Preparation method and application of Lactobacillus paracasei IOB413 metagenic powder

Abstract

The invention provides a preparation method and application of casei paracasei IOB413 after-generation powder, and relates to the field of microorganisms. Tartary buckwheat is used as a solid-state fermentation culture medium, and the lactobacillus paracasei IOB413 is fermented and then crushed to prepare the lactobacillus paracasei IOB413 after-generation powder with high flavone content. The flavonoid content of the metagenic powder prepared by the method is far higher than that in tartary buckwheat and casei paracasei IOB413, the levels of tumor necrosis factor alpha (TNF-alpha) and interleukin (IL-6) can be reduced, the level of anti-inflammatory factor interleukin (IL-10) can be increased, the gingival index and the dental plaque index can be reduced, the growth of streptococcus mutans can be inhibited to a certain extent, and the metagenic powder can be used for preventing and treating the dental plaque. The oral micro-ecology is improved.

Description

Technical Field

[0001] The invention relates to the field of microorganisms, and in particular to a preparation method and application of Lactobacillus paracasei IOB413 postbiotic powder. Background Art

[0002] Oral health is an important part of overall health. It is not only related to basic functions such as chewing, swallowing and speech, but also closely related to our social interactions and psychological state. However, oral problems are common among the population. Common oral diseases include caries, gingivitis, periodontitis, oral ulcers, etc. Oral diseases not only cause pain, difficulty chewing and other problems, but may also cause systemic diseases such as cardiovascular disease, diabetes, respiratory diseases, etc. Therefore, maintaining oral health is of great significance for preventing oral diseases.

[0003] Flavonoids are a class of natural polyphenols found in a variety of plants. They have attracted much attention due to their wide range of biological activities, including anti-inflammatory, antibacterial, and antioxidant effects. In the field of oral health, these properties of flavonoids may be important for the prevention and treatment of oral diseases. Flavonoids help maintain oral health by inhibiting the growth of oral pathogens such as Streptococcus mutans and Porphyromonas gingivalis. These compounds can inhibit bacterial adhesion and biofilm formation, reduce bacterial virulence, and thus reduce the risk of dental caries and periodontal disease.

[0004] As a grain with high nutritional value, buckwheat contains a variety of nutrients that are beneficial to health, including protein, fat, vitamins and minerals. Of particular note, buckwheat also contains flavonoids, which may have a positive effect on oral health. The flavonoids in buckwheat may have a protective effect on oral health through their anti-inflammatory and antibacterial properties, helping to reduce oral inflammation and inhibiting the growth of harmful bacteria.

[0005] Probiotics are a type of microorganism that is beneficial to human health. They can prevent oral diseases by regulating the balance of oral microbiota. A clinical trial evaluated the adjuvant treatment effect of probiotic tablets containing Lactobacillus plantarum, Lactobacillus brevis and Lactobacillus acidophilus on gingivitis. The results showed that although there was no significant difference in the improvement of gingival index between the probiotic group and the control group, the use of probiotics reduced the number of severe inflammatory sites and improved the microbial status. Compared with conventional drugs, the advantages of probiotics in regulating oral health are: higher safety, as part of the normal human microbiota, usually no serious side effects at normal doses, and safer for special populations; more gentle and lasting mode of action, through interaction with oral microorganisms, continuous regulation of microecological balance, the effect is better than conventional chemical sterilization products; more significant preventive effect, focusing on preventing the occurrence of oral diseases, can maintain a good balance of oral microorganisms; also contribute to overall health, can affect the intestinal microbial community, have a positive effect on intestinal health, and indirectly protect systemic health, conventional drugs usually only target local oral problems.

[0006] Therefore, it is of great research value to find a flavonoid-rich probiotic to improve the oral environment. Summary of the invention

[0007] In view of the technical problems existing in the prior art, the present invention aims to provide a preparation method of postbiotic powder of Lactobacillus paracasei IOB413 and application thereof.

[0008] The classification of Lactobacillus paracasei IOB413 is named Lacticaseibacillus paracasei , isolated from naturally fermented sourdough, and deposited in the General Microbiology Center of China Microorganism Culture Collection (CGMCC) on June 29, 2018. The deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, and its deposit number is CGMCC No.16022.

