Composition containing gamma-aminobutyric acid and application thereof

Abstract

The invention discloses a composition containing gamma-aminobutyric acid. The composition containing gamma-aminobutyric acid comprises a composition body with the sleep improving effect, and it is found through research that the composition body which is composed of gamma-aminobutyric acid, an elderberry extract, a lily extract, a spearmint extract and human milk oligosaccharide and has the sleep improving effect can remarkably prolong the sleep duration of a mouse with normal sleep; moreover, secretion of growth hormone and melatonin is remarkably improved, and the compound can be used for preparing health-care foods or medicines for improving insomnia and promoting growth and development.

Description

Technical Field

[0001] This invention relates to the technical field of improving sleep and promoting growth hormone secretion, and specifically to a composition containing γ-aminobutyric acid and its application. Background Technology

[0002] Sleep disorders have become a prominent global health problem. Chronic insomnia not only leads to cognitive decline but also inhibits the secretion of growth hormone (GH), causing irreversible damage, especially to children's growth and development. Currently, clinical treatment mainly relies on sedative drugs (such as benzodiazepines). Hypoglycemic agents (such as GH) and melatonin supplements can improve sleep, but the former has side effects such as addiction and drowsiness the next day, while the latter has limited effect on promoting GH secretion.

[0003] In the field of promoting growth hormone secretion, recombinant human growth hormone (rhGH) injection therapy is effective, but it is expensive and requires long-term invasive administration, resulting in poor patient compliance. Gamma-aminobutyric acid (GABA), as an endogenous neurotransmitter, has been shown to promote GH release by activating the hypothalamic-pituitary axis, but oral GABA alone has significant drawbacks: firstly, its bioavailability is low; secondly, while high doses can increase GH levels, they may cause side effects such as blood pressure fluctuations.

[0004] In recent years, natural compound compositions have become a research hotspot, but existing technologies still have bottlenecks such as insufficient synergy of components, process limitations, and limited functionality.

[0005] Therefore, there is an urgent need to develop a safe, multi-target synergistic compound composition that can simultaneously improve sleep quality and GH secretion levels by optimizing the ingredient ratio and key raw material processes, thus overcoming the limitations of existing technologies. Summary of the Invention

[0006] To overcome the aforementioned defects and shortcomings in the prior art, the present invention provides a composition containing γ-aminobutyric acid and its application.

[0007] In a first aspect, the present invention provides a composition containing γ-aminobutyric acid.

[0008] The composition contains the following parts by weight of raw materials:

[0009] Elderberry extract: 0.2-4 parts;

[0010] Lily extract: 1-6 parts;

[0011] Spearmint extract: 0.2-2 parts;

[0012] γ-Aminobutyric acid: 0.5-5 parts;

[0013] Human milk oligosaccharides: 0.1-1 part.

[0014] The composition prepared from the above-mentioned raw materials, which has the effects of improving sleep and promoting growth and development, can significantly increase the total sleep duration, significantly increase the melatonin content in serum, and significantly promote the secretion of growth hormone by pituitary cells.

[0015] The composition prepared from the above-mentioned raw materials, which has the effects of improving sleep and promoting growth and development, can more significantly increase the total sleep duration, more significantly increase the melatonin content in serum, and more significantly promote the secretion of growth hormone by pituitary cells.

[0016] Preferably, the preparation of the elderberry extract in the composition includes the following steps:

[0017] A1: Clean and dry the elderberries, crush them, and sieve them to obtain elderberry powder;

[0018] A2: Reflux extract elderberry powder with 80-90wt% ethanol-water solution at 40-50℃ for 0.5-1.5h, filter, and obtain the extract. Repeat 2-3 times, combine the extracts, and obtain elderberry extract.

[0019] A3: The elderberry extract was filtered using a cellulose dialysis bag to obtain a filtrate with a molecular weight cutoff of 500 Da-2 kDa;

[0020] A4: Freeze-dry the 500Da-2kDa filtrate to obtain elderberry extract powder.

