Lactobacillus plantarum GL-2 and application thereof

Abstract

The invention belongs to the technical field of microorganisms, and particularly relates to lactobacillus plantarum GL-2 and application thereof. The invention provides a strain of lactobacillus plantarum which is preserved in the 8th road of Wuchang District in Wuhan City, Hubei Province on March 15, 2023, and the preservation number is CCTCC M 2023328. The results of the embodiment show that the lactobacillus plantarum GL-2 provided by the invention has the effects of reducing blood sugar and blood fat, and the lactobacillus plantarum GL-2 provided by the invention also has the effects of repairing inflammation damaged organs and regulating intestinal microorganisms.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms, and particularly relates to a Lactobacillus plantarum GL-2 and an application thereof. Background Art

[0002] Lactic acid bacteria are probiotics widely found in the intestines of livestock and poultry, in foods and their products, and in a few clinical samples. They are not only abundant in foods like meat, dairy products, and vegetables, but are also widely distributed in the intestines and oral cavity of humans and other mammals. Lactic acid bacteria, named for their ability to ferment carbohydrates into lactic acid, are Gram-positive and have diverse morphologies, including cocci and bacilli.

[0003] Lactobacillus plantarum is a Gram-positive bacterium that ferments carbohydrates and produces large amounts of lactic acid. Its superior fermentation performance and diverse functionality have garnered widespread attention. Its fermented foods have a long history of consumption worldwide, and it is also a Generally Recognized as Safe (GRAS) edible fungus. Currently, Lactobacillus plantarum is widely used in food fermentation, probiotic preparations, and animal husbandry due to its lactic acid-producing properties and beneficial effects. However, the ubiquitous Lactobacillus plantarum still faces several challenges in practical application. Some Lactobacillus plantarum strains have poor colonization ability in the human intestine, making it difficult to fully exert their beneficial effects. In animal husbandry, some Lactobacillus plantarum strains are poorly adapted to the animal intestinal environment and are unable to effectively regulate the intestinal microecological balance. Furthermore, with growing market demand and increasingly widespread applications, the performance requirements for Lactobacillus plantarum are also becoming increasingly stringent. Therefore, there is an urgent need for a Lactobacillus plantarum strain with more stable performance, more diverse functions, and greater adaptability. Summary of the Invention

[0004] The object of the present invention is to provide a strain of Lactobacillus plantarum GL-2. The Lactobacillus plantarum GL-2 provided by the present invention not only has the effects of lowering blood sugar and blood lipids; the Lactobacillus plantarum GL-2 provided by the present invention also has the effects of repairing organs damaged by inflammation and regulating intestinal microorganisms.

[0005] In order to achieve the above object, the present invention provides the following technical solutions:

[0006] The present invention provides a strain of Lactobacillus plantarum GL-2, which was deposited in the China Center for Type Culture Collection on March 15, 2023, with a deposit number of CCTCC M 2023328.

[0007] The present invention provides a Lactobacillus plantarum GL-2, and the 16S rRNA sequence of the Lactobacillus plantarum GL-2 is shown as SEQ ID NO: 1.

[0008] The present invention provides a method for detecting the above-mentioned Lactobacillus plantarum, which comprises detecting whether a sample contains 16S rRNA with a sequence of SEQ ID NO: 1.

[0009] The present invention also provides a pharmaceutical preparation, the active ingredient of which includes the above-mentioned Lactobacillus plantarum GL-2.

[0010] The present invention provides an application of the aforementioned Lactobacillus plantarum GL-2 or the aforementioned pharmaceutical preparation in the preparation of a hypoglycemic drug.

[0011] The present invention provides an application of the aforementioned Lactobacillus plantarum GL-2 or the aforementioned pharmaceutical preparation in the preparation of a lipid-lowering drug.

[0012] The present invention provides an application of the aforementioned Lactobacillus plantarum GL-2 or the aforementioned pharmaceutical preparation in the preparation of a drug for regulating intestinal flora.

[0013] The present invention provides an application of the aforementioned Lactobacillus plantarum GL-2 or the aforementioned pharmaceutical preparation in preparing a drug for repairing organs damaged by inflammation.

[0014] Preferably, the organ is liver, pancreas, or kidney.

[0015] The present invention also provides a food composition comprising the above-mentioned Lactobacillus plantarum GL-2; the food composition is a beverage, a dairy product or a fermented food.

