CN114426938B

CN114426938B - Lactobacillus plantarum CCFM1202 with PCOS relieving function and application thereof

Info

Publication number: CN114426938B
Application number: CN202210120743.7A
Authority: CN
Inventors: 王刚; 贺禹丰; 赵建新; 张灏; 陈卫
Original assignee: Jiangnan University
Current assignee: Jiangnan University
Priority date: 2022-02-08
Filing date: 2022-02-08
Publication date: 2023-09-08
Anticipated expiration: 2042-02-08
Also published as: CN114426938A

Abstract

The application discloses lactobacillus plantarum CCFM1202 with a PCOS relieving function and application thereof, and belongs to the technical field of functional microorganisms. The lactobacillus plantarum CCFM1202 provided by the application has good tolerance to gastrointestinal fluid, and obviously improves abnormal testosterone level, abnormal luteinizing hormone level, abnormal triglyceride level and insulin resistance caused by polycystic ovary syndrome; the intestinal flora diversity reduction caused by polycystic ovary syndrome is remarkably recovered; in addition, lactobacillus plantarum CCFM1202 can relieve reproductive cycle disorder caused by polycystic ovary syndrome, and estrus interval ratio is reduced to 23.33+/-8.17%, and has no obvious difference with a normal group. Meanwhile, the bisphenol F has strong in-vitro adsorption capacity.

Description

Lactobacillus plantarum CCFM1202 with PCOS relieving function and application thereof

Technical Field

The application relates to lactobacillus plantarum CCFM1202 with a PCOS relieving function and application thereof, and belongs to the technical field of functional microorganisms.

Background

In recent years, economic development has led to changes in lifestyle of people in China, unreasonable dietary structure, and a significant increase in prevalence of polycystic ovary syndrome (polycystic ovary syndrome, PCOS) in women of child bearing age. Statistics from the National Institutes of Health (NIH) indicate that 4-10% of women of childbearing age worldwide have polycystic ovary syndrome. The ovulation failure caused by polycystic ovary syndrome severely threatens female fertility and costs over $1000 billion annually worldwide for diagnosing and treating the disease. Thus, there has been an urgent need to address the global burden caused by polycystic ovary syndrome.

The etiology of polycystic ovary syndrome is not clear, and its onset is associated with abnormal sex hormone levels, abnormal metabolism (e.g., insulin resistance), and localized lesions of the ovary. Typical symptoms of polycystic ovary syndrome include menoxenia, hyperandrogenism, and ovarian polycystic lesions. Furthermore, polycystic ovary syndrome is accompanied by a disturbance of intestinal microecology. At the same time, too high androgens and the resulting hirsutism can also lead to patients exhibiting emotional adverse effects. Bisphenol F is a common environmental pollutant and has close connection with the occurrence of abnormal sex hormone level and polycystic ovary syndrome.

Disclosure of Invention

This section is intended to outline some aspects of embodiments of the application and to briefly introduce some preferred embodiments. Some simplifications or omissions may be made in this section as well as in the description of the application and in the title of the application, which may not be used to limit the scope of the application.

In order to overcome the defects in the prior art, the application provides a lactobacillus plantarum (Lactobacillus plantarum) CCFM1202 which is preserved in the microorganism strain collection of Guangdong province in 2021, 12 months and 17 days, wherein the preservation address is 59 th floor 5 building Guangdong province microorganism research institute of Mitsui 100 in Guangzhou, and the preservation number is GDMCC No:62137.

the application also provides a probiotic preparation containing the lactobacillus plantarum CCFM1202.

In one embodiment, the content of Lactobacillus plantarum CCFM1202 in the probiotic preparation is more than or equal to 1×10 ⁶ CFU/mL or ≡1X10. ⁶ CFU/g。

The application also provides a pharmaceutical composition containing lactobacillus plantarum CCFM1202.

In one embodiment, the pharmaceutical composition further comprises a pharmaceutical carrier and/or pharmaceutical excipients.

The application also provides a fermented food which is produced by fermentation using lactobacillus plantarum CCFM1202.

In one embodiment, the fermented food comprises solid food, liquid food, semi-solid food.

In one embodiment, the fermented food comprises dairy products, soy products, fruit and vegetable products, the dairy products comprising milk, sour cream, cheese; the fruit and vegetable products comprise cucumber, carrot, beet, celery and cabbage products.

