Pediococcus pentosaceus and application thereof in accelerating metabolism of alcohol substances

Abstract

The invention provides pediococcus pentosaceus and application thereof in accelerating metabolism of alcohol substances. The strain can absorb and utilize alcohol substances, especially ethanol, n-propanol, isobutanol and isoamyl alcohol, and especially ethanol dehydrogenase and acetaldehyde dehydrogenase have high enzyme activity; the strain has very strong tolerance to cholate, artificial gastric juice and artificial intestinal juice, and has good self-aggregation property; a good colonization effect is achieved in the intestinal tract of the zebra fish; and the compound has relatively strong inhibition capability on escherichia coli, staphylococcus aureus and pseudomonas aeruginosa. The bacterium has a remarkable application value in improving intestinal flora and dispelling the effects of alcohol in intestinal probiotics.

Description

Technical Field

[0001] The present invention relates to the technical field of probiotics, and in particular to a strain of Pediococcus pentosaceus LTPP-271 and application thereof in accelerating the metabolism of alcohol substances. Background Art

[0002] Alcohol consumption is a leading cause of death and disease worldwide. Excessive drinking can lead to cardiovascular and cerebrovascular diseases, metabolic disorders, and is even closely linked to certain cancers. After drinking, approximately 10-20% of the alcohol that enters the stomach penetrates the gastric mucosa and enters the bloodstream through the capillaries. The alcohol then enters the small intestine through the pylorus. Due to the small intestine's inherent absorptive capacity and its larger surface area, approximately 80-90% of the alcohol is absorbed in the small intestine.

[0003] Alcohol metabolism occurs in three steps. Alcohol is converted to acetaldehyde by the enzyme alcohol dehydrogenase, which then converts it to acetic acid, which is then converted to carbon dioxide and water. Acetaldehyde, a Class I carcinogen designated by the World Health Organization, is broken down by alcohol into acetaldehyde, which poses serious risks to the human body. People deficient in acetaldehyde dehydrogenase experience symptoms of acetaldehyde poisoning, such as flushing and a rapid heartbeat. Studies have shown that ethanol and its homologues, such as n-propanol, isobutanol, and isopentanol, found in liquor, can exacerbate hangover symptoms by stimulating oxidative stress and promoting inflammation.

[0004] Probiotics can express alcohol dehydrogenase and acetaldehyde dehydrogenase in the intestines, thereby enabling the digestion and absorption of ethanol and effectively reducing alcohol damage to the liver and intestines. Pediococcus pentosaceus, a Gram-positive bacterium belonging to the Streptococcus family and the genus Pediococcus, is widely distributed in malt wort and fermented products such as pickles, sauerkraut, and cheese. It is approved as a new food ingredient and is one of the available food bacteria species. By screening for Pediococcus pentosaceus with alcohol-degrading properties, it is possible to degrade alcohol in the intestines, effectively reducing the burden of alcohol on the liver. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to provide a novel Pediococcus pentosaceus strain having the function of accelerating the metabolism of alcohol substances. The strain can absorb and utilize ethanol in an MRS culture medium containing 3% ethanol (v / v); it also has good absorption and utilization effects on n-propanol, isobutanol, and isopentanol; the enzyme activities of alcohol dehydrogenase and acetaldehyde dehydrogenase are both high, which has advantages in accelerating the alcohol metabolism function in the human body; at the same time, the probiotic has good tolerance to bile salts, artificial gastric juice, and artificial intestinal juice; it has strong inhibitory ability against Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa, and has significant application value in regulating intestinal health. Compared with other Pediococcus pentosaceus strains on the market, the strain has a better colonization effect in the zebrafish intestine.

[0006] The present invention provides a novel strain of Pediococcus pentosaceus, which is Pediococcus pentosaceus LTPP-271 was screened from homemade pickles made by farmers in Zunyi City, Guizhou Province. The fungus was deposited in the Guangdong Provincial Microbiological Culture Collection on June 4, 2024. The collection address is 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou City, and the collection number is GDMCC No. 64724.