[0009] One of the objects of the present invention is to provide a method for preparing postbiotic powder of Lactobacillus paracasei IOB413, the preparation method comprising: The secondary seed liquid of Lactobacillus paracasei IOB413 was solid-state fermented with tartary buckwheat as solid-state fermentation medium, and the fermented raw materials were dried and crushed to obtain Lactobacillus paracasei IOB413 postbiotic powder.

[0010] Preferably, during solid-state fermentation, the mass ratio of the secondary seed liquid to tartary buckwheat is 1:30-1:25.

[0011] Preferably, the solid culture medium material-water ratio is 1:1.75-1:1.25, and the inoculation amount is 1×10 7 ~5×10 7 cfu / mL, and the fermentation time was 22±2h.

[0012] Preferably, the method for preparing the secondary seed solution comprises: The IOB413 strain in the cryopreservation tube is inoculated into a slant culture medium and cultured at 35-39° C. for 24 hours, the strain is inoculated into an activation culture medium at a volume ratio of 1:30-1:20, and the culture is allowed to stand at 34-38° C. and sealed for 20-22 hours to obtain a first-level seed solution; the first-level seed solution is inoculated into an activation culture medium at a volume ratio of 1:30-1:20, and the culture is allowed to stand at 34-38° C. and sealed for 20-22 hours to obtain a second-level seed solution.

[0013] The second object of the present invention is to provide an application of postbiotic powder of Lactobacillus paracasei IOB413 in preparing medicines for improving oral environment.

[0014] The third object of the present invention is to provide an application of Lactobacillus paracasei IOB413 postbiotic powder in the preparation of mutans streptococcus inhibitors.

[0015] Beneficial effects of the present invention: The invention provides a method for preparing postbiotic powder of Lactobacillus paracasei IOB413, wherein tartary buckwheat is used as a solid fermentation medium, and Lactobacillus paracasei IOB413 is fermented and crushed to obtain postbiotic powder of Lactobacillus paracasei IOB413 with high flavonoid content. The flavonoid content of the postbiotic powder prepared by the method is much higher than that of tartary buckwheat and Lactobacillus paracasei IOB413, and can reduce the levels of tumor necrosis factor α (TNF-α) and interleukin (IL-6), increase the level of anti-inflammatory factor interleukin (IL-10), reduce the gingival index and dental plaque index, and can also inhibit the growth of Streptococcus mutans to a certain extent, and improve the oral microecology. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 is the flavonoid content; Figure 2 is the dental plaque index; Figure 3 is the gingival index; Figure 4 is the serum tumor necrosis factor α (TNF-α) level; Figure 5 is the serum interleukin-6 (IL-6) level; Figure 6 Serum interleukin 10 (IL-10) level.

[0017] The Lactobacillus paracasei IOB413 mentioned in the present invention is classified and named Lacticaseibacillus paracasei, isolated from naturally fermented sourdough, and deposited in the General Microbiology Center of China Microorganism Culture Collection (CGMCC) on June 29, 2018. The deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, and its deposit number is CGMCC No.16022. DETAILED DESCRIPTION

[0018] According to a first aspect of the present invention, a method for preparing postbiotic powder of Lactobacillus paracasei IOB413 is provided, wherein the preparation method comprises: The secondary seed liquid of Lactobacillus paracasei IOB413 was solid-state fermented with tartary buckwheat as solid-state fermentation medium, and the fermented raw materials were dried and crushed to obtain Lactobacillus paracasei IOB413 postbiotic powder.

[0019] In a preferred embodiment of the present invention, the mass ratio of the secondary seed liquid to tartary buckwheat during solid-state fermentation is 1:30-1:25.

