wheat bran extract: 5% (v / v);

[0148] VB1: 6mg / L;

[0149] Chitosan nanoparticles: 0.3 g / L;

[0150] Porous SiO2 nanoparticles: 0.1g / L.

[0151] Dissolve the above components in deionized water, dilute to 1 L, and stir evenly.

[0152] S2. Sterilize the prepared culture medium by filtering it with a 0.22 μm filter membrane and transfer it into a 5 L fermentation tank with a liquid volume of 3 L.

[0153] S3, inoculated with Sophora japonica mycelium, inoculum size of 10% (v / v), set fermentation parameters: pH 6.8 (dynamically adjusted by KH2PO4 buffer), temperature 30 ° C, rotation speed 180 r / min, dissolved oxygen level maintained at 30%-50%. Fermentation culture for 9 days, during which periodic sampling was performed to detect mycelial biomass, exopolysaccharide production and mycelial mortality.

[0154] Comparative Example 3

[0155] A method for culturing Sophora japonica mycelium, comprising the following steps:

[0156] S1. Culture medium preparation: Prepare the culture medium. The specific components and concentrations are as follows:

[0157] Soluble starch: 30 g / L (corn starch with amylopectin content ≥ 50%);

[0158] Maltose: 5g / L;

[0159] Peptone: 15 g / L;

[0160] Soybean cake powder hydrolyzate: 10g / L;

[0161] KH2PO4: 2.0g / L;

[0162] MgSO4·7H2O: 1.0g / L;

[0163] ZnSO4: 0.05g / L;

[0164] Na2SeO3: 0.02g / L;

[0165] Wheat bran extract: 5% (v / v);

[0166] VB1: 6mg / L;

[0167] Flake polylactic acid (PLA) nanomaterial: 0.8g / L (specific surface area 200m 2 / g, sheet thickness 30nm, surface not carboxylated).

[0168] Dissolve the above components in deionized water, dilute to 1 L, and stir evenly.

[0169] S2. Sterilize the prepared culture medium by filtering it with a 0.22 μm filter membrane and transfer it into a 5 L fermentation tank with a liquid volume of 3 L.

[0170] S3, inoculated with Sophora japonica mycelium, inoculum size of 10% (v / v), set fermentation parameters: pH 6.8 (dynamically adjusted by KH2PO4 buffer), temperature 30 ° C, rotation speed 180 r / min, dissolved oxygen level maintained at 30%-50%. Fermentation culture for 9 days, during which periodic sampling was performed to detect mycelial biomass, exopolysaccharide production and mycelial mortality.

[0171] Comparative Example 4

[0172] A method for culturing Sophora japonica mycelium, comprising the following steps:

[0173] S1. Prepare the culture medium. The specific components and concentrations are as follows:

[0174] Soluble starch: 25g / L (corn starch with amylose content ≥55%);

[0175] Flake polylactic acid (PLA) nanomaterial: 0.8g / L (specific surface area 200m 2 / g, sheet thickness 30nm, surface carboxylation treatment);

[0176] Peptone: 15 g / L;

[0177] Chitosan nanoparticles: 0.3 g / L (particle size 100 nm);

[0178] Potassium dihydrogen phosphate: 2.0 g / L;

[0179] Magnesium sulfate heptahydrate: 1.2 g / L;

[0180] Zinc sulfate: 0.05g / L;

[0181] Sodium L-lactate: 0.5 g / L;

[0182] Vitamin B1: 8 mg / L;

[0183] Porous silica (SiO2) nanoparticles: 0.1 g / L (porosity 85%, particle size 200 nm).

[0184] Dissolve the above components in deionized water, dilute to 1 L, and stir evenly.

[0185] S2. Sterilize the prepared culture medium by filtering it with a 0.22 μm filter membrane and transfer it into a 5 L fermentation tank with a liquid volume of 3 L.

[0186] S3, inoculated with Sophora japonica mycelium, inoculum size of 10% (v / v), set fermentation parameters: pH 6.8 (dynamically adjusted by KH2PO4 buffer), temperature 30 ° C, rotation speed 180 r / min, dissolved oxygen level maintained at 30%-50%. Fermentation culture for 9 days, during which periodic sampling was performed to detect mycelial biomass, exopolysaccharide production and mycelial mortality.

