Lactobacillus mucilaginosus and application thereof

Abstract

The invention discloses lactobacillus mucilaginosus and application thereof, and belongs to the technical field of microbial fermentation. The invention provides a lactobacillus mucus WX20 which is preserved in the Guangdong Microbial Culture Collection Center on September 12, 2024, and the preservation number of the lactobacillus mucus WX20 is GDMCC No: 65083. The lactobacillus mucus WX20 has the advantages that the lactobacillus mucus WX20 is preserved in the Guangdong Microbial Culture Collection Center on September 12, 2024; the strain obtained by screening can effectively degrade inosine and guanosine and inhibit the activity of xanthine oxidase, and has good tolerance in a simulated gastrointestinal fluid environment. When the strain is used for fermenting the leaf mustard, the fermentation period of the fermented leaf mustard can be shortened, a product prepared by utilizing the strain has the activity of degrading inosine and guanosine and inhibiting xanthine oxidase, a microbial inoculum and pickles can remarkably reduce the blood uric acid level of a mouse with hyperuricemia, and the strain has considerable application potential and high market value.

Description

Technical Field

[0001] The invention relates to a fermented mucus lactobacillus strain and application thereof, belonging to the technical field of microbial fermentation. Background Art

[0002] Uric acid is the final product of purine metabolism in the human body. When urate is over-secreted or under-excreted, it will cause hyperuricemia. As of now, there are more than 1 billion patients with hyperuricemia in the world, and the incidence rate is increasing year by year. Two-thirds of uric acid in the human body is endogenous, and the remaining one-third comes from the intake of exogenous high-purine and high-sugar foods. However, it is not enough to simply reduce the intake of high-purine foods to reduce uric acid levels. Intervention methods that inhibit uric acid synthesis and promote uric acid excretion are also needed. Xanthine oxidase inhibitors (allopurinol), uric acid dissolving drugs (pegylated mammalian uricase) and uricosuric drugs (benzbromarone) are currently the main drugs for the treatment of hyperuricemia. However, these drugs usually show certain side effects in the treatment of hyperuricemia. For example, allopurinol can cause severe allergic reactions and rashes, and benzbromarone may cause severe liver and kidney damage. Therefore, safe and effective methods are urgently needed to relieve hyperuricemia.

[0003] Lactic acid bacteria with uric acid lowering activity are applied to food fermentation to obtain foods with uric acid lowering activity for use in the daily diet of people with hyperuricemia to achieve the purpose of intervening in uric acid levels; at the same time, dietary supplementation of lactic acid bacteria can regulate the intestinal flora disorder caused by hyperuricemia; currently, there are still obvious deficiencies in the development and application of fermented foods using lactic acid bacteria with uric acid lowering activity; kimchi, as a traditional fermented food deeply loved by consumers, not only has a unique flavor and a refreshing taste, but also contains beneficial microorganisms such as lactic acid bacteria, which help to improve the intestinal environment and promote digestion. Compared with naturally fermented kimchi, inoculated fermentation can speed up the fermentation rate, shorten the fermentation cycle and thus improve production efficiency, and the quality of the finished kimchi is stable. Lactic acid bacteria with uric acid lowering activity are applied to kimchi fermentation, which plays a role in assisting the improvement of hyperuricemia and enhancing the edible value of kimchi. Summary of the invention

[0004] In order to solve the problems existing in the prior art, one of the purposes of the present invention is to provide a fermentation mucus lactobacillus

[0005] (Limosilactobacillusfermentum), the fermented mucus lactobacillus is fermented mucus lactobacillus WX20, which was deposited in the Guangdong Provincial Microbiological Culture Collection Center on September 12, 2024, with the address: 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou; the deposit number is: GDMCC No: 65083, and the taxonomic name is: Limosilactobacillusfermentum.

[0006] Another object of the present invention is to provide a bacterial agent prepared by fermenting Lactobacillus mucilaginosus WX20.

