CN110679392A

CN110679392A - Phellinus igniarius cultivation method

Info

Publication number: CN110679392A
Application number: CN201910913490.7A
Authority: CN
Inventors: 郭红伟; 马伟; 陈凯
Original assignee: Individual
Current assignee: Individual
Priority date: 2019-09-25
Filing date: 2019-09-25
Publication date: 2020-01-14

Abstract

The present disclosure relates to a method for cultivating phellinus igniarius, which includes the steps of: (1) culturing the cultivated species; (2) fruiting and culturing; (3) harvesting; (4) and (4) harvesting again: and (3) making a supplement liquid opening for the fungus bag picked in the step (3), connecting a supplement liquid to the supplement liquid opening, and then continuing culturing according to the conditions in the step (2), wherein the difference from the step (2) is as follows: increasing ventilation times, and picking after the sporocarps are mature; this is repeated for 2-3 times. The method can shorten Phellinus linteus production cycle, reduce pollution rate and increase fruiting body yield.

Description

Phellinus igniarius cultivation method

Technical Field

The present disclosure relates to a cultivation method of phellinus igniarius, which belongs to the cultivation of fungus organisms.

Background

The information in this background section is only for enhancement of understanding of the general background of the disclosure and is not necessarily to be construed as an admission or any form of suggestion that this information forms the prior art that is already known to a person of ordinary skill in the art.

Phellinus genus is Basidiomycetes, Polyporales, Polyporaceae, Phellinus genus (Phellinus), and is also called Morriscus, Tree chicken, Hoffonia simplicifolia, Phellinus linteus, Phellinus igniarius, Phellinus linteus, and Prunus mume. Modern medical research finds that phellinus igniarius has various pharmacological actions and has obvious curative effects in the aspects of antibiosis, immunoregulation, sweat gland secretion inhibition, tumor resistance, fibrosis resistance, inflammation diminishing, pain relieving, oxidation resistance and the like.

Modern researches have found that phellinus linteus contains polysaccharides, fatty acids, agaric acid, triterpenes, fusel alcohols, aromatic acids, amino acids, enzymes, flavones and polyphenols and other active ingredients, as well as iron, calcium, zinc, magnesium and other trace elements. These substances have effects of preventing and treating cardiovascular diseases, liver diseases, kidney diseases, nervous system diseases, etc., and have high medicinal value.

The types of Phellinus igniarius are various and comprise Phellinus baumii, Phellinus linteus, Phellinus igniarius, etc., wherein Phellinus igniarius (Phellinus igniarius) is one of Phellinus igniarius found to be high in medicinal value at present, but hyphae and fruiting bodies grow slowly, so that large-scale production and application of Phellinus igniarius are limited. For example, patent CN 108264390 a discloses a method for industrially cultivating phellinus linteus fruiting bodies, which requires inoculation for 6 months to pick the fruiting bodies, has a long cultivation period, and requires relatively expensive grain seeds as the main raw material of the cultivation medium; in addition, grain seeds are adopted as culture mediums, so that the grain seeds can grow and develop, the nutrition loss is very large, and the harvesting of fruit bodies for two times, three times and the like is not facilitated.

Disclosure of Invention

In view of the background technologies, the present disclosure provides a cultivation method of Phellinus igniarius, which has a short production period, a low pollution rate, a low cost of culture medium raw materials of cultivars, and a high yield of fruit bodies.

