KR102634302B1 - Composition for preventing or treating cognitive impairment comprising extract of iris lactea - Google Patents

Abstract

The present invention relates to a composition for the prevention or treatment of cognitive impairment containing an iris lactea extract or a fraction thereof, and specifically, a pharmaceutical composition for the prevention or treatment of cognitive impairment containing an iris lactea extract or a fraction thereof as an active ingredient; food composition; It relates to a feed composition and a method of treating cognitive impairment using the pharmaceutical composition.
The iris extract of the present invention exhibits an excellent normalizing effect and memory enhancing effect on brain damage such as the basal forebrain, white matter, and hippocampus, and thus reduces cognitive impairment related to cerebrovascular diseases such as vascular dementia and cerebral infarction and memory disorders caused by it. It can be useful in treatment.

Description

Composition for preventing or treating cognitive impairment comprising extract of iris lactea}

The present invention relates to a composition for the prevention or treatment of cognitive impairment containing iris lactea extract (ILE) or a fraction thereof, and specifically, to a composition for preventing, improving or treating cognitive impairment containing iris lactea extract (ILE) or a fraction thereof as an active ingredient. therapeutic pharmaceutical compositions; food composition; It relates to a feed composition and a method of treating cognitive impairment using the pharmaceutical composition.

Cognitive ability encompasses memory ability, spatial and temporal awareness, judgment, language ability, and calculation ability, and allows one to carry out daily life without the help of others. However, in all cases of brain damage, such as rapid neuronal death caused by stroke, trauma, etc., or slow neuronal death causing degenerative brain diseases, it immediately results in irreversible dysfunction of the neural network. Eventually, cognitive ability declines and people suffer from diseases such as forgetfulness or memory impairment, dementia, and Alzheimer's disease, and become unable to maintain their previous level of daily life. The number of patients related to the above-mentioned cognitive disorders, which not only causes personal suffering but also causes a decline in the working-age population, is expected to increase to more than 74.7 million by 2030 (Abate G, 2017, Oxid Med Cell Longev. 2017:10 ), the development of a treatment or treatment method is emerging as an urgent problem.

To date, many patents related to compositions with the effect of treating cognitive disorders and improving cognitive abilities have been disclosed and registered in Korea, and the representative published patents are as follows. Publication No. 2014-0144785 relates to a composition for improving memory and learning ability containing citron extract as an active ingredient; Registration No. 10-1837444 relates to a composition for preventing, improving, or treating cognitive dysfunction, containing an extract of prickly pear as an active ingredient; Registration No. 10-1823892 relates to Balsam extract that has the effect of improving memory, improving cognitive ability, and preventing, delaying, or treating dementia. However, although these chemical agents and natural compositions are available on the market, excellent preparations with guaranteed efficacy have not yet been developed. Additionally, commercially available drugs cause many side effects such as hepatotoxicity, insomnia, high blood pressure, and vomiting. Therefore, there is an urgent need to develop an agent that can effectively treat cognitive impairment with fewer side effects.

The present inventors have made extensive research efforts to develop a substance that can treat cognitive disorders such as vascular dementia and memory impairment (memory impairment), and as a result, iris lactea extract shows an excellent treatment effect on cognitive disorders caused by brain damage. was confirmed, and the present invention was completed.

One object of the present invention is to provide a pharmaceutical composition for preventing or treating cognitive impairment containing Iris Lactea Extract (ILE) or a fraction thereof as an active ingredient.

Another object of the present invention is to provide a method for preventing or treating cognitive impairment comprising administering the pharmaceutical composition to an entity other than a human.

Another object of the present invention is to provide a food composition for preventing or improving cognitive impairment containing iris extract or a fraction thereof as an active ingredient.

Another object of the present invention is to provide a feed composition for preventing or improving cognitive impairment containing iris extract or a fraction thereof as an active ingredient.

This is explained in detail as follows. Meanwhile, each description and embodiment disclosed in the present invention may also be applied to each other description and embodiment. That is, all combinations of the various elements disclosed in the present invention fall within the scope of the present invention. Additionally, the scope of the present invention cannot be considered limited by the specific description described below.

In one aspect of the present invention, in order to achieve the above-described object, the present invention provides a pharmaceutical composition for preventing or treating cognitive impairment containing Iris Lactea Extract (ILE) or a fraction thereof as an active ingredient.

Additionally, the present invention provides a use of iris extract or fractions thereof to prevent or treat cognitive impairment.

