CN104893992B - A kind of submerged fermentation method of Ganoderma Lucidum - Google Patents
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Abstract
The invention discloses a kind of submerged fermentation method of Ganoderma Lucidum, the natural Chinese medicinal herb raw materials such as ginseng, Bee Pollen, lucidum spore powder and Poria cocos are selected as Ganoderma Lucidum submerged fermentation culture medium, there is the yield height of purpose product, stable quality, mycelial growth is good, fermentation period is short, simple production process, extraction yield is high, nutriment enriches, cost is low, it is the medicinal fungi submerged fermentation technology of the environment-friendly type of a remarkable in economical benefits, there is significant development prospect.
Description
Technical field
The invention belongs to medicinal fungi submerged fermentation production technology and biomedicine field, and in particular to a kind of Ganoderma Lucidum
Submerged fermentation method.
Background technology
Ganoderma lucidum（Ganoderma lucidum）It is Basidiomycetes Polyporaceae（Polyporaceae）Ganoderma
（Ganodelma）The general name of fungi red sesame G.lucidumkarst and purple sesame G.japonicrnLloyd, have and strengthen the body resistance to consolidate the constitution
And other effects, quilt《This warp》Referred to as top grade.In recent years, it is a lot of to the chemical composition and clinical research of Ganoderma fungi, to ganoderma lucidum
Reproduction cell --- the research of spore medical value is also quite paid attention to.
Reishi sporule brown, avette, one end is truncate,（8.5~11.2）μm×（5.2~6.9）μm, have double wall.Every
The brown internal layer of a spore produces many needle-shaped kicks, stretches into the transparent outer layer of conidial cell wall dearly, and outer layer sporoderm is smooth.Spirit
Very tough and tensile, the general chemistry of two layers of walls of Ganoderma lucidum spore parcel, physical method is difficult to be broken, thus limit to its chemistry into
Divide, the research of bioactive ingredients.
Ganoderma lucidum is in national maximum virgin forest, Changbai Mountain and the growth of Natural Reserves in Hainan optimum.But due to more
The excessive picking of the mankind, the storage of Wild ganoderma are fewer and fewer over year.Ganoderma lucidum damages liver caused by a variety of physics and chemistry and biological factor
Wound has protective effect.No matter before liver damage occurs or after generation, liver can all be protected by taking ganoderma lucidum, mitigate hepatic injury.
Ganoderma lucidum can promote metabolism of the liver to medicine, poisonous substance, have the effect of definite for toxic hepatitis.Especially chronic hepatitis, spirit
Sesame can substantially eliminate the symptom such as dizzy, weak, nauseous, uncomfortable liver area, and can effectively improve liver function, tend to indices
Normally.So ganoderma lucidum can be used for treatment slow poisoning, all kinds of chronic hepatitis, hepatic sclerosis, hepatosis.Ganoderma lucidum can be effectively
Coronary artery dilator, increases coronary blood flow, improves Myocardial Microcirculation, strengthens the supply of myocardium oxygen and energy, therefore, to cardiac muscle
Ischemic has protective effect, can be widely used for the treatment and prevention of coronary heart diseases and angina pectoris etc..In addition, it may replace after taking ganoderma lucidum
Insulin suppresses disengaging for aliphatic acid, can improve the symptoms such as blood glucose, glucose in urine.Water-soluble polysaccharide in ganoderma lucidum, can mitigate non-pancreas islet
The occurring degree of plain dependent diabetes.
The chemical composition of Ganoderma is complex, and because of strain used, the strain place of production, cultural method, extraction process, preparation
Method is different and different.Contain polysaccharide, ucleosides, furan derivative, steroid in the fructification of Ganoderma, mycelium and spore
Ferment class, alkaloids, protein, polypeptide, amino acids, triterpenes, sequiterpene, organic germanium, inorganic salts etc..Ganoderma lucidum polysaccharide is spirit
One of principle active component of sesame.
