Composition helpful for improving sleep and preparation method thereof

Abstract

The invention relates to the technical field of pharmacy, and particularly provides a composition helpful for improving sleep and a preparation method thereof.The composition is prepared from spina date seeds, schisandra chinensis, fructus chebulae immaturus and melatonin, the sleep effect is improved in a Chinese and western combined mode, and compared with other products for improving sleep, the composition has the advantages that the composition is simple in preparation process and convenient to use. Natural products and melatonin are combined, so that the traditional Chinese medicine composition has the effects of strengthening body resistance to eliminate pathogenic factors, conditioning body functions and relieving side effects of western medicines while taking effect quickly. The combination of natural products used in the combination does not need chemical synthesis and structural modification, the process is simple, and industrial large-scale production is easy to realize.

Description

Technical Field

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

[0002] Insomnia is a subjective experience that patients cannot get a normal amount of sleep, have a shallow sleep depth, and cannot meet daytime social function activities. Its main manifestations are difficulty falling asleep, waking up early, waking up easily at night, difficulty falling asleep after waking up, or waking up from time to time. In severe cases, patients may not sleep all night. Although insomnia is not a fatal disease, it can lead to decreased resistance, body dysfunction, and a series of gastrointestinal, cardiovascular and cerebrovascular diseases. It is related to the rapid development of modern society, the accelerated pace of life, work pressure, living environment, diet and daily life, and the incidence rate is increasing.

[0003] At present, Western medicine drugs for the treatment of insomnia can be divided into four categories according to their pharmacodynamic effects: benzodiazepine receptor agonists, melatonin receptor agonists, histamine receptor antagonists, and orexin receptor antagonists. However, long-term oral Western medicine treatment is prone to adverse reactions such as withdrawal symptoms, rebound insomnia, and addiction. At present, Western medicine treatment for insomnia has disadvantages such as addiction, dependence, withdrawal reaction, and patients' difficulty in accepting it. The treatment of insomnia by traditional Chinese medicine has the following characteristics: (1) Oral administration of Chinese medicine is the main method of TCM treatment for insomnia. (2) Compared with decoction, Chinese patent medicine has the advantages of simplicity and convenience. It is increasingly widely used in clinical practice and is also accepted by more and more patients. (3) TCM acupuncture: The treatment of insomnia has a long history. It treats insomnia by dredging meridians, strengthening the body and eliminating evil, and harmonizing yin and yang, with significant clinical effects. However, the effect of TCM treatment for insomnia is relatively slow.

[0004] How to provide a combination of Chinese and Western medicine to improve sleep effect has become one of the technical problems that need to be solved urgently. Summary of the invention

[0005] In view of this, the present invention proposes a composition that helps improve sleep and a preparation method thereof, aiming to obtain a composition that improves sleep by combining traditional Chinese medicine with needs.

[0006] The technical solution of the present invention is achieved as follows: the present invention provides a composition that helps improve sleep, and the raw materials of the composition include: spiny jujube seeds, schisandra chinensis, citrus aurantium and melatonin.

[0007] Among them, sour jujube kernel is the dried mature seed of the Rhamnaceae plant sour jujube Ziziphus jujuba Mill.var.spinosa (Bunge) Huex HFChou, which was first recorded in "Shennong's Herbal Classic". It tastes sweet and is neutral in nature. It has the effects of calming the mind, promoting salivation, and nourishing the liver. Modern research shows that the active ingredients in sour jujube kernel are mainly terpenes, flavonoids, saponins, and alkaloids. The extraction of its active ingredients has been widely studied.

[0008] Schisandra chinensis (Turcz.) Baill. is the dried mature fruit of Schisandra chinensis, a plant of the Magnoliaceae family. It is divided into northern and southern varieties, with the northern variety being preferred. Northern Schisandra chinensis is sour and sweet, warm in nature, and enters the lung, heart, and kidney meridians. It has the effects of invigorating qi and promoting body fluid, astringing and consolidating, and tonifying the kidney and calming the mind. Modern pharmacological studies have shown that it has multiple effects such as antibacterial, anti-inflammatory, liver-protecting, and anti-tumor.

[0009] Xiqingguo is the dried young fruit of Terminalia chebula Retz., a plant of the Combretaceae family. It tastes bitter, sour, astringent, and flat. It enters the lung and large intestine meridians. It is mainly used to clear away heat, promote body fluid, and detoxify. It is used for diphtheria caused by yin deficiency.

[0010] Melatonin is one of the hormones secreted by the pineal gland in the brain. Melatonin belongs to the indole heterocyclic compound, and its chemical name is N-acetyl-5-methoxytryptamine, also known as pinealogen, melatonin, and melatonin. It plays an important role in regulating circadian rhythm and sleep-wakefulness.

