CN117327631B - Lactobacillus plantarum strain 10A-2 for improving anxiety and depression and application thereof - Google Patents
Lactobacillus plantarum strain 10A-2 for improving anxiety and depression and application thereof Download PDFInfo
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- CN117327631B CN117327631B CN202311605089.XA CN202311605089A CN117327631B CN 117327631 B CN117327631 B CN 117327631B CN 202311605089 A CN202311605089 A CN 202311605089A CN 117327631 B CN117327631 B CN 117327631B
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Abstract
The invention belongs to the field of probiotics, and provides a Lactobacillus plantarum strain 10A-2 for improving anxiety and depression and application thereof, wherein the Lactobacillus plantarum strain 10A-2 has been subjected to patent strain preservation in China general microbiological culture collection center (CCTCCNO): m2023621. The strain has the effects of regulating intestinal flora balance, promoting digestion and absorption of human body, and regulating and effectively relieving and treating insomnia, anxiety and depression and improving sleep quality. Daily administration of 60 million CFU and 100mg of metabolite of the strain of the present invention can provide the necessary endogenous amounts of cobalamin, phosphatidylserine, acetylcholine for sustainability during intestinal reproduction and growth. The composition has the effects of preventing, relieving and treating the anxiety depression, and can effectively maintain a healthy nervous system. Solves the problem that the bacterial agent with good improving effect on the anxiety depression is urgently needed at present.
Description
Technical Field
The invention belongs to the field of probiotics, and particularly relates to a lactobacillus plantarum strain 10A-2 for improving anxiety and depression and application thereof.
Background
The disclosure of this background section is only intended to increase the understanding of the general background of the invention and is not necessarily to be construed as an admission or any form of suggestion that this information forms the prior art already known to those of ordinary skill in the art.
Along with the increase of life rhythm and working pressure, the prevalence rate of anxiety and depression is increased, and serious influence is caused to life, study and work of people. Anxiety depression, also called anxiety neurosis, is a major clinical manifestation of generalized anxiety disorder and narcolepsy, often accompanied by dizziness, chest distress, palpitation, dyspnea, dry mouth, frequent urination, urgent urination, sweating, tremor, and restlessness, where anxiety is not caused by a real threat or where the degree of stress and panic is not commensurate with the reality. The experience of depressed mood can be reflected from the patient's speaking, expression, posture. Depressed patients often describe their mood experience over a longer period of time in terms of "low mood", "happy up", "depressed", "poor mood", "dysphoric", "uncomfortable mood", "no interest in anything" and the like.
Probiotics are a general term for a class of microorganisms that exert beneficial effects on host health. The probiotics can regulate emotion and relieve depression.
Microbiota generally refers to: all microorganisms living in or on the human body, such as intestinal flora. Enteric bacteria can affect the nervous system through metabolites. The brain and the intestine cooperate closely, and this connection is called the "gut-brain axis (GBA)", which connects the central nervous system and the brain. The human intestinal tract produces hundreds of neurochemicals, i.e., neurotransmitters, for use by the brain in psychological processes, such as: emotion, memory, and learning, etc. It has been found that administration of probiotics can improve anxiety and depression to some extent. However, there is an urgent need for a bacterial agent having a good ameliorating effect on anxiety depression.
Disclosure of Invention
In order to solve the problems, the invention provides a Lactobacillus plantarum strain 10A-2 for improving anxiety and depression and application thereof. The invention separates a lactobacillus plantarum from fresh mangosteen pulp in Guangxi nan in 10 months of 2007, and the lactobacillus plantarum is identified by a 16srRNA method (16S ribosomal RNA gene sequencing technology), and the lactobacillus plantarum has the effects of regulating intestinal flora balance, promoting human digestion and absorption, and simultaneously conditioning and effectively relieving and treating insomnia, anxiety, depression and improving sleep quality. The daily administration of 60 hundred million CFU and 100mg of metabolite of the strain can continuously provide the necessary endogenous quantity of cobalamin, phosphatidylserine and acetylcholine in the intestinal tract propagation and growth process, has the effects of preventing, relieving and treating the anxiety depression, and can keep the health of the nervous system.
In order to achieve the above purpose, the present invention adopts the following technical scheme:
in a first aspect, the invention provides a Lactobacillus plantarum strain for improving anxiety and depressionLactobacillus pLantarum) 10A-2, wherein the lactobacillus plantarum strain 10A-2 has been subjected to patent strain preservation in China Center for Type Culture Collection (CCTCCNO), and the preservation number is: m2023621, the preservation date is 25 days of 2023, 04 month and the preservation address is university of Wuhan.
