Enterococcus lactis W3F and application thereof
By screening Enterococcus lactis W3F from Yunnan Shilin milk cake, the problem of the lack of multifunctional blood pressure lowering, blood sugar lowering and uric acid lowering activities in the existing technology has been solved. It provides a safe and effective lactic acid strain for the preparation of fermented foods and health products, which can significantly reduce blood pressure, blood sugar and uric acid levels in middle-aged and elderly people.
Patent Information
- Application Number
- CN202511228439.4
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-08-29
- Publication Date
- 2025-12-02
AI Technical Summary
Existing technologies lack lactic acid bacteria strains that simultaneously possess strong antihypertensive, hypoglycemic, and uric acid-lowering activities, and traditional drugs may have side effects, making it difficult to effectively prevent and treat hypertension, hyperglycemia, and hyperuricemia symptoms in middle-aged and elderly populations.
Enterococcus lactis W3F was isolated and screened from milk cake in Shilin, Yunnan. It has a high ability to produce extracellular polysaccharides and can inhibit the activity of xanthine oxidase, α-amylase, α-glucosidase and angiotensin-converting enzyme. It can be prepared into fermented food or functional food/health product for lowering blood pressure, blood sugar and uric acid.
Enterococcus lactis W3F significantly inhibits the activities of xanthine oxidase, α-amylase and α-glucosidase, with an angiotensin-converting enzyme inhibition rate as high as 88%. It has good safety and no side effects, and is suitable for the preparation of drugs, functional foods or health products. It significantly reduces blood pressure, blood sugar and uric acid levels.
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Abstract
Description
Technical Field
[0001] This invention belongs to the field of microbial application technology, specifically relating to Enterococcus lactis W3F and its applications. Background Technology
[0002] Currently, my country's aging population continues to intensify, and the health problems of middle-aged and elderly people are gradually attracting attention. After middle age, with increasing age, activity levels gradually decrease, and the body gradually experiences various signs of decline, such as a decline in the physiological functions of tissues and organs, weakened glandular secretion, slowed metabolism, reduced resistance, and weakened immune function. Over time, a series of chronic diseases in middle-aged and elderly people, such as hypertension, hyperlipidemia, diabetes, gout, cardiovascular system diseases, and osteoarthritis, follow. As of 2024, the number of people with hypertension in China was approximately 245 million, the number of people with diabetes reached 148 million, and the number of people with gout exceeded 80 million; many of them suffer from two or more chronic diseases at the same time, causing enormous harm to their physical and mental health, and also placing a heavy disease burden on their families, society, and the country.
[0003] Besides medical interventions, consuming foods with specific health benefits plays a crucial role in promoting health and inhibiting disease. Lactic acid bacteria, Gram-positive bacteria that ferment sugars into lactic acid, are members of the normal gut microbiota and possess unique advantages in terms of safety and efficacy. They have been proven to effectively improve gastrointestinal health and enhance host immunity. Research has confirmed that some lactic acid bacteria also exhibit various effects in the pharmaceutical and food industries, including antibacterial, antioxidant, anti-inflammatory, cholesterol-lowering, blood pressure-lowering, and immunomodulatory properties. Therefore, isolating and screening lactic acid bacteria strains with strong hypoglycemic, hypotensive, and uric acid-lowering activities, especially those with multiple activities, and developing them into pharmaceuticals, functional foods, or health products, is of significant importance for better prevention and treatment of hypertension, diabetes, gout, and other diseases in middle-aged and elderly individuals, and for improving their health and quality of life. Summary of the Invention
[0004] The purpose of this invention is to overcome the deficiencies in the prior art and provide Enterococcus lactis W3F and its application, which is derived from Yunnan Shilin milk cake, has good safety and broad-spectrum antibacterial properties, and can inhibit the activity of xanthine oxidase, α-amylase, α-glucosidase and / or angiotensin-converting enzyme, thereby achieving the effects of lowering blood pressure, lowering blood sugar and / or lowering uric acid.