The application also provides application of lactobacillus plantarum CCFM1202 in preparing a product for relieving polycystic ovary syndrome.

In one embodiment, the product has at least one of the following effects:

(1) Has better tolerance to simulated gastrointestinal fluid;

(2) Restoring the reproduction cycle disorder of the polycystic ovary syndrome rats;

(3) Significantly reducing the testosterone level in polycystic ovary syndrome rats;

(4) Significantly reducing the luteinizing hormone level of polycystic ovary syndrome rats;

(5) Significantly reducing the triglyceride level in polycystic ovary syndrome rats;

(6) The insulin resistance index of the polycystic ovary syndrome rats is obviously reduced;

(7) The reduction of intestinal flora diversity of the polycystic ovary syndrome rats is obviously improved;

(8) Has remarkable in vitro adsorption capacity to bisphenol F.

In one embodiment, the amount of Lactobacillus plantarum CCFM1202 in the product is ≡1×10 ⁶ CFU/mL or ≡1X10. ⁶ CFU/g。

In one embodiment, the product comprises a food or pharmaceutical product.

In one embodiment, the pharmaceutical product comprises lactobacillus plantarum CCFM1202, a pharmaceutical carrier and/or a pharmaceutical adjuvant.

In one embodiment, the pharmaceutical carrier comprises microcapsules, microspheres, nanoparticles, and/or liposomes.

In one embodiment, the pharmaceutical product is in the form of powder, granule, capsule, tablet, pill or oral liquid.

In one embodiment, the food product comprises a solid food product, a liquid food product, a semi-solid food product.

In one embodiment, the food product comprises a dairy product, a soy product, a fruit and vegetable product.

The application has the beneficial effects that:

lactobacillus plantarum CCFM1202 can significantly improve abnormal testosterone levels, abnormal luteinizing hormone levels, abnormal triglyceride levels, and insulin resistance caused by polycystic ovary syndrome; the intestinal flora diversity reduction caused by polycystic ovary syndrome is remarkably recovered; lactobacillus plantarum CCFM1202 can relieve reproductive cycle disorder caused by polycystic ovary syndrome, and estrus interval is reduced to 23.33+/-8.17%, and has no obvious difference from a normal group. In addition, lactobacillus plantarum CCFM1202 has strong adsorption capacity to bisphenol F, has the capacity of relieving bisphenol F toxicity, and has an in vitro adsorption rate of 60 percent after 24 hours. The lactobacillus plantarum CCFM1202 can well endure simulated gastrointestinal fluid, can be used for preparing a pharmaceutical composition and fermented food for relieving polycystic ovary syndrome, improving intestinal flora, recovering physiological cycle and adsorbing bisphenol F, and has very wide application prospect.

Evidence of biological preservation

Lactobacillus plantarum (Lactobacillus plantarum) CCFM1202 deposited at the microorganism strain collection of Guangdong province at 12 months 17 of 2021 at accession number GDMCC No. 59 building 5, guangdong province, university of Mitsui 100, guangzhou City: 62137.

drawings

In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings that are needed in the description of the embodiments will be briefly described below, it being obvious that the drawings in the following description are only some embodiments of the present application, and that other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.

Wherein:

FIG. 1 is a colony morphology of Lactobacillus plantarum CCFM 1202;

FIG. 2 is the effect of Lactobacillus plantarum CCFM1202 on serum Testosterone (Teston) levels in polycystic ovary syndrome rats;

FIG. 3 is the effect of Lactobacillus plantarum CCFM1202 on serum Luteinizing Hormone (LH) levels in polycystic ovary syndrome rats;

FIG. 4 is the effect of Lactobacillus plantarum CCFM1202 on polycystic ovary syndrome rat serum Triglyceride (TG) levels;

FIG. 5 is the effect of Lactobacillus plantarum CCFM1202 on the level of insulin resistance index (HOMA-IR) in polycystic ovary syndrome rats;

FIG. 6 is the effect of Lactobacillus plantarum CCFM1202 on the Shannon index (Shannon) of the intestinal flora of polycystic ovary syndrome rats;

FIG. 7 is an in vitro adsorption of bisphenol F by Lactobacillus plantarum CCFM 1202; and (3) injection: * Indicating that the experimental group has significant differences (p < 0.05) compared to Letrozole building block (Letrozole). a, b represent significant differences in the groups represented by the different letters (p < 0.05).