Ganoderma lucidum is generally grown in the high and rather dark mountain forest of humidity, is mainly grown in rotten tree or the root of its trees
Portion.Ganoderma lucidum is not plant, itself cannot carry out photosynthesis, can only absorb nutriment from other organic matters or rotten tree.Ganoderma lucidum
It is the macro fungi of a kind of hard, more spore and micro-strip bitterness.When the ganoderma lucidum for reaching the maturity period will spray the spore of powdery, from
And bred.Now wild ganoderma lucidum is rarely found, and quality is not easily controlled.
Ganoderma lucidum and ganoderma lucidum bioactive substance become new because of its unique physicochemical property and magical biological function
Health food and medicine field research hotspot.However, the growth cycle length of Ganoderma Lucidum（More than 2~3 months）, by
Environment influences greatly, and low output, quality are unstable, and production cost is high, greatly affected ganoderma lucidum and its bioactive substance strong
Development in health industry.Therefore, in recent years with biotechnology, the research to Fermentation Process of Ganoderma Lucidum Mycelium Submerged fermentation, is obtained
Obtain ganoderma lucidum mycelium and its Active Growth metabolite（Ganoderic acid, ganoderma lucidum polysaccharide etc.）, and by the control of fermentation condition, shorten
Production cycle, reduces production cost, ganoderma lucidum and its bioactive substance is applied and is promoted in food and medicine.
Ginseng（Scientific name：Panax ginseng）, it is the root with meat, it is pharmaceutically acceptable.Ginseng belongs to Araliaceae, main raw
Grow in East Asia, particularly cold district.Ginseng is common medicinal material, for recovering, strengthening muscle power more afterwards, adjust hormone, reduction
Blood glucose and control blood pressure, control liver index and liver function health care etc..Saponin(e contained by ginseng root is its active ingredient, and China is long white
Hill ginseng saponin constituent is higher, but obtains and be not easy, and price is high.80.0 kilocalorie of ginseng nutrient component content (every 100 grams) heat
21.2 grams of carbohydrate, 0.7 gram of fat, 0.6 gram of protein, 3.5 grams of cellulose, is both to produce fermentation medium carbon source and nitrogen source
Active principle, and the metabolite of derivative nutritional ingredient can be promoted.
Bee Pollen（Scientific name POLLEN）, pollen contains a variety of nutriments, including 22 kinds of amino acid, 14 kinds of vitamins
With 30 various trace elements and substantial amounts of active protease, nucleic acid, flavone compound and other active materials.Pollen is rich in
Bak protein 20%-25%, total amino acid content more than 20%, free amino acid 1%-2%, carbohydrate 40%-
50%, fatty 5%-10%；Also abundant natural complex, trace element, enzyme, hormone, bioactive substance etc. are a variety of
Nutriment.Particularly the content of B family vitamin is extremely abundant.Every 100 grams of pollen, 600 microgram containing thiamine, riboflavin 1670
Microgram, 900 microgram of pyridoxol, 2700 microgram of pantothenic acid, 10000 microgram of nicotinic acid.It contains several mineral materials, in these mineral matters,
The content of some elements is respectively：Potassium 20~40%, magnesium 1~20%, calcium 1~15%, iron 1~12%, silicon 2~10%, phosphorus 1~20%.
And substantial amounts of active protease, nucleic acid, flavone compound and other active materials etc..Carried out as compounded carbons nitrogen source
During fermented and cultured, be conducive to mycelial growth.
Lucidum spore powder（English name:Ganoderma spore）.Lucidum spore powder crude protein content is 18.12%, amino
Acid content is 5.869%, crude fat content 20.39%.Unrighted acid is that advantage aliphatic acid is with oleic acid, linoleic acid
It is main.Also containing abundant triterpene compound, multiple beneficial mineral element.With lucidum spore powder deep glossy ganoderma fermenting culture medium
One of raw material, can promote the precursor inducer of mycelium transposon mutagenesis and promote a large amount of synthesis of exocellular polysaccharide.It is given birth to
Object amount, Thick many candies and thick ganoderic acid content are higher, are conducive to formation and the ganoderic acid content of ganoderma active material (ganoderic acid)
Raising.