[0005] To achieve the above objectives, the technical solution adopted by the present invention is as follows:
[0006] A strain of Enterococcus lactis W3F, capable of lowering blood pressure, blood sugar, and / or uric acid, is deposited at the China General Microbiological Culture Collection Center (CGMCC) under accession number CGMCC No. 32832.
[0007] As a further technical solution, the Enterococcus lactis W3F was isolated and screened from milk cake from Shilin, Yunnan.
[0008] As a further technical solution, the Enterococcus lactis W3F exhibits gastrointestinal tolerance.
[0009] As a further technical solution, the Enterococcus lactis W3F is a Enterococcus lactis W3F that produces a high amount of extracellular polysaccharides.
[0010] The use of Enterococcus lactis W3F and / or the fermentation product of Enterococcus lactis W3F in the preparation of antihypertensive drugs, hypoglycemic drugs and / or uric acid-lowering drugs.
[0011] The aforementioned Enterococcus lactis W3F is used as a fermentation agent. As a further technical solution, the Enterococcus lactis W3F and / or its fermentation products are used as functional additives in the preparation of functional foods or functional health products, which are used to lower blood pressure, lower blood sugar and / or lower uric acid.
[0012] As a further technical solution, the functional food includes any one of fermented milk, fermented soy milk, and fermented fish.
[0013] A fermented food that can lower blood pressure, blood sugar and / or uric acid is obtained by fermenting food raw materials with the aforementioned Enterococcus lactis W3F.
[0014] As a further technical solution, any one of the fermented milk, fermented soy milk, and fermented fish.
[0015] As a further technical solution, when the fermented food is fermented milk, the food raw material is one or more of goat milk, cow milk, and camel milk.
[0016] As a further technical solution, when the fermented food is fermented soy milk, the food ingredient is soy milk;
[0017] As a further technical solution, when the fermented food is fermented fish, the food ingredient is fish meat.
[0018] Compared with the prior art, the beneficial effects of the present invention are as follows:
[0019] This invention, based on an in vitro model of lowering blood pressure, blood sugar, and uric acid, screened Enterococcus lactis W3F from multiple strains of lactic acid bacteria from different food sources. Enterococcus lactis exhibits strong activities in lowering blood pressure, blood sugar, and uric acid simultaneously. It demonstrates excellent extracellular polysaccharide production, high safety, antibiotic sensitivity, and strong gastrointestinal tolerance. The strain or its fermentation products show inhibition rates of up to 42% on xanthine oxidase, 43% on α-amylase, 62% on α-glucosidase, and 88% on angiotensin-converting enzyme. It can be used to prepare drugs, functional foods, or functional health products for lowering blood pressure, blood sugar, and / or uric acid. It is particularly suitable for patients with hyperglycemia, hypertension, and hyperuricemia. Furthermore, fermented foods prepared using Enterococcus lactis W3F of this invention also have the effects of lowering blood pressure, blood sugar, and / or uric acid and can be used as functional foods or functional health products.
[0020] The Enterococcus lactis W3F strain of this invention was isolated from milk cake in Shilin, Yunnan Province. It is a probiotic and is safe, environmentally friendly, and has no side effects compared to traditional drugs for treating hyperglycemia, hypertension, and hyperuricemia. Attached Figure Description
[0021] Figure 1 This is a colony morphology diagram of Enterococcus lactis W3F.
[0022] Figure 2 Gram staining image of Enterococcus lactis W3F;
[0023] Figure 3 Phylogenetic tree of Enterococcus lactis W3F;
[0024] Figure 4 Image of the hemolytic zone of Enterococcus lactis W3F;
[0025] exist Figure 4 In the sample, A: Bacillus cereus ATCC 14579, B: Enterococcus lactis W3F. Detailed Implementation
[0026] The technical solution of the present invention will be clearly and completely described below with reference to specific embodiments. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.
