Lactobacillus pentosus HS01 for preventing obesity and application thereof

Abstract

The invention discloses lactobacillus pentosus HS01 for preventing obesity and application of the lactobacillus pentosus HS01, and belongs to the technical field of microorganisms. The lactobacillus pentosus HS01 is preserved in China General Microbiological Culture Collection Center (CGMCC), the preservation address is No.3, yard 1, Beichen West Road, Chaoyang District, Beijing, the preservation date is March 3, 2023, and the preservation number is CGMCC No.26736. The lactobacillus pentosus HS01 can be used for preparing the lactobacillus pentosus HS01. The 16SrDNA (Deoxyribose Nucleic Acid) sequence of the lactobacillus pentosus HS01 is as shown in SEQ ID NO. 1. The lactobacillus pentosus HS01 has good gastrointestinal tract tolerance and relatively strong oxidation resistance, can inhibit the growth of escherichia coli and staphylococcus aureus, and has the effects of regulating intestinal flora and relieving obesity and hyperlipidemia symptoms caused by high fat diet. By adopting the lactobacillus pentosus HS01 and the application thereof, the lactobacillus pentosus HS01 can be widely used for developing related probiotic functional products with the functions of inhibiting bacteria, preventing obesity and reducing blood fat.

Description

Technical Field

[0001] The invention relates to the technical field of microorganisms, in particular to Lactobacillus pentosus HS01 for preventing obesity and application thereof. Background Art

[0002] Obesity is defined as abnormal or excessive accumulation of body fat that may impair health. It is usually caused by an imbalance in lipid intake and metabolism, manifested as high body fat and blood lipid levels, and abnormal body fat metabolism. Obesity is spreading around the world at an alarming rate. Over the past 30-40 years, the trend of overnutrition and obesity has been on the rise, and has become a threat to the global public health system.

[0003] Rapid economic development has changed people's dietary structure, with an increase in the consumption of animal-derived foods, refined grains, and highly processed, high-sugar, and high-fat foods, while physical activity has decreased as sedentary behavior has increased. Dietary factors, lack of physical activity, and other individual-level risk factors are intertwined, making this growth trend unlikely to be reversed in the short term. Therefore, it is urgent to prevent or reduce the burden of obesity through multi-channel intervention measures.

[0004] At present, the most common means of obesity intervention are drug therapy, surgery and dietary intervention. However, long-term use of drugs can easily cause gastrointestinal adverse reactions such as nausea, vomiting, and diarrhea; although surgery to reduce gastric capacity can quickly achieve weight loss, the postoperative side effects caused by various risk factors and the possible cumulative complications make many patients daunted.

[0005] Lactic acid bacteria, as the most important type of Gram-positive bacteria among probiotics, are named for their ability to ferment sugars to produce lactic acid. Lactic acid bacteria are not only widely found in meat, dairy products and vegetables, but are also an important part of the oral and intestinal microbial communities of humans and other animals. As natural, safe and side-effect-free beneficial microorganisms, lactic acid bacteria have been widely used in many disease prevention and treatment aspects. In recent years, the role of lactic acid bacteria in improving intestinal flora and host metabolism in metabolic diseases such as obesity has been widely reported. However, the anti-obesity ability and mechanism of action of different types of lactic acid bacteria are different, and there are differences even between different strains of the same type. Therefore, it is of great significance to develop lactic acid bacteria with anti-obesity effects. The present invention provides a Lactobacillus pentosus HS01, which can be widely used in the development of related probiotic functional products with antibacterial, obesity prevention and lipid-lowering functions. Summary of the invention

[0006] The purpose of the present invention is to provide Lactobacillus pentosus HS01 for preventing obesity and its application, which can be widely used in developing related prebiotic functional products with antibacterial, obesity prevention and blood lipid lowering functions.

[0007] To achieve the above-mentioned purpose, the present invention provides Lactobacillus pentosus HS01 for preventing obesity. The Lactobacillus pentosus HS01 is deposited in the General Microbiological Center of China National Committee for the Preservation of Microorganisms, the preservation address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, the preservation date is March 3, 2023, and the preservation number is CGMCC No. 26736.

