Application of lactobacillus mucilaginosus strain A21196 in preparation of product for resisting anxiety and improving memory and concentration

Abstract

The invention provides an application of a fermentation lactobacillus mucus strain A21196 in preparation of a product for resisting anxiety and improving memory and concentration, and belongs to the technical field of biological medicines. Animal experiments show that the fermented lactobacillus mucus strain A21196 has the effects of resisting anxiety, resisting depression, improving cognitive impairment function and improving memory concentration, and provides a new means for preventing and / or treating nerve diseases related to anxiety disorder, depression and cognitive impairment.

Description

Technical Field

[0001] The invention belongs to the technical field of biological medicines, and particularly relates to an application of a fermented Lactobacillus fermentum strain A21196 in preparing products for resisting anxiety and improving memory and concentration. Background Art

[0002] Fermented mucus lactobacillus is a well-known intestinal probiotic. It is reported that the intestinal probiotic effects of fermented mucus lactobacillus include enhancing immunity, improving gastrointestinal function, exerting antioxidant function, lowering blood sugar and blood pressure, enhancing intestinal mucosal barrier function to maintain intestinal flora balance, degrading uric acid, inhibiting harmful bacteria and improving nutrient absorption, and alleviating alcoholic liver damage. It can be seen that fermented mucus lactobacillus plays an important role in promoting human health and preventing diseases. However, it should be noted that different strains may have different characteristics and effects. At present, there is no report on the field of fermented mucus lactobacillus in improving anxiety and improving cognitive functions such as memory. Summary of the invention

[0003] In view of this, the object of the present invention is to provide a use of fermented Lactobacillus mucilaginosus strain A21196 in the preparation of products for resisting anxiety and improving memory and concentration.

[0004] The invention provides an application of a fermented Lactobacillus mucilaginosus strain A21196 in preparing a medicine for preventing and / or treating anxiety or depression. The deposit number of the fermented Lactobacillus mucilaginosus strain A21196 is GDMCC No: 62959.

[0005] Preferably, the anxiety disorder includes at least one of the following anxiety behaviors and complications: acute anxiety, chronic anxiety, neurological dysfunction, cardiovascular and cerebrovascular diseases, and digestive system diseases.

[0006] The invention provides an application of a fermented Lactobacillus mucosa strain A21196 in preparing a medicine for preventing and / or treating cognitive dysfunction and / or improving memory. The deposit number of the fermented Lactobacillus mucosa strain A21196 is GDMCC No: 62959.

[0007] Preferably, the cognitive dysfunction includes at least one of the following behavioral disorders: memory impairment, aphasia, visuospatial disorder, executive dysfunction, calculation disorder, apraxia and agnosia.

[0008] Preferably, the fermented Lactobacillus mucilaginosus strain A21196 comprises at least one of the following cultures: bacterial bodies of the fermented Lactobacillus mucilaginosus strain A21196, metabolites of the fermented Lactobacillus mucilaginosus strain A21196, and fermentation broth of the fermented Lactobacillus mucilaginosus strain A21196.

[0009] Preferably, the fermented Lactobacillus mucilaginosus strain A21196 also exists in the form of a probiotic.

[0010] Preferably, the probiotic comprises at least one of the following: tablets, drops, capsules and powders.

[0011] Preferably, the viable bacterial concentration of the fermented Lactobacillus mucilaginosus strain A21196 in the probiotic is not less than 2×10 9 CFU / mL or 2×10 9 CFU / g.

[0012] Preferably, the viable bacterial concentration of the fermented Lactobacillus mucilaginosus strain A21196 is 5×10 9 CFU / mL~100×10 9 CFU / mL or 5×10 9 CFU / g~100×10 9 CFU / g.

[0013] Preferably, the drug comprises at least one of the following oral dosage forms: tablets, powders, capsules and oral liquids.

