WO2011002168A2 - Use of novel lactobacillus helveticus or a culture thereof for preventing or treating dementia or cognitive dysfunction - Google Patents

Abstract

The present invention relates to novel Lactobacillus helveticus, to a method for culturing same, and to the use thereof, and more particularly, to novel Lactobacillus helveticus which produces substances for inhibiting the production of amyloid-beta proteins, to pharmaceutical compositions containing the culture of Lactobacillus helveticus as an active ingredient for preventing or treating dementia or cognitive dysfunction, to the use thereof, to a method for preventing or treating dementia or cognitive dysfunction, and to a method for culturing same. The Lactobacillus helveticus or the culture thereof according to the present invention is effective in preventing or treating dementia, and in improving cognitive function or in preventing or treating cognitive dysfunction.

Description

 【Specification】

 [Name of invention]

 Use for the prevention or treatment of dementia or cognitive impairment of a novel Lactobacillus helvetica or its culture

 Technical Field

 <ι> This application claims the priority of Korean Patent Application No. 10-2009-0059051 filed on June 30, 2009, the entirety of which is a reference of the present application.

 <2> The present invention relates to the use of the novel Lactobacillus helbetticus or its culture for the prevention or treatment of dementia or cognitive dysfunction, and more particularly to produce a product that inhibits the production of beta amyloid protein. The present invention relates to a pharmaceutical composition for the prevention or treatment of dementia or cognitive dysfunction, comprising Lactobacillus helbetticus or its culture as an active ingredient, the use thereof, a method for preventing or treating dementia or cognitive dysfunction, and a culture method.

 <3>

 Background Art

 <4> According to the Korean Society of Dementia, dementia is a type of syndrome that causes people's intellectual and social activities to be lost due to a number of causal diseases, resulting in impairments in everyday life. In other words, dementia is a brain disease caused by damage and loss of nerve cells and is a disease in which severe disorders such as memory, concentration, language, and cognition progress gradually over a long period of time, leading to death. The causative diseases include Alzheimer's disease, Vascular dementia, Parkinson's disease, Lewy body dement ia, Huntington's disease, Creutzfeldt—Jacob's disease. Jacob disease, Pick's disease, and the like.

 <5>

The highest incidence of these causative diseases is Alzheimer's disease, which accounts for 50-70% of all dementia patients. Alzheimer's disease is classified into sporadic type and familial type according to the cause. More than 80% of cases are known to be sporadic and most occur after 65 years (Physiol Rev). 2001, 81: 741-66). Familiality accounts for less than 20% of the total, and the main causes of 100% mutations in genes such as amyloid precursor protein (APP), preseni 1 in l (PSl) and presenilin 2 (PS2). Degree It occurs quickly under 65 years old. Alzheimer's disease is a pathological feature in which senile plaques and neurofibrillary tangles accumulate outside and inside neurons, respectively. The senile plaques are generally spherical and are mainly found in the limbic system or neocortex, and peptides called beta amyloid (β-amyloid, Αβ) bind and deposit with each other to form nuclei and degenerated nerve fiber axons around them. Axons, dendrites, activated glial cells (microgl ia) and astrocytes are clusters of stones. Neuronal fiber concentrates are substances in neurons formed by binding to each other after abnormally large phosphorylation of a protein called tau is observed in various brain tissues of patients with Alzheimer's disease.

 The number of dementia patients, including Alzheimer's disease, continues to grow as the population ages. Alzheimer's disease is known to be around 26 million people worldwide as of 2008 and is expected to grow to more than 100 million by 2050 due to an increase in elderly population (Carlsson, J Alz Dis, 2008, 15: 327-). 338), according to the Korea Institute of Health and Human Services Research, 350,000 cases of dementia patients, or 8.3% of the elderly aged 65 or over, were steadily increasing with the increase of the elderly population. It is expected to be 10,000. Although the number of dementia patients is low in the total population, dementia is attracting attention because of the high socio-economic burden of caring for dementia patients. One year of treatment and care for a person with severe dementia costs more than 100 million won, which means that as of 2005, if all patients were treated with dementia, the cost would be about 35 trillion won. In addition to this enormous economic burden, the social life of families with dementia suffers a great deal. In general, to care for one dementia patient, two or more caregivers are needed, which leads to a limitation in the social life of the family and, furthermore, the loss of the economically active population. For these reasons, a number of studies for the treatment of dementia have been conducted at the national level in developed countries including the United States, and research on dementia has been actively conducted in Korea.