[0008] Preferably, the 16S rDNA sequence of the Lactobacillus pentosus HS01 is shown as SEQ ID NO.1.

[0009] The present invention also provides the use of the above-mentioned Lactobacillus pentosus HS01 in the preparation of bacterial agents, foods or medicines with antioxidant functions.

[0010] The present invention also provides the use of the above-mentioned Lactobacillus pentosus HS01 in the preparation of bacterial agents, foods or medicines with antibacterial functions.

[0011] The present invention also provides the use of the above-mentioned Lactobacillus pentosus HS01 in the preparation of bacterial agents, foods or medicines capable of preventing obesity and lowering blood lipids.

[0012] Preferably, the bacterial agent includes a bacterial powder preparation or an oral liquid solution.

[0013] Preferably, the food comprises fermented food or health care product.

[0014] Preferably, the medicine comprises powder, tablet or oral solution.

[0015] The advantages and positive effects of the Lactobacillus pentosus HS01 for preventing obesity and its application according to the present invention are:

[0016] 1. The Lactobacillus pentosus HS01 of the present invention has good gastrointestinal fluid tolerance, especially the survival rate in simulated gastric fluid is as high as 95.65%, and the tolerance is excellent.

[0017] 2. The bacterial suspension and fermentation broth of Lactobacillus pentosus HS01 in the present invention both showed strong reducing ability, DPPH scavenging ability and hydroxyl free radical scavenging ability.

[0018] 3. The Lactobacillus pentosus HS01 in the present invention can inhibit the growth of pathogenic bacteria Escherichia coli and Staphylococcus aureus, and also has the effect of regulating intestinal flora and alleviating obesity and hyperlipidemia symptoms caused by a high-fat diet. It can be widely used in the development of related prebiotic functional products with antibacterial, obesity prevention and lipid-lowering functions.

[0019] The technical solution of the present invention is further described in detail below through the accompanying drawings and embodiments. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1This is a colony morphology diagram of Lactobacillus pentosus HS01 in Example 1 of the present invention;

[0021] Figure 2 This is a morphological diagram under a microscope in Example 1 of the present invention;

[0022] Figure 3 This is a growth curve diagram of Lactobacillus pentosus HS01 in Example 1 of the present invention;

[0023] Figure 4 This is a graph showing pH changes in the fermentation culture broth of Lactobacillus pentosus HS01 in Example 1 of the present invention;

[0024] Figure 5 This is a graph showing the weight changes of each group of mice in Example 5 of the present invention;

[0025] Figure 6 This is the organ index diagram in Example 5 of the present invention;

[0026] Figure 7 This is a pathological analysis diagram of mouse liver sections in Example 5 of the present invention, wherein A is a normal group, B is a control group, and C is a Lactobacillus pentosus HS01 group;

[0027] Figure 8 The diversity and uniformity analysis diagram of the intestinal flora of mice in Example 5 of the present invention, wherein A is an observed-species index analysis diagram, and B is a Chao1 index analysis diagram;

[0028] Fig. 9 This is an analysis diagram of the abundance of the mouse intestinal flora in Example 5 of the present invention, wherein A is a bar graph of species abundance at the phylum level, B is a bar graph of species abundance at the class level, and C is a bar graph of species abundance at the order level. DETAILED DESCRIPTION

[0029] The technical solution of the present invention is further described below through the accompanying drawings and embodiments.

[0030] Unless otherwise defined, technical or scientific terms used in the present invention shall have the common meanings understood by one having ordinary skills in the field to which the present invention belongs.

[0031] Collection Information

[0032] Lactobacillus pentose HS01 (Lactobacillus pentose HS01), classification name: Lactobacillus pentosus; preservation unit: General Microbiology Center of China Microbiological Culture Collection Administration, preservation address: No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, preservation number: CGMCC NO: 26736, preservation time: March 3, 2023.