The term "Iris Lactea" of the present invention is a perennial plant of the Iridaceae family and is native to temperate and subtropical Asia, where it inhabits various temperate regions at an altitude of 600 to 380 m. Flowers come in a variety of colors, from pale blue to purple, white or yellow. In oriental medicine, it is used as a contraceptive and for fever, jaundice, sore throat, vomiting and blisters, and the leaves are used as animal feed.

In the present invention, the iris flowers may be purchased and used commercially, or those collected or cultivated in nature may be used, but are not limited thereto.

The term "extract" in the present invention refers to the resulting liquid component obtained by immersing the desired substance in various solvents and then extracting it at room temperature or at a heated state for a certain period of time, and the solid component obtained by removing the solvent from the liquid component. it means. In addition, in addition to the above results, it can be comprehensively interpreted to include all dilutions of the results, concentrates thereof, crude preparations, purifications, etc. of the above results. Accordingly, the iris lacteal extract (ILE) provided by the present invention is an extract obtained by extraction treatment, a diluted or concentrated liquid of the extract, a dried product obtained by drying the extract, a crude product or purified product of the extract, or It can be interpreted to include extracts of all formulations that can be formed using the extract itself and the extract, including mixtures thereof.

The extraction method for the iris flower extract of the present invention is not particularly limited, and may be extracted according to a method commonly used in the art. Non-limiting examples of the extraction method include hot water extraction, ultrasonic extraction, filtration, and reflux extraction, which may be performed alone or by combining two or more methods.

The type of solvent used for the extraction is not particularly limited, and any solvent known in the art can be used. Non-limiting examples of the extraction solvent include water, alcohol, or a mixed solvent thereof, and these may be used alone or in a mixture of one or more types, and specifically alcohol may be used, mainly having 1 to 4 carbon atoms. Alcohol can be used.

Additionally, the extract may be prepared and used in dry powder form after extraction, but this is not limited.

Additionally, in the present invention, the extract may be a seed extract of iris.

The term "cognitive disorder" of the present invention may include conditions resulting from reduced cognitive function, diseases resulting therefrom, and all diseases whose symptoms are loss of reasoning ability, forgetting, learning disabilities, concentration problems, and loss of intelligence. This may include depression and other decreases in mental function. In the present invention, the cognitive disorder may specifically mean vascular cognitive disorder. The vascular cognitive impairment refers to all cognitive dysfunction caused by cerebrovascular disease, and may be one or more selected from the group consisting of vascular dementia, vascular mild cognitive impairment, stroke, cerebral infarction, cerebral hemorrhage, or cerebral vasculopathy. . The vascular dementia includes multi-infarct dementia, strategic infarct dementia, subcortical vascular dementia, hypoperfusion dementia, and hemorrhagic dementia. , hereditary vascular dementia, or mixed dementia of Alzheimer's disease and vascular dementia, but is not limited thereto.

The term "prevention" in the present invention refers to all actions that suppress or delay the onset of cognitive impairment by administering the pharmaceutical composition according to the present invention.

The term "treatment" of the present invention refers to any action in which symptoms of a subject suspected of or suffering from cognitive impairment are improved or beneficially changed by administration of the pharmaceutical composition.

The pharmaceutical composition of the present invention may contain 0.001 to 80% by weight of the extract, specifically 0.001 to 70%, and more specifically 0.001 to 60% by weight based on the total weight of the composition, but is not limited thereto.

In addition, the pharmaceutical composition may further include a pharmaceutically acceptable carrier, excipient, or diluent commonly used in the preparation of the pharmaceutical composition, and the carrier may include a non-naturally occurring carrier. there is. The carriers, excipients, and diluents include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, and microcrystalline. Examples include cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil.

In addition, the pharmaceutical composition can be manufactured into tablets, pills, powders, granules, capsules, suspensions, oral solutions, emulsions, syrups, sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and transdermal preparations according to conventional methods. It can be formulated and used in the form of an absorbent, gel, lotion, ointment, cream, patch, cataplasma, paste, spray, skin emulsion, skin suspension, transdermal delivery patch, drug-containing bandage, or suppository. Specifically, when formulating, it may be prepared using commonly used diluents or excipients such as fillers, weighting agents, binders, wetting agents, disintegrants, and surfactants. Solid preparations for oral administration include, but are not limited to, tablets, pills, powders, granules, capsules, etc. These solid preparations may be prepared by mixing at least one excipient, such as starch, calcium carbonate, sucrose, lactose, gelatin, etc. Additionally, in addition to simple excipients, lubricants such as magnesium stearate and talc may also be used. In addition to oral liquid and liquid paraffin, it can be prepared by adding various excipients, such as wetting agents, sweeteners, fragrances, and preservatives. Preparations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations and suppositories. Non-aqueous solvents and suspensions may include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate. As a base for suppositories, Withepsol, Macrogol, Tween 61, cacao, laurin, glycerogelatin, etc. can be used.