[0021] The composition prepared from the above-mentioned raw materials, which has the effects of improving sleep and promoting growth and development, can more significantly increase the total sleep duration, more significantly increase the melatonin content in serum, and more significantly promote the secretion of growth hormone by pituitary cells.

[0022] Preferably, the composition having the effects of improving sleep and promoting growth and development also includes excipients acceptable for health products or pharmaceuticals.

[0023] Furthermore, the acceptable excipients in the health products include at least one of egg yolk powder, resistant dextrin, and maltodextrin.

[0024] In a second aspect, the present invention provides a health product containing a γ-aminobutyric acid composition, the health product comprising the following components in parts by weight:

[0025] The γ-aminobutyric acid-containing composition described in the first aspect: 10-11 parts;

[0026] Complex protein: 5-20 servings;

[0027] Multivitamin and mineral supplement 0.1-0.5 servings.

[0028] Furthermore, the complex protein is composed of whey protein, collagen, and colostrum basic protein in a mass ratio of 1:0.5-0.7:0.6-0.8; the complex vitamin and mineral is composed of milk calcium, vitamin D, and zinc lactate in a mass ratio of 1:0.2-0.4:0.1-0.3.

[0029] Compared with the prior art, the present invention has the following beneficial effects:

[0030] This invention discloses a composition containing γ-aminobutyric acid (GABA) and its applications. Research in this invention has found that the combined use of elderberry extract, lily extract, spearmint extract, GABA, and human milk oligosaccharides in the mass ratio specified in this application can significantly increase the total sleep duration in mice, increase melatonin levels in mouse serum, and promote the secretion of growth hormone by pituitary cells. This composition can be used to prepare pharmaceuticals and health products that improve insomnia and promote growth and development, and has broad application prospects in improving or treating sleep disorders and growth retardation. Detailed Implementation

[0031] The present invention will be further illustrated below with reference to specific embodiments, but the embodiments do not limit the present invention in any way. Unless otherwise specified, the reagents, methods, and equipment used in the present invention are conventional reagents, methods, and equipment in this technical field.

[0032] The raw materials used in this application and their sources are as follows:

[0033] Elderberry fruit: purchased from Guangdong Qingyunshan Pharmaceutical Co., Ltd.;

[0034] Lily extract: purchased from Guangdong Qingyunshan Pharmaceutical Co., Ltd.;

[0035] Spearmint extract: purchased from Kemin Foods, LC model: K110-42;

[0036] γ-Aminobutyric acid: purchased from Shandong Yangcheng Biotechnology Co., Ltd.;

[0037] Human milk oligosaccharides (HMOs): purchased from Hangzhou Xinfu Technology Co., Ltd.;

[0038] Elderberry extract: purchased from Ganzhou Helvkangjian Biotechnology Co., Ltd.;

[0039] Whey protein: purchased from Dairy Best Victoria Pty Ltd.

[0040] Unless otherwise specified, all reagents and materials used in the following experimental examples are commercially available.

[0041] A composition containing γ-aminobutyric acid, comprising the following raw materials in parts by weight:

[0042] Composition 1:

[0043] Elderberry extract: 3 parts;

[0044] Lily extract: 4 parts;

[0045] Spearmint extract: 1 part;

[0046] γ-Aminobutyric acid: 2 parts;

[0047] Human milk oligosaccharides: 0.5 parts;

[0048] The preparation method of elderberry extract includes the following steps:

[0049] A1: Clean and dry the elderberries, crush them, and pass them through an 80-mesh sieve to obtain elderberry powder;

[0050] A2: Elderberry powder was refluxed with 85wt% ethanol-water solution at 45℃ for 1h, filtered, and the extract was obtained. The extraction was repeated 3 times and the extracts were combined to obtain elderberry extract.

[0051] A3: The elderberry extract was filtered using a cellulose dialysis bag to obtain a filtrate with a molecular weight cutoff of 500 Da-2 kDa;

[0052] A4: Freeze-dry the 500Da-2kDa filtrate to obtain elderberry extract powder.