Detailed Description

In order that the above-recited objects, features and advantages of the present application will become more apparent, a more particular description of the application will be rendered by reference to specific embodiments thereof.

In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present application, but the present application may be practiced in other ways other than those described herein, and persons skilled in the art will readily appreciate that the present application is not limited to the specific embodiments disclosed below.

Further, reference herein to "one embodiment" or "an embodiment" means that a particular feature, structure, or characteristic can be included in at least one implementation of the application. The appearances of the phrase "in one embodiment" in various places in the specification are not necessarily all referring to the same embodiment, nor are separate or alternative embodiments mutually exclusive of other embodiments.

Lactobacillus plantarum CCFM1202 has the following biological properties:

(1) Characteristics of the cells: gram-positive, no spore formation, no movement of bacteria.

(2) Colony characteristics: obvious colonies are formed after aerobic or anaerobic culture for 36 hours, the diameter is between 0.5 and 2mm, the front surface shape is round, the side surface shape is in a protruding shape, the edge is neat, the cream is white, the surface is not transparent, the surface is moist and smooth, and pigment is not generated, see the attached figure 1.

(3) Growth characteristics: culturing in mMRS medium at 37deg.C under aerobic or anaerobic condition for about 16 hr to end logarithmic phase.

(4) Has better tolerance to simulated gastrointestinal fluid;

(5) Can obviously improve the abnormal reproduction cycle of the polycystic ovary syndrome rats;

(6) Can obviously improve abnormal testosterone levels of polycystic ovary syndrome rats;

(7) Can obviously improve abnormal luteinizing hormone level of the polycystic ovary syndrome rat;

(8) Can obviously improve abnormal triglyceride level of the polycystic ovary syndrome rats;

(9) Can obviously improve the insulin resistance of the polycystic ovary syndrome rats;

(10) Can obviously improve the reduction of intestinal flora diversity of rats with polycystic ovary syndrome;

(11) The bisphenol F has remarkable in-vitro adsorption capacity;

the extraction method of the strain comprises the following steps:

isolation and screening of lactic acid bacteria

(l) 1g of fresh faeces from healthy people was taken. Enriching the sample in a medium containing sorbitol GM17 at 35 ℃ for 12 hours;

(2) The enriched sample is coated on a GM17 solid flat plate added with 0.02 percent of olfactory cresol purple after gradient dilution, and is cultured for 24 to 48 hours;

(3) Selecting single bacterial colony with obvious color-changing ring and conforming to basic form of lactobacillus, carrying out flat plate streak purification, screening and separating lactobacillus;

(4) The single colony is cultivated in liquid GM17 culture solution for 24 hours, then gram staining is carried out, and gram positive bacteria are selected for subsequent experiments.

(II) preliminary identification of lactic acid bacteria for fermentation: method for measuring calcium dissolving ring

(l) Culturing the lactobacillus obtained by screening in the step (one) in a liquid sorbitol GM17 culture solution for 24 hours, and centrifuging the L-mL culture at 8000rpm for 2 minutes;

(2) With 0.05M KH ₂ PO ₄ Washing the solution twice;

(3) Resuspending the obtained bacterial sludge, streaking on sorbitol GM17-0.75% CaCO ₃ Culturing for 24 hours on a solid culture medium;

(4) And selecting bacterial colonies which are obvious in calcium dissolving ring, round in convex surface, fine, white in color and free of mycelium, and observing the bacterial colonies by a microscope after gram staining to perform preliminary judgment.

(III) molecular biological identification of lactic acid bacteria for fermentation:

(l) Single bacterial genome extraction:

A. culturing the lactobacillus obtained by screening in the step (II) overnight, taking the cultured overnight bacterial suspension l mL, centrifuging at 10000rpm for 2min in a 1.5mL centrifuge tube, and discarding the supernatant to obtain thalli;

B. washing the thalli with l mL of sterile water, centrifuging at 10000rpm for 2min, and discarding the supernatant to obtain thalli;

C. adding 200 mu LSDS lysate, and water-bathing at 80 ℃ for 30min;

D. 200 mu L of phenol-chloroform solution is added into the bacterial lysate, wherein the phenol-chloroform solution comprises the following components in percentage by volume: chloroform: isoamyl alcohol=25: 24:1, after being mixed reversely and evenly, centrifuging at 12000rpm for 5-10min, and taking 200 mu L of supernatant;

