Composition for improving sleep as well as preparation method and preparation thereof

Abstract

The invention belongs to the technical field of biological medicines, and particularly relates to a composition for improving sleep as well as a preparation method and a preparation thereof. The composition is prepared from the following raw materials: hydroxytyrosol and urolithin A, the mass ratio of the hydroxytyrosol to the urolithin A is 1: (0.1-5). The composition provided by the invention can improve the content of neurotransmitters 5-HT and GABA (gamma-aminobutyric acid), enhance the antioxidant capacity of the brain and further improve sleep. Moreover, the urolithin A and the hydroxytyrosol are small molecular substances, and can pass through a blood brain barrier to reach the brain, so that the effect of improving sleep is achieved. The composition for improving sleep, which is prepared by taking hydroxytyrosol and urolithin as raw materials, not only can be used for improving sleep, but also has no side effect and definite functional components.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and specifically relates to a composition for improving sleep and a preparation method and a preparation thereof. Background Art

[0002] The "2024 Chinese Residents' Healthy Sleep White Paper" jointly released by the China Sleep Research Society and other institutions shows that half of the residents do not get enough sleep at night, and 21% of the residents have poor sleep quality. Waking up easily / early and having difficulty falling asleep are the main sleep problems. 59% of people have symptoms of insomnia, and only 19% have no sleep disorders at all. Sleep problems bother many people, seriously affecting daily life, and even gradually leading to the occurrence of many chronic diseases.

[0003] At present, the drugs for improving sleep are mainly sedatives, hypnotic drugs, melatonin and traditional Chinese medicine. Sedatives and hypnotic drugs have large side effects, are addictive, and have obvious withdrawal symptoms, making it difficult to fundamentally solve the problem. Melatonin, when taken for a long time, can cause digestive tract reactions, endocrine disorders, and central nervous system reactions. Traditional Chinese medicine, because its effective ingredients are not clear, needs to be taken for a long time.

[0004] In summary, existing sleep-improving products have the disadvantages of large side effects and unclear efficacy ingredients. Therefore, it is very necessary to find a safe, clearly ingredient, and effective sleep-improving product. Summary of the invention

[0005] In order to solve the problem that the sleep-improving products in the prior art have large side effects and unclear functional components, the present invention provides a composition for improving sleep and a preparation method and preparation thereof. To achieve the above purpose, the present invention adopts the following technical solution.

[0006] The present invention provides a composition for improving sleep, which is made of the following raw materials: hydroxytyrosol and urolithin A; the mass ratio of the hydroxytyrosol to the urolithin A is 1:0.1-5. Mitochondria are one of the main sources of biological active oxygen, and improving mitochondrial function can reduce oxidative stress. Urolithin A inhibits neuroinflammation and neurotoxicity by inducing mitochondrial autophagy. Hydroxytyrosol can activate PGC-1α, induce the expression of nuclear respiratory factor NRF1 / 2, and then activate mitochondrial transcription factor Tfam to promote mitochondrial regeneration. Hydroxytyrosol assists the function of urolithin A from the perspective of improving mitochondria, thereby enhancing the antioxidant capacity of urolithin A and avoiding neuronal damage. Neurons can secrete neurotransmitters that regulate sleep, so hydroxytyrosol improves the effect of urolithin A on improving sleep by improving the antioxidant capacity of urolithin A; hydroxytyrosol can regulate intestinal microbiota to improve intestinal health. The intestine can produce a variety of neurotransmitters such as 5-HT and GABA, which affect the brain and sleep through the brain-gut axis. Therefore, hydroxytyrosol further increases the production of 5-HT and GABA neurotransmitters through its antioxidant properties or regulates the intestines, thereby improving sleep. And both raw materials are specific ingredients, present in natural plants or are metabolites of plants in the body, without side effects. Therefore, the composition for improving sleep prepared by using hydroxytyrosol and urolithin as raw materials in the present invention can not only be used to improve sleep, but also has no side effects and clear functional components. The composition for improving sleep provided by the present invention solves the problem that the side effects of sleep-improving products in the prior art are large and the functional components are unclear.