Poria cocos（Scientific name Wolfiporia cocos）.Tuckahoe nutrient enriches, the nutritional ingredient of every 100 grams of Poria cocos：Energy 16
Kilocalorie, 1.2 grams of protein, 0.5 gram of fat, 82.6 grams of carbohydrate, 80.9 grams of dietary fiber, 0.12 gram of riboflavin, potassium 58
Milligram, 1 milligram of sodium, 2 milligrams of calcium, 8 milligrams of magnesium, 9.4 milligrams of iron, 1.39 milligrams of manganese, 0.44 milligram of zinc, 0.23 milligram of copper P 32 milli
Gram, 4.55 microgram of selenium, 0.4 milligram of nicotinic acid etc..Using Poria cocos powder as Chinese-medicinal culture medium raw material, the conjunction of ganoderma lucidum polysaccharide can be promoted
Into being conducive to the raising of ganoderma polyoses content.
The content of the invention
It is an object of the invention to provide a kind of submerged fermentation method of Ganoderma Lucidum, and the yield with purpose product is high, matter
Amount is stablized, mycelial growth is good, fermentation period is short, simple production process, extraction yield height, nutriment enrich, cost is low
The features such as, be the environment-friendly type of a remarkable in economical benefits medicinal fungi submerged fermentation technology, have significant development and application before
Scape.
To achieve the above object, the present invention adopts the following technical scheme that：
A kind of submerged fermentation method of Ganoderma Lucidum, it is characterised in that：Comprise the following steps：
（1）The inclined-plane of inoculation is put at 26-28 DEG C and cultivates 5-7d；
（2）Mycelial inclined plane inoculating will be covered with fluid nutrient medium, in shaking speed 80-100r/min, stroke 7-8cm
Reciprocal concussion, 26-28 DEG C of shaken cultivation 2-3d, is made seed suspension liquid；
（3）Seeding tank seed culture, by kind of amount 15%-20%, 27-30 DEG C of a temperature, illumination condition：Continuous darkness condition, is stirred
Speed 160-180r/min is mixed, filtrated air throughput is 2.5m per minute3, tank pressure 0.04MPa, fermented and cultured 48-72h；pH
It is down to 4.5-5.0；
（4）Ferment tank culture：Inoculum concentration 25-30%, 27-30 DEG C of temperature, mixing speed 150-200r/min, nothing
Bacterium air vent amount is 2m per minute3, tank pressure 0.05MPa, cultivates 48-72h；Zymotic fluid retrogradation, thalline start self-dissolving, and pH is down to
3.0-3.5 residual sugar amount is down to 0.2-0.3%；
（5）Enzymolysis：Add compound protease 0.5%, amylase 0.75%, cellulase 0.75%；Protease is 2 × 105u/
Kg, amylase are 1.0 × 105U/kg, cellulase are 1.5 × 105u/kg；55-60 DEG C of temperature, time 3-4h.
Slant medium：Potato liquor 1000mL, skimmed milk power 30g, glucose 30g, agar 30g, peptone 30g,
Yeast extract 5g, magnesium sulfate 0.2g, vitamin B1 0.1g, pH are natural；
Submerged fermentation culture medium, percentage：Ginseng pulverate 2.0%, Bee Pollen 2.0%, lucidum spore powder 1.0%, Fu
Siberian cocklebur powder 1.0%, soy meal 2.0%, dusty yeast 0.5%, glucose 3.0%, magnesium sulfate 0.2%, dipotassium hydrogen phosphate 0.3%, sodium chloride
0.2%th, vitamin B1 0.1%, camellia oil 0.3%, surplus are water, pH=6-7, fineness of powder：80-100 mesh.
The extracting method of tunning comprises the following steps：
（1）High-pressure homogeneous processing：Zymotic fluid through enzymolysis processing is first through colloid mill homogeneous, rotation mill rotating speed 800-1000r/
Min, is disintegrated hyphostoma particle, reaches homogeneous, then through high pressure homogenizer, make zymotic fluid under the effect of 40MPa-60MPa high pressures
Pass through 200 mesh membrane filtrations；
（2）Supercritical CO2Extraction：In the supercritical state, supercritical carbon dioxide is contacted with material to be separated, made
The boiling point height component different with molecular size range is selectively extracted precipitation effective active matter by it successively；
（3）Zymotic fluid concentrates：Zymotic fluid is concentrated into concentration Brix=28-32%；
（4）Using vacuum freeze drying, condenser temperature≤- 60 DEG C, obtain tunning ganoderma lucidum polysaccharide and ganodenic acid is lived
Property material dry powder product.