[0011] In a second aspect, the present invention further provides a method for preparing the above-mentioned composition for improving sleep, comprising the following steps:

[0012] Step 1: Mix and crush the spiny jujube seeds, schisandra chinensis, and citronella, add an ethanol solution, perform ultrasonic extraction, and filter to obtain an extract;

[0013] Step 2, concentrating and drying the extract, re-dissolving, purifying and drying to obtain a Chinese medicine extract;

[0014] Step 3: After mixing the Chinese herbal extract with melatonin, a composition that helps improve sleep is obtained.

[0015] In some embodiments, the mass percentage of the spinulosae seeds is 20-50%, the mass percentage of the schisandrae chinensis is 20-50%, and the mass percentage of the citronella is 10-30%.

[0016] In some embodiments, in step 1, the mass ratio of the ethanol solution to the crushed medicinal material is (10-15):1, and the concentration of the ethanol solution is 20-50% vol.

[0017] In some embodiments, in step 1, the ultrasonic power is 450-500 W, the ultrasonic extraction time is 60-90 min, and the number of extractions is 2-3 times.

[0018] In some embodiments, in step 2, the temperature for concentrating and drying is 40-45°C.

[0019] In some embodiments, in step 2, the purification and drying method includes: dissolving the concentrated and dried extract with ethanol, adsorbing it on an AB-8 resin column for 2 hours, and then eluting it with 5 times the column volume of purified water for 2 hours, eluting it with 5 times the column volume of a low-concentration ethanol solution for 2 hours, and eluting it with a high-concentration ethanol solution for 2 hours, collecting the ultrapure water eluate and the low-concentration ethanol eluate, combining them, concentrating and drying them under reduced pressure, and obtaining a traditional Chinese medicine extract.

[0020] In some embodiments, in step three, the mass ratio of the Chinese herbal extract to melatonin is 1:(1-4).

[0021] In some embodiments, the concentration of the low concentration ethanol solution is 20-50% vol.

[0022] In some embodiments, the high concentration ethanol solution has a concentration of 95% vol.

[0023] In order to verify the effect of the above composition on improving sleep, the applicant used zebrafish as the experimental subject. The physiology and behavior of zebrafish have obvious circadian rhythms. Their behavior is characterized by being active during the day and resting at night and is easily affected by external stimuli such as light, temperature and sound. The level of melatonin secreted in their body also shows a certain circadian rhythm. Laboratories have proven the existence of circadian rhythms in the movement of zebrafish larvae. Zebrafish, like humans, are diurnal animals, unlike mice, which are nocturnal animals. This also shows that using zebrafish to study human circadian rhythms is more representative than using mice. Zebrafish larvae show obvious rhythmic behavior under normal light and dark conditions and under continuous low-intensity light conditions.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] The present invention adopts a combination of Chinese and Western medicine to improve the sleep effect. Compared with other sleep-improving products, the combination of natural products and melatonin used in this combination can not only achieve a fast onset of action, but also strengthen the body's health and eliminate evil, regulate body functions, and reduce the side effects of Western medicine. Moreover, the combination of natural products used in this combination does not require chemical synthesis and structural modification, and the process is simple and easy to industrialize and scale up. DETAILED DESCRIPTION

[0026] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0027] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the embodiments of the present invention belong. If the definitions set forth in this section are contrary to or otherwise inconsistent with the definitions set forth in the patents, patent applications, published patent applications, and other publications incorporated herein by reference, the definitions listed in this section take precedence over the definitions incorporated herein by reference.

[0028] Unless otherwise specified, the methods used in the following examples are all conventional methods. Unless otherwise specified, the materials, reagents and instruments used are all conventional materials, reagents and instruments in the art, and can be obtained by those skilled in the art through commercial channels.

[0029] When an amount, concentration or other value or parameter is expressed as a range, a preferred range or a range defined by a series of upper preferred values ​​and lower preferred values, this should be understood as specifically disclosing all ranges formed by any pairing of any range upper limit or preferred value with any range lower limit or preferred value, regardless of whether the range is disclosed separately. For example, when a range of "1 to 5" is disclosed, the described range should be interpreted as including the range "1 to 4", "1 to 3", "1 to 2", "1 to 2 and 4 to 5", "1 to 3 and 5", etc. When a numerical range is described herein, unless otherwise stated, the range is intended to include its end values ​​and all integers and fractions within the range. In the present specification and claims, range definitions can be combined and / or interchanged, and if not otherwise stated, these ranges include all subranges contained therein.