[0007] Furthermore, the 16s nucleotide sequence of the Pediococcus pentosaceus LTPP-271 is shown in SEQ ID NO.1.

[0008] Furthermore, the Pediococcus pentosaceus LTPP-271 is not hemolytic.

[0009] Furthermore, the survival rate of the Pediococcus pentosaceus LTPP-271 in simulated artificial gastric fluid was 100% after 3 hours; the survival rate exceeded 90% after 3 hours in simulated artificial intestinal fluid, and the survival rate exceeded 75% after 24 hours; and the survival rates exceeded 65%, 40% and 25% after culturing for 24 hours under conditions containing 0.1%, 0.2% and 0.3% bile salts, respectively.

[0010] Furthermore, the self-aggregation rate of the Pediococcus pentosaceus LTPP-271 in sterile PBS for 24 hours exceeds 55%; Furthermore, the Pediococcus pentosaceus LTPP-271 can colonize the zebrafish intestine for more than 48 hours.

[0011] Furthermore, the Pediococcus pentosaceus LTPP-271 has a good antibacterial effect on Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa, with the diameters of the inhibition zones being 11.6±0.1 mm, 13.8±0.1 mm, and 14.2±0.2 mm, respectively.

[0012] The present invention also provides a microbial preparation of the Pediococcus pentosaceus LTPP-271.

[0013] The present invention also provides a fermentation production method of the above-mentioned Pediococcus pentosaceus, which comprises: The above-mentioned Pediococcus pentosaceus LTPP-271 was inoculated into the culture medium at an inoculum size of 2%-5%, and cultured at 37° C. for 16 h-24 h to obtain a culture solution. The culture solution was centrifuged and freeze-dried to obtain bacterial cells.

[0014] In one embodiment of the present invention, the culture medium is MRS culture medium or a mixed carbon source obtained by adding electrochemically catalyzed CO2 to replace part of the glucose in the MRS culture medium.

[0015] Furthermore, the above-mentioned electrochemical mixed carbon source is a mixture of one-carbon and two-carbon products obtained by electrocatalytic CO2 studied by the applicant of the present invention, which contains at least ethanol, formate and acetate, such as the mixed carbon source obtained by the preparation method mentioned in patent application No. CN202211251789.9. According to the added amount of ethanol, formate and acetate in the designed and verified mixed carbon source, if the mixed carbon source obtained by electrocatalytic carbon dioxide exhaust does not meet the concentration and ratio requirements, the corresponding ethanol, formate or acetate is added to meet the fermentation culture requirements of Pediococcus pentosaceus.

[0016] The concentration of ethanol in the fermentation broth is preferably 1%-5% (v / v), the concentration of formate in the fermentation broth is preferably 2-10 g / L, and the concentration of acetate in the fermentation broth is preferably 5-15 g / L. The inventors have experimentally verified that the environment in which microorganisms grow is highly variable and uncertain, and the use of mixed carbon sources can reduce product inhibition under a single carbon source while improving carbon source utilization. At the same time, the use of mixed carbon sources enables microorganisms to grow rapidly under low substrate concentrations, with a higher specific growth rate than on any single substrate, making it easier for microorganisms to cope with malnutrition and highly variable living environments.

[0017] Furthermore, after replacing 50% of the glucose in the culture medium with the electrochemically mixed carbon source, the effective viable count of Pediococcus pentosaceus LTPP-271 was 4.91×10 9 ± 1.63×10 8 CFU / mL, and the effective viable bacteria count in the MRS medium of the control group was 3.23×10 9 ± 1.91×10 8 CFU / mL, the effective viable bacterial count increased by 52.1% after the electrochemical mixed carbon source replaced 50% of the glucose in the culture medium.

[0018] The present invention also provides a microbial preparation comprising the above-mentioned Pediococcus pentosaceus LTPP-271, wherein the viable cell count of the above-mentioned Pediococcus pentosaceus LTPP-271 is not less than 1×10 6 CFU / mL or 1×10 6 CFU / g.

