Fermented lactobacillus mucus LF-HF25 with effects of dispelling effects of alcohol, protecting liver, nourishing stomach and promoting digestion, and after-biogen microbial inoculum, preparation method and application of fermented lactobacillus mucus LF-HF25 and after-biogen microbial inoculum

Abstract

The invention discloses a strain of lactobacillus fermentum LF-HF25 obtained by screening sour cucumber, which is preserved in the China Center for Type Culture Collection, the preservation number is CCTCC No.M20241566, the classification name is the lactobacillus fermentum LF-HF25, the preservation date is July 15, 2024, and the preservation address is Wuhan University, China. The lactobacillus fermentum LF-HF25 has the advantages that the preservation number is CCTCC No.M20241566, the classification name is the lactobacillus fermentum LF-HF25, the preservation date is July 15, 2024, and the preservation address is Wuhan University, China. The lactobacillus mucilaginosus LF-HF25 or the metabiotic microbial inoculum prepared from the lactobacillus mucilaginosus LF-HF25 has the functions of dispelling the effects of alcohol, protecting the liver, promoting digestion and nourishing the stomach, also has the effect of improving sleep, and has a wide application prospect.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of microorganisms, in particular to a fermented Limosilactobacillus LF-HF25 with the effects of protecting liver from alcohol, promoting digestion and nourishing stomach, a postbiotic agent, a preparation method and application. BACKGROUND

[0002] With the acceleration of modern social life pace and the change of dietary structure, health problems such as alcoholic liver disease and gastrointestinal dysfunction have become increasingly common. These problems not only affect people's quality of life, but also pose a serious threat to health. Globally, a large number of deaths are caused by alcohol abuse, and the prevalence of chronic gastritis and indigestion is high. The alcohol-protecting liver drugs on the market currently rely on chemical synthesis, although they can accelerate the metabolism of ethanol, but long-term use may cause gastrointestinal irritation and increase the burden on liver and kidney.

[0003] Probiotics have gradually become the focus of gastrointestinal health research because of their ability to regulate intestinal flora and enhance intestinal barrier function. However, the survival rate of traditional active probiotics in gastric acid and bile salt environments is usually low, and different strains have different colonization abilities and functions. In recent years, postbiotics have attracted attention due to their high stability, good safety, and the need to consider live bacteria survival. Studies have shown that certain postbiotics can improve gastrointestinal health by regulating immune response and repairing intestinal mucosal damage, but their application in alcohol protection and liver protection still needs further exploration and development.

[0004] Therefore, it is necessary to develop a multi-effect strain with efficient ethanol metabolism, gastrointestinal mucosa repair and digestion promotion functions. It has been found that the fermented Limosilactobacillus LF-HF25 and postbiotic agent prepared by the present application also have the effect of improving sleep. SUMMARY

[0005] Therefore, the purpose of the embodiments of the present application is to broaden the research and understanding of fermented Limosilactobacillus, and to provide its application in multiple fields. A fermented Limosilactobacillus LF-HF25 with the effects of protecting liver from alcohol, promoting digestion and nourishing stomach and a postbiotic agent thereof are obtained by screening a large number of acid cucumbers.

[0006] In a first aspect, the present application discloses a fermented Limosilactobacillus LF-HF25, which is preserved in the China Center for Type Culture Collection, has a preservation number of CCTCC No. M 20241566, is classified and named as Limosilactobacillus fermentum LF-HF25, was preserved on July 15, 2024, and is preserved in Wuhan University, China.

[0007] In a second aspect, the application discloses the fermented Lactobacillus muiis LF-HF25 and the postbiotic agent thereof in the application of protecting liver, nourishing stomach and promoting digestion.

[0008] In a third aspect, the application discloses the fermented Lactobacillus muiis LF-HF25 and the postbiotic agent thereof in the application of improving sleep.

[0009] In a fourth aspect, the application discloses the postbiotic agent prepared from the fermented Lactobacillus muiis LF-HF25.

[0010] In a fifth aspect, the application discloses a preparation method of the postbiotic agent of the fermented Lactobacillus muiis LF-HF25.