[0007] Preferably, the concentration of the bacterial agent is 8 to 16 Lg CFU / g.

[0008] Preferably, the preparation method of the bacterial agent is as follows: fermented mucus lactobacillus WX20 is inoculated in MRS liquid culture medium to obtain a first-generation bacterial liquid, the first-generation bacterial liquid is again inoculated in MRS liquid culture medium to obtain a second-generation bacterial liquid, the second-generation bacterial liquid is centrifuged, the supernatant is discarded, the bacteria are washed with sterile water, the washed bacteria are resuspended with a skim milk powder solution, vacuum dried, and crushed to obtain a bacterial agent.

[0009] Preferably, the inoculation amount of the bacterial liquid is 1-5% of the volume of the MRS liquid culture medium.

[0010] Preferably, the culture time is 10 to 20 hours.

[0011] Preferably, the centrifugation conditions are: 2-6°C, 5000-8000 rpm for 3-20 min.

[0012] Preferably, the mass concentration of the skim milk powder solution is 5% to 35%.

[0013] The third object of the present invention is to provide an application of fermented Lactobacillus mucilaginosus WX20 or its bacterial agent, and the application includes the following two aspects:

[0014] (1) Application of fermented Lactobacillus mucilaginosus WX20 in the preparation of a drug for treating uric acid lowering;

[0015] (2) Application of fermented mucilaginous Lactobacillus WX20 in the preparation of direct-fermented mustard kimchi.

[0016] Preferably, the present invention provides an application method of fermented mucinous Lactobacillus WX20 bacterial agent in the preparation of direct-injection fermented mustard kimchi, specifically: wash and dry mustard, ginger slices, garlic slices and peppers, add them into a kimchi jar in sequence, add brine, dissolve 0.3-1.8g of the bacterial agent with sterile water and add it into the kimchi jar, ensure that the bacterial agent is evenly dispersed and adheres to the surface of the mustard, finally, compact the vegetables, cover with a lid, seal the kimchi jar, and place the kimchi jar in a light-proof environment at room temperature for fermentation for 5-9 days.

[0017] Preferably, the mass ratio of mustard greens, ginger slices, garlic slices and chili peppers is (50-500):(1-20):(1-20):(0.5-10)

[0018] Preferably, the amount of brine added is enough to cover the mustard greens, ginger slices, garlic slices and chili peppers.

[0019] Preferably, the brine comprises edible salt, liquor and sucrose, the mass concentration of edible salt in the brine is 2%-6%, the volume concentration of liquor is 0.6%-2%, and the mass concentration of sucrose is 1%-3.5%.

[0020] Technical effects of the present invention:

[0021] (1) The fermented mucus Lactobacillus WX20 provided by the present invention has good ability to degrade inosine and guanosine, and can inhibit the activity of xanthine oxidase. It has good survival ability in simulated gastrointestinal fluid, is sensitive to a variety of common antibiotics, and has high safety.

[0022] (2) The direct-fermented pickles prepared by the fermented Lactobacillus mucilaginosus WX20 of the present invention have degradation rates of inosine and guanosine of 75.64% and 71.13% respectively, and an inhibition rate of xanthine oxidase of 95.34%, and have excellent uric acid-lowering potential.

[0023] (3) The fermented Lactobacillus mucilaginosus WX20 starter of the present invention is used for direct fermentation, which shortens the pickling time of kimchi and greatly improves the production efficiency.

[0024] (4) The fermented Lactobacillus mucilaginosus WX20 provided by the present invention and the mustard kimchi fermented with the bacterial agent made from the strain of the present invention can significantly reduce the blood uric acid level of hyperuricemia mice. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 This is the result of microscopic examination after Gram staining of fermented Lactobacillus mucosa WX20.

[0026] Figure 2 The chromatograms and standard curves of inosine and guanosine are shown in Figure 2.

[0027] Figure 3 is the degradation rate of inosine and guanosine by fermentation Lactobacillus mucilaginosus WX20.