To date, research has shown that the main cause of Alzheimer's disease is AJ3, a constituent of the senile plaque, which causes nerve cell damage and causes Alzheimer's disease. Αβ is a peptide formed by the breakdown of ΑΡΡ, one of the main genes of familial Alzheimer's disease by β- and secretase, and consists of 40-42 amino acids (Nature, 1999, 402: 533-7). Dogs such as β- and secretase inhibitors and α-secretase activity promoters that inhibit the production of Aβ based on this mechanism of action Foot studies are underway, and other drugs that inhibit the accumulation of Aβ or promote the accumulation of Aβ are present in development (Nat Rev Neurosci, 2004, 5: 677–85). In recent years, efforts to develop A β as a vaccine have also been attempted by multinational pharmaceutical companies. However, the mechanism of action-based dementia treatment is a time-consuming step to commercialize, and the current marketed ones are Cognex, Donepezyl and Axellon (acetylcholine esterase inhibitors that have improved memory and short-term relief). Rivast igmin), reminil (Galantamine) and Memantine (Ebixa), the A receptor antagonist. In addition, blood circulation improvers such as Niserogoline, elcarnitine, and ginkgo biloba extract are indirectly confirmed to be effective for vascular dementia.

 As discussed above, current treatments for dementia mainly play a role in alleviating the symptoms of early dementia, but new drugs based on the mechanism of action and the mechanism of action are still required for development and release. It has failed in clinical practice because there is no therapeutic effect (J Alz Dis. 2008, 15: 327-338). For these reasons, there is an increasing demand for development of alternative substances, such as drugs or health functional foods, which can suppress the pathogenesis and prevent long-term effects due to the prevention of Alzheimer's disease.

 <ιο> Lactic acid bacteria have been used for a long time and it is recognized that Bulgarians who take Lactic Acid Bacteria fermented foods have a high rate of longevity. In addition, Mechinkov's research shows that harmful substances produced by intestinal harmful bacteria enter the blood and pose a threat to health, and many studies have been conducted on the function of lactic acid bacteria in attempts to replace the intestinal flora with beneficial lactic acid bacteria. Many beneficial functions such as intestinal action, anticancer effect, immune enhancing effect and anti-aging effect have been reported (Trends Biotechnol, 1997, 15; 270-274). Currently, lactic acid bacteria and fermented products are widely used as pharmaceuticals or health foods.

<ιι> In order to further expand the functions of these beneficial lactic acid bacteria and to use them for the purpose of preventing dementia, which has recently caused serious economic and social problems, studies on the protective effect of lactic acid bacteria on dementia have been carried out. Prior to conducting the study, the literatures and patents related to dementia of Lactobacillus were not found.

 <12>

 Detailed description of the invention

 [Technical problem]

<13> The incubators were investigating new substances for the treatment of dementia. Lactobacillus strain containing a substance that inhibits the production of beta amyloid by inhibiting the action of β-secretase, one of the causes of dementia, and its cultures are effective in preventing or treating dementia or cognitive dysfunction. Confirmed to complete the present invention.

 <14>

 It is therefore an object of the present invention to provide a Lactobacillus helvetticus which produces a product which inhibits the production of beta amyloid protein.

 <16>

 Another object of the present invention is to provide a use for the prevention or treatment of dementia of Lactobacillus helvetticus or a culture thereof.

 <18>

 Another object of the present invention is to provide a use for the prevention or treatment of cognitive dysfunction of Lactobacillus helvetticus or a culture thereof.

 <20>

 Another object of the present invention is to provide a food composition for improving cognitive function containing Lactobacillus helvetticus or a culture thereof as an active ingredient.