[0053] Composition 2:

[0054] Elderberry extract: 0.2 parts;

[0055] Lily extract: 6 parts;

[0056] Spearmint extract: 2 parts;

[0057] γ-Aminobutyric acid: 5 parts;

[0058] Human milk oligosaccharides: 1 part;

[0059] The preparation method of elderberry extract includes the following steps:

[0060] A1: Clean and dry the elderberries, crush them, and pass them through an 80-mesh sieve to obtain elderberry powder;

[0061] A2: Reflux extract elderberry powder with 85wt% ethanol-water solution at 45℃ for 1h, filter, and obtain extract. Repeat 3 times, combine extracts to obtain elderberry extract.

[0062] A3: The elderberry extract was filtered using a cellulose dialysis bag to obtain a filtrate with a molecular weight cutoff of 500 Da-2 kDa;

[0063] A4: Freeze-dry the 500Da-2kDa filtrate to obtain elderberry extract powder.

[0064] Composition 3:

[0065] Elderberry extract: 4 parts;

[0066] Lily extract: 1 part;

[0067] Spearmint extract: 0.2 parts;

[0068] γ-Aminobutyric acid: 0.5 parts;

[0069] Human milk oligosaccharides: 0.1 part;

[0070] The preparation method of elderberry extract includes the following steps:

[0071] A1: Clean and dry the elderberries, crush them, and pass them through an 80-mesh sieve to obtain elderberry powder;

[0072] A2: Reflux extract elderberry powder with 85wt% ethanol-water solution at 45℃ for 1h, filter, and obtain extract. Repeat 3 times, combine extracts to obtain elderberry extract.

[0073] A3: The elderberry extract was filtered using a cellulose dialysis bag to obtain a filtrate with a molecular weight cutoff of 500 Da-2 kDa;

[0074] A4: Freeze-dry the 500Da-2kDa filtrate to obtain elderberry extract powder.

[0075] Composition A:

[0076] Unlike composition 1, in step A2 of the method for preparing elderberry extract, 85wt% ethanol-water solution is replaced with 95wt% ethanol-water solution, while the remaining steps, raw materials and mass fractions are the same as those in composition 4.

[0077] Composition B:

[0078] Unlike composition 1, in step A2 of the method for preparing elderberry extract, 85wt% ethanol-water solution is replaced with 75wt% ethanol-water solution, while the remaining steps, raw materials and mass parts are the same as those in composition 4.

[0079] Composition C:

[0080] Unlike composition 1, in step A3 of the method for preparing elderberry extract, the molecular weight cutoff of the filtrate is >2kDa, while the remaining steps, raw materials and mass fractions are the same as those in composition 4.

[0081] Composition D:

[0082] Unlike composition 1, in step A3 of the method for preparing elderberry extract, the molecular weight cutoff of the filtrate is <500 Da, while the remaining steps, raw materials and mass fractions are the same as those in composition 4.

[0083] Composition E:

[0084] Unlike composition 1, the method for preparing elderberry extract does not perform step A3. Instead, the elderberry extract obtained in step A2 is freeze-dried to obtain elderberry extract powder. The remaining steps, raw materials, and mass fractions are the same as those in composition 4.

[0085] Composition F:

[0086] Unlike composition 1, the elderberry extract is a commercially available elderberry extract.

[0087] Preparation method of composition 1:

[0088] At room temperature, place all ingredients in a container, mix thoroughly, seal, and store away from light.

[0089] The preparation methods of compositions 2-3 and AF are the same as those of composition 1.

[0090] A health product with sleep-aiding effects comprises the following ingredients by weight:

[0091] Health Supplement 1:

[0092] Composition 1: 10.5 parts;

[0093] Complex protein: 15 servings;

[0094] Multivitamin and mineral supplement: 0.5 servings

[0095] The complex protein consists of whey protein, collagen, and colostrum basic protein in a mass ratio of 1:0.6:0.7.

