A probiotic composition and its application

Abstract

The present invention relates to the field of microbial technology, and in particular to a probiotic composition and its application. The present invention separates three probiotics that can effectively resist oxidation and delay aging from breast milk. The probiotics are natural products with high safety and no toxic side effects. After being compounded into a probiotic composition, it can reduce the level of active oxygen, increase the activity of antioxidant enzymes, reduce the accumulation of malondialdehyde, improve the body's antioxidant capacity, and prolong life by reducing lipofuscin accumulation. The probiotic composition of the present invention broadens the selectivity of antioxidant and anti-aging probiotic products and has broad application prospects.

Description

Technical Field

[0001] The present invention relates to the field of microbial technology, and in particular to a probiotic composition and application thereof. Background Art

[0002] Aging is considered a common correlate of many chronic diseases and functional decline, and is closely related to the homeostasis of the intestinal microbiome. As we age, the intestinal microbiome gradually becomes unbalanced or changes significantly, and probiotics are also considered a strategy to maintain the balance of the intestinal microbiome. Probiotics are microbiome regulators and are defined as "live microorganisms that, when ingested in sufficient amounts, confer health benefits on the host." Supplementation with probiotics can help prevent certain diseases, such as diabetes and hypertension. Probiotics have certain antioxidant properties and can help alleviate diseases such as cardiovascular, inflammatory, aging, and cancer caused by oxidative stress. Therefore, using probiotics to maintain the health of the intestinal microbiome and reduce oxidative stress to prolong life, reduce the risk of disease, and improve the quality of life has gradually become a popular choice.

[0003] With the isolation and screening of new probiotics, rapid and effective in vitro screening and in vivo testing (including safety, efficacy and toxicity studies) are essential, so it is very necessary to establish a low-cost in vivo model that can evaluate the characteristics of probiotics. Caenorhabditis elegans is widely used as an effective model for studying the protective effects of probiotics and host-probiotic interactions. Its advantages include feeding on bacteria, easy reproduction, relatively simple intestinal structure, short lifespan and no ethical issues, which greatly accelerated the research process of probiotics. The most important thing is that it has a complete genome sequence, with about 35~40% homology with the human genome in some genes, and a high degree of conservation with mammals. These advantages make Caenorhabditis elegans an ideal model for characterizing the anti-aging and antioxidant effects of probiotics.

[0004] As an important natural product, probiotics have received increasing attention in prolonging aging and resisting oxidative stress. Probiotics are beneficial microorganisms that exist naturally in the human intestine. They are harmless to the human body and can even help maintain the balance of intestinal flora, enhance immunity, and promote the absorption of nutrients. In contrast, although anti-aging drugs can relieve symptoms in the short term, long-term use may cause side effects and may even cause harm to the body. Studies have shown that a 2-week dietary intervention of polyphenol-rich probiotics in people aged 50 and above can reduce chronic low-grade inflammation, thereby preventing the progression of inflammatory aging.

[0005] Therefore, developing probiotics that can effectively resist oxidation and delay aging has become a technical problem that needs to be urgently solved in this field. Summary of the invention

[0006] In order to solve the above technical problems, the present invention provides a probiotic composition, including: Lactobacillus mucilaginosus B1-10, Lactobacillus paragaseri SYB-PG26 and Lactobacillus rhamnosus B2-1;

[0007] The deposit number of the Lactobacillus reuteri B1-10 is CGMCC No.26166;

[0008] The deposit number of the Lactobacillus paragaseri SYB-PG26 is CGMCC No.30095;

[0009] The deposit number of the Lactobacillus rhamnosus B2-1 is CGMCC No.26167.

[0010] In the present invention, Lactobacillus reuteri B1-10 is isolated from breast milk. The strain was deposited in the General Microbiology Center of the China Microbiological Culture Collection Administration on December 8, 2022. The address of the depository unit is: Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing; Postal Code: 100101, Classification Name: Lactobacillus reuteri Limosilactobacillus reuteri , the deposit number is CGMCC No.26166.

[0011] Lactobacillus paragaseri SYB-PG26 was isolated from breast milk. The strain was deposited in the General Microbiology Center of the China Microbiological Culture Collection Administration on March 22, 2024. The depository address is: Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing; Postal Code: 100101, Classification Name: Lactobacillus paragaseri Lactobacillus paragasseri , the deposit number is CGMCC No.30095.