[0014] The present invention provides an application of a fermented Lactobacillus mucoides strain A21196 in the preparation of a drug for preventing and / or treating anxiety or depression, and the deposit number of the fermented Lactobacillus mucoides strain A21196 is GDMCC No: 62959. The present invention establishes a chronic unpredictable mild stress model in mice, and uses a sucrose preference test, a tail suspension test, and a forced swimming test to evaluate the antidepressant efficacy of the fermented Lactobacillus mucoides strain A21196. The results of the sucrose preference experiment show that compared with the model group, the sugar water preference value of the mice in the strain A21196 group is significantly increased, indicating that the strain A21196 of the present invention has the function of improving the anhedonia of depressed mice. The results of the tail suspension experiment show that compared with the model group, the immobility time of the mice in the strain A21196 group is significantly shortened. At the same time, the results of the forced swimming experiment show that compared with the model group, the immobility time of the mice in the A21196 group is significantly shortened. This shows that the strain A21196 can effectively alleviate the depressive-like behavior of the model mice. At the same time, the present invention uses an open field test and an elevated plus maze test to evaluate the anti-anxiety behavior efficacy of fermented mucus lactobacillus strain A21196. The results of the open field test showed that compared with the model group, the time that the A21196 group mice entered the center of the open field was significantly increased; the results of the elevated plus maze test showed that compared with the model group, the number of times the A21196 group mice entered the open arm and the percentage of time in the open arm were significantly increased, and the time in the closed arm was significantly shortened. This shows that the fermented mucus lactobacillus strain A21196 can effectively alleviate the anxiety behavior of model mice, providing a new means for the treatment of depression and anxiety.

[0015] The present invention provides an application of a fermented Lactobacillus mucoides strain A21196 in the preparation of a drug for preventing and / or treating cognitive dysfunction and / or improving memory, and the deposit number of the fermented Lactobacillus mucoides strain A21196 is GDMCC No: 62959. The present invention uses a novel object recognition experiment to evaluate the efficacy of the fermented Lactobacillus mucoides strain A21196 in improving cognitive function and improving learning and memory. The results of the novel object recognition experiment showed that compared with the model group, the percentage of time that the mice in the A21196 group explored new objects was significantly increased, indicating that the A21196 strain can effectively improve the cognitive behavior of model mice and improve the memory of mice. The technical solution of the present invention provides a new means for the treatment of cognitive dysfunction and related neurological diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 This is the result of the effect of strain A21196 on the body weight of CUMS mice;

[0017] Figure 2 The test results show that strain A21196 significantly improves sucrose preference in CUMS mice;

[0018] Figure 3 The test results show that strain A21196 significantly shortens the tail suspension immobility time of CUMS mice;

[0019] Figure 4 The test results show that strain A21196 significantly shortens the forced swimming immobility time of CUMS mice;

[0020] Figure 5 The test results show that strain A21196 significantly increases the exploratory ability of CUMS mice in the open field;

[0021] Figure 6 The results showed that strain A21196 significantly increased the time that CUMS mice spent in the open arms;

[0022] Figure 7 The strain A21196 significantly increased the time CUMS mice spent exploring new objects;

[0023] Figure 8 The test results show that strain A21196 significantly increases the serum 5-HT level in mice;

[0024] Fig. 9 The strain A21196 significantly increased the level of BDNF in mouse serum;

[0025] Fig.10 The results show that strain A21196 significantly increases the BDNF level in the mouse hippocampus.

[0026] Biomaterial Deposit Information

[0027] Limosilactobacillusfermentum strain A21196 was deposited in the Guangdong Provincial Microbiological Culture Collection Center on November 11, 2022. The unit is abbreviated as GDMCC. The address is 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou. The deposit number is GDMCC No: 62959.

[0028] Lactobacillus plantarum strain A21337 was deposited in Guangdong Provincial Microbiological Culture Collection Center on May 23, 2024. The unit is abbreviated as GDMCC. The address is 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou. The deposit number is GDMCC No: 64677. DETAILED DESCRIPTION

[0029] The invention provides an application of a fermented Lactobacillus mucilaginosus strain A21196 in preparing a medicine for preventing and / or treating anxiety or depression. The deposit number of the fermented Lactobacillus mucilaginosus strain A21196 is GDMCC No: 62959.

[0030] In the present invention, the fermented mucus lactobacillus strain A21196 is isolated from a sample of intestinal microbial flora of centenarians. Gram staining shows that strain A21196 is a Gram-positive bacterium, the bacterial morphology is short rod-shaped, and the bacterial size (width×length) is (0.5~1.0) μm×(2.0~3.0) μm. 16S rDNA sequencing shows that strain A21196 belongs to fermented mucus lactobacillus.