[0020] When the mass ratio of the secondary seed liquid to the buckwheat culture medium is in the range of 1:30~1:25 during solid-state fermentation, the flavonoid content is further improved. Within this range, the supply and utilization of nutrients are further optimized. The secondary seed liquid contains a variety of nutrients required for microbial growth, which can make up for the nutritional deficiency of the buckwheat culture medium, adjust the carbon-nitrogen ratio, and provide a material basis for flavonoid synthesis; the growth environment is suitable, which promotes microbial growth and metabolism, and the active microbial cells in the seed liquid can enhance the overall activity, which is beneficial to flavonoid synthesis; the enzyme system is activated and a synergistic effect is produced, inducing the synthesis of more flavonoid-related enzymes, and at the same time different enzymes cooperate with each other to form a more efficient enzyme reaction system; In the present invention, when the mass ratio of secondary seed liquid and bitter buckwheat is lower than 1:35 during solid-state fermentation, the leavening agent is not enough to fully ferment bitter buckwheat, resulting in the flavonoids in bitter buckwheat failing to be fully released or converted, thereby affecting the improvement of flavonoid content. When the mass ratio of secondary seed liquid and bitter buckwheat is higher than 1:20 during solid-state fermentation, it causes waste of resources, and too much water will dilute the flavonoid content in bitter buckwheat, affecting the enzyme activity and microbial metabolism during the fermentation process, and then affecting the synthesis and accumulation of flavonoids. Therefore, the mass ratio of bacterial liquid to bitter buckwheat needs to be controlled within a suitable range to ensure the maximization of flavonoid content. Too high or too low mass ratio may be unfavorable for the improvement of flavonoid content.

[0021] In a preferred embodiment of the present invention, the solid culture medium material-water ratio is 1:1.75-1:1.25, and the inoculum size is 1×10 7 ~5×10 7 cfu / mL, and the fermentation time was 22±2h.

[0022] In a preferred embodiment of the present invention, the method for preparing the secondary seed solution comprises: The IOB413 strain in the cryopreservation tube is inoculated into a slant culture medium and cultured at 35-39° C. for 24 hours, the strain is inoculated into an activation culture medium at a volume ratio of 1:30-1:20, and the culture is allowed to stand at 34-38° C. and sealed for 20-22 hours to obtain a first-level seed solution; the first-level seed solution is inoculated into an activation culture medium at a volume ratio of 1:30-1:20, and the culture is allowed to stand at 34-38° C. and sealed for 20-22 hours to obtain a second-level seed solution.

[0023] According to a second aspect of the present invention, there is provided a use of postbiotic powder of Lactobacillus paracasei IOB413 in the preparation of a medicine for improving the oral environment.

[0024] According to a third aspect of the present invention, there is provided a use of postbiotic powder of Lactobacillus paracasei IOB413 in the preparation of a mutans Streptococcus inhibitor.

[0025] Example 1 Preparation of postbiotic powder and bacterial powder of Lactobacillus paracasei IOB413 1. Preparation of postbiotic powder (1) Activation and cultivation of strains: The IOB413 strain in the cryopreservation tube was inoculated into a slant culture medium and cultured at 37°C±2°C for 24 h. The strain was inoculated into an activation culture medium at a volume ratio of 1:30-1:20, and cultured at 36±2°C in a closed manner for 20-22 h to obtain a first-level seed solution. The first-level seed solution was inoculated into an activation culture medium at a volume ratio of 1:30-1:20, and cultured at 36±2°C in a closed manner for 20-22 h to obtain a second-level seed solution. The second-level seed solution was subjected to solid-state fermentation, and the mass ratio of the second-level seed solution to buckwheat was 1:25.

[0026] The solid fermentation medium was bitter buckwheat, the solid medium material-water ratio was 1:1.75-1:1.25, and the inoculation amount was 5×10 7 cfu / mL, and the fermentation time was 24h.

[0027] (2) Drying and crushing: Dry and crush the fermented raw materials. The drying temperature is 55-60℃, the crushing temperature is about -100℃, the main engine speed is 50Hz, the fan speed is 38Hz, the feed speed is 15Hz, the discharge speed is 25Hz, and the powder fineness is 300 to 500 mesh.

[0028] Among them, the components of the slant culture medium are: ingredients: 19g yeast powder, 10g glucose, 10g sucrose, 1ml Tween 80, 2g dipotassium hydrogen phosphate, 5g sodium acetate, 2g triammonium citrate, 0.2g magnesium sulfate, 0.05g manganese sulfate, 15g agar powder, and 1000ml distilled water.