[0030] The samples obtained from the following examples and comparative examples were used for zebrafish experiments. Zebrafish were raised in fish farming water at 28°C (water quality: 200 mg of instant sea salt was added to every 1L of reverse osmosis water, the conductivity was 450-550 μS / cm; pH was 6.5-8.5; hardness was 50-100 mg / L CaCO3), and were bred and provided by this laboratory. Wild-type AB strain zebrafish were bred in natural pairs. The experimental fish were zebrafish 5 days after fertilization (5dpf) for the evaluation test of improving sleep efficacy. From the beginning of fertilization, the zebrafish controlled the light cycle to 14±0.5 hours of light: 10±0.5 hours of darkness. 5dpf wild-type AB strain healthy, deformity-free, uniform zebrafish were randomly selected in a 6-well plate. After the sample was dissolved, high, medium and low dose groups were set according to the maximum tolerated dose. The positive control was Tianmeijian brand colostrum and jujube seed ganoderma lucidum capsule with a concentration of 1000 μg / mL. A normal control group and a model control group were also set up, with a capacity of 3 mL per well. After 24 hours of treatment at 28°C, all experimental groups except the normal control group were treated with pentylenetetrazol (PTZ) for 1 hour to establish a zebrafish insomnia model. Ten zebrafish were randomly selected from each experimental group and transferred to a 96-well plate, 1 tail / 200 μL / well.

[0031] Example 1

[0032] This embodiment provides a method for preparing a composition that helps improve sleep and its application in a zebrafish sleep experiment.

[0033] (1) Extraction of Chinese medicinal materials: 40% of Ziziphus jujuba seeds, 40% of Schisandra chinensis, and 20% of Cibotium barometz were weighed and crushed according to the mass percentage, and 10 times of 20% ethanol solution was added. Ultrasonic extraction was performed twice at 450W, each time for 80 minutes. After the extraction, the mixture was cooled to room temperature, filtered, and the filtrate was combined;

[0034] (2) Concentration and drying: The filtered extract is concentrated and dried under reduced pressure and vacuum at 45° C. to obtain a dry extract;

[0035] (3) Preparation and purification: The above dry extract was dissolved in ethanol solution, adsorbed on AB-8 resin column for 2 hours, eluted with 5 times ultrapure water for 2 hours, eluted with 5 times 20% ethanol solution for 2 hours, eluted with 95% ethanol solution for 2 hours, and the eluate was collected;

[0036] (5) Concentration and drying: The collected low-concentration ethanol and ultrapure water eluents are concentrated and dried under reduced pressure and vacuum to obtain the Chinese medicine extract.

[0037] (5) Preparation of the composition: The above-mentioned Chinese herbal extract and melatonin are mixed in a mass ratio of 1:4 to obtain a composition.

[0038] The zebrafish experiment was conducted on the samples prepared above. Zebrafish were raised in fish farming water at 28°C (water quality: 200 mg of instant sea salt was added to each 1L of reverse osmosis water, the conductivity was 450-550 μS / cm; pH was 6.5-8.5; hardness was 50-100 mg / LCaCO3), and were bred and provided by this laboratory. Wild-type AB strain zebrafish were bred in natural pairs. The experimental fish were zebrafish 5 days after fertilization (5dpf) for the evaluation of sleep improvement efficacy. From the beginning of fertilization, the zebrafish controlled the light cycle to 14±0.5 hours of light: 10±0.5 hours of darkness. 5dpf wild-type AB strain healthy, non-deformed, uniform zebrafish were randomly selected in a 6-well plate. The sample was dissolved in purified water and added to the experimental system at a final concentration of 1000 μg / mL of the composition. The positive control was Tianmeijian brand colostrum and jujube seed ganoderma lucidum capsule at a concentration of 1000 μg / mL. A normal control group and a model control group were set up at the same time, and the capacity of each well was 3 mL. After 24 hours of treatment at 28°C, except for the normal control group, all other experimental groups were treated with pentylenetetrazol (PTZ) for 1 hour to establish a zebrafish insomnia model. Ten zebrafish were randomly selected from each experimental group and transferred to a 96-well plate, 1 tail / 200 μL / well.

[0039] The final awakening activity data of the normal group, model group, positive group and experimental group are as follows:

[0040] Group Awakening activity mm Normal group <![CDATA[7699.505±179.971 *** ]]> Model Group 20806.991±824.511 Positive group <![CDATA[6258.445±202.676 *** ]]> Experimental Group <![CDATA[6870.281±324.229 *** ]]>

[0041] Compared with the model group, the awakening activity of the composition prepared in the above embodiment decreased by 67.0%, and the positive group decreased by 69.9%. It can be seen that the Chinese and Western medicine combination composition of the present application has a similar sleep-aiding effect as the commercially available sleep-improving products. Compared with the normal group, the awakening activity is lower, indicating a significant improvement effect.

[0042] Example 2

[0043] This embodiment provides a method for preparing a composition that helps improve sleep and its application in a zebrafish sleep experiment.

[0044] (1) Extraction of Chinese medicinal materials: 50% of Ziziphus jujuba seeds, 20% of Schisandra chinensis, and 30% of Cibotium barometz were weighed and crushed according to the mass percentage, and 10 times of 20% ethanol solution was added. Ultrasonic extraction was performed at 500W for 3 times, each time for 60 minutes. After the extraction, the mixture was cooled to room temperature, filtered, and the filtrate was combined;

[0045] (2) Concentration and drying: The filtered extract is concentrated and dried under reduced pressure and vacuum at 45° C. to obtain a dry extract;

[0046] (3) Preparation and purification: The above dry extract was dissolved in ethanol solution, adsorbed on AB-8 resin column for 2 hours, eluted with 5 times ultrapure water for 2 hours, eluted with 5 times 20% ethanol solution for 2 hours, eluted with 95% ethanol solution for 2 hours, and the eluate was collected;

[0047] (5) Concentration and drying: The collected low-concentration ethanol and ultrapure water eluents are concentrated and dried under reduced pressure and vacuum to obtain the Chinese medicine extract.