[0016] Compared with the prior art, the present invention has the following beneficial effects:

[0017] (1) The present invention provides a strain of Lactobacillus plantarum GL-2, which has significant effects on regulating blood sugar and blood lipids. It can play a positive role in lowering blood sugar and blood lipid levels for people with high blood sugar and blood lipids. Long-term intake of Lactobacillus plantarum GL-2 helps stabilize blood sugar and blood lipid indicators, reduce the risk of health problems such as cardiovascular disease caused by abnormal blood sugar and blood lipids, and provide strong protection for physical health.

[0018] (2) The present invention provides a strain of Lactobacillus plantarum GL-2 that has organ repair capabilities. Human organs are susceptible to damage from a variety of factors, and Lactobacillus plantarum GL-2 can participate in the organ repair process. It can promote organ cell regeneration, restore normal organ function, help the body return to a good physiological state, and improve overall health.

[0019] (3) In terms of intestinal microecological regulation, the plantarum Lactobacillus GL-2 provided by the present invention regulates the structure of intestinal microbial communities, inhibits the growth of harmful bacteria, creates a good growth environment for beneficial bacteria, maintains the balance of intestinal microecology, and thus promotes overall health of the body. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0021] Figure 1 This is an analysis diagram of the α-glucosidase inhibition activity assay results.

[0022] Figure 2 This is an analysis diagram of the α-amylase inhibitory activity assay results.

[0023] Figure 3 This is an analysis chart of the reducing ability measurement results.

[0024] Figure 4 This is an analysis chart of the oxygen ion scavenging ability test results.

[0025] Figure 5 This is an analysis chart of the DPPH scavenging ability test results.

[0026] Figure 6 This is an analysis chart of blood glucose changes in each group of mice for six consecutive weeks.

[0027] Figure 7 The graphs show the biochemical index results of mice in each group.

[0028] Figure 8 This is the organ index analysis chart of the liver, pancreas and kidney of each group of mice.

[0029] Figure 9 ACE, Chao1, Simpson and Shannon result analysis charts for each group.

[0030] Figure 10 This is the PCoA analysis diagram.

[0031] Figure 11 Schematic diagram of the composition of the top 10 species at the phylum level.

[0032] Figure 12 Schematic diagram of the composition of the top 20 species at the genus level.

[0033] Deposit information of biological materials

[0034] The strain Lactobacillus plantarum GL-2 of the present invention has the following deposit information:

[0035] Deposit name: Lactobacillus plantarum GL-2;

[0036] Depository: China Center for Type Culture Collection;

[0037] Deposit address: Bayi Road, Wuchang District, Wuhan City, Hubei Province;

[0038] Accession number: CCTCC M 2023328;

[0039] The preservation time is: March 15, 2023. DETAILED DESCRIPTION

[0040] In order to further illustrate the present invention, the technical solution provided by the present invention is described in detail below with reference to the accompanying drawings and embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0041] Unless otherwise specified, the production processes, experimental methods or detection methods involved in the embodiments of the present invention are all conventional methods in the prior art, and their names and / or abbreviations are conventional names in the field and are very clear and unambiguous in the relevant fields of use. Those skilled in the art can understand the conventional process steps based on the names and apply the corresponding equipment to implement them according to conventional conditions or the conditions recommended by the manufacturer.

[0042] The various instruments, equipment, raw materials or reagents used in the embodiments of the present invention are not particularly limited in their sources and are all conventional products that can be purchased through regular commercial channels or prepared according to conventional methods well known to those skilled in the art.

[0043] The composition of the oat and soybean meal culture medium for fermentation of strain GL-2 is 8% oat powder + 2% soybean meal + 6% cabbage juice + 4% soybean meal + 2% oat powder + 6% vegetable juice + 2% maltose + 2g / LK2PO4 + 2g / L sodium acetate + 2g / L ammonium citrate + 0.058g / L MnSO4 + 0.56g / LMgSO4 + 0.03g / L ammonium phenylpropionate + 0.01g / LVB1.

[0044] The strains LLY-002, LLY-003, LLY-007, LLY-008, and LLY-010 involved in Example 2 were all derived from laboratory cultures.