E. adding 400 μl of glacial ethanol or glacial isopropanol into 200uL of supernatant, standing at-20deg.C for 1h, centrifuging at 12000rpm for 5-10min, and discarding supernatant;

F. adding 500 μL70% (volume percent) of ice ethanol to resuspend the precipitate, centrifuging at 12000rpm for 1-3min, and discarding the supernatant;

oven drying at 60 deg.c or naturally drying;

h.50. Mu.LddH 2O was redissolved for PCR;

(2)16S rDNA PCR

A. bacterial 16S rDNA 50. Mu. LPCR reaction System:

10×Taq buffer, 5. Mu.L; dNTP, 5. Mu.L; 27F, 0.5. Mu.L; 1492R, 0.5. Mu.L; taq enzyme, 0.5. Mu.L; template, 0.5 μl; ddH2O, 38. Mu.L.

PCR conditions:

95℃5min；95℃10s；55℃30s；72℃30s；step2-430×；72℃5min；12℃2min；

(3) Preparing 1% agarose gel, mixing the PCR product with 10000×loading buffer, loading 5 μl, running at 120V for 30min, and performing gel imaging;

(4) Sequencing the PCR product of the 16S rDNA, searching and comparing the obtained sequence result in GeneBank by using BLAST, and selecting the sequencing result to identify a newly discovered strain belonging to lactobacillus plantarum, and preserving at-80 ℃ for later use.

Example 1: lactobacillus plantarum CCFM1202 has good tolerance to simulated gastrointestinal fluids

The frozen lactobacillus plantarum CCFM1202 is inoculated in an MRS culture medium, anaerobic cultured for 48 hours at the temperature of 37 ℃, subcultured for 2-3 times by the MRS culture solution, 1mL of the culture solution of the lactobacillus plantarum CCFM1202 is mixed with 9.0mL of artificial simulated gastric fluid (MRS culture medium containing 1% pepsin and pH=2.5) with the pH of 2.5, anaerobic cultured at the temperature of 37 ℃, sampled at 0 hours, 0.5 hours, 1 hour and 2 hours respectively, and subjected to plate colony count by casting culture by using the MRS agar culture medium, and the viable count is measured and the survival rate is calculated.

The survival rate is the ratio of the number of viable bacteria to the number of viable bacteria at the time of sampling in the culture solution to the number of viable bacteria at the time of 0h, expressed in%. 1mL of a culture solution of Lactobacillus plantarum CCFM1202 was added to 9mL of artificial simulated intestinal fluid (MRS medium containing 0.3% bovine bile salt, 1% trypsin, pH=8.0), anaerobic culture was performed at 37℃and samples were taken at 0h, 0.5h, 1h, 2h, 3h and 4h, respectively, plate colony counts were performed by casting culture with MRS agar medium, viable count was determined and survival rate was calculated. The survival rate is the ratio of the number of viable bacteria to the number of viable bacteria at the time of sampling in the culture solution to the number of viable bacteria at the time of 0h, expressed in%. The experimental results are shown in tables 1 and 2. The result shows that lactobacillus plantarum CCFM1202 has better tolerance to artificial gastrointestinal fluids.

TABLE 1 tolerance of Lactobacillus plantarum CCFM1202 in artificial simulated gastric fluid

TABLE 2 tolerance of Lactobacillus plantarum CCFM1202 in artificial simulated intestinal juice

Example 2: lactobacillus plantarum CCFM1202 has no toxic or side effect on SD rats

Inoculating cryopreserved Lactobacillus plantarum CCFM1202 into MRS medium, anaerobic culturing at 37deg.C for 48 hr, centrifuging to collect bacterial cells, and suspending Lactobacillus plantarum CCFM1202 bacterial in sterile physiological saline to give a concentration of 3.0X10 ⁹ CFU/mL of bacterial suspension.

Taking 8 healthy female SD rats with the weight of about 200-250g, after adapting to the environment for one week, taking 1mL of the concentration bacterial suspension for one time every day, and observing for one week, and recording death and weight conditions.