[0031] In the present invention, the fermented mucus lactobacillus strain A21196 preferably includes at least one of the following cultures: the bacterial body of the fermented mucus lactobacillus strain A21196, the metabolites of the fermented mucus lactobacillus strain A21196 and the fermented liquid of the fermented mucus lactobacillus strain A21196. The preparation method of the fermented liquid of the fermented mucus lactobacillus strain A21196 preferably inoculates the activated fermented mucus lactobacillus strain A21196 in MRS liquid culture to obtain seed liquid; inoculates the seed liquid in MRS liquid culture for expansion culture, and inoculates the fermented mucus lactobacillus strain A21196 bacterial liquid obtained by the expansion culture in MRS liquid culture again for fermentation culture to obtain fermented liquid. The activation method of the strain A21196 is preferably that the strain A21196 is inoculated on an MRS plate for streak culture, and a single colony is picked. The temperature of the streak culture is preferably 36-38°C, more preferably 37°C. The inoculation amount during the streak culture is preferably 0.5%-2%, more preferably 1%. The method for preparing seed liquid of strain A21196 is preferably to inoculate the above-picked single colony into MRS liquid culture medium for cultivation to obtain seed liquid. The culture temperature is preferably 36-38°C, more preferably 37°C. The culture time is preferably 2-4h, more preferably 3h. During the culture, the inoculation amount is preferably 3%-6%, more preferably 4%-5%. The temperature of the expanded culture is 36-38°C, more preferably 37°C. The inoculation amount during the expanded culture is preferably 5%-10%, more preferably 8%. The present invention does not impose any special restrictions on the formula of the MRS liquid culture medium and the MRS plate, and the MRS liquid culture medium and the MRS plate formula known in the art can be used. For example, the MRS liquid culture medium formula is 20g / L glucose, 10g / L beef extract, 10g / L peptone, 5g / L yeast powder, 2g / L dipotassium hydrogen phosphate, 2g / L diammonium citrate, 5g / L sodium acetate, 0.58g / L magnesium sulfate, 0.25g / L manganese sulfate, Tween-800.1%, and the pH value is 6.6-6.8. The difference between the MRS plate and the MRS liquid culture medium is that 1.5%-2% agar is added additionally. The method for preparing the bacterial body or metabolite of the fermented mucus lactobacillus strain A21196 is to separate the fermented liquid prepared above into solid and liquid, collect the precipitate as the bacterial body of the fermented mucus lactobacillus strain A21196, and collect the liquid as the metabolite of the fermented mucus lactobacillus strain A21196.

[0032] In the present invention, the fermented mucus lactobacillus strain A21196 preferably also exists in the form of a probiotic. The probiotic refers to supplementing the beneficial bacteria in the intestine so that the beneficial bacteria have sufficient quantity and exert unique biological effects, thereby achieving the purpose of health care or disease prevention and treatment. The probiotic preferably includes at least one of the following: tablets, drops, capsules and powders. The powder is processed by low-temperature freeze-drying technology to ensure the activity of the probiotics. The auxiliary materials of the powder preferably include a freeze-drying agent, such as skimmed milk powder, etc. The powder is suitable for warm water and is suitable for infants. The tablet is prepared by mixing the fermented mucus lactobacillus strain A21196 and the auxiliary materials and pressing them into tablets, which are suitable for chewing and taking. The auxiliary materials preferably include sweeteners or cocoa powder, etc. The capsule preferably contains the fermented mucus lactobacillus strain A21196 alone or mixed with auxiliary materials as the capsule core in the capsule shell. The drops are prepared by mixing fermented mucus lactobacillus strain A21196 with edible oil to form drops that isolate air and moisture. When taken, the dropper port is opened to squeeze the encapsulated oily substance into the mouth. The live bacteria concentration of fermented mucus lactobacillus strain A21196 in the probiotic is preferably not less than 1×10 10 CFU / mL or 1×10 10 CFU / g, can be 5×10 10 CFU / mL~100×10 10 CFU / mL or 5×10 10 CFU / g~100×10 10 CFU / g, or 10×10 10 CFU / mL, 20×10 10 CFU / mL, 10×10 10 CFU / mL, 30×10 10 CFU / mL, 40×10 10 CFU / mL, 50×10 10 CFU / mL, 60×10 10 CFU / mL, 70×10 10 CFU / mL or 10×10 10 CFU / g, 20×10 10 CFU / g, 10×10 10 CFU / g, 30×10 10 CFU / g, 40×10 10 CFU / g, 50×10 10 CFU / g, 60×10 10 CFU / g, 70×10 10 CFU / g.

