A composition, its preparation method and its use in promoting intellectual development

Abstract

The present application relates to the technical field of biotechnology, and particularly discloses a composition, a preparation method thereof and application thereof in promoting intelligence development. The present application finds that, when Lactobacillus plantarum HEAL9, oat green extract and sialic acid are compounded according to a specific ratio, a significant synergistic effect can be achieved in promoting intelligence development, and thus a composition is provided, which comprises: Lactobacillus plantarum HEAL9, oat green extract and sialic acid; and the mass ratio of the Lactobacillus plantarum HEAL9, the oat green extract and the sialic acid is (1-10):(0.7-30):(1-10). The present application has a strong effect in promoting intelligence development, and provides a new method for promoting intelligence development of infants / children.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, and more specifically, to a composition, a method for preparing the same, and its application in promoting intellectual development. Background Technology

[0002] Intellectual development encompasses key cognitive abilities such as memory, attention, logical reasoning, problem-solving, and critical thinking, which directly determine the speed and depth at which an individual learns new knowledge and masters new skills. Infancy and childhood are the critical window period for intellectual development.

[0003] Oat green extract is an active ingredient extracted from the whole plant (aerial parts - stems and leaves) of immature, still green oats (Latin name Avena sativa L., genus Avena in the family Poaceae) through processes such as extraction with 30% ethanol, concentration, drying, and grinding. It has the effects of improving concentration and cognitive performance, relieving stress and anxiety, and potential neuroprotective effects.

[0004] Sialic acid is a fundamental component of brain gangliosides and sialylated glycoproteins, and it participates in regulating important processes related to memory and cognition, including regulating neuronal myelination, synaptic transmission, intercellular interactions, and microglial cell immune responses.

[0005] HEAL9 is a plant-based lactobacillus. Current research has found that it can alleviate cognitive impairment and intestinal dysfunction in aging individuals by regulating the gut-microbiota axis, but there are no reports on its effects on intellectual development during adolescence.

[0006] Currently, most infant / children's intellectual development products are designed and developed by adding ingredients such as DHA, theanine, and sialic acid. However, it is still necessary to further improve the efficacy of these products and enrich their variety. Summary of the Invention

[0007] One of the objectives of this invention is to provide a new product that can effectively promote intellectual development.

[0008] The present invention provides a composition comprising: *Lactobacillus plantarum* HEAL9, oat extract and sialic acid; wherein the mass ratio of *Lactobacillus plantarum* HEAL9, oat extract and sialic acid is (1-10):(0.7-30):(1-10).

[0009] Currently, there are no reports on the application of oat green extract combined with probiotics in promoting intellectual development. This invention is the first to discover that when *Lactobacillus plantarum* HEAL9, oat green extract, and sialic acid are combined in a specific ratio, they exhibit a significant synergistic effect in promoting intellectual development. At the same dosage, the composition of this invention is significantly more effective than any single monomer or any combination of two. Furthermore, the composition of this invention is also more effective than other known combinations of components that promote intellectual development, achieving better results at the same dosage.

[0010] Furthermore, this invention has found that directly mixing sialic acid with probiotics can easily kill the probiotics, thus affecting the overall effect of the sialic acid-probiotic combination. However, by using the specific combination of substances in this invention, the compatibility of sialic acid and HEAL9 can be improved, thereby producing a synergistic effect in promoting intellectual development.

[0011] Preferably, the mass ratio of *Lactobacillus plantarum* HEAL9, oat extract and sialic acid is (1-10):(0.7-29.4):(1-10).

[0012] Preferably, in the composition of the present invention, the mass ratio of *Lactobacillus plantarum* HEAL9, oat extract and sialic acid is (1-10):(0.7-30):1, and more preferably (1-10):(0.7-29.4):1.

[0013] More preferably, in the composition of the present invention, the mass ratio of *Lactobacillus plantarum* HEAL9, oat extract and sialic acid is (9-10):(0.7-0.9):(1-1.2), and even more preferably the mass ratio is 10:0.7:1.

[0014] When the preferred material combination ratio of the present invention is used, even better results can be obtained.

[0015] In one specific embodiment, the mass ratio of *Lactobacillus plantarum* HEAL9, oat extract, and sialic acid in the composition of the present invention is 1:0.7:10, 1:0.7:1, or 2:29.4:1.

[0016] The present invention also provides a method for preparing the above composition, which includes the step of mixing Lactobacillus plantarum HEAL9, oat extract and sialic acid.

[0017] Those skilled in the art can use methods known in the art to mix HEAL9, oat green extract and sialic acid, and the present invention does not impose any special limitations on this.