[0027] In this invention,
[0028] mMRS medium: peptone 10 g / L, beef extract 10 g / L, yeast extract 5 g / L, sucrose 20 g / L, dipotassium hydrogen phosphate 2 g / L, sodium citrate 5 g / L, sodium acetate 5 g / L, magnesium sulfate heptahydrate 0.4 g / L, manganese sulfate tetrahydrate 0.1 g / L, and 20 g / L agar powder added to the solid medium; pH = 6.1.
[0029] Unless otherwise specified, all raw materials used in this invention are commercially available.
[0030] Example 1: Isolation of strains
[0031] 1. Isolation of strains
[0032] Strain Isolation: Weigh 10g of homemade milk cake from Shilin area of Yunnan, homemade fermented fish from Huangshan area of Anhui, and homemade pickled vegetables from Nanchang area of Jiangxi, respectively. Add 90mL of sterile water and shake on a shaker at 180r / min for 20min to prepare bacterial suspensions. Take 1mL of the bacterial suspension for serial dilution, and take 10g of each. -3 10 -4 10 -5 Three gradients were evenly spread on mMRS solid plates. After incubation at 37°C for 1-3 days, single colonies that grew rapidly, were moist and viscous, or could be drawn into threads were picked, purified to pure culture and stored.
[0033] Results: A total of 10 strains were isolated, including strains W3A, W3C, W3D, W3E and W3F from milk cake, strains ZXC-1, ZXC-4 and ZXC-5 from fermented fish, and strains CXY-1 and CXY-3 from sauerkraut.
[0034] Example 2: Study on the antihypertensive, hypoglycemic, and hypouric acid-lowering activities of the strain
[0035] 1. Preparation of sterile fermentation broth: Frozen bacterial cultures of W3A, W3C, W3D, W3E, W3F, ZXC-1, ZXC-4, ZXC-5, CXY-1, and CXY-3 were taken from a -80℃ freezer and streaked onto mMRS agar plates. The plates were incubated at 37℃ for 2 days. Single colonies were then transferred to 3 mL of mMRS liquid medium and incubated statically at 37℃ for 48 hours to obtain the seed culture. The seed culture was then transferred to 30 mL of mMRS liquid medium at a 2% inoculum level and incubated statically at 37℃ for 48 hours. The culture was then centrifuged at 10000 rpm for 10 minutes at 4℃. The supernatant was collected and filtered through a 0.22 μm filter to obtain the sterile fermentation broth, which is the sample to be tested.
[0036] 2. Detection indicators:
[0037] Uric acid-lowering activity was assessed by the inhibition rate of xanthine oxidase, hypoglycemic activity was assessed by the inhibition rates of α-amylase and α-glucosidase, and antihypertensive activity was assessed by the inhibition rate of angiotensin-converting enzyme.
[0038] 1) Determination of xanthine oxidase inhibitory activity: Mix 40 μL of the test sample with 20 μL of xanthine oxidase solution, shake for 30 s, incubate at 37℃ for 5 min, then add 40 μL of xanthine solution, shake for 30 s, react at 25℃ for 25 min, and measure the absorbance at 290 nm. The control group used phosphate buffer (pH 7.5) instead of the test sample, and the blank group used phosphate buffer (pH 7.5) instead of the xanthine oxidase solution. The formula for calculating the xanthine oxidase inhibition rate is as follows:
[0039] Xanthine oxidase inhibition rate (%) = [1 - (A1 - A2) / (A3 - A4)] × 100
[0040] In the formula: A1 represents the absorbance of the sample group; A2 represents the absorbance of the blank sample group; A3 represents the absorbance of the control group; A4 represents the absorbance of the blank control group.