[0033] In the present invention, the anxiety disorder preferably includes at least one of the following anxiety behaviors and complications: acute anxiety, chronic anxiety, neurological dysfunction, cardiovascular and cerebrovascular diseases, and digestive system diseases. The acute anxiety is mainly manifested as acute panic attacks, and patients often suddenly feel anxious, panicked or excited, which leads to hallucinations and delusions, and sometimes mild confusion of consciousness. The acute anxiety attack can generally last for a few minutes or hours, and the course of the disease is generally not long, and it will gradually tend to ease after a period of time. Acute anxiety patients almost always have more or less sleep disorders, most of which are manifested as difficulty in falling asleep, easy to wake up after falling asleep, often accompanied by nightmares, restless when waking up, and feeling very scared after waking up. The chronic anxiety refers to the anxiety that can last for a long time, and its anxiety level also fluctuates from time to time. Chronic anxiety in the elderly is usually manifested as being more sensitive and irritable at ordinary times, and being upset and inattentive when there is something unsatisfactory in life, and sometimes sulking and losing temper. The neurological dysfunction includes dizziness, vertigo, insomnia, nightmares, insomnia at night or excessive sleepiness during the day, confusion of right and wrong, tension, oppression, cold sweats or a lot of sweat, fear of cold and wind. The cardiovascular and cerebrovascular diseases include the impact of anxiety on blood pressure. Patients with common cerebrovascular diseases in clinical practice are not nervous. Once anxious and nervous, their blood pressure rises, which leads to cerebral hemorrhage and brings great risks. The indigestion is that long-term anxiety can cause colitis, stomach pain, chronic gastritis, and patients often have loss of appetite and irritability. Some patients will overeat, causing symptoms such as gastrointestinal indigestion.

[0034] In one embodiment of the present invention, an open field test and an elevated plus maze test were carried out to evaluate the anti-anxiety behavior efficacy of fermented mucus lactobacillus strain A21196 or its preparation. The results of the open field test showed that compared with the model group, the time taken by mice in the strain A21196 group to enter the center of the open field was significantly increased. The results of the elevated plus maze test showed that compared with the model group, the number of times the mice in the strain A21196 group entered the open arm and the percentage of time in the open arm were significantly increased, and the time in the closed arm was significantly shortened. This shows that the fermented mucus lactobacillus strain A21196 has the effect of improving the anxiety behavior of model mice.

[0035] In the present invention, the depression includes single-episode depression, bipolar depression and persistent depression (chronic depression). The single-episode depression refers to a patient who experiences only one significant depressive episode, which usually lasts for several months to about a year and then enters a remission period. Symptoms may include: persistent low mood and depression, loss of interest and pleasure in daily activities, lack of energy, fatigue and physical exhaustion, sleep problems (such as insomnia or excessive sleep), decreased attention and concentration, reduced self-worth, self-blame and inferiority, negative thinking, negative concepts and pessimistic attitudes, and possible anxiety, guilt and despair. The bipolar depression, also known as manic depression, is a mood disorder in which patients alternate between depression and mania. Manic phase symptoms may include: high mood, excitement and impulse, excessive self-confidence, impulsive behavior and high-risk decision-making, distraction, easy distraction and multitasking, and possible delusions or hallucinations. The persistent depression (chronic depression) refers to a depressive symptom duration of up to two years or longer. Symptoms may be mild, but persist, affecting daily life and function. Common symptoms include: depressed mood, loss of interest and pleasure, decreased energy or fatigue, changes in appetite, decreased libido, sleep disorders (such as insomnia or excessive sleep), decreased attention and concentration, low self-esteem, and pessimism about the future.

[0036] In the present invention, a chronic unpredictable mild stress model of mice was established, and the antidepressant efficacy of fermented mucus lactobacillus strain A21196 was evaluated using sucrose preference test, tail suspension test, and forced swimming test. The results of the sucrose preference experiment showed that compared with the model group, the sugar water preference value of mice in the strain A21196 group was significantly increased, indicating that the strain A21196 described in the present invention has the function of improving the anhedonia of depressed mice. The results of the tail suspension experiment showed that compared with the model group, the immobility time of mice in the strain A21196 group was significantly shortened. At the same time, the results of the forced swimming experiment showed that compared with the model group, the immobility time of mice in the strain A21196 group was significantly shortened. It shows that strain A21196 can effectively alleviate the depressive-like behavior of model mice.

