Composition for emotional relief and promoting neuron growth and use thereof

Abstract

A composition for emotional relief and promoting neuron growth includes lactic acid bacteria strains and a carrier. The lactic acid bacteria strains include isolated Bifidobacterium breveBBr-28, Bifidobacterium animalissubsp. lactisPS23, Bifidobacterium longumsubsp. infantisBR631, Bifidobacterium bifidumBD-188 and Lactiplantibacillus plantarumPQQ-1, all of which are deposited at the Food Industry Research and Development Institute. A use of a lactic acid bacteria strain in preparing a composition for emotional relief and promoting neuron growth is also disclosed.

Description

[Technical Field]

[0001] This invention relates to a composition and its use, and more particularly to a composition and its use for relieving mood and promoting neuronal growth. [Previous Technology]

[0002] The World Health Organization estimates that by 2030, depression will be the leading cause of socioeconomic burden globally. This means that with changing social structures, more people may suffer from depression, and the resulting socioeconomic burden will be even more severe. However, many people with depression may be unwilling to admit they have depression or resist treatment; therefore, most people with depression do not receive adequate medical care.

[0003] Depression is a normal human emotion. For example, when encountering setbacks or unpleasant situations, people inevitably feel down. However, if the low mood is severe and lasts for a long time without recovery, or even gets out of control, it may develop into depression. In addition, being in a state of high stress or depression for a long time may inhibit the secretion of brain-derived neurotrophic factor (BDNF), thereby affecting memory and cognitive function, and even leading to neurocognitive disorders such as Alzheimer's disease (AD).

[0004] In view of this, there is an urgent need to develop a safe and long-term edible nutritional supplement that can soothe emotions and promote neuronal growth. Since lactic acid bacteria are generally considered safe, identifying compositions containing lactic acid bacteria strains with soothing and neuronal growth-promoting activities is a pressing goal. [Summary of the Invention]

[0005] The present invention provides a composition containing a strain of lactic acid bacteria, which has physiological activity effects of soothing emotions and promoting neuronal growth. Therefore, the composition of the present invention can be used for soothing emotions and promoting neuronal growth, and exists in the form of a food composition or a pharmaceutical composition.

[0006] An embodiment of the present invention provides a composition for relieving mood and promoting neuronal growth, comprising a strain of lactic acid bacteria having the effects of relieving mood and promoting neuronal growth, and a carrier. Lactic acid bacteria strains with mood-soothing and neuronal growth-promoting effects include the isolated strains of *Bifidobacterium breve* BBr-28 (accession number BCRC 911254), *Bifidobacterium animalis subsp. lactis* PS23 (accession number BCRC 911251), *Bifidobacterium longum subsp. infantis* BR631 (accession number BCRC 911172), *Bifidobacterium bifidum* BD-188 (accession number BCRC 911252), and *Lactiplantibacillus plantarum* PQQ-1 (accession number BCRC 911262). All of these strains are registered with the Food Industry Development and Research Institute.

[0007] Another embodiment of the present invention is used in the preparation of a composition for soothing emotions and promoting neuronal growth, wherein the composition comprises a lactic acid bacteria strain having the activity of soothing emotions and promoting neuronal growth, and a carrier. Lactic acid bacteria strains with mood-soothing and neuronal growth-promoting effects include the isolated strains of *Bifidobacterium breve* BBr-28 (accession number BCRC 911254), *Bifidobacterium animalis subsp. lactis* PS23 (accession number BCRC 911251), *Bifidobacterium longum subsp. infantis* BR631 (accession number BCRC 911172), *Bifidobacterium bifidum* BD-188 (accession number BCRC 911252), and *Lactiplantibacillus plantarum* PQQ-1 (accession number BCRC 911262). All of these strains are registered with the Food Industry Development and Research Institute.

[0008] The following detailed description, with reference to specific embodiments and accompanying drawings, will make it easier to understand the purpose, technical content, features and effects of the present invention.