[0007] Preferably, it is made of the following raw materials in parts by weight: hydroxytyrosol and urolithin A; the mass ratio of the hydroxytyrosol to the urolithin A is 1:0.2-3.

[0008] Preferably, it is made of the following raw materials in parts by weight: hydroxytyrosol and urolithin A; the mass ratio of the hydroxytyrosol to the urolithin A is 1:1.

[0009] The present invention also provides a method for preparing the composition for improving sleep, comprising the following steps: respectively weighing hydroxytyrosol and urolithin A according to a mass ratio, and mixing them evenly to obtain the composition for improving sleep.

[0010] The present invention also provides a preparation for improving sleep, which comprises the composition for improving sleep and a pharmaceutically acceptable auxiliary material or carrier.

[0011] Preferably, the preparation is an internal preparation or an external preparation.

[0012] Preferably, the oral preparation is any one of tablets, powders, pills, capsules and oral liquids.

[0013] Preferably, the external preparation is any one of nasal drops, sprays, gels, emulsions, powders and aerosols.

[0014] Preferably, the auxiliary materials include any one or more of maltodextrin, sodium carboxymethyl cellulose, lactose, silicon dioxide, magnesium stearate, carbomer, chitosan, β-cyclodextrin and phospholipids.

[0015] Preferably, the carrier comprises any one or more of liposomes, cyclodextrin inclusion compounds, microspheres and nanoparticles.

[0016] After the sleep-improving composition provided by the present invention is embedded in a carrier, the solubility of the composition can be improved, the action time can be prolonged, and the absorption rate can be increased; the external dosage form of the product is mainly absorbed through the nose and directly enters the brain through the olfactory nerve or the trigeminal nerve, thereby avoiding the gastrointestinal tract from damaging the composition and avoiding the obstructive effect of the blood-brain barrier, and the absorption and onset of effect are faster than those of oral administration and other routes of administration.

[0017] Compared with the prior art, the present invention has the following beneficial effects:

[0018] 1. The present invention provides a composition for improving sleep. The raw materials used in the present invention are all natural ingredients or metabolites of natural ingredients in the body. The raw materials are safe, and the functional components are clear and effective. The composition for improving sleep provided by the present invention has the effects of regulating neurotransmitters, reducing oxidative stress, improving sleep, shortening sleep latency, and prolonging sleep time. The composition for improving sleep prepared by the present invention using hydroxytyrosol and urolithin as raw materials can not only be used to improve sleep, but also has no side effects and clear functional components.

[0019] 2. The sleep-improving composition provided by the present invention can improve neurotransmitters (GABA, 5-HT, etc.) and brain oxidative stress-related indicators (SOD, MDA), thereby improving sleep.

[0020] 3. The sleep-improving composition provided by the present invention also has the following effects:

[0021] (1) Urolithin A and hydroxytyrosol have synergistic effects, and the effect of the combination on improving sleep is better than that of a single substance.

[0022] (2) Urolithin A and hydroxytyrosol are both small molecules that can cross the blood-brain barrier, reach the brain, and improve sleep.

[0023] (3) The raw materials of the composition are natural ingredients or metabolites of natural ingredients in the body, the raw materials are safe, and the effective ingredients are clear. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 is the sleep latency and sleep time of each group of mice in the present invention; wherein, Figure 1 Figure A shows the sleep latency of mice in each group; Figure 1 Figure B shows the sleep time of mice in each group; where * indicates p < 0.05, and ** indicates p < 0.01.

[0025] Figure 2 is the neurotransmitter content in the brain tissue of each group of mice in the present invention; wherein, Figure 2 Figure A shows the 5-HT content in the brain tissue of mice in each group; Figure 2 Figure B shows the GABA content in the brain tissue of mice in each group; where * indicates p < 0.05, and ** indicates p < 0.01.