The results of these tests are shown in Table 3. These results indicate that the feeding concentration is 3.0X10 ⁹ CFU/mL of Lactobacillus plantarum CCFM1202 did not significantly affect ratsThe weight is not changed obviously, and no death phenomenon occurs. The appearance of the rat has no obvious pathological symptoms.

TABLE 3 variation of rat body weight and death

Time (Tian)	1	2	3	4	5	6	7
								Body weight (g)	203.1±2.5	205.2±3.1	206.6±3.7	207.7±4.2	208.8±4.6	209.7±3.5	211.3±5.3
Death condition	-	-	-	-	-	-	-

Note that: -: the rat does not die

Example 3: lactobacillus plantarum CCFM12022 has recovery effect on reproduction cycle disorder of polycystic ovary syndrome rats

30 healthy female SD rats weighing 200-250g are taken and are adapted to the environment for 1 week, and randomly divided into 5 groups: blank group (Control), letrozole-making group (Letrozole), dain-35 group (Diane 35), lactobacillus plantarum CCFM1202 group (CCFM 1202), lactobacillus plantarum N34 group (N34, strain is disclosed in golden star, etc.. Lactobacillus plantarum with high cell adhesion and high biofilm formation ability effectively inhibits transcription activity of Campylobacter jejuni virulence factor in mice [ J ]]The food and fermentation industry, 2020,46 (14): 12-18). Each group contained 6 rats, each rat had a lavage concentration of 3.0X10 ⁹ CFU/mL of bacterial suspension 1mL. The grouping and treatment methods of the experimental animals are shown in Table 4:

table 4 experimental animal groups

Week 1-2: normal rats were gavaged with 1% cmc solution and the remaining rats were gavaged with 1mg/kg/bw of letrozole (suspended in 1% cmc) for modeling polycystic ovary syndrome. And simultaneously processed in the manner of table 4.

Week 3-4: the treatment was carried out in the manner as shown in Table 4.

All experimental rats were examined daily for vaginal smears after stopping molding to observe changes in their reproductive cycle. The specific method comprises the following steps: the medical cotton swab is soaked in sterile physiological saline, inserted into the vagina of a rat for about 1cm, gently rotated and then taken out. A drop of sterile physiological saline and a clean glass slide are dripped, and the cotton swab is used for lightly smearing the cotton swab in the area dripped by the sterile physiological saline, and the cotton swab is naturally air-dried. And (3) dyeing the smear by using a Rayleigh dye solution, observing the vaginal cell morphology under an optical microscope after the smear is dried, and recording the reproduction period, wherein the reproduction period is divided into an estrus interval, an estrus prophase, an estrus period and an estrus postphase. Statistics were performed as a percentage of the last five day estrus interval of each rat.

And collecting fresh feces of the rats at the end of the test, freezing the fresh feces at-80 ℃, extracting the genome of the feces, and analyzing the intestinal flora structure by using a second-generation sequencer. At the end of the test, the rats are fasted without water inhibition for 12 hours, and after being drunk by injecting 0.5mL/10g 1% pentobarbital sodium solution into the abdominal cavity, the rats are subjected to heart blood sampling and are killed by a cervical dislocation method. The blood sample is centrifuged for 15min at 3000 Xg and 4 ℃, and the supernatant is taken and frozen at-80 ℃ for measuring relevant serum indexes. After the ovaries are collected, the ovaries are quickly rinsed in pre-cooled normal saline to remove blood, the ovaries are placed in paraformaldehyde for fixation, and the rest ovaries are frozen in liquid nitrogen and transferred to-80 ℃ for freezing.

Estrus cycle is an important index for evaluating the sexual cycle of rats, estrus of normal rats is 4-5 days, estrus later and estrus interval repeatedly appear, and estrus cycle disorder is an important characteristic of PCOS rats. Table 5 is the effect of Lactobacillus plantarum CCFM1202 on the reproductive cycle of polycystic ovary syndrome rats; table 5 records the change of the reproduction cycle of rats in different experimental groups in the last 5 days of animal experiments, and the letrozole modeling ensures that the reproduction cycle of the rats is always in an oestrus interval, and the gastric lavage lactobacillus plantarum CCFM1202 reduces the ratio of the oestrus interval in the reproduction cycle, so that the lactobacillus plantarum CCFM1202 can recover the disorder of the reproduction cycle of the rats with polycystic ovary syndrome, and the effect is stronger than that of lactobacillus plantarum N34.