[0096] The compound microbial minerals consist of milk calcium, vitamin D, and zinc lactate in a mass ratio of 1:0.3:0.2.

[0097] Health Supplement 2:

[0098] Composition 1: 11 parts;

[0099] Complex protein: 5 servings;

[0100] Multivitamin and mineral supplement: 0.3 servings

[0101] The complex protein consists of whey protein, collagen, and colostrum basic protein in a mass ratio of 1:0.7:0.6.

[0102] The compound microbial minerals consist of milk calcium, vitamin D, and zinc lactate in a mass ratio of 1:0.1:0.4.

[0103] Health Supplement 3:

[0104] Composition 1: 10 parts;

[0105] Complex protein: 20 servings;

[0106] Multivitamin and mineral supplement: 0.1 serving

[0107] The complex protein consists of whey protein, collagen, and colostrum basic protein in a mass ratio of 1:0.5:0.8.

[0108] The compound microbial minerals consist of milk calcium, vitamin D, and zinc lactate in a mass ratio of 1:0.2:0.3.

[0109] Preparation method of health products with sleep-aiding effects: Mix the composition, compound protein, compound vitamins and minerals evenly at room temperature, sterilize, pack in bags, and seal for storage.

[0110] Experimental Example 1: Sleep Induction Experiment

[0111] Experimental animals: Kunming mice, weighing 20±2g, half male and half female, clean grade.

[0112] Test samples: Test group: Compositions 1-3, composition AF, elderberry extract (self-made, preparation method is the same as composition 1), elderberry extract (commercially available), lily extract, spearmint extract, γ-aminobutyric acid, and human milk oligosaccharide were prepared into a 1 mg / mL concentration mixture using physiological saline, sealed and protected from light for later use.

[0113] Test method:

[0114] Animal grouping: The experimental animals were divided into a normal control group (1 group), a model control group (1 group), and an experimental group (15 groups), with 10 animals in each group.

[0115] Animal modeling: Mice that had been adapted to the diet, except for the normal control group, were injected intraperitoneally with chlorophenylalanine (PCPA) suspension for two consecutive days at a dose of 450 mg / kg body weight. The normal control group was injected intraperitoneally with the same volume of physiological saline. When mice other than the normal control group showed circadian rhythm disorder, significant weight loss, dry and frizzy fur, increased excitability and aggression, significantly increased sleep latency, and significantly shortened sleep time, it indicated that the insomnia model was successfully established.

[0116] Animal administration: The experimental group was given 5 mL / kg body weight of the test sample, while the model control group and the normal control group were given the same amount of physiological saline. The administration was carried out for 28 consecutive days.

[0117] Sleep test: 30 minutes after the last day of administration, sodium pentobarbital (45 mg / kg body weight) was injected intraperitoneally. The disappearance of the righting reflex was used as the standard. The sleep latency (the time from the injection of sodium pentobarbital to the disappearance of the righting reflex) and the sleep duration (the time from the disappearance of the righting reflex to its recovery) were measured and recorded. The test results are shown in Table 1.

[0118] Serum testing:

[0119] After the experiment, blood was collected from the decapitated experimental animals. After standing for 2 hours, the blood was centrifuged at 3000 rpm for 15 minutes to separate the serum. An appropriate amount of mouse serum was taken, and the serum MT (melatonin) level was detected according to the ELISA kit instructions. The MT content growth rate was calculated. MT content growth rate (%) = (experimental group measurement value - model control group measurement value) / model control group measurement value × 100%.

[0120] Table 1. Effects of the composition on sleep duration and serum MT levels in mice.

[0121]

[0122]

[0123] Note: "*" indicates comparison between the model group and the normal control group, p < 0.05; "#" indicates comparison between the experimental group and the model control group, p < 0.05; "a" indicates comparison between the experimental group and composition 1, p < 0.05; "b" indicates comparison between elderberry (homemade) and elderberry (commercially available), p < 0.05; "-" indicates no data.