[0033] The culture medium formula used in the embodiment is:

[0034] MRS liquid medium: yeast extract 5 g / L, peptone 10 g / L, beef extract 10 g / L, sodium acetate 5 g / L, glucose 20 g / L, diammonium citrate 2 g / L, K2HPO4·3H2O 2.68 g / L, MnSO4·H2O 0.05 g / L, MgSO4 0.048 g / L, Tween 80 1 mL / L, pH 6.2-6.4, sterilized (121°C, 1×10 5 Pa, 20min).

[0035] MRS solid medium: Add 15 g / L agar to MRS liquid medium, stir well, and sterilize (121°C, 1×10 5 Pa, 20min).

[0036] Preparation of LB liquid medium: tryptone 10g / L, yeast extract 5g / L, sodium chloride 10g / L, distilled water 1L, pH 7.2-7.4, sterilized (115℃, 1×10 5 Pa, 20min).

[0037] Preparation of LB solid medium: Add 15 g / L agar powder to the prepared LB liquid medium, stir evenly, and sterilize (115°C, 1×10 5 Pa, 20min).

[0038] Except for the Lactobacillus pentosus HS01 strain, the other strains in the examples were commercially available.

[0039] Example 1

[0040] Isolation, Screening and Identification of Lactobacillus pentosus HS01

[0041] 1. Screening of lactic acid bacteria

[0042] 1.1 Sample source

[0043] The present invention collects 35 portions of plant fermented foods (pickles, sauerkraut, sour bamboo shoots, etc.) from different regions for subsequent separation, purification and identification of lactic acid bacteria strains.

[0044] 1.2 Isolation and purification of strains

[0045] Under sterile conditions, 1 g of sample was added to 100 mL of MRS liquid culture medium and cultured in a constant temperature shaker at 37°C for 24 h. 1 mL of enriched culture medium was fully mixed with sterile saline using the doubling dilution method and then graded diluted. 10 -4 , 10 -5 , 10 -6The gradient dilutions were spread on MRS solid medium and inverted at 37°C for 48 hours. After the colonies were formed, single colonies of different sizes and shapes were selected and repeatedly streaked on MRS solid medium until the color, size and shape of the colonies on the same plate were consistent. Observe and record the colony morphology, including the shape of the top, side, size, color, transparency and surface roughness of the colonies.

[0046] Select the colonies with fine white round shape, raised middle part, neat edges, smooth and moist, transfer to 10mL MRS liquid medium, culture at 37℃ for 18h, take the bacteria for Gram staining, and observe the strain morphology under a high-power microscope. The rod-shaped Gram-positive bacteria under microscopy are preliminarily determined to be Lactobacillus, and are stored in glycerol tubes and frozen in a -80℃ refrigerator for future use.

[0047] 2. Identification of Lactobacillus pentosus HS01

[0048] 2.1 Colony characteristics

[0049] After culturing Lactobacillus pentosus HS01 on MRS solid medium for 48 hours, the colony diameter is between 0.5 and 2.0 mm, and it is milky white, with a raised round middle, neat edges, and smooth and moist. Figure 1 shown.

[0050] 2.2 Morphology under microscope

[0051] Lactobacillus pentosus HS01 is Gram-positive, round-ended, short rods, single, paired or short chains, without flagella, and does not produce spores, such as Figure 2 shown.

[0052] 2.3 16S rDNA identification

[0053] The Lactobacillus strains initially screened were identified by 16S rDNA gene sequencing. The target genomic DNA was extracted using a DNA extraction kit.

[0054] The PCR experiment of 16S rDNA was carried out using two universal primers, 27F: 5'-AGAGTTTGATCCTGGCTCAG-3' (SEQ ID NO.2) and 1492R: 5'-GGTTACCTTGTTACGACTT-3' (SEQ ID NO.3). The PCR product was sent to a third-party company for sequencing and compared with the NCBI BLAST sequence database to identify the strain at the species level. The results showed that the sequence (as shown in SEQ ID NO:1) had a homology of more than 99% with the 16S rDNA sequence of Lactobacillus pentosus.