[0029] The activation medium ingredients are: 25g yeast powder, 15g glucose, 10g sucrose, 1ml Tween 80, 2g dipotassium hydrogen phosphate, 5g sodium acetate, 2g triammonium citrate, 0.2g magnesium sulfate, 0.05g manganese sulfate, 15g agar powder, and 1000ml distilled water.

[0030] 2. Preparation of bacterial powder (1) Strain activation: Inoculate the strain into a liquid culture medium at a rate of 1%, and culture it at 37±2℃ in a closed state. Observe the colony morphology and bacterial characteristics at different culture times. The culture requires that the seeds be uniform, consistent in morphology, and strong. The seeds that have been cultured for 24 hours are more in line with the conditions, so the optimal culture time for seeds is determined to be 24 hours. The activated seed liquid is then inoculated into the activation culture medium at a rate of 3%, and the culture conditions are 37±2℃. Culture it in a closed state for 24 hours to obtain the fermented seed liquid. (2) Fermentation: The cultured secondary seed liquid was inoculated into the liquid culture medium at an inoculation rate of 3%. The fermentation tank was stirred at a speed of 50-70 r / min and cultured in a closed environment at 37°C for 12 h. During the fermentation process, food-grade sodium hydroxide was added to supplement the alkali to ensure that the pH of the fermentation liquid was maintained at 6.2±0.2. (3) Preparation of bacterial powder: After centrifugation of the fermentation medium, the bacterial sludge was obtained and vacuum freeze-dried to obtain the fermented mucus Lactobacillus IOB802 bacterial powder. The freeze-drying conditions were pre-freezing at -40°C for 6 hours, freeze-drying for 36±2 hours, and freeze-drying was completed when the vacuum degree dropped to 8.0Pa; Among them, the components of the activation culture medium are: 25g yeast powder, 15g glucose, 10g sucrose, 1ml Tween 80, 2g dipotassium hydrogen phosphate, 5g sodium acetate, 2g triammonium citrate, 0.2g magnesium sulfate, 0.05g manganese sulfate, 15g agar powder, and 1000ml distilled water.

[0031] The liquid culture medium composition is: 19g yeast powder, 10g glucose, 10g sucrose, 1ml Tween 80, 2g dipotassium hydrogen phosphate, 5g sodium acetate, 2g triammonium citrate, 0.2g magnesium sulfate, 0.05g manganese sulfate, and 1000ml distilled water.

[0032] Example 2 Rutin standard solution: Accurately weigh 40 mg of rutin standard that has been dried to constant weight, and add distilled water to prepare a 1.0 mg / mL rutin standard stock solution.

[0033] Flavonoid extraction: extraction temperature 40℃, extraction time 1.0h, ethanol concentration 60%, solid-liquid ratio 1:20, centrifugation at 10000r / min for 5min, and retain the supernatant.

[0034] Standard curve drawing: Take 7 10mL colorimetric tubes, add 0, 0.2, 0.4, 0.6, 0.8, 1.0, 1.2mL of 1.0mg / mL rutin standard stock solution, dilute to 2mL with 60% ethanol, then add 3mL of 60% ethanol, 0.3mL5% sodium nitrite, shake well, and let stand for 6min. Add 0.3mL10% aluminum nitrate, shake well, and let stand for 6min. Add 4.0mL of 4% sodium hydroxide, shake well, and let stand for 12min. Measure the absorbance at a wavelength of 510nm, and draw a standard curve with the rutin standard concentration value as the horizontal axis and the absorbance as the vertical axis.

[0035] Sample absorbance determination: Accurately pipette 2.0 mL of total flavonoids extract (diluted to an appropriate multiple) into a 10 mL colorimetric tube, add 3 mL of 60% ethanol and 0.3 mL of 5% sodium nitrite, shake well, and let stand for 6 minutes. Add 0.3 mL of 10% aluminum nitrate, shake well, and let stand for 6 minutes. Add 4.0 mL of 4% sodium hydroxide, shake well, and let stand for 12 minutes. Measure the absorbance at a wavelength of 510 nm, and substitute the absorbance into the standard curve to calculate the flavonoid content of the sample solution.