 <22>

 Another object of the present invention is to inoculate (a) inoculated with lactobacillus helveticus in a medium containing a dairy material; (b) removing the lactic acid bacteria from the culture;

(c) Lactobacillus helvetticus having a dementia prevention or therapeutic effect comprising the step of precipitating and separating the active ingredient by adding a solvent selected from the group consisting of acetone or an alcohol having 1 to 6 carbon atoms to the culture from which the lactic acid bacteria have been removed; It is to provide a method for producing a culture.

 <24>

 Technical Solution

 In order to achieve the above object, the present invention provides Lactobacillus helvetticus IDCC 3801 (Accession No .: KCTC-11332BP), which produces a product that inhibits the production of beta amyloid protein.

 <26>

In order to achieve another object of the present invention, the present invention provides a pharmaceutical composition for preventing or treating dementia, which contains Lactobacillus helvetticus or a culture thereof as an active ingredient. In order to achieve the another object of the present invention, the present invention provides a use for the production of a preventive or therapeutic agent for dementia of Lactobacillus helvetticus or its culture.

 In order to achieve another object of the present invention, the present invention provides a method for preventing or treating dementia, characterized by administering Lactobacillus helvetticus or a culture thereof to an individual in need thereof.

 <30>

 In order to achieve another object of the present invention, the present invention provides a pharmaceutical composition for preventing or treating cognitive dysfunction, containing Lactobacillus helvetticus or a culture thereof as an active ingredient.

 In order to achieve another object of the present invention, the present invention provides a use for the production of a preventive or therapeutic agent for cognitive dysfunction of Lactobacillus helvetica or its culture.

In order to achieve another object of the present invention, the present invention provides a method for preventing or treating cognitive dysfunction, characterized by administering Lactobacillus helvetticus or a culture thereof to an individual in need thereof in an effective amount.

 <34>

 In order to achieve the another object of the present invention, the present invention provides a food composition for improving cognitive function containing Lactobacillus helvetticus or a culture thereof as an active ingredient.

 <36>

 In order to achieve another object of the present invention, the present invention comprises the steps of: (a) inoculating the culture medium containing lactobacillus helvetikus incubation; (b) removing the lactic acid bacteria from the culture; (c) Lactobacillus helveti having a dementia preventive or therapeutic effect comprising the step of precipitating and separating the active ingredient by adding a solvent selected from the group consisting of acetone or an alcohol having 1 to 6 carbon atoms to which the lactic acid bacteria have been removed. Provided are methods of making Cous cultures.

 <38>

 Hereinafter, the present invention will be described in detail.

 <40>

 <4i> The Lactobacillus helvetticus IDCC 3801 strain of the present invention is a novel microorganism isolated from feces of breastfeeding infants.

The microorganism of the present invention is a strain belonging to the genus Lactobacillus (Lactobacillus) in molecular phylogenetic analysis based on the 16S rDNA nucleotide sequence Lactobacillus helvetticus A microorganism showing the highest molecular systemic relationship with the standard strain of Lactobacillus helveticus was identified as Lactobacillus helveticus IDCC 3801, and it is a Korean microorganism that is an international microorganism depositing institution for patent application. It was deposited on 8 May 2008 at the Conservation Center (www.kccm.or.kr) (accession number: CTC-11332BP).

 The inventors of the present invention have been found to inhibit the production of beta amyloid by inhibiting the action of β-secretase in the culture of Lactobacillus helvetticus, and this culture is used to prevent or treat dementia. Animal experiments confirmed that the effect is excellent. Such a dementia preventive or therapeutic effect of the Lactobacillus helvetticus culture of the present invention is an effect first known by the present invention.

 The lactobacillus helveticus according to the present invention can be cultured in large quantities by a conventional method of culturing Lactobacillus sp. As the culture medium, a medium consisting of a carbon source, a nitrogen source, vitamins and minerals, or whey medium using milk components, skim milk medium, and the like can be used. Available as the carbon source in the medium for the production of the culture of the present invention, at least one selected from the group consisting of glucose sucrose, maltose, fructose, lactose, xylose, galactose arabinose and combinations thereof, Preferably at least one selected from the group consisting of sucrose, fructose, glucose, galactose, arabinose and lactose, more preferably glucose. Available as the nitrogen source in the medium for the production of the culture of the present invention, yeast extract, soytone (peptone), beep extract (beef extract), tryptone, kathytone and selected from the group consisting of One or more, preferably one or more selected from the group consisting of yeast extract, peptone, tryptone and soyton, more preferably yeast extract or soyton.