[0055] The sequence (5' to 3') of the 16S rDNA of the strain to be identified is as follows (SEQ ID NO.1):

[0056]

[0057] Combined with the sequence alignment results and physiological and biochemical results of strain HS01, it was determined that the screened lactic acid bacteria HS01 was Lactobacillus pentose.

[0058] The Lactobacillus pentosus HS01 in the logarithmic growth phase was inoculated into MRS liquid culture medium at a 2% inoculation rate and cultured at 37°C for 2 hours, and then entered the logarithmic growth phase. After 14 hours of culture, it reached the stable phase. The growth curve of Lactobacillus pentosus HS01 is shown in Figure 2. Figure 3 shown.

[0059] 3. Preparation of Lactobacillus pentosus HS01 bacterial suspension and fermentation supernatant

[0060] The strains stored in the glycerol tube were streaked on the MRS agar plate, and single colonies were picked and expanded in MRS liquid medium for 18 hours. The 2% inoculum was transferred to MRS liquid medium, cultured at 37°C for 18 hours, and after 2-3 generations, centrifuged (4000r / min, 4°C, 10min). The supernatant was filtered through a 0.22μm filter membrane to remove impurities, and the resulting supernatant was the fermentation supernatant of Lactobacillus pentosus HS01; the precipitate was washed 3 times with 0.85% sterile saline and resuspended, and the concentration of the bacterial suspension was uniformly adjusted to 1×10 8 CFU / mL, and the obtained bacterial suspension is Lactobacillus pentosus HS01.

[0061] 24h pH change of Lactobacillus pentose HS01 fermentation broth before centrifugation Figure 4 shown.

[0062] Example 2

[0063] Determination of the tolerance of Lactobacillus pentosus HS01 to simulated gastrointestinal fluid

[0064] Simulated gastric fluid: adjust the pH of 0.1 mol / L potassium phosphate buffer to 2.5, add 10 g of pepsin after sterilization to fully dissolve it, and preheat at 37°C before use.

[0065] Simulated intestinal fluid: Adjust the pH of 0.1 mol / L potassium phosphate buffer to 8.0 and sterilize, add trypsin (10 g / L) and porcine bile salt (3 g / L), mix and fully dissolve to obtain simulated intestinal fluid. Preheat at 37°C before use.

[0066] After activating Lactobacillus pentosus HS01 for 2 to 3 generations, 1 mL of bacterial suspension was inoculated into 9 mL of simulated gastric fluid and simulated intestinal fluid, respectively. After incubation at 37°C for 3 h and 4 h, the number of live bacteria of the strain was determined by plate count method. The tolerance rate was calculated according to formula (1):

[0067]

[0068] Table 1 Tolerance of Lactobacillus pentosus HS01 to simulated gastric and intestinal fluids

[0069]

[0070] The results are shown in Table 1. Lactobacillus pentosus HS01 showed good tolerance in both pH 2.5 simulated gastric fluid and pH 8.0 simulated intestinal fluid containing bile salts, and the tolerance rate in simulated gastric fluid was significantly higher than that of the positive control Lactobacillus rhamnosus ATCC 53103. Therefore, Lactobacillus pentosus HS01 has good resistance to gastrointestinal adverse environment.

[0071] Example 3

[0072] Determination of Antioxidant Capacity of Lactobacillus pentosus HS01

[0073] 1. Determination of DPPH scavenging ability

[0074] 1 mL of bacterial suspension / fermentation liquid was fully mixed with 1 mL of DPPH solution (0.2 mmol / L, prepared with anhydrous ethanol), reacted at room temperature in the dark for 30 min, centrifuged (6 000 r / min, 10 min), and the absorbance value was measured at 517 nm and recorded as As. The absorbance value of the control group measured by physiological saline instead of the sample was recorded as Ac; an equal volume of physiological saline and anhydrous ethanol mixture was used as blank for zero adjustment. The DPPH free radical scavenging rate was calculated using formula (2).

