LU501506B1 - Application of a Composition in Preventing and Treating Alcoholic Brain Injury - Google Patents
Application of a Composition in Preventing and Treating Alcoholic Brain Injury Download PDFInfo
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Abstract
The invention discloses an application of a composition in preventing and treating alcoholic brain injury, which belongs to the technical field of biomedicine. The invention provides an application of the composition in preparing pharmaceutical compositions for preventing and treating alcoholic brain injury. The composition comprises the following components in parts by weight: 10-30 parts of algae polyphenol, 2-6 parts of sodium alginate, 30-55 parts of abalone peptide and 0.5-3 parts of spermidine; the algae polyphenol with antioxidant and antibacterial functions is combined with abalone peptide rich in various nutrients and multiple biological effects. Animal experiments show that the composition plays a multi-way, multi-target and synergistic role in the prevention and treatment of alcoholic brain injury. At the same time, the algae polyphenol has good product stability, high bioavailability, safety, no toxicity, convenient administration, no bad smell and good compliance.
Description
DESCRIPTION 507506 Application of a Composition in Preventing and Treating Alcoholic Brain Injury
TECHNICAL FIELD The invention relates to the technical field of biomedicine, in particular to the application of a composition in preventing and treating alcoholic brain injury.
BACKGROUND With the social progress and the rapid development of economic level, the living standard of our people has been significantly improved, and the demand for alcohol consumption is increasing. Chronic alcoholism has become one of the global public health problems. The influence of alcohol on body organs and their various systems mainly spread to the outside of the liver through liver metabolism, including the central nervous system, cardiovascular system, immune system, kidney, lung, gastrointestinal tract, pancreas and other organs. The cerebral cortex of people who drink alcohol for a long time will be in a state of anesthesia, and the damage to the nervous system will be manifested in a series of release states of thinking and behavior, such as the patient's intellectual disorder, impulsiveness, ataxia, incoherent speech, vomiting, lethargy, etc. In severe cases, it may cause epilepsy, convulsions and even death. Alcohol interferes with tissue oxidative stress defense system, causes structural changes of lipids and protein in the cell membrane, changes the fluidity of nerve cell membrane and the activity of adenosine triphosphate, hinders calcium transport, causes nerve cell dysfunction and leads to nerve cell death. As a lipophilic small molecule toxic substance, alcohol can cause damage to both central and peripheral nerves, especially through the blood-brain barrier, which easily acts on the brain, causing damage to brain nerve cells. At present, the incidence of brain diseases caused by alcohol continues to increase, but there are few studies on the mechanism of brain damage and protective effects of alcohol on the brain. Therefore, it is of market value to find an anti-hangover product that can effectively prevent and treat alcoholic brain damage. In the prior art, many substances for preventing and treating alcoholic brain injury are disclosed, for example, Chinese patent CN 201510956371.1 discloses a teabag composition for relieving alcoholic liver, brain and heart injury, which is mainly 507506 composed of Radix puerariae, Fructus crataegi, Hovenia dulcis Thunb, Flos chrysanthemi, Rhizoma phragmitis and Fructus chaenomelis, and has the function of protecting alcoholic liver and heart injury. Chinese patent CN 201310692332.6 discloses the application of resveratrol glycoside in preparing drugs for treating brain damage caused by chronic alcoholism, which is mainly composed of resveratrol glycoside and has a neuroprotective effect on chronic alcoholism. Chinese patent CN
201510915401.4 discloses the application of Chimaera phantasma extract in protective drugs for alcoholic brain injury, which is mainly composed of Chimaera phantasma extract, and has a protective effect on brain injury of alcoholic rats. Most of these substances have a wide curative effect, which can increase the activity of antioxidant enzymes in vivo and reduce the oxidative damage of the brain, but it is difficult to explain the protective mechanism of alcoholic brain injury, and the bioavailability of antioxidant active substances is low.
SUMMARY The purpose of the present invention is to provide an application of a composition in preventing and treating alcoholic brain injury to solve the above-mentioned problems existing in the prior art. The composition has a simple formula, good stability, strong oxidation resistance, good effect of preventing and treating alcoholic brain injury and convenient administration. To achieve the above objective, the present invention provides the following scheme: the invention provides an application of a composition in preparing a pharmaceutical composition for preventing and treating alcoholic brain injury, which comprises the following components in parts by weight: 10-30 parts of algae polyphenol, 2-6 parts of sodium alginate, 30-55 parts of abalone peptide and 0.5-3 parts of spermidine. Among them, algae polyphenols are polyphenol compounds extracted from algae, the main components are phloroglucinol and its derivatives, and its hydroxyl group is an important group with antioxidant activity. Algae polyphenols are considered as natural antioxidants to prevent or reduce chronic diseases because of their ability to scavenge free radicals and reactive oxygen species directly or indirectly.
However, polyphenols 507506 have poor stability and low bioavailability in vivo.
Sodium alginate is a kind of biopolymer material extracted from the sea, which has the advantages of biocompatibility, hydrophilicity and non-toxicity.
It can form hydrogels by ionization under mild conditions.
Alginate hydrogels are usually prepared by physical crosslinking, chemical crosslinking and enzyme crosslinking, and are widely used in the controlled release system of pharmaceutical active peptides and protein.