[0041] 2) Determination of α-amylase inhibitory activity: Add 50 μL of the test sample to 50 μL of α-amylase solution, shake for 30 s, and incubate at 37℃ for 5 min. Then add 50 μL of 1% starch solution, mix well, react at 37℃ for 15 min, then add 200 μL of 3,5-dinitrosalicylic acid, boil in a water bath for 5 min, and after cooling, measure the absorbance at 540 nm. The control group used phosphate buffer (pH 6.8) instead of the test sample, and the blank group used phosphate buffer (pH 6.8) instead of the α-amylase solution. The formula for calculating the α-amylase inhibition rate is as follows:
[0042] α-Amylase inhibition rate (%) = [1 - (A1 - A2) / (A3 - A4)] × 100
[0043] In the formula: A1 represents the absorbance of the sample group; A2 represents the absorbance of the blank sample group; A3 represents the absorbance of the control group; A4 represents the absorbance of the blank control group.
[0044] 3) Determination of α-glucosidase inhibitory activity: Add 20 μL of the test sample to 20 μL of α-glucosidase solution, shake for 30 s, incubate at 37℃ for 5 min, then add 20 μL of p-nitrophenol glucoside solution, react at 37℃ for 15 min, and finally add 80 μL of Na₂CO₃ to terminate the reaction. Measure the absorbance at 405 nm. The control group used phosphate buffer (pH 6.8) instead of the test sample, and the blank group used phosphate buffer (pH 6.8) instead of the α-glucosidase solution. The formula for calculating the α-glucosidase inhibition rate is as follows:
[0045] α-glucosidase inhibition rate (%) = [1 - (A1 - A2) / (A3 - A4)] × 100
[0046] In the formula: A1 represents the absorbance of the sample group; A2 represents the absorbance of the blank sample group; A3 represents the absorbance of the control group; A4 represents the absorbance of the blank control group.
[0047] 4) Assay for angiotensin-converting enzyme (ACE) inhibitory activity: Add 40 μL of the test sample, 50 μL of FAPPGG (N-[3-(2-furanyl)acryloyl]-L-phenylalanyl-glycyl-glycine) substrate, and 10 μL of angiotensin-converting enzyme sequentially to a 96-well plate. Immediately after mixing, measure the absorbance at 340 nm. Then, react at 37℃ for 30 min, and measure the absorbance at 340 nm again. The control group was replaced with HEPES buffer (pH 8.3) instead of the test sample. The ACE inhibition rate was calculated using the following formula:
[0048] ACE inhibition rate = [1 - (A1 - A2) / (B1 - B2)] × 100%
[0049] In the formula: A1 represents the absorbance value of the sample group after 0 min of reaction; A2 represents the absorbance value of the sample group after 30 min of reaction; B1 represents the absorbance value of the control group after 0 min of reaction; B2 represents the absorbance value of the control group after 30 min of reaction.
[0050] 3. Results
[0051] Table 1. Uric acid-lowering, blood glucose-lowering, and blood pressure-lowering activities of the strain.
[0052]
[0053] Hyperuricemia is a metabolic disease caused by excessive uric acid production or impaired excretion in the body, and is a prerequisite for gout. Xanthine oxidase is a key enzyme that promotes uric acid production, oxidizing purines to uric acid. Therefore, reducing the activity of xanthine oxidase can effectively reduce uric acid production, thereby lowering blood uric acid concentration. As shown in Table 1, among all strains, W3F showed the highest inhibition rate against xanthine oxidase, at 42.89 ± 1.22%.
[0054] α-Amylase and α-glucosidase are involved in the hydrolysis of carbohydrates and the absorption of glucose, respectively, and are important causes of postprandial hyperglycemia. Therefore, inhibiting their activity is an effective measure to control type II diabetes. As shown in Table 1, among all strains, W3F had the highest inhibition rates against α-amylase and α-glucosidase, at 44.20±1.75% and 62.48±1.57%, respectively.
[0055] Angiotensin-converting enzyme (ACE) is involved in the synthesis of angiotensin II and mainly acts on the body's pressor system. Inhibiting ACE activity prevents angiotensin I from being converted into angiotensin II, thereby lowering blood pressure. As shown in Table 1, among all strains, W3E had the highest inhibition rate of ACE at 91.70±3.00%, while W3F had a slightly lower inhibition rate than W3E, ranking second at 88.26±1.26%.