[0048] (5) Preparation of the composition: The above-mentioned Chinese herbal extract and melatonin are mixed in a mass ratio of 1:2.5 to obtain a composition.

[0049] The zebrafish experiment was conducted on the samples prepared above. Zebrafish were raised in fish farming water at 28°C (water quality: 200 mg of instant sea salt was added to each 1L of reverse osmosis water, the conductivity was 450-550 μS / cm; pH was 6.5-8.5; hardness was 50-100 mg / LCaCO3), and were bred and provided by this laboratory. Wild-type AB strain zebrafish were bred in natural pairs. The experimental fish were zebrafish 5 days after fertilization (5dpf) for the evaluation of sleep improvement efficacy. From the beginning of fertilization, the zebrafish controlled the light cycle to 14±0.5 hours of light: 10±0.5 hours of darkness. 5dpf wild-type AB strain healthy, non-deformed, uniform zebrafish were randomly selected in a 6-well plate. The sample was dissolved in purified water and added to the experimental system at a final concentration of 1000 μg / mL of the composition, and the positive control was 1000 μg / mL of Tianmeijian brand bovine colostrum and jujube seed ganoderma capsule. A normal control group and a model control group were set up at the same time, and the capacity of each well was 3 mL. After 24 hours of treatment at 28°C, except for the normal control group, all other experimental groups were treated with pentylenetetrazol (PTZ) for 1 hour to establish a zebrafish insomnia model. Ten zebrafish were randomly selected from each experimental group and transferred to a 96-well plate, 1 tail / 200 μL / well.

[0050] The final awakening activity data of the normal group, model group, positive group and experimental group are as follows:

[0051] Group Awakening activity mm Normal group <![CDATA[7699.505±179.971 *** ]]> Model Group 20806.991±824.511 Positive group <![CDATA[6258.445±202.676 *** ]]> Experimental Group <![CDATA[9876.375±465.129 *** ]]>

[0052] Compared with the model group, the composition prepared in the above embodiment decreased the amount of awakening activity by 52.5% and the positive group by 69.9%. It can be seen that the sleep-aiding effect of this embodiment is not as good as the composition formula of Example 1, but it still has a significant sleep-aiding effect. Compared with the normal group, the amount of awakening activity is higher, indicating that it has an improvement effect but the effect is not as good as that of Example 1 and the positive group.

[0053] Example 3

[0054] This embodiment provides a method for preparing a composition that helps improve sleep and its application in a zebrafish sleep experiment.

[0055] (1) Extraction of Chinese medicinal materials: 40% of Ziziphus jujuba seeds, 50% of Schisandra chinensis, and 10% of Cibotium barometz were weighed and crushed according to the mass percentage, and 10 times of 50% ethanol solution was added. Ultrasonic extraction was performed twice at 450W, each time for 90 minutes. After the extraction, the mixture was cooled to room temperature, filtered, and the filtrate was combined;

[0056] (2) Concentration and drying: The filtered extract is concentrated and dried under reduced pressure and vacuum at 50° C. to obtain a dry extract;

[0057] (3) Preparation and purification: The above dry extract was dissolved in ethanol solution, adsorbed on AB-8 resin column for 2 hours, eluted with 5 times ultrapure water for 2 hours, eluted with 5 times 50% ethanol solution for 2 hours, eluted with 95% ethanol solution for 2 hours, and the eluate was collected;

[0058] (5) Concentration and drying: The collected low-concentration ethanol and ultrapure water eluents are concentrated and dried under reduced pressure and vacuum to obtain the Chinese medicine extract.

[0059] (5) Preparation of the composition: The above-mentioned Chinese herbal extract and melatonin are mixed in a mass ratio of 1:1 to obtain a composition.

[0060] The zebrafish experiment was conducted on the samples prepared above. Zebrafish were raised in fish farming water at 28°C (water quality: 200 mg of instant sea salt was added to each 1L of reverse osmosis water, the conductivity was 450-550 μS / cm; pH was 6.5-8.5; hardness was 50-100 mg / LCaCO3), and were bred and provided by this laboratory. Wild-type AB strain zebrafish were bred in natural pairs. The experimental fish were zebrafish 5 days after fertilization (5dpf) for the evaluation of sleep improvement efficacy. From the beginning of fertilization, the zebrafish controlled the light cycle to 14±0.5 hours of light: 10±0.5 hours of darkness. 5dpf wild-type AB strain healthy, non-deformed, uniform zebrafish were randomly selected in a 6-well plate. The sample was dissolved in purified water and added to the experimental system at a final concentration of 1000 μg / mL of the composition, and the positive control was 1000 μg / mL of Tianmeijian brand bovine colostrum and jujube seed ganoderma capsule. A normal control group and a model control group were set up at the same time, and the capacity of each well was 3 mL. After 24 hours of treatment at 28°C, except for the normal control group, all other experimental groups were treated with pentylenetetrazol (PTZ) for 1 hour to establish a zebrafish insomnia model. Ten zebrafish were randomly selected from each experimental group and transferred to a 96-well plate, 1 tail / 200 μL / well.