Abalone peptide is made from abalone meat by enzymatic hydrolysis, and it is known as the "soft gold" of the sea because of its complete amino acids and reasonable proportion.
Abalone is rich in nutrients such as EPA, DHA, trace elements, taurine and superoxide dismutase, and has the functions of anti-oxidation, enhancing immunity and anti-fatigue.
Spermidine is a naturally occurring polyamine, a safe and efficient inducer of autophagy, which exists in almost all cells and has the functions of anti-aging, anti-cancer, cardiovascular and nerve protection, etc.
Spermidine is beneficial to assist drug delivery, improve the water solubility, stability and membrane permeability of drug molecules, and further improve the absorption of drugs in organisms.
According to the invention, alginate, spermidine and algae polyphenol with antioxidant and antibacterial functions are blended to prepare algae polyphenol particles.
The spermidine molecule contains three amino groups, and the amino groups at both ends and in the middle can be connected with sodium alginate and algae polyphenols respectively, forming an "umbrella" type protection structure for algae polyphenols and improving the stability of products.
Algae polyphenol particles and abalone peptides rich in various nutrients and multiple functions form a compound composition through multiple non-covalent bonds such as hydrogen bonding, static electricity, x-x stacking, etc., which has a synergistic effect on the prevention and treatment of alcoholic brain injury through various ways and multiple targets.
When the composition enters the stomach, sodium alginate forms alginic acid colloid under the action of gastric acid, which wraps alginic polyphenol and abalone peptide, thus avoiding the loss of activity in a complex environment in vivo; in the intestinal tract, alginic acid is converted into an ionic salt state, which can continuously release alginic polyphenols and abalone 507506 peptides, exert lasting activity and improve its bioavailability. The algae polyphenol is derived from edible macroalgae, including seaweed, kelp, gulfweed, Gracilaria lemaneiformis, Sargassum fusiforme, Undaria pinnatifida or Gracilaria. The extraction method of algae polyphenols can be carried out according to the conventional extraction method of algae polyphenols in the field. Preferably, the total phenol content in the algae polyphenols used in the invention is not less than 30% by weight. The abalone peptide is a compound peptide prepared by biological enzymolysis of abalone meat. The enzymolysis method of abalone peptide can be carried out according to the conventional enzymolysis method in the field. Preferably, the molecular weight distribution range of abalone peptides used in the present invention is: 2.15% of polypeptide with molecular weight > 3.0 kDa, 16.59% of polypeptide with molecular weight of 1.0-3.0 kDa and 81.26% of polypeptide with molecular weight < 1.0 kDa. Further, the composition comprises the following components in parts by weight: 20 parts of algae polyphenol, 4 parts of sodium alginate, 40 parts of abalone peptide and
1.5 parts of spermidine. Further, the preparation method of the composition comprises the following steps: S1, adding sodium alginate into water to prepare a sodium alginate solution, adjusting the pH of the solution to 6.0-7.5, adding spermidine, stirring at room temperature for 5-8 hours, treating with ice-water bath, adding algae polyphenols, stirring for 1-3 hours, freeze-drying, and crushing with liquid nitrogen to obtain algae polyphenol composite powder; S2, dissolving abalone peptide with water to prepare abalone peptide solution, adding the algae polyphenol composite powder under stirring, uniformly suspending, and freeze-drying to obtain the composition. During the preparation of the composition, the concentration of sodium alginate solution and abalone peptide solution has an important influence on the stability and synergy of the composition.
Further, the mass concentration of the sodium alginate solution is 1%-3.5%, preferably 507506
1.5%; if the mass concentration of sodium alginate solution is too large (> 3.5%), it is easy to agglomerate, difficult to form uniform algae polyphenol particles, and the efficacy is poor; if the mass concentration of sodium alginate solution is too small (< 1%), the interaction with spermidine is weak, and the protection effect of algae polyphenols cannot be achieved. Further, the mass concentration of the abalone peptide solution is 18%-26%, preferably 22%; if the mass concentration of abalone peptide solution is too large (> 26%), it will be easily infected by microorganisms and lose its activity; if the mass concentration of abalone peptide solution is too small (< 18%), the synergistic effect with algae polyphenols will be weakened, and the effect of preventing and treating alcoholic brain injury will not be achieved. Further, in step S1, it is necessary to strictly control the pH range of the solution. The method for adjusting the pH of the present invention is a conventional method. Because the sodium alginate solution is weakly alkaline, 0.1 mol/L HCI solution is used for adjusting the pH of the present invention; if the pH value of the solution is too low (<
6.0), the acidity is too strong, and sodium alginate generates alginic acid colloid and precipitates out; if the pH of the solution is too high (> 7.5), it is difficult for spermidine to form a composite umbrella structure with sodium alginate and algae polyphenols through electrostatic interaction. Therefore, in that present invention, the pH of the sodium alginate solution is preferably 6.0 to 7.5, more preferably 6.5. As an illustration, the room temperature involved in the present invention is also normal saline temperature or general temperature, generally defined as 25°C, which can be reasonably adjusted by the technicians in the field, and it is also within the scope of protection of the present invention. Furthermore, the pharmaceutical composition also includes pharmaceutically acceptable excipients. Further, the pharmaceutical composition is a tablet, and the auxiliary materials include corn starch, talcum powder and magnesium stearate; the mass fraction of each component in the tablet is as follows:
10-20 wt% of the composition, 65-75 wt% of corn starch, 11-17 wt% of talcum powder HUS0T506 and 0.5-2 wt% of magnesium stearate.