[0028] Figure 4 is the inhibition rate of fermented Lactobacillus mucilaginosus WX20 on xanthine oxidase.

[0029] Figure 5 The uric acid-lowering activity of fermented kimchi.

[0030] Figure 6 is the pH value of fermented kimchi.

[0031] Figure 7 is the number of viable bacteria in fermented kimchi. DETAILED DESCRIPTION

[0032] The present invention is further described in detail below in conjunction with the accompanying drawings and specific embodiments, but the protection scope of the present invention is not limited to the described contents.

[0033] The reagent preparation method used in the embodiment is as follows:

[0034] (1) The preparation method of MRS liquid medium is as follows: 10.0 g of peptone, 8.00 g of beef extract powder, 4.0 g of yeast extract powder, 20.0 g of glucose, 2.0 g of dipotassium hydrogen phosphate, 2.0 g of diammonium hydrogen citrate, 5.0 g of sodium acetate, 0.2 g of magnesium sulfate, 0.04 g of manganese sulfate, and 1.0 g of Tween 80 are dissolved in 1.0 L of distilled water and the pH value is adjusted to 6.5.

[0035] (2) The preparation method of MRS-CaCO3 solid culture medium is as follows: take 10.0 g of peptone, 8.0 g of beef extract powder, 4.0 g of yeast extract powder, 20.0 g of glucose, 2.0 g of dipotassium hydrogen phosphate, 2.0 g of diammonium hydrogen citrate, 5.0 g of sodium acetate, 0.2 g of magnesium sulfate, 0.04 g of manganese sulfate, 1.0 g of Tween 80, 15 g of calcium carbonate, and 20 g of agar; dissolve them in 1.0 L of distilled water and adjust the pH value to 6.5.

[0036] (3) The preparation method of 0.1 mol / L K3PO4 solution (pH = 7.0) is as follows: weigh 2.1227 g of K3PO4 and add it into 100 mL of ultrapure water.

[0037] (4) The preparation method of 1.25 mmol / L inosine solution is as follows: weigh 33.53 mg of inosine and add it into 100 mL of K3PO4 (pH = 7.0) solution.

[0038] (5) The preparation method of 1.25 mmol / L guanosine solution is as follows: weigh 35.41 mg of guanosine and add it to 100 mL of K3PO4 (pH = 7.0) solution.

[0039] (6) The preparation method of 0.2 mol / L phosphate buffer (pH = 7.5) is as follows: weigh 3.12 g of sodium dihydrogen phosphate powder and dissolve it in 100 mL of water to prepare a 0.2 mol / L sodium dihydrogen phosphate aqueous solution, weigh 7.16 g of disodium hydrogen phosphate powder and dissolve it in 100 mL of water to prepare a 0.2 mol / L disodium hydrogen phosphate aqueous solution, take 16 mL of the sodium dihydrogen phosphate aqueous solution and 84 mL of the disodium hydrogen phosphate aqueous solution and mix them to obtain a 0.2 mol / L phosphate buffer with a pH of 7.5.

[0040] (7) 0.5 U / mL xanthine oxidase was prepared as follows: 1 mg of xanthine oxidase with an enzyme activity of 8.3 U / mg was weighed and added to 16.6 mL of phosphate buffer (0.2 mol / L, pH = 7.5) to dissolve.

[0041] (8) 1 mmol / L xanthine solution was prepared as follows: 3.81 mg of xanthine was weighed into a beaker, 500 μL of 0.1 mol / L NaOH solution was added for ultrasonic dissolution, and then the volume was fixed to 25 mL with phosphate buffer (0.2 mol / L, pH = 7.5).

[0042] (9) The preparation steps of simulated gastric fluid are as follows: prepare 100 mL of 0.5% sodium chloride, add 0.35 g of pepsin, vortex mix, adjust the pH to 2 with concentrated hydrochloric acid, and filter with a sterile 0.22 μm filter membrane for sterilization.