[0061] The final awakening activity data of the normal group, model group, positive group and experimental group are as follows:

[0062] Group Awakening activity mm Normal group <![CDATA[7699.505±179.971 *** ]]> Model Group 20806.991±824.511 Positive group <![CDATA[6258.445±202.676 *** ]]> Experimental Group <![CDATA[12147.14±555.451 *** ]]>

[0063] Compared with the model group, the composition prepared in the above embodiment decreased the amount of awakening activity by 41.6% and the positive group by 69.9%. It can be seen that the sleep-aiding effect of this embodiment is not as good as the composition formula of Examples 1 and 2, but it still has a significant sleep-aiding effect. Compared with the normal group, the amount of awakening activity is higher, indicating that it has an improvement effect but the effect is not as good as that of Example 1, Example 2 and the positive group.

[0064] Comparative Example 1

[0065] This comparative example provides a preparation method of a composition and application thereof in a zebrafish sleep experiment.

[0066] (1) Extraction of Chinese medicinal materials: Weigh 50% of Ziziphus jujuba seeds and 50% of Schisandra chinensis chinensis by mass, grind them, add 10 times of 20% ethanol solution, and extract them twice at 450W ultrasonic, each time for 80 minutes. After the extraction, cool to room temperature, filter, and combine the filtrate;

[0067] (2) Concentration and drying: The filtered extract is concentrated and dried under reduced pressure and vacuum at 45° C. to obtain a dry extract;

[0068] (3) Preparation and purification: The above dry extract was dissolved in ethanol solution, adsorbed on AB-8 resin column for 2 hours, eluted with 5 times ultrapure water for 2 hours, eluted with 5 times 20% ethanol solution for 2 hours, eluted with 95% ethanol solution for 2 hours, and the eluate was collected;

[0069] (5) Concentration and drying: The collected low-concentration ethanol and ultrapure water eluents are concentrated and dried under reduced pressure and vacuum to obtain the Chinese medicine extract.

[0070] (5) Preparation of the composition: The above-mentioned Chinese herbal extract and melatonin are mixed in a mass ratio of 1:4 to obtain a composition.

[0071] The zebrafish experiment was conducted on the samples prepared above. Zebrafish were raised in fish farming water at 28°C (water quality: 200 mg of instant sea salt was added to each 1L of reverse osmosis water, the conductivity was 450-550 μS / cm; pH was 6.5-8.5; hardness was 50-100 mg / LCaCO3), and were bred and provided by this laboratory. Wild-type AB strain zebrafish were bred in natural pairs. The experimental fish were zebrafish 5 days after fertilization (5dpf) for the evaluation of sleep improvement efficacy. From the beginning of fertilization, the zebrafish controlled the light cycle to 14±0.5 hours of light: 10±0.5 hours of darkness. 5dpf wild-type AB strain healthy, non-deformed, uniform zebrafish were randomly selected in a 6-well plate. The sample was dissolved in purified water and added to the experimental system at a final concentration of 1000 μg / mL of the composition, and the positive control was 1000 μg / mL of Tianmeijian brand bovine colostrum and jujube seed ganoderma capsule. A normal control group and a model control group were set up at the same time, and the capacity of each well was 3 mL. After 24 hours of treatment at 28°C, except for the normal control group, all other experimental groups were treated with pentylenetetrazol (PTZ) for 1 hour to establish a zebrafish insomnia model. Ten zebrafish were randomly selected from each experimental group and transferred to a 96-well plate, 1 tail / 200 μL / well.

[0072] The final awakening activity data of the normal group, model group, positive group and experimental group are as follows:

[0073]

[0074] The composition prepared in this comparative example omitted the fruit of the Chinese prickly ash. Compared with the model group, the amount of awakening activity decreased by 30.7%, and the positive group decreased by 62.8%. It can be seen that the sleep-aiding effect of this comparative example is much lower than the composition formula of the embodiment, and the sleep-aiding effect is not good. Compared with comparative example 5, it can be seen that the improvement of the sleep-aiding effect is relatively limited.

[0075] Comparative Example 2

[0076] This comparative example provides a preparation method of a composition and application thereof in a zebrafish sleep experiment.