Further, the pharmaceutical composition is a capsule, and the auxiliary materials include lactose, corn starch and talcum powder; the mass fraction of each component in the capsule is: 10-30 wt% of the composition, 10-20 wt% of lactose, 50-60 wt% of corn starch and 5- wt% of talcum powder.
Further, the pharmaceutical composition is granules, and the auxiliary materials include corn starch, sodium hydroxymethylcellulose and magnesium stearate; the mass fraction of each component in the granule is as follows: 5-15 wt% of the composition, 74-84 wt% of corn starch, 5-15 wt% of sodium hydroxymethylcellulose and 0.5-2 wt% of magnesium stearate.
The invention discloses the following technical effects: (1) the invention combines algae polyphenols with antioxidant and antibacterial functions and abalone peptides rich in various nutrients and multiple biological effects, and forms a complex through multiple actions such as hydrogen bonding, static electricity, x-x stacking and the like; the animal model test shows that the compound plays a multi-way, multi-target and synergistic role in the prevention and treatment of alcoholic brain injury; (2) the present invention utilizes the hydrogel characteristics of alginate to form alginate colloid in the stomach at low pH, and wrap the algae polyphenol and abalone peptide complex, thus avoiding the loss of its activity in the complex environment in vivo; in the intestinal tract, alginic acid is converted into ionic salt state, and algae polyphenols and abalone peptides are released slowly and controlled, so as to exert lasting active effect; (3) the spermidine added in the invention is connected with alginate and algae polyphenol respectively by its "triamine" characteristic, forming an "umbrella" type protection structure for algae polyphenol, so that the stability, physical properties and taste of algae polyphenol are improved, which is beneficial to its prevention and treatment effect in alcoholic brain injury;
(4) the raw materials of the invention are all from natural food, which is green and safe; 507506 the preparation process is simple and easy for industrial production; it is also safe, non- toxic, convenient to take, and has no bad smell and good compliance.
BRIEF DESCRIPTION OF THE FIGURES In order to more clearly explain the embodiments of the present invention or the technical solutions in the prior art, the figures needed in the embodiments will be briefly introduced below. Obviously, the figures in the following description are only some embodiments of the present invention, and for ordinary technicians in the field, other figures can be obtained according to these figures without creative efforts. FIG. 1 shows the DPPH free radical scavenging ability of the composition for preventing and treating alcoholic brain injury prepared by the present invention. FIG. 2 shows the OH radical scavenging ability of the composition for preventing and treating alcoholic brain injury prepared by the present invention. FIG. 3 is the antioxidant stability of the composition for preventing and treating alcoholic brain injury prepared by the present invention. FIG. 4 is the influence of the composition for preventing and treating alcoholic brain injury prepared by the invention on the brain index of mice. FIG. 5 shows the influence of the composition for preventing and treating alcoholic brain injury prepared by the invention on the swimming route of mice after platform removal.
DESCRIPTION OF THE INVENTION Now, various exemplary embodiments of the present invention will be described in detail. This detailed description should not be considered as a limitation of the present invention, but should be understood as a more detailed description of some aspects, characteristics and embodiments of the present invention. It should be understood that the terms used in this invention are only for describing specific embodiments, and are not used to limit the invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper limit and the lower limit of the range is also specifically disclosed. Any stated value or intermediate value within the stated range and any other stated value or every smaller range between intermediate values within the stated range 507506 are also included in the present invention.
The upper and lower limits of these smaller ranges can be independently included or excluded from the range.
Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by the ordinary technicians in the field of this invention.
Although the present invention only describes the preferred methods and materials, any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention.
All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and/or materials related to the documents.
In case of conflict with any incorporated documents, the contents of this specification shall prevail.
Without departing from the scope or spirit of the present invention, it is obvious to those skilled in the art that many modifications and changes can be made to the specific embodiments of the present invention.
Other embodiments obtained from the description of the present invention will be obvious to the skilled person.
The specification and embodiments of this application are exemplary only.
The words "including", "comprising", "having" and "containing" used in this paper are all open terms, that is, they mean including but not limited to.
Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the technical field.
Unless otherwise specified, the reagents and materials used in the following embodiments are all commercially available.
As an illustration, the algae polyphenols and abalone peptides used in the present invention are prepared by conventional methods, and the algae polyphenols can also be purchased from Shandong Jiejing Group Corporation, and the abalone peptides can also be purchased from Shandong Donghai Longyuan Bio-tech Co., Ltd.
Embodiment 1 A composition for preventing and treating alcoholic brain injury comprises the following components in parts by weight: 20 parts of algae polyphenols, 4 parts of sodium alginate, 40 parts of abalone peptide and 1.5 parts of spermidine.
