Probiotic composition and application thereof

Abstract

The invention relates to the technical field of microbial medicinal preparations, in particular to a probiotic composition and application thereof. The probiotic composition comprises a probiotic composition formed by plant lactobacillus LP45, lactobacillus acidophilus La28, bifidobacterium animalis subsp. Lactis HN019, lactobacillus rhamnosus HN001, plant lactobacillus plantarum CN2018, lactobacillus rhamnosus LR863, lactobacillus rhamnosus LR519, bifidobacterium animalis subsp. Lactis BAL531 and bifidobacterium bifidum B11. The strains have a mutual synergistic effect, jointly regulate the balance of intestinal flora, and have positive effects on cognition, emotion, behaviors and the like of the brain through an intestinal tract-brain axis. According to the technical scheme, the technical problem that the probiotic preparation for improving the brain function is not ideal in action effect can be solved. The scheme not only opens up a new way for the application of the probiotics in the field of neuroscience, but also provides powerful support for brain health maintenance and disease treatment.

Description

Technical Field

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

[0002] Probiotics primarily affect brain function indirectly or directly through various means, including regulating the balance of intestinal flora, improving intestinal function, promoting nutrient digestion and absorption, regulating immune function, and preventing and improving certain diseases. The gut-brain axis, a core mechanism connecting the intestinal microbiome with the central nervous system, enables probiotics to positively influence aspects of the brain, including cognition, emotion, and behavior, by influencing the composition of intestinal microbes, the production of metabolites, and immune signaling.

[0003] Probiotics primarily improve brain function in the following ways: Probiotics can improve cognitive performance, including memory, attention, and learning ability. This may be related to probiotics regulating intestinal microbial metabolism, reducing inflammatory factors, and promoting neurotransmitter synthesis. Studies have shown that probiotics can regulate levels of neurotransmitters such as serotonin in the brain, thereby alleviating mood disorders such as anxiety and depression. Furthermore, probiotics can further protect the brain from damage by improving intestinal barrier function and reducing the penetration of harmful substances. Probiotics may improve sleep quality by regulating the synthesis and secretion of melatonin. Melatonin is an important sleep-regulating hormone, and its levels are affected by the intestinal microbiome. In response to stress, probiotics can modulate the activity of the hypothalamic-pituitary-adrenal axis (HPA axis), reducing stress responses and thereby enhancing an individual's ability to cope with stress.

[0004] Although many studies have shown that probiotics may have positive effects on cognitive function, mood disorders, sleep quality, and stress resistance, the drugs reported in the prior art to improve brain function still have shortcomings in their effects, and the effect on improving brain function is still limited. The combined use of multiple probiotics, the so-called "multi-strain probiotic preparations", has shown a broader potential in improving brain function than a single strain. This strategy is based on the fact that different strains may have complementary mechanisms of action and can have a positive impact on the gut-brain axis from multiple angles. Although "multi-strain probiotic preparations" theoretically have many potential health benefits, especially in improving brain function, their application also faces a series of challenges and problems. Not all strains are suitable for combined use, and current research on which specific strain combinations are most beneficial for specific health goals (such as improving brain function) is not sufficient. There may be complex interactions between different strains, including synergistic or antagonistic effects, which increases the difficulty of designing effective formulations.

[0005] It can be seen that although probiotics have broad application prospects and huge market potential in improving brain function, they still face many challenges and defects in the development of related drugs. In order to overcome these challenges, it is necessary to strengthen the scientificity and rigor of strain screening in the future, and conduct in-depth research on the interaction mechanism between intestinal microorganisms and the brain to determine microbial preparations suitable for clinical application of multiple probiotics for improving brain function. Summary of the Invention

[0006] The present invention aims to provide a probiotic composition to solve the technical problem that the probiotic preparations for improving brain function in the prior art have unsatisfactory effects.

[0007] In order to achieve the above object, the present invention adopts the following technical solutions:

[0008] A probiotic composition comprising probiotics: Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28;

[0009] Preferably, the ratio of the viable counts of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 is 1:1-4:1;

[0010] Preferably, the ratio of the viable cell counts of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 is 2:1.

[0011] Furthermore, a probiotic composition further comprises probiotics: Bifidobacterium animalis subsp. lactis HN019 and Lactobacillus rhamnosus HN001;

[0012] Preferably, the ratio of the viable cell counts of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. HN019, and Lactobacillus rhamnosus HN001 is 1-4:1:1:1;

[0013] Preferably, the ratio of the viable cell counts of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. lactis HN019, and Lactobacillus rhamnosus HN001 is 4:1:1:1.

[0014] Furthermore, a probiotic composition further comprises probiotics: Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519 and Bifidobacterium animalis subsp. lactis BAL531;

[0015] Preferably, the ratio of the viable cell count of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. lactis HN019, Lactobacillus rhamnosus HN001, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531 is 4:1:1:1:1:1:1:1:1.

[0016] Furthermore, a probiotic composition further comprises probiotics: Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519 and Bifidobacterium animalis subsp. lactis BAL531;

[0017] Preferably, the ratio of the viable cell counts of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519 and Bifidobacterium animalis subsp. lactis BAL531 is 4:1:1:1:1:1:1.