The remarkable advantage of the present invention is：Select the natural Chinese medicinal herb raw materials such as ginseng, Bee Pollen, lucidum spore powder and Poria cocos
As Ganoderma Lucidum submerged fermentation culture medium, yield height, stable quality, mycelial growth with purpose product are good, fermentation week
Phase is short, simple production process, the features such as extraction yield is high, nutriment enriches, cost is low, be a remarkable in economical benefits ring
The medicinal fungi submerged fermentation technology of guarantor's type, has significant development prospect.
Using Chinese herbal medicines such as natural plants ginseng pulverate, Bee Pollen, lucidum spore powder, Poria cocos powders as ganoderma lucidum deep-fermentation
The raw material of culture medium, Optimal Medium nutriment, makes full use of nitrogen source, carbon source, promotes ganoderma lucidum mycelium growth, improves and obtain
A large amount of thalline and metabolite, greatly improve the content of the effective active compositions such as ganoderma lucidum polysaccharide and triterpene.Using natural plants people
Join the Chinese herbal medicines such as powder, Bee Pollen, lucidum spore powder, Poria cocos powder as Ganoderma Lucidum submerged fermentation culture medium, and use supercritical CO2
Extraction, enzymolysis process, extract ganoderma lucidum polysaccharide and triterpene there is not yet relevant report.
Embodiment
Embodiment 1：
Strain：By the bacterial strain strain of Changbai Mountain in China Wild ganoderma tissue isolation and selection.
Submerged culturing method for making and tunning dry powder the extraction preparation method of Ganoderma Lucidum：
1st, slant medium is prepared：Potato liquor 1000ml, skimmed milk power 30g, glucose 30g, agar 30g, albumen
Peptone 30g, yeast extract 5g, magnesium sulfate 0.2g, vitamin B1 0.1g, pH are natural.
2nd, slant strains culture：Good quality strain is seeded in solid slope culture medium, is put at 26-28 DEG C and cultivates 5-
7d, mycelium cover with inclined-plane, and micro-strip is white, and growth is vigorous.
3rd, triangular flask Shaking culture：Slant strains are moved into the triangular flask for containing 200ml fluid nutrient mediums, turned using shaking table
Fast 80-100r/min, stroke 7-8cm back and forth shake, 26-28 DEG C of shaken cultivation 2-3d, cover with the mycelia of white, seed is made and hangs
Supernatant liquid.
4th, submerged fermentation culture medium is prepared.Fluid nutrient medium is prepared by following percentage by weight：Submerged fermentation culture medium（Press
Percentage by weight）：Ginseng pulverate 2.0%, Bee Pollen 2.0%, lucidum spore powder 1.0%, Poria cocos powder 1.0%, soy meal 2.0%, dusty yeast
0.5%th, glucose 3.0%, magnesium sulfate 0.2%, dipotassium hydrogen phosphate 0.3%, sodium chloride 0.2%, vitamin B1 0.1%, camellia oil
0.3%, surplus：Water, pH：6.5, fineness of powder：80-100 mesh.（Bee Pollen selects Flos Camelliae Japonicae pollen）
By above-mentioned percentage by weight, ginseng pulverate 6000g, Bee Pollen 6000g, lucidum spore powder 3000g, Poria cocos powder are taken
3000g, soy meal 6000g, dusty yeast 1500g, glucose 9000g, magnesium sulfate 600g, dipotassium hydrogen phosphate 900g, sodium chloride
600g, vitamin B1 300g, camellia oil 900g, surplus：Water, pH：6.5, fineness of powder：80-100 mesh.Prepare Liquid Culture
Base.Culture medium sterilizes 30min under the conditions of 120-125 DEG C, spare after cooling.（Bee Pollen selects Flos Camelliae Japonicae pollen）
5th, seeding tank seed expands culture（One grade fermemtation）：Seeding tank uses 500L stainless cylinder of steel, and feed intake 300L.By seed
Suspension moves into seeding tank, carries out seed and expands culture, by kind of amount 17.5%, 27-30 DEG C of a temperature, illumination condition：Continuous darkness bar
Part, mixing speed 160-180r/min, filtrated air throughput are per minute 1:0.4m3V/v, min, tank pressure 0.04MPa, fermentation
Cultivate 56h；PH is down to 4.7, and seeding tank Spawn incubation is ripe.