Specifically, the following technical scheme is adopted in the disclosure:

the present disclosure provides a cultivation method of Phellinus igniarius, which comprises the following steps:

(1) cultivation of cultivars: filling the culture medium of the cultivated species into a fungus bag, inoculating the stock species, carrying out dark culture, and ending fungus growing when the fungus bag is full of the strains;

wherein the culture medium of the cultivar consists of the following raw materials in percentage by mass: 20-25% of soybean straw, 20-25% of corncob, 3-5% of cane sugar, 2-3% of gypsum, 0.5-1% of cinnamon waste and the balance of mulberry sawdust; adding water to make the water content of the culture medium be 60-70 w/w%;

(2) fruiting and culturing:

continuing dark culture after spawn running is finished, opening openings at two sides of the fungus bag after hyphae in the fungus bag are completely changed into dark yellow from light yellow, and controlling the environment condition of fruiting;

(3) harvesting: picking up fruiting bodies at the base parts of the stalks after the fruiting bodies are mature, and then drying and storing the picked fruiting bodies;

(4) and (4) harvesting again: and (3) performing opening treatment on the fungus bags picked in the step (3) by using a supplementing liquid, connecting the supplementing liquid to the opening of the supplementing liquid, and then continuing culturing according to the step (2), wherein the difference from the step (2) is as follows: increasing ventilation times, and picking after the sporocarps are mature; repeating the above steps for 2-3 times;

wherein the supplementary liquid is prepared from the following raw materials in percentage by mass: 2.5-5 g/L of peptone, 5-10 g/L of glucose, 0.1-1 g/L of monopotassium phosphate, 0.1-1 g/L of magnesium sulfate, 0.1-1 g/L of zinc sulfate, 0.1-1 g/L of calcium sulfate and natural pH.

Compared with the related technology known by the inventor, one technical scheme of the present disclosure has the following beneficial effects:

(1) aiming at Phellinus igniarius, the inventor obtains the Phellinus igniarius cultivation method with short Phellinus igniarius production period, low pollution rate and high fruiting body yield through keen research.

(2) The cultivation method of Phellinus igniarius disclosed by the invention is high in production efficiency, the fungus bag can be reused, and three or four crops of sporocarp with the same weight can be obtained.

(3) The cultivation method of Phellinus igniarius disclosed by the invention has the advantages that the used raw materials of the culture medium are low in cost, and the large-scale production and application are facilitated.

Detailed Description

It should be noted that the following detailed description is exemplary and is intended to provide further explanation of the disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs.

It is noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of example embodiments according to the present disclosure. As used herein, the singular forms "a", "an" and "the" are intended to include the plural forms as well, and it should be understood that when the terms "comprises" and/or "comprising" are used in this specification, they specify the presence of the stated features, steps, operations, and/or combinations thereof, unless the context clearly indicates otherwise.

As described in the background art, the current artificial cultivation method of Phellinus igniarius (Phellinus igniarius) Phellinus igniarius is not ideal, and in order to solve the above technical problems, in an exemplary embodiment of the present disclosure, there is provided a cultivation method of Phellinus igniarius, which includes the steps of:

wherein the culture medium of the cultivar consists of the following raw materials in percentage by mass: 20-25% of soybean straw, 20-25% of corncob, 3-5% of cane sugar, 2-3% of gypsum, 0.5-1% of cinnamon waste and the balance of mulberry sawdust; adding water to make the water content of the culture medium be 60-70 w/w%.

(2) Fruiting and culturing:

and (3) continuing dark culture after spawn running is finished, opening two sides of the fungus bag after hyphae in the fungus bag are completely changed from light yellow to dark yellow, and controlling the fruiting environmental conditions: the day temperature is 28-32 ℃, the night temperature is 22-26 ℃, the temperature difference is 5-6 ℃, the humidity is 85-95%, scattered light is irradiated, and ventilation is carried out twice a day for 0.5-1 h each time;

(4) and (4) harvesting again: opening the fungus bag picked in the step (3), connecting a supplementing liquid to the opening, then continuing culturing according to the culture conditions in the step (2) (different from the step (2), increasing the ventilation frequency from two times per day to four times), and picking after the fruiting body is mature; repeating the above steps for 2-3 times;

wherein the supplementary liquid is prepared from the following raw materials in percentage by mass: 1.5-3 g/L of peptone, 4-6 g/L of glucose, 0.1-1 g/L of monopotassium phosphate, 0.1-1 g/L of magnesium sulfate, 0.1-1 g/L of zinc sulfate, 0.1-1 g/L of calcium sulfate and natural pH.

