CN107586727A - A kind of fermentation process of ganoderma lucidum mycelium - Google Patents
A kind of fermentation process of ganoderma lucidum mycelium Download PDFInfo
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Abstract
The present invention relates to a kind of fermentation process of ganoderma lucidum mycelium, belong to fungal culture technical field.The fermentation process of ganoderma lucidum mycelium provided by the invention comprises the following steps：1) lucidum strain after activation is inoculated in liquid fermentation medium, 23~26 DEG C of 5~7d of culture, throughput is 0.6~1V/Vmin, and air pressure is 0.03~0.06Mpa, obtains fermented bacterium；2) fermented bacterium for obtaining step 1), which is inoculated in solid fermentation culture medium, carries out light culture, and the time of the light culture is 30~40d.Solid fermentation is combined by glossy ganoderma mycelium fermentation method provided by the invention with liquid fermentation technology, has the advantages of production scale is big, with short production cycle, production technology is easy, active constituent content is high, product recoveries are high.
Description
Technical field
The present invention relates to fungal culture technical field, and in particular to a kind of fermentation process of ganoderma lucidum mycelium.
Background technology
GL-B has the different physiological roles such as antitumor, raising immunity, anti-aging, reducing blood lipid, there is larger medicine
With and health value, by current the world of medicine and the common concern of food industry, its manufactured medicine and health products are deep by people
Like that there is wide DEVELOPMENT PROSPECT.
The extracting mode of effective component of glossy ganoderma includes：Fructification extraction, mycelium extraction, the composition of the two extraction are poor
Different, component content respectively has height, there is the characteristics of it can not be substituted for each other.Wherein mycelium extraction uses cultural hypha technique
Comprising：Liquid fermentation and culture and solid fermentation culture, liquid fermentation be finally extract it is true in bacterium ball and leachate concentrate
The active ingredients such as granulose, albumen, advantage be extract purity or content it is higher, apply in terms of high potency drugs are prepared compared with
More, its shortcoming is zymotechnique requirement height, and input is high, and extraction process is complicated, recovery rate is relatively low.The advantages of solid fermentation is input
Low, simple process, product recoveries are high, and shortcoming is that extraction content is low, cultivation cycle length, can be only applied to often effect class health medicine
With the aspect of health products.Prior art does not have a kind of fermentation process of efficient ganoderma lucidum mycelium.
The content of the invention
It is an object of the invention to provide a kind of fermentation process of ganoderma lucidum mycelium.Glossy ganoderma fermentation method provided by the invention
Solid fermentation is combined with liquid fermentation technology, have production scale is big, with short production cycle, production technology is easy, effectively into
Divide the advantages of content is high, product recoveries are high.
The invention provides a kind of synthesis fermentation process of ganoderma lucidum mycelium, comprise the following steps：
1) lucidum strain after activation is inoculated in liquid fermentation medium, 23~26 DEG C of 5~7d of culture, throughput is
0.6~1V/Vmin, air pressure are 0.03~0.06Mpa, obtain fermented bacterium；
The liquid fermentation medium includes the component of following weight/mass percentage composition：Glucose 0.5~1%, dregs of beans 0.1~
0.2%, corn flour 0.1~0.2%, wheat flour 0.1~0.2%, yeast extract 0.1~0.3%, magnesium sulfate 0.003~
0.005%, potassium dihydrogen phosphate 0.08~0.1%, vitamin B compound 0.001% and defoamer 0.03~0.05%, surplus are
Water；
2) fermented bacterium for obtaining step 1), which is inoculated in solid fermentation culture medium, carries out light culture, the light culture
Time is 30~40d.
Preferably, inoculum concentration of the lucidum strain in the step 1) after activation in liquid fermentation medium is
0.1%~0.2%.
Preferably, the biomass for the fermented bacterium that the step 1) obtains is (2.00 ± 0.50) × 10-2G/mL, bacterium ball
Average diameter is 1.00 ± 0.10mm, and Peloton density is 120.00 ± 2.00/mL, and pH value is 5~6.5.
Preferably, the solid fermentation culture medium of the step 2) is bacterium bag bag solid fermentation culture medium.
Preferably, the inoculum concentration of fermented bacterium is 50~100mL/kg in the step 2).
Preferably, the carbon-nitrogen ratio in the solid fermentation culture medium is (20~28)：1, water content is 42~50%, is gone out
The pH value of culture medium is 7~8 before bacterium.
Preferably, the solid fermentation culture medium includes the component of following weight/mass percentage composition：Rice 5~10% is small
Wheat 10~15%, corn flour 35%~45%, corncob 5~8%, dregs of beans 5~10%, wheat bran 15~25%, calcium hydroxide 1~
2% and precipitated calcium carbonate 1~2%.
Preferably, the humidity of the step 2) light culture is 60~65%；The light culture includes following 4 stages：
1~9d, the temperature of the light culture is 25~26 DEG C, CO2Concentration is 1000~1500ppm；
10~15d, the temperature of the light culture is 23~24 DEG C, CO2Concentration is 1500~2000ppm；
16~30d, the temperature of the light culture is 24~25 DEG C, CO2Concentration is 2000~3000ppm；
30d is to completion is cultivated, and the temperature of the light culture is 25~26 DEG C, CO2Concentration is 1500~2000ppm.
Preferably, de- bag, section and baking process are also included after the step 2) light culture.