[0018] The present invention also provides the use of the above composition in the preparation of products that promote intellectual development.

[0019] The present invention also provides a product for promoting intellectual development, comprising the above-described composition.

[0020] The active ingredients in the products of the present invention may be only the above-described compositions of the present invention, or may also include other active ingredients.

[0021] The product of the present invention may also be a pharmaceutical product, which may include pharmaceutically acceptable excipients known in the art; the dosage form may be a tablet, capsule, granule, powder, pill, oral solution, syrup, emulsion, suspension, drop pill, enteric-coated preparation, or other pharmaceutical dosage forms known in the art.

[0022] The beneficial effects of this invention are at least as follows:

[0023] This invention provides a composition that promotes intellectual development by combining HEAL9, oat green extract and sialic acid in a specific ratio. This composition is highly effective and provides a new method for promoting the intellectual development of infants / children. Detailed Implementation

[0024] The preferred embodiments of the present invention will now be described in detail with reference to specific examples. It should be understood that the following examples are given for illustrative purposes only and are not intended to limit the scope of the invention. Those skilled in the art can make various modifications and substitutions to the present invention without departing from its spirit and essence.

[0025] Unless otherwise specified, the experimental methods used in the following examples are conventional methods. Unless otherwise specified, the materials and reagents used in the following examples are commercially available or prepared according to conventional methods in the art.

[0026] In this invention, *Lactobacillus plantarum* HEAL9 (hereinafter referred to as HEAL9) was purchased from Probi®, traded under the name Sensia, with a viable count of 6 × 10⁻⁶. 11cfu / g. The oat green extract described in the specific embodiments of this invention was purchased from IFF HealthSciences, traded under the name Neuravena® Oat Green Extract (trade name in China: Oat Green Extract Solid Beverage). The active ingredient, oat green extract, contains 70% (in the following examples and comparative examples, when using this raw material, its mass ratio needs to be multiplied by 70% to obtain the mass ratio of the active ingredient. For example, in Example 2, the mass ratio of HEAL9, oat green extract, and sialic acid is 1:1:1; after conversion, the mass ratio of the three active ingredients is 1:0.7:1). Sialic acid was purchased from Jiabiyou Biotechnology (Wuhan) Co., Ltd., traded under the name N-acetylneuraminic acid. Phosphatidylserine (PS) was purchased from Beijing Yifan International Trade Co., Ltd., produced by IFF, traded under the name Sharp PS®60P-FP CNPCN. It is a powdered phosphatidylserine product, with soybean lecithin as its main raw material, and a PS content of 60%. Theanine was purchased from Huisong Pharmaceutical, under the trade name "Tea Theanine," and is a yellow-brown amorphous powder extracted from tea leaves. Nervonic acid was purchased from Heze Zhonghe Jianyuan Biotechnology Co., Ltd., and is cis-15-tetracosenoic acid produced from rapeseed oil through processes such as saponification, acidification, extraction, separation, crystallization, and drying.

[0027] Example 1

[0028] This embodiment provides a composition comprising HEAL9, oat extract, and sialic acid in a mass ratio of 2:42:1. During preparation, the components are uniformly mixed according to the specified proportions.

[0029] Example 2

[0030] This embodiment provides a composition comprising HEAL9, oat extract, and sialic acid in a mass ratio of 1:1:1. During preparation, the components are uniformly mixed in the specified proportions.

[0031] Example 3

[0032] This embodiment provides a composition comprising HEAL9, oat extract, and sialic acid in a mass ratio of 1:1:10. During preparation, the components are uniformly mixed according to the specified proportions.

[0033] Example 4

[0034] This embodiment provides a composition comprising HEAL9, oat extract, and sialic acid in a mass ratio of 10:1:1. During preparation, the components are uniformly mixed according to the specified proportions.

[0035] Comparative Example 1

[0036] This comparative example provides a single-component substance, HEAL9.

[0037] Comparative Example 2

[0038] This comparative example provides a single-component substance, which is oat green extract.

[0039] Comparative Example 3

[0040] This comparative example provides a single-component substance, which is sialic acid.

[0041] Comparative Example 4

[0042] This comparative example provides a composition comprising HEAL9 and oat extract in a mass ratio of 14:1. During preparation, the components are uniformly mixed in the specified proportions.

[0043] Comparative Example 5

[0044] This comparative example provides a composition comprising oat green extract and sialic acid in a mass ratio of 5:1. During preparation, the components are uniformly mixed in the specified proportions.