[0011] S1, strain activation: the fermented Lactobacillus muiis LF-HF25 is taken out from a glycerol tube, and is quickly thawed in a water bath; the thawed bacterial liquid is streaked and inoculated into a plate containing a first culture medium, and is placed in an anaerobic tank and cultured at 34-36 DEG C for 50-52 h, and a single colony with a diameter of 1-3 mm is selected.

[0012] S2, seed liquid preparation: the activated colony is inoculated into 80 mL of a second culture medium, and is statically cultured at 34-36 DEG C for 14-16 h to obtain a seed liquid.

[0013] S3, fermentation culture: the seed liquid is transferred into 1.5 L of a third culture medium, the temperature is adjusted to 34-36 DEG C, the pH is adjusted to 6.0-6.5, and the fermentation is cultured at 80 r / min for 48-75 h, and then the temperature is immediately lowered to 4 DEG C to terminate the fermentation.

[0014] S4, the bacterial liquid containing the bacterial liquid and the metabolic product is inactivated at 121 DEG C for 25 min, and then is concentrated to 1 / 8 of the original volume at a low temperature in a vacuum, and is freeze-dried to obtain the postbiotic.

[0015] In the application, the temperature of the water bath thawing in step S1 is 37 DEG C, and the time is ≤2 min.

[0016] In the application, the first culture medium comprises the following components: soybean peptone 12 g, beef extract 8 g, yeast extract 6 g, lactose 15 g, Tween-20 1.5 g, sodium citrate 3 g, potassium dihydrogen phosphate 3 g, ferrous sulfate 0.1 g, L-cysteine 1 g, agar 18 g, and distilled water 1000 mL.

[0017] In the embodiment of the present application, the second culture medium comprises the following components: soybean peptone 12 g; beef extract 8 g; yeast extract 6 g; lactose 15 g; Tween-20 1.5 g; sodium citrate 3 g; potassium dihydrogen phosphate 3 g; ferrous sulfate 0.1 g; L-cysteine 1 g; 15 g calcium carbonate; and distilled water 1000 mL. The calcium carbonate in the second culture medium serves to neutralize the acid produced during fermentation to stabilize the pH of the culture medium, and to provide calcium ions to maintain the stability of the cell membrane.

[0018] In the embodiment of the present application, the concentration of the seed liquid in step S2 is 1.2-3.5 x 10 8 CFU / mL.

[0019] In the embodiment of the present application, the concentration of the bacterial strain in the bacterial liquid obtained after the fermentation in step S3 is 2.0-4.5 x 10 10 CFU / mL.

[0020] In the embodiment of the present application, the inoculation amount of the bacterial strain in step S3 is 3-3.5% (v / v).

[0021] In the embodiment of the present application, the third culture medium comprises the following components: soybean peptone 12 g; beef extract 8 g; yeast extract 6 g; lactose 15 g; Tween-20 1.5 g; sodium citrate 3 g; potassium dihydrogen phosphate 3 g; ferrous sulfate 0.1 g; L-cysteine 1 g; 20 g maltose; and distilled water 1000 mL. The maltose in the third culture medium serves as an additional carbon source to provide energy and carbon atoms for the growth of the bacterial strain to promote proliferation, and to stimulate metabolic activity to enhance the synthesis and accumulation of metabolic products of the bacterial strain.

[0022] The present application has the following beneficial effects:

[0023] 1. The fermented L. m. LF-HF25 and the postbiotic bacterial agent prepared in the present application have the effects of alcohol elimination and liver protection.

[0024] 2. The fermented L. m. LF-HF25 and the postbiotic bacterial agent prepared in the present application have the effects of promoting digestion and nourishing the stomach.

[0025] 3. The fermented L. m. LF-HF25 and the postbiotic bacterial agent prepared in the present application have the effect of improving sleep.

[0026] 4. The fermented L. m. LF-HF25 and the postbiotic bacterial agent prepared in the present application are highly safe and suitable for a wide range of people. BRIEF DESCRIPTION OF DRAWINGS

[0027] Figure 1 is a comparison chart of AST indicators of each group in Experiment 2.

[0028] Figure 2 is a comparison chart of ALT indicators of each group in Experiment 2.

[0029] Figure 3 is a comparison chart of TC indicators of each group in experiment 2.

[0030] Figure 4 is a comparison chart of TG indicators of each group in experiment 2.

[0031] Figure 5 is a comparison chart of ink advancing rate of each group in experiment 3.

