CN108410917A - The culture medium and its preparation of a kind of Bacillus acidi lactici production γ-aminobutyric acid and application - Google Patents
The culture medium and its preparation of a kind of Bacillus acidi lactici production γ-aminobutyric acid and application Download PDFInfo
- Publication number
- CN108410917A CN108410917A CN201810472692.8A CN201810472692A CN108410917A CN 108410917 A CN108410917 A CN 108410917A CN 201810472692 A CN201810472692 A CN 201810472692A CN 108410917 A CN108410917 A CN 108410917A
- Authority
- CN
- China
- Prior art keywords
- aminobutyric acid
- preparation
- culture
- medium
- yeast extract
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P13/00—Preparation of nitrogen-containing organic compounds
- C12P13/005—Amino acids other than alpha- or beta amino acids, e.g. gamma amino acids
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
Abstract
Include following components according to mass fraction the invention discloses a kind of culture medium of Bacillus acidi lactici production gamma aminobutyric acid and its preparation and application, the culture medium：Glucose 2%, yeast extract and rice bran mixture 3%, L glutamic acid 1.5%, Tween 80 0.1%, epsom salt 0.02%, four water manganese sulfates 0.005%, surplus is distilled water.Microorganism formulation containing gamma aminobutyric acid is made by tablet culture, shake-flask seed liquid, domestication culture and fermented and cultured are prepared, said preparation is applied to have to improve in cultural technique to be immunized, the effect of intestinal flora is adjusted, while there is the calm measuring body effect of gamma aminobutyric acid.
Description
Technical field
The invention belongs to microorganisms technical fields, and in particular to a kind of culture medium of Bacillus acidi lactici production γ-aminobutyric acid and
Its preparation and application.
Background technology
In animal husbandry, it is raw to show excited, neurotic, loss of appetite etc. for various reasons for the animal of different times
Reason state, for animal and group, especially rigid birth and the animal in the pregnancy period, are all unfavorable for this.It is anti-at present to answer
Sharp reaction mostly uses the drugs such as intramuscular injection antondin, dexamethasone sodium phosphate, antibiotic, there is and residual toxic to animal
Side effect.γ-aminobutyric acid is the naturally occurring nonprotein amino acid for having greater activity, is passed as a kind of nerve
Matter can play the calm, biological functions such as calm the nerves, and play and alleviate that animal is excited, unstrung physiological status, and to animal sheet
Body does not have apparent side effect.
γ-aminobutyric acid is widely distributed in animal and plant body, but since there are plants with reference state for γ-aminobutyric acid
It is interior, the difficulty of extraction is increased, concentration method is difficult with and obtains high concentration γ-aminobutyric acid.γ-aminobutyric acid is obtained at present
Method is mainly chemical synthesis and biological synthesis process, and a variety of toxic, corrosive chemicals are added to during chemical synthesis,
Severe reaction conditions, hardly possible extraction food-grade γ-aminobutyric acid.And chemical reagent is not added in biosynthesis substantially, and it is pollution-free, only need
Ensure that microbial metabolic products are within safe range, this will be as the following general orientation for preparing γ-aminobutyric acid.Micro- life
Object fermentation method is mainly the synthesis that γ-aminobutyric acid is catalyzed using the enzyme system of bacterial strain itself.Wherein glutamate decarboxylase is
Regulate and control the Major Enzymes of γ-aminobutyric acid synthesis.But generally existing glutamic acid decarboxylase enzyme activity is not high at present and strain is potential not
Safety issue, therefore the safety strain for finding high vigor glutamate decarboxylase produces γ-aminobutyric acid, it is still necessary to microorganism
The continuous effort of researcher.
Lactic acid bacteria has more OH free radical scavenging abilities, is generally recognized as safe microorganism in the world, be people and
Indispensable important physiology flora, is closely related with the vital movement of humans and animals, is widely used in food for a long time in animal body
Product are processed.Since some lactic acid bacterias can also generate γ-aminobutyric acid, thus had become using lactic acid bacteria production γ-aminobutyric acid
One safety and ideal approach.But conventional production γ-aminobutyric acid additive does not retain lactic acid bacteria living, the present invention goes out
Hair point is a kind of microorganism formulation of research, and the microorganism formulation includes the lactic acid bacteria of production γ-aminobutyric acid, makes it have lactic acid
Bacterium growth promotion is improved and is immunized, and has γ-aminobutyric acid calm measuring body effect while adjusting intestinal flora.
Invention content
For technological deficiency of the existing technology, present invention aims at provide a kind of Bacillus acidi lactici production gamma-amino fourth
The culture medium and its preparation of acid and application.
The present invention is realized especially by following technical scheme：
A kind of culture medium of Bacillus acidi lactici production γ-aminobutyric acid, includes following components according to mass fraction：Glucose 2%,
Yeast extract and rice bran mixture 3%, Pidolidone 1.5%, Tween-80 0.1%, epsom salt 0.02%, four water sulfuric acid
Manganese 0.005%, surplus are distilled water.
Preferably, a kind of culture medium of Bacillus acidi lactici production γ-aminobutyric acid, includes following components according to mass fraction：Portugal
Grape sugar 2%, yeast extract and rice bran mixture 3%, Pidolidone 1.5%, Tween-80 0.1%, epsom salt 0.02%,
Four water manganese sulfates 0.005%, surplus are distilled water.