[0045] Comparative Example 6

[0046] This comparative example provides a composition comprising oat green extract and sialic acid in a mass ratio of 1:14. During preparation, the components are uniformly mixed in the specified proportions.

[0047] Comparative Example 7

[0048] This comparative example provides a composition comprising HEAL9 and sialic acid in a mass ratio of 1:1. During preparation, the components are uniformly mixed in the specified proportions.

[0049] Comparative Example 8

[0050] This comparative example provides a composition comprising HEAL9, oat extract, and sialic acid in a mass ratio of 1:118:1. During preparation, the components are uniformly mixed according to the specified proportions.

[0051] Comparative Example 9

[0052] This comparative example provides a composition comprising HEAL9, oat extract, and sialic acid in a mass ratio of 118:1:1. During preparation, the components are uniformly mixed in the specified proportions.

[0053] Comparative Example 10

[0054] This comparative example provides a composition comprising HEAL9, oat extract, and sialic acid in a mass ratio of 1:1:118. During preparation, the components are uniformly mixed according to the specified proportions.

[0055] Comparative Example 11

[0056] This comparative example provides a composition comprising HEAL9, oat extract, and sialic acid in a mass ratio of 2:50:0.5. During preparation, the components are uniformly mixed according to the specified proportions.

[0057] Comparative Example 12

[0058] This comparative example provides a composition comprising HEAL9, oat extract, and sialic acid in a mass ratio of 2:76:1. During preparation, the components are uniformly mixed in the specified proportions.

[0059] Experimental Example 1

[0060] This experimental example demonstrates the effectiveness of combining the above-described embodiments and comparative examples in promoting intellectual development. Color preference tests are frequently used in zebrafish cognitive assessments. Zebrafish prefer shorter wavelengths of color, and compared to other colors (such as red, yellow, and green), they exhibit a strong preference for blue. This preference becomes more pronounced as brain development matures. In other words, the more movement a zebrafish makes in the blue area, the more cognitively mature its cognitive abilities.

[0061] The specific experimental method is as follows:

[0062] Animal intervention: Wild-type AB strain zebrafish, 5 days post-fertilization (5 dpf), were randomly selected and divided into groups of 30 fish each, placed in 20 mL beakers. Except for the normal control group, each group received a water-soluble sample. A normal control group and a positive control group were also established. The positive control group received edaravone at a concentration of 5 μg / mL; the remaining experimental groups received samples at a total concentration of 30 μg / mL.

[0063] Detection Method: After treatment at 28℃ for 1 day, zebrafish were randomly selected from each experimental group and placed into the "+" module of a zebrafish behavior analyzer. The module was divided into four areas: yellow, blue, red, and green. Six modules were placed in each group, with five zebrafish in each module. Data was collected using a behavior analyzer, and the percentage (%) of the total movement distance of the zebrafish in the blue area within 10 minutes was analyzed. This yielded six parallel experimental data points for each group (each data point was the sum of the test results of five fish in a single module). The statistical analysis results of this indicator were used to evaluate the sample's efficacy in promoting brain development. An increased percentage of movement in the blue area of ​​the cross maze indicated that the sample had the effect of promoting intelligence / brain development.

[0064] The experimental results are shown in Table 1: the combination of *Lactobacillus plantarum* HEAL9, oat extract, and sialic acid showed better results than the normal control and the combination of any two ingredients in pairs, demonstrating the effect of promoting intellectual development, specifically by increasing the proportion of motor activity in the blue area of ​​the cross maze. The efficacy of the examples was superior to that of the comparative examples.

[0065] Table 1

[0066]

[0067] Experimental Example 2

[0068] This experimental example further examines the zebrafish from different groups after sample treatment in Experiment 1. gdnfa Genes and bdnf Gene expression levels were measured to further confirm the efficacy of the composition in promoting intellectual development. The results are shown in Tables 2 and 3.

[0069] bdnf Genes can: (1) increase synaptic plasticity, thereby affecting long-term enhancement (nLTP), which is the basis of the learning process and memory formation (secondary memory); (2) promote neurogenesis, especially in the hippocampus; and (3) promote cell survival, mainly by maintaining and promoting the development, differentiation, growth and regeneration of various neurons, especially 5-hydroxytryptamine (5-HT) and dopaminergic (DA) neurons.

[0070] gdnfa The gene can promote the survival of different neuronal subsets at different stages of development in the central and peripheral nervous systems, supporting the generation of type 1 astrocytes, neuronal membrane cells, neurons, pineal gland cells, etc., and its survival-promoting efficiency is particularly significant for spinal motor neurons.