In the present invention, the treatment of cognitive impairment may include enhancing memory ability, normalizing memory-related factors, suppressing tau hyperphosphorylation, normalizing white matter damage, suppressing glial cell activity, and normalizing cholinergic neurons through a dementia-simulating animal model.

In a specific embodiment of the present invention, an animal model simulating dementia was produced and the effect of the extract on improving cognitive decline caused by brain damage was confirmed through an experiment of administering a seed extract of iris lactea to the model (Figure 3).

In addition, in one specific example, it was confirmed that the seed extract of iris lactea of the present invention increased and normalized memory-related factors in the hippocampus that were reduced by brain damage (FIGS. 4 and 5)

In addition, in one specific example, it was confirmed that the seed extract of iris lactea of the present invention normalized the expression of tau-related factors in the hippocampus hyperphosphorylated by brain damage (FIGS. 6 and 7).

In addition, in one specific example, it was confirmed that the seed extract of iris lactea of the present invention normalized white matter damage by effectively suppressing myelin myelin caused by brain damage (FIGS. 8 and 9).

In addition, in one specific example, it was confirmed that the seed extract of iris lactea of the present invention normalized hippocampal damage by effectively suppressing glial cells activated by brain damage (FIGS. 10 and 11).

In addition, in one specific example, it was confirmed that the seed extract of iris lactea of the present invention normalized cholinergic neurons in the basal forebrain by increasing the number of cholinergic neurons reduced by brain damage (FIGS. 12 and 13) .

Therefore, the seed extract of iris lactea provided by the present invention can be effectively used in the prevention or treatment of cognitive impairment.

Another aspect of the present invention for achieving the above object provides a method of treating cognitive impairment, including the step of administering the pharmaceutical composition to an individual other than a human suspected of having cognitive impairment.

At this time, the definitions of the terms “iris lactea”, “extract”, “prevention” and “treatment” are as described above.

As used herein, the term “administration” refers to the act of introducing a composition containing the extract into a subject by an appropriate method.

The term "individual" in the present invention refers to all animals such as rats, mice, and livestock, including humans, that have or may develop cognitive impairment. As a specific example, it may be a mammal, including humans.

The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. The term "pharmaceutically effective amount" means an amount sufficient to treat the disease with a reasonable benefit/risk ratio applicable to medical treatment, and the effective dose level is determined by the type and severity of the subject, age, sex, activity of the drug, It can be determined based on factors including sensitivity to the drug, time of administration, route of administration and excretion rate, duration of treatment, concurrently used drugs, and other factors well known in the field of medicine. For example, the iris extract can be administered at a dose of 0.01 to 5000 mg/kg per day, specifically 10 to 1000 mg/kg, and the administration may be administered once a day or divided several times.

The pharmaceutical composition may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with conventional therapeutic agents. And it can be administered single or multiple times. Considering all of the above factors, it is important to administer an amount that can achieve maximum effect with the minimum amount without side effects, and can be easily determined by a person skilled in the art.

In addition, the pharmaceutical composition can be administered orally or parenterally (e.g., intravenously, subcutaneously, intraperitoneally, or topically) depending on the desired method, and the dosage is determined by the patient's condition and weight, and the degree of the disease. , it varies depending on the drug form, administration route and time, but can be appropriately selected by a person skilled in the art.

Another aspect of the present invention for achieving the above object provides a food composition for preventing or improving cognitive impairment containing iris extract or a fraction thereof.

At this time, the definitions of the terms “iris lactea,” “extract,” and “prevention” are as described above.

As used herein, the term “improvement” refers to any action that results in at least a reduction in the severity of symptoms, for example, parameters associated with the condition being treated by administration of a composition comprising the extract.

As used herein, “food” refers to meat, sausages, bread, chocolate, candies, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, and alcoholic beverages. , vitamin complexes, health functional foods, and health foods, etc., and include all foods in the conventional sense.

