A segmented, regulated two-stage cultivation process for Ganoderma lucidum.

Abstract

This invention discloses a segmented, two-stage cultivation process for Ganoderma lucidum. The process first involves activating the fungal strain on a solid culture medium with specific components to ensure its robustness. Then, the activated strain is transferred to a liquid culture medium rich in nutrients such as corn flour and wheat bran. By precisely controlling temperature, humidity, and light cycle, and adding naphthaleneacetic acid (NAA) inducers in stages, fruiting bodies are induced. Finally, the fruiting bodies are harvested using vacuum low-temperature drying. This invention overcomes the bottlenecks of long solid-state cultivation cycles and poor quality in liquid-state cultivation. Through segmented and precise control combining physical environment and chemical inducers, the cultivation cycle is significantly shortened to 35-45 days. It can stably obtain Ganoderma lucidum fruiting bodies with regular morphology, high content of key active ingredients (polysaccharide content ≥15%, triterpenoid content ≥3%), and high yield, providing a reliable solution for the industrialized and standardized production of Ganoderma lucidum.

Description

Technical Field

[0001] This invention belongs to the field of microbial culture and bioengineering technology, specifically involving a method for the artificial and efficient cultivation of a rare edible and medicinal fungus—Ganoderma lucidum (Tai Sui), and particularly a two-stage cultivation process that promotes its growth and accumulation of active ingredients through segmented regulation. Background Technology

[0002] Reishi mushroom (commonly known as "Tai Sui") is a rare biological resource. While its taxonomic status remains controversial (it may be a large slime mold complex or a specific fungus), its bioactive components, such as polysaccharides, triterpenoids, and proteins, have been proven to have significant immunomodulatory and antioxidant effects, showing broad application prospects in functional foods and pharmaceutical raw materials. With surging market demand, achieving efficient and controllable artificial cultivation of Reishi mushroom has become a core issue for the industry's development.

[0003] However, current artificial cultivation techniques for Ganoderma lucidum are far from mature, facing a technological bottleneck in transitioning from "whether it can be cultivated" to "how to achieve high yield and quality." Existing methods can be broadly categorized into two types, but both have significant limitations: Traditional solid-state culture: This method mimics the natural growth environment and can produce fruiting bodies, but its cultivation cycle often lasts for months or even years, resulting in extremely high time costs. More critically, this process is essentially a "black box" that is difficult to control precisely: it not only requires a large area and is susceptible to contamination by other microorganisms leading to failure, but also suffers from huge fluctuations in the content of active ingredients in the final product and extremely poor batch stability due to the inability to uniformly regulate environmental factors such as light, temperature, and nutrient diffusion, completely failing to meet the requirements of standardized industrial production.

[0004] Preliminary liquid culture methods: To shorten the cycle, some studies have attempted to use liquid fermentation to culture mycelium. However, current technologies mostly remain at the level of a simple "one-pot" constant-temperature culture mode. This extensive process has fundamental flaws: First, it ignores the inherent laws governing the different physiological stages of Ganoderma lucidum growth and development (such as the rapid mycelial proliferation period and the fruiting body induction period), using a single culture medium and culture conditions, which cannot meet the specific needs of each stage, resulting in suboptimal levels of biomass accumulation and secondary metabolite synthesis. Second, the culture medium composition often simply adopts conventional microbial fermentation formulas (such as glucose-peptone systems), failing to specifically simulate and provide the optimal nutrient spectrum required for Ganoderma lucidum to synthesize specific active substances (such as high molecular weight polysaccharides and specific triterpenes). Finally, in the harvesting stage, high-temperature drying is often used, which easily leads to the degradation and inactivation of heat-sensitive active ingredients, resulting in a significant reduction in the efficacy of the final product.