[0032] Figure 6 is a fermentation Lactobacillus mui LF-HF25 colony chart.

[0033] In order to more clearly illustrate the technical solutions in the embodiments of the present application, the following will briefly introduce the drawings needed to be used in the embodiments or prior art description. Obviously, the drawings in the following description are only some embodiments of the present application, and other drawings can also be obtained by those skilled in the art without creative labor. DETAILED DESCRIPTION

[0034] In order to make the purposes, technical solutions and advantages of the present application more clear, the present application will be further described in detail below with examples. It should be understood that the specific embodiments described herein are only used to explain the present application and not to limit the present application.

[0035] Example 1: Screening of Fermented Lactobacillus mui LF-HF25

[0036] The applicant has carried out a large number of screening from sour cucumbers, and has obtained several strains of fermented Lactobacillus mui with good health care function, among which the present application involves a strain named fermented Lactobacillus mui LF-HF25.

[0037] Identification of strain LF-HF25 with reference to the prior art

[0038] Morphological identification

[0039] The strain LF-HF25 is short rod-shaped or slightly curved, arranged singly, in pairs or short chains, and the ends of some bacterial bodies are slightly swollen but do not form bifurcated structures, without flagella and motility. The colony of the strain is usually mucous (see Figure 6 The optimum growth temperature thereof is 30-37℃.

[0040] Physiological and biochemical identification

[0041] According to the "Berger's Bacterial Identification Manual" and "Common Bacterial System Identification Manual", the physiological and biochemical indicators of the strain LF-HF25 were determined, and the specific results are shown in Table 1. According to the bacterial strain identification manual, it can be known that the strain LF-HF25 meets the Lactobacillus mui, and it can be preliminarily determined as fermented Lactobacillus mui.

[0042] Table 1 physiological and biochemical characteristics identification results of strain LF-HF25

[0043]

[0044]

[0045] Molecular biology identification

[0046] The 16S rDNA of strain LF-HF25 was amplified by PCR using bacterial universal primers 27F / 1492R. The amplification product was detected by agarose gel electrophoresis, showing a specific band of about 1500 bp, which was consistent with the expected size. The PCR amplification product was sent to Shanghai SunGene Biological Engineering Co., Ltd. for sequencing. The obtained sequence was analyzed by BLAST comparison in the Gen Bank database. Sequence alignment results showed that the 16S rDNA sequence of LF-HF25 had 99.2% homology with the standard strain ATCC-14931 of L. fermentum. Combined with the morphological characteristics and physiological and biochemical characteristics of the strain, it was determined that strain LF-HF25 was L. fermentum, which was preserved in the China Center for Type Culture Collection, with the preservation number CCTCC No. M 20241566, the classification name L. fermentum LF-HF25, the preservation date July 15, 2024, and the preservation address Wuhan University, Wuhan, China.

[0047] Example 2 Preparation of bioinoculant

[0048] S1, strain activation: L. fermentum LF-HF25 was taken out from the glycerol tube, quickly thawed in a 37°C water bath for no more than 2 min; the thawed bacterial solution was streaked onto a plate containing the first culture medium and placed in an anaerobic jar at 35°C for 51 h, and a single colony with typical morphological characteristics and a diameter of 1-3 mm was selected.

[0049] S2, seed liquid preparation: the activated colony was inoculated into 80 mL of the second culture medium and incubated at 35°C for 15 h to obtain a seed liquid with a concentration of 2.3 x 10 8 CFU / mL.

[0050] S3, fermentation culture: the seed liquid was inoculated into 1.5 L of the third culture medium at an inoculation amount of 3.2%, and incubated at 35°C, pH 6.2, and 80 r / min for 50 h. The temperature was immediately lowered to 4°C and the fermentation was stopped. The strain concentration in the bacterial solution obtained after fermentation was 2.2 x 10 10 CFU / mL.

[0051] S4, the bacteria liquid containing bacteria liquid and its metabolites was inactivated at 121℃ for 25min, and then concentrated to 1 / 8 of the original volume at low temperature and vacuum, and then the bacteria liquid was divided into freeze-drying trays, cooled to-50℃ at a rate of 1℃ / min and maintained for 1h; primary drying was performed, the cold trap was controlled at-70℃, the vacuum degree was 35Pa, the shelf temperature was increased from-35℃ to-20℃, and the process lasted for 20h; secondary drying was then performed, the shelf temperature was increased to 25℃, the vacuum degree was≤10Pa, and the process lasted for 7h until the moisture content was≤5%, thereby obtaining the probiotic bacteria agent.