[0019] The microbial preparation may also contain other probiotics, such as Lactobacillus plantarum and Lactobacillus rhamnosus, and may also contain other auxiliary ingredients, such as vitamins, minerals, etc.

[0020] The present invention also provides a health product, food or medicine having the function of accelerating the metabolism of alcohol substances, which comprises the above-mentioned microbial preparation.

[0021] The dosage form of the health care product, food or medicine is preferably tablets, capsules, granules, oral liquid and the like.

[0022] The health care product, food or medicine may also contain other nutritional ingredients, such as turmeric powder, kudzu root powder and the like.

[0023] The health care product, food or medicine further comprises a carrier.

[0024] The beneficial effects of the present invention are as follows: the present invention provides a strain of Pediococcus pentosaceus LTPP-271 having the function of accelerating the metabolism of alcohol substances. When cultured in an MRS medium containing 3% (v / v) ethanol, the strain can absorb and utilize 64.6% of the ethanol in the medium, and the utilization rates of n-propanol, isobutanol, and isopentanol are 58.52%, 38.72%, and 46.30%, respectively. The alcohol dehydrogenase activity after fermentation is 18.38 U / mL, and the acetaldehyde dehydrogenase activity is 48.16 U / mL. The strain has good tolerance to artificial gastric juice, artificial intestinal juice, and bile salts: the survival rate in simulated artificial gastric juice is 100% after 3 hours; the survival rate after 3 hours in simulated artificial intestinal juice is 92.2%±3.2%, and the survival rate after 24 hours is 78.6%±2.4%; the strain can be cultured for 24 hours under conditions containing 0.1%, 0.2%, and 0.3% bile salts. h, with survival rates of 68.8%±2.6%, 41.3%±2.3% and 26.8%±2.8% respectively; it has good self-aggregation effect: the self-aggregation rate in sterile PBS for 24 h is 60.4%±6.5%; it can colonize in the zebrafish intestine for more than 48 h; it has strong inhibitory ability against Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa, and this bacterium has significant application value in regulating intestinal health and in the direction of intestinal probiotics for alcohol relief. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 Ethanol utilization standard curve; Figure 2 Microscopic examination results of Pediococcus pentosaceus LTPP-271 of the present invention; Figure 3 Columbia blood plate streak results of Pediococcus pentosaceus LTPP-271 of the present invention; Figure 4 Fluorescence image of Pediococcus pentosaceus LTPP-271 in the zebrafish intestine; Figure 5 Fluorescence value results of Pediococcus pentosaceus LTPP-271 in the zebrafish intestine; Figure 6 Fluorescence image of Pediococcus pentosaceus BNCC193259 in the zebrafish intestine; Figure 7 Fluorescence value results of Pediococcus pentosaceus BNCC193259 in the zebrafish intestine; Figure 8 The effective viable bacteria count of Pediococcus pentosaceus LTPP-271 after adding an electrochemical mixed carbon source. DETAILED DESCRIPTION

[0026] The following embodiments and drawings are used to describe the embodiments of the present invention in detail, so that the implementation process of how the present invention applies technical means to solve technical problems and achieve technical effects can be fully understood and implemented accordingly.

[0027] Example 1 Screening and Identification of Pediococcus Pentosaceus Using Ethanol 1.1 Screening, isolation and purification of strains Thirty samples of homemade kimchi were collected from farmers across the country. 1 mL of kimchi water from each sample was diluted and mixed in a test tube containing 9 mL of sterile saline (0.85%, %w / v). The dilution gradient was recorded as 10 -1 , and then draw 1 mL into a test tube containing 9 mL of sterile saline to dilute and mix, the dilution gradient is recorded as 10 -2 , and then dilute the kimchi water to 10 -6 The kimchi water solution and the aforementioned dilutions were spread onto MRS screening medium plates containing CaCO₃ and incubated at 37°C in an anaerobic workstation for 48 hours. Single colonies with distinct morphology, size, and color and a distinct calcium-dissolving zone were selected from the plates and streaked onto MRS plates to obtain single colonies. The resulting single colonies were subjected to Gram stain microscopy and catalase testing. Gram-positive, catalase-negative isolates were preliminarily identified as lactic acid bacteria, numbered, and stored for future use.