[0077] (1) Extraction of Chinese medicinal materials: 67% of Ziziphus jujuba seeds and 33% of Cibotium barometz were weighed and crushed according to the mass percentage, and 10 times of 20% ethanol solution was added. Ultrasonic extraction was performed twice at 450W, each time for 80 minutes. After the extraction, the mixture was cooled to room temperature, filtered, and the filtrate was combined;

[0078] (2) Concentration and drying: The filtered extract is concentrated and dried under reduced pressure and vacuum at 45° C. to obtain a dry extract;

[0079] (3) Preparation and purification: The above dry extract was dissolved in ethanol solution, adsorbed on AB-8 resin column for 2 hours, eluted with 5 times ultrapure water for 2 hours, eluted with 5 times 20% ethanol solution for 2 hours, eluted with 95% ethanol solution for 2 hours, and the eluate was collected;

[0080] (5) Concentration and drying: The collected low-concentration ethanol and ultrapure water eluents are concentrated and dried under reduced pressure and vacuum to obtain the Chinese medicine extract.

[0081] (5) Preparation of the composition: The above-mentioned Chinese herbal extract and melatonin are mixed in a mass ratio of 1:4 to obtain a composition.

[0082] The zebrafish experiment was conducted on the samples prepared above. Zebrafish were raised in fish farming water at 28°C (water quality: 200 mg of instant sea salt was added to each 1L of reverse osmosis water, the conductivity was 450-550 μS / cm; pH was 6.5-8.5; hardness was 50-100 mg / LCaCO3), and were bred and provided by this laboratory. Wild-type AB strain zebrafish were bred in natural pairs. The experimental fish were zebrafish 5 days after fertilization (5dpf) for the evaluation of sleep improvement efficacy. From the beginning of fertilization, the zebrafish controlled the light cycle to 14±0.5 hours of light: 10±0.5 hours of darkness. 5dpf wild-type AB strain healthy, non-deformed, uniform zebrafish were randomly selected in a 6-well plate. The sample was dissolved in purified water and added to the experimental system at a final concentration of 1000 μg / mL of the composition, and the positive control was 1000 μg / mL of Tianmeijian brand bovine colostrum and jujube seed ganoderma capsule. A normal control group and a model control group were set up at the same time, and the capacity of each well was 3 mL. After 24 hours of treatment at 28°C, except for the normal control group, all other experimental groups were treated with pentylenetetrazol (PTZ) for 1 hour to establish a zebrafish insomnia model. Ten zebrafish were randomly selected from each experimental group and transferred to a 96-well plate, 1 tail / 200 μL / well.

[0083] The final awakening activity data of the normal group, model group, positive group and experimental group are as follows:

[0084] Group Awakening activity mm Normal group <![CDATA[8130.976±534.005 *** ]]> Model Group 23852.772±545.439 Positive group <![CDATA[8863.737±463.315 *** ]]> Experimental Group <![CDATA[17298.897±484.330 * ]]>

[0085] The composition prepared in this comparative example omitted Schisandra chinensis, and compared with the model group, the amount of awakening activity decreased by 27.5%, and the positive group decreased by 62.8%. It can be seen that the sleep-aiding effect of this comparative example is much lower than the composition formula of the embodiment, and the sleep-aiding effect is not good. Compared with comparative example 5, it can be seen that the sleep-aiding effect is limited and is lower than comparative example 1.

[0086] Comparative Example 3

[0087] This embodiment provides a method for preparing a composition that helps improve sleep and its application in a zebrafish sleep experiment.

[0088] (1) Extraction of Chinese medicinal materials: 67% of Schisandra chinensis and 33% of Cibotium barometz were weighed and crushed according to the mass percentage, and 10 times of 20% ethanol solution was added. Ultrasonic extraction was performed twice at 450W, each time for 80 minutes. After the extraction, the mixture was cooled to room temperature, filtered, and the filtrate was combined;

[0089] (2) Concentration and drying: The filtered extract is concentrated and dried under reduced pressure and vacuum at 45° C. to obtain a dry extract;

[0090] (3) Preparation and purification: The above dry extract was dissolved in ethanol solution, adsorbed on AB-8 resin column for 2 hours, eluted with 5 times ultrapure water for 2 hours, eluted with 5 times 20% ethanol solution for 2 hours, eluted with 95% ethanol solution for 2 hours, and the eluate was collected;

[0091] (5) Concentration and drying: The collected low-concentration ethanol and ultrapure water eluents are concentrated and dried under reduced pressure and vacuum to obtain the Chinese medicine extract.

[0092] (5) Preparation of the composition: The above-mentioned Chinese herbal extract and melatonin are mixed in a mass ratio of 1:4 to obtain a composition.