[0012] Lactobacillus rhamnosus B2-1 was isolated from breast milk. The strain was deposited in the General Microbiology Center of the China Microbiological Culture Collection Administration on December 8, 2022. The address of the depository unit is: Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing; Postal Code: 100101, Classification Name: Lactobacillus rhamnosus Lacticaseibacillus rhamnosus , the deposit number is CGMCC No.26167.

[0013] Preferably, in a unit dose of the probiotic composition, the number of viable Lactobacillus reuteri B1-10 is 1×10 9 CFU or above.

[0014] Preferably, in a unit dose of the probiotic composition, the number of viable bacteria of Lactobacillus paragaseri SYB-PG26 is 1×10 9 CFU or above.

[0015] Preferably, in a unit dose of the probiotic composition, the number of viable Lactobacillus rhamnosus B2-1 is 1×10 9 CFU or above.

[0016] Preferably, the weight ratio of Lactobacillus reuteri B1-10, Lactobacillus paragaseri SYB-PG26 and Lactobacillus rhamnosus B2-1 is 1:(0.8~1.2):(0.8~1.2).

[0017] In some embodiments, the probiotic composition further comprises an excipient.

[0018] In the specific implementation process, the auxiliary materials include but are not limited to oligofructose or inulin, or other auxiliary materials well known to those skilled in the art. In the specific implementation process, the auxiliary materials may also include a probiotic protective agent.

[0019] In a specific implementation process, the probiotic composition is in the form of capsules, tablets, powders, drops, gels or suspensions.

[0020] Furthermore, the present invention provides a product containing the probiotic composition.

[0021] Furthermore, the present invention provides the use of the probiotic composition in preparing products.

[0022] Preferably, the product is used to improve the body's antioxidant capacity, delay aging or prolong life.

[0023] Preferably, the product extends lifespan by reducing lipofuscin accumulation.

[0024] Preferably, the product improves the body's antioxidant capacity by reducing the level of reactive oxygen species (ROS), increasing the activity of antioxidant enzymes, and reducing the accumulation of malondialdehyde (MDA).

[0025] Preferably, the antioxidant enzyme includes at least one of superoxide dismutase (SOD), catalase (CAT), and reduced glutathione (GSH).

[0026] In the specific implementation process, the product is a medicine.

[0027] Compared with the prior art, the present invention has the following beneficial effects:

[0028] The present invention separates three probiotics that can effectively resist oxidation and delay aging from breast milk. Probiotics are natural products with high safety and no toxic side effects. After being compounded into a probiotic composition, they can reduce the level of active oxygen, increase the activity of antioxidant enzymes, reduce the accumulation of malondialdehyde, improve the body's antioxidant capacity, and prolong life by reducing lipofuscin accumulation. The probiotic composition of the present invention broadens the selectivity of antioxidant and anti-aging probiotic products and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 This is a statistical graph of the lifespan of Caenorhabditis elegans.

[0030] Figure 2 It is a statistical graph of the measurement results of body length and width of Caenorhabditis elegans, where A is the measurement result of body width and B is the measurement result of body length.

[0031] Figure 3 This is a statistical graph showing the results of measuring the reproductive capacity of Caenorhabditis elegans.

[0032] Figure 4 It is a statistical graph of the results of the oxidative stress capacity measurement of Caenorhabditis elegans, wherein A is a line graph of the life span of Caenorhabditis elegans, and B is a statistical graph of the average life span of Caenorhabditis elegans.

[0033] Figure 5 This is a graph showing the results of measuring lipofuscin in Caenorhabditis elegans cells, where A is a fluorescence graph and B is the relative fluorescence intensity of lipofuscin.

[0034] Figure 6 This is a diagram showing the results of measuring ROS accumulation in Caenorhabditis elegans cells.

[0035] Figure 7 This is a graph showing the results of SOD activity measurement.

[0036] Figure 8 This is a graph showing the results of MDA content determination.

[0037] Fig. 9 This is a graph showing the results of CAT activity assay.