[0018] Furthermore, a probiotic composition further includes probiotics: Bifidobacterium bifidum B11;

[0019] Preferably, the ratio of the viable cell counts of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, Bifidobacterium animalis subsp. lactis BAL531, and Bifidobacterium bifidum B11 is 4:1:1:1:1:1:1:1.

[0020] This technical solution also provides an application of a probiotic composition in the preparation of a preparation for improving brain function.

[0021] Preferably, the improvement of brain function is at least manifested in improvement of memory ability and / or improvement of cognitive dysfunction.

[0022] Furthermore, the preparation further comprises at least one of a sweetener, a pigment, a dietary fiber supplement, a micronutrient, and a flavor;

[0023] The dosage form of the preparation is any one of lyophilized powder, enteric-coated capsules, chewable tablets or liquid suspension;

[0024] The total number of viable probiotic bacteria per unit dose is 1×10 9 CFU to 5×10 11 CFU.

[0025] Furthermore, the preparation is prepared from the following raw materials in parts by mass: 20-30 parts of inulin, 15-25 parts of isomalt, 5-15 parts of isomaltooligosaccharide, 5-10 parts of fructooligosaccharide, 1-5 parts of stachyose and 10-40 parts of the probiotic composition.

[0026] Furthermore, the composition of the probiotic composition is as follows:

[0027] It is composed of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 in a ratio of 1:1-4:1 in terms of viable bacteria count, preferably 2:1 in terms of viable bacteria count;

[0028] Or, it is composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. HN019, and Lactobacillus rhamnosus HN001 in a ratio of 1-4:1:1:1 in terms of viable cell count, preferably, the ratio of 4:1:1:1;

[0029] or, composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. lactis HN019, Lactobacillus rhamnosus HN001, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531 in a ratio of 4:1:1:1:1:1:1:1:1;

[0030] or, composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531 at a ratio of 4:1:1:1:1:1:1 in terms of viable cell count;

[0031] Or, it is composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, Bifidobacterium animalis subsp. lactis BAL531 and Bifidobacterium bifidum B11 with a ratio of 4:1:1:1:1:1:1:1 in terms of live cell count.

[0032] The technical principle of this solution is:

[0033] The present invention focuses on the field of microbial pharmaceutical preparations, particularly the use of probiotic compositions in improving brain function. The gut-brain axis theory emphasizes the complex interaction between the gut microbiome and the central nervous system. Gut microbes not only affect intestinal health but also profoundly influence the brain's cognitive, emotional, and behavioral functions through pathways such as metabolites, neurotransmitters, immune factors, and endocrine signaling. The core of the present invention lies in screening and formulating a series of probiotic strains to achieve optimal gut microbiome regulation. These strains include, but are not limited to, Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium animalis subsp. lactis HN019, Lactobacillus rhamnosus HN001, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, Bifidobacterium animalis subsp. lactis BAL531, and Bifidobacterium bifidum B11. Each strain possesses unique biological properties, such as regulating the balance of the intestinal microbiome, inhibiting the growth of harmful bacteria, promoting nutrient absorption, modulating immune responses, and synthesizing neuroactive substances.

[0034] Extensive experimental research has revealed that Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 can synergistically improve brain function. The addition of Bifidobacterium animalis subsp. lactis HN019 and Lactobacillus rhamnosus HN001 further enhances the effectiveness of the probiotic composition. Furthermore, the addition of strains such as Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863 and LR519, and Bifidobacterium animalis subsp. lactis BAL531 further strengthens the probiotic composition's efficacy in improving learning and memory, and alleviating cognitive impairment.

[0035] In addition, the probiotic composition is supplemented with a variety of prebiotic ingredients, including inulin, isomalt, isomaltooligosaccharide, oligofructose, and stachyose. These prebiotics provide the probiotics with a rich supply of nutrients, aiding their growth, reproduction, and colonization in the intestines, thereby enhancing their beneficial effects. Furthermore, prebiotics are fermented and utilized by beneficial bacteria in the intestines, producing metabolites such as short-chain fatty acids, which are important for maintaining intestinal health, regulating intestinal pH, and promoting intestinal motility.

[0036] The beneficial effects of this program are:

[0037] (1) Significantly improves cognition and memory: Through classic animal behavioral tests such as the Morris water maze, the platform jumping test, and novel object recognition, the present invention demonstrates the significant effects of the probiotic composition in improving the cognition and memory abilities of experimental animals. These improvements may be closely related to the mechanisms by which probiotics regulate intestinal microbial metabolism, reduce inflammatory factors, promote neurotransmitter synthesis, and enhance neural connectivity in the brain.

[0038] (2) Synergistic and multi-target effects: The probiotic composition of the present invention exhibits significant synergistic effects, whereby different strains interact with each other to enhance regulatory effects on the gut-brain axis. This multi-targeted mode of action not only improves therapeutic efficacy but also reduces the risk of side effects that may be associated with a single strain.

[0039] (3) Personalized formulation and customized treatment: Based on a deep understanding of the biological characteristics of different strains, the present invention provides a variety of probiotic formulations to meet the personalized treatment needs of different populations and disease states. This customized treatment strategy is expected to achieve more precise and effective brain function improvement in the future.