The invention provides a kind of synthesis fermentation process of ganoderma lucidum mycelium.Glossy ganoderma mycelium fermentation side provided by the invention
Solid fermentation is combined by method with liquid fermentation technology, combines the advantage of solid, liquid fermentation, has production scale big, raw
Produce the advantages of cycle is short, production technology is easy, pollution rate is low, active constituent content is high, product recoveries are high.Be advantageous to subsequently have
Imitate the high efficiency extraction of composition and the recovery of product.The fermentation process of the present invention has production to advise compared to existing solid fermentation
The advantages of mould is big, with short production cycle, pollution rate is low, compared to liquid fermentation and culture, there are production technology simplicity, active constituent content
High, the advantages of product recoveries are high.The mycelium mixture of the comprehensive fermentation process culture, ganoderma volatile oil recovery rate are up to
0.03%, Ganoderma Lucidum acid extraction rate reached 3.2%, containing sour solid product acid purity up to more than 25%, GL-B recovery rate can
Up to 10.1%, purity of polysaccharide is up to more than 45%.
Embodiment
The invention provides a kind of synthesis fermentation process of ganoderma lucidum mycelium, comprise the following steps：
1) lucidum strain after activation is inoculated in liquid fermentation medium, 23~26 DEG C of 5~7d of culture, throughput is
0.6~1V/Vmin, air pressure are 0.03~0.06Mpa, obtain fermented bacterium；
The liquid fermentation medium includes the component of following weight/mass percentage composition：Glucose 0.5~1%, dregs of beans 0.1~
0.2%, corn flour 0.1~0.2%, wheat flour 0.1~0.2%, yeast extract 0.1~0.3%, magnesium sulfate 0.003~
0.005%, potassium dihydrogen phosphate 0.08~0.1%, vitamin B compound 0.001% and defoamer 0.03~0.05%, surplus are
Water；
2) fermented bacterium for obtaining step 1), which is inoculated in solid fermentation culture medium, carries out light culture, the light culture
Time is 30~40d.
What the present invention was carried out first is the preparation of lucidum strain after activating.In the present invention, the activation number of the strain
Preferably 2 times, by being cultivated to obtain first class inoculum and second class inoculum in activation medium, the present invention chooses second class inoculum
It is used for liquid fermentation and culture as the lucidum strain after activation.In the present invention, the first class inoculum activation medium is preferably adopted
With well known to those skilled in the art plus rich potato culture, second class inoculum activation medium is preferably that every liter of water includes Portugal
Grape sugar：10~20g, peptone：2~4g, yeast extract：1~2g, potassium dihydrogen phosphate：1~2g, epsom salt：0.75~1.5g
And vitamin B compound：0.25~0.5g.The present invention does not have special restriction to the source of the lucidum strain, using this area
Conventional commercial lucidum strain known to technical staff, preferred choose of the present invention derive from China General Microbiological culture presevation
Administrative center, bacterium numbering are CGMCC5.743 lucidum strain.
The preferred mycelia for choosing the speed of growth as 5.5~6.5mm/d of the invention is as first class inoculum.The present invention is in one-level bacterium
During the selection of kind, neat, the pure white and strong bacterial strain of colony edge is preferably chosen.The preparation of first class inoculum of the present invention can
The weaker original strain of growth is screened out out, ensures the activity of strain.The condition of first class inoculum culture of the present invention is preferably 22
~28 DEG C of 5~8d of light culture, more preferably 23 DEG C of light culture 7d.
After obtaining first class inoculum, the preferred mycelia for choosing 1.5~2.5mm of first class inoculum colony edge of the present invention is placed in two level
Cultivated in strain activation and culture base, the condition of the activation culture is preferably 5~7d of constant-temperature shaking culture, chooses biomass
For 1.5~2.5 × 10-2G/mL strain, as second class inoculum.The present invention is before constant-temperature shaking culture, preferably by first class inoculum
Stand to mycelia block and sprout in two level activation medium, the condition stood is preferably 23~25 DEG C of standings under dark condition
28~32h, more preferably 24 DEG C standing 30h.In the present invention, the temperature of the shaken cultivation is preferably 23~26 DEG C, more excellent
Elect 24 DEG C as.Shaken cultivation of the present invention is preferably carried out using constant-temperature table, and the rotating speed of the shaking table is preferably 140~160r/
Min, more preferably 150r/min.In the present invention, second of activation culture can ensure the homology and synchronism of strain.
In the present invention, second activate after obtain the preferred size of strain, be evenly distributed, do not sink, sphere covers with fine hair, culture medium face
Substantially shoaled after color relatively sterilizing and transparent.In existing Ganoderma lucidum submerged fermentation technology, secondary metabolite is easily produced, and it is our
Method strictly by the activation of strain twice, screening, can ensure the homology and synchronism of strain, while inoculum concentration is according to we
Method ensures 0.1~0.2%, can greatly shorten the liquid fermentation time, ensures that the activity of tunning synchronously reaches peak value, have
Beneficial to follow-up solid fermentation.