 On the other hand, the present invention provides a pharmaceutical composition for preventing or treating dementia or cognitive impairment or a food composition for enhancing cognitive function, containing Lactobacillus helvetticus or a culture thereof as an active ingredient.

 <46>

The culture of the present invention can also be used by removing the cells with a culture medium cultured Lactobacillus helveticus. The Lactobacillus helvetticus can be any of the same kind of lactic acid bacteria, and the range includes all subspecies or variants of the Lactobacillus helveticus species. Preferably it may be Lactobacillus helveticus IDCC 3801 of accession number KCTC-11332BP. In order to prepare a culture medium of the present invention having an effect of inhibiting the production of beta amyloid, the lactobacillus helveticus of the present invention may be cultured into a culture medium containing dairy material. Dairy ingredients are ¾ components derived from mammals. The dairy ingredients are sterilized in a liquid state and can be used as they are or powdered by lyophilization. Dairy ingredients can be taken directly from animals or purchased from commercially available products. Examples of dairy ingredients include, but are not limited to, milk, goat's milk, salt-owned, camel oil or their skim milk, whole milk.

 <49>

 For example, a liquid medium to which glucose and milk are added may be used to prepare beta amyloid protein and lactobacillus helveticus strain culture that inhibits production. The culture can be carried out under conventional lactic acid culture conditions, for example,

Incubate at 371 for about 24 hours. Centrifugation or filtration may be performed to recover the cells in the culture and to separate the culture medium, and this step may be performed according to the needs of those skilled in the art. Concentrated cells can be preserved so as not to lose their activity by vibrating or kneading according to conventional methods. For example, it can be stored ^at minus 80 ^° C, containing glycerol, or lyophilized by suspending in sterile 10% skim milk. In addition, the microorganism according to the present invention may be improved or improved to have an equivalent activity or better activity by a conventional physicochemical mutation method known in the art.

<51> A culture that inhibits the production of beta amyloid protein of the present invention has high efficacy

It may be used after purification to make it usable as a high pharmaceutical composition. The purification process can be performed by any method as long as the activity of the culture of the present invention is maintained. Preferably, the active ingredient is included by precipitation by organic solvent after removing impurities through centrifugation, filtration and ultrafiltration. Fractions can be precipitated. The organic solvent used may be any one as long as it can precipitate the active ingredient in the culture of the present invention, preferably acetone or an alcohol having 6 to 6 carbon atoms. The amount of organic solvent used for precipitation may be any amount as long as the activity of the culture is maintained and may vary depending on the type of precipitation solvent, but preferably, if the precipitation solvent is acetone or alcohol, it is more than 1 times the volume of the culture. It can be added up to 4 times and mixed, and centrifuged to separate the active ingredient. Precipitation using an organic solvent may be performed once, or may be repeated one or more times with the same or different types of organic solvents or with the same or different volume ratio with the culture. When repeated, the target is the previous needle It may be a dissolved solution of the sediment from the whole process or a supernatant from a previous sedimentation process. The precipitated active ingredient may be dissolved in itself or in another solvent.

 <52>

On the other hand, the present invention comprises the steps of (a) inoculating the culture medium containing lactobacillus helvetikus cultured; (b) removing the lactic acid bacteria from the culture; (C) Lactobacillus helveti having a dementia prophylactic or therapeutic effect comprising the step of precipitating and separating the active ingredient by adding a solvent selected from the group consisting of acetone or alcohol having 1 ^' to 6 carbon atoms to remove the culture lactic acid bacteria Provided are methods of making Cous cultures.