[0187] Performance measurement

[0188] The cultures in Examples 1-5 and Comparative Examples 1-4 were subjected to the following performance tests:

[0189] Mycelial biomass: measured by centrifugal drying method, and the average value was taken;

[0190] Extracellular polysaccharide production and composition: polysaccharide content was determined by phenol-sulfuric acid method, and β-glucan ratio was analyzed by HPLC;

[0191] Mycelial mortality was determined by trypan blue staining and the average value was taken.

[0192] The test data is shown in Table 3:

[0193] Table 3 Test data table

[0194]

[0195]

[0196] Summary and Analysis

[0197] From the comparative data of Examples 1-5 and Comparative Examples 1-4, it can be seen that the present invention significantly improves the culture efficiency and product quality of Sophora japonica mycelium by adding sheet-like polylactic acid (PLA) nanomaterials to the culture medium and optimizing its surface treatment and the synergistic effect of other components.

[0198] Comparing Examples 1-5 with Comparative Examples 1-4: In Example 1, the mycelial biomass reached 30.1 g / L, which was significantly higher than 24.5 g / L of Comparative Example 1. At the same time, the data of other Examples were significantly better than those of the Comparative Examples. The reasons for this are:

[0199] First, sheet-like polylactic acid (PLA) nanomaterials are more easily combined with starch with a high branched content. The large number of α-1,6 glycosidic bonds in the branched starch molecules form a branched structure, constructing a highly branched three-dimensional network. This structure gives it high solubility and low retrogradation, so it is easy to form a colloid in liquid culture medium and is not easy to age and precipitate. It can be evenly dispersed and form a stable composite sustained-release carrier with PLA nanosheets. In contrast, amylose is mainly composed of linear α-1,4 glycosidic bonds, which makes its molecules tightly curled into a spiral structure and easily form a semi-solid gel. In liquid fermentation, this gelation will hinder oxygen dissolution and material transfer, and the rapid retrogradation characteristics of amylose will lead to a decrease in carbon source utilization and increase process complexity. Therefore, corn starch with a higher branched starch content shows significant advantages in the cultivation of Sophora japonica mycelium and is superior to high-amylose corn starch.

[0200] Second, the addition of flaky PLA nanomaterials in Example 1 synergistically interacted with chitosan nanoparticles and porous SiO2 nanoparticles in the culture medium to form a three-dimensional porous network structure. This structure provided physical support for the mycelium, enabling it to better adhere and grow, effectively promoting its growth and reproduction.

[0201] Third, PLA nanomaterials form hydrogen-bonded adsorption complexes with starch molecules through surface carboxyl groups, gradually releasing glucose under the action of amylase, providing a continuous carbon source for the mycelium. Under the adsorption of PLA nanomaterials and the stirring of the fermentation tank, the glucose is dispersed more evenly, avoiding local mycelium necrosis caused by uneven glucose distribution. At the same time, PLA itself degrades to produce lactic acid monomers, which together with sodium L-lactate activate the mycelial metabolic pathway, forming a slow-release and adsorption dual-mode nutrient supply method. This method ensures that the mycelium can obtain sufficient nutrition throughout the growth cycle, thereby increasing the mycelial growth rate and biomass.

[0202] Fourthly, chitosan (positively charged) and PLA (negatively charged) combine with each other through electrostatics to form a "core-shell" structure, which increases the loading capacity of carbon and nitrogen sources. This surface charge regulation not only improves the adsorption capacity of nutrients, but also optimizes the mass transfer process, enabling nutrients to be more efficiently delivered to the mycelial cells. In Example 1, the extracellular polysaccharide yield reached 2.2 g / L, and β-glucan accounted for 72%, both of which were better than 1.5 g / L and 62% in Comparative Example 1. This indicates that surface charge regulation and mass transfer optimization significantly improved the synthesis efficiency of polysaccharides.

[0203] Fifth, in Examples 1-5, the three-dimensional network formed by the PLA sheets and SiO2 inhibited hyphae aggregation and reduced feedback inhibition caused by localized accumulation of metabolites. This physical barrier drag-reducing effect enabled more uniform distribution and growth of hyphae, improving the metabolic efficiency of the entire culture system. In Example 1, the hyphae mortality rate was less than 8% throughout the entire process, while in Comparative Example 1 it was as high as 21%, indicating that the physical barrier drag-reducing effect effectively maintained the vitality of the hyphae.