[0040] (4) Safety and long-term benefits: All strains are derived from nature and have undergone rigorous screening and safety assessments to ensure product safety. The use of the probiotic composition of the present invention not only helps maintain intestinal microecological balance but may also have a positive impact on preventing neurodegenerative diseases, enhancing immunity, and improving quality of life.

[0041] In summary, this invention achieves significant improvements in brain function by deeply exploring the mechanisms of the gut-brain axis and selecting and formulating probiotic strains. This innovative achievement not only opens up new avenues for the application of probiotics in neuroscience but also provides strong support for future efforts to maintain brain health and treat diseases. DETAILED DESCRIPTION

[0042] The following examples are intended only to more clearly illustrate the technical solutions of the present invention and are therefore provided as examples only and are not intended to limit the scope of protection of the present invention. The specific embodiments listed herein are merely exemplary of the present invention and are not intended to limit the scope of protection of the present invention. For those skilled in the art, any equivalent modifications and substitutions made to the embodiments described below are also within the scope of the present invention. Therefore, equivalent transformations and modifications made without departing from the spirit and scope of the present invention should be encompassed within the scope of the present invention. To better illustrate the present invention, numerous specific details are provided in the following specific embodiments. Those skilled in the art should understand that the present invention can be implemented without certain specific details. In other embodiments, methods, means, equipment, and steps well known to those skilled in the art are not described in detail in order to highlight the main purpose of the present invention. Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those commonly understood by those skilled in the art. Unless otherwise specified, the units used in this specification are international standard units, and the numerical values and numerical ranges appearing in the present invention should be understood to include the inevitable systematic errors in industrial production.

[0043] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods; the materials, reagents or instruments used without indicating the manufacturer are all reagents and materials that can be obtained from commercial channels. If no specific conditions are specified in the examples, they are all carried out according to conventional conditions or conditions recommended by the manufacturer. At the same time, the present invention does not limit the sources of the raw materials used. Unless otherwise specified, the raw materials used in the present invention are all common commercial products in the field of this technology. Unless otherwise specified, the "ratios" referred to in the following examples are all ratios by mass.

[0044] The following is a brief description of the strains used in this patent:

[0045] Lactobacillus plantarum LP45 is a microorganism in the prior art and is commercially available. References and patents include: Li Sitong, "Mechanisms of Lactobacillus and Bifidobacterium in Relieving Constipation," Chinese Journal of Microecology, Vol. 35, No. 7, July 2023; CN112094790B, "A Live Lactobacillus plantarum LP45 Preparation for Regulating Intestinal Flora and Its Applications"; and CN103642716B, "A Lactobacillus plantarum and Its Applications (Accession No. CGMCC NO. 8072).

[0046] Lactobacillus plantarum CN2018 is a prior art microorganism that can be obtained commercially. Reference patent: CN102174450B, a Lactobacillus plantarum strain for resisting Helicobacter pylori infection and its uses (accession number: CGMCC No. 4286).

[0047] Lactobacillus acidophilus La28 is a microorganism in the prior art and is commercially available. References and patents include: Li Sitong, "Mechanisms of Lactobacillus and Bifidobacterium in Relieving Constipation," Chinese Journal of Microecology, Vol. 35, No. 7, July 2023; CN113893267B, "New Application of Lactobacillus acidophilus La28 in Anti-Influenza Virus Treatment"; and CN106011006A, "A Strain of Lactobacillus acidophilus La28 with Immunomodulatory and Anti-Allergic Functions and Its Applications (Deposit Number: CGMCC No. 11506).

[0048] Lactobacillus rhamnosus LR863 is a prior art microorganism that is commercially available. Reference patent: CN108048347B Lactobacillus rhamnosus, Lactobacillus rhamnosus preparations, and uses thereof; deposited with the General Microbiology Center of the China Culture Collection Administration (CGMCC No. 14410).

[0049] Bifidobacterium animalis subsp. lactis HN019 is a microorganism currently available and commercially available. It has been widely used in commercial activities. For example, see patents CN114568701B, Compositions containing oligofructose and probiotics and their use; and CN1264422A, Lactic acid bacteria for enhancing immunity (this strain is deposited with the Australian Government Analytical Laboratory (AGAL) under the accession number NM97 / 09513 and the American Type Culture Collection under the accession number ATCC SD5674).

[0050] Lactobacillus rhamnosus HN001 is a microorganism known in the art and commercially available. It has been widely used in commercial activities. For example, see patents: CN118058469A: A composite probiotic powder for assisting in improving vaginitis symptoms, its preparation, and use; CN118319959A: A probiotic powder for regulating allergic constitutions, and its preparation method; and CN1150314C: An immune-enhancing lactic acid bacterium. (HN001 was originally isolated from cheddar cheese produced in New Zealand. It has strong tolerance to gastric acid and bile salts, can survive and colonize in the intestines, and metabolizes to produce substances such as lactic acid and short-chain fatty acids, maintaining intestinal microecological balance. This strain is deposited with the Australian Government Analytical Laboratory (AGAL) under accession number NM97 / 09514 and with the American College of Tissue Cultures Collection under registration number ATCC SD5675.)