[0045] Example 1

[0046] Obtaining Lactobacillus plantarum GL-2

[0047] The process for obtaining Lactobacillus plantarum GL-2 is as follows: fresh kimchi is diluted with sterile water as a sample, the sample is graded and spread on MRS medium for primary screening, typical colonies are picked and transferred to MRS plates for further purification, single colonies are selected on the purified plates, PCR amplified, and then sequenced. The identified and purified Lactobacillus plantarum is stored in a glycerol tube for future use, thereby obtaining Lactobacillus plantarum GL-2.

[0048] Extract plant lactobacillus GL-2 genomic DNA, PCR amplification (PCR experiment of 16S rRNA), bacterial strain inoculation in MRS liquid nutrient medium, bacterial strain inoculum size is 1%, temperature 37 ℃ of fermentation culture 18h, get the bacterial strain of fermentation and carry out 6000r / min centrifugal 10min, abandon supernatant, with normal saline, unnecessary substratum is cleaned up, resuspend with normal saline again and prepare bacterial suspension, put 4 ℃ for standby use.Get part bacterial suspension and extract bacterial strain genome by test kit requirement and send company to sequence order.

[0049] The sequence of 16S rRNA is shown in SEQ ID NO. 1:

[0050]

[0051] Example 2 In vitro test verification

[0052] The plant lactobacillus GL-2 provided by the present invention was activated and verified by in vitro experiments, mainly from the digestive juice tolerance (artificial gastrointestinal juice), α-glucosidase inhibitory activity, α-amylase inhibitory activity, reducing power determination, O 2- The six in vitro indicators of scavenging ability determination and scavenging rate of free radical DPPH were verified, mainly based on the strain's α-glucosidase inhibitory activity and α-amylase inhibitory activity as the main reference indicators.

[0053] Digestive fluid tolerance (artificial gastrointestinal fluid)

[0054] Artificial gastric juice was prepared by adjusting the pH to 2 with 0.5% physiological saline, adding 0.6 g of pepsin to a volume of 200 ml, and sterilizing and filtering with a 0.22 μm filter membrane (prepared and used immediately).

[0055] The cultured bacterial suspension was centrifuged at 4000 r / min, 4°C, for 10 min. When the OD value was adjusted to 1, the bacterial suspension was added to artificial gastric fluid. After 3 h, the suspension was diluted, smeared, and counted. The survival rate of the gastric fluid was calculated.

[0056] Preparation of artificial intestinal fluid: pH 6.8, take 6.8g of potassium dihydrogen phosphate, add 500ml of water, and dissolve it. Use 0.1mol of sodium hydroxide to adjust the pH to 6.8. Take 10g of pancreatic enzyme, dissolve it in water, mix it, dilute it to 1000ml, filter it with a 0.22mm filter membrane, and set aside.

[0057] The initial concentration of each strain was adjusted to 2×10 9 cfu / ml, and then added to artificial gastric fluid at a ratio of 1:1. The initial bacterial concentration was 1×10 9 cfu / ml, and the plates were smeared after 3 h of culture to determine the bacterial concentration in each group.

[0058] Add the bacteria into artificial intestinal fluid at a ratio of 1:1, culture for 3 hours, spread on the plate, record the bacterial count, and calculate the survival rate of each strain in artificial gastric fluid and intestinal fluid respectively.

[0059] Table 1 Tolerance of artificial gastric juice

[0060]

[0061]

[0062] From the results of artificial gastric juice in Table 1, it can be seen that GL-2 has the highest survival rate in artificial gastric juice, which is 43%. It can be seen that strain GL-2 has relatively good tolerance to gastric juice.

[0063] Table 2 Tolerance to artificial intestinal fluid

[0064] strain 0h colony count (cfu / ml) 3h colony count (cfu / ml) Survival rate (%) LLY-002 <![CDATA[1.6×10 7 ]]> <![CDATA[4.3×10 5 ]]> 2.7 LLY-003 <![CDATA[1.4×10 7 ]]> <![CDATA[5.6×10 6 ]]> 40 LLY-007 <![CDATA[2.7×10 6 ]]> <![CDATA[3.3×10 5 ]]> 12.2 LLY-008 <![CDATA[2.1×10 7 ]]> <![CDATA[5.6×10 6 ]]> 26.7 LLY-010 <![CDATA[2.75×10 6 ]]> <![CDATA[3.3×10 5 ]]> 12 GL-2 <![CDATA[2.15×10 8 ]]> <![CDATA[2.7×10 7 ]]> 12.6

[0065] According to the results of artificial intestinal fluid, the survival rate of GL-2 in artificial intestinal fluid can reach 12.6%. After being cultured in simulated gastric fluid and intestinal fluid, 2.7×10 7 cfu / ml can reach the small intestine.