Implementation Method

[0009] The various embodiments of the present invention will be described in detail below, with illustrations provided. In addition to these detailed descriptions, the present invention can be widely implemented in other embodiments. Any easy substitutions, modifications, or equivalent changes to the described embodiments are included within the scope of the present invention and are subject to the claims. In the description of the specification, many specific details are provided to give the reader a more complete understanding of the present invention; however, the present invention may still be implemented even if some or all of the specific details are omitted. Furthermore, well-known steps or elements are not described in the details to avoid unnecessarily limiting the present invention. Identical or similar elements in the drawings will be represented by the same or similar symbols. It should be noted that the drawings are for illustrative purposes only and do not represent the actual size or number of elements; some details may not be fully drawn for the sake of simplicity.

[0010] There is extensive research on the effects of probiotics on human health. Generally speaking, strains that have special effects on human health are called functional probiotics (Guidelines for the evaluation of probiotics in food; Report of joint FAO / WHO working group on drafting guidelines for the evaluation of probiotics in food; London Ontario, Canada April 30 and May 1, 2002: 1-7). It should be noted that the special effects of the above-mentioned functional probiotics lie in the specificity of the strain rather than the species. That is, even the same species may have different effects, or even opposite effects, if they are different strains.

[0011] The freeze-dried culture of the lactic acid bacteria strain described in this invention has been deposited with the Food Industry Development & Research Institute, located at No. 331, Food Rd., Hsinchu City, Taiwan. Detailed information regarding the deposit is shown in Table 1.

[0012] Table 1. Preservation data of lactic acid bacteria strains strain name Classification Deposit number Deposit Date BBr-28 Bifidobacterium breve Bifidobacterium breve BCRC 911254 2024 / 12 / 10 PS23 Bifidobacterium animalis subsp. lactis Bifidobacterium animalissubsp.lactis BCRC 911251 2024 / 12 / 10 BR631 Bifidobacterium longum infant subspecies Bifidobacterium longumsubsp.infantis BCRC 911172 2023 / 02 / 20 BD-188 Bifidobacterium bifidum Bifidobacterium bifidum BCRC 911252 2024 / 12 / 10 PQQ-1 Lactobacillus plantarum Lactiplantibacillus plantarum BCRC 911262 2025 / 01 / 09

[0013] The combination of lactic acid bacteria strains listed in Table 1 has physiological activity functions of soothing emotions and promoting neuronal growth. Therefore, the combination of the above-mentioned lactic acid bacteria strains can be used for the purpose of soothing emotions and promoting neuronal growth.

[0014] One embodiment of the present invention provides a composition for relieving mood and promoting neuronal growth, comprising a strain of lactic acid bacteria having the effects of relieving mood and promoting neuronal growth, and a carrier. In one embodiment, the lactic acid bacteria strains include the following isolated strains: Bifidobacterium breve BBr-28 (accession number BCRC 911254); Bifidobacterium animalis subsp. lactis PS23 (accession number BCRC 911251); Bifidobacterium longum subsp. infantis BR631 (accession number BCRC 911172); Bifidobacterium bifidum BD-188 (accession number BCRC 911252); and Lactiplantibacillus plantarum PQQ-1 (accession number BCRC 911262). All of these strains are deposited at the Food Industry Development Institute. For example, the ratio of each lactic acid bacteria strain can be 1:1:1:1:1, but is not limited to this; the ratio of each lactic acid bacteria strain can be designed to an appropriate ratio based on systematic experimental results. In one embodiment, the carrier can be a physiologically acceptable excipient or diluent. Alternatively, the carrier can be a pharmaceutically acceptable excipient or diluent.

[0015] In one embodiment, the effective viable count of the lactic acid bacteria strain in the composition of the present invention is greater than or equal to 1 × 10⁶ CFU / g; in a preferred embodiment, the effective viable count of the lactic acid bacteria strain in the composition of the present invention is equal to or greater than 5 × 10⁹ CFU / g. In one embodiment, the dosage form of the composition of the present invention may be tablets, capsules, or powder for convenient oral administration. Characteristics of the lactic acid bacteria strain of the present invention:

[0016] The taxonomic characteristics of the strain were confirmed based on 16S rRNA sequence analysis and API bacterial identification system analysis results. The morphological and general characteristics of the lactic acid bacteria strain of this invention are detailed in Table 2: Table 2 Morphological and General Characteristics of the Lactic Acid Bacteria Strain of this Invention strain name Morphological characteristics Bifidobacterium breve BBr-28 strain 1. When cultured in MRS medium, the bacteria exhibit irregular shapes such as Y-shape, V-shape, or curved shape. The bacteria are small and usually appear alone, not in chains. 2. Gram-positive bacilli that do not produce spores, do not possess catalase, oxidase, or motility, grow in an absolutely anaerobic environment, with an optimal growth temperature of 37±1℃, and are facultative heterogeneous fermenting strains that do not produce gas during glucose metabolism. Bifidobacterium animalis subsp. lactis PS23 strain 1. When cultured in MRS medium, the bacteria exhibit irregular shapes such as Y-shape, V-shape, or curved shape, and usually appear alone rather than in chains. 2. Gram-positive bacilli that do not produce spores, do not possess catalase, oxidase, or motility, grow in an absolutely anaerobic environment, with an optimal growth temperature of 37±1℃, and are facultative heterogeneous fermenting strains that do not produce gas during glucose metabolism. Bifidobacterium longum infant subspecies BR631 strain 1. When cultured in MRS medium, the bacterial cells are short rod-shaped with an oval tail, and usually appear singly or in pairs of short chains. 2. Gram-positive bacilli that do not produce spores, do not possess catalase, oxidase, or motility, can grow in both aerobic and anaerobic environments, with an optimal growth temperature of 37±1℃. They are facultative heterogeneous fermenting strains that do not produce gas during glucose metabolism. Bifidobacterium bifidum BD-188 1. When cultured in MRS medium, the bacterial cell morphology is medium to short rod-shaped with an oval tail. They usually appear alone or in pairs of short chains. 2. Gram-positive bacilli that do not produce spores, do not possess catalase, oxidase, or motility, can grow in both aerobic and anaerobic environments, with an optimal growth temperature of 37±1℃. They are facultative heterogeneous fermenting strains that do not produce gas during glucose metabolism. Lactobacillus plantarum PQQ-1 1. When cultured in MRS medium, the colonies that grow are white, solid, and round, with short rod-shaped cells and rounded tails, and usually appear alone. 2. Gram-positive bacilli that do not produce spores, do not possess catalase, oxidase, or motility, can grow in both aerobic and anaerobic environments, with an optimal growth temperature of 37±1℃. They are facultative heterogeneous fermenting strains that do not produce gas during glucose metabolism. Preparation of the lactic acid bacteria powder and composition of the present invention:

[0017] The *Bifidobacterium breve* BBr-28 strain, *Bifidobacterium animalis* subsp. *lactobacter* PS23 strain, *Bifidobacterium longum* subsp. *infant* BR631 strain, and *Bifidobacterium bifidum* BD-188 strain of the present invention were isolated from breast milk; the *Lactobacillus plantarum* PQQ-1 strain was isolated from *Saussurea involucrata*. The isolated lactic acid bacteria strains were stored at -80°C with 20% glycerol. In use, 1% (v / v) inoculum was inoculated into Difco TMMRS (De Man, Rogosa and Sharpe) medium and cultured at 37°C for 20 hours. Subsequently, the five cultures were centrifuged at 4°C for 20 minutes, the supernatant was discarded, and the resulting precipitates (pellets) were freeze-dried to obtain the bacterial powder of each lactic acid bacteria strain. In one embodiment, the precipitate after centrifugation could be mixed with a protective carrier before freeze-drying to maintain better bacterial activity.