[0204] Comparing Examples 1-3, the concentrations of the flaky PLA nanomaterials were 0.8 g / L, 0.6 g / L, and 1.0 g / L, respectively. From the experimental data, it can be seen that when the concentration of the flaky PLA nanomaterials was 0.8 g / L, the mycelial biomass and the extracellular polysaccharide yields both reached the highest values (30.1 g / L and 2.2 g / L). As the concentration decreased or increased, the mycelial biomass and the extracellular polysaccharide yields both decreased. This indicates that there is an optimal range for the concentration of the flaky PLA nanomaterials, within which it is The synergistic effect of other components in the culture medium is the strongest, and they can maximize their effect in promoting mycelial growth and polysaccharide synthesis; the reason for the existence of the optimal range is that when the number of PLA sheets is insufficient, they cannot form a continuous three-dimensional porous network with chitosan and SiO2 nanoparticles, resulting in room for improvement in the uniformity of the dispersion of mycelial attachment points and the efficiency of nutrient transfer in local areas. When the number of PLA sheets is too high, the excessive stacking of PLA sheets causes the network pore size to shrink, hindering mycelial penetration and oxygen diffusion, which is also not conducive to mycelial growth.

[0205] Comparing Example 1, Example 4 and Example 5, the concentrations of soluble starch were 25 g / L, 20 g / L and 30 g / L, respectively. It can be seen from the experimental data that the concentration of soluble starch has a significant effect on the growth efficiency and product synthesis of Sophora auricularia mycelium. Similarly, there is a clear threshold for the synergistic effect with the flaky PLA nanomaterials for carbon source supply and physical structure balance. Amylopectin (≥50%) and carboxylated PLA form a porous slow-release carrier through hydrogen bonding and hydrophobic interaction. When the amount of starch is appropriate, the carrier porosity is large, and the glucose produced in the carrier can be released in sufficient amount to promote the growth of Sophora auricularia mycelium. When the starch is insufficient, the carrier structure becomes loose, manifested as a large porosity and an excessively fast glucose release rate. When the starch is excessive, excessive cross-linking will block the carrier pores, reducing the glucose slow-release efficiency and affecting mycelial growth.

[0206] It can be seen from the data of Example 1 and Comparative Example 1 that the traditional culture medium mainly contains components such as soluble starch, maltose, peptone, soybean cake powder hydrolyzate, KH2PO4, MgSO4·7H2O, ZnSO4, Na2SeO3, wheat bran extract and VB1, while the culture medium of the present application adds flaky PLA nanomaterials, chitosan nanoparticles and porous SiO2 nanoparticles on this basis, and optimizes the concentration and ratio of each component. Through the three-dimensional network structure and slow-release-adsorption dual-mode nutrient supply, it effectively promotes the growth of mycelium, significantly improves the synthesis efficiency of polysaccharides through metabolic signal cascade amplification and physical barrier drag reduction effect, and effectively reduces the mortality rate of mycelium and maintains the vitality of mycelium by optimizing the physical structure and nutrient supply.

[0207] Example 1 was compared with Comparative Example 2. Chitosan nanoparticles and porous SiO2 nanoparticles were added to Example 1, while no flaky PLA nanomaterial was added to Comparative Example 2, and only chitosan and porous SiO2 nanoparticles were added. Experimental data showed that the mycelial biomass and extracellular polysaccharide yield of Example 1 were both higher than those of Comparative Example 2, indicating that the flaky PLA nanomaterial, when acting synergistically with chitosan and porous SiO2 nanoparticles, can more effectively promote the growth of mycelium and the synthesis of polysaccharides.

[0208] Comparing Example 1 with Comparative Example 3, the flaky PLA nanomaterial in Example 1 was subjected to carboxylation surface treatment, while the flaky PLA nanomaterial in Comparative Example 3 was not carboxylation treated. Experimental data showed that the mycelial biomass and extracellular polysaccharide yield of Example 1 were both higher than those of Comparative Example 3, indicating that the carboxylation surface treatment enhanced the hydrogen bond adsorption capacity of the flaky PLA nanomaterial with starch molecules, further optimized the slow-release-adsorption dual-mode nutrient supply, and increased the mycelial growth rate and polysaccharide synthesis efficiency.