[0124] By comparing the model control group with the normal control group in Table 1, the sleep latency was significantly increased and the sleep duration was significantly shortened, indicating that the model was successfully established.

[0125] Comparison of the results between the model control group and the experimental group showed that there were significant changes in sleep latency, sleep duration, and serum MT levels in both groups, indicating that the experimental sample had a significant impact on sleep latency, sleep duration, and serum MT levels.

[0126] Comparing the results of elderberry extract (commercially available) and elderberry extract (self-made) in Table 1, it can be seen that the elderberry extract prepared by the preparation method defined in this invention has better sleep-aiding effects and can increase serum MT levels compared to commercially available elderberry extract.

[0127] Comparing the results of Elderberry Extract (commercially available), Elderberry Extract (self-made), Lily Extract, Spearmint Extract, γ-aminobutyric acid (GABA), Human Milk Oligosaccharide, and Composition 1 in Table 1, it can be seen that the combined use of Elderberry Extract (self-made), Lily Extract, Spearmint Extract, GABA, and Human Milk Oligosaccharide in the mass ratios specified in this application has a more significant effect in reducing sleep latency, increasing sleep time, and increasing the MT content in mouse serum. This indicates that Elderberry Extract, Lily Extract, Spearmint Extract, GABA, and Human Milk Oligosaccharide have a significant synergistic effect.

[0128] Comparison of the results of composition 1 and composition AF shows that the elderberry extract prepared using the preparation steps specified in this application can significantly reduce sleep latency and increase the total sleep duration and MT secretion in experimental animals.

[0129] Experimental Example 2: Effect of the composition on promoting growth hormone secretion by GH3 cells in rat pituitary tumors

[0130] Test samples: Test group: Composition 1, Composition AF, elderberry extract (self-made, preparation method is the same as Composition 1), elderberry extract (commercially available), lily extract, spearmint extract, γ-aminobutyric acid, and human milk oligosaccharide were prepared into a 1 mg / mL concentration mixture using cell culture medium, sealed and protected from light for later use; blank group was cell culture medium.

[0131] Cells: Rat pituitary tumor GH3 cells;

[0132] Culture medium: 90 v / v % DMEM medium + 10 v / v % FBS;

[0133] Culture conditions: Gas phase: 95% air; 5% carbon dioxide; Temperature: 37℃; Humidity: 70-80%.

[0134] Methods: Rat pituitary adenoma GH3 cells in logarithmic growth phase were injected with 1×10⁻⁶ cells. 3 Cells were seeded in 96-well plates. After cell attachment, 20 μL of cell culture medium per well was discarded, and 20 μL of experimental sample per well was added. The plates were incubated at 37°C for 48 h. After incubation, the concentration of growth hormone in the cell supernatant was measured using an ELISA kit (Rat Growth Hormone (GH) ELISA Kit, ELISA Biotechnology, ml002921), strictly following the instructions in the kit's manual. Results are expressed as mean ± standard deviation, with six replicates per well.

[0135] Growth hormone secretion growth rate (%) = (Experimental group growth hormone measurement value - Control group growth hormone measurement value) / Control group growth hormone measurement value × 100%

[0136] Table 2. Effects of the composition on promoting growth hormone secretion by GH3 pituitary tumor cells in rats (N=6)

[0137]

[0138]

[0139] Note: "*" indicates that compared with composition 1, p < 0.05.

[0140] The comparison of the results of elderberry extract (self-made), elderberry extract (commercially available), lily extract, spearmint extract, γ-aminobutyric acid (GABA), human milk oligosaccharides and composition 1 in Table 2 shows that the combined use of elderberry extract, lily extract, spearmint extract, GABA and human milk oligosaccharides in the mass ratios specified in this application has a more significant effect on increasing growth hormone secretion, indicating that elderberry extract, lily extract, spearmint extract, GABA and human milk oligosaccharides have a significant synergistic effect.