[0018] The above-mentioned lactic acid bacteria powder and carrier of the present invention are mixed evenly to form the composition of the present invention, wherein the effective viable count of the lactic acid bacteria strain is equal to or greater than 5 × 10⁹ CFU / g. In one embodiment, the carrier may contain oligosaccharides, sugar alcohols, sodium carboxymethyl cellulose, ethyl cellulose, galactooligosaccharides, stearic acid, magnesium stearate, potato starch, or a combination thereof. For example, oligosaccharides may contain fructooligosaccharides, isomaltose oligosaccharides, xylooligosaccharides, stachyose, mannose, arabinose, or a combination thereof; sugar alcohols may contain erythritol, sorbitol, isomaltitol, xylitol, mannitol, or a combination thereof. Physiological activity analysis of the composition of the present invention:

[0019] The 30 participants in the experiment were divided into two groups. One group took the capsules of the composition of the present invention twice a day, once between 2-3 pm and once 2 hours before bedtime, for 2 weeks. The other group took the capsules containing only the powder of Bifidobacterium animalis subsp. lactis PS23, serving as a control group. The other administration methods were the same. Before taking the composition of the present invention and the control group, and after 2 weeks of taking the composition, blood samples were collected from the subjects to detect changes in serotonin. In the group of the present invention, changes in dopamine, brain-derived neurotrophic factor (BDNF), and melatonin were also detected. (I) Effect of taking the composition of one embodiment of the present invention on serotonin:

[0020] Serotonin, also known as 5-hydroxytryptamine (5-HT), is a monoamine neurotransmitter responsible for transmitting mood-related signals in the brain. Serotonin regulates emotional responses; a deficiency can lead to feelings of depression, sadness, and anxiety. Conversely, sufficient serotonin levels result in more stable moods, easier relaxation, and even feelings of happiness and well-being.

[0021] Before and 2 weeks after taking the compositions of the embodiments and control groups of the present invention, approximately 5 ml of blood was collected from the subjects for serotonin detection. The collected blood was centrifuged to separate the serum. The serum was processed using serotonin ELISA kits (abnova) according to the manufacturer's instructions, and after HRP (horseradish peroxidase) catalyzed TMB (3,3',5,5'-Tetramethyl-benzidine) color development, the absorbance was measured using an ELISA Reader at OD 450 nm, and the serotonin concentration was calculated using a standard curve.

[0022] Figure 1a shows the test results of serotonin changes in subjects before and after taking the composition of an embodiment of the present invention. Figure 1b shows the test results of serotonin changes in subjects before and after taking the composition of the control group. Figure 2 shows the comparison results of serotonin changes in subjects after taking the composition of an embodiment of the present invention and the control group. The values ​​are presented as Mean ± SD. Statistical analysis was performed using paired t-tests, and p < 0.05 was considered statistically significant.

[0023] As can be seen from Figures 1a and 1b, after 2 weeks of taking the composition of the present invention and the control group, the serotonin levels in the blood of the subjects increased significantly compared to before taking the composition (p < 0.001). Furthermore, comparing the change in serotonin levels (i.e., the difference between serotonin levels before and after taking the composition of the present invention) with the control group, the increase in serotonin levels after taking the composition of the present invention was superior to that of the control group, as shown in Figure 2, and the difference was significant (p = 0.0119). Therefore, the composition of the present invention containing five strains of lactic acid bacteria significantly increased serotonin levels compared to the control group containing only a single strain of lactic acid bacteria. (II) Effect of taking the composition of the present invention on dopamine:

[0024] Dopamine is a neurotransmitter that the human body can produce on its own. The nervous system uses dopamine to transmit messages between nerve cells. Dopamine plays an important role in making people feel happy, and it helps people to strive, concentrate, and be interested in things. When dopamine is insufficient, people may experience problems such as fatigue, poor concentration, lack of motivation, and mood swings.

[0025] Blood samples were collected from subjects before and two weeks after taking the composition of the present invention for dopamine detection. The collected blood was centrifuged to separate serum. The serum was processed using Dopamine ELISA kits (abnova, KA1887) according to the manufacturer's manual. After color development, the absorbance was measured using an ELISA Reader at OD 450 nm, and the dopamine concentration was calculated using a standard curve.