[0043] (10) The preparation steps of simulated intestinal fluid are as follows: 1.216 g of potassium dihydrogen phosphate and 0.3 g of pig bile salt are dissolved in 100 mL of water, the pH value is adjusted to 7.8 with 0.1 mol / L sodium hydroxide solution, and then 0.1 g of trypsin is added. Sterilize by filtering with a sterile 0.22 μm filter membrane.

[0044] Example 1

[0045] Strain isolation and identification

[0046] (1) Strain isolation: 1 mL of kimchi brine stored in 50% glycerol was taken and added to 15 mL of MRS liquid culture medium, and cultured at 37°C until the test tube became turbid and there was no wrinkled substance covering the surface of the culture medium. At this time, the first-generation bacterial solution was obtained. After that, continuous subculture was performed every 12 hours to obtain the third-generation bacterial solution. 1 mL of the third-generation bacterial solution was centrifuged to remove the supernatant and washed twice with physiological saline before resuspending. Gram staining was performed for microscopic examination, and the strains with purple microscopic fields were selected for the next screening. The third-generation bacterial solution of the strains with good microscopic examination results was diluted to 10 with physiological saline. -5 , 10 -6 , draw 100 μL, spread on MRS-CaCO3 solid plate, invert and culture at 37℃ for 48h, select single colonies with calcium dissolution circle and streak purification on MRS-CaCO3 solid plate, purify 2-3 times until there are more single colonies on the plate; select single colonies for Gram staining and microscopic examination, select single colonies for enrichment culture in MRS liquid medium and store in 50% glycerol, freeze at -80℃, named WX20, WX20 Gram staining and microscopic examination as follows Figure 1 Shown, the microscope field shows purple rod-shaped bacteria.

[0047] (2) Extract genomic DNA from the WX20 strain obtained in step (1), then amplify the genomic DNA of the strain using 16S rDNA universal primers 36F and 1492R, and then sequence and identify the strain species.

[0048] The primer sequences of 16S rDNA universal primers 36F and 1492R are as follows:

[0049] 36F: 5'-TCTGGCTCAGGATGAACGC-3';

[0050] 1492R: 5'-TACGGCTACCTTGTTACGACTT-3';

[0051] PCR system components: 1 μL DNA template (bacterial solution), 1 μL 36F, 1 μL 1492R, 22 μL sterile water, 2× TaqPCR Master Mix (PCR premix).

[0052] The strain was identified as Lactiplantibacillus by morphology and 16S rDNA. The results of 16S rDNA homology analysis showed that the homology with the known fermenting mucus lactobacillus (Limosilactobacillusfermentum) in the GeneBank database reached 100%. The isolated fermenting mucus lactobacillus was named WX20, and its 16S rDNA sequence is shown in SEQ ID NO.1.

[0053] Example 2

[0054] Evaluation of the uric acid-lowering activity of fermented Lactobacillus mucilaginosus WX20 in vitro

[0055] The strain was inoculated into MRS medium at 2% inoculation volume and incubated overnight at 37°C to activate the first generation bacterial solution. The first generation bacterial solution was subcultured at 2% inoculation volume and cultured at 37°C for 15 h. 1 mL of the second generation bacterial solution was taken and centrifuged at 5000 rpm for 5 min at 4°C. The precipitate was washed three times with sterile water, and then the precipitated bacteria were resuspended in 5 mL of 1.25 mmol / L inosine and guanosine solution to make the concentration of fermented mucus Lactobacillus WX20 reach 10 9 CFU / mL. The suspension was fully mixed, shaken at 200 rpm for 1 h at 37°C, and then 0.1 mL of perchloric acid solution (0.1 mol / L) was added to stop the reaction. The mixture was centrifuged at 5000 rpm for 5 min, and the supernatant was filtered with a 0.22 μm membrane filter and then measured on a machine. The concentrations of inosine and guanosine in the supernatant were determined by high performance liquid chromatography. The chromatographic conditions were Agilent TC-C18 (2) column (4.6×250 mm, 5 μm), flow rate of 0.8 mL / min, column temperature of 30°C, mobile phase of methanol: water = 10:90, detection wavelength of 248 nm, injection volume of 10 μL, and standard curves of inosine and guanosine were drawn according to peak area and concentration. These standard curves were used to calculate the concentrations of inosine and guanosine in the solution after lactic acid bacteria degradation.