[0141] The results of comparing composition 1 and composition AF show that the elderberry extract extracted by the elderberry extract extraction method defined in this application significantly increases the amount of growth hormone secreted.

[0142] Test Example 3: Acute Toxicity Test

[0143] Test sample: Health product 1 was prepared into a 250mg / mL mixture using deionized water.

[0144] Experimental Methods: Thirty male SD rats weighing 200g-220g were selected. The rats were administered oral gavage twice daily at 6-hour intervals, with each gavage volume being 20mL / kg body weight, equivalent to a dose of 10.00g / kg body weight. The rats were fasted for 6 hours before the first gavage. They were observed for two weeks after gavage, and the signs of poisoning and mortality were recorded.

[0145] Experimental Results: No obvious symptoms of poisoning were observed in SD rats after gavage, and no deaths were observed during the 14-day observation period. At the end of the observation period, the test animals were euthanized and dissected. No obvious abnormalities were found in the major organs such as the liver, spleen, kidneys, intestines, heart, and lungs. The median lethal dose (LD50) of the health product described in this invention for SD rats was greater than 10.00 g / kg·bw. According to the acute toxicity dose grading standard in GB15193.3-2014, it belongs to the practically non-toxic category.

[0146] The embodiments described above are some, but not all, of the embodiments of this application. The detailed description of the embodiments of this application is not intended to limit the scope of the claimed application, but merely to illustrate selected embodiments. All other embodiments obtained by those skilled in the art based on the embodiments of this application without inventive effort are within the scope of protection of this application.

Claims

1. A γ-aminobutyric acid composition, characterized in that, Includes the following quantities of raw materials: Elderberry extract: 0.2-4 parts; Lily extract: 1-6 parts; Spearmint extract: 0.2-2 parts; γ-Aminobutyric acid: 0.5-5 parts; Human milk oligosaccharides: 0.1-1 part.

2. The composition according to claim 1, characterized in that, Includes the following quantities of raw materials: Elderberry extract: 2-4 parts; Lily extract: 3-4 parts; Spearmint extract: 0.5-1.5 parts; γ-Aminobutyric acid: 1-3 parts; Human milk oligosaccharides: 0.1-0.5 parts.

3. The composition according to claim 1 or 2, characterized in that, The elderberry extract includes the following preparation steps: A1: Clean and dry the elderberries, crush them, and sieve them to obtain elderberry powder; A2: Reflux extract elderberry powder with 80-90wt% ethanol-water solution at 40-50℃ for 0.5-1.5h, filter, and obtain the extract. Repeat 2-3 times, combine the extracts, and obtain elderberry extract. A3: The elderberry extract was filtered using a cellulose dialysis bag to obtain a filtrate with a molecular weight cutoff of 500 Da-2 kDa; A4: Freeze-dry the 500Da-2kDa filtrate to obtain elderberry extract powder.

4. The composition according to any one of claims 1-3, characterized in that, The composition also includes excipients acceptable for health products or pharmaceuticals.

5. The composition according to claim 4, characterized in that, The health supplement may accept at least one of the following excipients: egg yolk powder, resistant dextrin, and maltodextrin.

6. The use of the composition according to any one of claims 1-3 in the preparation of a product having sleep-improving effects.

7. The use of the composition according to any one of claims 1-3 in the preparation of a product having the function of promoting growth and development.

8. A health product containing a γ-aminobutyric acid composition, characterized in that, The health supplement contains the following components by weight: The composition according to any one of claims 1-3: 10-11 parts; Complex protein: 5-20 servings; Multivitamin and mineral supplement 0.1-0.5 servings.

9. The health product according to claim 8, characterized in that, The complex protein is composed of whey protein, collagen, and colostrum basic protein in a mass ratio of 1:0.5-0.7:0.6-0.

8.

10. The health product according to claim 8, characterized in that, The compound vitamin and mineral mixture consists of milk calcium, vitamin D, and zinc lactate in a mass ratio of 1:0.2-0.4:0.1-0.3.

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