[0093] The zebrafish experiment was conducted on the samples prepared above. Zebrafish were raised in fish farming water at 28°C (water quality: 200 mg of instant sea salt was added to each 1L of reverse osmosis water, the conductivity was 450-550 μS / cm; pH was 6.5-8.5; hardness was 50-100 mg / LCaCO3), and were bred and provided by this laboratory. Wild-type AB strain zebrafish were bred in natural pairs. The experimental fish were zebrafish 5 days after fertilization (5dpf) for the evaluation of sleep improvement efficacy. From the beginning of fertilization, the zebrafish controlled the light cycle to 14±0.5 hours of light: 10±0.5 hours of darkness. 5dpf wild-type AB strain healthy, non-deformed, uniform zebrafish were randomly selected in a 6-well plate. The sample was dissolved in purified water and added to the experimental system at a final concentration of 1000 μg / mL of the composition, and the positive control was 1000 μg / mL of Tianmeijian brand bovine colostrum and jujube seed ganoderma capsule. A normal control group and a model control group were set up at the same time, and the capacity of each well was 3 mL. After 24 hours of treatment at 28°C, except for the normal control group, all other experimental groups were treated with pentylenetetrazol (PTZ) for 1 hour to establish a zebrafish insomnia model. Ten zebrafish were randomly selected from each experimental group and transferred to a 96-well plate, 1 tail / 200 μL / well.

[0094] The final awakening activity data of the normal group, model group, positive group and experimental group are as follows:

[0095] Group Awakening activity mm Normal group <![CDATA[8130.976±534.005 *** ]]> Model Group 23852.772±545.439 Positive group <![CDATA[8863.737±463.315 *** ]]> Experimental Group <![CDATA[19779.040±637.764 * ]]>

[0096] The composition prepared in this comparative example omitted the Chinese date kernel, and compared with the model group, the amount of awakening activity decreased by 17.1%, and the positive group decreased by 62.8%. It can be seen that the sleep-aiding effect of this comparative example is much lower than the composition formula of the embodiment, and the sleep-aiding effect is not good. Compared with comparative example 5, it can be seen that the sleep-aiding effect is worse, indicating that the composition of this comparative example does not achieve a better sleep-aiding effect.

[0097] Comparative Example 4

[0098] This embodiment provides a method for preparing a composition that helps improve sleep and its application in a zebrafish sleep experiment.

[0099] (1) Extraction of Chinese medicinal materials: 40% of Ziziphus jujuba seeds, 40% of Schisandra chinensis, and 20% of Cibotium barometz were weighed and crushed according to the mass percentage, and 10 times of 20% ethanol solution was added. Ultrasonic extraction was performed twice at 450W, each time for 80 minutes. After the extraction, the mixture was cooled to room temperature, filtered, and the filtrate was combined;

[0100] (2) Concentration and drying: The filtered extract is concentrated and dried under reduced pressure and vacuum at 45° C. to obtain a dry extract;

[0101] (3) Preparation and purification: The above dry extract was dissolved in ethanol solution, adsorbed on AB-8 resin column for 2 hours, eluted with 5 times ultrapure water for 2 hours, eluted with 5 times 20% ethanol solution for 2 hours, eluted with 95% ethanol solution for 2 hours, and the eluate was collected;

[0102] (5) Concentration and drying: The collected low-concentration ethanol and ultrapure water eluents are concentrated and dried under reduced pressure and vacuum to obtain the Chinese medicine extract, i.e., the composition.

[0103] The zebrafish experiment was conducted on the samples prepared above. Zebrafish were raised in fish farming water at 28°C (water quality: 200 mg of instant sea salt was added to each 1L of reverse osmosis water, the conductivity was 450-550 μS / cm; pH was 6.5-8.5; hardness was 50-100 mg / LCaCO3), and were bred and provided by this laboratory. Wild-type AB strain zebrafish were bred in natural pairs. The experimental fish were zebrafish 5 days after fertilization (5dpf) for the evaluation of sleep improvement efficacy. From the beginning of fertilization, the zebrafish controlled the light cycle to 14±0.5 hours of light: 10±0.5 hours of darkness. 5dpf wild-type AB strain healthy, non-deformed, uniform zebrafish were randomly selected in a 6-well plate. The sample was dissolved in purified water and added to the experimental system at a final concentration of 1000 μg / mL of the composition, and the positive control was 1000 μg / mL of Tianmeijian brand bovine colostrum and jujube seed ganoderma capsule. A normal control group and a model control group were set up at the same time, and the capacity of each well was 3 mL. After 24 hours of treatment at 28°C, except for the normal control group, all other experimental groups were treated with pentylenetetrazol (PTZ) for 1 hour to establish a zebrafish insomnia model. Ten zebrafish were randomly selected from each experimental group and transferred to a 96-well plate, 1 tail / 200 μL / well.

[0104] The final awakening activity data of the normal group, model group, positive group and experimental group are as follows:

[0105] Group Awakening activity mm Normal group <![CDATA[8130.976±534.005 *** ]]> Model Group 23852.772±545.439 Positive group <![CDATA[8863.737±463.315 *** ]]> Experimental Group <![CDATA[18954.663±389.671 * ]]>

[0106] The composition prepared in this comparative example omitted the western medicine component melatonin. Compared with the model group, the amount of wakefulness activity decreased by 20.5%, and the positive group decreased by 62.8%. It can be seen that the sleep-aiding effect of this comparative example is much lower than the composition formula of the embodiment, and the sleep-aiding effect is not good. Compared with comparative example 5, it can be seen that the sleep-aiding effect is worse, indicating that the sleep-aiding effect of the composition in this comparative example is far less good than that of melatonin.