The preparation method of the composition comprises the following steps: 507506 S1, adding sodium alginate into distilled water to prepare a 1.5 wt% sodium alginate solution according to the parts by weight, adjusting the pH value of the solution to 6.5, adding spermidine, stirring at room temperature for 6.5 hours, treating with ice water bath, adding algae polyphenol after the solution system is cooled to 8 °C, continuously stirring for 2 hours, freeze-drying, and crushing with liquid nitrogen to obtain algae polyphenol composite powder; S2, dissolving abalone peptide with distilled water according to the parts by weight to prepare 22 wt% abalone peptide solution, adding the algae polyphenol composite powder in S1 under stirring, uniformly suspending, and freeze-drying to obtain the composition.
Embodiment 2 A composition for preventing and treating alcoholic brain injury comprises the following components in parts by weight: 15 parts of algae polyphenols, 3 parts of sodium alginate, 35 parts of abalone peptide and 1 part of spermidine.
The preparation method of the composition comprises the following steps: S1, adding sodium alginate into distilled water to prepare a 2.0 wt% sodium alginate solution according to the parts by weight, adjusting the pH of the solution to 7.0, adding spermidine, stirring at room temperature for 7 hours, treating in an ice-water bath, adding algae polyphenols after the solution system is cooled to 6°C, stirring for 1.5 hours, freeze-drying, and crushing with liquid nitrogen to obtain algae polyphenol composite powder; S2, dissolving abalone peptide with distilled water according to the parts by weight to prepare 24 wt% abalone peptide solution, adding the algae polyphenol composite powder in S1 under stirring, uniformly suspending, and freeze-drying to obtain the composition.
Embodiment 3 A composition for preventing and treating alcoholic brain injury comprises the following components in parts by weight: 10 parts of algae polyphenol, 2 parts of sodium alginate, 30 parts of abalone peptide and 0.5 part of spermidine.
The preparation method of the composition comprises the following steps: 507506 S1, adding sodium alginate into distilled water to prepare a 1 wt% sodium alginate solution according to the parts by weight, adjusting the pH of the solution to 6.0, adding spermidine, stirring at room temperature for 5 hours, treating with an ice water bath, adding algae polyphenols after the solution system is cooled to 5°C, stirring for 1 hour, freeze-drying, and crushing with liquid nitrogen to obtain algae polyphenol composite powder; S2, dissolving abalone peptide with distilled water according to the parts by weight to prepare 18 wt% abalone peptide solution, adding the algae polyphenol composite powder in S1 under stirring, uniformly suspending, and freeze-drying to obtain the composition.
Embodiment 4 A composition for preventing and treating alcoholic brain injury comprises the following components in parts by weight: 30 parts of algae polyphenols, 6 parts of sodium alginate, 55 parts of abalone peptide and 3 parts of spermidine.
The preparation method of the composition comprises the following steps: S1, adding sodium alginate into distilled water to prepare a 3.5 wt% sodium alginate solution according to the parts by weight, adjusting the pH of the solution to 7.5, adding spermidine, stirring at room temperature for 8 hours, treating in an ice water bath, adding algae polyphenol after the solution system is cooled to 10°C, continuously stirring for 3 hours, freeze-drying, and crushing with liquid nitrogen to obtain algae polyphenol composite powder; S2, dissolving abalone peptide with distilled water according to the parts by weight to prepare 26 wt% abalone peptide solution, adding the algae polyphenol composite powder in S1 under stirring, uniformly suspending, and freeze-drying to obtain the composition.
Comparative embodiment 1 The only difference from Embodiment 1 is that vitamin C is used instead of algae polyphenols.
Comparative embodiment 2
The only difference from Embodiment 1 is that spermine is used instead of spermidine. 507506 Comparative embodiment 3 The difference from Embodiment 1 only lies in the direct blending of raw materials, specifically as follows: a composition for preventing and treating alcoholic brain injury comprises the following components in parts by weight: 20 parts of algae polyphenols, 4 parts of sodium alginate, 40 parts of abalone peptide and 1.5 parts of spermidine. The preparation method of the composition comprises the following steps: According to the parts by weight, taking sodium alginate, spermidine, algae polyphenol and abalone peptide as raw materials, adding distilled water, uniformly suspending, and freeze-drying to obtain the composition. Comparative embodiment 4 The only difference from Embodiment 1 is that oyster peptide is used instead of abalone peptide. Application embodiment 1 Antioxidation experiment of composition for preventing and treating alcoholic brain injury
1. Experimental method (1) Determination of free radical scavenging capacity of 1,1-diphenyl-2- trinitrophenylhydrazine (DPPH)
2.0 mL of sample solutions with different mass concentrations (the compositions prepared in Embodiments 1-4 and Comparative Embodiments 1-4) are mixed with 2 mL DPPH absolute ethyl alcohol solution, and then the absorbance of the solution is measured at 517 nm after standing in the dark for 60 min at room temperature. Using distilled water instead of sample solution as blank control, calculate the scavenging rate of DPPH free radicals according to the following formula (1), and use ICso to express the scavenging effect.
Scavenging rate (%) = on * 100 (1) Wherein Ay is the absorbance of the sample and Ay is the absorbance of the blank control.