[0028] MRS medium components include: peptone 10 g, yeast extract 5 g, beef extract 10 g, glucose 20 g, sodium acetate 5 g, diammonium citrate 1.5 g, Tween 80 1 mL, magnesium sulfate 0.5 g, manganese sulfate 0.05 g, and potassium dihydrogen phosphate 2 g. The volume is adjusted to 1 L. For solid medium, add 15-20 g agar powder. CaCO3 should be added to the screening medium at a rate of 1%-2% (w / v).

[0029] 1.2 In vitro ethanol utilization assay The above lactic acid bacteria were cultured in MRS medium containing 3% and 5% (v / v) ethanol, and the OD 600 Strains with no significant decrease in ethanol utilization were tested. The ethanol content in the fermentation broth was determined using the potassium dichromate colorimetric method. Finally, a lactic acid bacteria strain, designated LTPP-271, was isolated from homemade pickles from farmers in Zunyi, Guizhou Province, demonstrating an ethanol utilization efficiency of 64.6% in 3% ethanol. The specific testing method is as follows: Ethanol standard curve determination: Prepare ethanol standard solutions with concentrations of 0.0%, 0.4%, 0.8%, 1.2%, 1.6%, and 2.0% by volume. Add 0.3 mL of the ethanol standard solution and 3 mL of the acidic potassium dichromate solution to a 15 mm x 15 cm test tube. Add a stopper to the test tube and incubate in a 100°C water bath for 15 minutes. Cool to room temperature with running water and measure the absorbance at 590 nm. Figure 1 , ethanol has a good linear relationship with absorbance in the concentration range of 0.0% to 2%, and the linear regression equation is y=0.3903x+0.0024, where x represents the ethanol concentration and y is the absorbance value.

[0030] In vitro ethanol utilization test of LTPP-271: The seed liquid of Pediococcus pentosaceus LTPP-271 cultured overnight was inoculated into MRS, MRS + 3% ethanol, and MRS + 5% ethanol culture media at an inoculum size of 2%. MRS, MRS + 3% ethanol, and MRS + 5% ethanol blank culture media without bacterial liquid were used as controls.

[0031] The above six groups of shake flasks, each with three biological parallels, were cultured for 20 hours and then centrifuged at 12,000 rpm for 10 minutes. The supernatant after centrifugation was diluted 6 times with deionized water for later use. The ethanol utilization test method is the same as the sample test method in the standard curve test. Because the organic carbon source in the MRS culture medium will also produce a color reaction with the potassium dichromate solution, the absorbance corresponding to the ethanol concentration needs to be subtracted from the absorbance of the MRS blank control group. The formula for calculating the ethanol utilization concentration is: Ethanol utilization concentration (%, v / v) = [(OD 590 I-OD 590 II)-0.0024] / 0.3903×dilution factor, where OD 590 I = MRS supplemented with ethanol blank group - MRS supplemented with ethanol inoculated group; OD 590 II = MRS blank group-MRS inoculated bacteria group.

[0032] The final test results are shown in Table 1. It can be seen that when 3% ethanol is added, the ethanol utilization rate of LTPP-271 in the culture medium is 64.6%. In the MRS culture medium supplemented with 5% ethanol, the ethanol utilization rate of LTPP-271 in the culture medium is 45.0%.

[0033] Table 1 Ethanol concentration corresponding to OD measured by potassium dichromate method 590 value

[0034] 1.3 Microscopic examination of strains and 16S identification The above-mentioned ethanol-using strain Pediococcus pentosaceus LTPP-271 was subjected to Gram staining, and the microscopic examination results were as follows: Figure 2The cultured bacterial liquid was sent for testing for 16 seconds, and the identification sequence was shown in SEQ ID NO. 1. Comparison confirmed that it was Pediococcus pentosaceus. Pediococcus pentosaceus LTPP-271 was deposited with the Guangdong Provincial Microbiological Culture Collection on June 4, 2024, at Building 59, 5th Floor, No. 100 Xianlie Middle Road, Guangzhou, with the deposit number GDMCC No. 64724.