[0066] α-Glucosidase inhibitory activity assay

[0067] The fermentation supernatant and fermentation broth were reacted, and 150 uL of PBS solution (concentration 0.1 mol / L, pH = 6.8) was added, and then 75 uL of 20 mmol / L 4-nitrophenyl-β-D-pyranoside (PN PG) solution and 25 uL of the sample to be tested were added. The mixture was placed in a 37°C water bath for 10 minutes, 50 uL of 0.2 u / mL α-glucosidase solution was added and the reaction was continued for 10 minutes. 1 ml of 1 mol / L Na2CO3 was added as a reaction termination solution, and the absorbance of the reaction solution was measured at 405 nm. The absorbance is proportional to the free amount of p-nitrophenol PNP. A PBS solution with a pH value of 6.8 and a concentration of 0.1 mol / L was used as a blank control for α-glucosidase and the sample to be tested in the reaction system. After the reaction, the inhibitory activity of lactic acid bacteria on α-glucosidase was calculated.

[0068] The absorbance is measured, and the absorbance is proportional to the amount of free p-nitrophenol PNP. In the reaction system, a PBS solution with a pH of 6.8 and a concentration of 0.1 mol / L is used as a blank control for the α-glucosidase solution and the sample to be tested. Figure 1 As shown,

[0069] Depend on Figure 1 As shown in Figure 2, the fermentation broth of strain GL-2 showed good α-glucosidase activity inhibition ability, while its supernatant had no inhibition ability. Compared with other strains, its fermentation broth had relatively good α-glucosidase activity inhibition ability.

[0070] α-Amylase inhibitory activity assay

[0071] Take 2 mL of α-amylase solution prepared with a pH of 7.0 phosphate buffer at a concentration of 2 mg / mL, add 2 mL of sample solution (i.e., lactic acid bacteria suspension, fermentation supernatant) (blank is replaced by normal saline). After reacting at 37°C for 30 minutes, add 2 mL of 1% soluble starch, react at 37°C for 15 minutes, add iodine solution for color development, and measure the oD value at 660 nm. The results are as follows. Figure 2 As shown,

[0072] Depend on Figure 2As shown, the fermentation broth and supernatant of strain GL-2 have strong α-amylase activity inhibition ability, with the fermentation broth being about 80% and the supernatant being about 90%. It has a good effect in inhibiting the hydrolysis of starch into glucose, especially in controlling blood sugar.

[0073] Reducing power determination

[0074] Take 0.5mL of sample solution, add 0.5mL of PBS solution (concentration is 0.2mol / L, pH=6.6), then add 0.5mL of 1% potassium ferricyanide, and place in a 50℃ water bath for 20min. Take it out and add 0.5mL of 10% trichloroacetic acid, shake well, centrifuge at 3000r / min for 10min, take 11mL of supernatant and add 1mL of 0.1% FeCl3, shake well and let it stand for 10min. Use blank control and measure the OD value at 700nm. The greater the absorbance, the greater the reducing power. The results are as follows Figure 3 shown.

[0075] Depend on Figure 3 As shown in the figure, the fermentation liquid of strain GL-2 has the strongest reducing ability, which shows that strain GL-2 has a strong effect on repairing organ damage.

[0076] Oxygen ion scavenging ability determination

[0077] Take 1mL of sample and add 4mL (pH=8.2) of Tris-HCl-EDTA buffer. After the sample and buffer are incubated at 25℃ for 25min, add 0.5mL of freshly prepared 5mmol / L pyrogallol solution and incubate at 25℃ for 10min. Finally, add 0.1mL of 8mol / L HCl to terminate the reaction. Since the self-oxidation rate of pyrogallol is too fast, it has a relatively stable absorbance value at 420nm. Therefore, after terminating the reaction, measure its absorbance at 420nm. Figure 4 shown.