[0005] In summary, existing technologies either have excessively long and uncontrollable cycles, or while shortening the time, they sacrifice product quality and efficacy. The core problem lies in the lack of a process concept based on the biological characteristics of Ganoderma lucidum that allows for precise segmented control of strain activation, vegetative growth, and reproductive growth. Therefore, developing a staged cultivation process that balances high efficiency and high quality, and enables the targeted enrichment of active ingredients, is crucial for overcoming the bottlenecks in the industrialization of Ganoderma lucidum. Summary of the Invention

[0006] To address the above problems, this invention discloses a segmented, controlled two-stage cultivation process for Ganoderma lucidum.

[0007] The objective of this invention is achieved through the following technical solution.

[0008] A segmented, regulated two-stage cultivation process for Ganoderma lucidum includes the following steps: (1) First stage - strain activation culture: The Ganoderma lucidum strain is inoculated onto a primary solid culture medium, which contains the following components by weight: 150-200 parts potato extract, 15-20 parts glucose, 5-10 parts peptone, 15-20 parts agar, and 1000 parts water; it is cultured for 10-15 days at a temperature of 18-22℃, a humidity of 75-80%, and in the dark until the colony diameter reaches 2-3 cm. (2) Second stage - fruiting body induction culture: The inoculum blocks cultured in the first stage are transferred to a secondary liquid culture medium containing the following components by weight: 30-50 parts corn flour, 20-30 parts wheat bran, 5-10 parts yeast extract, 1-2 parts potassium dihydrogen phosphate, 0.5-1 parts magnesium sulfate, and 1000 parts water; cultured for 25-30 days under the conditions of temperature 25-28℃, humidity 85-90%, light intensity 500-1000 lux, and photoperiod of 12 hours light / 12 hours dark, and 0.1-0.3 mg / L naphthaleneacetic acid is added on the 10th and 20th days of culture, respectively; (3) Harvesting and post-processing: When the diameter of the fruiting body of Ganoderma lucidum reaches 5-8 cm, harvest the fruiting body, rinse it with sterile water, and then dry it at 40-50℃ until the moisture content is less than 10%.

[0009] Furthermore, in the above cultivation process, in step (1), the potato extract is prepared by boiling and extracting fresh potatoes, and its soluble solids content is 10%-15%.

[0010] Furthermore, in the above cultivation process, in step (1), the cultivation conditions are: 12 days of cultivation at 20°C, 78% humidity, and in darkness.

[0011] Furthermore, in the above-mentioned cultivation process, in step (2), the weight ratio of corn flour to wheat bran in the secondary liquid culture medium is (35-45):(22-28).

[0012] Furthermore, in the above cultivation process, in step (2), the naphthaleneacetic acid is added by dissolving it in sterile water and applying it evenly to the surface of the culture medium in the form of a spray.

[0013] Furthermore, in the above cultivation process, in step (2), the light conditions are: light intensity of 800 lux, light cycle of 12 hours of light / 12 hours of darkness, and cultivation temperature of 26℃.

[0014] Furthermore, in the above cultivation process, in step (3), the drying is carried out by vacuum drying method, with a vacuum degree of 0.08-0.10 MPa and a drying time of 4-6 hours.

[0015] The present invention also discloses a fleshy Ganoderma lucidum fruiting body obtained by the above-mentioned cultivation process, wherein the polysaccharide content is not less than 15% and the triterpenoid content is not less than 3%.

[0016] Furthermore, the above-mentioned Ganoderma lucidum fruiting bodies, after being stored at room temperature for 12 months, retain more than 90% of the polysaccharides and show no signs of mold or insect infestation.

[0017] The present invention also discloses the application of the above-mentioned Ganoderma lucidum fruiting bodies in the preparation of health products or anti-tumor adjuvant drugs.

[0018] Compared with existing technologies, the present invention has the following advantages and beneficial effects: 1) High efficiency and high yield: The cultivation cycle is significantly shortened from the traditional months or even years to 35-45 days, and the biological yield of fruiting bodies is significantly improved, meeting the efficiency requirements of industrialization.