[0075]

[0076] Table 2 DPPH clearance rate of Lactobacillus pentosus HS01

[0077]

[0078] The results are shown in Table 2. Both the bacterial suspension and fermentation supernatant of Lactobacillus pentosus HS01 can effectively scavenge DPPH free radicals, and the scavenging effect of the fermentation broth on DPPH is significantly better than that of the positive control strain. The antioxidant activity of fermentation metabolites is a very important aspect of the antioxidant effect of lactic acid bacteria. There is no significant difference in the scavenging rate of DPPH free radicals between the bacterial suspension of Lactobacillus pentosus HS01 and the control strain.

[0079] 2. Determination of reducing power

[0080] Take 0.5 mL of potassium ferricyanide (1%), PBS (pH 6.6) and bacterial suspension / fermentation liquid, mix well, incubate at 50°C for 20 min and cool quickly. Add 0.5 mL of trichloroacetic acid (10%), centrifuge at 3 000 r / min for 10 min, take 1 mL of supernatant and react with 1 mL of FeCl3 (0.1%) at room temperature for 10 min, and measure the absorbance value A of the sample group at 710 nm. s . Use physiological saline instead of sample to measure the absorbance A c The difference in absorbance between the sample group and the control group is used to characterize the strength of the reducing ability. The reducing power is calculated according to formula 3.

[0081] Reduction power = A s -A c (3)

[0082] Table 3 Reducing ability of Lactobacillus pentosus HS01

[0083]

[0084] After red blood salt (potassium ferrocyanide) is reduced to yellow blood salt, it reacts with Fe3+ to generate Prussian blue, and the amount of Prussian blue generated is used as an indicator to evaluate the reducing ability of the strain. As shown in Table 3, the reducing ability of the fermentation broth of Lactobacillus pentosus HS01 is significantly higher than that of the control positive strain, indicating that it has good antioxidant activity.

[0085] 3. Determination of hydroxyl radical scavenging rate

[0086] Take 1 mL of 1,10-phenanthroline (2.5 mmol / L), PBS (0.02 mmol / L, pH 7.4) and distilled water, mix well, then add 1 mL of bacterial suspension / fermentation broth, FeSO4 (2.5 mmol / L) and H2O2 (20 mmol / L) and incubate at 37°C for 1.5 h. Measure the absorbance of the mixture at 536 nm. b ; Replace distilled water with an equal volume of sample to measure the absorbance A s ; Replace H2O2 with an equal volume of distilled water and measure the absorbance A c The hydroxyl radical scavenging rate was calculated according to formula (4).

[0087]

[0088] Table 4 Hydroxyl radical scavenging rate of Lactobacillus pentosus HS01

[0089]

[0090] The inhibition of hydroxyl radicals by lactic acid bacteria can effectively reduce or alleviate DNA degradation and damage to biological cell membranes, so the hydroxyl radical scavenging rate is an important indicator for evaluating antioxidant capacity. As shown in Table 4, Lactobacillus pentosus HS01 has a high scavenging ability for hydroxyl radicals, and the scavenging rates of its bacterial suspension and fermentation supernatant for hydroxyl radicals are significantly higher than those of the positive control strain.

[0091] Example 4

[0092] Inhibitory Effects of Lactobacillus pentosus HS01 on Escherichia coli and Staphylococcus aureus

[0093] This example is used to illustrate that Lactobacillus pentosus HS01 has the ability to significantly inhibit the growth of Escherichia coli and Staphylococcus aureus.

[0094] The Escherichia coli and Staphylococcus aureus stored in the glycerol tube were transferred to Luria-Bertani (LB) solid medium by streaking method and cultured in a constant temperature incubator at 37°C for 48 hours. A single colony was picked and inoculated into LB liquid medium and cultured in a constant temperature shaker at 37°C for 18 hours. The culture solution was transferred to LB liquid medium again with an inoculation volume of 2% (v / v) and cultured in a constant temperature shaker at 37°C for 18 hours to improve the activity of the strain and to be used for experiments.