[0036] The calculation formula of flavonoid content is:

[0037] Where: C represents the corresponding mass concentration calculated by the standard curve, mg / mL; N represents the dilution factor; V represents the total volume of the extracted sample, mL; M represents the sample mass, g.

[0038] The results are as follows Figure 1 As shown, the flavonoid content in IOB413 powder is 0.02 mg / g, the flavonoid content in buckwheat is 132.25 mg / g, and the flavonoid content in the raw material after fermentation is 240.37 mg / g. The flavonoid content increases significantly after fermentation, which is higher than the sum of the contents in buckwheat and powder, indicating that the strain IOB413 can significantly increase the flavonoid content by fermenting with buckwheat as solid culture medium.

[0039] Example 3 Antibacterial experiment Streptococcus mutans is a Gram-positive bacterium that lives in the oral cavity and salivary glands of normal people. The toxins and proteases it produces can damage periodontal tissues, causing red, swollen and painful gums, even suppuration, and a foul odor.

[0040] Inoculate mutans Streptococcus in BHI liquid culture medium and culture at 30℃ for 12h; activate Lactobacillus paracasei IOB413 powder and postbiotic powder in MRS liquid culture medium. Take a small amount of bacterial solution and adjust the OD value of the bacterial solution to 1 with sterile water. Weigh 0.05g of Lactobacillus paracasei IOB413 powder and its postbiotic powder and dissolve them in sterile water. Use the Oxford cup method and filter paper method to perform antibacterial experiments: prepare a double-layer plate, the lower layer is MRS culture medium, and the upper layer is BHI culture medium containing mutans Streptococcus. For probiotics and their compound bacterial solutions, the Oxford cup method is used for antibacterial experiments, and for probiotic postbiotics and their compound postbiotic solutions, the filter paper cup method is used for antibacterial experiments. After completion, the plate is cultured at 30℃ for 48h.

[0041] Observe the growth of colonies on each plate. Record the size of the inhibition zone, which is the area around the Oxford cup where no colonies grow. Evaluate the antibacterial effect of the substance to be tested based on the size of the inhibition zone.

[0042] BHI liquid culture medium was purchased from Tianjin Liyuan Chemical Co., Ltd. MRS liquid culture medium was purchased from Qingdao Haibo Biotechnology Co., Ltd.

[0043] Streptococcus mutans can produce sticky polysaccharides to adhere to teeth to form dental plaque, metabolize sucrose to produce lactic acid to corrode tooth enamel, and is the main pathogen of dental caries. Therefore, the ability to inhibit the growth of Streptococcus mutans is the core characteristic of anti-caries pathogens. The diameter of the inhibition zone of Lactobacillus paracasei IOB413 is 32.62±1.52mm, and the diameter of the inhibition zone of Lactobacillus paracasei IOB413 postbiotics is 34.96±1.19mm. Both Lactobacillus paracasei IOB413 and its postbiotics have a certain inhibitory effect on Streptococcus mutans, among which the IOB413 postbiotic group has a better inhibitory effect and has a stronger ability to inhibit the growth of Streptococcus mutans. This is related to the production of flavonoids in the experimental group. Studies have shown that flavonoids can inhibit the growth of bacteria associated with dental caries and periodontal diseases, such as Streptococcus mutans and Streptococcus violaceus. They reduce the number of these bacteria through antibacterial, bactericidal and anti-biofilm activities, thereby helping to maintain oral health.