[0078] Depend on Figure 4 As shown, both the fermentation broth and the supernatant of strain GL-2 have strong oxygen ion scavenging capabilities, with the fermentation broth being about 90% and the supernatant being about 80%, indicating that strain GL-2 has a strong effect in protecting cells from damage caused by superoxide anions.

[0079] DPPH scavenging ability assay

[0080] Take 2mL of sample and add 2mL of 0.2mmol / L DPPH (prepared by dissolving 0.0079g in 100ml of water). Set up sample blank group and control group, replace DPPH with equal amount of methanol and replace sample with equal amount of methanol respectively. Methanol is used as blank control. React in the dark at 37℃ for 20min. After centrifugation at 4000r / min, measure absorbance at 519nm. The results are as follows. Figure 5 shown.

[0081] Depend on Figure 5 As shown, there is no significant difference in the scavenging ability among the strains, and the scavenging ability is all above 80%. The strain GL-2DPPH has a relatively strong scavenging ability.

[0082] Combined with the results of intestinal and gastric juice tolerance, three redox abilities and α-glucosidase and α-amylase inhibition, it can be seen that strain GL-2 can not only effectively regulate blood sugar levels, but also has a significant protective and repair effect on organ damage.

[0083] Example 2

[0084] Testing of Lactobacillus plantarum GL-2 on blood sugar regulation

[0085] The present invention selected 60 5-week-old C57BL / 6J mice and randomly divided them into 12 groups, with 5 mice in each group. After the mice were allowed to grow adaptively for one week, different treatments were applied to each group.

[0086] High-fat and low-fat diabetic mice were fed a high-fat diet for 5 weeks. At week 6, mice were intraperitoneally injected with 200 mg / kg of STZ (streptozotocin) after fasting for 16 hours, divided into two 100 mg / kg injections 72 hours apart. Seventy-two hours after the second injection, blood glucose levels were measured to determine if the 12-hour fasting period was greater than 11.1 mmol / l. A blood glucose level greater than 11.1 mmol / l was considered a successful model.

[0087] After the model was successfully established, mice were fed with a high-fat diet to obtain a high-fat diabetic model, and mice were fed with a low-fat diet to obtain a low-fat diabetic model.

[0088] The diabetic model mice successfully established above were used, and different treatments were applied to each group of model mice. Each treatment consisted of two groups, as follows:

[0089] Healthy group: Two groups of 10 mice in total, which consumed normal feed and had free access to food and water.

[0090] Live bacteria intervention group: fermentation broth was centrifuged at 6000 rap / min for 8 min, the supernatant was removed, and an equal amount of normal saline was added for resuspending;

[0091] In the dead bacteria intervention group, the fermentation liquid was centrifuged at 6000 rap / min for 8 minutes, the supernatant was removed, and an equal amount of physiological saline was added for resuspending, and the cells were inactivated at 100°C for 5 minutes.

[0092] Supernatant intervention group: fermentation broth was centrifuged at 6000 rap / min for 8 min and the precipitate was removed;

[0093] Metformin positive control group: Metformin solution was prepared at a dose of 0.2 g / kg and administered orally;

[0094] Model group: After the model was successfully established, no intervention was performed.

[0095] The model mice treated in the above groups were placed under the same conditions:

[0096] Culture medium: fermented oat and soybean meal cultured milk of strain GL-2; oral administration volume: 200ul; administration dose: metformin 0.2g / kg; intervention dose of Lactobacillus plantarum GL-2: 10 9 CFU / mouse; intervention method: gavage; blood collection from mice, which were fasted for 16 hours without water, and the end of the mouse vein was cut off about 1-2 mm, and the second drop of blood was taken for blood glucose measurement.

[0097] Before the intervention, and during the six-week intervention period, tail vein blood was collected to measure the fasting blood glucose values ​​of mice at a fixed time each week. The measurement process was as follows: at 5 pm the day before the experiment, the mice were placed in a clean cage and fasted, with normal drinking water, and fasted for 16 hours. At 9 am the next day, the glucose tolerance test was started. After weighing each mouse, about 1-2 mm of the end of the mouse's tail was cut off with scissors, and the tail of the mouse was gently squeezed to allow the blood to accumulate into a drop. The first drop of blood was removed with a paper towel, and the second drop of blood was measured with a blood glucose meter. The measured value was determined as the blood glucose value at 0 min. The operation was as gentle as possible so that the mice would not be overly frightened. The fasting blood glucose value measurement results are shown in Table 3 and Figure 6 shown.