[0038] Fig.10 This is a graph showing the results of GSH content determination. DETAILED DESCRIPTION

[0039] In order to make the purpose, technical solution and advantages of the present invention clearer, the technical solution of the present invention will be described clearly and completely below. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0040] In the examples provided in this specification, if no specific techniques or conditions are specified, the techniques or conditions described in the literature in this field or the product instructions are used. If the manufacturer of the reagents or instruments used is not specified, they are all conventional products that can be purchased through regular channels.

[0041] The strains and bacterial strains in the following embodiments: wild-type Caenorhabditis elegans ( Caenorhabditis elegans ) and Escherichia coli ( Escherichia coli elegans Genetics Center (University of Minnesota, Minneapolis, MN, USA) and Fujian Shangyuan Biotechnology Co., Ltd. (Fuzhou, Fujian, China), respectively; the deposit number of Lactobacillus reuteri B1-10 is CGMCC No.26166; the deposit number of Lactobacillus paragaseri SYB-PG26 is CGMCC No.30095; the deposit number of Lactobacillus rhamnosus B2-1 is CGMCC No.26167.

[0042] In the following examples, all data are expressed as mean ± standard error. .

[0043] Example 1 Probiotics Culture

[0044] The probiotic strains Lactobacillus mucilaginosus B1-10, Lactobacillus paragaseri SYB-PG26, and Lactobacillus rhamnosus B2-1 were cultured in MRS broth (Beijing Luqiao Technology Co., Ltd.) at 37 °C, and the Escherichia coli OP50 strain was cultured in Luria-Bertani medium (Beijing Luqiao Technology Co., Ltd.) at 37 °C and shaken at 220 rpm for 12 h. All strains were subcultured three times before the experiment. The bacteria were collected by centrifugation at 8000 rpm for 2 min, washed three times with sterile M9 buffer, and centrifuged at 8000 rpm for 2 min to remove the supernatant. The bacterial sludge was adjusted to a final concentration of 0.1 g / ml with M9 buffer for subsequent experiments.

[0045] Example 2 Cultivation and synchronization of Caenorhabditis elegans

[0046] The Caenorhabditis elegans was transferred to NGM medium containing Escherichia coli OP50 and cultured at 20°C to meet the normal growth and development of nematodes for subsequent experiments. Then the Caenorhabditis elegans was synchronized, and the worm plate with enough eggs was selected. The plate was washed with M9 buffer, centrifuged in a 15ml centrifuge tube, and the supernatant was removed to keep 4ml. Lysis buffer (0.5ml of 5% sodium hydroxide, 1ml of 10% sodium hypochlorite) was added, mixed and shaken for 3min, and M9 buffer was added to 15ml, and the supernatant was removed by centrifugation. Repeat this operation 3 times to obtain synchronized eggs. The obtained eggs were placed in M9 buffer overnight so that all the eggs were hatched and kept in the same period, and then transferred to a culture dish inoculated with Escherichia coli OP50 for culture. After culture to the L4 stage, they were used for experiments. Except for reproductive experiments, all L4 stage Caenorhabditis elegans were transferred to NGM medium containing 300mM 5-fluorouracil to prevent interference caused by nematode egg laying.

[0047] Example 3 Life span determination of Caenorhabditis elegans

[0048] The synchronized Caenorhabditis elegans were cultured to the L4 stage and transferred to NGM culture plates covered with Escherichia coli OP50 (OP50 group), Escherichia coli OP50 + Lactobacillus mucilaginosus B1-10 (OP50 + B1-10 group), Escherichia coli OP50 + Lactobacillus paragaseri SYB-PG26 (OP50 + SYB-PG26 group), Escherichia coli OP50 + Lactobacillus rhamnosus B2-1 (OP50 + B2-1 group) and Escherichia coli OP50 + probiotic composition (OP50 + composition group), and probiotics alone or probiotic composition were mixed with Escherichia coli OP50 strains, which were the food source of Caenorhabditis elegans itself, at a weight ratio of 1:3. The total weight of the probiotics alone and the probiotic composition was the same. In the probiotic composition, the weight ratio of Lactobacillus reuteri B1-10, Lactobacillus paragaseri SYB-PG26 and Lactobacillus rhamnosus B2-1 is 1:1:1, and the number of viable bacteria of Lactobacillus reuteri B1-10 is 1×10 9 CFU, the viable count of Lactobacillus paragaseri SYB-PG26 was 1×10 9 CFU, the number of viable bacteria of Lactobacillus rhamnosus B2-1 is 1×10 9 CFU.