[0026] Figure 3 is the SOD enzyme activity and MDA content in the brain tissue of each group of mice in the present invention; wherein, Figure 3 Figure A shows the SOD enzyme activity in the brain tissue of mice in each group; Figure 3 Figure B shows the MDA content in the brain tissue of mice in each group; where * indicates p < 0.05, and ** indicates p < 0.01. DETAILED DESCRIPTION

[0027] The present invention is described in detail below in conjunction with the accompanying drawings and specific examples, but should not be construed as limiting the present invention. Unless otherwise specified, the technical means used in the following examples are conventional means well known to those skilled in the art, and the materials, reagents, etc. used in the following examples, unless otherwise specified, can be obtained from commercial sources.

[0028] The composition of the present invention can be prepared into any conventional oral preparations, including any one of tablets, powders, pills, capsules and oral liquids, by conventional methods of pharmaceutical preparations, and can also be prepared into any conventional external preparations, including any one of nasal drops, sprays, gels, emulsions, powders and aerosols. In order to realize the above dosage forms, pharmaceutically acceptable excipients such as fillers, disintegrants, lubricants, suspending agents, adhesives, flavoring agents, preservatives, etc. need to be added when preparing these dosage forms.

[0029] Example 1

[0030] A composition for improving sleep is prepared from the following raw materials: hydroxytyrosol and urolithin A, wherein the mass ratio of hydroxytyrosol to urolithin A is 1:1.

[0031] The preparation method of the above-mentioned composition for improving sleep comprises the following steps:

[0032] Hydroxytyrosol and urolithin A are weighed respectively according to the above mass ratio, and mixed evenly to obtain the composition for improving sleep.

[0033] Example 2

[0034] A composition for improving sleep is prepared from the following raw materials: hydroxytyrosol and urolithin A, wherein the mass ratio of hydroxytyrosol to urolithin A is 1:5.

[0035] The preparation method of the above-mentioned composition for improving sleep comprises the following steps:

[0036] Hydroxytyrosol and urolithin A are weighed respectively according to the above mass ratio, and mixed evenly to obtain the composition for improving sleep.

[0037] Example 3

[0038] A composition for improving sleep is prepared from the following raw materials: hydroxytyrosol and urolithin A, wherein the mass ratio of hydroxytyrosol to urolithin A is 10:1.

[0039] The preparation method of the above-mentioned composition for improving sleep comprises the following steps:

[0040] Hydroxytyrosol and urolithin A are weighed respectively according to the above mass ratio, and mixed evenly to obtain the composition for improving sleep.

[0041] Example 4

[0042] A composition for improving sleep is prepared from the following raw materials: hydroxytyrosol and urolithin A, wherein the mass ratio of hydroxytyrosol to urolithin A is 1:3.

[0043] The preparation method of the above-mentioned composition for improving sleep comprises the following steps:

[0044] Hydroxytyrosol and urolithin A are weighed respectively according to the above mass ratio, and mixed evenly to obtain the composition for improving sleep.

[0045] Example 5

[0046] A composition for improving sleep is prepared from the following raw materials: hydroxytyrosol and urolithin A, wherein the mass ratio of hydroxytyrosol to urolithin A is 5:1.

[0047] The preparation method of the above-mentioned composition for improving sleep comprises the following steps:

[0048] Hydroxytyrosol and urolithin A are weighed respectively according to the above mass ratio, and mixed evenly to obtain the composition for improving sleep.

[0049] Example 6

[0050] A composition for improving sleep is prepared from the following raw materials: hydroxytyrosol and urolithin A, wherein the mass ratio of hydroxytyrosol to urolithin A is 1:4.

[0051] The preparation method of the above-mentioned composition for improving sleep comprises the following steps:

[0052] Hydroxytyrosol and urolithin A are weighed respectively according to the above mass ratio, and mixed evenly to obtain the composition for improving sleep.

[0053] In order to illustrate the effect of a composition for improving sleep provided by the present invention, taking Example 1 as an example, the following study was conducted.

[0054] 1. Experimental Methods

[0055] 1. Experimental Animals

[0056] SPF male ICR mice (5-6 weeks old) weighing 20±2 g were purchased from the Experimental Animal Center of Air Force Medical University.

[0057] 2. Drugs and reagents

[0058] Pentobarbital sodium was purchased from Beijing Chemical Reagent Company.