[0095] The antibacterial activity of Lactobacillus pentosus HS01 was determined by the internationally accepted agar diffusion method. 7 Add 1% of the indicator bacterial suspension of CFU / mL to the sterilized LB agar medium cooled to about 45°C and mix thoroughly. After the plate solidifies, place the Oxford cup on the surface of the plate, add 200μL of the fermentation supernatant sample, diffuse in the refrigerator at 4°C for 12h, and culture at 37°C for 18h to observe its antibacterial effect. Nisin and Lactobacillus rhamnosus ATCC 53103 were used as positive controls. Measure the diameter of the inhibition zone, and repeat each three times.

[0096] Table 5 Determination of antibacterial ability of Lactobacillus pentosus HS01 fermentation broth

[0097]

[0098]

[0099] As shown in Table 5, the fermentation supernatant of Lactobacillus pentosus HS01 has a good inhibitory effect on both Staphylococcus aureus and Escherichia coli, and is better than the antibacterial effect of the positive control nisin (P<0.05); compared with Lactobacillus rhamnosus ATCC 53103, the fermentation supernatant of Lactobacillus pentosus HS01 has a better antibacterial effect on Escherichia coli (P<0.05), and the antibacterial effects of the two on Staphylococcus aureus are comparable. It can be seen that the metabolites of Lactobacillus pentosus HS01 of the present invention have good antibacterial ability.

[0100] Example 5

[0101] Lactobacillus pentosus HS01 regulates intestinal flora and improves obesity and hyperlipidemia symptoms caused by high-fat diet

[0102] This example is used to illustrate that Lactobacillus pentosus HS01 has a significant effect on improving obesity and hyperlipidemia symptoms caused by a high-fat diet.

[0103] The experimental animal room maintains a temperature of 22±2℃ and a humidity of 55%±5% all year round, and strictly follows the 12h day and 12h night cycle standard. 36 male 4-week-old SPF-grade C57BL / 6J mice were selected and randomly divided into 3 groups after adaptive feeding with basic feed for one week (as shown in Table 6). In the following 8 weeks, except for the normal group fed with basic feed, the other two groups were fed with high-fat feed. During these 8 weeks, the mice in the Lactobacillus pentosus HS01 group were gavaged with Lactobacillus pentosus HS01 (bacterial suspension suspended in sterile saline) on a daily basis. The gavage volume was 200μL / mouse, and the bacterial content of the Lactobacillus pentosus HS01 group was 1×10 9 CFU / mouse; the normal group (CN) and the control group (HFD) were gavaged with 200 μL of normal saline at the same time. All mice were free to eat during the experiment, and their body weights were measured regularly every week. Figure 5 As shown. Figure 5 It can be seen that after 8 weeks of intragastric administration of the Lactobacillus pentosus HS01 described in the present invention, the body weight of mice in the Lactobacillus pentosus group was significantly reduced compared with that in the control group (P<0.05).

[0104] Table 6 Grouping of mice

[0105]

[0106]

[0107] At the end of the experimental period, all mice were fasted but not watered for 12 hours. After anesthesia, blood was collected from the eyeballs, and then the mice were immediately killed by dislocation. The blood was centrifuged at 3500r / min for 15 minutes to obtain serum, which was frozen and stored at -20℃ for later use. The mice were dissected on a sterile operating table, and the kidneys, livers, and spleens of the mice were removed and rinsed with refrigerated sterile saline (0.9%), and the water was absorbed and weighed and recorded; the liver was immersed in 4% paraformaldehyde solution for histopathological sections; the colon feces were collected on ice and then quickly transferred to -80℃ for freezing and storage. The organ index was calculated according to formula (5).

[0108]

[0109] The organ index results are as follows Figure 6 As shown. Figure 6 It can be seen that the kidney and liver indexes of the control group increased significantly, indicating that the high-fat diet caused fat accumulation and enlargement of the organs, while the organ indexes of the mice in the Lactobacillus pentosus HS01 group remained at the same level as those of the normal group mice, indicating that Lactobacillus pentosus HS01 can alleviate organ lesions caused by a high-fat diet.