[0049] The life span was measured with the time of transfer to the culture plate containing probiotics as d0. The vitality of the worms was observed every 24 hours, and the number of live or dead worms was scored. The criterion for the death of C. elegans was no response to the touch of the platinum wire, no head swing, no pharyngeal pump movement or body stiffness. All culture plates were incubated at 20°C. The worms were transferred to fresh NGM culture plates every 2-3 days to ensure that the worms had enough food to eat. There were 50 worms per 60 mm culture plate, and each group was repeated 3 times for independent experiments. The survival rate of C. elegans was examined using Kaplan-Meier survival analysis, and its average life span was calculated.

[0050] Test results such as Figure 1 As shown, the intervention of probiotics prolonged the lifespan of nematodes. Compared with the lifespan of C. elegans fed with Escherichia coli OP50 alone, the lifespan of nematodes fed with Lactobacillus mucilaginosus B1-10 increased by 11.33% (p<0.001), the lifespan of nematodes fed with Lactobacillus paragaseri SYB-PG26 increased by 2% (p<0.01), and the lifespan of nematodes fed with Lactobacillus rhamnosus B2-1 and the probiotic combination increased by 8.67% (p<0.0001).

[0051] The average lifespan of C. elegans was then calculated, as shown in Table 1. The average lifespans of the OP50+SYB-PG26 group, OP50+B1-10 group, OP50+B2-1 group, and OP50+combination group were 3.04d, 1.21d, 3.09d, and 3.18d longer than those of the OP50 group, respectively. The results showed that probiotic intervention could significantly prolong the lifespan of C. elegans, especially Lactobacillus mucilaginosus B1-10, Lactobacillus rhamnosus B2-1, and the probiotic combination.

[0052] Table 1 Lifespan of Caenorhabditis elegans

[0053]

[0054] Example 4 Measurement of body length and width of Caenorhabditis elegans

[0055] The synchronized Caenorhabditis elegans were cultured to the L4 stage and transferred to NGM culture plates of different groups in Example 3, with at least 20 worms per 60 mm culture plate, and the culture plates were cultured at 20°C. The body length and width of Caenorhabditis elegans in different groups were recorded on the 2nd, 4th, and 8th days, and images of Caenorhabditis elegans adults were taken with a stereo microscope and measured using the Zhiwei Image software. At least 20 worms were used in each group to determine the body size. One-way analysis of variance (ANOVA) was used to analyze the differences between groups, followed by Bonferroni's post-test. Three independent experiments were repeated for each group.

[0056] Test results such as Figure 2As shown in A, for the effect on body width, we can observe that on the 2nd, 4th, and 8th day of adulthood, the intervention of probiotics made the body width of C. elegans smaller than that of the OP50 group, and on the 4th day it was significantly smaller than that of the OP50 group (p<0.05). However, on the 8th day, there was no significant difference in the body width of adults between the groups. The effect on body length was the same as that of C. elegans body width, as shown in Figure 2 As shown in B, at 2d and 4d, the body width of C. elegans was significantly reduced after intervention with Lactobacillus reuteri B1-10 (p<0.01), but there was no significant difference in the C. elegans groups at 8d of adulthood. In general, probiotic intervention slowed down the growth and development of C. elegans compared with the OP50 control group, especially in the early development stage, when the development of body width and length was significantly slowed down.

[0057] Example 5 Determination of reproductive capacity of Caenorhabditis elegans

[0058] The synchronized Caenorhabditis elegans were cultured to the L4 stage and transferred to NGM culture plates of different groups in Example 3, with 20 worms in each group, and each worm was placed on a culture dish, and the culture dish was cultured at 20°C. The Caenorhabditis elegans were transferred to a culture dish containing fresh bacteria every 24 hours, and the previous culture dish was cultured for 2 days. When it was in the L4 stage, the number of offspring was counted using a stereomicroscope. This process was repeated for 6 days until the mother worm stopped laying eggs. One-way analysis of variance (ANOVA) was used to analyze the differences between the groups, followed by Bonferroni's post-test. Three independent experiments were repeated for each group.