[0056] Comprehensive analysis of the inhibition rates of the four enzymes revealed that, among all strains, W3F exhibited inhibition rates exceeding 40% for all four enzymes. Specifically, it showed the highest inhibition rates against xanthine oxidase, α-amylase, and α-glucosidase, and the second highest inhibition rate against angiotensin-converting enzyme. Other strains showed inhibition rates of one or more enzymes below 10%. Therefore, strain W3F, screened from milk cake, possesses strong antihypertensive, hypoglycemic, and uric acid-lowering activities.
[0057] Example 3: Identification of strain W3F
[0058] 1. Morphological and Gram staining identification: such as Figure 1 As shown, after strain W3F was cultured on mMRS plates at 37°C for 2 days, the colonies were milky white, round and convex, with smooth edges, moist surface, and viscous and stringy when picked up; Figure 2 As shown, Gram staining is blue, and the bacterial cells are spherical, indicating that they are Gram-positive bacteria.
[0059] 2. Molecular identification and phylogenetic tree of 16S rRNA
[0060] Strain W3F was identified using 16S rRNA sequencing. Universal primers 27F (5′-GTTTGATCMTGGCTCAG-3′) and 1492R (5′-TACGGYTACCTTGTTACGACTT-3′) were used for PCR amplification. The PCR products were sequenced and analyzed. The nucleotide sequence of the 16S rDNA of strain W3F is shown in SEQ ID NO.1. A phylogenetic tree was constructed using MEGA 5.1 software, and the results are shown below. Figure 3 .
[0061] SEQ ID NO.1:
[0062]
[0063] The 16S rDNA gene sequence of strain W3F is 1463 bp in length.
[0064] After identification, strain W3F was determined to belong to Enterococcus lactis, and was named Enterococcus lactis W3F. Enterococcus lactis W3F was deposited at the China General Microbiological Culture Collection Center, located at No. 1 Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, on November 28, 2024; accession number CGMCC No. 32832.
[0065] Example 4: Safety Study of Enterococcus lactis W3F
[0066] 1. Hemolytic assay
[0067] Hemolysis is an important indicator for evaluating the safety of bacterial strains. W3F was spot-inoculated onto Columbia blood agar plates and incubated at 37°C for 2 days to observe the hemolytic zone. Bacillus cereus ATCC 14579 was used as a positive control. Results are shown below. Figure 4 .
[0068] 2. Antibiotic susceptibility testing
[0069] Antibiotic susceptibility was determined using commercially available antimicrobial susceptibility testing discs. Enterococcus lactis W3F bacterial suspensions were serially diluted with physiological saline to a concentration of 1 × 10⁻⁶. 8 CFU / mL, then take 50μL of the diluted bacterial solution and spread it evenly on mMRS solid medium. After drying, place the antibacterial susceptibility test discs of 14 antibiotics into the plates and incubate at 37℃ for 1 day. Observe the antibacterial effect and measure the diameter of the inhibition zone. The results are shown in Table 2.
[0070] 3) Hemolytic assay results: From Figure 4 It can be seen that the positive control Bacillus cereus ATCC 14579 formed a hemolytic zone and showed hemolysis, while Enterococcus lactis W3F did not form a hemolytic zone and showed no hemolysis. Therefore, Enterococcus lactis W3F can be used safely.
[0071] 4) Results of antibiotic susceptibility testing:
[0072] Table 2. Antibiotic susceptibility of Enterococcus lactis W3F
[0073]
[0074] Antibiotic susceptibility is also an important indicator for evaluating the safety of bacterial strains. In this experiment, 14 common antibiotics, including penicillin (10 μg), ampicillin (10 μg), piperacillin (100 μg), erythromycin, cefazolin (30 μg), cefoperazone (75 μg), amikacin (30 μg), streptomycin (10 μg), cephalexin (30 μg), minocycline (30 μg), tetracycline (30 μg), lincomycin (2 μg), vancomycin (30 μg), and doxycycline (30 μg), all formed inhibition zones on the W3F strain plate. Therefore, Enterococcus lactis W3F is sensitive to all 14 antibiotics and can be used safely.