[0071] therefore, bdnf Genes and gdnfa The upregulation of gene expression levels indicates that the sample has the effect of promoting intelligence / brain development.

[0072] In this embodiment, zebrafish from each group were collected, total RNA was extracted, and detected by Real-time RT-PCR. gdnfa Genes and bdnf Relative gene expression level (internal reference) β-actin Gene). gdnfa Genes and bdnf The higher the relative expression level of a gene, the stronger its effect on promoting intellectual development.

[0073] Table 2

[0074]

[0075] Table 3

[0076]

[0077] In Tables 2 and 3 (if applicable), compare with the normal control group. p<0.05, p<0.01, p<0.001;

[0078] Compared with HEAL9+ oat green extract (14:1), # p<0.05, ## p<0.01, ### p<0.001;

[0079] Compared with oat green extract + sialic acid (5:1),

[0080] Compared with oat green extract + sialic acid (1:14), % p<0.05, %% p<0.01, %%% p<0.001;

[0081] Compared to HEAL9+ sialic acid (1:1), & p<0.05, && p<0.01, &&& p<0.001.

[0082] Experimental results: The combination of *Lactobacillus plantarum* HEAL9, oat extract, and sialic acid showed better effects than the normal control and the combination of any two ingredients in pairs, demonstrating a promotion of intellectual development, specifically by enhancing... gdnfa Genes and bdnf The relative expression level of genes.

[0083] Experimental Example 3

[0084] In this experiment, known active ingredients that promote intellectual development (theanine, nervonic acid, and phosphatidylserine) were used to replace each ingredient in the optimal combination of *Lactobacillus plantarum* HEAL9, oat extract, and sialic acid (Example 4; the ratio of the three ingredients was 10:1:1) in equal proportions. The proportion of movement in the blue area of ​​the zebrafish cross maze was measured in each group according to the method in Example 1 (except for the normal control group, the positive control was given edaravone at a concentration of 5 μg / mL; the other experimental groups were given samples at a total concentration of 30 μg / mL of the composition) to compare the effects of each group on promoting intellectual development.

[0085] The experimental results are shown in Table 4: Compared with Example 4 of the present invention, after replacing any one of the components in the three-component combination of Bacillus plantarum HEAL9 + oat extract + sialic acid with known active ingredients (theanine, nervonic acid, phosphatidylserine), the efficacy of the three-component combination in promoting intellectual development was significantly reduced. Specifically, the proportion of movement in the blue area of ​​the cross maze was significantly reduced, and it no longer had the synergistic effect of the two-component combination.

[0086] Table 4

[0087]

[0088] In Table 4, compared with HEAL9 + oat green extract + sialic acid (10:1:1), p<0.001.

[0089] Although the present invention has been described in detail above with general descriptions and specific embodiments, modifications or improvements can be made to it, which will be obvious to those skilled in the art. Therefore, all such modifications or improvements made without departing from the spirit of the present invention fall within the scope of protection claimed by the present invention.

Claims

1. A composition, characterized in that, include: The ingredients are *Lactobacillus plantarum* HEAL9, oat green extract, and sialic acid; the mass ratio of *Lactobacillus plantarum* HEAL9, oat green extract, and sialic acid is (1-10):(0.7-30):(1-10); the oat green extract is made from the stems and leaves of the immature oat plant, using 30% ethanol extraction, concentration, drying, and grinding processes.

2. The composition according to claim 1, characterized in that, The mass ratio of *Lactobacillus plantarum* HEAL9, oat extract, and sialic acid is (1-10):(0.7-30):

1.

3. The composition according to claim 1, characterized in that, The mass ratio of *Lactobacillus plantarum* HEAL9, oat extract, and sialic acid is (9-10):(0.7-0.9):(1-1.2).

4. The composition according to claim 3, characterized in that, The mass ratio of *Lactobacillus plantarum* HEAL9, oat extract, and sialic acid is 10:0.7:

1.

5. The composition according to claim 1, characterized in that, The mass ratio of *Lactobacillus plantarum* HEAL9, oat extract, and sialic acid is 1:0.7:10, 1:0.7:1, or 2:29.4:

1.

6. A method for preparing the composition according to any one of claims 1-5, characterized in that, This includes the step of mixing Lactobacillus plantarum HEAL9, oat extract, and sialic acid.

7. The use of the composition according to any one of claims 1-5 in the preparation of products that promote intellectual development.

8. A product that promotes intellectual development, characterized in that, Includes the composition according to any one of claims 1-5.

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