[0051] Bifidobacterium animalis subsp. lactis BAL531 is a microorganism in the prior art and can be obtained commercially. It has been widely used in commercial activities. For example, reference can be made to the following literature and patents: Li Sitong, Mechanism of Lactobacillus and Bifidobacteria in Relieving Constipation, Chinese Journal of Microecology, Vol. 35, No. 7, July 2023; CN117653672A, Application of Bifidobacterium animalis subsp. lactis BAL531 in the Preparation of Digestive Products (Deposit Number: CGMCC No. 17329).

[0052] Lactobacillus rhamnosus LR519 is a commercially available microorganism. Reference patents include: CN117256844A, a fermented product of a medicinal and edible composition for improving menopausal symptoms and its preparation method; and CN112481156B, a strain of Lactobacillus rhamnosus LR519 and its composition for alleviating diarrhea (deposit number CGMCC No. 15969).

[0053] Bifidobacterium bifidum B11 is a microorganism in the prior art and can be obtained commercially. For reference, see patent CN117653671A, "Use of Bifidobacterium bifidum B11 in the Preparation of Products for Improving Functional Dyspepsia" (Deposit Number: CGMCC No. 24381).

[0054] Example 1

[0055] This embodiment provides a probiotic composition, designated as experimental group 1, which is composed of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 at a ratio of 4:1 in terms of viable bacteria count.

[0056] At the same time, this embodiment also provides a preparation comprising the aforementioned probiotic composition and a preparation method thereof, specifically:

[0057] The formulation is: 25 parts inulin, 20 parts isomalt, 10 parts isomaltooligosaccharide, 7 parts fructooligosaccharide, 3 parts stachyose, and 35 parts probiotic composition. In practical applications, the formulation can be made from the following raw materials in parts by weight: 20-30 parts inulin, 15-25 parts isomalt, 5-15 parts isomaltooligosaccharide, 5-10 parts fructooligosaccharide, 1-5 parts stachyose, and 10-40 parts probiotic composition.

[0058] In the preparation, prebiotics are added to the probiotic composition. The prebiotics can be selected from a mixture of any one or more of galacto-oligosaccharides (GOS), resistant dextrin / resistant starch, lactulose, xylo-oligosaccharides (XOS), polydextrose, arabinogalactan, β-glucan, manno-oligosaccharides (MOS), inulin, isomalt, isomaltooligosaccharide, fructo-oligosaccharide, and stachyose.

[0059] The preparation method of the preparation is:

[0060] S1: The bacterial strains required in the formulation are activated and cultured in liquid culture medium by conventional means in the prior art, and the bacterial bodies are collected by centrifugation and freeze-dried to obtain bacterial powder;

[0061] S2: Mix the bacterial powder according to the ratio of live bacteria count in the preparation formula, then weigh the bacterial powder corresponding to the formula according to the mass ratio, and add other auxiliary agents at the same time. The auxiliary agents include 25 parts of inulin, 20 parts of isomalt, 10 parts of isomaltooligosaccharide, 7 parts of fructooligosaccharide, and 3 parts of stachyose in terms of mass ratio.

[0062] In addition, at least one of sweeteners, pigments, dietary fiber supplements, micronutrients, and flavors may be added to the preparation to improve the taste or appearance of the preparation or to enrich its nutritional value. In practical applications, the preparation may be in the form of a lyophilized powder, enteric-coated capsules, chewable tablets, or liquid suspension. In practical applications, the total viable count of probiotics per unit dose is 1×10 9 CFU to 5×10 11 CFU.

[0063] Example 2

[0064] This example provides a probiotic composition, designated as experimental group 2, which is composed of live bacteria of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. lactis HN019, and Lactobacillus rhamnosus HN001 in a ratio of 4:1:1:1.

[0065] At the same time, this embodiment also provides a preparation containing the aforementioned probiotic composition and a preparation method thereof. The formulation and preparation method thereof refer to the aforementioned Example 1, and only the probiotic combination method is different from Example 1.

[0066] Example 3

[0067] This embodiment provides a probiotic composition, recorded as experimental group 3, which is composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. HN019, Lactobacillus rhamnosus HN001, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531 in a ratio of 4:1:1:1:1:1:1:1:1 of live bacteria.

[0068] At the same time, this embodiment also provides a preparation containing the aforementioned probiotic composition and a preparation method thereof. The formulation and preparation method thereof refer to the aforementioned Example 1, and only the probiotic combination method is different from Example 1.

[0069] Comparative Example 1

[0070] Please refer to the aforementioned Example 1. This example provides a comparative example of Example 1. This example provides:

[0071] The control group 1A is a probiotic composition composed of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 at a ratio of 1:1 in terms of viable bacteria count.

[0072] The control group 1B is a probiotic composition composed of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 at a ratio of 2:1 in terms of viable bacteria count.

[0073] The control group 1C is a probiotic composition composed of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 at a ratio of 6:1 in terms of viable bacteria count.

[0074] The control group 1D is a probiotic composition composed of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 at a ratio of 1:2 in terms of viable bacteria count.

[0075] The control group 1E is a probiotic composition composed of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 at a ratio of 1:4 in terms of viable bacteria count.