[0035] SEQ ID NO.1: 1.4 In vitro ethanol homologue utilization assay The main ethanol homologues in liquor are n-propanol, isobutanol, isopentanol, etc. n-propanol, isobutanol, and isopentanol were added to MRS medium at a volume of 500 mg / mL and inoculated. 600 The values did not decrease significantly. GC / MS was used to determine the contents of volatile substances such as n-propanol, isobutanol, and isopentanol in the samples.

[0036] GC conditions: inlet temperature 250°C, carrier gas high-purity helium at a flow rate of 1.0 mL / min, split injection, split ratio 30:1. Column: TG-WAXMS (0.25 μm, 30 m × 0.25 mm). Initial temperature 40°C, ramped to 100°C at a rate of 5°C / min, then to 230°C at a rate of 40°C / min, held for 5 min.

[0037] Mass spectrometry conditions: ionization mode: EI; emission current: 50 μA; electron energy: 70 eV; ion source temperature: 250°C; transfer line temperature: 240°C; mass scan range: 33–350 amu.

[0038] The utilization rates of Pediococcus pentosaceus LTPP-271 for n-propanol, isobutanol, and isopentanol were 58.52%, 38.72%, and 46.30%, respectively, which can accelerate the metabolic utilization of alcohol substances.

[0039] Table 2 Content (mg / mL) and utilization rate of ethanol homologues in Pediococcus pentosaceus LTPP-271

[0040] 1.5 Alcohol dehydrogenase and acetaldehyde dehydrogenase activity assay The activity of alcohol dehydrogenase and acetaldehyde dehydrogenase was determined according to the kits of Beijing Solebaugh Technology Co., Ltd. Bacterial sample processing: After activation of Pediococcus pentosaceus LTPP-271, the culture was shaken for 20 h, and the fermentation broth was centrifuged at 8000 rpm for 5 min. 6 Resuspend the bacteria at a ratio of 10 CFU of bacteria to 1 mL of extract. Disrupt the resuspension by ultrasonication in an ice bath at 300 W for 3 seconds, 7 seconds between intervals, for 3 minutes. Centrifuge at 12,000 rpm at 4°C for 20 minutes. Place the supernatant on ice for testing.

[0041] Alcohol dehydrogenase activity assay: Follow the instructions in the kit, adding the reagents sequentially and measuring the absorbance change at 340 nm over a period of 15 to 75 seconds. For a blank control, replace the corresponding supernatant with distilled water. The final alcohol dehydrogenase activity, calculated using the formula in the kit, was 18.38 U / mL.

[0042] Acetaldehyde dehydrogenase activity: Add the above reagents separately to a 1 mL quartz cuvette, mix thoroughly, and measure the absorbance value A1 at 340 nm after 1 minute. Immediately place in a 37°C water bath for 30 minutes, take out, wipe dry, and measure the absorbance value A2 at 31 minutes. According to the formula given in the kit, the acetaldehyde dehydrogenase activity was measured to be 48.16 U / mL.

[0043] Comparative Example 1 In vitro ethanol utilization test of Pediococcus pentosaceus BNCC193259 The same method as in Example 1.2 was used, except that the strain used was Pediococcus pentosaceus BNCC193259, purchased from the Guangdong Provincial Microbial Culture Bank. Ethanol utilization by Pediococcus pentosaceus BNCC193259 was tested. The test results are shown in Table 3. Substituting the absorbance into the formula, the results show that when supplemented with 3% ethanol, Pediococcus pentosaceus BNCC193259 utilized 1.7% of the ethanol in the culture medium. In MRS culture supplemented with 5% ethanol, the ethanol utilization rate was -0.5%. The calculated results for the utilization of ethanol in the culture medium by Pediococcus pentosaceus BNCC193259 were within a 5% error margin, indicating that Pediococcus pentosaceus BNCC193259 does not have the ability to utilize ethanol.