The yeast extract is with rice bran according to mass ratio 1:1 mixing.
Bacillus acidi lactici of the present invention is Lactobacillus brevis (Lactobacillus brevis) P-14 bacterial strains, deposit number
For CGMCC NO.9029, it is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on April 9th, 2014,
Preservation address：Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Institute of Microorganism, Academia Sinica.
The present invention also provides a kind of preparation methods of the microorganism formulation containing γ-aminobutyric acid, include the following steps：
1) tablet culture：Lactobacillus brevis P-14 is inoculated on solid activation medium tablet using streak inoculation method, in
30 DEG C~37 DEG C cultures 36~48h, recovery Lactobacillus brevis P-14 form single bacterium colony；
2) shake-flask seed liquid makes：With oese by the free of contamination Lactobacillus brevis P-14 single bacteriums that grow fine in step (1)
It falls and is inoculated into liquid activation medium, 30 DEG C~37 DEG C 36~48h of static gas wave refrigerator obtain shake-flask seed liquid；
3) domestication culture：The shake-flask seed liquid that access step (2) in culture medium is tamed in liquid carries out domestication culture, is inoculated with
Amount is 2~5%, and condition of culture is 30 DEG C~37 DEG C, static gas wave refrigerator, and 36~48h passed on for two generations；
4) fermented and cultured：Fermentation medium is added in fermentation tank (1 ton), takes the Lactobacillus brevis after domestication in step (3)
P-14 bacterium solutions are inoculated with, inoculum concentration 0.1%, and ferment 72~96h at 30 DEG C, and microorganism formulation is obtained after fermentation.
The solid activation medium is prepared according to the following recipe：Peptone 10g, beef extract 10g, yeast extract
5g, dipotassium hydrogen phosphate 2g, Triammonium citrate 2g, sodium acetate 5g, glucose 20g, Tween 80 1mL, epsom salt 0.5g, four
Water manganese sulfate 0.25g, agar 20g, distilled water 1L, 121 DEG C of sterilizing 20min, cooling are spare.
The liquid activation medium is prepared according to the following recipe：Peptone 10g, beef extract 10g, yeast extract
5g, dipotassium hydrogen phosphate 2g, Triammonium citrate 2g, sodium acetate 5g, glucose 20g, Tween 80 1mL, epsom salt 0.5g, four
Water manganese sulfate 0.25g, distilled water 1L, 121 DEG C of sterilizing 20min, cooling are spare.
The domestication culture medium is prepared according to the following recipe：Concentration of glucose 20g, yeast extract and the dense mixture of rice bran
30g, Pidolidone concentration 15g, Tween-80 1.0mL, epsom salt 0.2g, four water manganese sulfate 0.05g, distilled water 1L, 121
DEG C sterilizing 20min, cooling it is spare.
The fermentation medium is prepared according to the following recipe：Glucose 15Kg, yeast extract and rice bran mixture
22.5Kg, Pidolidone 11.25Kg, Tween-80 750mL, epsom salt 150g, four water manganese sulfate 37.5g, distilled water
750Kg, 75 DEG C of 2h that sterilize, it is spare after being quickly cooled to 35 DEG C.
Effective total viable count >=1.5 in the microorganism formulation containing γ-aminobutyric acid obtained according to above-mentioned preparation method ×
109CFU/mL, alpha-aminobutyric acid content 335mg/100mL.
The microorganism formulation containing γ-aminobutyric acid that above-mentioned preparation method obtains is also within protection scope of the present invention.
In addition, the application the present invention also provides the microorganism formulation containing γ-aminobutyric acid in aquaculture.
The application process of mentioned microorganism preparation is specifically included is diluted the microorganism formulation in 0.1% ratio in drinking-water
It is drunk for cultivated animals；The microorganism formulation is pressed into 0.1% ferment complete feed, is compared by certain with the complete feed not fermented
Example mixed feeding.
The program of the ferment complete feed is：The microorganism formulation is inoculated in the complete of 30% water content by 0.1%
Valence feed is sealed and is placed 3-7 days under room temperature, and viable count reaches 5.33 × 108CFU/g, GABA content 0.015%.
Beneficial effects of the present invention are：
Rice bran contains the important physiological activator such as glutamate decarboxylase, oryzanol, lipopolysaccharides.Therefore using in rice bran meal
Glutamic acid and glutamate decarboxylase, by lactic acid bacteria carry out biofermentation can using bacterial strain glutamate decarboxylase decarboxylation
Rice bran meal Glutamic Acid is converted into high activity γ-aminobutyric acid, the feed that can improve animal anti-stress ability is prepared
Additive makes rice bran meal obtain increment and utilizes, and for rationally utilizing grain processing by-product resource, ensures animal products health peace
Entirely, grain processing enterprise synergy is promoted to have important meaning.The present invention is with paddy by-product-rice bran rich in GAD and glutamic acid
Partial medium nitrogen source is substituted, rice bran added value is not only increased, also reduces fermentation medium cost.That prepares is rich in
The microorganism formulation of γ-aminobutyric acid and lactic acid bacteria viable is used to add in cultivated animals drinking-water or ferment complete feed,
Production performance can not only be improved, meat is improved by optimizing meat nutrition component and aliphatic acid composition, height can also be reduced
Temperature stress be to negative effect that cultivated animals are brought.