[0019] 2) Excellent and controllable quality: Through the two-stage design of "solid first and then liquid" and the precise intervention of light and plant growth regulators, the formation of fruiting bodies is effectively induced, ensuring the product has a regular shape and significantly increasing the content of core active substances such as polysaccharides and triterpenes, and the batch-to-batch stability is good.

[0020] 3) Sufficient activity retention: The low-temperature vacuum drying post-processing process minimizes the degradation of heat-sensitive active ingredients, thus better maintaining the product's bioavailability and appearance quality.

[0021] 4) Strong process controllability: The entire process parameters are clear, making it easy to standardize operations and scale up, freeing it from excessive dependence on the natural environment and laying a solid foundation for the production of high-quality, standardized Ganoderma lucidum products. Attached Figure Description

[0022] Figure 1 Comparison of total polysaccharide content (% dry weight); Figure 2 Comparison of total triterpenoid content (% dry weight); Figure 3 Comparison of the effects of ConA-induced proliferation of mouse spleen lymphocytes (proliferation index SI); Figure 4 Comparison of DPPH free radical scavenging rates (%). Detailed Implementation

[0023] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below. However, it should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the scope of the invention. Furthermore, descriptions of well-known structures and technologies are omitted in the following description to avoid unnecessarily obscuring the concept of the invention. All raw materials used in the embodiments of this invention are commercially available.

[0024] To meet the requirement of full disclosure, the raw materials and their key parameters involved in this invention are hereby specified as follows. 1. Ganoderma lucidum strain Raw material: Ganoderma lucidum (obtained according to prior patent CN202210520319.1 (A Ganoderma lucidum, a high-concentration oral liquid containing Ganoderma lucidum and its preparation method), or commercially available).

[0025] Pretreatment: Cut the raw material into small pieces of 0.5 cm × 0.5 cm × 0.5 cm, surface sterilize with 0.1% mercuric chloride solution for 3-5 minutes, then rinse with sterile water 3-5 times and set aside.

[0026] 2. Key Raw Materials for Primary Solid Culture Media Potato extract: prepared by boiling fresh potatoes in water after cutting them into chunks, with the soluble solids content controlled at 10%-15% (measured by refractometer).

[0027] Glucose: food grade or analytical grade.

[0028] Peptone: biological reagent grade, total nitrogen content ≥12%.

[0029] Agar: Bacterial culture grade, gel strength ≥800 g / cm².

[0030] 3. Key raw materials for secondary liquid culture media Corn flour: food grade, passed through an 80-mesh sieve, starch content ≥60%.

[0031] Wheat bran: Fresh, free from mold, with a crude protein content of ≥12%.

[0032] Yeast extract: biological reagent grade, total nitrogen content ≥9.0%.

[0033] Potassium dihydrogen phosphate: analytical grade, content ≥99.0%.

[0034] Magnesium sulfate: analytical grade, content ≥99.0%.

[0035] 4. Inducing agents Naphthaleneacetic acid: Plant hormone grade, purity ≥95.0%. Dissolve in a small amount of anhydrous ethanol before use, then dilute with sterile water to the required concentration.

[0036] 5. Post-processing reagents Sterile water: Meets the standards for water for injection in the Chinese Pharmacopoeia.

[0037] All other chemical reagents not specifically mentioned, such as acids and bases used to adjust pH, are of analytical grade.

[0038] Example 1 A segmented, regulated two-stage cultivation process for Ganoderma lucidum includes the following steps: (1) First stage - strain activation culture: The Ganoderma lucidum strain is inoculated onto a primary solid culture medium containing the following components: 180 parts of potato extract (soluble solids content 12%), 18 parts of glucose, 8 parts of peptone, 18 parts of agar, and 1000 parts of water; cultured at 20℃, 78% humidity and in the dark for 12 days until the colony diameter reaches about 2.5 cm.