The food composition of the present invention can be expected to have a high lung cancer improvement effect because it is derived from the daily ingestible porridge, and can therefore be very useful for health promotion purposes.

The above-mentioned health functional food (functional food) is the same term as food for special health use (FoSHU), and is a medicine processed to efficiently exhibit bioregulatory functions in addition to nutritional supply, with high medical effects. It means food. Here, ‘function’ means controlling nutrients for the structure and function of the human body or obtaining useful effects for health purposes, such as physiological effects. The food of the present invention can be manufactured by methods commonly used in the industry, and can be manufactured by adding raw materials and ingredients commonly added in the industry. Additionally, the food formulation can be manufactured without limitation as long as it is a formulation recognized as a food.

The food composition of the present invention can be manufactured in various types of formulations, and unlike general drugs, it is made from natural products and has the advantage of not having side effects that may occur when taking the drug for a long period of time. It is also highly portable, so it can be used as a raw material. The food of the invention can be consumed as an adjuvant to improve the effect of cognitive impairment.

The above-mentioned health food refers to food that has a more active health maintenance or promotion effect compared to general food, and health supplement food refers to food for the purpose of health supplementation. In some cases, the terms health functional food, health food, and health supplement are used interchangeably.

Specifically, the health functional food is a food prepared by adding the composition of the present invention to food materials such as beverages, teas, spices, gum, and confectionery, or by encapsulating, powdering, or suspending it, and consuming it may cause certain health problems. It means bringing about an effect, but unlike regular drugs, it has the advantage of not having any side effects that may occur when taking the drug for a long time since it is made from food.

The food composition may further include a physiologically acceptable carrier. The type of carrier is not particularly limited and any carrier commonly used in the art can be used.

Additionally, the food composition may contain additional ingredients that are commonly used in food compositions to improve smell, taste, vision, etc. For example, it may include vitamins A, C, D, E, B1, B2, B6, B12, niacin, biotin, folate, pantothenic acid, etc. Additionally, minerals such as zinc (Zn), iron (Fe), calcium (Ca), chromium (Cr), magnesium (Mg), manganese (Mn), copper (Cu), and chromium (Cr); and amino acids such as lysine, tryptophan, cysteine, and valine.

In addition, the food composition contains preservatives (potassium sorbate, sodium benzoate, salicylic acid, sodium dehydroacetate, etc.), disinfectants (bleaching powder, high bleaching powder, sodium hypochlorite, etc.), antioxidants (butylhydroxyanisole (BHA), butylhydroxide) roxitoluene (BHT), etc.), colorants (tar color, etc.), coloring agents (sodium nitrite, sodium nitrite, etc.), bleaching agents (sodium sulfite), seasonings (MSG monosodium glutamate, etc.), sweeteners (dulcine, cyclemate, saccharin) , sodium, etc.), flavorings (vanillin, lactones, etc.), leavening agents (alum, D-potassium hydrogen tartrate, etc.), strengtheners, emulsifiers, thickeners (grease), coating agents, gum base agents, anti-foam agents, solvents, improvers, etc. May contain food additives. The additives can be selected depending on the type of food and used in an appropriate amount.

As an example of the food composition of the present invention, it can be used as a health drink composition, and in this case, it can contain various flavoring agents or natural carbohydrates as additional ingredients, like ordinary drinks. The above-mentioned natural carbohydrates include monosaccharides such as glucose and fructose; Disaccharides such as maltose and sucrose; polysaccharides such as dextrins and cyclodextrins; It may be a sugar alcohol such as xylitol, sorbitol, or erythritol. Sweeteners include natural sweeteners such as thaumatin and stevia extract; Synthetic sweeteners such as saccharin and aspartame can be used. The ratio of the natural carbohydrate may be generally about 0.01 to 0.04 g, specifically about 0.02 to 0.03 g, per 100 ml of the health drink composition of the present invention.

In addition to the above, the health drink composition includes various nutrients, vitamins, electrolytes, flavors, colorants, pectic acid, salts of pectic acid, alginic acid, salts of alginic acid, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, It may contain alcohol or carbonating agent. Additionally, it may contain pulp for the production of natural fruit juice, fruit juice beverage, or vegetable beverage. These ingredients can be used independently or in combination.

The ratio of these additives is not very important, but is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the health drink composition of the present invention.

Another aspect of the present invention for achieving the above object provides a feed composition for preventing or improving cognitive impairment containing iris extract or a fraction thereof.