[0056] The degradation rates of inosine and guanosine by fermented Lactobacillus mucilaginosus WX20 are as follows Figure 2 shown.

[0057] The standard curve of inosine is y=7997.8x+111.03, R 2 =0.9996, the standard curve of guanosine is y=7557.5x+197.81, R 2 =0.9995, such as Figure 3 As shown, the degradation rate of inosine by fermented Lactobacillus mucosa WX20 was 39.69%, and the degradation rate of guanosine was 98.11%, which was considered to have good in vitro uric acid-lowering activity.

[0058] Example 3

[0059] Determination of the inhibition rate of xanthine oxidase by fermented Lactobacillus mucilaginosus WX20

[0060] The preparation of strain samples was as follows: the strain was inoculated into MRS medium at a 2% inoculation rate, incubated at 37°C overnight to obtain the first-generation bacterial solution, the first-generation bacterial solution was further subcultured and cultured at 37°C for 15 h to obtain the second-generation bacterial solution, and the OD of the second-generation bacterial solution was calculated. 600 Adjusted to 1.1±0.5. Take 2mL of the adjusted second-generation bacterial solution and centrifuge at 4°C, 5000rpm for 5min to obtain a cell-free supernatant (CFS). Wash the precipitate three times with sterile water and resuspend it with 2mL of sterile water to make a cell suspension (CS). Ultrasonicate the CS at a power of 200W for 10min and then centrifuge to obtain a cell-free extract (CFE). Incubate the CS at 37°C for 12h and then centrifuge to obtain a cell-free metabolite (CFM).

[0061] The 96-well plate includes control wells (sterile water), positive control wells (allopurinol, 0.2 mg / L) and sample wells (CFS, CFE and CFM). First, add 75 μL of xanthine at a concentration of 1 mmol / L to each group of wells, then add 75 μL of sterile water, allopurinol and bacterial sample to CFS, CFE and CFM respectively; finally, add 75 μL of xanthine oxidase solution at a concentration of 0.5 U / mL and preheated at 25°C for 20 minutes; use an ELISA reader to record the changes in the absorbance of each group of wells at 295 nm within 6 minutes (measured once per minute), and calculate the inhibition rate of each sample on xanthine oxidase. The results are as follows: Figure 4 shown.

[0062]

[0063] In formulas (1) and (2): k 样品 The slope of the uric acid production curve for different samples; k 空白 The slope of the uric acid production curve drawn for the blank group; k 对照The slope when the uric acid production curve is plotted for allopurinol.

[0064] Depend on Figure 4 It can be seen that the inhibition rate of the supernatant (CFS) of fermented Lactobacillus mucilaginosus WX20 on xanthine oxidase is 56.89%, the inhibition rate of the cell-free fragment (CFE) on xanthine oxidase is 14.30%, and the inhibition rate of the cell-free metabolite (CFM) on xanthine oxidase is 19.32%.

[0065] Example 4

[0066] Production of fermented mucus lactobacillus WX20 bacterial agent

[0067] The fermented mucus lactobacillus WX20 was inoculated with an inoculum volume of 2% in MRS liquid medium for activation culture, and the second generation bacterial solution after 15 hours of culture was centrifuged at 4°C and 5000rpm for 5 minutes. After collecting the bacterial bodies, they were washed 3 times with sterile water. A 10% skim milk powder solution (w / v) was sterilized at 95°C for 15 minutes and then cooled to room temperature. The washed bacterial bodies were then resuspended with a skim milk powder solution, pre-frozen at -80°C for 1 hour, and then vacuum freeze-dried for 48 hours. The viable bacterial count of the bacterial agent was 10.20lg CFU / g.