6th, fermentation tank Submerged fermentation（Second order fermentation）：Fermentation tank uses 5000L stainless cylinder of steel, and feed intake 3000L.By kind
Amount 27.5%, 27-30 DEG C of temperature, illumination condition：Continuous darkness condition, mixing speed 160-180r/min, filtrated air throughput
For per minute 1:0.4m3V/v, min, tank pressure 0.04MPa, fermented and cultured 60h；PH is down to 3.3, and residual sugar amount is down to 0.25%, fermentation
Terminate, form submerged fermentation mycelia liquid.
7th, digest：By weight percentage, compound protease 0.5%, amylase 0.75%, cellulase 0.75%, albumen are added
Enzyme is 2 × 105U/kg, amylase are 1.0 × 105U/kg, cellulase are 1.5 × 105u/kg.Temperature is maintained at 55-60 DEG C,
Enzymolysis time 3.5h.
8th, high-pressure homogeneous processing：Zymotic fluid through enzymolysis processing is first through colloid mill homogeneous, rotation mill rotating speed 800-1000r/
min.The material particles such as mycelium are disintegrated, reach homogeneous, then through high pressure homogenizer（40MPa-60MPa）, make zymotic fluid in height
Pressure effect is lower to pass through 200 mesh membrane filtrations.
9th, supercritical CO2Extraction, using the relation of the solvability and its density of supercritical carbon dioxide, in supercritical state
Under state, supercritical carbon dioxide is contacted with material to be separated, makes it selectively boiling point height and molecular size range
Different components extracts precipitation effective active matter successively.
10th, zymotic fluid concentrates, using dual-effect concentrator, 55-70 DEG C of evaporating temperature, steam pressure 0.25MPa, by zymotic fluid
Liquid concentration is to concentration Brix=30%.
11st, vacuum freeze drying, using working vacuum degree≤130Pa, the vacuum freeze drying of condenser temperature≤- 60 DEG C is set
Tunning active material dry powder that is standby, obtaining.
12nd, tunning active material dry powder meter 106.5kg, yield 35.5g/L are obtained through measuration.（Dry powder is real
Measured moisture content is 5.0%）.
Production, embodiment 2 and embodiment 3 are repeated by above-mentioned implementation and proportioning：
Embodiment 2, tunning active material dry powder meter 108.5kg, yield 36.2g/L are obtained through measuration.（It is dry
Powder observed watercut is 5.0%）.
Embodiment 3, tunning active material dry powder meter 107.8kg, yield 35.9g/L are obtained through measuration.
（Dry powder observed watercut is 5.0%）.
1 Ganoderma Lucidum fermentation dry powder main component of table and content（G/100g dry weights）
Above-mentioned main component examination criteria and method：
1st, polysaccharide body（With glucose meter）, phend-sulphuric acid.
2nd, total triterpene（In terms of ursolic acid）, ultraviolet spectrophotometry.
3rd, protein（In terms of butt, N × 6.25）, using GB5009-2010 standard detections.
4th, the method that amino acid measures amino acid in food using GB/T 5009.124-2003 automatic amino acid analyzers
Measure.
5th, soluble dietary fiber, using GB/T5009.88 standard determination methods.
The foregoing is merely presently preferred embodiments of the present invention, all equivalent changes done according to scope of the present invention patent with
Modification, should all belong to the covering scope of the present invention.