After lucidum strain after being activated, the lucidum strain after activation is inoculated in liquid fermentation medium by the present invention
In, 23~26 DEG C of 5~7d of culture, throughput is 0.6~1V/Vmin, and air pressure is 0.03~0.06Mpa, obtains liquid fermentation
Strain.In the present invention, the liquid fermentation medium includes the component of following weight/mass percentage composition：Glucose 0.5~1%,
Bean cake powder 0.1~0.2%, corn flour 0.1~0.2%, wheat flour 0.1~0.2%, yeast extract 0.1~0.3%, magnesium sulfate
0.003~0.005%, potassium dihydrogen phosphate 0.08~0.1%, vitamin B compound 0.001% and defoamer 0.03~0.05%,
Surplus is water.In the present invention, the glucose and corn flour mainly provide the carbon source of culture medium, bean cake powder, wheat flour and ferment
Female cream mainly provides the nitrogen source of culture medium, and the pH in potassium dihydrogen phosphate major control culture matrix, potassium ion provides metabolism environment,
Magnesium ion is the activator of a variety of enzymes in magnesium sulfate.In the culture medium of the present invention, the preferred Angel Yeast LM800 of yeast extract, grape
The pure grade of chemicals Optimization Analysis, the sources of remaining each component such as the preferred food grade of sugar, magnesium sulfate, urea do not have special limit
It is fixed, using the conventional commercial product of each component well known to those skilled in the art.In the present invention, it is living in the step 1)
Inoculum concentration of the lucidum strain in liquid fermentation medium after change is 0.1%~0.2%.In the present invention, the liquid obtained
The biomass of fermented bacterium is (2.00 ± 0.50) × 10-2G/mL, bacterium mean diameter of a ball are 1.00 ± 0.10mm, and Peloton density is
120.00 ± 2.00/mL, pH value is 5~6.5.In the present invention, the cultivation temperature of the liquid fermentation and culture is 23~26
DEG C, more preferably 24 DEG C.In the present invention, the time of the liquid fermentation and culture is 5~7d, more preferably 6d.In the present invention
In, the throughput of the liquid fermentation and culture is 0.6~1V/Vmin, more preferably 0.8V/Vmin.Present invention ventilation is excellent
Choosing is handled air using filtrated air system, and cardinal principle is that pure air carries out air compression, tool by air compressor machine
Body, gases at high pressure are preserved by air accumulator, dehumidified by freeze drier, dedusting, cooling air to 22~24 DEG C, by double
The microorganisms such as 0.1 μm of micro-pore-film filtration bacterium of layer, are finally passed through culture apparatus.In the present invention, the liquid fermentation and culture
Air pressure is 0.03~0.06Mpa, more preferably 0.05Mpa.In the present invention, the gas being passed through in the fermentation process is preferably
Cold dry filtrated air.
The present invention does not have special restriction to the specific operation process of the liquid fermentation and culture, using people in the art
The known liquid fermentation method of member.Specifically fermented using following methods：In the present invention, the liquid fermentation and culture
Zymolysis Equipment specifically preferably includes：Steam sterilizing system, cold cycling water system, filtrated air system and sterile pipes inoculation system
System.In the present invention, the steam sterilizing system is used for the sterilizing of installation for fermenting, and formula sterilizing, steam pressure are passed through using steam
Preferably 0.12Mpa.Installation for fermenting of the present invention is preferably fermentation tank, and material is preferably SUS304 stainless steels, and volume is preferred
For 800L, described fermentation tank includes interlayer, and the interlayer is act as by recirculated water thermostatic control fermentation temperature.This hair
The bright sterilization method to fermentation medium does not have special restriction, using sterilizing methods well known to those skilled in the art.
Specifically, the raw material in addition to defoamer is preferably stirred when water temperature rises to 75 DEG C, waits fermentation tank by the present invention with hot water dissolving
Interior water temperature rises to 90 DEG C, adds raw material and defoamer, 121 DEG C of sterilizing 45min, after the completion of sterilizing, utilizes cold and hot water circulation system
Cold water cooling processing is carried out by interlayer, be inoculated with again after culture medium is cooled to 24 DEG C.The present invention is in liquid fermentation mistake
Cheng Zhong, the data such as fermentation pressure tank, temperature, smell, throughput are preferably observed per 2h and are recorded.In the present invention, fermented and cultured
During it is preferred carry out two sub-sampling detections, to judge whether bacterium solution is contaminated, detection method is preferably the purpose of detection：Smell
Judge that bacterium solution has free from extraneous odour, naked eyes to judge bacterium solution whether there is discoloration muddiness, Gram's staining microscope inspection whether there is bacterium infection, flat board
Line detects the presence of bacterium and other fungal contaminations.
After obtaining fermented bacterium, fermented bacterium is inoculated in solid fermentation culture medium by the present invention by sterile pipes to be carried out
Light culture.The strain seeded process is preferably inoculated with using sterile pipes inoculation system, and specifically, the inoculation pipeline is adopted
With high pressure steam sterilization, sterilising conditions are preferably 121 DEG C, 45min, are inoculated with after tube-cooled.In the present invention, it is described
The time of light culture is 30~40d；In the present invention, the carbon-nitrogen ratio in the solid fermentation culture medium is (20~28)：1, contain
Water is 42~50%, and the pH value of culture medium is 7~8 before sterilizing.In the present invention, the time of the light culture is preferably 35d,
Carbon-nitrogen ratio in the solid medium is preferably 25：1, water content is preferably 48%, and the setting of the water content can ensure
The completeness of sterilizing, the requirement of the elasticity and mycelia of bacterium bag to moisture, the pH value of culture medium is preferably before present invention sterilizing
7.5.In the present invention, the setting of carbon-nitrogen ratio can ensure the growth of ganoderma lucidum mycelium, and energy in the solid fermentation culture medium
Ensure that mycelium growth vigor is normal on the premise of cost of material is minimum.In the present invention, the solid fermentation culture medium of the step 2)
Preferably bacterium bag bag solid fermentation culture medium, the i.e. solid culture of ganoderma lucidum mycelium are carried out in bacterium bag bag.The present invention is to bacterium bag
The material of bag does not have special restriction, preferred using PP well known to those skilled in the art (polypropylene) bacterium bag bag, specification
For 36cm × 15cm × 0.5mm's.The bacterium bag bag, which often wraps, preferably fills 2.5 ± 0.5 jin of culture matrixes, and the height of the bacterium bag is excellent
Elect 17.5~18cm as.In the present invention, the sterilising conditions after the bacterium bag sacked material are preferably 123 DEG C of sterilizing 200min.
In the present invention, the inoculum concentration of the fermented bacterium is 50~100mL/kg, more preferably 60mL/kg.The present invention
There is no special restriction to the inoculation method of fermented bacterium, be inoculated with i.e. using classification inoculation apparatus well known to those skilled in the art
Can.Specifically, it is 17.5cm that the present invention, which chooses length, the inoculating gun of four eight fluid holes of row is averagely set to be inoculated with, it is more excellent
Choosing, the present invention utilizes the single output of prset counter and solenoid valve control strain, specifically, is inoculated with gnotobasis,
Inoculating gun is inserted in plup inlet during inoculation, is stepped on switch and is carried out material spray, strain is uniformly sprayed to charge level, in material and expects bottom.