[0075] Example 5: Gastrointestinal tolerance of Enterococcus lactis W3F
[0076] 1. Test method:
[0077] Strain strain W3F was inoculated into 50 mL of liquid mMRS medium and incubated at 37 °C for 24 h. After centrifugation at 10000 r / min for 10 min, the supernatant was discarded and the precipitate was resuspended in physiological saline to obtain the bacterial suspension.
[0078] 1 mL of bacterial suspension was inoculated into 9 mL of simulated gastric juice, simulated intestinal juice, and mMRS medium at pH 2.5, respectively, and incubated statically at 37°C. The bacterial count at 0 h and 3 h was determined using the mMRS agar plate counting method. The survival rate was used to assess the tolerance of the strain, and the calculation formula is as follows: Survival rate (%) = (Nt / N0) × 100, where N0 (log CFU / mL) and Nt (log CFU / mL) are the bacterial counts at 0 h and 3 h, respectively.
[0079] 2. Results Analysis:
[0080] Table 3. Gastrointestinal tolerance of Enterococcus lactis W3F
[0081] strain name pH 2.5 Simulated gastric juice Simulated intestinal fluid W3F (%) 93.52±0.98 81.95±0.75 88.92±0.44
[0082] High survival rates of probiotics, either directly or after entering the gastrointestinal tract with food, are prerequisites for their ability to exert functions such as lowering blood pressure, blood sugar, and uric acid. As shown in Table 3, *Enterococcus lactis* W3F survived for 3 hours under simulated gastric and intestinal fluid conditions at pH 2.5, with survival rates of 93.52±0.98%, 81.95±0.75%, and 88.92±0.44%, respectively. This demonstrates that the W3F strain provided by this invention exhibits strong gastrointestinal tolerance and significant effects. The strong gastrointestinal tolerance of strain W3F creates the conditions and foundation for its in vivo application in terms of its blood pressure, blood sugar, and uric acid-lowering activities.
[0083] Example 6: Study on the ability of Enterococcus lactis W3F to produce extracellular polysaccharides
[0084] 1. Preparation of sterile fermentation broth: Same as in Example 2.
[0085] 2. Extraction of extracellular polysaccharides: Trichloroacetic acid was added to the sterile fermentation broth to a final concentration of 4%. After standing at 4℃ for 12 h, the mixture was centrifuged at 10,000 r / min for 10 min. The supernatant was added with 2 volumes of anhydrous ethanol, and after standing at 4℃ for 12 h, it was centrifuged at 10,000 r / min for 10 min. The precipitate was reconstituted with sterile deionized water and dialyzed for 24 h, with the water changed every 6 h during this period, to obtain a crude polysaccharide solution. The crude polysaccharide was obtained by vacuum freeze-drying.
[0086] 3. Determination of extracellular polysaccharide content: The extracellular polysaccharide yield was determined using the phenol-sulfuric acid method, with glucose as the standard to plot a standard curve. 100 μL of glucose standard solutions at concentrations of 50, 100, 200, 400, 600, 800, and 1000 mg / L were respectively added to 300 μL of deionized water, followed by 200 μL of 6% phenol. The mixture was then quickly mixed, and 1 mL of concentrated sulfuric acid was added immediately. The mixture was incubated at room temperature for 15 min, and the absorbance at 490 nm was measured. A standard curve was plotted with glucose concentration on the x-axis and absorbance on the y-axis, and the extracellular polysaccharide yield was calculated.