[0039] (2) Second stage – Fruiting body induction culture: The robust inoculum blocks (approximately 1cm × 1cm) from the first stage culture were transferred to a 5L fermenter containing 2L of secondary liquid culture medium, which contained the following components: 40 parts corn flour, 25 parts wheat bran, 8 parts yeast extract, 1.5 parts potassium dihydrogen phosphate, 0.8 parts magnesium sulfate, and 1000 parts water. The culture was carried out for 28 days at a temperature of 26℃, humidity of 88%, light intensity of 800 lux, and a photoperiod of 12 hours light / 12 hours darkness. On the 10th and 20th days of culture, naphthaleneacetic acid was added by spraying with a sterile aqueous solution to achieve a final concentration of 0.2 mg / L in the culture medium.

[0040] (3) Harvesting and post-processing: When the diameter of the fruiting body of Ganoderma lucidum reaches 6-7 cm, harvest the fruiting body, rinse it with sterile water, and then place it in a vacuum drying oven and dry it for 5 hours at a vacuum degree of 0.09 MPa and a temperature of 45℃ until the moisture content is 8.5%.

[0041] Example 2 A segmented, regulated two-stage cultivation process for Ganoderma lucidum includes the following steps: (1) First stage - strain activation culture: The Ganoderma lucidum strain is inoculated onto a primary solid culture medium containing the following components: 150 parts of potato extract (soluble solids content 10%), 15 parts of glucose, 5 parts of peptone, 15 parts of agar, and 1000 parts of water; cultured at 18℃, 75% humidity and in the dark for 15 days until the colony diameter reaches about 2 cm.

[0042] (2) Second stage – Fruiting body induction culture: The robust inoculum blocks from the first stage culture were transferred to a 3L fermenter containing 1.5L of secondary liquid culture medium, which contained the following components: 30 parts corn flour, 20 parts wheat bran, 5 parts yeast extract, 1 part potassium dihydrogen phosphate, 0.5 parts magnesium sulfate, and 1000 parts water. The culture was carried out for 30 days at a temperature of 25℃, humidity of 85%, light intensity of 500 lux, and a photoperiod of 12 hours light / 12 hours darkness. On the 10th and 20th days of culture, naphthaleneacetic acid was added by spraying with a sterile aqueous solution to achieve a final concentration of 0.1 mg / L in the culture medium.

[0043] (3) Harvesting and post-processing: When the diameter of the fruiting body of Ganoderma lucidum reaches 5-6 cm, harvest the fruiting body, rinse it with sterile water, and then place it in a vacuum drying oven and dry it for 6 hours at a vacuum degree of 0.08 MPa and a temperature of 40℃ until the moisture content is 9.0%.

[0044] Example 3 A segmented, regulated two-stage cultivation process for Ganoderma lucidum includes the following steps: (1) First stage - strain activation culture: The Ganoderma lucidum strain is inoculated onto a primary solid culture medium containing the following components: 200 parts of potato extract (soluble solids content 15%), 20 parts of glucose, 10 parts of peptone, 20 parts of agar, and 1000 parts of water; cultured for 10 days at a temperature of 22℃, a humidity of 80%, and in the dark until the colony diameter reaches about 3 cm.

[0045] (2) Second stage – Fruiting body induction culture: The robust inoculum blocks from the first stage culture were transferred to a 5L fermenter containing 2.5L of secondary liquid culture medium, which contained the following components: 50 parts corn flour, 30 parts wheat bran, 10 parts yeast extract, 2 parts potassium dihydrogen phosphate, 1 part magnesium sulfate, and 1000 parts water. The culture was carried out for 25 days at a temperature of 28℃, humidity of 90%, light intensity of 1000 lux, and a photoperiod of 12 hours light / 12 hours darkness. On the 10th and 20th days of culture, naphthaleneacetic acid was added by spraying with a sterile aqueous solution to achieve a final concentration of 0.3 mg / L in the culture medium.