In the present invention, the solid fermentation culture medium preferably includes the component of following weight/mass percentage composition：Rice 5~10%, wheat 10
~15%, corn flour 35%~45%, corncob 5~8%, dregs of beans 5~10%, wheat bran 15~25%, calcium hydroxide 1~2%
With precipitated calcium carbonate 1~2%.In the present invention, the solid fermentation culture medium preferably also include weight/mass percentage composition be 0.1~
0.2% urea.In the present invention, the particle diameter of the corn flour is preferably 0.6~1.0mm, and the particle diameter of the dregs of beans is preferably
0.8~1.2mm, the particle diameter of the rice is preferably 2.0~2.5mm, and the particle diameter of the wheat is preferably 2.4~2.8mm.At this
In invention, the corn flour, dregs of beans, the particle diameter of rice and wheat can ensure bacterium bag unitary physical structure, and balance bacterium bag is breathed freely
Property with sterilizing completeness.In the present invention, the wheat preferably soaks 36h before use with 25 DEG C of water, and the rice is excellent
Select 25 DEG C of water immersion 12h.
In the present invention, the humidity of the step 2) light culture is 60~65%；The light culture includes following 4 ranks
Section：
1~9d, the temperature of the light culture is 25~26 DEG C, CO2Concentration is 1000~1500ppm；
10~15d, the temperature of the light culture is 23~24 DEG C, CO2Concentration is 1500~2000ppm；
16~30d, the temperature of the light culture is 24~25 DEG C, CO2Concentration is 2000~3000ppm；
30d is to completion is cultivated, and the temperature of the light culture is 25~26 DEG C, CO2Concentration is 1500~2000ppm.
In the present invention, de- bag, section and baking process are also included after the light culture, obtains sheet fungus block.In this hair
In bright, the complete ganoderma lucidum mycelium of bacterium bag bag solid fermentation medium culture preferably carries out de- bag, takes off the mycelium after bag and preferably enters
The progress of slicer tool is preferably made in row section, in the present invention, the slicing processes by oneself, and the thickness of the section is preferably 1~
1.5cm, present invention preferably employs drip pan with holes to be dispensed.In the present invention, baking, which is preferable in drying room, is carried out, the drying room
The preferred 360m of volume3, the baking preferably uses jet chimney space heating, preferably power 2.2kw, air quantity 9000m3/ h axle
Flow fan air inducing is dehumidified by the auxiliary heat of jet chimney fin, preferably power 0.55kw, 3000~4000m of air quantity3/ h centrifugation
Blower fan circulated heat and hydrofuge, baking temperature are preferably between 70~75 DEG C.The baking method of the present invention can ensure：Total steam
Flow is 1.25~1.5 tons/time, and single drying output is 1.8 tons, and baking time is less than 22h, and fungus block moisture is not after baking
7%.After the present invention obtains sheet fungus block, there is no special restriction to the following process process of sheet fungus block, using this area skill
Art personnel's conventional machining mode, after pulverization process, obtains ganoderma lucidum powder；Or with supercritical CO2Fluid extraction bacterium powder obtains
To ganoderma volatile oil, GL-B and ganoderic acid.
A kind of fermentation process of ganoderma lucidum mycelium of the present invention is done with reference to specific embodiment further in detail
Introduction, technical scheme includes but is not limited to following examples.
Embodiment 1
Ganoderma lucidum original strain is inoculated in multiple flat board culture dishes equipped with culture medium and cultivated.
The specific preparation method of the ganoderma lucidum one-level activated spawn comprises the following steps：
Ganoderma lucidum original strain derives from China General Microbiological culture presevation administrative center, and bacterium numbering is
CGMCC5.743。
The ganoderma lucidum original strain is transferred in 24 DEG C of constant incubator under 4 DEG C of storage conditions, recovers biology enzyme
Activity, be placed in after 24h stand-by in superclean bench.
For the ganoderma lucidum one-level activated spawn using rich potato culture is added, specific manner of formulation is as follows：
(1) potato 200g is cleaned, remove the peel, removes eye, the thickness for the 0.2~0.3cm that cuts into slices；
(2) gained potato is placed in hot water, first boils 3~5 minutes to softening with big fire, then be changed to be cooked by slow fire 10 points
Clock, fill potato nutritional and be dispersed in water；
(3) with 6~8 layers of filtered through gauze, leachate is retained；
(4) yeast extract 1g, peptone 2g, potassium dihydrogen phosphate 1g, epsom salt are added in leachate obtained by step (3)
0.5g, vitamin B compound 1g, glucose 20g, agar powder 20g, 1000mL are settled to water；
(5) mixed liquor of step (4) is sub-packed in the culture dish of multiple 10cm specifications, culture medium thickness is 5mm or so.
Dispense culture dish brown paper and 5, rubber band after culture medium to prick a bundle, lain against in vertical high-pressure sterilizing pot,
121 DEG C sterilize 45 minutes, during must not tilt culture dish.It is placed in after cooling stand-by in superclean bench.
In ganoderma lucidum original strain described in picking in superclean bench in culture dish.Specifically, cut length and width with inoculation hook
For 5mm original strain block, culture dish center is placed in, after being inoculated with 15~20 culture dishes, with sealed membrane good seal, carries out mark
Note, 23 DEG C of dark culturing 7d.Culture dish is inverted after mycelium germination, and sealed membrane is pricked into hole, each culture with aseptic inoculation pin
Ware pricks 20 holes, continues to cultivate.
After colony growth to culture dish edge 1.5cm or so, the selection speed of growth is 5.5~6.5mm/d, colony edge
Neatly, the mycelia of the pure white and strong bacterial strain of mycelia is as ganoderma lucidum one-level activated spawn.