[0098] Table 3 Fasting blood glucose measurement results of experimental group and blank group

[0099]

[0100] Results Analysis: Comparison of the test results in Table 3 shows that the blood glucose levels of mice all showed a downward trend, with significant decreases in the fermentation broth group, live bacteria group, supernatant group, and metformin group. The results also showed a significant effect on maintaining blood glucose stability in all groups of mice. Compared with the negative control model group and the positive control metformin group, the live bacteria group showed a superior effect. As can be seen from the results in Table 1, the live bacteria group showed a 51.6% decrease in average blood glucose compared to the initial modeling blood glucose, which was higher than the 42.6% decrease in the positive control metformin group. The average blood glucose levels in the supernatant group and fermentation broth group decreased by 18.7% and 32.8%, respectively. The blood glucose levels of mice in the killed bacteria group and model group showed little change.

[0101] from Figure 6 It can be seen that the blood glucose dispersion of mice in the metformin group of the positive control group tended to be stable after the fifth week of gavage, while the blood glucose dispersion of mice in the live bacteria group, fermentation liquid group and supernatant group decreased and tended to be stable after the fourth week of gavage, one week earlier than the positive control group. The blood glucose dispersion of mice in the model group and the dead bacteria group of the negative control group always changed greatly, indicating that live bacteria and their metabolites have a good effect on maintaining blood glucose stability in diabetes.

[0102] Comprehensive analysis showed that the live bacteria intervention group had a significant effect on maintaining blood sugar stability and lowering blood sugar, and was better than the metformin group, followed by the fermentation broth group and the supernatant group.

[0103] Example 3

[0104] Testing of Lactobacillus plantarum GL-2 on blood lipid regulation

[0105] At the end of the experiment in Example 3, the eyeballs of the mice in each group that had been intervened for 6 weeks were removed and blood was collected. After being placed at 4°C for 3 hours, the blood was centrifuged at 3500 rpm for 10 minutes, and the supernatant was collected. The supernatant was collected again by centrifugation at 3000 rpm for 5 minutes. The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglycerides (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) in the serum were detected. The results are shown in Table 4 and Figure 7 shown.

[0106] Table 4 Serum ALT, AST, TG, TC, HDL-C, LDL-C test results

[0107]

[0108] Results: Comparison of TG and TC levels across groups revealed that mice fed a high-fat diet showed a significant increase in triglyceride levels. Compared to the healthy control group, all groups showed significant differences (P>0.05), except for the live and dead bacteria groups, where TG levels did not reach a significant difference (P<0.05). The dead bacteria group showed the greatest decrease in TG levels, followed by the live bacteria group. TG levels in the other groups decreased slightly compared to the model group, but the decrease was modest. Regarding TC (total cholesterol), TC levels increased slightly in all groups, with the model group showing a significant difference from the healthy control group (P>0.05). The remaining groups showed no significant differences (P<0.05), indicating that each group demonstrated significant control over TC levels. The fermentation broth and supernatant groups showed the greatest effect in controlling TC levels.

[0109] Comparing the HDL-C results of each group, it can be seen that compared with the healthy group, only the HDL-C content of the live bacteria group increased, while the HDL-C content of the dead bacteria group decreased, and the HDL-C content of other groups did not change much, indicating that live bacteria have a good effect on regulating cholesterol levels in the body and reducing cardiovascular diseases.

[0110] In terms of LDL-C control, compared with the healthy group, the LDL-C content in each group increased, among which the model group, fermentation liquid group and metformin group reached significant differences (P>0.05), and there was no significant difference between the other experimental groups and the healthy group (P<0.05). Among them, the supernatant group had the best LDL control effect, followed by the live bacteria group and the dead bacteria group.

[0111] Example 4

[0112] Determination of organ repair effects of Lactobacillus plantarum GL-2

[0113] Long-term hyperglycemia and metabolic disorders activate oxidative stress and inflammatory signaling pathways, prompting the release of pro-inflammatory factors such as TNF-α and IL-6. Inflammation, as a key intermediary, damages the vascular endothelium, induces tissue fibrosis and cell dysfunction, and causes complications in multiple organs such as the heart, kidneys, eyes, and nerves. At the end of the experiment in Example 3, the present invention removed the eyeballs of each group of mice that had been intervened for 6 weeks and collected blood. The liver, pancreas, and kidney were then dissected and washed with physiological saline to remove blood from the tissues. The tissues were then washed with absorbent paper. The weight of the organs of each group of mice was weighed and combined with the body weight to calculate the organ index, as shown in Table 5 and Figure 8 shown.