[0044] Example 4 Establishment and treatment of halitosis model in rats Thirty-two 4-week-old SPF Sprague Dawley rats were randomly divided into 4 groups after 1 week of adaptive feeding in the animal room, with 7-8 rats in each group. The grouping and treatment methods were as follows: (1) The blank control group rats were smeared with 200 μL 0.9% saline at the same time every day for 11 weeks. (2) The model group rats were smeared with 200 μL 3×10 6CFU / ml mutans Streptococcus solution was applied to the tooth roots of rats for 7 weeks. (3) Rats in the IOB413 group were first smeared with 200 μL of 3×10 6 CFU / ml mutans Streptococcus solution was applied to the tooth roots of rats for 7 weeks to complete the modeling. Then, 200 μL of 3×10 9 CFU / ml of Lactobacillus paracasei IOB413 solution was applied to the tooth roots of rats for 4 weeks. (4) Rats in the IOB413 postbiotic group were first smeared with 200 μL of 3×10 6 CFU / ml mutans Streptococcus solution was applied to the tooth roots of rats for 7 weeks to complete the modeling. 200 μL of 3×10 9 CFU / ml of Lactobacillus paracasei 413 postbiotic solution was applied to the tooth roots of rats for 4 weeks. The solution formulation of each group is shown in Table 1. The weight of rats in each group was roughly the same. The specific grouping method and application method are shown in Table 2.

[0045] Table 1 Preparation of solvents

[0046] Table 2 Grouping of rats

[0047] Example 5 Plaque index measurement Dental plaque is the main cause of gingivitis. It is a film-like substance composed of bacteria, saliva, food debris and dead cells that is constantly deposited on the teeth. Plaque and tartar can irritate the gums, causing them to become red, swollen and bleed easily. The plaque index is a measure of the accumulation of dental plaque and is a key indicator for evaluating oral hygiene and preventing periodontal disease. Flavonoids can inhibit the growth of many types of bacteria in dental plaque, including those associated with caries and periodontal disease. They work by destroying the bacterial cell membrane, inhibiting the activity of enzymes or interfering with the metabolic processes of bacteria.

[0048] Plaque index: 0 = no plaque; 1 = scattered plaque is present at the cervical margin of the tooth; 2 = a continuous thin band of plaque is visible at the cervical margin; 3 = The plaque zone at the cervical region is larger than 1 mm, but ≤ 1 / 3 of the tooth surface; 4 = plaque covers >1 / 3 and ≤2 / 3 of the tooth surface; 5=Plaque covers > 2 / 3 of the tooth surface.

[0049] The teeth were examined on the buccal and lingual surfaces, with a maximum of 10 counts for each tooth.

[0050] The evaluation of dental plaque index revealed that the oral health of rats in the IOB413 and IOB413 postbiotic groups was enhanced. Figure 2 As shown, the dental plaque index of rats was significantly reduced after treatment, among which IOB413 postbiotics had the most obvious effect. IOB413 postbiotics can produce a large amount of flavonoids, reduce the formation of dental plaque, improve oral health, and relieve gingival inflammation.

[0051] Example 6 Gingival index measurement The combination of the gingival index and the dental plaque index can effectively monitor the health of the gums, guide treatment and preventive measures, and is of great significance to the maintenance of oral health. The gingival index quantifies the degree of inflammation by observing the color, shape, and bleeding of the gums, which helps to identify early signs of periodontal disease. There is a direct relationship between gingivitis and the gingival index, and the gingival index can be used as a quantitative tool to assess the degree of gingivitis. Using the WHO-recommended CPI periodontal probe, check the mesial buccal, buccal, distal buccal and lingual surfaces of each tooth, score them separately, and divide the sum of the 4 scores by 4 to get the score of the tooth. The gingival index indicators are as follows: 0 = normal gingival appearance, no bleeding with sulcus probe; 1 = normal gingival appearance, but bleeding on sulcus probe; 2 = bleeding on probe, gingival color change but no swelling; 3 = bleeding on probe, gingival color change with mild swelling; 4 = bleeding on probe with marked gingival swelling, with or without color change; 5 = Bleeding on probe, spontaneous bleeding, gingival color change and obvious swelling, with or without ulceration.

[0052] The results are as follows Figure 3 As shown, the gingival index of the model group was significantly higher than that of the blank group, indicating that the rats' gums were significantly damaged and severely inflamed. After treatment with IOB413 and postbiotics, the gingival index showed a downward trend, which was consistent with the results of inflammatory factors and dental plaque. The IOB413 postbiotic group was able to significantly reduce the gingival index of the model group and had a significant improvement effect on gingival inflammation.