The fungus block of 1.5~2.5mm of inoculation first class inoculum colony edge mycelia is cultivated in two level activation medium.
The specific preparation method of the ganoderma lucidum two level activated spawn comprises the following steps：
The ganoderma lucidum two level activation medium includes following components：Glucose：20g, peptone：3g, yeast extract：1g, phosphorus
Acid dihydride potassium：2g, epsom salt：1g, vitamin B compound：0.5g.
Above-mentioned each group material is dissolved with 75 DEG C of water, and is settled to 1000mL.
It is sub-packed in the straight mouth triangular flasks of multiple 500mL, each bottled 250mL of triangle, one is placed in each triangular flask
2.5cm magnetic stir bar, and covered tightly with self-control cotton plug, triangle bottleneck separately is wrapped up with brown paper, bottleneck is tightened with rope.
The self-control cotton plug uses the longer non-absorbent cotton of fiber, and specific production method is：Cut and triangular flask bottleneck
The cotton block of size matching, cotton block is wrapped with left hand tiger's jaw, hand thumb compresses side, and opposite side ensures that smooth is in U-shaped,
Clip wraps cotton mass with the medical gauze piece that cotton mass size matches, and compresses, and in cotton mass U-shaped opening rope system
Tight feeder cloth is indeformable.
The triangular flask dispensed is placed in vertical high-pressure sterilizing pot, and 121 DEG C sterilize 45 minutes.Ultra-clean work is placed in after cooling
It is stand-by in platform.
The ganoderma lucidum one-level activated spawn is placed in superclean bench, selection one-level activated spawn colony edge 1.5~
2.5mm mycelia block is inoculated with, and the inoculation quantity of every bottle of triangular flask is 6 ferfas silk blocks.Using diameter 6.0mm card punch system
Standby mycelia block.The strain block is to float on media surface.
Triangular flask is placed in the lower 24 DEG C of constant incubators of dark condition and stands 30h after the completion of inoculation, is shifted after mycelium germination
To constant-temperature table, rotating speed 150r/min, 24 DEG C of dark culturing 7d, choose bacterium ball size, be evenly distributed, do not sink, sphere length
Full fine hair, culture medium color after sterilizing compared with substantially shoaling and transparent strain is as ganoderma lucidum two level activated spawn.
Second class inoculum is seeded in fermentation tank, inoculum concentration 0.1%, 24 DEG C incubated, and throughput is 0.6~1V/
Vmin, air pressure are 0.03~0.06Mpa, and biomass is chosen as 2.00 × 10 after cultivating 7d-2G/mL, bacterium bulb diameter are
1.00mm, Peloton density are 122/mL, and pH is 6.2 liquid fermentation strain.
The specific preparation method of the Ganoderma lucidum submerged fermentation strain comprises the following steps：
The Ganoderma lucidum submerged fermentation culture medium includes the component of following weight/mass percentage composition：Glucose 1%, bean cake powder
0.2%, corn flour 0.2%, wheat flour 0.2%, yeast extract 0.1%, magnesium sulfate 0.004%, potassium dihydrogen phosphate 0.08% is compound
Vitamin B 0.001% and defoamer 0.03%, surplus are water.
Installation for fermenting of the present invention is fermentation tank, and material is SUS304 stainless steels, volume 800L, described fermentation tank
Include interlayer, the interlayer is act as by recirculated water thermostatic control fermentation temperature.
The total amount of liquid spawn, adds the water of equivalent in fermentation tank according to needed for calculating inoculum concentration and inoculation bacterium bag number,
The steam that tank body interlayer is passed through 0.12Mpa is heated up, when staying in water temperature and rising to 75 DEG C, by the raw material heat in addition to defoamer
Water dissolving stirring, waits water temperature in fermentation tank to rise to 90 DEG C, adds raw material and defoamer.When water temperature rises to 121 DEG C, interlayer is closed
Steam cock, open steam cock in tank and steam is passed directly into tank body, and filter sterilised line switching is opened, enter
Sterilizing program.121 DEG C of sterilizing 45min.After the completion of sterilizing, carried out using the cold water of cold and hot water circulation system by interlayer at cooling
Reason, is inoculated with after culture medium is cooled to 24 DEG C.
Placed before the ganoderma lucidum two level activated spawn is inoculated with and 12h is stirred on magnetic stirring apparatus so as to smash bacterium ball, it is described
It is that multiple spot is sprouted to smash purpose, shortens the time that Ganoderma lucidum mycelium is sprouted in fermentation tank, while prevents from blocking inoculation pipeline.
Fermentation tank intake valve is closed during inoculation pressure inside the tank is down to blast pipe is micro- a gas discharge, close air bleeding valve.Will
Flame snare to inoculation mouth is lighted, and after opening inoculation mouth, makes inoculation mouth is micro- to have gas discharge by air inlet switch is crack.By triangular flask
Bottleneck is opened under the protection of flame ring, is poured into tank.Wet towel knock down the flame circle is used after tightening inoculation lid.Open into outlet
Valve, regulation pressure inside the tank to 0.05Mpa, throughput are that 0.8V/Vmin enters cultivation stage.Increase by one before the air bleeding valve
Brass Vertical Lift Check Valves prevent air suck-back.
During the Ganoderma lucidum submerged fermentation, per 2h observation fermentation pressure tank, temperature, smell, throughput, whether there is discoloration it is muddy
The data such as turbid simultaneously record.4d carries out sampling detection twice before being inoculated with fermentation completion, the purpose of detection is whether to judge bacterium solution
Contaminated, detection method uses detection method well-known to those skilled in the art, is：Gram's staining sediments microscope inspection whether there is
Bacterium infection, plate streaking detect the presence of bacterium and other fungal contaminations.
Selection biomass is (2.00 ± 0.50) × 10-2G/mL, bacterium mean diameter of a ball are 1.00 ± 0.10mm, Peloton density
For 120.00 ± 2.00/mL, the single batch fermentation product that pH value is 5~6.5 is as Liquid Strain of Ganoderma Lucidum.