In the step (1), the stock is prepared by the following method:

A. activating the parent strain:

inoculating the mother seeds to a PDA comprehensive culture medium for activation, and carrying out dark culture at a constant temperature of 25-28 ℃ for 8-12 days;

the PDA comprehensive culture medium comprises: 200g of potato (peeled), 20g of glucose, 3g of monopotassium phosphate, 1.5g of magnesium sulfate, 18 g of vitamin B, 20g of agar and 1000mL of water, wherein the pH value is natural;

B. stock culture:

putting the stock culture medium into a strain bag, sealing, sterilizing at high pressure, inoculating the activated strain into the stock culture medium, performing dark culture at a constant temperature of 25-28 ℃, growing the strain in the strain bag after 25-40 days, and refrigerating the strain bag full of the strain (namely the stock) at 4 ℃ for standby or immediate use.

The method aims to screen and optimize the formula of the stock culture medium, and the stock culture medium obtained in a short time is thick in hypha, and is prepared from the following raw materials in percentage by mass through a large number of tests: 20-25% of soybean straw, 20-25% of corncob, 2-3% of gypsum, 4-6% of glucose, 0.4-0.6% of potassium hydrogen phosphate, 0.4-0.6% of magnesium sulfate and the balance of mulberry sawdust. Adding water to make the water content of the culture medium be 55-65 w/w%.

In the step (1), in the culture medium of the cultivar, the cinnamon waste refers to cinnamon residue, cinnamon leaves and other waste generated after the production of cinnamon oil. Tests prove that the pollution rate of fungus sacks and phellinus igniarius sporocarp can be effectively reduced by adding a small amount of cinnamon waste into the culture medium. The using amount of the compound is 0.5-1%, and the compound contains aromatic oil substances and terpenoids, has a certain inhibiting effect on the growth of phellinus igniarius hyphae, so the content is not suitable to be high.

The cooperation of the mulberry sawdust, the soybean straw and the corncob can keep the proper carbon-nitrogen ratio for the growth of the phellinus igniarius sporocarp, prevent the growth of hypha without limitation, advance the fruiting period, meet the requirement of rapid growth of phellinus igniarius, and ensure that the fungus bag at the later stage of the culture medium adopting the formula is not easy to shrink, so that the phellinus igniarius sporocarp can be cultured and harvested for many times. The granularity of solid matters (mulberry sawdust, soybean straws, corncobs and cinnamon waste) in the culture medium is 0.1-2 cm. Tests prove that the culture medium of the cultivar can obviously reduce the pollution rate of fungus bags and phellinus igniarius sporocarp under the condition of providing sufficient nutrient precursors for phellinus igniarius.

In the step (1), the dark culture temperature is 25-28 ℃, and the culture time is 28-35 days.

The inventor researches the influence of environmental conditions such as temperature, humidity and illumination on the growth of the phellinus igniarius sporocarp, and tests show that the specific day-night temperature difference is more beneficial to the rapid growth of the phellinus igniarius sporocarp. Therefore, in the step (2), the environmental conditions for fruiting are selected as follows: the day temperature (6: 00-18: 00) is 28-32 ℃, the night temperature (18: 00-6: 00 in the next day) is 22-26 ℃, the temperature difference is 5-6 ℃, the humidity is 85-95%, the light is scattered and irradiated, and the ventilation is carried out twice a day for 0.5-1 h each time.

In the step (3), drying treatment is carried out until the water content in the fruit body is 1-3 (w/w)%.

In the step (4), tests prove that 400-500 mL of supplement liquid is dripped into each fungus bag every month, so that the growth and development of sporocarp can be effectively promoted, and the growth period is shortened. During dripping, the dripping speed should be strictly controlled, which cannot be too fast, so as to prevent the water content of the culture medium of the cultivated species from being larger.

In the step (4), the supplementary liquid is prepared by using distilled water.