[0052] Comparative Example 1: Fermented L. mesenteroides LF-HF25 was replaced by fermented L. mesenteroides ATCC-14931 (purchased from Gray Algae Biotechnology Co., Ltd.), and other conditions were the same as those in Example 1.

[0053] Experiment 1

[0054] Preparation of experimental animals: 40 SPF Kunming mice, half male and half female, weighing 20±2g, were provided by Beijing Vital River Laboratory Animal Technology Co., Ltd. The mice were adaptively fed in a light environment with a temperature of 23±2℃, a relative humidity of 55±10% and a 12h light-dark cycle for 7d. During the feeding period, the mice were free to eat.

[0055] Grouping: 40 Kunming mice were selected and randomly divided into 4 groups, namely, a blank control group, a model control group, an Example 1 group and a Comparative Example 1 group, 10 mice in each group. Before the experiment, all mice were fasted for 12h, and free water was provided during the period.

[0056] Modeling and administration: the mice in the blank control group were given 10mL / kg of normal saline by gavage, and the other three groups were given 10mL / kg of 52° Baijiu to establish an acute alcohol poisoning model. After 30min of alcohol gavage, the mice in the Example 1 group and the Comparative Example 1 group were given fermented L. mesenteroides LF-HF25 and fermented L. mesenteroides ATCC-14931 probiotic bacteria powder, respectively, 100mg / kg (gavaged after being dissolved in normal saline), once a day; the blank control group and the model control group were given an equal dose of normal saline, once a day.

[0057] Measurement of sober-up time: the sober-up time was determined by the recovery of the righting reflex. The mice were placed on their backs on a water platform. If the mice could not right themselves within 30s, it was determined that the righting reflex was lost and the mice were drunk. If the mice could right themselves within 30s, it was determined that the righting reflex was recovered and the mice were sober. The time from alcohol gavage to righting reflex recovery was recorded as the sober-up time. The entire observation process was carried out in a quiet and well-lit environment to reduce the influence of external interference on the experimental results. The experimental results are shown in Table 2.

[0058] Table 2

[0059] Group Awakening time / min Blank control group 0 (not drunk) Model control group 69.12±2.32 Example 1 group 34.84±2.15 Comparative example 1 group 51.62±3.29

[0060] Table 2 is a comparison table of the sober-up time of mice in each group. As can be seen from the figure, the blank control group did not consume alcohol, and the sober-up time was 0, which was the sober state and could be regarded as the normal state reference. The model control group had a sober-up time of 69.12±2.32 min, which was significantly increased compared with the blank control group, and more than 30 seconds was a deep drunk state. The sober-up time of the Example 1 group was 34.84±2.15 min, which was significantly shorter than that of the model control group, indicating that the intervention measures of this group could effectively accelerate sober-up. The sober-up time of the Comparative Example 1 group was 51.62±3.29 min, which was shorter than that of the model control group but longer than that of the Example 1 group. It can be seen that the Example 1 group is the most outstanding in shortening the sober-up time, and has the best sober-up effect, which confirms that the fermented Lactobacillus mucus LP-HF25 probiotic powder has the effect of alcoholism relief.

[0061] Experiment 2

[0062] Preparation of experimental animals: SPF Kunming mice, half male and half female, weighing 20±2 g, were provided by Beijing Vantoll Life-Science Experimental Animals Technology Co., Ltd. The mice were placed in a light environment with temperature of 23±2℃, relative humidity of 55±10% and 12h light-dark alternation for adaptive feeding for 7d. During the feeding period, the mice were free to eat.

[0063] Grouping: 40 Kunming mice were selected and randomly divided into 4 groups, namely blank control group, model control group, Example 1 group and Comparative Example 1 group, 10 mice in each group. Before the experiment, all mice were fasted for 12h, and free water was provided during the period.