Specific implementation mode
Below in conjunction with specific embodiment of the present invention, technical solution of the present invention is clearly and completely described, is shown
So, described embodiments are only a part of the embodiments of the present invention, instead of all the embodiments.Based on the reality in the present invention
Example is applied, every other embodiment obtained by those of ordinary skill in the art without making creative efforts all belongs to
In the scope of protection of the invention.
The culture medium prescription of 1 Bacillus acidi lactici highly producing gamma-aminobutyric acid of embodiment optimizes
Using paddy by-product-rice bran as nitrogen source, rice bran contains abundant GAD and glutamic acid for this research, not only improves fermentation
γ-aminobutyric acid yield in liquid, and the zymotic fluid that the reaction was complete can also be added as feed as high protein substance, reduce
Pollution to environment, reduces fermentation medium cost, improves rice bran added value.
About optimal components and suitable concentration in culture medium is determined, experiment is first with single factor experiment determination to γ-
The notable factor of aminobutyric acid yield effect.In order to make full use of the value of byproduct rice bran, experiment to choose rice bran and replace part egg
White peptone is nitrogen source.Before experiment, rice bran was smashed into 80 mesh sieve with pulverizer, microorganism can be promoted preferably to utilize rice in this way
The nutriment of chaff.According to single factor experiment as a result, being carried out to glucose, yeast extract and rice bran, Pidolidone principal element
Orthogonal test designs Three factors-levels table, optimal medium constituent concentration is determined according to orthogonal result.
Obtaining culture medium optimum combination by orthogonal experiment is：Concentration of glucose 20g/L, yeast extract+rice bran concentration (1：
1) 30g/L, Pidolidone concentration 15g/L, Tween-80 1.0mL, epsom salt 0.2g/L, four water manganese sulfate 0.05g/L, hair
Bacteria concentration is up to 1.910 in zymotic fluid, and γ-aminobutyric acid production quantity is up to 355mg/100ml.
Medium optimization initial incubation based component：Glucose 20g, peptone 10g, beef extract 10g, yeast extract 5.0g are spat
Temperature -801.0mL, epsom salt 0.2g, four water manganese sulfate 0.05g, distilled water 1L.
The determination program of above-mentioned nitrogen source is：It is respectively 1 by unique sole nitrogen source rice bran and ratio：1 yeast extract and rice bran
Compound nitrogen source is added in initial medium, adds a concentration of 10g/L, 20g/L, 30g/L respectively, investigate its to strain growth and
Target product influences, and the results are shown in Table 1.
The influence of 1 nitrogen source type of table and concentration to strain growth and γ-aminobutyric acid yield
The determination program of above-mentioned carbon source is：Respectively selection 20g/L glucose, soluble starch, maltose, fructose, sucrose,
Lactose is added in initial medium, provides strain growth and needs sole carbon source.Different carbon source is had studied to strain growth and γ-
The influence of aminobutyric acid yield, the results are shown in Table 2.On the basis of most suitable carbon source is determined, 5g/L, 10g/L, 15g/L are added respectively,
20g/L, 25g/L, 30g/L determine most suitable carbon source concentration, the results are shown in Table 3 to initial medium.
Influence of 2 carbon source kind of table to strain growth and γ-aminobutyric acid yield
Influence of 3 concentration of glucose of table to strain growth and γ-aminobutyric acid yield
The determination program of the above-mentioned other ingredients of culture medium is：Add a concentration of 0g/L, 5g/L, 10g/ respectively in the medium
L, the Pidolidone of 15g/L, 20g/L, 25g/L, 30g/L investigates substrate Pidolidone concentration to strain growth and gamma-amino fourth
Acid yield influences, and the results are shown in Table 4；Add a concentration of 0ml/l, 0.5ml/l, 1ml/l, 1.5ml/l, 2ml/ respectively in the medium
The Tween-80 of l investigates Tween-80 concentration to strain growth and γ-aminobutyric acid yield effect, the results are shown in Table 5；In culture medium
The middle phosphopyridoxal pyridoxal phosphate for adding a concentration of 0g/L, 0.1g/L, 0.15g/L, 0.2g/L, 0.25g/L, 0.3g/L respectively investigates phosphorus
Sour pyridoxal concentration the results are shown in Table 6 to strain growth and γ-aminobutyric acid yield effect.
Influence of 4 concentration of substrate of table to strain growth and γ-aminobutyric acid yield
Influence of the 5 Tween-80 concentration of table to strain growth and γ-aminobutyric acid yield
Influence of the 6 phosphopyridoxal pyridoxal phosphate concentration of table to strain growth and γ-aminobutyric acid yield
Orthogonality analysis method Optimal Medium program is：It is formed using orthogonal analysis Optimal Medium, according to single factor test
Test result chooses glucose, yeast extract and rice bran, progress 3 factor, the 3 horizontal quadrature experiment of Pidolidone principal element, with
Bacteria concentration is index in zymotic fluid, the results are shown in Table 7.