[0044] Table 3 OD values corresponding to ethanol concentration measured by potassium dichromate method 590 value

[0045] By comparing the ethanol utilization capabilities of different Pediococcus pentosaceus in Comparative Example 1 and Example 1 of the present application, it can be seen that not all Pediococcus pentosaceus have the ability to utilize ethanol.

[0046] Example 2 Determination of probiotic properties of Pediococcus pentosaceus LTPP-271 2.1 Verification of hemolytic activity of Pediococcus pentosaceus LTPP-271 Pediococcus pentosaceus was streaked on Columbia blood agar plates and cultured at 37°C in an anaerobic workstation for 48 h. Figure 3 , Pediococcus pentosaceus LTPP-271 is not hemolytic.

[0047] 2.2 Simulated gastric and intestinal fluid tolerance test Sterile simulated gastric and intestinal fluids were purchased from Shanghai Yuanye Biotechnology Co., Ltd. After streaking and activation, single colonies were inoculated into MRS liquid medium. After 20 hours of incubation, 5 mL of the suspension was centrifuged and resuspended in sterile saline. The resuspended suspension was mixed with simulated gastric fluid at a ratio of 1:9 and incubated at 37°C. Serial dilutions were performed at 0 and 3 hours, with triplicate counts per strain. The survival rate was calculated as the number of viable cells at 3 hours / the number of viable cells at 0 hours × 100%. The resuspended suspension was mixed with simulated intestinal fluid at a ratio of 1:9 and incubated at 37°C. Serial dilutions were performed at 0, 3, and 24 hours, with triplicate counts per strain. The survival rate was calculated as the number of viable cells at sampling time / the number of viable cells at 0 hours × 100%. As shown in Tables 3 and 4 below, Pediococcus pentosaceus LTPP-271 exhibited good tolerance to simulated gastric acid and intestinal fluid. After 3 hours in simulated artificial gastric fluid, the number of viable bacteria did not decrease compared to 0 hours, thus the survival rate of Pediococcus pentosaceus LTPP-271 in simulated artificial gastric fluid was 100% (Table 4). Table 5 shows that the survival rate of Pediococcus pentosaceus LTPP-271 was 92.2% ± 3.2% after 3 hours and 78.6% ± 2.4% after 24 hours.

[0048] Table 4 Effective viable counts of Pediococcus pentosaceus LTPP-271 in simulated gastric fluid

[0049] Table 5 Effective viable counts of Pediococcus pentosaceus LTPP-271 in simulated intestinal fluid

[0050] 2.3 Bile salt tolerance After streaking, single colonies were selected and inoculated into MRS liquid medium for 16 hours. A 5% inoculum of the bacterial suspension was then inoculated into MRS medium supplemented with 0.1%, 0.2%, and 0.3% bile salts. Anaerobically incubated at 37°C for 24 hours, the number of viable cells was determined under different bile salt concentrations to calculate the strain's tolerance to bile salts. Bile salt tolerance = the number of viable cells of Pediococcus pentosaceus in the presence of different bile salt concentrations / the number of viable cells in the medium without bile salts × 100%.

[0051] As shown in Table 6 below, Pediococcus pentosaceus LTPP-271 has good bile salt tolerance. When cultured for 24 h under conditions containing 0.1%, 0.2%, and 0.3% bile salts, the survival rates were 68.8%±2.6%, 41.3%±2.3%, and 26.8%±2.8%, respectively.

[0052] Table 6 Effective viable counts of Pediococcus pentosaceus LTPP-271 in bile salt-containing medium

[0053] 2.4 Determination of strain self-agglutination ability After activating the standby strain, the strain was inoculated into liquid culture medium and cultured at 37°C for 24 h. The strain was then centrifuged at 10,000 rpm / min for 5 min. The strain was washed three times with sterile PBS and then resuspended and the OD value of the bacterial solution was adjusted. 600 Record the initial OD between 0.6 and 0.8 600 The value is A0. After 4 mL of the suspension is allowed to stand for 24 h, the supernatant is slowly aspirated to measure the OD 600 The value is recorded as A, and the self-agglutination rate is calculated. Self-agglutination rate = [(A0-A) / A0] × 100%. The initial OD of the suspension of Pediococcus pentosaceus LTPP-271 was tested. 600 The A0 value was 0.767±0.03, and the OD 600 The value A is 0.303±0.02, so the self-aggregation rate of Pediococcus pentosaceus LTPP-271 is 60.4%±6.5%, which has a good self-aggregation effect.