[0107] Comparative Example 5

[0108] In this comparative example, melatonin was used alone for zebrafish experiments. Zebrafish were raised in fish farming water at 28°C (water quality: 200 mg of instant sea salt was added to every 1L of reverse osmosis water, the conductivity was 450-550μS / cm; pH was 6.5-8.5; hardness was 50-100 mg / L CaCO3), and were bred and provided by this laboratory. Wild-type AB strain zebrafish were bred in natural pairs. The experimental fish were zebrafish 5 days after fertilization (5dpf) for the evaluation of the efficacy of improving sleep. From the beginning of fertilization, the zebrafish controlled the light cycle to 14±0.5 hours of light: 10±0.5 hours of darkness. 5dpf wild-type AB strain healthy, deformity-free, uniform zebrafish were randomly selected in a 6-well plate. The sample was dissolved in purified water, and the final concentration of melatonin added to the experimental system was 1000μg / mL, and the positive control Tianmeijian brand bovine colostrum and jujube seed ganoderma lucidum capsule was 1000μg / mL. A normal control group and a model control group were set up at the same time, and the capacity of each well was 3mL. After 24h of treatment at 28℃, except for the normal control group, the remaining experimental groups were treated with pentylenetetrazol (PTZ) for 1h to establish the zebrafish insomnia model. Ten zebrafish were randomly selected from each experimental group and transferred to a 96-well plate, 1 tail / 200μL / well.

[0109]

[0110]

[0111] This comparative example only uses melatonin. Compared with the model group, the amount of wakefulness activity decreased by 22.5%, and the positive group decreased by 71.0%. It can be seen that the sleep-inducing effect of the simple melatonin is far lower than that of the combination formula in the embodiment, and the sleep-inducing effect is not good.

[0112] In summary, the Chinese and Western combination of sleep-improving composition formula used in this application has a significant improvement in sleep effect, and has a sleep-aiding effect close to that of commercially available sleep-aiding products. It is not difficult to see from the sleep-aiding data of the comparative example that the components in this application have a synergistic effect. It can be seen from the data comparison of Examples 1-3 that when the mass ratio of Example 1 is adopted, the sleep-aiding effect is the best.

[0113] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A composition for improving sleep, characterized in that: The raw materials of the composition include: spiny jujube seeds, schisandra chinensis, citrus aurantium and melatonin.

2. The method for preparing a composition for improving sleep according to claim 1, characterized in that: The steps include: Step 1: Mix and crush the spiny jujube seeds, schisandra chinensis, and citronella, add an ethanol solution, perform ultrasonic extraction, and filter to obtain an extract; Step 2, concentrating and drying the extract, re-dissolving, purifying and drying to obtain a Chinese medicine extract; Step 3: After mixing the Chinese herbal extract with melatonin, a composition that helps improve sleep is obtained.

3. The method for preparing a composition for improving sleep according to claim 2, characterized in that: The mass percentage of the spiny jujube kernel is 20-50%, the mass percentage of the schisandra chinensis fruit is 20-50%, and the mass percentage of the citronella fruit is 10-30%.

4. The method for preparing a composition for improving sleep according to claim 2, wherein: In step 1, the mass ratio of the ethanol solution to the crushed medicinal material is (10-15):1, and the concentration of the ethanol solution is 20-50% vol.

5. The method for preparing a composition for improving sleep according to claim 2, characterized in that: In step 1, the ultrasonic power is 450-500W, the ultrasonic extraction time is 60-90min, and the extraction times are 2-3 times.

6. The method for preparing the composition for improving sleep according to claim 2, characterized in that: In step 2, the temperature for concentration and drying is 40-45°C.

7. The method for preparing a composition for improving sleep according to claim 2, characterized in that: In step 2, the purification and drying method includes: dissolving the concentrated and dried extract with ethanol, adsorbing it on an AB-8 resin column for 2 hours, and then eluting it with 5 times the column volume of purified water for 2 hours, eluting it with 5 times the column volume of a low-concentration ethanol solution for 2 hours, and eluting it with a high-concentration ethanol solution for 2 hours, collecting the ultrapure water eluate and the low-concentration ethanol eluate, combining them, concentrating and drying them under reduced pressure, and obtaining a traditional Chinese medicine extract.

8. The method for preparing a composition for improving sleep according to claim 2, wherein: In step 3, the mass ratio of the Chinese herbal extract to melatonin is 1:(1-4).

9. The method for preparing a composition for improving sleep according to claim 7, wherein: The concentration of the low concentration ethanol solution is 20-50% vol.

10. The method for preparing the composition for improving sleep according to claim 7, characterized in that: The concentration of the high concentration ethanol solution is 95% vol.

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