[0059] p-Chlorophenylalanine (PCPA) was purchased from Shanghai Bevanta Biotechnology Co., Ltd.

[0060] Hydroxytyrosol (mass fraction 98%) was purchased from Hubei Hengjingrui Chemical Co., Ltd.

[0061] Urolithin A was purchased from Shanghai Standard Company.

[0062] Sodium carboxymethyl cellulose (CMC-Na) was purchased from Merck.

[0063] Polysorbate 80 was purchased from Xi'an Tianzheng Pharmaceutical Excipients Co., Ltd.

[0064] Physiological saline was purchased from Shijiazhuang Siyao Co., Ltd.

[0065] Total superoxide dismutase (SOD) assay kit and malondialdehyde (MDA) assay kit were purchased from Nanjing Jiancheng Bioengineering Institute.

[0066] ELISA kits for GABA and 5-HT were purchased from Ruixin Biotechnology Co., Ltd.

[0067] 3. Grouping, modeling and drug administration

[0068] Fifty ICR mice were randomly divided into five groups after 7 days of adaptive feeding, with 10 mice in each group, namely, normal group (NC group), model group (INS group), hydroxytyrosol group (HT group), urolithin A group (UA group), and combination group of hydroxytyrosol and urolithin A (MIX group). Except for the normal group, the mice in the other groups were established with sleep disorder models.

[0069] Among them, the combination group of hydroxytyrosol and urolithin A is the combination group for improving sleep prepared in the example.

[0070] The model group is the sleep disorder model group.

[0071] The sleep disorder model of the model group was established as follows:

[0072] The mice were intraperitoneally injected with para-chlorophenylalanine (PCPA) suspension every morning for 3 consecutive days at a dose of 300 mg / kg. When the mice showed restlessness, agitation, and increased daytime activity, the model was successfully established.

[0073] After successful modeling, mice were gavaged with drugs the next morning for 7 consecutive days, with a gavage volume of 1 mL / 100 g. The normal group and the model group were gavaged with the same amount of distilled water. The dosage design and grouping are shown in Table 1.

[0074] The concentration of the p-chlorophenylalanine suspension is 0.03 g / mL, and the preparation method is to weigh 3 g of PCPA powder and dissolve it in 80 mL of normal saline, add two drops of 2% (v / v) polysorbate-80 to assist dissolution, and adjust the volume to 100 mL to obtain a 0.03 g / mL PCPA suspension. The injection dose is 1 mL / 100 g.

[0075] Table 1 Animal experiment groups and dosages

[0076]

[0077] Note: The preparation method of hydroxytyrosol solution is to weigh 0.5g hydroxytyrosol, dissolve it in distilled water and dilute it to 100mL to obtain 5mg / mL hydroxytyrosol solution. The preparation method of 0.5% (w / v) CMC-Na is to weigh 0.5g CMC-Na and dissolve it in 80mL distilled water, stir and ultrasonically dissolve it, and then dilute it to 100mL to obtain 0.5% (w / v) CMC-Na solution. The preparation method of urolithin A suspension is to weigh 0.5g urolithin A powder and dissolve it in 80mL 0.5% (w / v) CMC-Na solution, stir and ultrasonically dissolve it, and then dilute it to 100mL with 0.5% (w / v) CMC-Na to obtain 5mg / mL urolithin A suspension. The preparation method of the mixed suspension of hydroxytyrosol and urolithin A is as follows: 0.5 g of the composition for improving sleep in Example 1 is weighed and dissolved in 80 mL of 0.5% (w / v) CMC-Na solution, and then the mixture is stirred and ultrasonically dissolved, and then diluted to 100 mL with 0.5% (w / v) CMC-Na to obtain a mixed suspension of hydroxytyrosol and urolithin A.

[0078] 4. Mouse weight changes

[0079] The body weights of mice in each group were measured before modeling, after modeling and 7 days after oral administration.

[0080] 5. Statistics of autonomous activities

[0081] 40 minutes after the last administration, mice in each group were placed in the YLS-1A multifunctional mouse voluntary activity recorder chamber and allowed to adapt to the environment for 5 minutes. The voluntary activity recorder was then used to continuously record the number of voluntary activities of the mice (number of times the forelimbs were lifted upward) within 5 minutes.