<54>

 In step (a), lactobacillus helveticus is inoculated into a medium containing dairy material, followed by fermentation. The Lactobacillus helvetticus can be any of the same kind of lactic acid bacteria, the range of which includes all subspecies or variants of the Lactobacillus helveticus species. Preferably Lactobacillus helveticus IDCC 3801 of accession number KCTC-11332BP. Inoculation of Lactobacillus helveticus is performed by adding the appropriate amount of Lactobacillus helvetus to the culture medium so that Lactobacillus helvetus can fully grow under culture conditions. The inoculated amount of Lactobacillus helvetticus may be added to the culture medium by about 1 to 5% (v / v) of the pre-cultured Lactobacillus helvetticus culture. Cultivation and fermentation of Lactobacillus helveticus can be made according to the culture medium and the culture medium containing milk materials known in the art. This process can be easily adjusted and used by those skilled in the art according to the strain selected. These various methods are described in various documents (eg, James et al., Biochemical Engineering, Prentice-Hal 1 International Editions). Depending on the cell growth method, suspension culture and adhesion culture are divided into batch, fed-batch and continuous culture according to the culture method.

In addition, during the culture, antifoaming agents such as fatty acid polyglycol esters can be used to suppress bubble generation. In addition, in order to maintain an aerobic state, oxygen or an oxygen-containing gas (eg, air) may be injected. The temperature at the time of culture is usually 25 ^° C to 40 V, preferably 35 ° C to 40 ° C, can be incubated for 7 to 40 hours.

<57>

 In step (b), the lactic acid bacteria are removed from the fermented product fermented and cultured in the step (a). Centrifugation to separate and remove the lactic acid bacteria cells from the culture after the end of the culture

may be subjected to centrifugation or filtration. The vendor can carry out as needed. At this time, the removal of the cells shows that the lactic acid bacteria cells have been substantially removed through a method for removing conventional cells.

 <59>

 Step (C) may be precipitated and separated as described above.

 <61>

 On the other hand, the production method of the lactic acid bacteria culture of the present invention may further comprise the step of ultrafiltration (ultrafiltration) of a substance having a molecular weight of 5,000 or less for smooth separation and concentration of the active ingredient to obtain an ultrafiltration solution. have. Therefore, the production method of the present invention comprises the steps of: (a) inoculating Lactobacillus helvetikus in culture medium containing the milk raw material; (b) centrifuging the culture to remove lactic acid bacteria; (c) obtaining a substance having a molecular weight of 5,000 or less in the culture in which the lactic acid bacteria were removed; And (d) adding a solvent selected from the group consisting of acetone and alcohol having 1 to 6 carbon atoms to the obtained material to precipitate and isolate the active ingredient, thereby preventing lactobacillus hel. It may be a production recipe of Beticus culture.

 Steps (a), (b), and (d) can be performed as described above.

 In the above (c) step (b) to obtain a substance with a molecular weight of 5,000 or less in the culture from the lactic acid bacteria is removed through step (b). This may be preferably performed by a method of ultrafiltration using a filter that filters only a substance having a molecular weight of 5,000 or less, to obtain an ultrafiltration solution, but is not limited thereto.

 <65>

 In an embodiment of the present invention, nine types of lactic acid bacteria are isolated from fecal breastfeeding infants and precultured in MRS medium, using 37% SG medium containing 10% skim milk powder and 0.5> glucose. , And cultured for 25 hours. The culture was centrifuged at 15,000 rpm for 10 minutes to remove the strain by the method of taking the supernatant, and then ultrafiltration using a filter membrane to filter first with a 0.22um syringe filter and filter the molecular weight less than 5,000. The filtered solution was added with acetone or alcohol 1 times to the volume, and then centrifuged to secure a 1-fold precipitate, and to the remaining supernatant, 1-fold volume of the ultrafiltrate was added to secure a 2-fold precipitate in the same manner. . A four-fold precipitate was also obtained in the same way. The recovered precipitate was dissolved in 0.5% TFMtrifluoroacetic acid and used (see Examples 1-1 and 1-2).

<67> Recovered acetone and sediment (precipitate obtained by adding twice the acetone compared to ultrafiltrate) were injected with β-secretase and wild-type ΑΡΡ695 gene, and they were overexpressed. Lactobacillus strains having β-secretase activity inhibitory activity were selected by measuring the amount of ΑΡΡα and ΑΡΡβ in the culture and colonies after treatment with cell line BA-3 (see Example 1-3). . As a result, it was confirmed that the Lactobacillus Helveticus IDCC 3801 strain of the present invention has an excellent effect of inhibiting β-secretase activity, and the strain was identified as belonging to the Lactobacillus Helveticus species in May 2008. It was deposited on the 8 th of Korea microbial conservation center (Accession No .: KCTC-11332BP Example 1-4).