[0037] The invention provides an application of a fermented Lactobacillus mucosa strain A21196 in preparing a medicine for preventing and / or treating cognitive dysfunction and / or improving memory. The deposit number of the fermented Lactobacillus mucosa strain A21196 is GDMCC No: 62959.

[0038] In the present invention, the source of the fermented Lactobacillus mucosa strain A21196 and the type of probiotics are the same as those in the above technical solution, which will not be described in detail here.

[0039] In the present invention, the cognitive dysfunction preferably includes at least one of the following behavioral disorders: memory impairment, aphasia, visuospatial disorder, executive dysfunction, calculation disorder, apraxia and agnosia.

[0040] In one embodiment of the present invention, a novel object recognition experiment was conducted to evaluate the improvement effect of fermented Lactobacillus mucilaginosus strain A21196 on the cognitive function, especially memory, of model mice. The experimental results showed that compared with the model group, the percentage of time spent exploring new objects in the strain A21196 group of mice increased significantly, indicating that strain A21196 can effectively improve the cognitive behavior of model mice and improve the memory of mice.

[0041] In another embodiment of the present invention, in order to further explore the molecular mechanism of fermented mucus lactobacillus strain A21196 to play an anti-anxiety and improve cognitive ability and memory function, 5-hydroxytryptamine (5-HT) and brain-derived neurotrophic factor (BDNF) in the serum of mice in the model group and the drug-treated group were detected respectively. The results showed that compared with the model group, the 5-HT content of mice in the strain A21196 group was significantly increased. At the same time, compared with the model group, the BDNF content of mice in the strain A21196 group was significantly increased. 5-hydroxytryptamine (5-HT) is an important neurotransmitter in the body, widely distributed in the central nervous system, and is involved in regulating emotions, sleep, appetite and behavior. Brain-derived neurotrophic factor (BDNF) is a protein that promotes neuronal growth and survival, participates in cognitive processes such as synaptic plasticity, learning and memory, and plays an important role in the development and functional maintenance of the nervous system. 5-HT and BDNF synergistically regulate cognitive function. The present invention increases the levels of 5-HT and BDNF in model mice through strain A21196, so that 5-HT regulates the activity of neurons by binding to its receptor, thereby promoting the expression and secretion of BDNF, increasing cognitive processes such as anti-anxiety behavior and learning and memory, and enhancing neural network function.

[0042] The following is a detailed description of the application of a fermented Lactobacillus mucilaginosus strain A21196 provided by the present invention in the preparation of products for anti-anxiety and improving memory and concentration, in conjunction with the examples, but they should not be construed as limiting the scope of protection of the present invention.

[0043] Example 1

[0044] A preparation method of fermented Lactobacillus mucilaginosus strain A21196 probiotic agent

[0045] The frozen strain A21196 was inoculated on an MRS plate at a 1% inoculation rate and streaked at 37°C, and a single colony was picked to obtain an activated strain A21196. The single colony picked above was inoculated in an MRS liquid culture medium and cultured at 37°C for 2 hours to obtain a seed solution. The seed solution was inoculated in an MRS liquid culture medium at a 5% inoculation rate and expanded at 37°C to the logarithmic phase to obtain a bacterial solution containing the fermented mucus Lactobacillus strain A21196, and a 12% skim milk aqueous solution was added as a protective agent at a volume ratio of 1:1, and vacuum freeze-dried to obtain a probiotic containing the fermented mucus Lactobacillus strain A21196.

[0046] The MRS liquid culture medium contains 20 g / L glucose, 10 g / L beef extract, 10 g / L peptone, 5 g / L yeast powder, 2 g / L dipotassium hydrogen phosphate, 2 g / L diammonium citrate, 5 g / L sodium acetate, 0.58 g / L magnesium sulfate, 0.25 g / L manganese sulfate, and 0.1% Tween-80, and the pH value is 6.6 to 6.8. The difference between the MRS plate and the MRS liquid culture medium is that 1.5% to 2% agar is added additionally.