In order to produce the sporocarp with high efficiency, the method for culturing and harvesting the sporocarp for multiple times is adopted in the present disclosure, but in the experimental research process, the inventor finds that when the sporocarp is harvested for the first time and is not added with any nutrition, the growth cycle of the sporocarp is prolonged, the growth vigor is poor, and the quality is low when the sporocarp is harvested for the second time. Meanwhile, the ventilation frequency is increased, so that the water content of the culture medium of the cultivated species is kept at a specific level, and excessive water is prevented from inhibiting hypha from absorbing nutrition, thereby preventing nutrition from being transmitted to the sporocarp. In addition, if nutrition is added in an additional opening, the risk of infecting mixed bacteria is increased, therefore, cinnamon waste is added into a culture medium of a cultivated species, and the probability of pollution of fungus bags and fruiting bodies is greatly reduced.

Secondly, the inventor screens and optimizes the raw material components and the content of the supplementary liquid, and the finally obtained formula is as follows: 1.5-3 g/L of peptone, 4-6 g/L of glucose, 0.1-1 g/L of monopotassium phosphate, 0.1-1 g/L of magnesium sulfate, 0.1-1 g/L of zinc sulfate, 0.1-1 g/L of calcium sulfate and natural pH. The peptone is a mixture containing various nutrients such as peptone, peptide and amino acid, and is more beneficial to the utilization and absorption of phellinus linteus hyphae compared with other nitrogen sources through experimental verification. The addition of glucose to the supplement solution can improve energy and promote the rapid growth of Phellinus linteus fruiting body. Phosphorus ions, potassium ions, magnesium ions, zinc ions and calcium ions are involved in the composition of some enzyme proteins in the phellinus igniarius, and are essential in the growth process of the phellinus igniarius.

The phellinus igniarius culture vessel according to the present disclosure uses a fungus bag, and the material of the fungus bag may be polyethylene or polypropylene.

The inoculation procedure referred to in this disclosure should be strictly performed aseptically to prevent contamination.

Regarding the inoculation amount related to the present disclosure, 1.0-1.2 g of the culture medium seeds with each 100g of the culture medium are used, if the inoculation amount is small, the entering mixed bacteria can be rapidly propagated in the culture medium, the inoculation operation is a conventional operation in the field, and no special description is provided here.

In order to make the technical solutions of the present disclosure more clearly understood by those skilled in the art, the technical solutions of the present disclosure will be described in detail below with reference to specific embodiments.

Unless otherwise specified, the following test materials and test methods were used in the following examples.

Test materials:

(1) the strain source is as follows: the species Phellinus linteus used in this disclosure is Phellinus igniarius, No. BNCC231138, stored on PDA slant medium, which is available through conventional commercial routes.

(2) Stock activation medium (PDA integrated medium): 200g of potato (peeled), 20g of glucose, 3g of monopotassium phosphate, 1.5g of magnesium sulfate, 18 g of vitamin B, 20g of agar and 1000mL of water, and the pH value is natural. The culture medium is mainly used for strain activation.

(3) The stock culture medium is composed of the following raw materials in percentage by mass: 20% of soybean straw, 20% of corncob, 2% of gypsum, 4% of glucose, 0.6% of potassium hydrogen phosphate, 0.6% of magnesium sulfate and the balance of mulberry sawdust. Adding water to make the water content of the culture medium 60%.

(4) The culture medium for the cultivars comprises the following raw materials in percentage by mass: 20% of soybean straw, 20% of corncob, 4% of sucrose, 2% of gypsum, 0.5% of cinnamon waste and the balance of mulberry sawdust. Adding water to make the water content of the culture medium 60%.

(5) The supplementary liquid consists of the following raw materials: peptone 2g/L, glucose 6g/L, potassium dihydrogen phosphate 0.2g/L, magnesium sulfate 0.2g/L, zinc sulfate 0.2g/L, calcium sulfate 0.2g/L, and pH is natural.