At this time, the definitions of the terms “iris lactea”, “extract”, “cognitive disorder”, “prevention” and “improvement” are as described above.

Since the iris extract according to the present invention shows an excellent therapeutic effect for cognitive disorders, it can be included in a feed composition for the purpose of preventing or improving vascular cognitive function, and since the feed composition can be consumed by animals on a daily basis, vascular cognitive impairment High effectiveness in preventing or improving function can be expected.

As used herein, the term "feed" means any natural or artificial diet, meal, etc., or a component of the meal, for or suitable for eating, ingestion, and digestion by animals.

The type of feed is not particularly limited, and feed commonly used in the art can be used. Non-limiting examples of the feed include plant feed such as grains, roots and fruits, food processing by-products, algae, fiber, pharmaceutical by-products, oils and fats, starches, gourds or grain by-products; Examples include animal feeds such as proteins, inorganic substances, fats and oils, mineral oils, oils and fats, single-cell proteins, zooplanktons or food. These may be used alone or in combination of two or more types.

The Iris Lactea Extract of the present invention exhibits an excellent normalizing effect and memory enhancing effect on brain damage such as the basal forebrain, white matter, and hippocampus, and thus causes cerebrovascular diseases such as vascular dementia and cerebral infarction and memory disorders caused by it. It can be useful in the treatment of cognitive disorders related to back.

Figure 1 is a diagram showing an experimental design to measure the effect of Iris Lactea Extract on object and location recognition memory.
Figure 2 is a diagram showing the inherent object detection behavior of a mouse, and the Y-axis represents the total time (object exploration time) during which the mouse approaches an object and explores the object during the training period. At this time, Sham+VEH (physiological saline, saline) is the normal control group as the simulation treatment group; UCCAO+VEH (saline) is a negative control group treated with saline in the brain injury group; UCCAO+ILE100 is an experimental group administered 100 mg/kg of iris extract to the brain-damaged group; UCCAO+ILE200 is an experimental group administered 200 mg/kg of iris extract to the brain-damaged group; UCCAO+ILE400 refers to the experimental group in which 400 mg/kg of iris extract was administered to the brain damage group, and the description of the above categories applies equally to the drawings below.
Figure 3 is a diagram showing the pharmacological effect of iris extract on the decline in object and location recognition memory in the brain damage group. The Y-axis represents a scale (recognition index) that quantifies and compares the search time for a new object compared to an existing object or a new location compared to an existing location.
Figure 4 is a diagram showing the normalizing effect of memory-related factors (PI3K-Akt-CREB-BDNF pathway) of iris extract in the hippocampus, showing the phosphorylation of PI3K, Akt, and CREB proteins and the Western blot results for BDNF protein.
Figure 5 is a graph quantifying Western blot bands for phospho-PI3K, phospho-Akt, phospho-CREB, and BDNF proteins according to Figure 4.
Figure 6 is a diagram showing the normalizing effect of tau-related factors (PI3K-Akt-GSK3β-tau) of iris extract in the hippocampus, showing Western blot results for phosphorylation of GSK3β and tau proteins.
Figure 7 is a graph quantifying Western blot bands for phospho-GSK3β (ser9), phospho-Tau (Thr212), and phospho-Tau (Ser199/202) proteins according to Figure 6.
Figure 8 is a diagram showing the effect of iris extract on normalizing demyelination of myelin in corpus callosum tissue within the white matter, showing the results of immunofluorescence staining for MBP protein.
Figure 9 is a graph quantifying the results of immunofluorescence staining for the MBP protein according to Figure 8.
Figure 10 is a diagram showing the effect of iris extract on suppressing gliosis in hippocampus tissue, showing the results of immunofluorescence staining for Iba-1 protein.
Figure 11 is a graph quantifying the results of immunofluorescence staining for the Iba-1 protein according to Figure 10.
Figure 12 is a diagram showing the normalizing effect of iris extract on cholinergic neurons in the basal forebrain (Substantia innominata-nucleus basalis magnocellularis) tissue, showing the results of immunohistochemical staining for ChAT protein.
Figure 13 is a graph quantifying the results of immunohistochemical staining for ChAT protein according to Figure 12.

Hereinafter, the present invention will be described in more detail through examples. These examples are for illustrating the present invention in more detail, and the scope of the present invention is not limited by these examples.