In a second aspect, the invention provides application of the lactobacillus plantarum strain 10A-2 in preparation of a microbial inoculum for improving anxiety depression.
In a third aspect, the invention provides an application of the lactobacillus plantarum strain 10A-2 in preparing a microbial inoculum for improving sleep disorder.
Preferably, the Lactobacillus plantarum strain 10A-2 is ingested in an amount of greater than 60 hundred million CFU/day during consumption.
Preferably, the improvement comprises: has at least one of preventing, relieving and treating effects on related symptoms.
In a fourth aspect, the invention provides a method for preparing a bacterial agent for improving anxiety depression, comprising the following steps:
performing expansion culture, primary seed culture, seed tank culture and fermentation culture on the lactobacillus plantarum strain 10A-2 to obtain fermentation broth;
centrifuging the fermentation bacteria liquid to respectively collect fermented lactobacillus plantarum bacterial mud and centrifuged fermentation liquid, and respectively freeze-drying and crushing the fermented lactobacillus plantarum bacterial mud and the centrifuged fermentation liquid to obtain fermented lactobacillus plantarum bacterial powder and fermented lactobacillus plantarum metabolite powder;
mixing lactobacillus plantarum powder and lactobacillus plantarum metabolite powder with a bacteria powder carrier to obtain the lactobacillus plantarum.
Preferably, 9-12 parts of the lactobacillus plantarum powder, 4-6 parts of the lactobacillus plantarum metabolite powder and 114-166 parts of the fungus powder carrier.
Preferably, the fungus powder carrier consists of the following raw materials in parts by weight: 18-22 parts of microcrystalline cellulose, 80-120 parts of medicinal corn starch, 8-12 parts of inulin and 8-12 parts of magnesium stearate.
Preferably, the inoculation amount of the primary seed culture is 0.5-0.6% in terms of volume ratio.
Preferably, the inoculation amount of the seed tank culture is 0.3-0.4% in terms of volume ratio.
Preferably, the inoculation amount of the fermentation is 0.3-0.4% by volume.
Preferably, the centrifugal speed is 10000 r r/min-12000 r/min.
Preferably, the moisture content of the microbial inoculum is less than or equal to 4 percent in mass ratio.
The beneficial effects of the invention are that
(1) The lactobacillus plantarum strain 10A-2 has the functions of regulating intestinal flora balance and promoting human digestion and absorption, and simultaneously has the effects of conditioning, effectively preventing, relieving or treating anxiety and depression and improving sleep quality. The daily administration of 60 hundred million CFU and 100mg of metabolite of the strain can continuously provide the necessary endogenous quantity of cobalamin, phosphatidylserine and acetylcholine in the intestinal tract propagation and growth process, and has the effects of preventing, relieving and treating the anxiety depression, and better maintaining the health of the nervous system.
(2) The lactobacillus plantarum viable bacteria and the metabolites thereof have the advantages of simple production process, short period, easy operation, easy acquisition of used raw materials, no three-waste emission in production, and meeting the requirements of green and environment-friendly production.
Detailed Description
It should be noted that the following detailed description is exemplary and is intended to provide further explanation of the invention. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.
The invention will now be described in further detail with reference to the following specific examples, which should be construed as illustrative rather than limiting.
Equipment used in production: strain incubator, ultra-clean bench, constant temperature incubator, various test tubes, triangular flask, seed tank, fermenter, vacuum freeze dryer, three-dimensional stirrer, low temperature pulverizer, and vacuum packaging machine.
Example 1 identification of strains
The lactobacillus plantarum is separated from fresh mangosteen pulp, and identified as lactobacillus plantarum 10A-2 by a 16srRNA method, and the sequence of the strain is shown as SEQ ID NO: 1.
EXAMPLE 2 Strain culture
1. Enlarged strain culture
The lactobacillus plantarum 10A-2 obtained in example 1 preserved in a refrigerator at-80 ℃ is subjected to expansion culture, and a culture medium is prepared: stachyose 15g, maltitol 3g, beef extract powder 4g, sodium acetate 1.5g, tween-800.5 mL, monopotassium phosphate 0.8g, tap water 1000mL, pH value adjusted to 7 (alkali liquor adjustment).