[0054] 2.5 Intestinal colonization ability Various microorganisms (bacteria) often fall from different environments to different parts of the host, and can settle in these parts and continue to grow and reproduce offspring. This phenomenon is usually called "colonization." The genetic similarity of the fecal microbial communities of zebrafish, mice and humans is 50%, and these genes account for 99% of the gene expression abundance of each metagenome. CM-DiI or Dio labels bacteria by binding to the lipid molecules of the membrane structure. It is stably expressed in the bacteria and has strong and stable fluorescence. The dye is passed into the daughter cells as the bacteria divide, but it will not enter adjacent bacteria. The target probiotics are labeled with fluorescent dyes. As the zebrafish ingest them, the fluorescently labeled target probiotics can adhere to and colonize in the intestines of the zebrafish, showing strong fluorescence in the intestines. Taking advantage of the transparent characteristics of zebrafish embryos, the growth of probiotics in zebrafish can be observed. The specific experimental process and results are as follows: Table 7 Results of the experimental evaluation of the stable colonization time of the intestine after the removal of Pediococcus pentosaceus LTPP-271 (n = 10)

[0055] Comparative Example 2: Zebrafish colonization experiment with Pediococcus pentosaceus BNCC193259 The same method as in Example 2.5 was used, except that the strain used was Pediococcus pentosaceus BNCC193259 purchased from the Guangdong Provincial Microbial Culture Bank, to test the colonization of Pediococcus pentosaceus BNCC193259 in the zebrafish intestine. Figure 6 and Figure 7 It can be seen that the stable colonization time of Pediococcus pentosaceus BNCC193259 is 4 h.

[0056] Table 8 Evaluation of the stable intestinal colonization time after removal of Pediococcus pentosaceus BNCC193259 (n = 10)

[0057] 2.6 Antibacterial experiment Escherichia coli ( Escherichia coli ATCC25922), Staphylococcus aureus ( Staphylococcus aureus ATCC6538) and Pseudomonas aeruginosa ( Pseudomonas aeruginosa CMCCB10104) were inoculated into nutrient agar medium to revive and activate the strains. The activated pathogenic bacteria were inoculated into broth medium and the bacterial solution concentration was adjusted to 1×10 8 CFU / mL. Pipette 0.5 mL of the above pathogenic bacteria culture solution and add it to 200 mL of sterilized nutrient agar medium that has not yet solidified (the temperature has been cooled to about 40°C). After fully mixing, prepare nutrient agar plates at a volume of 16 mL per culture dish. After the culture medium has cooled and solidified, place a sterilized Oxford cup in the culture medium and press it gently so that there is no gap between the Oxford cup and the plate. Each plate corresponds to one strain of bacteria, and three Oxford cups are set as replicates. Resuscitate Pediococcus LTPP-271 and activate it, and adjust the concentration of the cultured bacterial solution to 1×10 8 CFU / mL. 100 μL of the bacterial broth was added to an Oxford cup and incubated at 37°C for 24 h. After incubation, the diameter of the inhibition zone was measured using a vernier caliper and recorded. The results showed that the fermentation broth of Pediococcus pentosaceus LTPP-271 had strong inhibitory activity against the growth of pathogenic bacteria such as Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa (Table 9).