(2) Determination of hydroxyl radical scavenging capacity 7501506 Add 1 mL HO; solution (8.8 mmol/L), 1 mL FeSO; solution (9 mmol/L) and 12 mL sample solutions with different mass concentrations (the compositions prepared by Embodiments 1-4 and Comparative Embodiments 1-4) into the same colorimetric tube, then add 1 mL salicylic acid ethanol solution (9 mmol/L), shake well, and place in a water bath at 37°C for 30 min. Use distilled water instead of sample solution as blank control. The hydroxyl radical scavenging rate is calculated according to formula (1), and the scavenging effect is expressed by ICso.
2. Experimental results (1) DPPH free radical scavenging effect The smaller the ICso value, the stronger the ability to scavenge free radicals. Vitamin C (Vo) is a strong antioxidant, which is used as positive control. The DPPH free radical scavenging effect of the composition for preventing and treating alcoholic brain injury is shown in FIG. 1. The ICso of Embodiments 1 to 4 groups are close to that of Vc group, showing strong DPPH free radical scavenging effects, especially the DPPH free radical scavenging ability of Embodiment 1 is the strongest. Although the group of Comparative embodiment 1 also showed DPPH radical scavenging effect close to those of Embodiments 1-4, the DPPH radical scavenging abilities of Comparative embodiments 2-4 are far inferior to that of Embodiments 1-4. (2) Hydroxyl radical scavenging effect Vitamin C (Vc) is a strong antioxidant, which is used as positive control. The hydroxyl radical scavenging effect of the composition for preventing and treating alcoholic brain injury is shown in FIG. 2. The ICso of Embodiments 1-4 groups is almost close to that of Vc group, showing a strong hydroxyl radical scavenging effect, especially in Embodiment 1. Although the hydroxyl radical scavenging effect of Comparative embodiment 1 is similar to that of Embodiments 1-4, the hydroxyl radical scavenging abilities of Comparative embodiment 2-4 are far less than that of Embodiments 1-4. Based on the above two experimental results, embodiments 1-4 all showed strong free radical scavenging effects, which indicates that the composition for preventing and treating alcoholic brain injury prepared by the invention had strong antioxidant activity,
especially in Embodiment 1. The formula or preparation method of the composition of 507506 the invention is changed, and the expected antioxidant effect can not be achieved. Application embodiment 2 Experiment on antioxidant stability of the composition for preventing and treating alcoholic brain injury Ferric reducing ability of power (FRAP) test is used to detect the total antioxidant capacity of the composition during storage. Sample storage: the sample is stored in a brown bottle and placed at room temperature. Samples are sampled at regular intervals to test the total antioxidant capacity. Test method: take 0.02 mL FeSO4 solution, add 0.18 mL FRAP solution (ready-to-use), mix well, take a water bath at 37°C for 10 min, and measure the absorbance at 593 nm by UV-Vis spectrophotometer. The FeSO, equivalent standard curve is established with FeSO; solution concentration as abscissa and absorbance as ordinate. With Vc solution as positive control, FeSO4 equivalent is calculated according to the standard curve. The relative percentage of Vc of each sample is calculated according to FeSO4 equivalent. The results of antioxidant stability of the composition for preventing and treating alcoholic brain injury are shown in FIG. 3. Because of the instability of polyphenol properties, polyphenols are easily oxidized and easily deteriorated during long-term storage. The total antioxidant capacity of the composition of Comparative embodiment 3 decreased almost linearly during storage, while the total antioxidant capacity of the composition of Embodiment 1 remained almost stable with the extension of storage time within 5 days in the same environment. Therefore, the formula and preparation method of the composition can effectively improve the antioxidant stability of the composition. Application embodiment 3 Determination of brain protection efficacy of the composition for preventing and treating alcoholic brain injury
1. Molding method Experiments: Kunming male mice, aged 6-8 weeks, weighing 18-22 grams, SPF grade, animal feeding conditions: (25+1)°C, relative humidity: (55+5)%.
Experiment: 140 mice are randomly divided into 10 groups, with 14 mice in each group, 507506 which are divided into saline group, model group, Embodiments 1-4 groups and Comparative embodiments 1-4 groups. Each group of mice is gavaged with the corresponding volume of 56-degree Red Star Erguotou Baijiu (saline group is gavaged with the corresponding volume of saline) at a graded concentration daily, and 1 hour after gavage, each group is gavaged with 10 g/kg-BW (body weight), while the saline and model groups are gavaged with saline. Gavage is continued for 8 weeks. The mice are weighed weekly to adjust the dose of wine and the test substance. The gavage dose is 2 mL/kg-BW for the first week in order to cause persistent alcohol damage, after which the gavage dose is increased according to the mortality of the mice each week, etc.