[0064] The mice in the Example 1 group and the Comparative Example 1 group were respectively given fermented Lactobacillus mucus LF-HF25 and fermented Lactobacillus mucus ATCC-14931 probiotic powder by gavage, 100mg / kg (gavaged after being prepared with normal saline), once a day. The blank control group and the model control group were given the same dose of normal saline by gavage every day, 30min after each gavage, the blank control group was given 10mL / kg of normal saline, and the other three groups were given 10mL / kg of 52° Baijiu (Chinese liquor). The mouse chronic alcoholism model was established, and the body weight was measured every 5d for 6 weeks, and the gavage amount was adjusted according to the body weight change.

[0065] Determination of AST, ALT, TC and TG in serum: at the end of the 6th week of the experiment, the mice were weighed after fasting for 12h, and the blood was collected from the eye orbit and left for 30min. The serum was separated by centrifugation at 3000r / min for 15min. The content of ALT, AST, TC and TG in the serum was determined according to the instructions of the detection kit (ALT, AST, TC and TG kits were purchased from Shanghai Enzyme-linked Biological Co., Ltd.), and the experimental results are shown in Table 3. Figures 1-4 ​

[0066] Figure 1 is a comparison chart of AST indicators of each group. As can be seen from the chart, the AST level of Example 1 group is lower than that of model control group and Comparative Example 1 group, and the gap with the blank control group is relatively small. This indicates that Example 1 group has a better regulatory effect on the elevated AST caused by liver damage, and is closer to the normal level, suggesting that it has a protective effect on the liver at the AST-related level.

[0067] Figure 2 is a comparison chart of ALT indicators of each group. As can be seen from the chart, the ALT level of Example 1 group is lower than that of model control group and Comparative Example 1 group, and is closer to the blank control group. ALT is an important indicator of liver function, and its level is reduced, indicating that Example 1 group performs better in maintaining normal liver function and reducing liver damage.

[0068] Figure 3 is a comparison chart of TC indicators of each group. As can be seen from the chart, the TC content of Example 1 group is closer to the blank control group than the model control group and Comparative Example 1 group. This indicates that Example 1 group can effectively regulate liver cholesterol metabolism and reduce the impact of abnormal cholesterol deposition on the liver.

[0069] Figure 4 is a comparison chart of TG indicators of each group. As can be seen from the chart, the TG content of Example 1 group is closer to the blank control group than the model control group and Comparative Example 1 group. This indicates that Example 1 group can better regulate triglyceride metabolism and reduce liver fat synthesis and accumulation.

[0070] In summary, Example 1 group has the best liver protection effect, confirming that the fermented L. muceris LF-HF25 probiotic bacteria powder has a liver protection effect.

[0071] Experiment 3: Test of digestive function

[0072] Preparation of experimental animals: SPF Kunming mice, half male and half female, weighing 20±2g, provided by Beijing Vito Lihua Experimental Animal Technology Co., Ltd. The mice were placed in a temperature of 23±2℃, relative humidity of 55±10%, and 12h light-dark alternating light environment for 7d of adaptive feeding. During the feeding period, the mice were free to eat.

[0073] Forty mice were randomly divided into four groups (blank control group, model control group, Example 1 group and Comparative Example 1 group), 10 mice in each group. During the experiment, the Example 1 group and the Comparative Example 1 group were respectively given fermented Lactobacillus mucus LP-HF25 and fermented Lactobacillus mucus ATCC-14931 postbiotic powder of 100 mg / kg per day, and the blank control group and the model control group were given the same volume of normal saline, and the intervention lasted for 30 days. Before the end of the experiment, all animals were fasted for 16 h (free water), and then the model control group, the Example 1 group and the Comparative Example 1 group were given 5 mg / kg compound diphenoxylate by gavage (gavage volume 20 mL / kg). 30 min later, each dose group was given ink containing the corresponding test substance (containing 5% activated carbon powder and 10% gum arabic), and the blank control group and the model control group were only given ink without medicine (containing 5% activated carbon powder and 10% gum arabic). 25 min later, the animals were sacrificed, the small intestine from the pylorus to the ileocecal junction was separated, the total length of the small intestine and the propulsion distance of the ink front from the pylorus were measured, and the ink propulsion rate was calculated, and the experimental results are shown in Table 1. Figure 5

[0074] The ink propulsion rate = ink propulsion length / total length of small intestine x 100%.