[0047] Example 2

[0048] 1. Establishment of a chronic unpredictable mild stress model in mice

[0049] 48 healthy 6-week-old SPF male C57BL / 6J mice, weighing 18-22g, were purchased from Sibeifu (Beijing) Biotechnology Co., Ltd., and the experimental animal production license is SCXK (Beijing) 2019-0010. The mice were kept in a constant temperature and humidity barrier environment (temperature 22±1℃, humidity 55±10%), with a light-dark alternation time of 12h:12h, and free food and water. After a week of adaptation, the mice were randomly grouped according to their weight, and the grouping is shown in Table 1. Every day, 1 to 3 different stimulations were used: restraint for 4h, crowding for 4h, tail clamping for 1min, ice water swimming for 5min, wet bedding for 24h, lighting all night for 24h, continuous darkness for 24h, no bedding for 24h, fasting for 24h, water deprivation for 24h, cage tilting for 24h, noise stimulation for 10min, and foreign body stimulation for 4h; the stimulation was irregular every day, and the model was continuously established for 4 weeks, and the mice were weighed once a week. After the modeling, the mice were subjected to behavioral tests to evaluate whether the modeling was successful. The antidepressant behavioral efficacy of the fermented Lactobacillus mucin strain A21196 was evaluated using the sucrose preference test (SPT), tail suspension test (TST), and forced swim test (FST). The open field test (OFT) and elevated plus maze test (EPM) were used to evaluate the anti-anxiety behavioral efficacy of the fermented Lactobacillus mucin strain A21196. The novel object recognition test (NOR) was used to evaluate the efficacy of the fermented Lactobacillus mucin strain A21196 in improving cognitive dysfunction and improving memory and concentration.

[0050] Table 1 Treatment methods of mice in each group

[0051] Group Number deal with Normal group 12 Daily intragastric administration of normal saline, 0.2 ml / rat CUMS Group 12 CUMS model was established, and normal saline was gavaged daily, 0.2 ml / rat A21196 Group 12 <![CDATA[CUMS model establishment, intragastric administration of A21196 at 0.2 ml / rat per day, 1×10 10 CFU / mL]]> A21337 Group 12 <![CDATA[Modeling by CUMS, intragastric administration of A21337 at 0.2 ml / rat per day, 1×10 10 CFU / mL]]>

[0052] 2. Mouse Weight Statistics

[0053] The mice were modeled according to the above modeling method, and the weight of the mice was counted once a week. The results are as follows: Figure 1 shown.

[0054] The experimental results show that ( Figure 1 As shown in Figure 2, compared with the normal group, the weight of mice in the CUMS model group decreased significantly (p < 0.0001). Compared with the model group, there was no significant change in the weight loss trend of mice after oral administration of A21196 or A21337, indicating that the strain A21196 and the control strain A21337 had no effect on the weight of mice.

[0055] 3. Sucrose preference experiment

[0056] Before the formal test, all mice were first trained with sugar water for 3 days, that is, they were given a bottle of drinking water containing 1% sucrose and a bottle of regular drinking water, and the positions of the two bottles were swapped from time to time. After the training period, all mice were fasted and deprived of water, and a formal sugar water test was conducted. Two bottles of drinking water containing 1% sucrose and regular drinking water of the same volume were prepared. After 24 hours, the sugar water consumption (g) of each mouse was counted, and the sugar water preference value was calculated according to Formula I:

[0057] Sugar water preference value = (mass of sugar water consumed / total mass of liquid consumed) × 100% Formula I.

[0058] The results are as follows Figure 2 As shown, compared with the normal group, the sugar preference value of mice in the CUMS model group was significantly reduced (p < 0.0001), indicating that the model was successfully constructed. In addition, after intragastric administration of A21196, the sugar preference value of mice was significantly increased (p < 0.0001), indicating that the strain A21196 of the present invention has the function of improving the anhedonia of depressed mice.

[0059] 4. Tail suspension test

[0060] The mouse tail was fixed with tape at the 1 / 3 of the back, and hung on a stand. The timing and video recording were started immediately, and the video recording was stopped after 6 minutes of observation. The immobility time of the mouse in the last 4 minutes was counted using Smart 3.0 analysis software. Immobility refers to the state in which the animal gives up active struggle and is completely immobile.

[0061] The results are as follows Figure 3 As shown, compared with the normal group, the immobility time of mice in the CUMS model group was significantly increased (p < 0.001), indicating that the model was successfully constructed. Compared with the model group, the immobility time of mice in the A21196 group was significantly shortened (p < 0.05), indicating that the A21196 strain can effectively alleviate the depressive-like behavior of mice caused by CUMS.