The operation is as follows: adding the raw materials in the formula into 1000mL of water before and after the raw materials are not separated, stirring for 10min to dissolve the raw materials, then subpackaging into test tubes, filling 5mL of each test tube with kraft paper, sealing, sterilizing in a medical sterilizer at 0.1-0.12 MPa for 20min, and finishing the preparation of the culture medium.
Inoculating: under aseptic condition (super clean bench), opening a freezer at-80deg.C for preserving two Lactobacillus plantarum 10A-2, thawing at room temperature, taking 1 test tube culture medium, adding 0.5mL of each of the two thawed bacteria into the test tube, sealing the tube orifice with kraft paper, slightly shaking with hands, placing into a constant temperature incubator at 37deg.C, and standing for 16 hr to obtain the final product.
2. First-stage seed culture
The primary seed culture formula comprises: 5g of isomaltulose, 3g of peptone, 2g of beef extract powder, 1.2g of arginine, 0.5g of sodium acetate, 1.5g of threonine, 0.3g of tyrosine, 1mL of tween-80 and 1000mL of tap water, and the pH value is natural.
The operation is as follows: the above raw materials are added into 1000mL tap water before and after the raw materials are not separated, a stirrer is started, and the raw materials are stirred for 10min to dissolve all the raw materials, so that the raw materials are called primary seed culture solution.
Adding the stirred and melted primary seed culture solution into a triangular flask according to the amount, wrapping a bottleneck by using 6 layers of gauze and adding a layer of kraft paper, placing the flask into a sterilizer for sterilization, opening an exhaust valve on the sterilizer and the like at first, closing the flask when a small amount of steam is discharged, opening the exhaust valve on the medical sterilizer for exhausting for 6min-8min when the pressure on the medical sterilizer reaches 0.05MPa, then closing the exhaust valve, continuing to heat, starting timing when the steam pressure in the medical sterilizer reaches 0.1MPa-0.12MPa, sterilizing for 20min, leaving a heat source after the process is finished, naturally cooling, opening the medical sterilizer when a pressure meter on the medical sterilizer returns to zero, taking out a sterilized culture medium, placing the sterilized culture medium into an ultra-clean workbench for natural cooling, transferring the cultured expanded solution into the sterilized culture solution of the triangular flask when the temperature is reduced to 37 ℃, and inoculating strains with the inoculum size of 0.5%, namely: 100mL of primary seed culture solution is inoculated into 0.5mL of expansion culture solution, after the completion, the bottle mouth is sealed by original gauze and kraft paper, the primary seed culture solution and the kraft paper are mixed uniformly by slightly shaking the primary seed culture solution and the primary seed culture solution, and then the primary seed culture solution is placed in an incubator for culturing for 18h at 37 ℃, thus the primary seed culture solution is called primary strain solution.
3. Seed tank culture
Sterilization of fermentation equipment, tubing, and sterile filtration systems:
the operation is as follows: firstly, opening valves of each inlet and outlet pipeline and a sterile air pipeline, introducing steam of 0.12-0.14 MPa, enabling the steam to be communicated with the pipeline valves and a small amount of steam to be discharged, ventilating for 40min, and then closing the valves of each inlet and outlet pipeline for standby.
Empty pot sterilization of seed pot and fermentation pot:
closing each valve, opening the drain valve at the bottom of the tank and the drain valve in the interlayer of the tank, opening the direct steam valve to introduce steam of 0.12MPa-0.14MPaMPa into the pipe, starting timing when the temperature in the tank reaches 121 ℃, sterilizing for 40min, and then closing the drain valve at the bottom of the tank and the drain valve in the interlayer, and naturally cooling the temperature in the tank to 37 ℃ for standby.
Seed pot culture:
the formula comprises the following components: 2% of isomaltulose, 0.7% of casein peptone, 0.1% of dipotassium hydrogen phosphate, 0.5% of serine, 0.1% of sodium acetate, 0.8% of arginine, 0.6% of phenylalanine, 0.1% of tween-80, 0.1% of polyether defoamer, 92.4% of tap water and natural pH value, wherein the percentages are calculated according to mass ratio.