7 orthogonal experiments of table and range analysis
10% or so byproduct rice brans are generated during China's paddy processing, rice bran annual output is about at 13,000,000 tons or so.
Also there are upper 1,000,000 tons of rice brans to generate every year in Sichuan.This is a very huge renewable resource.However China only has 10%
Left and right rice bran is fully and rationally used far away for oil expression and deep processing, resource.Rice bran contains glutamate decarboxylase, Gu Wei
The important physiological activator such as element, lipopolysaccharides.Therefore using the glutamic acid and glutamate decarboxylase in rice bran meal, pass through lactic acid bacteria
Rice bran meal Glutamic Acid can be converted into high activity γ-using bacterial strain glutamate decarboxylase decarboxylation by carrying out biofermentation
Aminobutyric acid prepares the feed addictive that can improve animal anti-stress ability, so that rice bran meal is obtained increment and utilizes, for closing
Reason utilizes grain processing by-product resource, ensures that animal products is healthy and safe, grain processing enterprise synergy is promoted to have important meaning
Justice.
2 microorganism formulation preparation method of embodiment
The preparation method of microorganism formulation is as follows：
1) tablet culture, using solid activation medium, tablet activates the Lactobacillus brevis
It is inoculated on solid activation medium tablet using streak inoculation method by γ-aminobutyric acid Lactobacillus brevis is produced, in 30-
37 DEG C of 36~48h of culture, make its recovery, form single bacterium colony.
Wherein solid activation medium is prepared according to the following recipe：Peptone 10g, beef extract 10g, yeast extract 5g,
Dipotassium hydrogen phosphate 2g, Triammonium citrate 2g, sodium acetate 5g, glucose 20g, Tween 80 1mL, epsom salt 0.5g, four water sulphur
Sour manganese .25g, agar 20g, distilled water 1L, 121 DEG C of sterilizing 20min, cooling are spare.
2) shake-flask seed makes, using shaking flask and liquid activation medium, the Bacillus acidi lactici in incubation step (1).
The free of contamination strain that grows fine in step (1) is inoculated into respectively in liquid activation medium with oese, with
36~48h of static gas wave refrigerator under the conditions of 30 DEG C~37 DEG C of 500mL triangular flasks obtains shake-flask seed liquid, and sterile working sampling carries out
Bacterial strain is identified, it is ensured that bacterial strain is correct.
Wherein liquid activation medium is prepared according to the following recipe：Peptone 10g, beef extract 10g, yeast extract 5g,
Dipotassium hydrogen phosphate 2g, Triammonium citrate 2g, sodium acetate 5g, glucose 20g, Tween 80 1mL, epsom salt 0.5g, four water sulphur
Sour manganese 0.25g, distilled water 1L, 121 DEG C of sterilizing 20min, cooling are spare.
3) domestication culture tames culture medium, the Bacillus acidi lactici in domestication incubation step (2) using liquid.
Liquid is added in the big triangular flasks of 1L and tames culture medium, the bacterium solution of access step (2) carries out domestication culture, inoculum concentration
It is 2~5%, condition of culture is 36 DEG C, static gas wave refrigerator, and 48h passed on for two generations.
Wherein domestication culture medium is prepared according to the following recipe：Glucose 20g, yeast extract+rice bran concentration (1：1)30g、L-
Glutamic acid 15g, Tween-80 1.0ml, epsom salt 0.2g, four water manganese sulfate 0.05g, distilled water 1L, 121 DEG C of sterilizings
20min, cooling are spare.
4) fermented and cultured, using stainless steel fermentation tank and fermentation medium, the galactopoiesis after being tamed in fermented and cultured step (3)
Acidfast bacilli prepares microorganism formulation.
Fermentation medium is added in 1 ton of stainless steel fermentation tank, the cultured liquid bacterial strain of domestication in step (3) is taken to carry out
Inoculation, inoculum concentration 0.1%, 30 DEG C of 72~96h of fermentation calculate viable count after 72h under the microscope, when effective total viable count >=
1.5×109When CFU/mL, that is, prepare microorganism formulation.
Fermentation medium is prepared according to the following recipe：Glucose 15Kg, yeast extract and rice bran mixture 22.5Kg, L- paddy
Propylhomoserin 11.25Kg, Tween-80 750mL, epsom salt 150g, four water manganese sulfate 37.5g, distilled water 750Kg, 75 DEG C of sterilizings
2h, it is spare after being quickly cooled to 35 DEG C.
The technique effect of technical solution of the present invention is further detailed with reference to specific Application Example.
Embodiment 3
60 male mices are randomly divided into 5 groups by weight requirement, every group 12, i.e. negative control；Positive controls；It presses
Microorganism formulation 0.05%, 0.1%, 0.2% 3 kind of concentration are divided into experiment 1-3 groups, number, sub-cage rearing.Raise room temperature 33
DEG C, relative humidity:50%-60%.