2. Morris water maze experiment Alcoholic brain injury can obviously reduce the learning and memory ability of mice. The ability of mice to learn and remember the position and direction of space is tested by searching for the platform hidden under water in the water maze. Morris water maze test is performed on mice after 8 weeks of alcohol injury. Mice swim in a black circular pool with a diameter of 120 cm and a height of 40 cm. The pool is filled with water and contained a movable platform with a diameter of 8 cm. The experiment lasts for five days, four times a day, and the mouse is put into water from four different starting points. Record the time when the mouse found the platform (escape latency). On the 6 day, the original platform is removed, and the mice are put into the water at any water entry point. The residence time of the mice in the original platform, the times and paths of crossing the original platform are recorded. Samples are collected after fasting for 24 hours on the second day of the experiment: (1) after dissecting the mouse, take out the brain tissue, wash away the blood on the surface with precooled normal saline, suck it dry with filter paper, quickly weigh it, and calculate the brain index with the formula: Brain index (%)=brain weight (g)/body weight (g)*100% (2) quickly dissect brain tissue, store at -80°C, and prepare 10% brain homogenate; according to the requirements of corresponding kits, strictly operate, and measure the contents of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and 507506 malondialdehyde (MDA) in brain homogenate.
2. Experimental results (1) Brain index If the brain index increases, it means congestion of organs, edema, hyperplasia and hypertrophy, etc. If the brain index decreases, it indicates atrophy and other degenerative changes of organs. The experimental results of brain index of the composition for preventing and treating alcoholic brain injury are shown in FIG. 4. After the mouse model is established, compared with the normal saline group, the brain index of the model group is increased significantly (p<0.01), which indicates that excessive drinking would cause brain edema in mice. Compared with the model group, the brain index of mice in Embodiments 1-4 groups decreased significantly with a statistical difference (p< 0.01), among which the effect of Embodiment 1 group is the best, and the brain index is almost close to that of normal saline group, which indicate that the composition for preventing and treating alcoholic brain injury prepared by the invention could effectively improve the brain index, reduce edema, hyperplasia, hypertrophy and other pathological changes in brain tissue, and had significant preventive and therapeutic effects on alcoholic brain injury. Compared with the model group, although the brain index of mice in Comparative embodiments 1-4 is decreased, there is no statistical difference, which showed that the change of the formula or preparation method of the patent composition could not effectively reduce the brain index induced by alcohol, nor could it prevent and treat the brain injury induced by alcohol. (2) Analysis of SOD, GSH-Px and MDA results The damage of alcohol to organism cells is mainly through inducing oxidative stress. Excessive long-term drinking will lead to the changes of SOD and GSH activities in brain cells of the organism, which will further lead to oxidative stress to damage the central nervous system. Therefore, alcoholic brain injury can be prevented by regulating the activity of antioxidant enzymes in vivo. The effect of the composition for preventing and treating alcoholic brain injury on oxidase activity is shown in Table 1. Compared with the normal saline group, the contents of SOD and GSH-Px in the brain tissue of 507506 the model group decreased significantly, with statistical difference (p< 0.01), which indicate that the abnormal saline oxidative stress in the brain of mice caused by continuous excessive drinking is one of the important factors that caused brain damage in mice.
Compared with the model group, the contents of SOD and GSH-Px in the brain tissue of mice in Embodiments 1-4 are significantly increased (p< 0.01), especially in Embodiment 1, which show that the composition for preventing and treating alcoholic brain injury prepared by the invention could significantly reduce the level of peroxide in the brain of mice caused by alcohol and enhance the antioxidant capacity of brain tissue.
MDA is the product of free radicals acting on lipid peroxidation, and the degree of lipid peroxidation in vivo can be reflected by measuring MDA content.
The MDA contents in the brain tissue of mice in Embodiments 1-4 groups are significantly lower than that in the model group (p< 0.01), among which the MDA content in Embodiment 1 group is the lowest.
The results show that the composition for preventing and treating alcoholic brain injury prepared by the invention can increase the activity of SOD and GSH-Px, enhance the antioxidant capacity of mouse brain tissue, correct the balance disorder of oxidation and antioxidant, and reduce the damage of free radicals to brain cells, thus protecting brain injury caused by chronic alcoholism.
Table 1 Contents of SOD, GSH-Px and MDA in brain tissue of mice in each group © Gow SOD GSHPx MDA (U/mg-protein) (U/mg-protein) (nmol/mg-protein) normal saline 119.25+£8.37 55.30+2.34 4.51+0.94 Model 78.03+8.09" 25.3042.34% 7.11£0.91% Embodiment 1 115.57+9.62" 51.11+5.06"" 4,54+0.88" Embodiment 2 109.97+9.12* 49.5245.07" 4.77+0.83" Embodiment 3 103.43+11.46"" 45.23+4.78”" 4.89+0.94™ Embodiment 4 98.67+8.49" 42.8842 85" 4.93+0.98"* Somparaive 01.87+7.62* 37.63+4.12* 6.83+1.02 opera 83.80+11.10 31.26+4.74 6.91+0.99 Somparaiive 85.07+10.42 33.48+2.87 6.54+0.84"
Comparative LUS01506 embodiment 4 80.08+11.14 28.00+2.12 7.02+0.95 Note: Compared with normal saline group, *p<0.01; compared with the model group, “p<0.05, “p<0.01. (3) Experimental results of Morris water maze in mice Morris water maze experiment includes two parts: positioning navigation and space search.
The results of escape latency in positioning navigation experiment represent the ability of brain memory acquisition, while the results of target quadrant residence time and times of crossing platform in space search experiment represent the ability of brain memory retention.
The experimental results in Table 2-3 correspond to the results of positioning navigation and spatial search, respectively.