The assay method of the strain bio amount is：Part bacterium solution is taken, 5min, repeated washing 3 are centrifuged through 5000r/min
It is secondary, it is freeze-dried under the conditions of being 15Pa in -45 DEG C, vacuum, hypha biomass is determined after 48h.The measure of the bacterium bulb diameter
Method is：By microscope ocular micrometer, 10 bacterium bulb diameters of random measurement in same sample, its arithmetic mean of instantaneous value is taken to make
For the average diameter of bacterium ball.The measuring method of the Peloton density is：1mL zymotic fluids are accurately drawn with pipette in culture dish
Counted, calculate the number of bacterium ball in every milliliter.
Glossy ganoderma fermentation strain is inoculated in solid fermentation culture medium by aseptic inoculation pipeline, culture room is transferred to and carries out
Light culture, mycelia obtain integrating tunning after covering with bacterium bag.
10 × 20mm of aseptic inoculation pipeline internal-and external diameter specification high pressure resistant silica gel tube, inoculation end connect stainless steel inoculation
Rifle.The inoculating gun length 17.5cm, it is uniformly distributed four rows, eight fluid holes.The inoculating gun connection 20cm stainless steels tubulose folder
Set, the other end set a ball valve switch, connection draining, discharge duct.The fermenter base discharging opening connects a four-way,
It is divided into (1) tank body discharging opening, (2) sample tap, (3) inoculation mouth of pipe, (4) water inlet pipe mouth or the steam mouth of pipe.
The inoculation pipeline sterilization method is as follows：
After the completion of Liquid Strain of Ganoderma Lucidum fermentation, inoculation pipeline is connected to the mouth of pipe, and the mouth of pipe connects water inlet pipe, opens
Water pipe and inoculation pipeline valve, inoculation pipeline, washing time 20min, after the completion of flushing, by the mouth of pipe are rinsed with running water
Water pipe is replaced with jet chimney connection.
Steam valve is opened, high steam is passed through in inoculated tube, the other end opens the ball valve switch of inoculating gun chuck, will
Steam is discharged.Sterilising temp is that 121 DEG C, sterilization pressure 0.12Mpa, sterilization time 45min sterilizings are completed, and treats tube-cooled extremely
Less than 25 DEG C are inoculated with.
The purpose of the flushing is to rinse out remaining bacterium ball, the nutriment in pipeline and inoculating gun, prevent miscellaneous
The growth of bacterium.
The preparation of the ganoderma lucidum solid fermentation culture matrix：
The ganoderma lucidum solid fermentation culture matrix includes the component of following weight/mass percentage composition：Rice 10%, wheat 15%,
Corn flour 40%, corncob 8%, dregs of beans 5%, wheat bran 20%, calcium hydroxide 1% and precipitated calcium carbonate 1%.The culture matrix
Carbon-nitrogen ratio is 25：1.
The setting of carbon-nitrogen ratio can ensure the growth of Ganoderma lucidum mycelium in the solid fermentation culture medium, and can guarantee that in raw material
Mycelium growth vigor is normal on the premise of cost is minimum.
Before use, all raw materials need to pass through pretreatment, including crush, soak.The particle diameter of the corn flour be 0.6~
1.0mm, the particle diameter of the dregs of beans is 0.8~1.2mm, and the particle diameter of the rice is 2.0~2.5mm, and the particle diameter of the wheat is
2.4~2.8mm.The wheat soaks 36h, 25 DEG C of rice, corncob water immersion 12h before use with 25 DEG C of water.
The corn flour, dregs of beans, the particle diameter of rice and wheat can ensure bacterium bag unitary physical structure, and balance bacterium bag is breathed freely
Property with sterilizing completeness.In order to adjust the gas permeability of bacterium bag, the granularity of setting is larger, leads for the wheat, rice, corncob
Cause water absorption big and not hygroscopic, immersion purpose is to prevent the dry core of the raw material so as to cause sterilizing not thorough.
The culture matrix each component is poured into 10.4m3Agitated kettle in, be 1000kg/ pots, be stirring evenly and then adding into
Water, ensure that water content is 48% in matrix before packing, pH value 7.5.It is sent to after stirring in automatic packing machine, ensures bacterium
Bag height 17cm, per packed 2.4 jin of culture matrixes.Punching press twice is carried out in the automatic packing machine packing process, second of punching
It is pressed in the punching of bacterium bag matrix center, depth 15.5cm.
The setting of the water content can ensure the completeness of sterilizing, the requirement of the elasticity and mycelia of bacterium bag to moisture.
Carried out in the solid fermentation high temperature resistant bacterium bag bag, the bacterium bag bag uses PP well known to those skilled in the art
(polypropylene) bacterium bag bag, specification are 36cm × 15cm × 0.5mm.
After bacterium bag completes, it is positioned in supporting high-temperature resistance plastice basket, 12 bags every basket, plastic crate is positioned over vehicle of sterilization
Interior, push-in high pressure steam sterilization cabinet is sterilized.
The sterilization process vacuumize 2 times, heating and heat preservation 1 set 105 DEG C of 30min, heating and heat preservation 2 set 115 DEG C
30min, sterilizing set 123 DEG C of 200min, stewing put to be vented setting 30min.After the completion of sterilizing, bacterium bag is cooled to less than 25 DEG C progress
Inoculation.
The vaccination ways are specific as follows：
The bacterium bag that cooling is completed is sent in transfer room of the cleanliness factor less than 100,000 grades and is inoculated with." one is followed during inoculation
Hand takes rifle, takes lid on the other hand " principle.Being inoculated with operating process is：1. uncap；2. inoculating gun insertion bacterium bag material hole, gun breech disk block
Untill the collar；3. pin steps on floor push, spray strain；4. stopping makes strain spray for 1~2 second to the greatest extent, while opens next bacterium
Clad；5. inoculating gun closes the lid immediately after extracting bacterium bag out, according still further to 2. 3. 4. 5. flow is inoculated with successively.Finished bacterium bag in
Transfer room is sent out on conveyer belt.