Claims (2)
- A kind of 1. submerged fermentation method of Ganoderma Lucidum, it is characterised in that：Comprise the following steps：（1）The inclined-plane of inoculation is put at 26-28 DEG C and cultivates 5-7d；（2）Mycelial inclined plane inoculating will be covered with fluid nutrient medium, it is past in shaking speed 80-100r/min, stroke 7-8cm Multiple concussion, 26-28 DEG C of shaken cultivation 2-3d, is made seed suspension liquid；（3）Seeding tank seed culture, 27-30 DEG C of inoculum concentration 15%-20%, temperature, illumination condition：Continuous darkness condition, stirring speed 160-180r/min is spent, filtrated air throughput is 2.5m per minute3, tank pressure 0.04MPa, fermented and cultured 48-72h；PH drops To 4.5-5.0；（4）Ferment tank culture：Inoculum concentration 25-30%, 27-30 DEG C of temperature, mixing speed 150-200r/min, sterile sky Gas throughput is 2m per minute3, tank pressure 0.05MPa, cultivates 48-72h；Zymotic fluid retrogradation, thalline start self-dissolving, and pH is down to 3.0-3.5 residual sugar amount is down to 0.2-0.3%；（5）Enzymolysis：Add final concentration of 0.5% compound protease, final concentration of 0.75% amylase, final concentration of 0.75% Cellulase；Protease is 2 × 105U/kg, amylase are 1.0 × 105U/kg, cellulase are 1.5 × 105u/kg ；Temperature 55-60 DEG C of degree, time 3-4h；Slant medium：Potato liquor 1000mL, skimmed milk power 30g, glucose 30g, agar 30g, peptone 30g, yeast Cream 5g, magnesium sulfate 0.2g, vitamin B1 0.1g, pH are natural；Submerged fermentation culture medium, percentage：Ginseng pulverate 2.0%, Bee Pollen 2.0%, lucidum spore powder 1.0%, Poria cocos powder 1.0%th, soy meal 2.0%, dusty yeast 0.5%, glucose 3.0%, magnesium sulfate 0.2%, dipotassium hydrogen phosphate 0.3%, sodium chloride 0.2%, dimension Raw element B1 0.1%, camellia oil 0.3%, surplus are water, pH=6-7, fineness of powder：80-100 mesh, is made fluid nutrient medium.
- 2. the submerged fermentation method of the Ganoderma Lucidum according to claim 1, it is characterised in that：It is further comprising the steps of：（1）High-pressure homogeneous processing：Zymotic fluid through enzymolysis processing first grinds rotating speed 800-1000r/min through colloid mill homogeneous, rotation, Hyphostoma particle is disintegrated, reaches homogeneous, then through high pressure homogenizer, zymotic fluid is passed through under the effect of 40MPa-60MPa high pressures 200 mesh membrane filtrations；（2）Supercritical CO2Extraction：In the supercritical state, supercritical carbon dioxide is contacted with material to be separated, there is it The boiling point height component different with molecular size range is optionally extracted precipitation effective active matter successively；（3）Zymotic fluid concentrates：Zymotic fluid is concentrated into concentration Brix=28-32%；（4）Using vacuum freeze drying, condenser temperature≤- 60 DEG C, obtain tunning ganoderma lucidum polysaccharide and ganodenic acid active matter Matter dry powder product.
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| CN105935143B (en) * | 2016-06-29 | 2019-02-26 | 浙江蕴本生物科技有限公司 | The production method of ganoderma lucidum ferment |
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| CN111304125A (en) * | 2020-03-02 | 2020-06-19 | 广州金塞迩生物医学科技有限公司 | A strain compound for high-efficiency Chinese medicinal fermentation, its preparation method, application, and Chinese medicinal fermented product fermented by the same |
| CN113303419A (en) * | 2021-06-15 | 2021-08-27 | 山东效峰生物科技股份有限公司 | Preparation process of medicinal and edible fungus mixed fermentation mycelium powder |
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| CN103966107A (en) * | 2014-05-13 | 2014-08-06 | 大连佳腾生物科技有限公司 | Culture medium and method for producing multi-component high-activity ganoderma lucidum powder by using same |
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