[0062] 5. Forced swimming test

[0063] The mice were placed in a transparent experimental bucket (filled with sterile water about 30 cm high) for a 6-minute swimming test. The Smart 3.0 analysis software was used for real-time video recording and analysis, and the immobility time of the mice in the last 4 minutes (immobility, the judgment standard was immobility of the limbs or only slight movement of the hind limbs) was calculated.

[0064] The results are as follows Figure 4As shown in the figure, compared with the normal group, the immobility time of mice in the CUMS model group was significantly increased (p < 0.0001), indicating that the model was successfully constructed. Compared with the model group, the immobility time of mice in the A21196 group was significantly shortened (p < 0.05), indicating that strain A21196 can effectively alleviate the depressive-like behavior of mice caused by CUMS.

[0065] 6. Open field experiment

[0066] The mouse was placed in a corner of the open field, and the timing and recording were started immediately. The recording was stopped after 10 minutes of observation. When replacing the animal, in order to avoid affecting the next test result, the surface needs to be wiped with 75% alcohol to remove the residual information of the previous animal (such as urine, feces, odor, etc.). Smart 3.0 analysis software was used for real-time recording and analysis, and the time (s) that the mouse stayed in the center area of ​​the open field was counted.

[0067] The results are as follows Figure 5 As shown, compared with the normal group, the time for mice in the CUMS model group to enter the center of the open field was significantly shortened (p < 0.01). Compared with the model group, the time for mice in the A21196 group to enter the center of the open field was significantly increased (p < 0.01), and the strain A21196 was more effective than the control strain A21337. This shows that the A21196 strain can effectively alleviate the anxiety behavior of mice accompanied by depression-like symptoms caused by CUMS.

[0068] 7. Elevated plus maze test

[0069] Place the mouse in the central area of ​​the elevated plus maze (the mouse faces the open arm), start timing and recording immediately, and stop recording after 5 minutes of observation. When replacing the animal, in order to avoid affecting the next test result, it is necessary to wipe the surface with 75% alcohol to remove the residual information of the previous animal (such as urine, feces, odor, etc.). Use Smart 3.0 analysis software for real-time recording and analysis, and count the number of times the mouse enters the open arm, and the percentage of time in the open arm and the closed arm (%).

[0070] The results are as follows Figure 6 As shown, compared with the normal group, the number of times the mice in the CUMS model group entered the open arm and the percentage of time in the open arm were significantly shortened (p < 0.05). Compared with the model group, the number of times the mice in the A21196 group entered the open arm and the percentage of time in the open arm were significantly increased (p < 0.0001), and the time in the closed arm was significantly shortened (p < 0.0001), and the strain A21196 was better than the control strain A21337. It shows that the A21196 strain can effectively alleviate the anxiety behavior of mice accompanied by depression-like symptoms caused by CUMS.

[0071] 8. New object recognition experiment

[0072] The novel object recognition test (NOR) is mainly used to evaluate the cognitive function, especially memory, of rodents (such as mice and rats). The core of this test is the difference in the animals' exploration behavior towards new and old objects, which is used to evaluate their memory and cognitive abilities.

[0073] Before the formal test, the mice were trained for 2 days. On the first day, the mice were allowed to freely explore and adapt in the open field for 10 minutes. On the second day, two identical objects A were placed in diagonal positions, and the mice were allowed to explore freely for 5 minutes. One hour later, one of the objects A was replaced with another new object B, and the mice were allowed to explore freely for 5 minutes. It should be noted that each time the animal is replaced, in order to avoid affecting the results of the next test, it is necessary to wipe the open field and the surface of the object with 75% alcohol to remove the residual information of the previous animal (such as urine, feces, odor, etc.). In the formal experiment, object A was retained in the open field, and object B was replaced with object C. Each mouse was placed in the same direction and allowed to explore freely for 5 minutes. Smart 3.0 analysis software was used for real-time video recording and analysis, and the percentage (%) of time the mouse explored the new object C was counted.

[0074] The results are as follows Figure 7 As shown in the figure, compared with the normal group, the percentage of time spent exploring new objects in the CUMS model group was significantly shortened (p < 0.01). Compared with the model group, the percentage of time spent exploring new objects in the A21196 group was significantly increased (p < 0.001), indicating that strain A21196 can effectively improve the cognitive behavior of CUMS depressed mice and improve the memory of mice.