[0026] Figure 3 shows the experimental results of changes in dopamine levels in subjects before and after taking the composition of an embodiment of the present invention. Statistical analysis was performed using a paired t-test, with p < 0.05 considered statistically significant. As can be seen from Figure 3, after taking the composition of an embodiment of the present invention, the dopamine levels in the blood of most subjects increased significantly, and there was a significant difference compared to before taking the composition of an embodiment of the present invention (p = 0.0073). Therefore, the composition of an embodiment of the present invention has the physiological activity of promoting the production of dopamine in the human body, thereby having the effect of soothing the user's emotions, especially the problem of low mood caused by dopamine deficiency. (III) Effect of taking the composition of an embodiment of the present invention on brain-derived neurotrophic factor (BDNF):

[0027] Brain-derived neurotrophic factor (BDNF) is the most abundant nerve growth factor in the brain. It promotes the growth of neurons (nerve cells) and the formation of synapses in brain nerve cells, and regulates the transmission of neurotransmitters and intracellular information. Studies have shown that a lack of BDNF may cause some cognitive impairments, such as Alzheimer's disease, one of the common degenerative dementias.

[0028] Blood samples were collected from subjects before and two weeks after taking the composition of the present invention for BDNF detection. The collected blood was centrifuged to separate serum. The serum was processed using BDNF ELISA kits (Aviva, OKBB00127) according to the manufacturer's instructions. After color development, the absorbance was measured using an ELISA Reader at OD 450 nm, and the BDNF concentration was calculated using a standard curve.

[0029] Figure 4 shows the experimental results of changes in BDNF in subjects before and after taking the composition of an embodiment of the present invention. Statistical analysis was performed using a paired t-test, with p < 0.05 considered statistically significant. As can be seen from Figure 4, after taking the composition of an embodiment of the present invention, the BDNF levels in the blood of most subjects increased significantly, and the difference was statistically significant compared to before taking the composition of an embodiment of the present invention (p = 0.0111). Therefore, the composition of an embodiment of the present invention has the physiological activity of promoting the production of BDNF in the human body, and thus has the effect of promoting neuronal growth, thereby reducing neurocognitive dysfunction diseases such as Alzheimer's disease. (IV) Effect of taking the composition of an embodiment of the present invention on melatonin:

[0030] Patients with depression often experience sleep disturbances and exhibit circadian rhythm disruptions. Clinically, poor sleep often exacerbates the depression in patients with depression. Melatonin plays a role in regulating the biological clock; generally, melatonin secretion is lower during the day and increases rapidly at night. Higher melatonin secretion during the day may lead to poor daytime mental state.

[0031] Blood samples were collected from subjects before and two weeks after taking the composition of this invention for melatonin testing. It should be noted that the blood samples were collected during the day. The collected blood was centrifuged to separate the serum. The serum was processed using Melatonin ELISA kits (Aviva, OKEH02566) according to the manufacturer's instructions. After color development, the absorbance was measured using an ELISA Reader at OD 450 nm, and the melatonin concentration was calculated using a standard curve.

[0032] Figure 5 shows the experimental results of changes in melatonin levels in subjects before and after taking the composition of an embodiment of the present invention. Statistics were calculated using a paired t-test, with p < 0.05 considered statistically significant. As can be seen from Figure 5, after taking the composition of an embodiment of the present invention, melatonin levels in the blood of most subjects were significantly reduced, and the difference was statistically significant compared to before taking the composition of an embodiment of the present invention (p = 0.0028), indicating that better sleep quality was achieved after taking the composition of an embodiment of the present invention. Therefore, the composition of an embodiment of the present invention has a physiological activity effect of adjusting the biological clock, and thus has the effect of improving sleep quality and relieving mood.

[0033] In summary, the compositions of the embodiments of the present invention have physiological activities that soothe emotions and promote neuronal growth. It should be noted that, compared with compositions containing only a single lactic acid bacteria strain, the compositions of the embodiments of the present invention containing five lactic acid bacteria strains can significantly increase serotonin, thereby soothing emotions. Thus, it can be seen that the combination of lactic acid bacteria strains of the embodiments of the present invention has unexpected effects.

[0034] The above-described embodiments are only for illustrating the technical ideas and features of the present invention. Their purpose is to enable those skilled in the art to understand the content of the present invention and implement it accordingly. They should not be used to limit the patent scope of the present invention. That is, all equivalent changes or modifications made in accordance with the spirit disclosed in the present invention should still be covered within the patent scope of the present invention. [Simplified Explanation of the Diagram]

[0035] Figure 1a shows the test results of changes in serotonin levels in subjects before and after taking the composition of an embodiment of the present invention.