Example 1. Production of a dementia simulating animal model and establishment of an oral drug administration method

Example 1-1. Method for producing a dementia simulating animal model

To confirm the effect of iris extract on inhibiting the progression of vascular dementia or memory loss, an animal model simulating dementia was first created.

Specifically, the experimental animals used were 8-week-old C57BL/6 mice supplied by Orient Co., Ltd. (Jungwon-gu, Seongnam-si, Gyeonggi-do). Animals were acclimated for 7 days. The temperature of the animal room was 23±3℃, relative humidity 50±10%, lighting was on a 12-hour light/dark cycle (lights on at 7:00, lights off at 19:00), and the illumination level was 150~300 Lx. Adjustments were made to maintain consistent breeding environmental conditions. Additionally, water and solid feed (PMI nutrition, USA) were allowed to be freely consumed for 24 hours.

To create an animal model simulating vascular dementia, chronic cerebral hypoperfusion was induced by unilateral common carotid artery occlusion (hereinafter referred to as “brain injury (UCCAO)”) (Yosizaki et al. al., 2008). The mouse was anesthetized with anesthetic gas (isoflurane, 2.0%), laid flat on the operating table, and the skin was incised approximately 1.5 cm above the point where the upper limb and the midline meet to expose the unilateral common carotid artery without damaging the tissue. After separating the tissues and nerves attached to the exposed carotid artery, the carotid artery was ligated using nylon thread to cause cessation of blood supply, thereby producing brain damage-induced experimental animals. Afterwards, the incision area was closed with sutures and the patient was observed for about 20 minutes to see if he or she woke up from anesthesia and recovered.

Example 1-2. Iris extract oral administration method

After the procedure in Example 1-1, 14 days had passed and the iris extract was diluted with physiological saline and administered orally for 60 days. They were divided into a total of 5 groups listed in [Table 1] below, and the same volume of physiological saline (Saline) was orally administered to the control group. Oral administration was set at three concentrations: low concentration (100 mg/kg), medium concentration (200 mg/kg), and high concentration (400 mg/kg).

group Cognitive function and neurobiological verification
(number of animals) Normal control group Simulation group (Sham) + Vehicle (physiological saline solution) 12 Negative control group Brain injury group (UCCAo) + Vehicle (physiological saline solution) 14 experimental group Brain injury group (UCCAo) + iris extract 100 mg/kg 9 Brain injury group (UCCAo) + iris extract 200 mg/kg 11 Brain injury group (UCCAo) + iris extract 400 mg/kg 11

Example 2. Confirmation of memory enhancement effect of iris extract

In order to confirm the effect of iris extract on improving memory ability on a dementia simulating animal model, iris extract was administered according to the method of Example 1 above and then a novel object and location recognition test was performed.

Specifically, a white box (40 One day later, objects of the same size and shape were placed in the same box, and the mice were allowed to adapt to the two objects for 10 minutes (training). Another day later, one of the two objects was replaced with a new object and the mouse was placed so that it could approach both objects for 5 minutes (Novel object test). Finally, one day later, one of the two objects was moved to a new location, and the mouse was also allowed to approach both objects for 5 minutes (Novel location test).

Object and location recognition ability (recognition index) was compared by quantifying the degree of exploration (time) for a new object compared to an existing object or a new location compared to an existing location.

As a result, as can be seen in Figure 2, it was confirmed that there was no difference between the groups in the inherent behavior of detecting objects.

In addition, as can be seen in Figure 3, the group administered iris extract showed a higher ability to recognize new objects and locations compared to the group administered physiological saline, which was the negative control. In particular, the groups administered 200 and 400 mg/kg of iris extract were similar to the normal control group. It was confirmed that new objects and locations were recognized to this degree. Through these results, it was found that iris extract has the effect of improving cognitive decline caused by brain damage in mice, and can therefore be usefully used in the prevention and treatment of cognitive disorders such as memory impairment and dementia.

Example 3. Confirmation of the effect of iris extract on normalizing memory-related factors

In order to confirm the effect of normalizing memory-related factors on a dementia simulating animal model, the expression of memory-related factors in the hippocampus was measured after administering iris extract according to the method of Example 1 above.

Specifically, the hippocampus of mice that had completed the administration of iris extract and the cognitive test was extracted, and then PI3K (Phosphoinositide 3-kinase), phospho-PI3K, Akt (Protein kinase B), phospho-Akt, and CREB ( The expression levels of cAMP-response element binding protein), phospho-CREB, and BDNF (Brain-derived neurotrophic factor) proteins were analyzed. β-Actin was used as a loading control.