 <68>

 In another embodiment of the present invention, the conditions for obtaining a culture that inhibits the β_secretase activity of the present invention were investigated. As a result, it was confirmed that the Lactobacillus helveticus of the present invention has an effect of inhibiting β-secretase activity when cultured in SG medium containing skim milk powder rather than cultured in MRS which is a lactic acid bacteria culture medium. Precipitation is also possible using alcohol, which is harmless to human body compared to setone, and it was confirmed that β-secretase activity of the precipitate was best suppressed, especially when precipitated by adding 2 times the volume of the culture (Example) 2).

 <70>

 In another embodiment of the present invention, in order to determine whether the culture of the present invention is effective against Alzheimer's disease, which is related to dementia and related diseases, the memory test through the murine-maze and the test of Aβ40 in brain tissue using Alzheimer's disease animal model. The amount was confirmed. As a result, it was found that when the fermentation product of the present invention shows that the 44-% memory improvement effect, the Aβ40 was reduced by about 20%. As a result, it can be seen that the culture of Lactobacillus helvetticus of the present invention showed an effect of improving dementia in an animal model of Alzheimer's disease (see Example 4).

 <72>

 On the other hand, the present invention provides a pharmaceutical composition for the prevention or treatment of dementia containing Lactobacillus helvetticus or a culture thereof as an active ingredient. The Lactobacillus helveticus can be any of the same kind of lactic acid bacteria that inhibit the production of beta amyloid protein disclosed in the present invention, and the range includes all subspecies or variants of the Lactobacillus helveticus species. Preferably it may be Lactobacillus helveticus IDCC 3801 of accession number KCTC-11332BP.

 <74>

In the growth phase of dementia of the present invention, the normal intellectual level is maintained and acquired. Refers to a disorder in which cognitive impairment and personality change occur. Dementia causes the brain nerves to be destroyed by various causes, resulting in general disorders of mental function such as memory impairment, speech impairment, urinary incontinence, paranoid thinking, aphasia, and psychiatry such as depression, personality disorder, and aggression. It may be accompanied by symptoms. Diseases classified as dementia include, but are not limited to, Alzheimer's dementia, Vascular dementia, Parkinson's disease dementia, Lewy body dementia, Huntington's disease dementia, Creutzfeldt-Jakob dementia, and Pixar disease dementia includes dementia due to senile dementia and head injury.

 <76>

 The composition of the present invention inhibits the activity of β-secretase, promotes the activity of α-secretase, and ultimately has the activity of inhibiting the production of beta amyloid. Turned out for the first time.

 <78>

 In Alzheimer's disease, Αβ and tau proteins are the main causes of senile plaques and nerve fibers, which are formed inside and outside nerve cells, respectively, causing brain tissue atrophy and memory disorders. Aβ and tau are closely related to direct neuronal cell death, which suggests that activating the pathway in which Αβ is produced in ΑΡΡ leads to early symptoms and that the tau protein is involved in the subsequent acceleration of the disease. (Small and Duff, Neuron, 2008, 5: 591-598). Therefore, in order to prevent, improve symptoms or treat Alzheimer's disease, blocking of Aβ production pathway is considered to be important. Αβ was cleaved by β-secretase into a C-terminal fragment β (CTFp) containing ΑΡΡβ and Αβ amino acid sequences, and then the γ-secretase acted on the CTFP to react with ACIDCAPP intracellular domine. Since Aβ is formed, inhibiting the activity of β-secretase is one method of preventing, ameliorating or treating Alzheimer's disease. In addition, d-secretase decomposes ΑΡΡ competitively with β-secretase, inhibits the production of Αβ and α-secretase, which is one of the products degraded by α-secretase, induces ΑΡΡα production. Increasing the activity of is another way of preventing, ameliorating or treating Alzheimer's disease.