[0076] Comparative Example 2

[0077] Please refer to the aforementioned Example 2. This example provides a comparative example of Example 2. This example provides:

[0078] Control group 2A is a probiotic composition composed of Bifidobacterium lactis subsp. HN019 and Lactobacillus rhamnosus HN001 in a ratio of 1:1 in terms of viable bacteria count; for this bacterial species combination (Bifidobacterium lactis subsp. HN019 and Lactobacillus rhamnosus HN001), the prior art has been extensively studied, including a probiotic composition and its application disclosed in the Chinese invention patent application with document number: CN118389362A. This patented technology solves the problems of postpartum hemorrhage, weight gain during pregnancy, and increased vitamin D levels by using a probiotic composition of Lactobacillus acidophilus La28, Bifidobacterium lactis BAL531, and Lactobacillus plantarum LP45, thereby achieving the effect of reducing postpartum hemorrhage and body fat percentage. At the same time, the Chinese invention patent application with document number CN103637219A discloses a probiotic composition and its application, and infant food. By adding a probiotic composition of Bifidobacterium HN019 and Lactobacillus rhamnosus HN001 to infant food, it solves the high incidence of infant eczema, improves intestinal microecology, reduces the incidence of eczema and enhances immunity, which is close to the effect of breastfeeding. However, the document does not mention whether the combination of Bifidobacterium lactis subspecies HN019 and Lactobacillus rhamnosus HN001 has a certain effect on improving brain function.

[0079] Control group 2B, which is a probiotic composition composed of the viable bacteria count of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. lactis HN019, and Lactobacillus rhamnosus HN001 in a ratio of 1:1:1:1;

[0080] Control group 2C, which is a probiotic composition composed of the viable bacteria count of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. HN019, and Lactobacillus rhamnosus HN001 in a ratio of 2:1:1:1;

[0081] Control group 2D, which is a probiotic composition composed of the viable bacteria count of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. HN019, and Lactobacillus rhamnosus HN001 in a ratio of 6:1:1:1;

[0082] Comparative Example 3

[0083] Please refer to the aforementioned Example 3. This example provides a comparative example with respect to Example 2. This example provides:

[0084] Control group 3A, which is a probiotic composition composed of Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531 in a ratio of 1:1:1:1 in terms of viable cell count;

[0085] Control group 3B, which is a probiotic composition composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531 in a ratio of 4:1:1:1:1:1:1 in terms of viable cell count;

[0086] Control group 3C, which is a probiotic composition, composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, Bifidobacterium animalis subsp. lactis BAL531, and Bifidobacterium bifidum B11 (Note: the preservation number of Bifidobacterium bifidum B11 is CGMCC No. 24381, and its classification name is Bifidobacterium bifidum) in a ratio of 4:1:1:1:1:1:1:1 in terms of viable cell count;

[0087] Control group 3D, which is a probiotic composition, composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, Bifidobacterium animalis subsp. lactis BAL531, and Bifidobacterium bifidum B11 (Note: the preservation number of Bifidobacterium bifidum B11 is CGMCC No. 24381, and its classification name is Bifidobacterium bifidum) according to the ratio of viable cell count of 4:1:1:1:1:1:1:4;

[0088] Comparative Example 4

[0089] Please refer to the aforementioned Example 1. This example provides a comparative example of Example 1. This example provides:

[0090] Control group 4A is a probiotic composition consisting solely of Lactobacillus plantarum LP45. Regarding this strain LP45, Chinese invention patent CN111110705A discloses the use of Lactobacillus plantarum LP45 in preparing products that improve memory, relieve anxiety, and alleviate depression. This Lactobacillus plantarum LP45 addresses the problem of memory impairment caused by modern life by improving the abnormal expression of BDNF and GABA, providing food and pharmaceutical preparations that improve memory, relieve anxiety and depression, and achieve the effects of memory enhancement and mood stabilization.

[0091] Control group 4B is a probiotic composition consisting only of Lactobacillus acidophilus La28. For this strain, the Chinese invention patent application with document number CN114392281A discloses a new application of Lactobacillus acidophilus La28 for inhibiting Helicobacter pylori as a gastric mucosal protection product. The Lactobacillus acidophilus La28 solves the problem that the existing probiotic composition cannot protect the gastric mucosa alone, achieves inhibition of Helicobacter pylori and multiple protection of the gastric mucosa, enhances gastric mucosal cell activity and ulcer healing ability, and is suitable for a variety of people. The Chinese invention patent with document number CN113893267B discloses a new application of Lactobacillus acidophilus La28 in anti-influenza virus. It uses live bacteria and inactivated strains of Lactobacillus acidophilus La28 to solve the problems of side effects and poor effects of existing anti-influenza drugs, achieves non-toxic and effective influenza prevention and treatment, and is particularly suitable for high-risk populations through multiple anti-influenza mechanisms and diversified dosage forms. However, the use of La28 alone in improving brain function has not been reported.

[0092] Test Example 1

[0093] The purpose of this experimental example is to test the efficacy of the aforementioned probiotic compositions. The experimental methods and experimental results are as follows:

[0094] 1.1 Experimental Animal Grouping and Dosage Scheme

[0095] Animal Grouping:

[0096] SPF male C57BL / 6 mice (8 weeks old, weighing 20-25 g) were selected and, after 7 days of adaptive feeding, randomly divided into groups (n=10), including all combinations of experimental groups 1-3, comparative examples 1-4, and a blank control group (normal saline).