Every intragastric administration on mice 0.2mL physiological saline of negative control group；Every intragastric administration on mice 0.2ml 20mg/ of positive controls
L γ-aminobutyric acid solution；Low concentration group, every middle concentration group, high concentration group mouse distinguish gavage 0.05%, 0.1%, 0.2%
Microorganism formulation (bacteria containing amount be 1.5 × 109CFU/mL)0.2ml.Once a day, continuous gavage 30d, each gavage time one
It causes.
On-test and mouse feed to each group mouse empty stomach 8h when 30d, and the 2h that cuts off the water supply calculates mouse daily gain by only weighing
And feed-weight ratio, after gavage, observe the general state of animal, observation period 30d.Experiment overall process and observed content do detailed note
Record.
Influence of 8 microorganism formulation of table to mouse growth in conventional environment
Test result shows 3 test groups weightening more apparent than control group after 30d, average weight gain 14.63g/, than control
Group improves 21.4%.Two groups of control group daily gain gaps are little.The sample of various concentration is also different to mouse daily gain effect, sample
Product to the effect of mouse daily gain be successively the sample of 0.2% sample=0.1% >=0.05% sample >=saline control group=
γ-aminobutyric acid group, 0.1% or more concentration have remarkable effect.
The feed-weight ratio of test group is higher than control group, and the feed-weight ratio highest of experiment 2 and experiment 3 shows that sample has increase feed
The effect of utilization rate, 0.1% or more concentration have remarkable effect.
Embodiment 4
40 male mices are randomly divided into 4 groups by weight requirement, every group 10, are divided into blank control group, gavage 0.2ml
Physiological saline.The microorganism of low concentration group, every middle concentration group, high concentration group mouse difference gavage 0.05%, 0.1%, 0.2%
(bacteria containing amount is 1.5 × 10 to preparation9CFU/mL)0.2ml.Once a day, continuous gavage 14d, each gavage time consistency.Raising
33 DEG C of room temperature, humidity 50%-60%.
It is pre- raise test specimen 14d after, mouse is placed in 37 DEG C, in the environment of humidity 70%-80%, observation is moved in 30 minutes
The death toll and record of object.
Influence of 9 microorganism formulation of table to survival ability under mouse hot environment
Test result shows that the 0.1% and 0.2% concentration samples death rate is 50%, and the death rate is minimum, 0.05% death rate
70%, it is consistent with blank control group, illustrate that a certain concentration microorganism formulation can reduce the high temperature lethality of mouse, produces γ-ammonia
Base butyric acid lactic acid bacterial liquid has certain protective role to heat-stressed mice.
Embodiment 5
By 300 growing and fattening pigs, 3 processing groups are randomly divided into, 5 repetitions of each processing group are each to repeat 20.Control group
(100% basal diet of feeding)；30% fermentation group (+30% fermented feed of 70% basal diet of feeding)；(the feeding of 50% fermentation group
+ 50% fermented feed of 50% basal diet).
The preparation method of fermented feed is：The basal diet of 0.1% microorganism formulation fermentation 30% moisture of addition, sealing are normal
Temperature fermentation 3-7 days, viable count reaches 5.33 × 108CFU/g, GABA content 0.015%.It is freely eaten and drinks during experiment
Water manages feeding, drinking-water and the health condition of observation swinery daily according to pig farm conventinal breeding method, records feed consumption rate and morbidity
Death condition.Experimental period is 47 days.Record all stage feed consumption rate, on-test and and at the end of weigh on an empty stomach, calculate average
Daily ingestion amount, average daily gain and feed-weight ratio.Off-test each repeats the similar test pig of 1 weight of middle selection respectively
It is butchered, acquisition longissimus dorsi muscle is measured meat.
Table 10 produces influence of the γ-aminobutyric acid lactobacillus fermentation feed to Performance of Finishing Pigs
γ-aminobutyric acid has the function of increasing appetite, promotes growth as feed addictive；Bacillus acidi lactici is as prebiotic
Bacterium also has the function of improving feed quality in animal-breeding, improves the price of deed, promotes growth of animal, while having research to send out
Existing, lactobacillus fermentation feed influences the growth performance of fattening period pig to be better than growth phase.As can be seen from Table 10, this experiment
By adding the complete feed of production γ-aminobutyric acid lactobacillus fermentation in finishing pigs diets, its growth promoting function is carried out
Research, the results showed that, production γ-aminobutyric acid lactobacillus fermentation feed can significantly reduce growing and fattening pigs feedstuff-meat ratio, and 50% fermentation is raised
Expect that additive amount effect is better than 30% fermented feed additive amount.
Table 11 produces influence of the γ-aminobutyric acid lactobacillus fermentation feed to fattening meat quality
Table 12 produces influence (mg/100g) of the γ-aminobutyric acid lactobacillus fermentation feed to growing and fattening pigs muscle fatty acid
The evaluation of meat includes organoleptic feature, nutritive value and flavor substance etc., these are affected to varying degrees
The quality of meat.Yellowish pink is the important content of muscle appearance evaluation, depends mainly on myoglobins in muscle and hemoglobin
Content, Bacillus acidi lactici can inhibit Cell membrane lipids peroxidization, and the myoglobins in muscle is delayed to be oxidized to siderosis
Lactoferrin improves yellowish pink brightness to slow down yellowish pink dimmed speed.Redness a* values be influence muscle color it is main because
Element, a* values are higher, and meat quality is better, the display of this test result, and 50% production γ-aminobutyric acid lactobacillus fermentation feed can
Pork redness a* values are significantly improved, pork yellowing b* values are reduced, are improved yellowish pink.Lactobacillus fermentation feed has reduce brightness simultaneously
The trend of L* values, L* values are lower to show that muscular system waterpower is higher, and drip loss is smaller, this is consistent with experiment drip loss result.