The composition of the present invention is an A [beta] production blocker, which effectively blocks the A [beta] production pathway and promotes the production of A [beta] A, which exhibits neuroprotective effects.

<81> In the present invention and one embodiment, the composition of the present invention promotes the production of ΑΡΡα,

Inhibition of the production of ΑΡΡβ was confirmed by Western blot through the cell experiment (see Example 2).

 <83>

<84> In another embodiment of the present invention was confirmed in this animal model of Alzheimer's composition of the present invention as the amount of beta amyloid point kit to reduce beta amyloid levels in the brain via the ^"animal testing (see Examples 4-2) . In addition, the composition of the present invention was confirmed to improve or treat memory and cognitive decline, which is one of the main symptoms of dementia in animal models of dementia, through a U-shaped maze test in dementia-induced rats to which the present invention and the composition were administered. (See Example 4-1).

 <85>

 As a result, the composition of the present invention effectively blocked the Aβ production pathway, promoted the production of AΑΡΡα showing a neuroprotective effect, and confirmed that it has an excellent effect on preventing, improving or treating dementia.

 <87>

 In addition, the present invention provides a pharmaceutical composition for preventing or treating cognitive dysfunction, containing Lactobacillus helvetticus or a culture thereof as an active ingredient.

The lactobacillus helveticus can be any of the same types of lactic acid bacteria that are efficacious in cognitive dysfunction symptoms, and the range includes all subspecies or strains of the Lactobacillus helveticus species. Preferably it may be Lactobacillus helveticus IDCC 3801 of accession number KCTC-11332BP.

 <90>

<9i> Cognitive impairment refers to mental and behavioral disorders due to brain disease, brain injury, addiction, endocrine disorders, metabolic or nutritional abnormal it ies, and drugs. covering due to mental ^illness, and is as a kind of brain disorders, as well as forgetfulness (anmesia) corresponding to the temporary short-term memory impairment, depending on the extent appearing in dementia (dementi a) causing overall disorder of orientation and judgment of symptoms . .

 <92>

Cognitive deficits or cognitive impairments include cognitive function or cognitive domains, such as work memory, attention and vigilance, language learning and memory, visual learning and memory, reasoning and problem solving such as practice. Deterioration in function, processing speed or social awareness Included. In particular, cognitive deficits or cognitive impairments may include attention deficit, disrupted thinking, slow thinking, difficulty understanding, poor concentration, problem solving disorder, poor memory, difficulty in expressing thoughts and / or thoughts, feelings and Difficulties in incorporating behavior or the disappearance of irrelevant thoughts. The terms "cognitive defect" and "cognitive impairment" are intended to refer to the same and are used interchangeably.

 Diseases classified as cognitive impairment include, but are not limited to, cognitive deficits associated with schizophrenia, cognitive deficiencies associated with age-induced memory impairment or mental disorders, cognitive deficiencies associated with diabetes, and post-stroke recognition Defects, memory deficits associated with hypoxia, cognitive and attention deficits associated with senile dementia, memory problems associated with mild cognitive impairment, impaired cognitive function associated with dementia, impaired cognitive function associated with Alzheimer's disease, impaired cognitive function associated with Parkinson's disease, vasculature Impaired cognitive function associated with dementia, cognitive problems associated with brain tumors, Pick's disease, cognitive deficits due to autism, cognitive deficits after electroconvulsive therapy, cognitive deficits associated with traumatic brain injury, memory loss disorders and delirium Can be.

 <95>

 In one embodiment of the present invention confirmed that the composition of the present invention exhibits excellent cognitive improvement effect in the underwater maze and manual avoidance test in the scopolamine induced memory impaired animal model administered the composition of the present invention (See Example 5).

 <97>

 The pharmaceutical composition according to the present invention may contain Lactobacillus helveticus or its culture alone or may further contain one or more pharmaceutically acceptable carriers, excipients or diluents. The pharmaceutical composition according to the present invention may have a composition comprising 0.001 to 99.999% by weight of Lactobacillus helvetticus or its culture and a balance of the carrier.