[0097] Dosage regimen:

[0098] Probiotic intervention phase (D1-D28):

[0099] Each group of mice was gavaged with the corresponding probiotic composition (bacterial powder dissolved in physiological saline, with a total viable count of 1×10 9 CFU / mL), and the mice were dosed at a dose of 10 mL / kg according to their body weight for 28 consecutive days; the blank control group was given an equal volume of normal saline.

[0100] Model construction (D29):

[0101] On the day after the end of drug administration (D29), all mice were intraperitoneally injected with scopolamine (5 mg / kg / d) to induce an acute cognitive dysfunction model, and then behavioral tests were performed starting on D30, and the experimental results were recorded and analyzed.

[0102] 1.2 Experimental results and analysis

[0103] 1.2.1 Novel Object Recognition Experiment (D30-D32)

[0104] D30 (adaptation period): Mice were placed in an open behavioral box (40×40×40 cm) and allowed to explore freely for 10 minutes.

[0105] D31 (training period): Two identical objects (A1, A2) were placed and the exploration time of the mice was recorded within 10 minutes.

[0106] D32 (test period): Replace one of the objects with a new object (B), and record the exploration time of the new and old objects within 5 minutes (use 75% ethanol to wipe the objects and behavior box after each experiment to eliminate odor interference).

[0107] The experimental results are expressed as recognition index, which is calculated as follows:

[0108] Recognition index = new object exploration time / (new object exploration time + old object exploration time) × 100%;

[0109] A larger recognition index (over 50%) indicates that the mouse spends more time exploring the new object, indicating that it can distinguish between new and old objects, suggesting that long-term memory is well retained (dependent on the function of the hippocampus and perirhinal cortex).

[0110] 1.2.2 Platform test (D33-D34)

[0111] D33 (training phase): Mice were placed in the reaction box to adapt for 5 minutes. After the power was turned on, if the mouse jumped off the platform, it was considered to have escaped successfully. The latency of the first jump off the platform (learning ability) was recorded. The training lasted for 5 minutes. If the mouse did not jump off the platform, the latency was recorded as 300 seconds.

[0112] D34 (test phase): After 24 hours, the platform jumping latency (time to first jump) and the number of errors within 3 minutes were tested.

[0113] The experimental results are expressed as test latency and number of errors. The test latency is the time it takes for the mouse to jump off the safety platform for the first time. The longer the latency, the deeper the mouse's memory of the electric shock and the stronger its long-term memory retention ability (dependent on the amygdala and prefrontal cortex). The fewer the number of errors, the stronger the mouse's associative memory of the electric shock and the stronger its ability to inhibit erroneous responses.

[0114] 1.2.3 Morris water maze test (D35-D40)

[0115] Days 35-39 (positioning navigation test): The platform was fixed in quadrant II of the pool, and mice were trained daily in quadrant order. The escape latency from entering the water until finding the platform was recorded (upper limit 60 seconds). Those that failed to find the platform were guided to the platform and remained there for 15 seconds.

[0116] D40 (Space Exploration Experiment):

[0117] The platform was removed and the mouse was placed in the water from quadrant IV. The number of times the mouse crossed the original platform position and the time it stayed in the target quadrant within 60 seconds were recorded. The more times the mouse crossed the original platform position, the more accurate the mouse's spatial memory of the platform position was. The longer the total time the mouse stayed in the quadrant where the original platform was located, the higher the mouse's dependence on spatial cues and the more stable the long-term spatial memory retention.

[0118] 1.3 Data collection and statistical analysis

[0119] Data Records:

[0120] Behavioral data were automatically collected using EthoVision software. Experimental results were analyzed using SPSS 26.0 software, and quantitative data are presented as mean ± standard deviation (mean ± SD). Comparisons among multiple groups were performed using one-way analysis of variance (ANOVA). If homogeneity of variance was established (Levene's test, P > 0.05), a post hoc test using the Tukey method was performed. In cases of heterogeneity of variance, the Welch correction and the Games-Howell test were used.

[0121] Significance threshold: *P<0.05, **P<0.01. Experimental results are shown in Tables 1-3.

[0122] Table 1: Experimental results of experimental group 1, control groups 1A-1E, and control groups 4A-B

[0123]

[0124] *: Compared with the blank control group, *P<0.05, **P<0.01; #: Compared with the control group 4, #P<0.05, ##P<0.01.

[0125] Table 2: Experimental results of experimental group 2 and control groups 2A-2D

[0126]

[0127]

[0128] *: Compared with the blank control group, *P<0.05, **P<0.01; #: Compared with the control group 2A, #P<0.05, ##P<0.01.

[0129] Table 3: Experimental results of experimental groups 3A-3D

[0130]

[0131] *: Compared with the blank control group, *P<0.05, **P<0.01; #: Compared with the control group 3B, #P<0.05, ##P<0.01.