Volatile materials such as amino acid, aliphatic acid etc. is to form the precursor substance of fragrance, and muscle fat can influence in meat
The ingredient of volatile materials is the main reason for influencing flavor, if intramuscular fat content increases, taste can also increase,
The succulence and tenderness of muscle can also be improved.This test result is also shown, and production γ-aminobutyric acid lactobacillus fermentation feed carries
High fat content increases the trend of pork succulence and flavor.Aliphatic acid forms other than influencing meat flavor in meat, and
The main source of aliphatic acid, especially saturated fatty acid in diet, it can influence many diseases in life, including cancer
Disease and angiocardiopathy etc.；And unsaturated fatty acid can then reduce the risk of these diseases.This test result is shown, produces γ-
Aminobutyric acid lactobacillus fermentation feed has increase unsaturated fatty acid, reduces the trend of saturated fatty acid, meanwhile, produce γ-ammonia
Base butyric acid section lactobacillus ferment feed also significantly improves the content of the palmitoleic acid with chronic disease therapy effect in muscle, carries
High muscle fat oleic acid/linoleic ratio, muscle fat oleic acid/linoleic ratio are also react meat flavor one
Index, ratio is higher, and flavour is with regard to dense.
Both at home and abroad about producing research of the γ-aminobutyric acid Bacillus acidi lactici to meat quality, this experiment from organoleptic feature,
Its influence to meat quality is had studied in terms of nutritive value and flavor substance, the results show that production γ-aminobutyric acid Bacillus acidi lactici
Fermented feed can improve meat quality in terms of improving yellowish pink, optimization nutritional ingredient and muscle fatty acid composition.
Embodiment 6
By 180 head growth growing and fattening pigs, it is randomly divided into control group and test group, each organizes 4 repetitions, each repeatedly 22-23
Head.100% basal diet of control group fed；Test group feeds+50% fermented feed of 50% basal diet.
The preparation method of fermented feed is：The basal diet of full bacterium solution fermentation 30% moisture of addition of 0.1% Bacillus acidi lactici, it is close
Seal normal temperature fermentation 3-7 days, viable count reaches 5.33 × 108CFU/g, GABA content 0.015%.During experiment be freely eaten and
Drinking-water manages feeding, drinking-water and the health condition of observation swinery daily according to pig farm conventinal breeding method, records feed consumption rate and hair
Sick death condition.Experimental period is the hot weather in October in July-, is amounted to 102 days.All stage is recorded using automatic feed system
Feed consumption rate, on-test and and at the end of early morning on an empty stomach weigh, weigh evening before that day 20：00 stops feed, does not cut off the water supply, and calculates
Average daily gain, average daily gain, feed-weight ratio.
Table 13 produces influence of the γ-aminobutyric acid lactobacillus fermentation feed to growing-finishing pig production performance under hot environment
Influence of the heat stress to cultivated animals body is very widely, instead by the various physiology of change body, biochemistry
It answers, reduces feed intake, the speed of growth, feed conversion rate and product quality, serious economic loss is brought to aquaculture.Lactic acid bar
Bacterium is body normal intestinal flora, by adjusting intestinal microecology balance, generates carbohydrate, albumen in special enzyme system decomposition food
Matter and fat improve food digestion rate, and synthesis vitamin promotes the approach such as Intestinal Mucosal Tissues development to play growth promoting function,
Simultaneously it has also been found that it can offset the negative effect that stress be brought.γ-aminobutyric acid has as feed addictive in excited feeding
Pivot, the effect for increasing appetite, promoting growth, and it is mainly also embodied in heat resistanceheat resistant in the effect for influencing animal body anti-stress ability
It stress aspect.The complete feed of γ-aminobutyric acid lactobacillus fermentation is produced in this experiment by being added in growing-finishing pigs diets,
It is studied in high-heat environment growth promoting function, the results showed that, production γ-aminobutyric acid lactobacillus fermentation Feed Energy is notable
Growing-finishing pig feedstuff-meat ratio is reduced, has the tendency that increasing average daily gain.
The preferred embodiment of the present invention has been described in detail above and comparative example.It should be appreciated that the common skill of this field
Art according to the present invention can be conceived without creative work makes many modifications and variations.Therefore, it is all in the art
Technical staff can be obtained by logical analysis, reasoning, or a limited experiment on the basis of existing technology under this invention's idea
The technical solution arrived, all should be in the protection domain being defined in the patent claims.
Claims (9)
1. a kind of culture medium of Bacillus acidi lactici production γ-aminobutyric acid, which is characterized in that according to mass fraction include following components：
Glucose 2%, yeast extract and rice bran mixture 3%, Pidolidone 1.5%, Tween-80 0.1%, epsom salt
0.02%, four water manganese sulfates 0.005%, surplus is distilled water.