[0046] (3) Harvesting and post-processing: When the diameter of the fruiting body of Ganoderma lucidum reaches 7-8 cm, harvest the fruiting body, rinse it with sterile water, and then place it in a vacuum drying oven and dry it for 4 hours at a vacuum degree of 0.10 MPa and a temperature of 50℃ until the moisture content is 8.0%.

[0047] Comparative Example 1 A method for cultivating Ganoderma lucidum, compared to Example 1, differs in that it does not involve a defined two-stage culture; instead, it uses a single liquid culture medium (with the same composition as the secondary liquid culture medium in Example 1) for isothermal shaking culture. Unactivated Ganoderma lucidum inoculum blocks are directly inoculated into the liquid culture medium and cultured at 26°C in the dark with shaking at 120 rpm for 28 days. Naphthaleneacetic acid is not added. At the end of the culture, the mycelium is flocculent and fails to form regular fruiting bodies, resulting in a low biomass (by dry weight).

[0048] Comparative Example 2 A method for cultivating Ganoderma lucidum, compared with Example 1, differs in that naphthaleneacetic acid (NAA) is not added as an inducer during the second stage of fruiting body induction culture, while all other conditions remain the same as in Example 1. After cultivation, the fruiting body formation rate slows down, and the average diameter of the fruiting bodies at harvest is only 4 cm, with significantly lower triterpenoid content than in Example 1.

[0049] Comparative Example 3 A method for cultivating Ganoderma lucidum, compared with Example 1, differs in that the light condition during the second stage of fruiting body induction culture is changed to complete darkness, while all other conditions remain the same as in Example 1. After the culture is completed, the mycelium overgrows and forms clumps, but the fruiting body differentiation is not obvious, the morphology is irregular, and the polysaccharide content is lower than that in Example 1.

[0050] Comparative Example 4 A method for cultivating Ganoderma lucidum, compared with Example 1, differs in that the post-harvest drying process uses traditional hot air drying, drying in a 70°C forced-air drying oven until the moisture content is similar to that of Example 1. The dried fruiting bodies are dark in color, and tests show that the retention rate of heat-sensitive active ingredients (such as certain specific polysaccharide components) is significantly lower than that of the vacuum drying method in Example 1.

[0051] Comparative Example 5 A method for cultivating Ganoderma lucidum (meat-like fungus) is described, referencing the traditional solid-state culture method. The fungal strain is inoculated onto a solid culture medium composed of sawdust, wheat bran, and sucrose, and cultured for 6 months at room temperature (fluctuating between 20-25℃) and natural humidity. The resulting fruiting bodies are uneven in size, have an extremely long cultivation period, and are susceptible to contamination by other microorganisms.

[0052] Test Example 1 Determination of Key Active Ingredient Content Objective: To verify the effect of the segmented two-stage cultivation process of the present invention on increasing the content of core bioactive substances in Ganoderma lucidum fruiting bodies.

[0053] method: Polysaccharide content determination: The phenol-sulfuric acid method was used. 50 mg of each of the dried Ganoderma lucidum fruiting body powder obtained in Examples 1-3 and Comparative Examples 1-5 were accurately weighed, extracted with hot water, and then reacted with phenol and concentrated sulfuric acid. The absorbance was measured at a wavelength of 490 nm, and the total polysaccharide content was calculated using glucose as a standard.

[0054] Determination of triterpenoid content: The vanillin-glacial acetic acid method was used. 20 mg of each sample powder was accurately weighed, extracted with anhydrous ethanol, and then reacted with vanillin-glacial acetic acid solution and perchloric acid. The absorbance was measured at 550 nm, and the total triterpenoid content was calculated using oleanolic acid as a standard.

[0055] Fruiting body yield calculation: At harvest, weigh the total weight of fresh fruiting bodies in each treatment group, and weigh the dry weight after drying. Fruiting body yield (%) = (Dry weight of harvested fruiting bodies / Dry weight of inoculated spawn blocks) × 100%.

[0056] Results: See Table 1 below and Figure 1 , Figure 2 As shown.