[0068] Example 5

[0069] Evaluation of the probiotic properties of fermented Lactobacillus mucilaginosus WX20

[0070] (1) Evaluation of tolerance to simulated gastrointestinal fluid

[0071] Weigh the fermented Lactobacillus mucilaginosus WX20 freeze-dried powder to a concentration of 1×10 9 The CFU / mL bacterial suspension was added to 9 mL of simulated gastric fluid and incubated at 37°C for 3 h. Subsequently, 1 mL of the bacterial suspension digested in artificial gastric fluid for 3 h was taken and added to 9 mL of artificial intestinal fluid and cultured at 37°C for 3 h. The plate count method was used to determine the number of viable bacteria in the simulated gastric fluid and simulated intestinal fluid at 0 and 3 h, and the survival rate was calculated. The tolerance of Lactobacillus fermentans WX20 in simulated gastrointestinal fluid is shown in Table 1.

[0072] Table 1 Tolerance of Lactobacillus mucilaginosus WX20 to simulated gastrointestinal fluid

[0073] 0h(1gCFU / mL) 3h(lgCFU / mL) Survival rate Simulated gastric juice 8.19±0.02 8.11±0.03 83.55％ Simulated intestinal fluid 7.80±0.03 7.66±0.03 72.63％

[0074] As shown in Table 1, the survival rates of Lactobacillus mucosa WX20 in simulated gastric juice and simulated intestinal juice were 83.55% and 72.63%, respectively, and it had good tolerance to gastrointestinal juice.

[0075] (2) Antibiotic sensitivity test

[0076] 100 μL of 1×10 8 CFU / mL of bacterial suspension was applied on MRS agar plates. When there was no visible liquid on the surface of the culture medium, the antibiotic susceptibility paper was placed in the center of the culture dish. After incubation at 37°C for 48 hours, the diameter of the antibiotic inhibition zone was measured with a vernier caliper to evaluate its drug resistance.

[0077] Table 2 Evaluation of antibiotic susceptibility of fermented Lactobacillus mucilaginosus WX20

[0078]

[0079] As shown in Table 2, fermented mucus lactobacillus is sensitive to eight antibiotics, including rifampicin, gentamicin, penicillin, cefoperazone, clindamycin, tetracycline, chloramphenicol, and erythromycin, but is insensitive to ciprofloxacin and vancomycin, and has high safety.

[0080] Example 6

[0081] Application of fermented mucus lactobacillus WX20 in preparation of direct-throw fermented kimchi

[0082] Take 1000g mustard, 15g ginger slices, 15g garlic slices and 7.5g chili peppers and add them to a kimchi jar. In addition, add 3L brine (the mass concentration of edible salt in the brine is 5%, the volume concentration of white wine is 1.5%, and the mass concentration of sucrose is 1.5%). Then dissolve 0.3g of fermented mucus lactobacillus freeze-dried powder in water (just dissolve the lactobacillus freeze-dried powder) and add it to the kimchi jar. Turn the vegetables to make the strains adhere evenly to the surface of the mustard, then press the vegetables with pebbles, cover the lid and seal the jar with water, and place the kimchi jar at room temperature and ferment for 8 days in the dark.

[0083] Comparative Example 1

[0084] The difference between this comparative example 1 and example 6 is that no leavening agent is inoculated and natural fermentation is adopted.

[0085] Effect Analysis of Example 6 and Comparative Example 1

[0086] (1) Determination of uric acid-lowering activity of kimchi

[0087] A potassium phosphate solution containing inosine and guanosine was prepared using kimchi water as a solvent. In addition, the kimchi homogenate was mixed with an equal amount of the prepared inosine and guanosine solution (w / v=1:1), incubated at 37°C for 1 hour and then centrifuged. The content of inosine and guanosine in the kimchi water and kimchi was determined and their degradation rates were calculated. The kimchi was homogenized, filtered and the filtrate was collected. The kimchi water was adjusted to pH 7.5 with 0.1 mol / L NaOH and filtered. The collected filtrate was filtered with a filter membrane with a pore size of 0.22 μm. The inhibition rate of kimchi and kimchi water on xanthine oxidase was determined, and the results are as follows: Figure 5 shown.