[0132] According to the experimental data of table 1, embodiment 1 (experimental group 1) and comparative example 1 (control group 1A-1E), comparative example 4 (control group 4A, B) are analyzed.For experimental group 1 (combination using two kinds of probiotics to form), control group 4A (LP45 is used alone), control group 4B (La28 is used alone, and experimental data shows that it does not have the effect of significantly improving brain function), the total amount of probiotics used in three test groups is consistent, and difference is only the composition of probiotics. If there is no synergistic effect between two kinds of probiotics, according to conventional reasoning, the effect of experimental group 1 should be between the effect of control group 4A and the effect of control group 4B. But, in fact, the technical effect of plant lactobacillus LP45, lactobacillus acidophilus La28 two kinds of probiotics are used in conjunction (experimental group 1), is better than the effect of plant lactobacillus LP45 being used alone and lactobacillus acidophilus La28 being used alone. This explanation plant lactobacillus LP45 and lactobacillus acidophilus La28 are used in conjunction, and on the learning and memory ability that promotes experimental animals, there is the effect of synergistic effect. More specifically, in the Morris water maze test, experimental group 1 used a combination of two probiotics. The escape latency of the experimental mice was significantly lower than that of the control groups 4A and B, and the number of crossings was significantly higher than that of the control groups 4A and B, indicating that the mice in experimental group 1 had stronger learning ability (improved cognitive dysfunction) and memory ability after drug treatment. In the platform jumping test, experimental group 1 used a combination of two probiotics. The test latency of the experimental mice was significantly higher than that of the control groups 4A and B, and the number of errors was significantly lower than that of the control groups 4A and B, indicating that the mice in experimental group 1 had stronger learning ability (improved cognitive dysfunction) and memory ability after drug treatment. In the novel object recognition test, experimental group 1 used a combination of two probiotics. The recognition index of the experimental mice was significantly higher than that of the control groups 4A and B, indicating that the mice in experimental group 1 had stronger memory ability after drug treatment.

[0133] In addition to experimental group 1, the synergistic effect of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 can also be observed in control groups 1A (LP45 and La28 ratio 1:1) and 1B (LP45 and La28 ratio 2:1). The technical effects of control groups 1A and 1B are significantly better than those of control groups 4A and B. Therefore, the combined ratio of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 should be 1:1-4:1 (among which 2:1 has the best effect). In control groups 1C-1E, the ratios of LP45 and La28 were 6:1, 1:2, and 1:4, respectively. The effects of these three control groups did not show any significant improvement over those of control groups 4A and B. Control group 1E had the worst effect, suggesting that an excessively high proportion of Lactobacillus acidophilus La28 may weaken the effect of the combination. This also shows that the synergistic effect of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 needs to be achieved under a specific ratio.

[0134] According to the experimental data of Table 1 and Table 2, embodiment 1 (experimental group 1), embodiment 2 (experimental group 2) and comparative example 2 (control group 2A-2D) are analyzed.Experimental group 2 added bifidobacterium lactis subspecies HN019 and rhamnosus lactobacillus HN001 on the basis of experimental group 1, further promoted the effect that medicine improves experimental animal learning ability (improving cognitive dysfunction) and memory ability. For the combination of these two probiotics of bifidobacterium lactis subspecies HN019 and rhamnosus lactobacillus HN001, the effect of itself is not too outstanding (control group 2A), has only the effect of improving brain function by a small margin relative to blank control. But, use does not possess the probiotic combination (HN019 and HN001) of the improvement effect of significant amplitude, after being used in conjunction with plant lactobacillus LP45 and lactobacillus acidophilus La28 in experimental group 1 (experimental group 2), aspect improving experimental animal learning ability (improving cognitive dysfunction) and memory ability, but demonstrated very significant improvement, exceeded the expectation of those skilled in the art.

[0135] The control group 2D, which contained a slightly higher content of Lactobacillus plantarum LP45, showed less than ideal improvements in learning and memory. Control groups 2B, 2C, and experimental group 2 all demonstrated significant improvements in memory and learning abilities. This suggests that a 1-4:1:1:1 ratio of the four probiotics, including Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. lactis HN019, and Lactobacillus rhamnosus HN001, significantly improved drug efficacy. The optimal ratio was 4:1:1:1 (experimental group 2).

[0136] According to the experimental data of table 2 and table 3, embodiment 2 (experimental group 2), embodiment 3 (experimental group 3) and comparative example 3 (control group 3A-3D) are analyzed.Experimental group 3 adds plant lactobacillus CN2018, rhamnosus lactobacillus LR863, rhamnosus lactobacillus LR519, animal bifidobacterium lactis subspecies BAL531 these four kinds of probiotics on the basis of experimental group 2, further improves the effect that medicine improves experimental animal learning ability (improving cognitive dysfunction) and memory ability.For the combination of plant lactobacillus CN2018, rhamnosus lactobacillus LR863, rhamnosus lactobacillus LR519, animal bifidobacterium lactis subspecies BAL531 these four kinds of probiotics, its action effect is not too outstanding (control group 3A), and its action effect is compared with blank control group, and there is not too big difference. The inventor has also attempted to combine the use (control group 3B) of plant lactobacillus LP45 and lactobacillus acidophilus La28 of four kinds of probiotics with experimental group 1, and its effect does not also appear to promote significantly relative to experimental group 1, but, relative to blank, its effect has significantly promoted.This shows that, in order to promote the effect of probiotic composition, it is necessary to, on the basis of plant lactobacillus LP45 and lactobacillus acidophilus La28, add bifidobacterium lactis subspecies HN019 and rhamnosus lactobacillus HN001; In order to further promote effect, it is possible to, on the basis of above-mentioned four kinds of probiotics, further add plant lactobacillus CN2018, rhamnosus lactobacillus LR863, rhamnosus lactobacillus LR519, animal bifidobacterium lactis subspecies BAL531.