Example 1

Obtaining the stock, comprising the following steps:

A. activation of the preserved strain:

inoculating the stored slant strain (mother strain) BNCC231138 to a PDA comprehensive culture medium for activation, and carrying out dark culture at a constant temperature of 25 ℃ for 8 days;

B. stock culture:

the strain bags are 15 x 40cm polyethylene bags, the stock culture medium is filled into the strain bags, the strain bags are compacted while being filled, each strain bag weighs about 3.0-3.5 kg, then sealing is carried out, after autoclaving, activated strains are inoculated to the stock culture medium, dark culture is carried out at the constant temperature of 25 ℃, the strain bags are full of the strains after 30 days, and the strain bags full of the strains (namely the stock strains) are refrigerated at 4 ℃ for standby.

Example 2

A cultivation method of Phellinus igniarius comprises the following steps:

(1) cultivation of cultivars: the culture medium of the cultivar is put into a polyethylene bag of 15X 40cm, compacted while being filled, then sealed, autoclaved, inoculated with the stock of example 1, dark cultured at a constant temperature of 25 ℃, the humidity in the culture room is controlled to be 65%, and the strain grows over the fungus bag after about 30 days.

(2) Fruiting and culturing:

continuously carrying out constant-temperature dark culture at 25 ℃, observing color change of hyphae in the fungus bag, opening rectangular openings (with the size of about 3-4 multiplied by 1-2 cm) at two sides of the fungus bag when the hyphae in the fungus bag are basically completely changed into dark yellow from the previous light yellow, controlling the environmental conditions of a fruiting culture room, wherein the day temperature (6: 00-18: 00) is 30 ℃, the night temperature (18: 00-6: 00) is 25 ℃, the humidity is about 90 percent, irradiating by scattered light, ventilating twice a day for 0.5h every time, and regularly disinfecting the fruiting culture room to prevent infection.

(3) Harvesting: harvesting at the stage when the fruiting bodies are about 9 mature at 90-92 days after opening, the weight of the fruiting bodies is basically highest, shearing the fruiting bodies at the base parts of the stalks by using scissors, observing the shapes, and drying the picked fruiting bodies until the moisture content is below 1.5%, weighing, and storing after 70g per fungus bag on average, wherein the width of each pileus is about 6-15 cm and the thickness of each pileus is 6-8 cm; thus completing the harvest of the first crop of sub-entities.

(4) Harvesting in the second crop: opening the fungus bag picked in the step (3), connecting a supplementing liquid to the opening, and then continuing culturing according to the conditions in the step (2), wherein the difference from the step (2) is that: and (4) increasing the ventilation frequency from two times to four times, harvesting the second batch of phellinus igniarius sporocarp in the 4 th month after the first batch of phellinus igniarius sporocarp is harvested according to the method in the step (3), wherein the harvested sporocarp has good growth vigor, and the average dry weight is about 70 g/fungus bag.

Example 3

A cultivation method of Phellinus igniarius comprises the following steps:

(1) cultivation of cultivars: the culture medium of the culture is put into a polyethylene bag with the diameter of 15 multiplied by 40cm, the bag is compacted while being filled, then the bag is sealed, after high-pressure sterilization, the stock seeds in the example 1 are inoculated, dark culture is carried out at the constant temperature of 28 ℃, the humidity in a culture room is 70 percent, and the strain grows over the fungus bag after 28 days.

(2) Fruiting and culturing:

continuously culturing at constant temperature of 28 ℃ in the dark, observing color change of hyphae in the fungus bag, when the hyphae in the fungus bag is changed from light yellow to dark yellow basically, opening rectangular openings (the size is about 3-4 multiplied by 1-2 cm) at two sides of the fungus bag, controlling the environmental conditions of a fruiting culture room, wherein the day temperature (6: 00-18: 00) is 30 ℃, the night temperature (18: 00-6: 00 in the second day) is 25 ℃, the humidity is about 90%, irradiating by scattered light, ventilating twice a day, and ventilating for 0.5h each time. And regularly sterilizing the fruiting culture chamber to prevent infection.