[0088] Depend on Figure 5 It can be seen that the degradation of inosine and guanosine by kimchi and brine in Example 6 first increased, then decreased, and finally stabilized. The degradation rate of inosine and guanosine by kimchi in Comparative Example 1 did not change significantly. At the end of fermentation, the degradation rates of inosine and guanosine by kimchi in Example 6 reached 44.74% and 53.25%, respectively, and the degradation rates of inosine and guanosine by kimchi water reached 75.14% and 76.13%, respectively. The inhibition rates of xanthine oxidase by kimchi and brine in Example 6 and Comparative Example 1 increased.

[0089] At the end of fermentation, the inhibition rate of kimchi on xanthine oxidase in Example 6 increased from 23.26% to 47.81%, and the inhibition rate of kimchi water on XOD was as high as 95.34%, which was higher than the inhibition rate of kimchi and kimchi water on XOD in Comparative Example 1.

[0090] (2) Determination of kimchi pH value

[0091] The pH value of kimchi brine was measured with a calibrated pH meter. Figure 6 shown.

[0092] Depend on Figure 6 It can be seen that the pH value of the kimchi in Example 6 during fermentation first decreased and then stabilized, and the final pH value was 3.34. The pH value of the kimchi in Comparative Example 1 was higher than that of the kimchi in Example 6 throughout the fermentation process. At the end of the fermentation time, the pH value was still decreasing and had not reached the late stage of fermentation.

[0093] (3) Determination of viable bacterial count in kimchi

[0094] The kimchi water was diluted 10 times with sterile water, and 10 -4 , 10 -5 , 10 -6 100 μL of kimchi water was spread on MRS agar medium and incubated at 37°C for 48 h before plate counting to obtain the change curve of total lactic acid bacteria count during kimchi fermentation. Figure 7 shown.

[0095] Depend on Figure 7 It can be seen that the number of lactic acid bacteria in kimchi in Example 6 first increased, then decreased and stabilized, while the number of viable bacteria in kimchi in Comparative Example 1 first increased and then decreased. The number of viable bacteria in Example 6 reached a peak of 8.54 lg CFU / mL on the 4th day of fermentation. At the end of fermentation, the colony count of kimchi in Example 6 was 7.87 lg CFU / mL, and the colony count of kimchi in Comparative Example 1 was only 5.59 lg CFU / mL, which was lower than that of kimchi in Example 6.

[0096] (5) Determination of volatile flavor substances in kimchi

[0097] 5g kimchi homogenate was placed in a 20mL headspace bottle and 20μL of 50mg / L cyclohexanone was added as an internal standard. After the sample bottle was equilibrated at 65℃ for 20min, it was extracted at 65℃ for 50min using a solid phase extraction needle. After the extraction, the extraction needle was inserted into the GC-MS system and thermal desorption was retained at 240℃ for 5min. The chromatographic conditions were HP-5MS gas chromatography column (30×0.25μm×0.25μm), helium as carrier gas, and the flow rate was 1mL / min. The initial temperature of the column oven was 40℃ for 3min, then heated to 150℃ at a rate of 4℃ / min, then heated to 230℃ at a rate of 7℃ / min, and finally held for 10min. The mass spectrometry conditions were: scanning range 30-400m / z, ionization voltage 70eV; ion source temperature 250℃. The qualitative and quantitative analysis of the volatile compounds was performed by comparing the retention times of NIST14, and the results are shown in Table 3.