TABLE 5

Group of	Estrus days (%)
		Control	30.00±10.95*
Letrozole	100.00±0.00
		Diane35	70.00±20.98*
CCFM1202	23.33±8.17*
		N34	96.67±8.17

Example 4: lactobacillus plantarum CCFM1202 reduces testosterone levels in polycystic ovary syndrome rat serum

SD rats were grouped, modeled and processed as in example 3. At the end of the test, rats were fasted without water for 12h, and were anesthetized with 0.5mL/10g of 1% sodium pentobarbital solution, and were heart-sampled. The blood sample is centrifuged for 10min at 3000 Xg and 4 ℃, the supernatant is taken, and the testosterone content in the serum is measured according to the detection method of the kit.

The experimental results are shown in figure 2, the testosterone level in the serum of the letrozole building block rat is obviously increased, and the serum testosterone level of the rat can be obviously reduced by the Lactobacillus plantarum CCFM1202. This demonstrates that lactobacillus plantarum CCFM1202 can significantly improve hyperandrogenism caused by polycystic ovary syndrome and has a stronger effect than lactobacillus plantarum N34.

Example 5: lactobacillus plantarum CCFM1202 reduces the luteinizing hormone level in polycystic ovary syndrome rats

SD rats were grouped, modeled and processed as in example 3. At the end of the test, rats were fasted without water for 12h, and were anesthetized with 0.5mL/10g of 1% sodium pentobarbital solution, and were heart-sampled. The blood sample is centrifuged for 10min at 3000 Xg and 4 ℃, the supernatant is taken, and the content of the xanthogen in the serum is measured according to the detection method of the kit.

The experimental results are shown in figure 3. The serum level of the flavopogenin in the rat serum of the letrozole building module is obviously increased, and the serum luteinizing hormone level of the rat can be obviously reduced by the Lactobacillus plantarum CCFM1202. This demonstrates that lactobacillus plantarum CCFM1202 can significantly improve the imbalance of the levels of xanthogen induced by polycystic ovary syndrome and has a stronger effect than lactobacillus plantarum N34.

Example 6: lactobacillus plantarum CCFM1202 reduces triglyceride levels in polycystic ovary syndrome rats

SD rats were grouped, modeled and processed as in example 3. At the end of the test, rats were fasted without water for 12h, and were anesthetized with 0.5mL/10g of 1% sodium pentobarbital solution, and were heart-sampled. The blood sample is centrifuged for 10min at 3000 Xg and 4 ℃, the supernatant is taken, and the content of triglyceride in serum is measured according to the detection method of the kit.

The experimental results are shown in figure 4. The triglyceride level in the serum of the letrozole building block rat is obviously increased, and the triglyceride level of the serum of the rat can be obviously reduced by the Lactobacillus plantarum CCFM1202. This demonstrates that lactobacillus plantarum CCFM1202 is able to significantly improve the imbalance in triglyceride levels caused by polycystic ovary syndrome and has a greater efficacy than lactobacillus plantarum N34.

Example 7: lactobacillus plantarum CCFM1202 reduces polycystic ovary syndrome rat insulin resistance

SD rats were grouped, modeled and processed as in example 3. At the end of the test, rats were fasted without water for 12h, and were anesthetized with 0.5mL/10g of 1% sodium pentobarbital solution, and were heart-sampled. Blood samples were centrifuged at 3000 Xg at 4℃for 10min, and the supernatant was taken and assayed for fasting blood glucose and fasting insulin levels by the assay method of the kit. An insulin resistance index is calculated from fasting blood glucose and fasting insulin levels.

Insulin resistance index = fasting glycemia (mmol/L) ×fasting insulin (mIU/L)/22.5

The experimental results are shown in figure 5. The insulin resistance index in the serum of the letrozole building block rat is obviously increased, and the lactobacillus plantarum CCFM1202 can obviously reduce the insulin resistance index of the rat. This demonstrates that lactobacillus plantarum CCFM1202 is able to significantly improve insulin resistance induced by polycystic ovary syndrome and has a stronger effect than lactobacillus plantarum N34.