The specific operation is as follows: firstly, putting tap water used in a formula into a seed tank sterilized by an empty tank, starting a stirrer, respectively adding raw materials required by the formula (the adding sequence is not divided into front and rear), continuously stirring, introducing steam, heating to 95 ℃ by utilizing tank interlayer steam, heating by direct steam, keeping the temperature in the tank to 121 ℃ for 20min to achieve a sterilization effect, closing the steam, starting tap water cooling through the tank interlayer, and standing by when the temperature in the tank reaches 37 ℃, thus the seed culture medium is obtained. Inoculating the cultured primary strain liquid into a seed tank under the aseptic condition, wherein the inoculum size is 3% of the medium in the tank, and 3kg of the primary strain liquid is respectively inoculated into 100kg of the seed tank medium, and then, the inoculating cap on the tank is covered to start stirring culture, and the culture conditions are as follows: the temperature is 37 ℃, the pH value is natural, the stirring rotation speed is 120r/min, the tank pressure is 0.05MPa, the culture time is 12 hours, and the requirements can be met, if the tank pressure is reduced, sterile air can be introduced to keep the tank pressure, and the tank pressure is excessively high and can be adjusted by a deflation valve at the top of the tank, so that the seed tank is called as seed tank strain.
4. Fermentation culture
The formula comprises the following components: 2% of isomaltulose, 0.5% of casein peptone, 1% of xylooligosaccharide, 1.2% of stachyose, 3% of wheat oligopeptide, 2% of yeast extract powder, 1% of sodium acetate, 0.2% of tween-80, 0.3% of leucine, 0.2% of proline, 0.5% of dipotassium hydrogen phosphate, 0.1% of polyether defoamer and 69.2% of tap water.
The specific operation is as follows: firstly, tap water used in the formula is put into a fermentation tank sterilized by an empty tank, a stirrer is started, raw materials used in the formula are put into the fermentation tank (the adding sequence is not divided into the front and the back), and then steam is introduced for heating. Firstly, heating to 95 ℃ by using interlayer steam, then, heating by using direct steam, starting timing when the temperature in the tank reaches 121 ℃, keeping for 30min to achieve a sterilization effect, then, cooling by using interlayer tap water, starting inoculating strains when the temperature in the tank is reduced to 37 ℃, firstly, increasing the tank pressure of a seed tank to 0.1MPa before inoculating, keeping the tank pressure of a fermentation tank to 0.05MPa, then, opening an inoculating pipeline valve, conveying the seed tank strains cultured in the seed tank into the fermentation tank in a pressure difference mode, and then closing an inoculating channel valve.
Inoculating 3kg of seed tank strain into each 100kg of culture medium in a fermentation tank, and culturing: culturing at 37deg.C for 0-5 hr, stirring at 120r/min, maintaining the pressure at 0.05MPa, maintaining pH at 6 by automatic acid and alkali regulating system during fermentation, introducing sterile air if the pressure is reduced, maintaining the pressure until fermentation is completed, fermenting for 25 hr, and cooling to 20deg.C.
5. Freeze drying of lactobacillus fermentum mud and fermentation broth
(1) Pumping the fermentation liquor into a storage tank for centrifugation, and operating: starting a centrifugal machine, adjusting the rotation speed to 10000 r/min, centrifuging at a feeding speed of 6kg/min, collecting wet bacterial sludge after centrifugation, and freeze-drying the collected wet bacterial sludge.
(2) Freeze drying of bacterial mud metabolites
Preparation of a suspension: 1000g of inactivated wet bacterial sludge is taken, 4000 ml of sterile physiological saline, 300g of maltodextrin, 1500g of anhydrous glucose, 10g of zinc sulfate and 40g of sodium gluconate are added.
The operation is as follows: firstly 2500mL of physiological saline in the formula is taken, maltodextrin is added, stirring is carried out, 1000g of inactivated wet bacterial mud is added, the rest physiological saline, anhydrous glucose and auxiliary materials in the formula are added, stirring is fully and uniformly carried out, the suspension is called suspension, freeze drying is carried out on the suspension, and finally the moisture content is controlled to be less than or equal to 4%, thus obtaining the lactobacillus plantarum fermented powder.