[0057] Table 1: Results of determination of key active ingredient content and fruiting body yield

[0058] Conclusion: The key active ingredients (total polysaccharides and total triterpenes) content and fruiting body yield of the products in the embodiments of this invention (Examples 1-3) are significantly higher than those of all comparative examples. In particular, compared with Comparative Example 1 (single liquid culture) which did not form regular fruiting bodies, the polysaccharide and triterpenoid contents of Example 1 increased by approximately 77% and 89%, respectively, and the yield increased by approximately 200%. This indicates that the two-stage culture strategy of this invention (solid activation followed by liquid induction) can efficiently promote the formation of Ganoderma lucidum fruiting bodies and the simultaneous accumulation of target active substances, and the product quality is stable between batches, which is far superior to traditional solid culture (Comparative Example 5).

[0059] Test Example 2 In vitro assessment of immunomodulatory activity (lymphocyte proliferation model) Objective: To evaluate the potential promoting effect of the Ganoderma lucidum product obtained in this invention on the body's specific immune function.

[0060] method: A mouse spleen lymphocyte proliferation model (MTT assay) was used. Lymphocyte suspensions were prepared from aseptically isolated mouse spleens, and after adjusting cell density, they were seeded into 96-well plates. A blank control group, a ConA (concanavalin A, 5 μg / mL) stimulation group, and a ConA+ sample group (containing aseptically treated aqueous extracts of each sample, with a final concentration of 100 μg / mL) were set up. After incubation at 37℃ and 5% CO2 for 48 hours, MTT solution was added to each well for an additional 4 hours of incubation. After termination of incubation, DMSO was added to dissolve formazan crystals, and absorbance (OD value) was measured at 570 nm. The lymphocyte proliferation index (SI) was calculated as: (OD value of ConA+ sample group - OD value of blank control group) / (OD value of ConA stimulation group - OD value of blank control group).

[0061] Results: See Table 2 below and Figure 3 As shown.

[0062] Table 2: Effects on ConA-induced proliferation of mouse spleen lymphocytes (proliferation index SI)

[0063] Conclusion: The product of Example 2 of this invention significantly enhances ConA-induced lymphocyte proliferation, with a proliferation index far exceeding that of other comparative examples. This effect is superior to that of Comparative Example 2 (lacking naphthaleneacetic acid induction) and Comparative Example 3 (lacking light regulation), suggesting that the light and hormone induction in the second stage of the process of this invention are crucial for stimulating the production of substances that enhance immune activity in the fruiting body of Ganoderma lucidum.

[0064] Test Example 3 In vitro antioxidant capacity assay Objective: To compare the overall antioxidant capacity of extracts from different samples.

[0065] method: DPPH free radical scavenging rate: Prepare a 0.1 mM DPPH ethanol solution, mix it with an equal volume of sample water extract (2 mg / mL), react in the dark for 30 minutes, and then measure the absorbance at 517 nm to calculate the scavenging rate.

[0066] Hydroxyl radical (·OH) scavenging rate: The Fenton reaction system was used. o-phenanthroline, PBS buffer, sample extract, FeSO4, and H2O2 were added sequentially. After reacting for 60 minutes, the absorbance was measured at 536 nm, and the scavenging rate of ·OH was calculated.

[0067] Total reducing power: The sample extract was mixed with phosphate buffer and potassium ferricyanide solution, and then incubated in a water bath at 50°C for 20 minutes. Trichloroacetic acid was added, and the supernatant was centrifuged and reacted with ferric chloride solution. The absorbance was measured at 700 nm. The higher the absorbance value, the stronger the reducing power.

[0068] Results: See Table 3 below and Figure 4 As shown.

[0069] Table 3: Results of in vitro antioxidant capacity assay

[0070] Conclusion: The product of Example 2 of this invention exhibits the strongest in vitro antioxidant capacity, with all three indicators significantly higher than those of the comparative example. This indicates that the Ganoderma lucidum fruiting bodies cultivated using the segmented regulation process of this invention contain richer antioxidant substances (such as polysaccharides and polyphenols) and may have a superior structure.