[0114] Table 5. Body weight and organ index of mice in each group

[0115]

[0116] Result analysis: According to the organ index measurement results of each group, the weight of mice in each group decreased significantly compared with the healthy group. Diabetes can cause weight loss in mice.

[0117] The liver, kidney, and pancreas organ indices of all groups of mice increased to varying degrees. With the exception of the model group, no significant differences were found in the liver organ indices of the other experimental groups compared to the healthy group. This suggests that after oral gavage intervention, liver enlargement caused by secondary inflammation of diabetes in mice was significantly improved. The overall liver index of the dead bacteria group was smaller, while the liver indices of the other groups were similar to those of the healthy group.

[0118] In the comparison of kidney organ indexes, the model group and the sterilized bacteria group showed significant differences compared with the healthy group (P>0.05), while the other groups showed no significant differences (P<0.05). However, compared with the healthy group, the kidney organ indexes of all groups were increased, indicating that the kidney enlargement caused by secondary inflammation of diabetes was significantly improved after oral gavage intervention.

[0119] Comparing the changes in the pancreatic organ index of each group, it was found that the pancreatic organ index of mice in the model group and metformin drug group was significantly different from that of the healthy group (P>0.05), and the pancreatic organ index of mice in other intervention groups had no significant difference from that of the healthy group (P<0.05). Among them, the live bacteria group and supernatant group had the best improvement effect on the pancreas, and the pancreas recovered to the same level as the healthy group.

[0120] Based on the above analysis, the live bacteria group, fermentation liquid group and supernatant group had good improvement effects on repairing organ damage of the liver, kidney and pancreas caused by secondary inflammation of diabetes, among which the live bacteria group performed the best.

[0121] Example 5

[0122] Determination of the regulatory effect of Lactobacillus plantarum GL-2 on intestinal flora

[0123] The cecal contents of the mice in each group that were intervened for 6 weeks at the end of the experiment in Example 3 were analyzed for α-diversity and β-diversity. The species richness of the intervention group and the experimental group were compared by measuring the indicators: Chao1, Ace, Shannon, Simpson, Coverage, and PD_whole_tree, as shown in Table 6. Figure 9 The present invention also performs microbial diversity analysis by measuring the differences in species diversity, and the results are shown in FIG. Figure 10 As shown in the figure, the changes in the composition of the microbial flora of each group were analyzed at the phylum and genus levels.

[0124] Table 6. ACE, Chao1, Simpson and Shannon results for each group

[0125]

[0126]

[0127] Result analysis: According to the comparison of Chao1 and Ace index of each group, the Chao1 and Ace index of the healthy group (H) and the live bacteria group (A) were the largest, indicating that the species richness of these two groups was the largest. The Chao1 and Ace index of the dead bacteria group (D) was the smallest, indicating that its species richness was the smallest, followed by the model group. The Student t test found that the Chao1 and Ace indexes of the model group and the dead bacteria group were significantly different (P>0.05). From the results, it can be seen that the species richness of the intestinal flora of diabetic mice will decrease relatively. Through gavage intervention, the richness of the flora in the intestine of mice will be increased. The intervention effect is in the order of live bacteria group>metformin group>supernatant group>fermentation broth group. The flora richness of the dead bacteria group is lower than that of the model group.

[0128] Comparison of the Shannon index and Simpson index among the groups revealed no significant differences in the Shannon index among the groups (P<0.05). The Simpson index of the live bacteria group (A) and the fermentation broth group (F) was significantly different from that of the healthy group (H) (P>0.05). The species diversity of the intervention groups was slightly lower than that of the healthy group (H). The dead bacteria group had the lowest species diversity, the metformin group had the highest microbial diversity, followed by the supernatant group, the fermentation broth group, and the live bacteria group.

[0129] Comprehensive comparative analysis of Chao1, Ace, Shannon and Simpson indexes showed that gavage intervention can improve the reduction in intestinal flora richness caused by diabetes.