[0137] Control group 3C, based on control group 3B, further added Bifidobacterium bifidum B11, which further enhanced the probiotic composition's effectiveness in improving learning ability (improving cognitive dysfunction) and memory in mice. Control group 3D, based on control group 3C, further increased the proportion of Bifidobacterium bifidum B11, but its effectiveness was somewhat weakened.

[0138] In addition, regarding the dosage ratio of the eight probiotics, experimental group 3 (8-bacteria combination, ratio 4:1:1:1:1:1:1:1:1) had the most outstanding effect (escape latency 21.6 seconds, recognition index 82.4%), indicating that the synergy of multiple bacterial species can significantly enhance cognitive protection.

[0139] The above is only an embodiment of the present invention, and the common knowledge such as the specific technical solutions and / or characteristics in the solution are not described in detail here. It should be pointed out that for those skilled in the art, without departing from the technical solution of the present invention, several variations and improvements can be made, which should also be regarded as the scope of protection of the present invention, and these will not affect the effect of the implementation of the present invention and the practicality of the patent. The scope of protection required by this application shall be based on the content of its claims, and the specific implementation methods and other records in the description can be used to interpret the content of the claims.

Claims

1. A probiotic composition, characterized in that It includes probiotics: Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28; Preferably, the ratio of the viable counts of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 is 1:1-4:1; Preferably, the ratio of the viable cell counts of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 is 2:

1.

2. A probiotic composition according to claim 1, characterized in that Also included are probiotics: Bifidobacterium animalis subsp. lactis HN019 and Lactobacillus rhamnosus HN001; Preferably, the ratio of the viable cell counts of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. HN019, and Lactobacillus rhamnosus HN001 is 1-4:1:1:1; Preferably, the ratio of the viable cell counts of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. lactis HN019, and Lactobacillus rhamnosus HN001 is 4:1:1:

1.

3. A probiotic composition according to claim 2, characterized in that Also included are probiotics: Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531; Preferably, the ratio of the viable cell count of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. lactis HN019, Lactobacillus rhamnosus HN001, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531 is 4:1:1:1:1:1:1:1:

1.

4. A probiotic composition according to claim 1, characterized in that Also included are probiotics: Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531; Preferably, the ratio of the viable cell counts of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519 and Bifidobacterium animalis subsp. lactis BAL531 is 4:1:1:1:1:1:

1.

5. A probiotic composition according to claim 4, characterized in that, Also includes probiotics: Bifidobacterium bifidum B11; Preferably, the ratio of the viable cell counts of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, Bifidobacterium animalis subsp. lactis BAL531, and Bifidobacterium bifidum B11 is 4:1:1:1:1:1:1:

1.

6. Use of the probiotic composition according to any one of claims 1 to 5 in preparing a preparation for improving brain function, characterized in that: Preferably, the improvement of brain function is at least manifested in improvement of memory ability and / or improvement of cognitive dysfunction.

7. The use according to claim 6, wherein the preparation further comprises a prebiotic; Preferably, the prebiotics are selected from a mixture of any one or more of galacto-oligosaccharides, resistant dextrin / resistant starch, lactulose, xylo-oligosaccharides, polydextrose, arabinogalactan, β-glucan, oligomannose, inulin, isomalt, isomaltooligosaccharide, oligofructose, and stachyose.

8. The use according to claim 7, wherein the preparation further comprises at least one of a sweetener, a pigment, a dietary fiber supplement, a micronutrient, and a flavor; The dosage form of the preparation is any one of lyophilized powder, enteric-coated capsules, chewable tablets or liquid suspension; The total number of viable probiotic bacteria per unit dose is 1×10 9 CFU to 5×10 11 CFU.

9. The use according to claim 6, characterized in that The preparation is prepared from the following raw materials in parts by mass: 20-30 parts of inulin, 15-25 parts of isomalt, 5-15 parts of isomaltooligosaccharide, 5-10 parts of fructooligosaccharide, 1-5 parts of stachyose and 10-40 parts of a probiotic composition.

10. The use according to claim 6, characterized in that The composition of the probiotic composition is as follows: It is composed of Lactobacillus plantarum LP45 and Lactobacillus acidophilus La28 in a ratio of 1:1-4:1 in terms of viable bacteria count, preferably 2:1 in terms of viable bacteria count; Or, it is composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. HN019, and Lactobacillus rhamnosus HN001 in a ratio of 1-4:1:1:1 in terms of viable cell count, preferably, the ratio of 4:1:1:1; or, composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Bifidobacterium lactis subsp. lactis HN019, Lactobacillus rhamnosus HN001, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531 in a ratio of 4:1:1:1:1:1:1:1:1; or, composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, and Bifidobacterium animalis subsp. lactis BAL531 at a ratio of 4:1:1:1:1:1:1 in terms of viable cell count; Or, it is composed of Lactobacillus plantarum LP45, Lactobacillus acidophilus La28, Lactobacillus plantarum CN2018, Lactobacillus rhamnosus LR863, Lactobacillus rhamnosus LR519, Bifidobacterium animalis subsp. lactis BAL531 and Bifidobacterium bifidum B11 with a ratio of 4:1:1:1:1:1:1:1 in terms of live cell count.

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