[0071] Test Example 4 Product stability assessment Objective: To evaluate the stability of the product of this invention under accelerated storage conditions, with a focus on the effect of drying methods.

[0072] method: The fruiting body powders from Example 2 (vacuum drying) and Comparative Example 4 (hot air drying) were stored for 3 months under accelerated testing conditions (temperature 40°C ± 2°C, relative humidity 75% ± 5%). Samples were taken at the end of January, February, and March for testing. Retention rate of active ingredients: The content of polysaccharides and triterpenes was determined according to the method in Test Example 1, and the retention percentage relative to the content of 0 was calculated.

[0073] Color stability: The lightness (L), red-green value (a), and yellow-blue value (b*) of the powder were measured using a colorimeter. The total color difference ΔE after 3 months of storage was calculated as √[(ΔL*)]. 2 + (Δa) 2 + (Δb) 2 ].

[0074] Results: As shown in Table 4 below.

[0075] Table 4: Changes in indicators after 3 months of accelerated testing (40°C / 75% RH)

[0076] Conclusion: After 3 months of accelerated testing, the product of Example 2, using the vacuum drying process of this invention, showed significantly higher retention rates of polysaccharides and triterpenes compared to Comparative Example 4 (hot air drying), and exhibited less color change (ΔE). This indicates that the low-temperature vacuum drying process employed in this invention can effectively reduce the degradation of heat-sensitive active ingredients and non-enzymatic browning, better maintaining the product's bioactivity and appearance quality, and thus extending its shelf life.

[0077] Test Example 5 Determination of physical properties of fruiting bodies (rehydration and leachate content) Objective: To evaluate the physical properties of the sub-entities obtained by this invention and to provide a reference for their application in subsequent processing (such as brewing and extraction).

[0078] method: Rehydration test: Accurately weigh 1.0 g (denoted as W_d) each of the dried fruiting body slices from Example 2 and Comparative Example 5 (conventional solid culture), place them in excess distilled water, and soak at 25°C. After soaking for 30, 60, and 120 minutes, remove them, blot off the surface moisture with filter paper, and weigh them (denoted as W). _w ). Calculate the rehydration ratio (RR) = W _w / W _d .

[0079] Determination of water-soluble extract content: Accurately weigh 2.0 g of each sample powder (denoted as W_s), place it in a Soxhlet extractor, and extract with distilled water under reflux until colorless. Collect the extract, evaporate to dryness, and dry at 105℃ to constant weight. Weigh the residue (denoted as W). _e Water-soluble extract content (%) = (W _e / W _s ) × 100%.

[0080] result: Rehydration: The rehydration rate of the sample in Example 2 reached 6.8% after soaking for 120 minutes, while the rehydration rate of the sample in Comparative Example 5 was 4.5%. The sample in Example 2 rehydrated faster and had a fuller morphology after rehydration.

[0081] Water-soluble extract content: The water-soluble extract content of the sample in Example 2 was 42.5%, while that of the sample in Comparative Example 5 was 35.8%.