[0053] Example 7 Serum inflammatory factor level determination Measuring inflammatory factors is crucial for evaluating and monitoring the occurrence, development, and treatment effects of inflammatory diseases. Inflammatory factors are a class of proteins that play a key role in inflammatory responses, they can indicate or monitor the disease and its progression, and serve as clinically applicable parameters for treatment.

[0054] The serum of each group of rats in Example 4 was placed at room temperature for 2 hours, and the levels of tumor necrosis factor α, interleukin 6 and interleukin 10 were determined according to the instructions of the test kit. The levels of tumor necrosis factor α (TNF-α), interleukin (IL-6) 6 and interleukin 10 (IL-10) reflect the oral status of rats to a certain extent. The results of the determination of the levels of tumor necrosis factor α (TNF-α) in the serum of each group of rats are shown in FIG. Figure 4 As shown, the level of the model group increased compared with the blank group, indicating that the application of mutans streptococci caused an imbalance of oral bacteria in rats. Compared with the model group, the levels of tumor necrosis factor α (TNF-α) in the probiotic group and the postbiotic group decreased, and the effect of the postbiotic group was more obvious. This is because the IOB413 postbiotic produces more flavonoids, This study showed that compared with the blank control group, tumor necrosis factor α (TNF-α) in the model group would cause a certain degree of imbalance in oral health, accompanied by bleeding and swollen gums, while the intake of appropriate probiotics and postbiotics could effectively reduce the level of tumor necrosis factor α (TNF-α).

[0055] The results of IL-6 and IL-10 were as follows Figure 5 and Figure 6 As shown, the model group showed a significant increase in interleukin 6 (IL-6) and a significant decrease in IL-10 compared with the blank group, indicating that mutans streptococci caused an inflammatory response in the rats, and the rats had gingival bleeding, redness, and swelling accompanied by vesicles. The level of IL-6 in the IOB413 group and the IOB413 postbiotic group was lower than that in the model group, while the level of IL-10 was significantly increased in the model group. The effect of the postbiotic group was more significant, indicating that the oral health and gingival inflammation of rats were improved after treatment with IOB413 powder and postbiotics. The reason for the better effect of postbiotics is the increase in flavonoid content. Flavonoids can have a potential protective effect on the periodontal membrane and alveolar bone tissue by regulating the inflammatory response in the periodontal components, thereby further regulating the oral environment.

Claims

1. A method for preparing postbiotic powder of Lactobacillus paracasei IOB413, characterized in that, The preparation method comprises: The secondary seed liquid of Lactobacillus paracasei IOB413 was solid-state fermented with tartary buckwheat as solid-state fermentation medium, and the fermented raw materials were dried and crushed to obtain Lactobacillus paracasei IOB413 postbiotic powder.

2. The preparation method according to claim 1, characterized in that During the solid-state fermentation, the mass ratio of the secondary seed liquid to tartary buckwheat is 1:30-1:

25.

3. The preparation method according to claim 1, characterized in that: The solid culture medium material-water ratio is 1:1.75-1:1.25, and the inoculation amount is 1×10 7 ~5×10 7 cfu / mL, and the fermentation time was 22±2h.

4. The preparation method according to claim 1, characterized in that: The preparation method of the secondary seed solution comprises: The IOB413 strain in the cryopreservation tube was inoculated into the slant culture medium and cultured at 35-39°C for 24 hours. The strain was inoculated into the activation culture medium at a volume ratio of 1:30-1:20, allowed to stand at 34-38°C, and cultured in a closed manner for 20-22 hours to obtain the first-level seed solution. The first-level seed solution was inoculated into the activation culture medium at a volume ratio of 1:30-1:20, allowed to stand at 34-38°C, and cultured in a closed manner for 20-22 hours to obtain the second-level seed solution.

5. Application of the postbiotic powder of Lactobacillus paracasei IOB413 obtained by the preparation method as claimed in claim 1 in preparing medicines for improving oral environment.

6. Application of the postbiotic powder of Lactobacillus paracasei IOB413 obtained by the preparation method as claimed in claim 1 in preparing a mutans Streptococcus inhibitor.

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