[0087] 4. Results Analysis:
[0088] Extracellular polysaccharides are important metabolites of lactic acid bacteria, possessing not only antibacterial, anti-inflammatory, antioxidant, anti-diabetic, cholesterol-lowering, and immunomodulatory functions, but also interacting with food components to enhance their nutritional, sensory, and rheological properties. They have broad application prospects in functional foods, pharmaceuticals, and biotechnology. Therefore, this invention investigated the extracellular polysaccharide production capacity of *Enterococcus lactis* W3F. The results showed that the extracellular polysaccharide yield of *Enterococcus lactis* W3F was high, at 1097.10 ± 7.32 mg / L, creating conditions for the development of functional foods, health products, and pharmaceuticals.
[0089] Example 7: Study on the antihypertensive, hypoglycemic, and hypouric acid-lowering activities of functional foods prepared by fermentation of Enterococcus lactis W3F
[0090] 1. Preparation of fermented goat milk, fermented fish, and fermented soy milk
[0091] 1) Preparation of fermented goat milk: Add 100mL of fresh goat milk to a glass bottle, sterilize at 100℃ for 15min, and after cooling, add 2.0mL of 2×10 to the sample group in a clean bench. 8 CFU / mL of Enterococcus lactis W3F was added, shaken well, and fermented at 37℃ for 15 hours. After fermentation, the fermented goat milk was placed at 4℃ for 24 hours. 1g of the fermented goat milk that had been left to stand at 4℃ for 24 hours was taken, 5mL of sterile water was added, and the mixture was sonicated for 15 minutes. It was then centrifuged at 8000rpm for 15 minutes at 4℃. The supernatant was transferred to a new centrifuge tube, and the pH was adjusted to 7.0. It was then centrifuged again at 8000rpm for 15 minutes at 4℃. The antihypertensive, hypoglycemic, and hypouric acid-lowering activities of the supernatant were measured. The control group consisted of unfermented raw goat milk containing 2mL of sterile saline solution without Enterococcus lactis W3F.
[0092] 2) Preparation of fermented fish meat: Stun fresh grass carp, remove head, tail, scales, and internal organs. Take a whole piece of back and abdominal muscle, remove the skin, rinse thoroughly with sterile water, drain, and mince in a meat grinder for 2 minutes. Simultaneously, add 50 mL of physiological saline to a glass bottle and sterilize at 100℃ for 15 minutes. After cooling, add 50 g of minced fish meat to a clean bench. Also, add 2.0 mL of 2×10⁻⁶ saline solution to the sample group. 8CFU / mL of Enterococcus lactis W3F was added, shaken well, and fermented at 15°C for 30 hours. After fermentation, the fermented fish was stored at 4°C for 24 hours. 1 g of the fermented fish stored at 4°C was taken, 5 mL of sterile water was added, and the mixture was sonicated for 15 min. It was then centrifuged at 8000 rpm for 15 min at 4°C. The supernatant was transferred to a new centrifuge tube, the pH was adjusted to 7.0, and the mixture was centrifuged again at 8000 rpm for 15 min at 4°C. The antihypertensive, hypoglycemic, and hypouricemic activities of the supernatant were measured. The control group consisted of unfermented raw fish containing 2 mL of sterile saline solution without Enterococcus lactis W3F.
[0093] 3) Preparation of fermented soybean milk: Weigh 50g of plump soybeans, wash them clean, add 450mL of tap water, and grind them in a blender for 2 minutes. Then, add 100mL of the fermented soybean milk to a glass bottle and sterilize at 100℃ for 15 minutes. After cooling, add 2.0mL of 2×10⁻⁶ mol / L slurry to the sample group in a clean bench. 8 CFU / mL of Enterococcus lactis W3F was added, shaken well, and fermented at 37℃ for 15 h. After fermentation, the fermented soy milk was stored at 4℃ for 24 h. 1 g of the fermented soy milk stored at 4℃ was taken, 5 mL of sterile water was added, and the mixture was sonicated for 15 min. It was then centrifuged at 8000 rpm for 15 min at 4℃. The supernatant was transferred to a new centrifuge tube, the pH was adjusted to 7.0, and the mixture was centrifuged again at 8000 rpm for 15 min at 4℃. The antihypertensive, hypoglycemic, and hypouricemic activities of the supernatant were measured. The control group consisted of unfermented raw soy milk supplemented with 2 mL of sterile saline solution without Enterococcus lactis W3F.