As a result, as can be seen in Figures 4 and 5, the expression levels of phospho-PI3K, phospho-Akt, phospho-CREB and BDNF proteins in the iris extract administered group were found to be significantly increased compared to the negative control group. In particular, it was confirmed that the groups administered 200 and 400 mg/kg of iris extract recovered to a similar extent as the normal control group. Through these results, it was found that iris extract increases and normalizes memory-related factors in the hippocampus that have been reduced by brain damage in mice, so it can be useful in the prevention and treatment of cognitive disorders such as memory impairment and dementia.

Example 4. Confirmation of tau hyperphosphorylation inhibition effect of iris extract

In order to confirm the effect of suppressing tau hyperphosphorylation in a dementia-simulating animal model, the expression of tau-related factors in the hippocampus was measured after administering iris extract according to the method of Example 1 above. Tau hyperphosphorylation is a substance that causes filament-shaped neurofibrillary tangles within the brain nerve cells (neurons) of patients with cognitive impairment, and is known to be a key phenomenon causing cognitive impairment.

Specifically, the hippocampus of mice that had completed the administration of iris extract and the cognitive test was extracted, and GSK3β (Glycogen synthase kinase 3 beta), phospho-ser9-GSK3β, Tau, phospho-Thr212-tau, and phospho- The expression level of Ser199/202-tau protein was analyzed.

As a result, as can be seen in Figures 6 and 7, it was found that the expression level of phospho-ser9-GSK3β protein, which is known to inhibit tau phosphorylation, was significantly increased in the iris extract administered group compared to the negative control group. In addition, the expression levels of phospho-Thr212-tau and phospho-Ser199/202-tau proteins, which represent phosphorylated tau, were confirmed to be significantly reduced, showing that iris extract effectively inhibits tau phosphorylation. In particular, it was confirmed that the group administered 400 mg/kg of iris extract recovered to a similar extent as the normal control group. Through these results, it was found that iris extract normalizes the expression of tau-related factors in the hippocampus hyperphosphorylated due to brain damage in mice, so it can be usefully used in the prevention and treatment of cognitive disorders such as memory impairment and dementia. .

Example 5. Confirmation of the effect of iris extract on normalizing white matter damage

In order to confirm the effect of normalizing white matter damage in a dementia simulating animal model, the degree of myelin demyelination in the corpus callosum was analyzed after administering iris extract according to the method of Example 1 above. Since myelin sheath plays a role in mediating signal transmission between nerve cells, the so-called demyelination phenomenon, in which the myelin sheath is peeled off, interrupts these signal transmissions, increasing the risk of cognitive impairment.

Specifically, mice that had completed administration of iris extract and cognitive tests were perfused and fixed with 4% paraformaldehyde, and then brain tissue was cut into 40 μm sections and subjected to immunofluorescence staining. The sections were reacted with normal goat serum and then stained using MBP (Myelin basic protein, 1:1000) as the primary antibody and goat anti-mouse (1:200) as the secondary antibody. To stain brain structures (counterstain), the cells were reacted with DAPI (4',6-diamidino-2-phenylindole, 1:2500) for 10 minutes.

As a result, as can be seen in Figures 8 and 9, the density of MBP protein in the corpus callosum was confirmed to be significantly increased in the iris extract administered group compared to the negative control group, confirming that the iris extract effectively inhibits demyelination of myelin. . In particular, it was confirmed that the groups administered 200 and 400 mg/kg of iris extract showed a similar effect to the normal control group. Through these results, it can be seen that iris extract normalizes white matter damage by effectively suppressing myelin demyelinated due to brain damage in mice, so it can be useful in the prevention and treatment of cognitive disorders such as memory impairment and dementia. there was.

Example 6. Confirmation of gliosis inhibition effect of iris extract

In order to confirm the effect of suppressing glial activity on a dementia-simulating animal model, the level of microglia expression in the hippocampus was analyzed after administering iris extract according to the method of Example 1 above. Microglial cells act as phagocytes for waste products in nervous tissue and play a role in removing degenerated neurons and foreign substances within the tissue. Therefore, an increase in microglial cells indicates that brain damage has occurred.