2. Low-temperature concentration of the centrifugal fermentation liquor: concentrating under vacuum degree of 0.06MPa at 55-65deg.C until the solid content is 80% and the water content is 20% (by mass ratio), adding medicinal corn starch as carrier, mixing, and vacuum drying. Taking 1000g of fermentation liquor concentrated extract, adding 200g of medicinal corn starch, uniformly mixing, and drying in a vacuum drying oven under the condition of: vacuum degree 0.06MPa, temperature 57 deg.C, drying time 45h, moisture content 3% is qualified, then pulverizing to 100 mesh powder, sealing and packaging for use to obtain fermented lactobacillus plantarum metabolite powder. Wherein, the mass fraction ratio of the fermentation liquor concentrated extract to the medicinal corn starch is 4.5:1.
compounding of a microbial inoculum: 9g of 100-mesh fermented lactobacillus plantarum bacterial powder, 4g of fermented lactobacillus plantarum metabolite powder, 18g of microcrystalline cellulose, 80g of medicinal corn starch, 8g of inulin and 8g of magnesium stearate are taken, wherein microcrystalline cellulose, medicinal corn starch, inulin and magnesium stearate are taken as bacterial powder carriers. The materials are put into a solid stirring mixer to be mixed uniformly, then the materials are packaged into capsules under the aseptic condition, and the finished product of the composition is obtained after bottling or tabletting.
Example 3
The difference from example 2 is that the microbial inoculum is compounded: taking 12g of fermented lactobacillus plantarum bacterial powder of 100 meshes, 6g of fermented lactobacillus plantarum metabolite powder, 22g of microcrystalline cellulose, 120g of medicinal corn starch, 12g of inulin and 12g of magnesium stearate.
Example 4
The difference from example 2 is that the inoculation amount of the primary seed culture is 0.6% by volume.
Example 5
The difference from example 2 is that the inoculum size of the seed tank culture was 0.4% by volume.
Example 6
The difference from example 2 is that the inoculum size of the fermentation culture is 0.4% by volume.
Example 7
The difference from example 2 is that the rotational speed of the centrifugation is 12000r/min.
Example 8
Clinical trial
1 general data
A total of 600 cases (i.e., anxiety-depressive patients) were selected from 7/2020 and 8/2021 who received the subject pharmaceutical composition and had sleep disorders, depression, and anxiety symptoms, wherein treatment 1: middle-aged and elderly male 200, ages 34-80, and average age 55; treatment group 2: middle-aged and elderly female 200, ages 33-82, and average age 56; treatment 3 groups: teenager male 100, age 15-18 years, average age 15 years; treatment 4 groups: teenager females 100, 16-18 years of age, with an average age of 16 years; the control group was 200 persons with an average age of 30 years, as subjects. The patients in each group have no statistical significance (P is more than 0.05) in comparison with the basic data such as gender, age, disease type and the like, and have comparability.
2 evaluation criteria
Improvement: the emotional response is weakened, the sleep disorder is relieved, and the somatic symptoms are relieved.
The obvious improvement is as follows: the emotion is controlled, sleep disorder is reduced, and somatic symptoms are reduced.
The improvement is extremely obvious: mood tends to stabilize, sleep is basically stable, and a few somatic symptoms exist.
The complete improvement is as follows: stable emotion, regular sleep and disappearance of somatic symptoms.
The improvement is not: the symptoms were not significantly reduced or aggravated.
3 methods of treatment
Treatment 1-4 groups: the Lactobacillus plantarum lyophilized powder prepared in example 2 was administered to patients, respectively, 0.3g of the powder was administered once a day before sleeping, and each 0.3g of the powder contained 60 hundred million CFU of live Lactobacillus plantarum and 100mg of metabolite.
Results 4 results
The results of the comparison of the four groups of clinical treatment effects after 20 days of treatment are shown in Table 1.
Table 1 effects of improving anxiety and depression
Description: the statistics result is started after the male 200 people take the medicine for 2 days, the condition of the anxiety and depression of 110 people with different ages is improved after the male 200 people take the medicine for 2 days, and the anxiety and depression of 152 people are obviously improved after the male 200 people take the medicine for 8 days. The anxiety depression of 164 people is extremely obvious to be improved after 14 days of taking the medicine. The anxiety and depression conditions of 190 people are completely improved after 20 days of taking, the mood of things is increased, the interests of all aspects in life are increased, and the pleasure and comfort of the mood can be maintained; a total of 10 people cannot improve anxiety and depression after taking lactobacillus plantarum.
Female: 200 people start counting results after taking 2 days, 112 people with different ages feel anxiety and depression conditions are improved after taking 2 days, and 159 people feel anxiety and depression conditions are obviously improved after taking 8 days. The improvement of the anxiety depression condition of 169 people is extremely obvious after 14 days of taking the medicine. The anxiety and depression conditions of 191 people are completely improved after 20 days of taking, the mood of things is increased, the interests of all aspects in life are increased, and the pleasure and comfort of the mood can be kept; a total of 9 people cannot improve anxiety depression after taking lactobacillus plantarum.