[0130] Analysis of species diversity Figure 10 As shown, the PC1 variation was 7.01% and the PC2 variation was 6.08%. These two principal axes together explained 13.09% of the total variation. The samples in the supernatant group had a larger degree of variation and a lower similarity. The species composition of each intervention group was highly similar, and was significantly different from that of the healthy group (P>0.05).

[0131] according to Figure 11 and Figure 12As shown, the order of bacterial composition is Firmieutes (Firmicutes), Bacteroidota (Bacteroidetes), Deferribacterota (Deferribacterota), Actinobacteriota (Actinobacteria), and Desulfobacterota (Proteobacteria). Compared with the control group, Bacteroidota and Firmieutes showed the greatest differences in each intervention group. Except for the dead bacteria group, where the F / B ratio decreased, the F / B ratios of the other groups increased, consistent with the results of high-fat and high-sugar feeding intervention. Compared with the model group, the F / B ratios of the live bacteria group, supernatant group, fermentation broth group, and metformin group increased, indicating that oral gavage intervention can enhance the microbial composition in the mouse intestine and increase the F / B ratio, thereby enhancing the anti-inflammatory effect and immune function of the mouse body.

[0132] Subordinate level analysis showed that compared with the healthy group, all intervention groups had significantly increased Lactobacillus (Lactobacillus) (P>0.05), with the fermentation broth group having the largest proportion, followed by the metformin group, and the dead bacteria group having the lowest proportion. Compared with the healthy groups, the metformin group had a significantly higher proportion of Biffidobacterium (Bifidobacterium) than the healthy group (P>0.05), while the live bacteria group and fermentation broth group had a slightly higher proportion of Biffidobacterium than the healthy group but did not reach a significant difference (P<0.05). The model group, dead bacteria group, and supernatant group had a significantly lower proportion of Biffidobacterium than the healthy group (P>0.05).

[0133] In summary, metformin is effective in improving the intestinal microbial structure of mice, increasing Lactobacillus and Biffidobacterium, and improving immune disorders and inflammatory responses caused by diabetes. The fermentation broth group and the live bacteria group also have relatively good effects.

[0134] Although the above embodiment provides a detailed description of the present invention, it is only a part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on this embodiment without creativity, and these embodiments all fall within the scope of protection of the present invention.

[0135] Although the above embodiment provides a detailed description of the present invention, it is only a part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on this embodiment without creativity, and these embodiments all fall within the scope of protection of the present invention.

Claims

1. A strain of Lactobacillus plantarum GL-2, characterized in that The plant acid acid bacteria Lactobacillus GL-2 was deposited in the China Center for Type Culture Collection on March 15, 2023, with the deposit number being CCTCC M 2023328.

2. A strain of Lactobacillus plantarum GL-2, characterized in that The 16S rRNA sequence of the Lactobacillus plantarum GL-2 is shown in SEQ ID NO:

1.

3. A method for detecting Lactobacillus plantarum according to claim 1 or 2, characterized in that, The method comprises detecting whether a sample contains 16S rRNA with a sequence of SEQ ID NO:

1.

4. A pharmaceutical preparation, characterized in that The active ingredient of the pharmaceutical preparation includes the Lactobacillus plantarum GL-2 according to claim 1 or 2.

5. Use of the Lactobacillus plantarum GL-2 according to any one of claims 1 to 2 or the pharmaceutical preparation according to claim 4 in the preparation of a hypoglycemic drug.

6. Use of the Lactobacillus plantarum GL-2 according to any one of claims 1 to 2 or the pharmaceutical preparation according to claim 4 in the preparation of a lipid-lowering drug.

7. Use of the Lactobacillus plantarum GL-2 according to any one of claims 1 to 2 or the pharmaceutical preparation according to claim 4 in the preparation of a medicament for regulating intestinal flora.

8. Use of the Lactobacillus plantarum GL-2 according to any one of claims 1 to 2 or the pharmaceutical preparation according to claim 4 in the preparation of a drug for repairing organs damaged by inflammation.

9. The use according to claim 8, characterized in that The organs are liver, pancreas and kidney.

10. A food composition, characterized in that The food composition comprises the Lactobacillus plantarum GL-2 according to any one of claims 1 to 2; and the food composition is a beverage, a dairy product or a fermented food.

Images