Example 8: lactobacillus plantarum CCFM1202 can restore the reduction of intestinal flora diversity caused by polycystic ovary syndrome

SD rats were grouped, modeled and processed as in example 3. Near the end of the test, fresh faeces from rats were collected and immediately the faeces were frozen in a-80 ℃ freezer. The fecal genome was then extracted and the intestinal flora was analyzed for changes using a second generation sequencing technique.

As shown in figure 6, the experimental results of the flora analysis show that the shannon index of the letrozole making model rat is 5.80+/-0.19, the shannon index of the CCFM1202 group is 6.14+/-0.17, the shannon index of the intestinal flora of the polycystic ovary syndrome rat is obviously reduced, and the intake of the lactobacillus plantarum CCFM1202 can obviously return to the shannon index of the intestinal flora. This shows that the screened lactobacillus plantarum CCFM1202 has the capability of regulating the reduction of intestinal flora diversity caused by polycystic ovary syndrome, and has stronger action and effect than lactobacillus plantarum N34.

Example 9: lactobacillus plantarum CCFM1202 has strong in vitro adsorption capacity for bisphenol F

Purification and activation culture of lactic acid bacteria, inoculating to MRS liquid culture medium according to 1% (v/v) inoculum size, and culturing at 37deg.C for 18 hr. And then centrifuging at 8000r/min for 5min to collect thalli, cleaning the precipitate with normal saline, centrifuging at 8000r/min for 5min, and removing the precipitate to obtain living thalli cells, namely wet thalli. The wet cells were resuspended in 100mg/L bisphenol F physiological saline to a final cell concentration of 1X 10 ⁹ CFU/L. Culturing at 37deg.C with shaking table 120r/min for 24 hr. Taking out bisphenol F culture solution, centrifuging at 4deg.C and 8000rpm for 10minTransferring the supernatant for later use.

The bisphenol F culture supernatant is subjected to HPLC sample introduction pretreatment by adopting a solid phase extraction mode. HPLC analysis conditions are that the instrument model is SHIMADZULC-20A, the detector is a secondary array tube detector, the chromatographic column model is SinoChrom ODS-BP (4.6X250 mm,5 um), the mobile phase is methanol and water (7:3), the column temperature is 40 ℃, the flow rate is 0.5mL/min, the sample injection volume is 10uL, and the detection wavelength of BPF is 277nm. And calculating the adsorption quantity of the lactobacillus to the bisphenol F according to the concentration difference of the bisphenol F before and after adsorption. Average of 3 replicates. As shown in FIG. 7, the in vitro adsorption rate of Lactobacillus plantarum CCFM1202 on bisphenol F for 24h is up to 60% and is much higher than that of Lactobacillus plantarum N34 of the control strain.

While the application has been described with reference to the preferred embodiments, it is not limited thereto, and various changes and modifications can be made therein by those skilled in the art without departing from the spirit and scope of the application as defined in the appended claims.

Claims

1. Lactobacillus plantarum strainLactobacillus plantarum) CCFM1202, deposited at the microorganism strain collection center of Guangdong province at 12 months 17 of 2021, deposited at the 5 th floor Guangdong province microorganism institute of Hirschq 100 th university, guangzhou City under the accession number GDMCC No:62137.

2. a probiotic formulation comprising lactobacillus plantarum CCFM1202 of claim 1.

3. The probiotic preparation according to claim 2, characterized in that the content of lactobacillus plantarum CCFM1202 in the probiotic preparation is not less than 1 x 10 ⁶ CFU/mL or ≡1X10. ⁶ CFU/g。

4. A pharmaceutical composition comprising lactobacillus plantarum CCFM1202 of claim 1.

5. The pharmaceutical composition according to claim 4, wherein the pharmaceutical composition further comprises a pharmaceutical carrier and/or pharmaceutical excipients.

6. A fermented food product, characterized in that it is produced by fermentation using the lactobacillus plantarum CCFM1202 of claim 1; the fermented food contains the lactobacillus plantarum CCFM1202.

7. The fermented food according to claim 6, wherein the fermented food is selected from the group consisting of solid food, liquid food, semi-solid food.

8. Use of lactobacillus plantarum CCFM1202 according to claim 1 for the manufacture of a medicament for alleviating polycystic ovary syndrome.

9. The use according to claim 8, wherein the amount of Lactobacillus plantarum CCFM1202 in the medicament is not less than 1X 10 ⁶ CFU/mL or ≡1X10. ⁶ CFU/g。