[0209] Example 1 was compared with Comparative Example 4. Unlike Example 1, Comparative Example 4 used high-amylose corn starch with an amylose content of ≥55%. The experimental data show that the test data of Comparative Example 4, which used a higher amylose content, was significantly inferior to that of Example 1. This is because the sheet-like PLA nanomaterial is more easily combined with starch with a higher branched content because the branched structure formed by the large number of α-1,6 glycosidic bonds in the amylopectin molecules forms a highly branched three-dimensional network. This structure gives it high solubility and low retrogradation, making it easy to form a colloid while not easily aging and precipitating. It can be uniformly dispersed in liquid culture medium and form a stable composite sustained-release carrier with the PLA nanosheets. In contrast, the linear α-1,4 glycosidic bond structure in amylose causes the molecules to tightly curl into a helix, easily forming a semi-solid gel. During liquid fermentation, gelation hinders dissolved oxygen and mass transfer. In addition, the rapid retrogradation property of amylose leads to a decrease in carbon source utilization, reducing the growth efficiency of Sophora japonica mycelium.

[0210] In summary, the present application significantly improves the culture efficiency and product quality of Sophora japonica mycelium by adding flaky PLA nanomaterials to the culture medium and optimizing its surface treatment and the synergistic effect of other components. Compared with traditional culture media, the culture medium of the present invention can construct a three-dimensional porous network structure, realize slow-release-adsorption dual-mode nutrient supply, optimize the mass transfer process, and significantly improve the growth rate of mycelium and the synthesis efficiency of polysaccharides through metabolic signal cascade amplification and physical barrier drag reduction effect. This innovative technology not only provides a new technical solution for the efficient cultivation of Sophora japonica mycelium, but also provides an important theoretical basis and practical guidance for the innovation and development of fungal fermentation technology.

[0211] The above description is based on the ideal embodiment of the present invention. Based on the above description, relevant personnel can make various changes and modifications without departing from the technical scope of this invention. The technical scope of this invention is not limited to the content of the specification and must be determined according to the scope of the claims.

Claims

1. A locust ear mycelium culture medium, characterized in that In terms of mass percentage, it includes the following ingredients: Soluble starch 2.00%–3.00%; 0.03%–0.10% of sheet-like polylactic acid nanomaterials; Peptone 1.00%–1.80%; Chitosan nanoparticles 0.01%–0.05%; Potassium dihydrogen phosphate 0.15%–0.30%; Magnesium sulfate heptahydrate 0.08%–0.15%; Zinc sulfate 0.002%–0.010%; Sodium L-lactate 0.03%–0.10%; Vitamin B1 0.0005%–0.001%; Porous silica nanoparticles 0.005%–0.020%; The balance is water.

2. A Sophora japonica mycelium culture medium according to claim 1, characterized in that: The specific surface area of the sheet-like polylactic acid nanomaterial is 150-250 m 2 / g, flake thickness 10-50nm.

3. The method for culturing Sophora japonica mycelium according to claim 1, wherein: The soluble starch is corn starch or cassava starch with an amylopectin content of ≥50%.

4. The method for culturing Sophora japonica mycelium according to claim 1, wherein: The particle size of the chitosan nanoparticles is 50-150 nm.

5. The method for culturing Sophora japonica mycelium according to claim 1, wherein: The porosity of the porous SiO2 nanoparticles is ≥80%, and the particle size is 100-300nm.

6. The method for culturing Sophora japonica mycelium according to claim 1, wherein: The surface of the sheet-like polylactic acid nanomaterial is carboxylated to enhance hydrogen bond adsorption capability with starch molecules.

7. A method for culturing Sophora japonica mycelium, characterized in that: Fermentation culture is carried out using the liquid culture medium according to any one of claims 1 to 6, wherein the fermentation process includes the following parameters: pH value: 6.5-7.0; Speed: 180r / min; Temperature: 30℃; Fermentation time: 8-10 days.

8. The method for culturing Sophora japonica mycelium according to claim 6, wherein: The pH value was dynamically adjusted by KH2PO4 buffer.

9. The method for culturing Sophora japonica mycelium according to claim 1, wherein: The culture medium was sterilized by filtration through a 0.2-0.25 μm filter membrane before use to ensure a sterile environment.

10. The method for culturing Sophora japonica mycelium according to claim 1, wherein: During the fermentation process, the dissolved oxygen level was maintained at 30%-50% by a dissolved oxygen controller.

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