[0082] Among them, the YLS-1A multifunctional mouse autonomous activity recorder was purchased from Chengdu Taimeng Technology Co., Ltd.

[0083] 6. Pentobarbital sodium sleep synergy experiment

[0084] 40 minutes after the last administration, the sodium pentobarbital sleep synergy test was performed. The specific method is as follows:

[0085] Each group of mice was intraperitoneally injected with sodium pentobarbital 0.01 mL / g, with a dose of 50 mg / kg. The sleep latency and sleep duration of the mice were observed and recorded through the righting reflex test.

[0086] Among them, the method of righting reflex experiment is as follows:

[0087] When the mouse is placed in dorsal recumbency, it can turn right immediately. If it cannot turn right for more than 1 minute, it is considered that the righting reflex has disappeared and the animal has entered sleep. The recovery of the righting reflex means that the animal has awakened. The time from intraperitoneal injection of sodium pentobarbital to the disappearance of the righting reflex is taken as the sleep latency, and the time from the disappearance to the recovery of the righting reflex is taken as the sleep time.

[0088] 7. Synergistic subthreshold dose sodium pentobarbital sleep experiment

[0089] 40 minutes after the last oral administration, all mice were intraperitoneally injected with sodium pentobarbital 0.01 mL / g (dose 28 mg / kg). If the righting reflex of the mouse disappeared for more than 1 minute, the mouse was judged to be asleep. The sleeping rate of mice in each group was calculated according to the sleeping rate calculation formula.

[0090] The calculation formula of the sleeping rate is: sleeping rate = (number of sleeping mice / total number of mice) × 100%.

[0091] 8. Effects on neurotransmitter content and oxidative stress levels in sleep-disordered mice

[0092] 60 minutes after the last oral administration, all mice were decapitated, the mouse brain tissue was dissected and taken out, and the homogenate was prepared with physiological saline at a mass ratio of 1:9, and then centrifuged at 4°C, 5000r / min for 10 minutes; the supernatant was taken after centrifugation, and the SOD enzyme activity in the brain tissue of each group of mice was detected according to the requirements of the total superoxide dismutase (SOD) assay kit, the malondialdehyde (MDA) content in the brain tissue of each group of mice was detected according to the requirements of the MDA assay kit, the GABA content in the brain tissue of each group of mice was detected according to the requirements of the GABA ELISA kit, and the 5-HT content in the brain tissue of each group of mice was detected according to the requirements of the 5-HT ELISA kit.

[0093] 2. Results

[0094] 1. Effects of hydroxytyrosol, urolithin A and their combination on sodium pentobarbital-induced sleep in sleep-disordered mice

[0095] The results of the effects of hydroxytyrosol, urolithin A and their combination on the sodium pentobarbital-induced sleep experiment in sleep-disordered mice are as follows: Figure 1 shown.

[0096] Depend on Figure 1 It can be seen that compared with the NC group, the sleep latency of the INS group was significantly prolonged and the sleep time was significantly shortened, both of which were extremely significant differences (p < 0.01), indicating that the PACA-induced sleep disorder mouse model was successful.

[0097] Compared with the INS group, the sleep latency and sleep time of the HT group, UA group, and MIX group were significantly different (p < 0.01). In terms of sleep latency, the UA group had a significant difference compared with the MIX group (p < 0.05), and the HT group had a significant difference compared with the MIX group (p < 0.01); in terms of sleep time, the MIX group had a significant difference compared with the HT group and the UA group (p < 0.01). The above results show that hydroxytyrosol, urolithin A and the combination of the two can significantly shorten the sleep latency of the model group mice and prolong their sleep time, and the combination of hydroxytyrosol and urolithin A has the best effect.

[0098] 2. Effects of hydroxytyrosol, urolithin A and their combination on neurotransmitters in brain tissue of sleep-disordered mice

[0099] The results of the effects of hydroxytyrosol, urolithin A and their combination on neurotransmitters in the brain tissue of sleep-disordered mice are as follows: Figure 2 shown.