2. a kind of culture medium of Bacillus acidi lactici production γ-aminobutyric acid according to claim 1, which is characterized in that described
Yeast extract is with rice bran according to mass ratio 1:1 mixing.
3. a kind of preparation method of the microorganism formulation containing γ-aminobutyric acid, which is characterized in that include the following steps：
1) tablet culture：Lactobacillus brevis P-14 is inoculated on solid activation medium tablet using streak inoculation method, in 30 DEG C
~37 DEG C of cultures 36~48h, recovery Lactobacillus brevis P-14 form single bacterium colony；
2) shake-flask seed liquid makes：The free of contamination Lactobacillus brevis P-14 single bacterium colonies that grow fine in step (1) are connect with oese
In kind to liquid activation medium, 30 DEG C~37 DEG C 36~48h of static gas wave refrigerator obtain shake-flask seed liquid；
3) domestication culture：The shake-flask seed liquid that access step (2) in culture medium is tamed in liquid carries out domestication culture, and inoculum concentration is
2~5%, condition of culture is 30 DEG C~37 DEG C, static gas wave refrigerator, and 36~48h passed on for two generations；
4) fermented and cultured：Fermentation medium is added in fermentation tank, take Lactobacillus brevis P-14 bacterium solutions in step (3) after domestication into
Row is inoculated with, inoculum concentration 0.1%, and ferment 72~96h at 30 DEG C, and microorganism formulation is obtained after fermentation.
4. preparation method according to claim 3, which is characterized in that the solid activation medium is according to the following recipe
It prepares：Peptone 10g, beef extract 10g, yeast extract 5g, dipotassium hydrogen phosphate 2g, Triammonium citrate 2g, sodium acetate 5g, grape
Sugared 20g, Tween 80 1mL, epsom salt 0.5g, four water manganese sulfate 0.25g, agar 20g, distilled water 1L, 121 DEG C sterilize
20min, cooling are spare.
5. preparation method according to claim 3, which is characterized in that the liquid activation medium is according to the following recipe
It prepares：Peptone 10g, beef extract 10g, yeast extract 5g, dipotassium hydrogen phosphate 2g, Triammonium citrate 2g, sodium acetate 5g, grape
Sugared 20g, Tween 80 1mL, epsom salt 0.5g, four water manganese sulfate 0.25g, distilled water 1L, 121 DEG C of sterilizing 20min are cooling
It is spare.
6. preparation method according to claim 3, which is characterized in that the domestication culture medium is matched according to the following recipe
System：Glucose 20g, yeast extract and rice bran mixture 30g, Pidolidone 15g, Tween-80 1.0mL, epsom salt
0.2g, four water manganese sulfate 0.05g, distilled water 1L, 121 DEG C of sterilizing 20min, cooling are spare.
7. preparation method according to claim 3, which is characterized in that the fermentation medium is matched according to the following recipe
System：Glucose 15Kg, yeast extract and rice bran mixture 22.5Kg, Pidolidone 11.25Kg, Tween-80 750mL, seven water sulphur
Sour magnesium 150g, four water manganese sulfate 37.5g, distilled water 750Kg, 75 DEG C of sterilizing 2h are spare after being quickly cooled to 35 DEG C.
8. the microorganism formulation containing γ-aminobutyric acid that claim 3 preparation method is prepared.
9. application of the microorganism formulation according to any one of claims 8 containing γ-aminobutyric acid in aquaculture.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810472692.8A CN108410917A (en) | 2018-05-17 | 2018-05-17 | The culture medium and its preparation of a kind of Bacillus acidi lactici production γ-aminobutyric acid and application |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810472692.8A CN108410917A (en) | 2018-05-17 | 2018-05-17 | The culture medium and its preparation of a kind of Bacillus acidi lactici production γ-aminobutyric acid and application |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN108410917A true CN108410917A (en) | 2018-08-17 |
Family
ID=63139769
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810472692.8A Pending CN108410917A (en) | 2018-05-17 | 2018-05-17 | The culture medium and its preparation of a kind of Bacillus acidi lactici production γ-aminobutyric acid and application |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108410917A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109479619A (en) * | 2019-01-22 | 2019-03-19 | 缙云县珍稀食用菌专业合作社 | A method of improving 4-Aminobutanoicacid content in elegant precious mushroom fructification |
| CN110339101A (en) * | 2019-06-14 | 2019-10-18 | 广东萱嘉医品健康科技有限公司 | A kind of fermentation of seaweed liquid magma and the preparation method and application thereof containing γ-aminobutyric acid |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1392262A (en) * | 2002-06-08 | 2003-01-22 | 江南大学 | Process for preparing food function factor gamma-amino-butyric acid |
| WO2004112816A1 (en) * | 2003-05-21 | 2004-12-29 | Ryukyu Bio-Resource Development Co., Ltd. | Fermentation product and process for producing the same |
| CN101928679A (en) * | 2009-09-22 | 2010-12-29 | 江南大学 | Breeding for efficiently converting L-glutamate into gamma-amino butyric acid lactobacillus |
| CN102212564A (en) * | 2010-04-02 | 2011-10-12 | 财团法人中华谷类食品工业技术研究所 | Fermentation method for producing gamma-aminobutyric acid and fermentation culture medium thereof |