[0075] Figure 5 is a comparison chart of the ink propulsion rate of each group. In the ink propulsion rate index for measuring the intestinal propulsion function, the model control group is much lower than the blank control group, showing that the model treatment damages the intestinal propulsion function. The ink propulsion rate of the Example 1 group is significantly higher than that of the model control group, and is close to that of the blank control group, while the Comparative Example 1 group is lower than the Example 1 group; at the same time, the fermented Lactobacillus mucus LF-HF25 postbiotic powder prepared by the present application can support the stomach function health care by promoting the efficiency of gastrointestinal digestion and reducing the burden on the stomach, thereby having the effect of nourishing the stomach. Therefore, the Example 1 group can effectively improve the intestinal propulsion function, and performs better than other groups, proving that the fermented Lactobacillus mucus LF-HF25 postbiotic powder has the effects of promoting digestion and nourishing the stomach.

[0076] Experiment 4

[0077] Preparation of experimental animals: SPF grade C57BL / 6J mice, half male and half female, weighing 20±2g, provided by Beijing Vital River Laboratory Animal Technology Co., Ltd. The mice were placed in a temperature of 23±2℃, relative humidity of 55±10% and 12h light-dark alternating light environment for adaptive feeding for 7d, and the mice were free to eat during feeding.

[0078] ​Fifty mice were randomly divided into a blank control group, a model control group, a positive control group (diazepam, 1.3 mg / kg), an Example 1 group, and a Comparative Example 1 group, with 10 mice in each group. Except for the blank control group, the other groups were given PCPA (300 mg / kg) by gavage for 3 consecutive days to establish an insomnia model. During modeling, the diazepam group was injected intraperitoneally with 0.2 mL of diazepam, the blank control group and the model control group were injected with an equal amount of 0.9% sodium chloride solution, and the Example 1 group and the Comparative Example 1 group were given fermented Lactobacillus muci laplantarum LF-HF25 and fermented Lactobacillus muci laplantarum ATCC-14931 postbiotic powder, respectively, at 100 mg / kg per day for 10 days.

[0079] After the experiment, all mice were injected intraperitoneally with pentobarbital sodium (4.5 mg / mL), and then placed flat on a flat plate to observe the sleep state. The sleep latency (the time from injection of pentobarbital sodium to the disappearance of the righting reflex) and sleep time (the time from the disappearance of the righting reflex to recovery) of the mice were recorded to evaluate the effect of different treatments on sleep.

[0080] Table 3

[0081] Group Sleep latency / min Sleep time / min Blank control group 3.23±0.16 37.23±5.72 Model control group 6.01±0.21 14.28±2.16 Positive control group 4.06±0.49 50.18±6.37 Example 1 group 4.69±0.27 29.42±3.26 Comparative example 1 group 5.76±0.41 15.66±2.39

[0082] Sleep latency refers to the time required from turning off the light and going to bed to falling asleep, and the shorter the time, the easier it is to fall asleep. The sleep latency of the blank control group was 3.23 ± 0.16 min, and that of the model control group was extended to 6.01 ± 0.21 min, indicating that the model treatment made it more difficult to fall asleep. The positive control group was 4.06 ± 0.49 min, the Example 1 group was 4.69 ± 0.27 min, and the Comparative Example 1 group was 5.76 ± 0.41 min. The sleep latency of the Example 1 group was significantly shorter than that of the model control group and the Comparative Example 1 group, although it was slightly longer than that of the positive control group, but the difference was not large, indicating that the Example 1 group could effectively shorten the sleep time and had a better effect on promoting sleep.

[0083] In terms of sleep time, the blank control group was 37.23 ± 5.72 min, the model control group was only 14.28 ± 2.16 min, showing that the model treatment significantly shortened the sleep time. The positive control group reached 50.18 ± 6.37 min, the Example 1 group was 29.42 ± 3.26 min, and the Comparative Example 1 group was 15.66 ± 2.39 min. The sleep time of the Example 1 group was significantly longer than that of the model control group and the Comparative Example 1 group, although it was not as good as the positive control group, but it also significantly prolonged the sleep time.

[0084] Comprehensive sleep latency and sleep time two indicators, the implementation of example 1 group in the model control group and the comparative example 1 group advantage compared to the obvious, although compared with the positive control group has a gap, but the overall effect is outstanding, shows that the implementation of example 1 group in improving sleep effect best, confirmed the fermentation of mucus lactobacillus LF-HF25 after the effect of biofermentation bacteria powder has improved sleep.