[0036] Figure 1b shows the test results of changes in serotonin levels before and after the subjects took the control group's composition.

[0037] Figure 2 shows the comparison results of serotonin changes after subjects took the composition of an embodiment of the present invention and the control group.

[0038] Figure 3 shows the test results of dopamine changes in subjects before and after taking the composition of an embodiment of the present invention.

[0039] Figure 4 shows the experimental results of changes in brain-derived neurotrophic factor (BDNF) in subjects before and after taking the composition of an embodiment of the present invention.

[0040] Figure 5 shows the test results of changes in melatonin levels in subjects before and after taking the composition of an embodiment of the present invention. [Biomaterial Storage]

[0042] 1. Taiwan, Republic of China, Food Industry Development Institute, 2024 / 12 / 10, BCRC 911254 2. Taiwan, Republic of China, Food Industry Development Institute, 2024 / 12 / 10, BCRC 911251 3. Taiwan, Republic of China, Food Industry Development Institute, 2023 / 02 / 20, BCRC 911172 4. Taiwan, Republic of China, Food Industry Development Institute, 2024 / 12 / 10, BCRC 911252 5. Taiwan, Republic of China, Food Industry Development Institute, 2025 / 01 / 09, BCRC 911262

Claims

1. A composition for soothing emotions and promoting neuronal growth, comprising: a strain of lactic acid bacteria having the effects of soothing emotions and promoting neuronal growth, the lactic acid bacteria strain comprising: isolated *Bifidobacterium breve* BBr-28 strain, accession number BCRC 911254; *Bifidobacterium animalis subsp. lactis* PS23 strain, accession number BCRC 911251; *Bifidobacterium longum subsp. infantis* BR631 strain, accession number BCRC 911172; *Bifidobacterium bifidum* BD-188 strain, accession number BCRC 911252; and *Lactiplantibacillus plantarum* PQQ-1 strain, accession number BCRC 911262, the above strains are all stored at the Food Industry Development Institute; and the carrier contains oligosaccharides, sugar alcohols, sodium carboxymethyl cellulose, ethyl cellulose, galactooligosaccharides, stearic acid, magnesium stearate, potato starch or a combination thereof.

2. The composition for relieving mood and promoting neuronal growth as described in claim 1, wherein the effective viable count of the lactic acid bacteria strain is equal to or greater than 5 × 10⁹ CFU / g.

3. The composition for relieving mood and promoting neuronal growth as described in claim 1, wherein the carrier is a physiologically acceptable excipient or diluent.

4. The composition for relieving mood and promoting neuronal growth as described in claim 1, wherein the carrier is a pharmaceutically acceptable excipient or diluent.

5. The composition for relieving mood and promoting neuronal growth as described in claim 1, wherein the dosage form is a tablet, capsule or powder.

6. Use of a lactic acid bacteria strain in the preparation of a composition for soothing mood and promoting neuronal growth, wherein the composition comprises: a lactic acid bacteria strain having activity for soothing mood and promoting neuronal growth, the lactic acid bacteria strain comprising the isolated *Bifidobacterium breve* strain BBr-28, accession number BCRC 911254; *Bifidobacterium animalis subsp. lactis* strain PS23, accession number BCRC 911251; *Bifidobacterium longum subsp. infantis* strain BR631, accession number BCRC 911172; *Bifidobacterium bifidum* strain BD-188, accession number BCRC 911252; and *Lactiplantibacillus plantarum*. The PQQ-1 strain, with accession number BCRC 911262, was deposited at the Food Industry Development Institute; along with the carrier.

7. The use as described in claim 6, wherein the effective viable count of the lactic acid bacteria strain is equal to or greater than 5 × 10⁹ CFU / g.

8. The use as described in claim 6, wherein the carrier is a physiologically acceptable excipient or diluent.

9. The use as described in claim 6, wherein the carrier is a pharmaceutically acceptable excipient or diluent.

10. For the purposes described in claim 6, the dosage form is tablets, capsules or powder.

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