[0100] Figure 2 In the INS group, compared with the NC group, 5-HT and GABA were significantly reduced, with extremely significant differences (p < 0.01).

[0101] Compared with the INS group, the HT group, UA group, and MIX group all increased the content of 5-HT and GABA in brain tissue, and there was a significant difference (p < 0.05); indicating that hydroxytyrosol, urolithin A, and the combination of the two can significantly increase the content of 5-HT and GABA in brain tissue of the model group mice. Compared with the HT group and UA group, the MIX group can significantly increase the content of 5-HT and GABA in brain tissue, with a very significant difference (p < 0.01), indicating that the combination of hydroxytyrosol and urolithin A is higher than a single substance in increasing the content of 5-HT and GABA in brain tissue.

[0102] 3. Effects of hydroxytyrosol, urolithin A and their combination on oxidative stress levels in brain tissue of sleep-disordered mice

[0103] The results of the effects of hydroxytyrosol, urolithin A and their combination on the oxidative stress level in the brain tissue of sleep-disordered mice are as follows Figure 3 shown.

[0104] Figure 3 In the INS group, compared with the NC group, the SOD activity was significantly decreased, and the MDA content was significantly increased, both with extremely significant differences (p < 0.01).

[0105] Compared with the INS group, the HT group, UA group, and MIX group all increased the SOD activity in brain tissue and reduced the MDA content, and there was a significant difference (p < 0.05); this shows that hydroxytyrosol, urolithin A, and the combination of the two can significantly increase the SOD activity in the brain tissue of the model group mice and reduce the MDA content. And compared with the HT group and UA group, the MIX group can significantly increase the SOD activity in the brain tissue of mice and reduce the MDA content, with a significant difference (p < 0.05), indicating that the combination of hydroxytyrosol and urolithin A is superior to a single substance in increasing the SOD activity in the brain tissue of mice and reducing the MDA content.

[0106] 4. Effects of hydroxytyrosol, urolithin A and their combination on the body weight of mice

[0107] The results of the effects of hydroxytyrosol, urolithin A and the combination of the two on the body weight of mice are shown in Table 2.

[0108] Table 2 Body weight of mice in each group

[0109] Experimental Grouping Body weight before modeling (g) Body weight after modeling (g) Body weight after treatment (g) NC 20.01±1.32 21.82±0.93 23.55±1.32# INS 20.11±1.06 20.24±1.40* 22.05±1.19 HT 19.55±1.17 19.59±1.14** 23.68±1.16# HA 20.58±1.14 20.28±1.08* 23.89±0.84## MIX 20.25±1.40 19.66±1.23** 23.99±1.01##

[0110] Note: * indicates p < 0.05 compared with the NC group, ** indicates p < 0.01 compared with the NC group; # indicates p < 0.05 compared with the INS group, ## indicates p < 0.01 compared with the INS group.

[0111] In Table 2, there was no significant difference in the body weight of mice in each group before modeling (p>0.05). After modeling, compared with the NC group, the circadian rhythm of mice in the INS group, HT group, UA group, and MIX group was disturbed or even disappeared, and the body weight was significantly reduced, and the difference was statistically significant (p<0.05). After 7 days of treatment with the corresponding drugs, the body weight of mice in each treatment group was significantly improved compared with the INS group, and the difference was statistically significant (p<0.05).

[0112] 5. Effects of hydroxytyrosol, urolithin A and their combination on the number of spontaneous activities in sleep-disordered mice

[0113] The results of the effects of hydroxytyrosol, urolithin A and the combination of the two on the number of spontaneous activities of sleep-disordered mice are shown in Table 3.

[0114] Table 3 The number of spontaneous activities of mice in each group

[0115] Experimental Grouping Number of independent activities NC 157±10 INS 204±15a HT 166±7bc HA 162±7bc MIX 146±9b

[0116] Note: a means compared with the NC group, p < 0.01; b means compared with the INS group, p < 0.01; c means compared with the MIX group,

[0117] p<0.01.