Teenagers' men: 100 people start counting results after taking the medicine for 2 days, 59 people with different ages feel anxiety and depression conditions are improved after taking the medicine for 2 days, and 69 people feel anxiety and depression conditions are obviously improved after taking the medicine for 8 days. The anxiety depression of 75 people is extremely obvious to be improved after 14 days of taking the medicine. The anxiety and depression conditions of 92 people are completely improved after 20 days of taking, the mood of things is increased, the interests of all aspects in life are increased, and the pleasure and comfort of the mood can be kept; a total of 8 people can not improve anxiety and depression after taking lactobacillus plantarum.
Teenagers and women: 100 people start counting results after taking the medicine for 2 days, 60 people with different ages feel anxiety and depression conditions are improved after taking the medicine for 2 days, and 70 people feel anxiety and depression conditions are obviously improved after taking the medicine for 8 days. The anxiety depression of 76 people is extremely obviously improved after 14 days of taking the medicine. 93 people can completely improve anxiety and depression conditions after taking the medicine for 20 days, the mood of things is increased, the interests of all aspects in life are increased, and the pleasure and comfort of the mood can be kept; a total of 7 people cannot improve anxiety and depression after taking Lactobacillus plantarum.
The above description is only of the preferred embodiments of the present invention and is not intended to limit the present invention, but various modifications and variations can be made to the present invention by those skilled in the art. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (10)
1. Lactobacillus plantarum strain 10A-2 for improving anxiety and depression, which is classified and named as Lactobacillus plantarumLactobacillus pLantarum) The strain is preserved in China Center for Type Culture Collection (CCTCC) for 4 and 25 days in 2023, and the biological preservation number is CCTCCNO: m2023621.
2. Use of lactobacillus plantarum strain 10A-2 according to claim 1 for the preparation of a bacterial agent for improving anxiety depression.
3. Use of lactobacillus plantarum strain 10A-2 according to claim 1 for the preparation of a microbial agent for improving sleep disorders.
4. A method for preparing a microbial agent for improving anxiety and depression, which is characterized by comprising the following steps:
performing expansion culture, primary seed culture, seed tank culture and fermentation culture on the lactobacillus plantarum strain 10A-2 of claim 1 to obtain fermentation broth;
centrifuging the fermentation bacteria liquid to respectively collect fermented lactobacillus plantarum bacterial mud and centrifuged fermentation liquid, and respectively freeze-drying and crushing the fermented lactobacillus plantarum bacterial mud and the centrifuged fermentation liquid to obtain fermented lactobacillus plantarum bacterial powder and fermented lactobacillus plantarum metabolite powder;
mixing lactobacillus plantarum powder and lactobacillus plantarum metabolite powder with a bacteria powder carrier to obtain the lactobacillus plantarum.
5. The method for preparing the bacterial agent for improving anxiety and depression as claimed in claim 4, wherein 9-12 parts of the lactobacillus plantarum bacterial powder, 4-6 parts of the lactobacillus plantarum metabolite powder and 114-166 parts of the bacterial powder carrier are fermented.
6. The preparation method of the microbial inoculum for improving anxiety and depression as claimed in claim 4, wherein the microbial powder carrier consists of the following raw materials in parts by weight: 18-22 parts of microcrystalline cellulose, 80-120 parts of medicinal corn starch, 8-12 parts of inulin and 8-12 parts of magnesium stearate.
7. The method for preparing a bacterial agent for improving anxiety and depression as claimed in claim 4, wherein the inoculation amount of the primary seed culture is 0.5% -0.6% by volume.
8. The method for preparing the microbial inoculum for improving anxiety depression as claimed in claim 4, wherein the inoculation amount of the seed tank culture is 0.3% -0.4% by volume ratio.
9. The method for preparing a microbial agent for improving anxiety and depression as claimed in claim 4, wherein the inoculation amount of the fermentation is 0.3% -0.4% by volume.
10. The method for producing a bacterial agent for improving anxiety and depression as claimed in claim 4, wherein the rotational speed of the centrifugation is 10000 r/min-12000 r/min.
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| WO2018034430A1 (en) * | 2016-08-16 | 2018-02-22 | 주식회사 엠디헬스케어 | Composition for preventing or treating mental disorder, containing lactobacillus plantarum-derived vesicle |
| CN112204129A (en) * | 2018-02-02 | 2021-01-08 | Ko生物技术有限公司 | Lactobacillus plantarum KBL396 strain and application thereof |
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