[0058] Table 9 Evaluation of antibacterial activity of Pediococcus pentosaceus LTPP-271 strain (diameter: mm)

[0059] Example 3 Growth of Pediococcus pentosaceus LTPP-271 in electrochemical mixed carbon source The streaked and activated Pediococcus pentosaceus LTPP-271 colonies were picked and inoculated into MRS liquid culture medium with an electrochemical mixed carbon source added, wherein the electrochemical mixed carbon source replaced 50% of the glucose in the MRS culture medium. The specific formula is shown in Table 10 below. The control group was MRS culture medium. The final concentrations of the main components of the electrochemical mixed carbon source were: 20 g / L ethanol, 3 g / L sodium formate, and 8 g / L sodium acetate. The effective viable bacteria count in the culture medium was determined after culturing at 37°C for 16 hours in an anaerobic incubator. The results are shown below. Figure 8After the electrochemical mixed carbon source replaced 50% of the glucose in the culture medium, the effective viable count of Pediococcus pentosaceus LTPP-271 was 4.91×10 9 ± 1.63×10 8 CFU / mL, and the effective viable bacteria count in the MRS medium of the control group was 3.23×10 9 ± 1.91×10 8 CFU / mL, the effective viable bacteria count increased by 52.1%.

[0060] Table 10 Culture medium components

[0061] All of the above are intended to be primary implementations of this intellectual property and do not constitute limitations on other implementations of such new products and / or methods. Those skilled in the art will utilize this important information and modify the above to achieve similar implementations. However, all modifications or adaptations based on this invention to new products are reserved.

[0062] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any other manner. Any person skilled in the art may utilize the above-disclosed technical content to modify or modify the present invention into equivalent embodiments. However, any simple modifications, equivalent variations, and modifications to the above embodiments that do not depart from the technical content of the present invention and are based on the technical essence of the present invention remain within the scope of protection of the present invention.

Claims

1. A Pediococcus pentosaceus having the function of accelerating the metabolism of alcohol substances, characterized in that: The Pediococcus pentosaceus was named LTPP-271 and its deposit number was GDMCC No. 64724.

2. The Pediococcus pentosaceus having the function of accelerating alcohol metabolism according to claim 1, characterized in that: The 16s nucleotide sequence of Pediococcus pentosaceus LTPP-271 is shown in SEQ ID NO.

1.

3. The Pediococcus pentosaceus having the function of accelerating alcohol metabolism according to claim 1 or 2, characterized in that: The Pediococcus pentosaceus LTPP-271 is cultured in an MRS medium containing 3% (v / v) ethanol and absorbs and utilizes 64.6% of the ethanol in the medium, with utilization rates of n-propanol, isobutanol, and isopentanol being 58.52%, 38.72%, and 46.30%, respectively.

4. The Pediococcus pentosaceus having the function of accelerating alcohol metabolism according to claim 1 or 2, characterized in that: The Pediococcus pentosaceus LTPP-271 can colonize the zebrafish intestine for more than 48 hours.

5. The fermentation production method of Pediococcus pentosaceus according to any one of claims 1 to 4, characterized in that: include: The Pediococcus pentosaceus LTPP-271 is inoculated into a culture medium at an inoculum size of 2% to 5%, and cultured at 37° C. for 16 to 24 hours to obtain a culture solution. The culture solution is centrifuged and freeze-dried to obtain bacterial cells.

6. The fermentation production method of Pediococcus pentosaceus according to claim 5, characterized in that: The culture medium is MRS culture medium or a mixed carbon source obtained by adding electrochemically catalyzed CO2 to replace part of the glucose in the MRS culture medium, the mixed carbon source contains at least ethanol, and the concentration of the ethanol in the fermentation broth is 1%-5% (v / v).

7. A microbial preparation having the function of accelerating the metabolism of alcohol substances, characterized in that: The method comprises the Pediococcus pentosaceus according to any one of claims 1 to 4, wherein the viable cell count is not less than 1×10 6 CFU / mL or 1×10 6 CFU / g.

8. The microbial preparation having the function of accelerating the metabolism of alcohol substances according to claim 7, characterized in that: Also contains other probiotics.

9. A health product, food or medicine having the function of accelerating the metabolism of alcohol substances, characterized in that: Comprising the microbial preparation according to claim 7 or 8.

10. Use of the Pediococcus pentosaceus according to any one of claims 1 to 4 in the preparation of health products, foods or medicines having the function of accelerating the metabolism of alcohol substances.

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