[0082] Conclusion: The product of Example 2 of this invention exhibits superior rehydration properties and a higher content of water-soluble extracts. This indicates that the fruiting bodies cultured using the liquid-induced method of this invention may have a more porous and loose structure, which is beneficial for water penetration and the dissolution of contents. This characteristic has positive implications for developing ready-to-eat products or improving the extraction efficiency of active ingredients. Summarize: The above test examples demonstrate that the process of this invention exhibits the following advantages: Regarding active ingredients, the polysaccharide content of the products in the examples reached 15.2%-17.5%, the triterpenoid content reached 3.1%-3.6%, and the fruiting body yield reached 260%-298%, all significantly higher than the comparative examples (e.g., polysaccharide content in single liquid culture was only 9.5%). In terms of bioactivity, the product of Example 2 showed a stimulation index as high as 1.65 in the lymphocyte proliferation model, far superior to the comparative examples (1.12-1.41); its DPPH free radical scavenging rate reached 85.3%, and its hydroxyl free radical scavenging rate reached 76.8%, demonstrating strong antioxidant capacity. In terms of stability, after 3 months of accelerated testing, the polysaccharide and triterpenoid retention rates of Example 2 (vacuum dried) both exceeded 93%, and the color difference ΔE was only 2.1, significantly better than the hot-air dried comparative example 4 (retention rate approximately 87%, ΔE=5.3). Furthermore, the product of this invention also exhibits superior rehydration properties (rehydration rate 6.8) and a higher content of water-soluble extractives (42.5%). These data fully demonstrate the outstanding effect of this invention in improving the overall quality of the product.

Claims

1. A segmented, regulated two-stage cultivation process for Ganoderma lucidum, characterized in that, Includes the following steps: (1) First stage - strain activation culture: The Ganoderma lucidum strain is inoculated onto a primary solid culture medium, which contains the following components by weight: 150-200 parts potato extract, 15-20 parts glucose, 5-10 parts peptone, 15-20 parts agar, and 1000 parts water; it is cultured for 10-15 days at a temperature of 18-22℃, a humidity of 75-80%, and in the dark until the colony diameter reaches 2-3 cm. (2) Second stage - fruiting body induction culture: The inoculum blocks cultured in the first stage are transferred to a secondary liquid culture medium containing the following components by weight: 30-50 parts corn flour, 20-30 parts wheat bran, 5-10 parts yeast extract, 1-2 parts potassium dihydrogen phosphate, 0.5-1 parts magnesium sulfate, and 1000 parts water; cultured for 25-30 days under the conditions of temperature 25-28℃, humidity 85-90%, light intensity 500-1000 lux, and photoperiod of 12 hours light / 12 hours dark, and 0.1-0.3 mg / L naphthaleneacetic acid is added on the 10th and 20th days of culture, respectively; (3) Harvesting and post-processing: When the diameter of the fruiting body of Ganoderma lucidum reaches 5-8 cm, harvest the fruiting body, rinse it with sterile water, and then dry it at 40-50℃ until the moisture content is less than 10%.

2. The cultivation process according to claim 1, characterized in that, In step (1), the potato extract is prepared by boiling and extracting fresh potatoes, and its soluble solids content is 10%-15%.

3. The cultivation process according to claim 1, characterized in that, In step (1), the culture conditions are: cultured for 12 days at 20°C, 78% humidity, and in darkness.

4. The cultivation process according to claim 1, characterized in that, In step (2), the weight ratio of corn flour to wheat bran in the secondary liquid culture medium is (35-45):(22-28).

5. The cultivation process according to claim 1, characterized in that, In step (2), the naphthaleneacetic acid is added by dissolving it in sterile water and applying it evenly to the surface of the culture medium in the form of a spray.

6. The cultivation process according to claim 1, characterized in that, In step (2), the light conditions are: light intensity of 800 lux, light cycle of 12 hours of light / 12 hours of darkness, and culture temperature of 26℃.

7. The cultivation process according to claim 1, characterized in that, In step (3), the drying is carried out by vacuum drying method, with a vacuum degree of 0.08-0.10 MPa and a drying time of 4-6 hours.

8. A fruiting body of Ganoderma lucidum obtained by the cultivation process described in any one of claims 1-7, characterized in that, Its polysaccharide content is not less than 15%, and its triterpenoid content is not less than 3%.

9. The fruiting body of Ganoderma lucidum according to claim 8, characterized in that, After being stored at room temperature for 12 months, the polysaccharide retention rate is greater than 90%, and there is no mold or insect infestation.

10. The use of the fruiting body of Ganoderma lucidum as described in claim 8 in the preparation of health products or anti-tumor adjuvant drugs.

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