[0075] Example 3

[0076] Detection of 5-HT levels in mouse serum

[0077] The mice were treated according to the method of Example 2, and appropriate amount of serum was taken from each group of mice and tested according to the instructions of the mouse 5-HT ELISA kit (provided by Jianglai Biotechnology).

[0078] The results of 5-HT detection in mouse serum are as follows Figure 8 As shown, compared with the normal group, the 5-HT level in the serum of the mice in the CUMS model group was significantly decreased (p<0.001). Compared with the model group, the 5-HT content in the mice in the strain A21196 group was significantly increased (p<0.001).

[0079] Example 4

[0080] Detection of BDNF levels in mouse serum and hippocampus

[0081] The model mice were treated with drugs according to the method of Example 2, and appropriate amounts of serum or hippocampal tissue were taken from each group of mice, and the detection was performed according to the instructions of the mouse BDNF ELISA kit (provided by Lianke Bio) and the total RNA extraction kit (provided by Promega Bio), the single-stranded synthesis kit (provided by Quanshijin), and the 2X SYBR Green qPCR MiX (provided by Si Kejie). The primers used for qPCR are shown in Table 2.

[0082] Table 2 Primers for qPCR detection

[0083]

[0084] The results of mouse serum BDNF detection are as follows Fig. 9 As shown, compared with the normal group, the BDNF level in the serum of mice in the CUMS model group was significantly decreased (p<0.0001). Compared with the model group, the BDNF content in mice in the strain A21196 group was significantly increased (p<0.05).

[0085] The results of BDNF detection in mouse hippocampus are as follows Fig.10 As shown in the figure, compared with the normal group, the BDNF level in the hippocampus of mice in the CUMS model group was significantly decreased (p < 0.001). Compared with the model group, the BDNF content in mice in the strain A21196 group was significantly increased (p < 0.001).

[0086] In summary, strain A21196 may alleviate the depressive behavior of mice by increasing the level of BDNF, and can effectively improve the cognitive behavior of CUMS depressed mice and improve the memory of mice. It is comparable to the control strain A21337 group.

[0087] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A use of a fermented Lactobacillus fermentum strain A21196 in the preparation of a medicament for preventing and / or treating anxiety or depression, wherein the deposit number of the fermented Lactobacillus fermentum strain A21196 is GDMCC No: 62959.

2. The use according to claim 1, characterized in that: The anxiety disorder includes at least one of the following anxiety behaviors and complications: acute anxiety, chronic anxiety, neurological dysfunction, cardiovascular and cerebrovascular diseases, and digestive system diseases.

3. Use of a fermented Lactobacillus fermentum strain A21196 in the preparation of a drug for preventing and / or treating cognitive dysfunction and / or improving memory, wherein the deposit number of the fermented Lactobacillus fermentum strain A21196 is GDMCC No: 62959.

4. The use according to claim 3, characterized in that: The cognitive dysfunction includes at least one of the following behavioral disorders: memory impairment, aphasia, visuospatial disorder, executive dysfunction, calculation disorder, apraxia and agnosia.

5. The use according to any one of claims 1 to 4, characterized in that: The fermented Lactobacillus mucilaginosus strain A21196 comprises at least one of the following cultures: bacterial bodies of the fermented Lactobacillus mucilaginosus strain A21196, metabolites of the fermented Lactobacillus mucilaginosus strain A21196, and fermentation liquid of the fermented Lactobacillus mucilaginosus strain A21196.

6. The use according to claim 5, characterized in that: The fermented Lactobacillus mucilaginosus strain A21196 also includes being present in the form of a probiotic.

7. The use according to claim 6, characterized in that: The probiotics include at least one of the following: tablets, drops, capsules and powders.

8. The use according to claim 6, characterized in that: The live bacteria concentration of the fermented Lactobacillus mucilaginosus strain A21196 in the probiotic is not less than 1×10 10 CFU / mL or 1×10 10 CFU / g.

9. The use according to claim 8, characterized in that: The live bacteria concentration of the fermented Lactobacillus mucilaginosus strain A21196 is 5×10 10 CFU / mL~100×10 10 CFU / mL or 5×10 10 CFU / g~100×10 10 CFU / g.

10. The use according to any one of claims 1 to 4, characterized in that: The drug comprises at least one of the following oral dosage forms: tablets, powders, capsules and oral liquids.

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