[0118] In Table 3, the number of activities in the INS group increased significantly compared with the NC group, with a very significant difference (p < 0.01).

[0119] Compared with the INS group, the HT group, UA group, and MIX group all reduced the number of mouse activities, and there was a very significant difference (p < 0.01); compared with the HT group and UA group, the MIX group could significantly reduce the number of mouse activities, and there was a very significant difference (p < 0.01). This shows that hydroxytyrosol, urolithin A, and the combination of the two can significantly reduce the number of spontaneous activities of the model group mice, and the combination group has the best effect.

[0120] 6. Effects of hydroxytyrosol, urolithin A and their combination on subthreshold dose sodium pentobarbital sleep test in sleep-disordered mice

[0121] Table 4 Sleeping rate of mice in each group

[0122] Experimental Grouping Sleep rate NC 10％ INS 0％ HT 40％ HA 50％ MIX 70％

[0123] The effects of hydroxytyrosol, urolithin A and the combination of the two on the subthreshold dose of sodium pentobarbital sleep test in sleep-disordered mice are shown in Table 4. The sleep rate in the model group was the lowest, and the sleep rates in each test group were improved.

[0124] From the above experimental results, it can be seen that hydroxytyrosol, urolithin A and the combination of the two can shorten the sleep latency of sleep-disordered mice and prolong their sleep time. That is, the sleep-improving composition provided by the present invention can be used to improve sleep.

[0125] 5-HT can inhibit the excitability of the brain, reduce anxiety and tension, and thus help to fall asleep. It is also involved in regulating the sleep cycle, including sleep duration and sleep quality. GABA is the most important inhibitory neurotransmitter in the brain, which can promote the transformation of wakefulness to sleep. At present, the research on improving insomnia is mainly focused on the research of neurotransmitters, hormones, etc. Positive regulation of the levels of neurotransmitters 5-HT and GABA can positively regulate sleep. However, insomnia can cause oxidative stress, and whether improving oxidative stress can improve insomnia has not yet been clarified. The present invention proves that urolithin A, hydroxytyrosol and its composition can increase the content of 5-HT and GABA in the brain, improve oxidative stress and improve sleep.

[0126] Urolithin A and hydroxytyrosol both have antioxidant and anti-inflammatory effects, and it can be estimated that the combination of the two can also reduce the level of oxidative stress. However, at the same dose, the oxidative stress level of the combination group was significantly lower than that of the single substance group, indicating that urolithin A and hydroxytyrosol have a synergistic effect in reducing the level of oxidative stress. Urolithin A and hydroxytyrosol can both improve brain neurotransmitters, but the effect of their individual raw materials on improving neurotransmitters is less than that of their combination (the dose of the combination is consistent with the dose of the individual raw materials). This shows that hydroxytyrosol enhances the effect of urolithin A on improving the content of 5-HT and GABA in brain tissue. The reasons may be: ① Hydroxytyrosol further enhances the antioxidant capacity of UA, and the strong antioxidant capacity avoids damage to neuronal cells (the transmission of sleep neural signals requires the participation of neurons, and neurons can secrete neurotransmitters that regulate sleep), thereby improving sleep; ② Studies have shown that hydroxytyrosol can regulate intestinal microbiota and improve intestinal health. The intestine can produce a variety of neurotransmitters such as 5-HT and GABA, which affect the brain and sleep through the brain-gut axis. Hydroxytyrosol further increases the production of 5-HT and GABA neurotransmitters through its antioxidant properties or regulation of the intestine. Therefore, hydroxytyrosol can improve the antioxidant properties and neurotransmitter levels of urolithin A, synergistically improving sleep.

[0127] The present invention uses PACA to prepare a mouse sleep disorder model, and detects the body weight, the number of autonomous activities, the neurotransmitter in brain tissue and the oxidative stress level after oral administration; performs a sodium pentobarbital sleep synergistic experiment to detect the sleep latency and sleep time of the mouse; and performs a subthreshold dose sodium pentobarbital sleep experiment to detect the sleep onset rate of the mouse; it is shown that hydroxytyrosol, urolithin A and the combination thereof can improve the neurotransmitter and oxidative stress level in the mouse brain tissue, thereby improving the sleep latency and sleep time of the mouse, and the two substances synergistically enhance the effect, and the effect of the combination is better than that of any one component.