[0098] Table 3 Types and contents of volatile flavor substances in kimchi

[0099]

[0100]

[0101]

[0102] As shown in Table 3, the volatile flavor compounds of kimchi can be mainly divided into seven categories, namely phenols, ethers, sulfides, aldehydes, ketones, isothiocyanates, esters, and other compounds. Isothiocyanates in kimchi are the main volatile compounds of mustard, among which allyl isothiocyanate is the main source of the spicy taste of mustard. As the fermentation proceeds, isothiocyanates can be degraded into sulfides. The content of allyl isothiocyanate in the kimchi of Example 6 is lower than that in Comparative Example 1. Esters are produced by esterification of free acids and alcohols in the late fermentation of kimchi, and have floral or fruity aromas. Terpenes are mainly produced in the late fermentation. The types and contents of esters and terpenes in kimchi in Example 6 are higher than those in kimchi in Comparative Example 1. Aldehydes and ketones have lower thresholds and contribute more to the flavor of kimchi. The types of aldehydes and ketones in kimchi in Example 6 are richer. In general, the flavor compounds of the kimchi in Example 6 are better than those in Comparative Example 1, and the kimchi in Example 6 has a good aroma.

[0103] Example 7

[0104] Intervention effects of fermented Lactobacillus mucilaginosus WX20 and its fermented mustard kimchi on serum uric acid levels in mice

[0105] After 1 week of adaptive feeding, 40 6-8 week old male C57BL / 6J mice (weight 20-25g) were randomly divided into normal group, model group, allopurinol group, fermented Lactobacillus mucus WX20 group (LF group), and kimchi group, with 8 mice in each group. Except for the normal group, mice in each group were gavaged with 100 mg / kg adenine and 300 mg / kg potassium oxonate every day. Both drugs were dissolved in 0.5% sodium carboxymethyl cellulose solution. At the same time, they were fed with 12% yeast feed to induce the establishment of a hyperuricemia mouse model. The mice in the normal group were gavaged with the same volume of 0.5% sodium carboxymethyl cellulose solution and fed with ordinary feed. The therapeutic drugs were gavaged after an interval of 1 hour. The control group and the model group were gavaged with 100 μL sterile water, the allopurinol group was gavaged with 5 mg / kg allopurinol, and the LF group was gavaged with 10 9 CFU / mL of fermented mucus lactobacillus WX20, the gavage dose was 100μL, and the kimchi group was gavaged with 50g / 60kg of fermented kimchi. The experimental period was 21 days. The uric acid level of mice was measured using a uric acid detector on the 1st, 7th, 14th and 21st days. The uric acid levels of mice are shown in Table 4:

[0106] Table 4 Uric acid values ​​of mice under the intervention of fermented mucus lactobacillus WX20 and its fermented kimchi

[0107]

[0108] As shown in Table 4, starting from the seventh day, the uric acid level of mice in the model group was higher than that in other groups, and the uric acid level continued to increase, reaching 341.78 μmol / L on the twenty-first day, indicating that the uric acid value of the mice reached a high uric acid level. The uric acid level of mice in the allopurinol group remained below 173 μmol / L. After intervention with fermented mucus Lactobacillus WX20, the uric acid level of mice was significantly lower than that of the model group. After gavage with fermented kimchi, the uric acid level of mice was also significantly lower than that of the model group. The uric acid values ​​of mice in the kimchi group gavaged with fermented mucus Lactobacillus WX20 and fermented with bacterial agents were lower than the high uric acid level.

Claims

1. A strain of Lactobacillus fermentum, characterized in that: The fermented mucus lactobacillus is fermented mucus lactobacillus WX20, and its preservation number is GDMCC No: 65083.

2. A bacterial agent prepared by fermenting mucus lactobacillus according to claim 1.

3. Use of the fermented mucus lactobacillus according to claim 1 or the bacterial agent according to claim 2 in the preparation of a medicament for treating uric acid lowering.

4. Use of the fermented mucus lactobacillus according to claim 1 or the bacterial agent according to claim 2 in the preparation of direct-injection fermented mustard kimchi.

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