The inoculum concentration of the Liquid Strain of Ganoderma Lucidum is 60mL/kg.The inoculating gun length is 17.5cm, averagely sets four rows
Eight fluid holes, the control device prset counter and magnetic valve of the inoculum concentration.
The Ganoderma lucidum mycelium culture concrete mode is as follows：
After the completion of inoculation, ganoderma lucidum bacterium bag is transferred to culture room and carries out dark culturing.The humidity of the light culture be 60~
65%；The light culture includes following 4 stages：
First stage is 1~9d, belongs to the mycelium germination phase, and mycelia self-heating amount is low, needs equipment to provide cultivation temperature
To 25~26 DEG C, because mycelia respiratory capacity is smaller, CO2Concentration is 1000~1500ppm；
Second stage is 10~15d, belongs to mycelia and is colonized a phase, and mycelia self-heating amount is maximum, and oxygen consumption starts
Increase, cultivation temperature need to be down to 23~24 DEG C, CO2Concentration is controlled in 1500~2000ppm；
Phase III is 16~30d, belongs to the mycelia field planting second phase, mycelia self-heating amount is larger, CO2Accumulated concentrations are most
Height, cultivation temperature are arranged on 24~25 DEG C, CO2Concentration is 2000~3000ppm；
The fourth stage, which is 30d, to be completed to cultivating, and belongs to mycelia latter stage of ripening, mycelia caloric value and the first stage
Remain basically stable, cultivation temperature is arranged to 25~26 DEG C, CO2Accumulated concentrations are more than the first stage, are 1500~2000ppm.
The present invention carries out fermented and cultured, solid fermentation with liquid fermentation combination technology using solid fermentation to ganoderma lucidum mycelium
Difference, the present invention increase liquid to water content of substrate using solid medium moisture is reduced with traditional handicraft with inoculum concentration
The mode of body strain inoculum concentration.The solid medium water content standard is accounting, the granularity size according to formula each component
And water absorbing properties formulation, on the premise of sterilizing completeness principle is ensured, the culture matrix is more dry, and liquid spawn is oozed
Permeability is better, and the principle standard of a variety of amounts of liquid bacteria is that strain amount is got on the premise of bottom is particularly in culture matrix without ponding
Greatly, strain radix is more, and mycelial growth is faster.And bacterium bag part ponding instead results in mycelia local growth slowly or is difficult to give birth to
It is long, while a variety of amounts of liquid bacteria are bigger, the later stage fungi polysaccharide content of extraction is higher.
By contrast, solid fermentation raw materials formula described in this method and each component ratio, granular size it is specific under the premise of,
Solid fermentation matrix water content is 48%, inoculum concentration 60ml/kg, and mycelium growth vigor is most fast.
Originally the present invention makes ganoderma lucidum mycelium using liquid fermentation mode, single fermentation amount is bigger, and extraction cost is lower,
But on the premise of fermentation tank blade diameter length ratio is equivalent, Zymolysis Equipment is bigger, Zymolysis Equipment technique requires higher with corollary equipment,
Operating procedure requires more harsh with zymotechnique, expands numerous number increase, fermenting, total cycle is longer, and risk factor is bigger, required
Early investment is also higher, and extraction process and equipment requirement are also higher.The application method uses 800L fermentation tanks to make
Liquid fermentation strain, is not directly used in extraction, but expands in numerous matrix to solid fermentation, there is that early investment is low, pollution rate
Low, the ability to ward off risks is strong, the advantages of production technology relative ease.
Embodiment 2
With the working process of glossy ganoderma fermentation product：
The Ganoderma lucidum mycelium tunning of maturation is obtained using embodiment 1, is cut into slices, drying and processing, obtains sheet fungus block；
After sheet fungus block pulverization process, ganoderma lucidum powder is obtained；With supercritical CO2Fluid extraction bacterium powder obtains ganoderma volatile oil, ganoderma lucidum
Polysaccharide and ganoderic acid.
The section drying of ganoderma lucidum maturation hypha fermentation product, specifically includes following steps：
The ganoderma lucidum bacterium bag that solid fermentation culture is completed carries out manually taking lid, takes ring, de- bag processing, takes off the tunning after bag
Mechanical section is carried out, the single-sheet thickness of the section is 1~1.5cm.
The section machinery is self-control slicer, and inspiration comes from paper mill cutting machine, set one can 90 ° of closures consolidate
Determine plug-in strip handle, the subordinate stainless steel blade for welding 13 maximum gauge 3mm, the blade spacing is 1.25cm.Under blade
Square table face sets a 17cm length, the stainless steel cylinder fixed card slot of 5cm diameters, and the neck sets empty bar in 13 transverse directions
Line.Empty fringe spacing 3.35mm, can prevent bacteria residue from residuing in gap blade, lower half when plug-in strip blade above carries in neck upper semi-circle
Circle neck fringe spacing 8mm, sheet fungus block can pass through neck after the completion of section.One table that can place drip pan is set below neck
Face, desktop are directly dropped down onto in drip pan from neck height 20cm, the fungus block by neck.Neck one end connection is one long
50cm, diameter identical arc pan feeding cylinder, cylinder ends are in 15 ° with desktop, are easy to put into bacterium bag.The feeding cylinder, neck,
Blade is all retractable or dismantles, the cleaning being easy to after the completion of section.
The drip pan drip pan with holes improves drying efficiency, aperture 3mm, 25/dm of drip pan hole density2.The drip pan 80 ×
50 × 2cm specifications, thickness 1mm stainless steel, each drip pan put 3 ganoderma lucidum bacterium bags, put 3 × 14 altogether.