[0059] The test results are as follows Figure 3 As shown in the figure, compared with the group fed only with Escherichia coli OP50, the probiotic intervention group produced fewer offspring, and there was no significant difference. Among them, the intervention of Lactobacillus rhamnosus B2-1 produced the least offspring of nematodes. Different probiotic interventions resulted in the number of offspring of most C. elegans being 280-320. Overall, the reproduction time of C. elegans in the probiotic intervention group seemed to be delayed, which may be related to the effect of probiotics on metabolic regulation or growth and development.

[0060] Example 6 Determination of Oxidative Stress Capacity of Caenorhabditis elegans

[0061] The synchronized Caenorhabditis elegans were cultured to the L4 stage and transferred to different groups of NGM culture plates in Example 3, with at least 50 worms per 60 mm plate, and cultured at 20°C for 4 days. Afterwards, the Caenorhabditis elegans in the culture plates of different groups were transferred to blank NGM medium containing 0.03% H2O2, and the live worms were counted every 1 h, and each group was repeated 3 independent experiments. The survival rate of Caenorhabditis elegans was examined using Kaplan-Meier survival analysis.

[0062] The test results are as follows Figure 4 As shown in A, the probiotic intervention group significantly prolonged the lifespan of Caenorhabditis elegans under oxidative stress conditions, and the probiotic combination prolonged the lifespan of nematodes more significantly. Figure 4 In B, the average lifespan of the control group OP50 was 6.92±0.47h, and the average lifespan of the Lactobacillus mucilaginosus B1-10, Lactobacillus paragaseri SYB-PG26, Lactobacillus rhamnosus B2-1 and probiotic combination groups increased to 8.09±0.84h, 9.49±0.45h (p<0.05), 9.00±0.98h (p<0.05) and 10.84±1.16h (p<0.001), respectively. Overall, compared with feeding Escherichia coli OP50 alone, probiotic intervention significantly enhanced the tolerance of Caenorhabditis elegans to oxidative stress and improved the host's adaptability to environmental stress.

[0063] Example 7 Lipofuscin Determination

[0064] Intracellular lipofuscin is an autofluorescent compound that accumulates in senescent cells and is a marker of cell damage during aging. Synchronized C. elegans were cultured to the L4 stage, transferred to NGM culture plates of different groups as in Example 3, and cultured at 20°C for 4 days. C. elegans from each group were collected, washed three times with M9 buffer, and transferred to a slide containing 2% (w / v) agarose. C. elegans were anesthetized with NaN3 and observed under an inverted fluorescence microscope (DMI8, Leica, Germany). At least 15 worms in each group were quantified using ImageJ software.

[0065] The test results are as follows Figure 5 As shown in A, compared with the control group, the intervention of probiotics reduced the accumulation of lipofuscin in C. elegans. Quantitative analysis showed that the accumulated lipofuscin in C. elegans decreased by 58.4%, 33.9%, 72.0% and 77.6% in the Lactobacillus mucilaginosus B1-10, Lactobacillus paragastris SYB-PG26, Lactobacillus rhamnosus B2-1 and probiotic combination groups, respectively ( Figure 5 B). Although the reduction of lipofuscin in the Lactobacillus paragaseri SYB-PG26 group was not significant, the probiotic combination showed the most significant inhibitory effect (p<0.0001).

[0066] Example 8 Determination of Reactive Oxygen Species (ROS) and Antioxidant Enzyme Activity

[0067] Determination of reactive oxygen species (ROS): The synchronized C. elegans were cultured to the L4 stage, transferred to NGM culture plates of different groups as in Example 3, and cultured at 20°C for 4 days. The C. elegans of each group were collected, washed 3 times with M9 buffer, and 100 μL H2DCF-DA (purchased from Nanjing Jiancheng Bioengineering Institute) was added to a final concentration of 100 μM. The worms were incubated at 20°C in the dark for 2 h, then washed 3 times with M9 buffer, transferred to a slide containing 2% (w / v) agarose, anesthetized with NaN3, and observed under an inverted fluorescence microscope (DMI8, Leica, Germany). At least 15 worms in each group were quantified using ImageJ software.