After 5 days of Morris experiment, the escape latency of mice in each experimental group is gradually shortened, and the escape latency of mice in the model group is significantly longer than that in the saline group (p< 0.05). Compared with the model group, the escape latency of mice in Embodiments 1-4 and Comparative embodiments 1-4 decreases day by day from the 2"* day, especially in Embodiments 1-4 (p< 0.01). Compared with the model group, there are significant differences in the time of staying in the target quadrant and the times of crossing the platform after removing the original platform of the mice in Embodiments 1-4 (p< 0.01), among which the mice in Embodiment 1 have the longest time of staying in the target quadrant and the most times of crossing the platform.
In addition, it can be seen from the swimming route map 5 of mice after removing the platform that the purpose of searching for the original platform of mice in the model group is not strong, but the purpose of mice in Embodiment 1 group and saline group is strong.
The results show that the brain memory acquisition function and brain memory retention function of mice are damaged after continuous excessive drinking of alcohol until the eighth week of the experiment, and the composition for preventing and treating alcoholic brain injury prepared by the invention has a remarkable improvement effect on the brain injury of mice caused by alcohol.
Table 2 Results of Morris navigation test of mice (x+s) Escape latency Group _— mm 1d 2d 3d 4d 5d + normal 513310 42344127 35.0141.15 32334451 27.67+4.04 saline 0
79.93+4.4 73.40+3.40 69.63£3.80 68.70+3.45 65.12+£3.95 Model + ## ## ## ## Embodiment 53.33+2.3 43.00+2.65 36.33+1.53 33.06=1.59 28.12+3.04 1 6 Fk Fk Fk Fk Embodiment 54.30+2.3 43.67+£0.58 38.33+1.53 35.33+£0.86 29.32+3.61 2 1 Fk Fk * Fk Embodiment 56.00+2.1 44.82+0.52 37.33£1.19 35.99+3.33 29.87+2.33 3 1 Fk Fk Fk Fk Embodiment 57.26+2.1 46.00+£3.17 40.03£2.66 37.29+3.35 30.19+0.89 4 8 Fk Fk Fk Fk Comparative
73.11+£3.2 58.37+3.12 57.07+£2.11 embodiment 0 65.34+2.29 63.11+3.23 ; ; 1 Comparative 7623427 embodiment 0 68.26+2.29 64244383 63.06+2.73 61.67+1.53 2 Comparative 71 63439 58.22+2.45 embodiment ‘ 2 ‘ 66.00+2.34 63.45+£2.58 61.52+3.36 a 3 Comparative 74 86+1.6 embodiment 4 66.74+3.50 63.83+3.18 62.33+£3.67 61.75+£2.51
A Note: compared with normal saline group, p< 0.01; compared with the model group, “p<0.05, “p<0.01. Table 3 Results of Morris space search test in mice (X+s) Target drant Resid i . Group ABC ame CIEE Times of crossing platform normal saline 23.50+2.54 5.71+0.09 Model 11.50+2.54% 1.68+0.09% Embodiment 1 22.30+2.95”” 4.51+1.03" Embodiment 2 21.50+1.88" 4.13+0.41 Embodiment 3 20.00+1.48" 3.96+0.25" Embodiment 4 19.33+3.34"" 3.78+0.29""
"Comparative 11 LU501506 embodiment 1 13.06+1.24 1.96+0.11 Somparaive 14.26+1.85 1.86+0.12 Somparaiive 13.26+2.18 2.25+0.19" Comparative 12.45+1.93 1.81+0.11 Note: compared with normal saline group, p< 0.01; compared with the model group, “p<0.05, “p<0.01. Embodiment 4 Acute toxicity test Animals: 20 healthy mice, weighing 18-22 g, half male and half female, are randomly divided into experimental group and blank control group.
Subject: the composition of Embodiment 1 1s used as the subject.
Experimental method: In the pre-experiment, the half lethal dose could not be measured, so the above experimental dosage is selected for the experiment.
Taking the weight of the mice as the standard, the mice in the experimental group are given a dose of 10 g/kg once every 24 hours for two weeks, while the mice in the blank control group are given distilled water, and they are all free to drink water and feed.
During the experiment, the activity, diet, feces, respiration, weight and death of the mice are observed.
Results: there is no death in the experimental group within 2 weeks after oral administration.
Compared with the control group, the experimental group has no significant difference in weight change, eating and drinking, and the mice are in good mental state and active.
After execution of the animals, there 1s no pathological changes in the main organs with naked eyes.
The results show that the oral application of the composition for preventing and treating alcoholic brain injury has no obvious damage to animals, no toxicity to the organism, and is safe and nontoxic.