| CN102925504A (en) * | 2012-08-23 | 2013-02-13 | 浙江师范大学 | Method and fermentation culture medium used for synthesizing gamma-aminobutyric acid through microbial fermentation |
-
2018
- 2018-05-17 CN CN201810472692.8A patent/CN108410917A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1392262A (en) * | 2002-06-08 | 2003-01-22 | 江南大学 | Process for preparing food function factor gamma-amino-butyric acid |
| WO2004112816A1 (en) * | 2003-05-21 | 2004-12-29 | Ryukyu Bio-Resource Development Co., Ltd. | Fermentation product and process for producing the same |
| CN101928679A (en) * | 2009-09-22 | 2010-12-29 | 江南大学 | Breeding for efficiently converting L-glutamate into gamma-amino butyric acid lactobacillus |
| CN102212564A (en) * | 2010-04-02 | 2011-10-12 | 财团法人中华谷类食品工业技术研究所 | Fermentation method for producing gamma-aminobutyric acid and fermentation culture medium thereof |
| CN102925504A (en) * | 2012-08-23 | 2013-02-13 | 浙江师范大学 | Method and fermentation culture medium used for synthesizing gamma-aminobutyric acid through microbial fermentation |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109479619A (en) * | 2019-01-22 | 2019-03-19 | 缙云县珍稀食用菌专业合作社 | A method of improving 4-Aminobutanoicacid content in elegant precious mushroom fructification |
| CN110339101A (en) * | 2019-06-14 | 2019-10-18 | 广东萱嘉医品健康科技有限公司 | A kind of fermentation of seaweed liquid magma and the preparation method and application thereof containing γ-aminobutyric acid |
| CN110339101B (en) * | 2019-06-14 | 2022-06-10 | 广东萱嘉医品健康科技有限公司 | Seaweed fermentation broth protoplasm containing gamma-aminobutyric acid and preparation method and application thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103766650B (en) | A kind of feed addictive of layer chicken and preparation thereof | |
| CN109251874A (en) | A kind of probiotics preparation and its preparation method and application | |
| CN102823726B (en) | It is a kind of improve cottonseed meal content and can detoxification biofermentation method | |
| CN101919494B (en) | Complex microorganism additive as well as preparation method and application thereof | |
| CN101278702B (en) | Novel microbial feed additive and method of producing the same | |
| AU2020104245A4 (en) | A Biological Fermentation Preparation and Its Preparation Method | |
| CN1813562A (en) | Compound microbial fermented fodder nutrient additive and its preparing method | |
| CN107535724B (en) | Flammulina velutipes foot ferment and application thereof | |
| CN107373022B (en) | Pig feed | |
| CN106974063B (en) | Method for producing high-efficiency protein feed by using feed enzyme in cooperation with bacillus coagulans | |
| CN103392910A (en) | Method for preparing microbial fermented feed for effectively improving mutton quality | |
| CN106962594A (en) | A kind of Selenium-enriched fermentation dregs of beans, preparation method and application | |
| CN109315586A (en) | A kind of preparation method and application of ruminant fermentation of Chinese herbal medicine biological feedstuff | |
| CN107041483A (en) | A kind of laying cycle of laying hens feed for improving premunition and preparation method thereof | |
| CN112175846B (en) | Candida utilis strain UCY-11 and application thereof in preparation of fermented hybrid broussonetia papyrifera feed | |
| CN112136965B (en) | Immunity-enhancing and growth-promoting fermented Chinese herbal medicine feed additive and preparation method thereof | |
| CN103289907A (en) | A candida utilis protein product and applications thereof in breeding industry | |
| CN109315605A (en) | A kind of fermentation process of peanut meal, the fermented peanut meal of this method preparation and its application | |
| CN107937311A (en) | The streptococcus thermophilus of one plant height production gamma aminobutyric acid, preserve cultural method and the method that acidified milk is prepared using the streptococcus thermophilus | |
| CN102577839B (en) | Method for producing medicinal or medicine-food dual-purpose fruit body by utilizing yellow wine grains as solid culture medium | |
| CN108432979A (en) | A kind of Australia freshwater lobster ecological feed | |
| CN103952336A (en) | Bacillus licheniformis-Bacillus subtilis-Lactobacillus casei preparation and preparation method thereof | |
| CN108410917A (en) | The culture medium and its preparation of a kind of Bacillus acidi lactici production γ-aminobutyric acid and application | |
| CN101690541B (en) | Method for preparing feed protein from microbial fermented silkworms | |
| KR102492755B1 (en) | Method for preparing fermented total mixed ration using microbial strain complex and steam treatment |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| TA01 | Transfer of patent application right |
Effective date of registration: 20200106 Address after: 610000 No. 588, east route, Longquanyi District, Chengdu, Sichuan Province Applicant after: Sichuan Jinke pharmaceutical limited liability company Address before: 610100 77 B101, Vanke City Garden, Jinjiang District, Chengdu, Sichuan Applicant before: Song Yan |
|
| TA01 | Transfer of patent application right | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20180817 |
|
| WD01 | Invention patent application deemed withdrawn after publication |