[0110] After the intervention period, the liver sections of mice were pathologically analyzed (HE staining). Figure 7 As shown in the figure, the mice in the control group had severe fatty liver deformation, the boundaries between liver cell cords were not clear; the liver cells were highly swollen, the cytoplasmic boundaries were no longer obvious, the sinusoids almost disappeared, and large vacuoles (formed by the fusion of many vacuoles after the dissolution of small lipid droplets) appeared in the cytoplasm of most cells. However, no severe fatty liver degeneration was observed in the livers of mice in the Lactobacillus pentosus HS01 group, which was similar to that of the normal group.

[0111] The levels of triglyceride (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LCL-C) were determined according to the method described in the instructions of the kit (commercially purchased).

[0112] Table 7 The lipid-lowering effect of Lactobacillus pentosus HS01

[0113]

[0114]

[0115] The results are shown in Table 7. The high-fat diet caused a significant increase in the levels of triglycerides, total cholesterol, and low-density lipoprotein cholesterol in the serum of the control group of mice, and a significant decrease in the level of high-density lipoprotein cholesterol; however, the Lactobacillus pentosus HS01 intervention group could reduce the levels of triglycerides, total cholesterol, and low-density lipoprotein cholesterol in the serum of mice on a high-fat diet, and increase the level of high-density lipoprotein cholesterol, making the blood lipid metabolism indicators of mice on a long-term high-fat diet closer to those of mice in the normal group. Therefore, Lactobacillus pentosus HS01 has a good effect on lowering blood lipids.

[0116] The total genomic DNA of mouse colon feces was extracted and collected by CTAB / SDS method, and its purity and concentration were detected by 1% agarose gel. The DNA concentration was adjusted to 1 μg / μL (diluted with sterile water) as a substrate template for PCR amplification. The primer sequences are shown in Table 8 (SEQ ID NO.4-5). The product was purified by gel extraction kit and used The sequencing library was generated using a DNA PCR-Free sample preparation kit (Illumina, USA), evaluated using a Qubit@2.0 Fluorometer (ThermoScientific) and an Agilent Bioanalyzer 2100 system, and finally subjected to paired-end sequencing based on the Illumina NovaSeq sequencing platform.

[0117] Table 8 PCR primer sequences

[0118]

[0119] The raw PE data obtained by Illumina NovaSeq sequencing were spliced ​​and quality controlled, and the obtained clean tags were filtered for chimeras to obtain effective data (Effective Tags) that can be used for subsequent analysis. QIIME2 was used to calculate observed_otus and chao1 indices to evaluate species diversity. In order to further explore the differences in community structure between grouped samples, the LEfSe (LDA effect size) statistical analysis method was used to test the significance of differences in species composition and community structure of grouped samples.

[0120] chao1 and observed-species were selected to characterize the diversity and uniformity of species distribution in mouse colon feces samples. The results are shown in Figure 8As shown. Compared with the CN group, all indexes in the HFD group decreased, indicating that the high-fat diet did reduce the species richness and diversity of the intestinal flora of mice. After the intervention of Lactobacillus pentosus HS01, the downward trend of the observedspecies index and chao1 index of the intestinal flora of mice was alleviated, indicating that Lactobacillus pentosus HS01 has the effect of regulating the diversity and richness of the intestinal flora of mice fed a high-fat diet.

[0121] Fig. 9 This is a graph showing the abundance of intestinal flora in mice. Fig. 9 It can be seen that the abundance of Firmicutes in the intestinal flora of mice in the HFD group increased significantly, while the abundance of Bacteroidetes decreased significantly, indicating that the high-fat diet caused intestinal flora disorders in mice. The abundance of Firmicutes and Bacteroidetes in the Lactobacillus pentosus HS01 group was similar to that in the CN group, indicating that Lactobacillus pentosus HS01 alleviated the imbalance of intestinal flora caused by a high-fat diet.

[0122] The above results show that Lactobacillus pentosus HS01 can significantly reduce the body weight and organ index of mice fed a high-fat diet, and has a significant effect on alleviating obesity, liver fatty degeneration, dyslipidemia, and intestinal flora imbalance caused by a high-fat diet, and has great development value.