Embodiment 5 A tablet for preventing and treating alcoholic brain injury The tablet for preventing and treating alcoholic brain injury comprises the following components by mass fraction: wt% of the composition prepared in Embodiment 1; 70 wt% of corn starch;
14 wt% of talcum powder; HUS01508 1 wt% of magnesium stearate. Embodiment 6 A tablet for preventing and treating alcoholic brain injury The tablet for preventing and treating alcoholic brain injury comprises the following components by mass fraction: wt% of the composition prepared in Embodiment 2; 75 wt% of corn starch; 13 wt% of talcum powder; 2 wt% of magnesium stearate. Embodiment 7 A tablet for preventing and treating alcoholic brain injury The tablet for preventing and treating alcoholic brain injury comprises the following components by mass fraction: 20 wt% of the composition prepared in Embodiment 3; 65 wt% of corn starch;
14.5 wt% of talcum powder;
0.5 wt% of magnesium stearate. Embodiment 8 A capsule for preventing and treating alcoholic brain injury The capsule for preventing and treating alcoholic brain injury comprises the following components by mass fraction: 20 wt% of the composition prepared in Embodiment 1; wt% of lactose; 55 wt% of corn starch; 10 wt% of talcum powder. Embodiment 9 A capsule for preventing and treating alcoholic brain injury The capsule for preventing and treating alcoholic brain injury comprises the following components by mass fraction: 12 wt% of the composition prepared in Embodiment 3; wt% of lactose; 53 wt% of corn starch; 15 wt% of talcum powder.
Embodiment 10 A capsule for preventing and treating alcoholic brain injury 7501506 the capsule for preventing and treating alcoholic brain injury comprises the following components by mass fraction: 27 wt% of the composition prepared in Embodiment 4; 10 wt% of lactose; 58 wt% of corn starch; wt% of talc powder. Embodiment 11 A granule for preventing and treating alcoholic brain injury The capsule for preventing and treating alcoholic brain injury comprises the following components by mass fraction: wt% of the composition prepared in Embodiment 1; 79 wt% of corn starch; 10 wt% of sodium carboxymethyl cellulose; 1 wt% of magnesium stearate. Embodiment 12 A granule for preventing and treating alcoholic brain injury The capsule for preventing and treating alcoholic brain injury comprises the following components by mass fraction: 7 wt% of the composition prepared in Embodiment 3; 84 wt% corn starch; 7 wt% of sodium carboxymethyl cellulose; 2 wt% of magnesium stearate. Embodiment 13 A granule for preventing and treating alcoholic brain injury The capsule for preventing and treating alcoholic brain injury comprises the following components by mass fraction:
13.5 wt% of the composition prepared in Embodiment 2; 74 wt% corn starch; 12 wt% sodium carboxymethyl cellulose;
0.5 wt% of magnesium stearate. The above-mentioned embodiments only describe the preferred mode of the present invention, and do not limit the scope of the present invention. Without departing from the design spirit of the present invention, all kinds of modifications and improvements 507506 made by ordinary technicians in the field to the technical scheme of the present invention should fall within the protection scope determined by the claims of the present invention.
Claims (9)
1. An application of a composition in preparing a pharmaceutical composition for preventing and treating alcoholic brain injury, which comprises the following components in parts by weight: 10-30 parts of algae polyphenols, 2-6 parts of sodium alginate, 30-55 parts of abalone peptide and 0.5-3 parts of spermidine.
2. The application according to claim 1, characterized in that the composition comprises the following components in parts by weight: 20 parts of algae polyphenol, 4 parts of sodium alginate, 40 parts of abalone peptide and 1.5 parts of spermidine.
3. The application according to claim 1 or 2, characterized in that the preparation method of the composition comprises the following steps: S1, adding sodium alginate into water to prepare a sodium alginate solution, adjusting the pH of the solution to 6.0-7.5, adding spermidine, stirring at room temperature for 5-8 hours, treating with ice-water bath, adding algae polyphenols, stirring for 1-3 hours, freeze-drying, and crushing with liquid nitrogen to obtain algae polyphenol composite powder; S2, dissolving abalone peptide with water to prepare abalone peptide solution, adding the algae polyphenol composite powder under stirring, uniformly suspending, and freeze-drying to obtain the composition.
4. The application according to claim 3, characterized in that the mass concentration of the sodium alginate solution is 1%-3.5%; the mass concentration of the abalone peptide solution is 18%-26%.
5. The application according to claim 3, characterized in that the pH of the sodium alginate solution is adjusted to 6.5.
6. The application according to claim 1, characterized in that the pharmaceutical composition further comprises pharmaceutically acceptable excipients.
7. The application according to claim 6, characterized in that the pharmaceutical composition is a tablet, and the auxiliary materials include corn starch, talcum powder and magnesium stearate; the mass fraction of each component in the tablet is as follows:
10-20 wt% of the composition, 65-75 wt% of corn starch, 11-17 wt% of talcum powder 7501506 and 0.5-2 wt% of magnesium stearate.
8. The application according to claim 6, characterized in that the pharmaceutical composition is a capsule, and the auxiliary materials include lactose, corn starch and talcum powder; the mass fraction of each component in the capsule is: 10-30 wt% of the composition, 10-20 wt% of lactose, 50-60 wt% of corn starch and 5- wt% of talcum powder.
9. The application according to claim 6, characterized in that the pharmaceutical composition is granules, and the auxiliary materials include corn starch, sodium hydroxymethylcellulose and magnesium stearate; the mass fraction of each component in the granule is as follows: 5-15 wt% of the composition, 74-84 wt% of corn starch, 5-15 wt% of sodium hydroxymethylcellulose and 0.5-2 wt% of magnesium stearate.
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