Described dry is carried out in drying room, drying room volume 360m3, the drying room is front and rear up and down respectively to set 4 temp probes
With moisture probe.The drying room rear end sets power 0.55kw, 3000~4000m of air quantity3/ h centrifugal blower, utilized to improve
Rate, the centrifugal blower set circulation and the dual-purpose passage of outer row, toast the normally opened outer multiple rows of moisture of row's passage early stage, and the later stage opens more and followed
Ring passage opens air exhaust passage less, can shorten baking time, save the energy.The drying room jet chimney space heating, the steam
Pipe drainage drain tap is placed in outside drying room, diameter 25mm iron pipes, is placed in baking rack lower end with coil form, the baking is set up
8 layers, individual layer height 30cm are put, width 1.6m, every layer sets 4 coil pipes to heat.Another every drying room front end 2 power 2.2kw, wind
Measure 9000m3/ h auxiliary heat the dehumidifying of axial flow blower air inducing.
The drying course is divided into 3 steps：Heating, wet down and insulation.After drip pan piles drying room, coil pipe steam valve is opened
Door carry out drying room heating, suitably turns coil pipe quantity of steam down after temperature rises to 70 DEG C, opens fin steam valve, and open axle
Flow fan and centrifuge, air inducing hydrofuge.When space average humidity is less than 5%, into the holding stage, centrifugal blower is set
Circulated air starts 45min per hour, and outer air draft starts 15min, toasts and terminate after insulation 6h.When baking temperature is less than 65 for a long time
DEG C, hot and humid high nutrition environment meeting mould growth, baking temperature can destroy polysaccharide material structure higher than 80 DEG C.During heating, protect
It is sufficient to demonstrate,prove steam pressure, the heating-up time is no more than 3 hours；During baking, strict control enters drying room quantity of steam, and baking temperature is 70
Between~75 DEG C.
This section baking method can ensure：Single toasts the total dosage of steam in 1.25~1.5 tons/time, single drying finished product
Measure as 1.8 tons, baking time is less than 22h, and fungus block moisture is not higher than 7% after baking.
Being dispensed after the completion of baking, be weighed into 50 jin/bag, sewing machine seals, the packaging bags with film inner bag,
It is stand-by that storage at shady and cool, aeration-drying is placed in after packaging.
The mycelium mixture of the comprehensive fermentation process culture, after measured, ganoderma volatile oil recovery rate is up to 0.03%,
Ganoderma Lucidum acid extraction rate reached 3.2%, containing sour solid product acid purity up to more than 25%, GL-B recovery rate according to kind not
With up to 10.1%, purity of polysaccharide is up to more than 45%.
Described above is only the preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, under the premise without departing from the principles of the invention, some improvements and modifications can also be made, these improvements and modifications also should
It is considered as protection scope of the present invention.
Claims (9)
1. a kind of synthesis fermentation process of ganoderma lucidum mycelium, comprises the following steps：
1) lucidum strain after activation is inoculated in liquid fermentation medium, 23~26 DEG C of 5~7d of culture, throughput 0.6
~1V/Vmin, air pressure are 0.03~0.06Mpa, obtain fermented bacterium；
The liquid fermentation medium includes the component of following weight/mass percentage composition：Glucose 0.5~1%, dregs of beans 0.1~
0.2%, corn flour 0.1~0.2%, wheat flour 0.1~0.2%, yeast extract 0.1~0.3%, magnesium sulfate 0.003~
0.005%, potassium dihydrogen phosphate 0.08~0.1%, vitamin B compound 0.001% and defoamer 0.03~0.05%, surplus are
Water；
2) fermented bacterium for obtaining step 1), which is inoculated in solid fermentation culture medium, carries out light culture, the time of the light culture
For 30~40d.
2. fermentation process according to claim 1, it is characterised in that the lucidum strain after being activated in the step 1) is in liquid
Inoculum concentration in body fermentation medium is 0.1%~0.2%.
3. fermentation process according to claim 1, it is characterised in that the biomass for the fermented bacterium that the step 1) obtains
For (2.00 ± 0.50) × 10-2G/mL, bacterium mean diameter of a ball are 1.00 ± 0.10mm, Peloton density is 120.00 ± 2.00/
ML, pH value are 5~6.5.
4. fermentation process according to claim 1, it is characterised in that the solid fermentation culture medium of the step 2) is bacterium bag
Bag solid fermentation culture medium.
5. the fermentation process according to claim 1 or 4, it is characterised in that the inoculum concentration of fermented bacterium in the step 2)
For 50~100mL/kg.
6. fermentation process according to claim 1, it is characterised in that the carbon-nitrogen ratio in the solid fermentation culture medium is
(20~28)：1, water content is 42~50%, and the pH value of culture medium is 7~8 before sterilizing.
7. fermentation process according to claim 1, it is characterised in that the solid fermentation culture medium includes following quality hundred
Divide the component of content：Rice 5~10%, wheat 10~15%, corn flour 35%~45%, corncob 5~8%, dregs of beans 5~
10%, wheat bran 15~25%, calcium hydroxide 1~2% and precipitated calcium carbonate 1~2%.
8. fermentation process according to claim 1, it is characterised in that the humidity of the step 2) light culture be 60~
65%；The light culture includes following 4 stages：
1~9d, the temperature of the light culture is 25~26 DEG C, CO2Concentration is 1000~1500ppm；
10~15d, the temperature of the light culture is 23~24 DEG C, CO2Concentration is 1500~2000ppm；
16~30d, the temperature of the light culture is 24~25 DEG C, CO2Concentration is 2000~3000ppm；
30d is to completion is cultivated, and the temperature of the light culture is 25~26 DEG C, CO2Concentration is 1500~2000ppm.
9. fermentation process according to claim 1, it is characterised in that also include de- bag after the step 2) light culture, cut
Piece and baking process.
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