[0123] Example 6

[0124] Preparation of Antioxidant Fermented Coconut Milk by Lactobacillus pentosus HS01

[0125] Coconut milk was collected from fresh coconuts and filtered to remove insoluble impurities. It was then sterilized (115°C, 1×10 5 Pa, 20min), and then quickly cooled to about 37°C for use. The viable count of Lactobacillus pentosus HS01 was adjusted to 1.0×10 8 CFU / mL, inoculated into the sterilized raw materials at a 4% inoculum, fermented at a constant temperature for 24 h to obtain fermented coconut milk, and then freeze-dried at -45 °C to obtain freeze-dried powder for standby use.

[0126] Take 1g of fermented coconut milk and 1g of unfermented coconut milk (not inoculated with Lactobacillus) freeze-dried powder and dissolve them in 10mL of distilled water respectively, centrifuge (10,000r / min, 10min), take 1mL of supernatant respectively to measure the antioxidant capacity (DPPH scavenging rate and hydroxyl radical scavenging rate) of fermented coconut milk and unfermented coconut milk, the measuring method is the same as Example 3.

[0127] Table 9 Antioxidant capacity of coconut milk fermented by Lactobacillus pentosus HS01

[0128]

[0129] The results are shown in Table 9. It can be seen from Table 9 that after coconut milk was fermented with Lactobacillus pentosus HS01, the DPPH and hydroxyl radical scavenging ability of coconut milk was significantly improved, indicating that Lactobacillus pentosus HS01 can be used to develop antioxidant fermented foods.

[0130] All experiments in all embodiments of the present invention were repeated at least 3 times, and the data are expressed as mean ± standard deviation ( ) indicates that the inter-group comparison was performed using SPSS 25 software, and one-way analysis of variance was performed. Paired t-test was used for analysis within the group. Different letters represent significant differences between the two groups (p<0.05).

[0131] In summary, Lactobacillus pentosus HS01 has good gastrointestinal tolerance, antioxidant ability and antibacterial ability. Intervention in high-fat diet mice can significantly reduce the body weight and organ index of mice, alleviate fatty liver symptoms, dyslipidemia and intestinal flora disorders, showing its excellent performance in preventing obesity and lowering blood lipids, and can be used for the development of antioxidant, antibacterial and blood lipid-lowering products.

[0132] Therefore, the present invention adopts the above-mentioned Lactobacillus pentosus HS01 for preventing obesity and its application, which can be widely used in developing related prebiotic functional products with antibacterial, obesity prevention and blood lipid lowering functions.

[0133] Finally, it should be noted that the above embodiments are only used to illustrate the technical solution of the present invention rather than to limit it. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that they can still modify or replace the technical solution of the present invention with equivalents, and these modifications or equivalent replacements cannot cause the modified technical solution to deviate from the spirit and scope of the technical solution of the present invention.

Claims

1. Lactobacillus pentosus HS01 for preventing obesity, characterized by: The Lactobacillus pentosus HS01 is deposited in the General Microbiology Center of the China Microbiological Culture Collection Administration, the deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, the deposit date is March 3, 2023, and the deposit number is CGMCC No.26736.

2. The Lactobacillus pentosus HS01 for preventing obesity according to claim 1, characterized in that: The 16S rDNA sequence of the Lactobacillus pentosus HS01 is shown in SEQ ID NO.

1.

3. Use of Lactobacillus pentosus HS01 as described in any one of claims 1-2 in the preparation of bacterial agents, foods or medicines with antioxidant function.

4. Use of Lactobacillus pentosus HS01 as described in any one of claims 1 to 2 in the preparation of a bacterial agent, food or medicine with antibacterial function.

5. Use of Lactobacillus pentosus HS01 as described in any one of claims 1 to 2 in the preparation of a bacterial agent, food or medicine for preventing obesity and lowering blood lipids.

6. The use according to any one of claims 3 to 5, characterized in that: The bacterial agent includes a bacterial powder preparation or an oral liquid solution.

7. The use according to any one of claims 3 to 5, characterized in that: The food includes fermented food or health care product.

8. The use according to any one of claims 3 to 5, characterized in that: The medicine includes powder, tablet or oral solution.

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