Specifically, mice that had completed administration of iris extract and cognitive tests were perfused and fixed with 4% paraformaldehyde, and then brain tissue was cut into 40 μm sections and subjected to immunofluorescence staining. The sections were reacted with normal goat serum and then stained using Iba-1 (ionized calcium-binding adapter molecule, 1:1000) as the primary antibody and donkey anti-rabbit (1:200) as the secondary antibody. To stain brain structures (counterstain), the cells were reacted with DAPI (4',6-diamidino-2-phenylindole, 1:2500) for 10 minutes.

As a result, as can be seen in Figures 10 and 11, the density of Iba-1 protein in the hippocampus was confirmed to be significantly reduced in the iris extract administered group compared to the negative control group, confirming that the iris extract effectively inhibits microglia. . In particular, it was confirmed that the groups administered 200 and 400 mg/kg of iris extract showed a similar effect to the normal control group. Through these results, it was found that iris extract normalizes hippocampal damage by effectively suppressing glial cells activated by brain damage in mice, so it can be useful in the prevention and treatment of cognitive disorders such as memory impairment and dementia.

Example 7. Confirmation of the cholinergic neuron normalizing effect of iris extract

In order to confirm the effect of normalizing cholinergic neurons on a dementia simulating animal model, the number of cholinergic neurons in the basal forebrain was analyzed after administering iris extract according to the method of Example 1 above. Cholinergic neurons play a key role in learning and memory by secreting neurotransmitters, and it is known that cholinergic neurons degenerate in patients with brain damage such as dementia.

Specifically, mice that had completed administration of iris extract and cognitive tests were perfused and fixed with 4% paraformaldehyde, and then brain tissue was cut into 40 μm sections and immunohistochemical staining was performed. The sections were reacted with normal horse serum and stained using ChAT (Choline acetyltransferase, 1:1000) as the primary antibody and rabbit anti-goat (1:500) as the secondary antibody.

As a result, as can be seen in Figures 12 and 13, it was confirmed that the number of ChAT-positive cells in the basal forebrain of the iris extract-administered group significantly increased compared to the negative control group. In particular, the iris extract 400 mg/kg administered group was compared to the normal control group. It was confirmed that a similar degree of effect was observed. Through these results, iris extract normalizes cholinergic neurons in the basal forebrain by increasing the number of cholinergic neurons reduced by brain damage in mice, so it can be useful in the prevention and treatment of cognitive disorders such as memory impairment and dementia. And it was found.

From the above description, those skilled in the art to which the present invention pertains will understand that the present invention can be implemented in other specific forms without changing its technical idea or essential features. In this regard, the embodiments described above should be understood in all respects as illustrative and not restrictive. The scope of the present invention should be construed as including the meaning and scope of the patent claims described below rather than the detailed description above, and all changes or modified forms derived from the equivalent concept thereof are included in the scope of the present invention.

Claims (10)

A pharmaceutical composition for preventing or treating cognitive impairment containing Iris lactea extract or a fraction thereof as an active ingredient,
A pharmaceutical composition wherein the part of the iris flower is a seed.
delete According to clause 1,
A pharmaceutical composition, wherein the cognitive disorder is vascular cognitive disorder. According to paragraph 3,
A pharmaceutical composition, wherein the vascular cognitive impairment is caused by cerebrovascular disease. According to clause 4,
The pharmaceutical composition, wherein the cerebrovascular disease is selected from the group consisting of vascular dementia, vascular mild cognitive impairment, stroke, cerebral infarction, cerebral hemorrhage, or cerebral aneurysm.
According to clause 1,
The extract is a pharmaceutical composition obtained by extraction with one selected from the group consisting of water, alcohols having 1 to 4 carbon atoms, and combinations thereof. According to clause 1,
The iris extract is a pharmaceutical composition that improves memory ability, normalizes memory-related factors, inhibits tau hyperphosphorylation, normalizes white matter damage, inhibits glial cell activity, or normalizes cholinergic neurons,
A pharmaceutical composition wherein the memory-related factors are phospho-PI3K, phospho-Akt, phospho-CREB, and BDNF proteins.
A method for preventing or treating cognitive impairment, comprising administering the pharmaceutical composition of any one of claims 1 and 3 to 6 to an entity other than a human.
A food composition for preventing or improving cognitive impairment containing Iris lactea extract or a fraction thereof as an active ingredient,
A food composition wherein the part of the iris flower is a seed.
A feed composition for preventing or improving cognitive impairment containing Iris lactea extract or a fraction thereof as an active ingredient,
A feed composition wherein the part of the iris flower is a seed.