[0094] 2. Detection indicators: Uric acid-lowering activity was assessed by the inhibition rate of xanthine oxidase, blood glucose-lowering activity was assessed by the inhibition rates of α-amylase and α-glucosidase, and blood pressure-lowering activity was assessed by the inhibition rate of angiotensin-converting enzyme. For specific measurement methods, please refer to Example 2.
[0095] 3. Results
[0096] Table 4: Study on the antihypertensive, hypoglycemic, and hypouric acid-lowering activities of fermented goat milk, fermented fish, and fermented soy milk
[0097]
[0098] As shown in Table 4, the inhibition rates of xanthine oxidase, α-amylase, α-glucosidase, and angiotensin-converting enzyme in the three unfermented foods ranged from 5.04±1.30% to 10.93±2.78%. After fermentation with *Enterococcus lactis* W3F, the inhibition rates of xanthine oxidase, α-amylase, α-glucosidase, and angiotensin-converting enzyme in the three foods increased to between 27.26±1.12% and 63.71±1.49%, respectively, with increases exceeding 4.9 times. This indicates that fermented goat milk, fermented fish, and fermented soy milk prepared by fermentation with *Enterococcus lactis* W3F all possess strong hypoglycemic, hypotensive, and uric acid-lowering activities.
[0099] The embodiments described above are merely preferred embodiments of the present invention, and not an exhaustive list of all possible implementations of the present invention. Any obvious modifications made by those skilled in the art without departing from the principles and spirit of the present invention should be considered to be included within the scope of protection of the claims of the present invention.
Claims
1. A type of Enterococcus lactis W3F capable of lowering blood pressure, blood sugar, and / or uric acid, characterized in that, It is deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 32832.
2. The Enterococcus lactis W3F as described in claim 1, characterized in that, The Enterococcus lactis W3F was isolated and screened from milk cake from Shilin, Yunnan.
3. The Enterococcus lactis W3F as described in claim 1, characterized in that, The Enterococcus lactis W3F strain exhibits gastrointestinal tolerance.
4. The Enterococcus lactis W3F as described in claim 3, characterized in that, The Enterococcus lactis W3F is a high-extracellular polysaccharide-producing Enterococcus lactis W3F.
5. The use of Enterococcus lactis W3F as described in any one of claims 1-4 and / or the fermentation product of Enterococcus lactis W3F as described in any one of claims 1-4 in the preparation of antihypertensive drugs, hypoglycemic drugs and / or uric acid-lowering drugs.
6. The application of *Enterococcus lactis* W3F as described in any one of claims 1-4 as a starter culture, and *Enterococcus lactis* W3F as described in any one of claims 1-4 and / or its fermentation products as functional additives in the preparation of functional foods or functional health products, characterized in that... The functional foods or functional health products mentioned above are used to lower blood pressure, lower blood sugar, and / or lower uric acid.
7. The application as described in claim 6, characterized in that, The functional foods include any one of fermented milk, fermented soy milk, and fermented fish.
8. A fermented food that can lower blood pressure, blood sugar, and / or uric acid, characterized in that, It is obtained by fermenting food raw materials with Enterococcus lactis W3F as described in any one of claims 1-4.
9. The fermented food according to claim 8, characterized in that, Any one of the fermented milk, fermented soy milk, and fermented fish.
10. The fermented food according to claim 8, characterized in that, When the fermented food is fermented milk, the food ingredient is one or more of goat milk, cow milk, and camel milk; When the fermented food is fermented soy milk, the food ingredient is soy milk; When the fermented food is fermented fish, the food ingredient is fish meat.