[0085] It is speculated that the post-fermentation regulates the key pathway of the gut-brain axis, promotes tryptophan metabolism and 5-hydroxytryptamine synthesis. The short-chain fatty acids contained in it can enhance the intestinal barrier function, reduce the competitive consumption of tryptophan, and activate the synthesis of 5-HT by intestinal chromaffin cells, thereby increasing the level of 5-HT in the brain through blood circulation, thereby shortening the sleep latency. In addition, the post-fermentation may reduce the excitability of the nervous system by inhibiting inflammatory factors, prolong the slow wave sleep time, and ultimately achieve the dual improvement of sleep efficiency and sleep duration.

[0086] The above is only the preferred specific embodiment of the present application, but the protection scope of the present application is not limited to this. Any person skilled in the art can easily think of changes or replacements within the technical scope disclosed in the present application, which should be covered within the protection scope of the present application.

Claims

1. A strain of Lactobacillus muci genis LF-HF25, characterized by: The strain is preserved in China Center for Type Culture Collection, with the preservation number CCTCC No. M 20241566, the preservation date of July 15, 2024, and the preservation address of Wuhan University, China.

2. Use of the fermented Lactobacillus muiis LF-HF25 or the prepared probiotic agent thereof in preparation of a preparation for protecting liver, promoting digestion and improving sleep.

3. Use of the fermented Lactobacillus muiis LF-HF25 or the prepared probiotic agent thereof in preparation of a preparation for improving sleep.

4. The probiotic agent prepared from the fermented Lactobacillus muiis LF-HF25.

5. The method of preparing the fermentable L. m. LF-HF25 probiotic bioinoculant according to claim 4, characterized in that: The specific steps are as follows: S1, strain activation: the fermented Lactobacillus muiis LF-HF25 is taken out from the glycerol tube, quickly thawed in a 37℃ water bath; the bacterial liquid is streaked into a plate containing the first culture medium, placed in an anaerobic tank, and cultured at 34-36℃ for 50-52h; 1-3mm single colonies with typical morphological characteristics are selected for standby; S2, seed liquid preparation: the activated colonies are inoculated into 80mL of the second culture medium, and cultured at 34-36℃ for 14-16h to obtain the seed liquid for standby; S3, fermentation culture: the seed liquid is inoculated into 1.5L of the third culture medium, and cultured at 34-36℃ and pH 6.0-6.5 for 48-75h; after the end, the temperature is immediately lowered to 4℃ and the fermentation is stopped for standby; S4, the bacterial liquid containing the bacterial liquid and its metabolites prepared in step S3 is inactivated at 121℃ for 25min, concentrated, freeze-dried to obtain the probiotic.

6. The method of claim 5, wherein: The first culture medium comprises the following components: soybean peptone 12g; beef extract 8g; yeast extract 6g; lactose 15g; Tween-20 1.5g; sodium citrate 3g; potassium dihydrogen phosphate 3g; ferrous sulfate 0.1g; L-cysteine 1g; agar 18g; distilled water 1000mL. The second culture medium comprises the following components: soybean peptone 12g; beef extract 8g; yeast extract 6g; lactose 15g; Tween-20 1.5g; sodium citrate 3g; potassium dihydrogen phosphate 3g; ferrous sulfate 0.1g; L-cysteine 1g; 15g calcium carbonate; distilled water 1000mL.

7. The method of claim 5, wherein: The inoculation amount of the strain in step S3 is 3-3.5%; the third culture medium comprises the following components: soybean peptone 12g; beef extract 8g; yeast extract 6g; lactose 15g; Tween-20 1.5g; sodium citrate 3g; potassium dihydrogen phosphate 3g; ferrous sulfate 0.1g; L-cysteine 1g; 20g maltose; distilled water 1000mL.

8. The method of claim 5, wherein: ​ 9. The preparation method according to claim 5, wherein the seed liquid concentration in step S2 is 1.2-3.5 x 10 8 CFU / mL.

10. The preparation method according to claim 5, wherein the concentration of the strain in the bacterial solution obtained after the fermentation is completed in step S3 is 2.0-4.5 x 10 10 CFU / mL.

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