(3) Harvesting: harvesting at the stage when the fruiting bodies are about 9 mature in 95-100 th day after opening, the weight of the fruiting bodies is basically highest, shearing the fruiting bodies at the base parts of the stalks by using scissors, observing the shapes, and drying the picked fruiting bodies until the moisture content is below 1.5%, weighing, and storing after 70g per fungus bag on average, wherein the width of each pileus is about 6-15 cm and the thickness of each pileus is 6-8 cm; thus completing the harvest of the first crop of sub-entities.

Experimental example 1

The differences from example 2 are: the culture medium of the culture of the experimental example is different from the culture medium of the culture of the following: 20% of rice bran, 20% of bran, 4% of sucrose, 2% of gypsum and the balance of mulberry sawdust. Adding water to make the water content of the culture medium 60%.

Other methods and conditions were the same as in example 2.

The conventional culture medium is adopted, the pollution rate is calculated when the sporocarp is harvested in the first crop, the pollution rate reaches 15%, and the culture medium is seriously shrunk and is not suitable for secondary harvesting.

By using the method and culture medium of example 1, the contamination rate was calculated at the time of harvesting fruiting bodies in the first crop, and was only 6%, which was 9% lower than that of example 1.

The above embodiments are preferred embodiments of the present disclosure, but the embodiments of the present disclosure are not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present disclosure should be regarded as equivalent replacements within the scope of the present disclosure.

Claims

1. A cultivation method of Phellinus igniarius is characterized by comprising the following steps:

(2) fruiting and culturing:

(4) and (4) harvesting again: and (3) making a supplement liquid opening for the fungus bag picked in the step (3), connecting a supplement liquid to the supplement liquid opening, and then continuing culturing according to the conditions in the step (2), wherein the difference from the step (2) is as follows: increasing ventilation times, and picking after the sporocarps are mature; repeating the above steps for 2-3 times;

2. The method of claim 1, wherein in step (1), said stock is prepared by a process comprising the steps of:

A. activating the parent strain:

B. stock culture:

putting the stock culture medium into a strain bag, sealing, sterilizing, inoculating the activated strain to the stock culture medium, performing dark culture at a constant temperature of 25-28 ℃, growing the strain in the strain bag after 25-40 days, and refrigerating the strain bag at 4 ℃ for later use or immediately using;

further, the stock culture medium is composed of the following raw materials in percentage by mass: 20-25% of soybean straw, 20-25% of corncob, 2-3% of gypsum, 4-6% of glucose, 0.4-0.6% of potassium hydrogen phosphate, 0.4-0.6% of magnesium sulfate and the balance of mulberry sawdust; adding water to make the water content of the culture medium be 55-65 w/w%.

3. The method according to claim 1, wherein the dark culture temperature in step (1) is 25 to 28 ℃ and the culture time is 28 to 35 days.

4. The method as claimed in claim 1, wherein in the step (2), the environmental conditions for fruiting are as follows: the day temperature is 28-32 ℃, the night temperature is 22-26 ℃, the temperature difference is 5-6 ℃, the humidity is 85-95%, the light is scattered and irradiated, and the ventilation is carried out twice a day for 0.5-1 h each time.

5. The method according to claim 1, wherein in the step (3), the fruit body is dried preferably to have a water content of 1 to 3 (w/w)% in the fruit body.

6. The method according to claim 1, wherein in the step (4), 400-500 mL of the supplement liquid is dripped per month per fungus pack.

7. The method as claimed in claim 1, wherein in the step (4), different from the step (2), there are: the number of ventilation increases from two to four times a day.

8. The method according to claim 1, wherein in step (4), the makeup solution is prepared using distilled water.

9. The method of claim 1, wherein the material of the fungus sack is polyethylene or polypropylene.

10. The method as claimed in claim 1, wherein 1.0-1.2 g of seed is used per 100g of cultivation material.