[0128] The raw materials used in the present invention are all natural ingredients or metabolites of natural ingredients in the body, the raw materials are safe, and the functional components are clear and effective. The composition for improving sleep provided by the present invention has the effects of regulating neurotransmitters, reducing oxidative stress, improving sleep, shortening sleep latency, and prolonging sleep time. The composition for improving sleep prepared by the present invention using hydroxytyrosol and urolithin A as raw materials can not only be used to improve sleep, but also has no side effects and clear functional components.

[0129] The present invention also provides a preparation for improving sleep, which is an oral preparation or an external preparation. The oral preparation is any one of tablets, powders, pills, capsules and oral liquids. The external preparation is any one of nasal drops, sprays, gels, emulsions, powders and aerosols.

[0130] The above-mentioned drug is prepared by adding different adjuvants or carriers to the above-mentioned composition for improving sleep according to different dosage forms, and processing it into a dosage form that can be used by animals through the preparation process of the corresponding dosage form. The drug adjuvant includes any one or more of maltodextrin, sodium carboxymethyl cellulose, lactose, silicon dioxide, magnesium stearate, carbomer, chitosan, β-cyclodextrin and phospholipids; the carrier includes any one or more of liposomes, cyclodextrin inclusion compounds, microspheres and nanoparticles.

[0131] The effect verification of the above-mentioned preparation for improving sleep was carried out in the same experiment as the verification experiment of the composition for improving sleep. The results showed that the preparation for improving sleep has the same effect of improving sleep as the composition for improving sleep.

[0132] It should be noted that when the present invention involves a numerical range, it should be understood that the two endpoints of each numerical range and any numerical value between the two endpoints can be selected. In order to avoid redundancy, the present invention describes a preferred embodiment.

[0133] Although preferred embodiments of the present invention have been described, those skilled in the art may make additional changes and modifications to these embodiments once they understand the basic inventive concepts, and all such changes and modifications fall within the scope of the present invention.

Claims

1. A composition for improving sleep, characterized in that: The invention is prepared from the following raw materials: hydroxytyrosol and urolithin A; the mass ratio of the hydroxytyrosol to the urolithin A is 1:0.1-5.

2. A composition for improving sleep according to claim 1, characterized in that: The invention is prepared from the following raw materials: hydroxytyrosol and urolithin A; the mass ratio of the hydroxytyrosol to the urolithin A is 1:0.2-3.

3. A composition for improving sleep according to claim 2, characterized in that: The invention is prepared from the following raw materials: hydroxytyrosol and urolithin A; the mass ratio of the hydroxytyrosol to the urolithin A is 1:

1.

4. The method for preparing the composition for improving sleep according to any one of claims 1 to 3, characterized in that: The method comprises the following steps: hydroxytyrosol and urolithin A are weighed respectively according to a mass ratio, and mixed evenly to obtain the composition for improving sleep.

5. A preparation for improving sleep, characterized in that: The sleep improving composition comprises the sleep improving composition according to claim 1, and a pharmaceutically acceptable auxiliary material or carrier.

6. The preparation for improving sleep according to claim 5, characterized in that: The preparation is an internal preparation or an external preparation.

7. The preparation for improving sleep according to claim 6, characterized in that: The oral preparation is any one of tablets, powders, pills, capsules and oral liquids.

8. The preparation for improving sleep according to claim 6, characterized in that: The external preparation is any one of nasal drops, sprays, gels, emulsions, powders and aerosols.

9. The preparation for improving sleep according to claim 5, characterized in that: The auxiliary materials include any one or more of maltodextrin, sodium carboxymethyl cellulose, lactose, silicon dioxide, magnesium stearate, carbomer, chitosan, beta-cyclodextrin and phospholipids.

10. The preparation for improving sleep according to claim 5, characterized in that: The carrier includes any one or more of liposomes, cyclodextrin inclusion compounds, microspheres and nanoparticles.

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