[0068] Determination of antioxidant enzyme activity: The synchronized Caenorhabditis elegans were cultured to the L4 stage, transferred to NGM culture plates of different groups in Example 3, and cultured at 20°C for 4 days. Washed 3 times with M9 buffer to eliminate the influence of bacteria on the surface of Caenorhabditis elegans as much as possible, then centrifuged and the supernatant was removed. C. elegans was broken using a cryogenic freezing grinder (Sevier Biotechnology Co., Ltd.), accompanied by intermittent grinding (grinding 90s, stopping 30s) to prevent enzyme inactivation. The homogenate was centrifuged at 2500rpm and 4°C for 10min, and the supernatant was collected for enzyme activity determination. The enzyme activity determinations included superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA) and reduced glutathione (GSH), which were determined using the corresponding kits (purchased from Nanjing Jiancheng Bioengineering Research Institute). The protein content was determined using the BCA protein analysis kit (purchased from Nanjing Jiancheng Bioengineering Research Institute).

[0069] The above tests were repeated three times for each group. The differences among the groups were analyzed by one-way analysis of variance (ANOVA), followed by Bonferroni's post-test.

[0070] The results of the determination of reactive oxygen species (ROS) are as follows Figure 6 The OP50 group in the control group had the highest ROS accumulation level. The different probiotic intervention groups significantly reduced the accumulation of ROS. The Lactobacillus mucilaginosus B1-10, Lactobacillus paragaseri SYB-PG26, Lactobacillus rhamnosus B2-1 and probiotic combination groups decreased by 9.7% (p<0.05), 29.3% (p<0.0001), 39% (p<0.0001), and 44.5% (p<0.0001), respectively. The probiotic combination had the best effect.

[0071] The test results of SOD, MDA, CAT and GSH are as follows: Figure 7~Figure 10As shown, after feeding with different probiotics, the activities of SOD, MDA, CAT and GSH in C. elegans also increased significantly (p<0.01). Among them, the nematodes fed with Lactobacillus rhamnosus B2-1 had the highest levels of SOD and GSH, which increased by 0.46 times and 1.71 times respectively compared with the control group OP50 group (p<0.0001). The MDA content in the probiotic composition group was the lowest, which was 0.51 times that of the control group OP50 group, and the CAT activity was the highest, which was 1.76 times that of the control group OP50 group (p<0.0001). These results further confirmed that the probiotic composition of the present invention has the activity of enhancing the antioxidant state in C. elegans and reducing oxidative damage.

[0072] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or make equivalent replacements for some of the technical features therein. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. Use of a probiotic composition in the preparation of a medicine, the probiotic composition comprising: Lactobacillus mucilaginosus B1-10, Lactobacillus paragaseri SYB-PG26, and Lactobacillus rhamnosus B2-1; The deposit number of the Lactobacillus reuteri B1-10 is CGMCC No.26166; The deposit number of the Lactobacillus paragaseri SYB-PG26 is CGMCC No.30095; The deposit number of the Lactobacillus rhamnosus B2-1 is CGMCC No.26167; The medicine is used for improving antioxidant capacity or delaying aging.

2. The use according to claim 1, characterized in that: In a unit dose of the probiotic composition, the number of viable Lactobacillus reuteri B1-10 is 1×10 9 CFU or above.

3. The use according to claim 1, characterized in that: In a unit dose of the probiotic composition, the number of viable bacteria of Lactobacillus paragaseri SYB-PG26 is 1×10 9 CFU or above.

4. The use according to claim 1, characterized in that: In a unit dose of the probiotic composition, the number of viable bacteria of Lactobacillus rhamnosus B2-1 is 1×10 9 CFU or above.

5. The use according to claim 1, characterized in that: The weight ratio of Lactobacillus reuteri B1-10, Lactobacillus paragaseri SYB-PG26 and Lactobacillus rhamnosus B2-1 is 1: (0.8~1.2): (0.8~1.2).

6. The use according to claim 1, characterized in that: The probiotic composition further comprises auxiliary materials.

7. The use according to claim 1, characterized in that: The probiotic composition is in the form of capsules, tablets, powders, drops, gels or suspensions.

Images