 <99>

 <100> The above pharmaceutically acceptable means `` physiologically acceptable '' when administered to humans, does not inhibit the action of the active ingredient and usually does not cause allergic reactions such as gastrointestinal disorders, dizziness or similar reactions. Toxic composition.

Pharmaceutically acceptable carriers may further include, for example, carriers for oral administration or carriers for parenteral administration. Carriers for oral administration may include lactose, starch, cellulose derivatives, magnesium stearate and stearic acid. In addition, peptides It may include a variety of drug ^' transfers used for oral administration of the formulation. In addition, carriers for parenteral administration may include water, suitable oils, saline, aqueous glucose and glycols, and the like, and may further include stabilizers and preservatives. Suitable stabilizers include antioxidants such as sodium hydrogen sulfite, sodium sulfite or ascorbic acid. Suitable preservatives include benzalkonium chloride, methyl- or propyl-paraben and chlorobutane. The pharmaceutical composition of the present invention may further include a lubricant, a humectant, a sweetener, a flavoring agent, an emulsifier, a suspension agent, and the like, in addition to the above components. Other pharmaceutically acceptable carriers may be referred to those described in the following literature (Remington * s Pharmaceutical Sciences, 19th ed., Mack Publishing Company, East on, PA, 1995).

<102> A pharmaceutical composition comprising Lactobacillus helvetticus or a culture thereof according to the present invention as an active ingredient may include a pharmaceutically effective amount of Lactobacillus helvetticus or a culture thereof alone or one or more pharmaceuticals. It may further include a carrier that is acceptable. The pharmaceutically effective amount '' refers to an amount that exhibits more reaction than the negative control, and preferably an amount sufficient to treat or prevent dementia or cognitive dysfunction. At this time, the dementia is Alzheimer's disease (Alzheimer's disease) dementia, vascular dementia (Vascular dementia), Parkinson's disease (Parkinson 's disease) dementia, ^ruyiche' dementia (Lewy body dement ia), chorea (Hunt ington 's disease ) Dementia, Creutzfeldt-Jacob disease dementia or Pick's disease dementia, which may be cognitive deficits associated with schizophrenia, age-induced memory impairment, or Cognitive defects associated with mental disorders, Cognitive defects associated with diabetes, Cognitive defects associated with post-adolescence, Memory defects associated with hypoxia, Cognitive and attention deficits associated with senile dementia, Memory problems associated with mild cognitive impairment, Impaired cognitions associated with dementia Function, impaired cognitive function associated with Alzheimer's disease, impaired cognitive function associated with Parkinson's disease, impaired cognitive function associated with vascular dementia, brain tumor It may be related cognitive problems, Pick's disease, cognitive deficits due to autism, cognitive deficits after electroconvulsion therapy, cognitive deficits associated with traumatic brain injury, amnesia, or delirium.

 <103>

The composition of the present invention can be administered to any mammal, including humans. For example, it can be administered orally or parenterally. Parenteral administration methods include, but are not limited to, intravenous, intramuscular, intraarterial, intramedullary, intradural, intracardiac, It may be transdermal, incised, intraperitoneal, intranasal, intestinal, topical, sublingual or rectal.

 <105>

 The pharmaceutical composition of the present invention may be formulated into a preparation for oral or parenteral administration according to the route of administration as described above.

 <107>

 In the case of preparations for oral administration, the composition of the present invention may be a powder, granules, tablets, pills, sugar tablets, capsules, solutions, gels, syrups, slurries, suspensions, etc. Can be formulated. For example, oral formulations can be obtained by tablets or dragees by combining the active ingredient with a solid brother and then grinding it, adding the appropriate adjuvant and processing it into a granular mixture. Examples of suitable excipients include sugars, corn starch, wheat starch, rice starch and potato starch, including lactose, dextrose, sucrose, solbi, manny, xili, erysri and maltitol. Fillers such as starch, cellulose, methyl salose, sodium carboxymethyl cellulose and hydroxypropylmethyl-cellulose, including cellulose, gelatin, polyvinylpyridone and the like. In some cases, crosslinked polyvinylpyrrolidone, agar, alginic acid or sodium alginate may be added as a disintegrant. Furthermore, the pharmaceutical composition of the present invention may further include anticoagulants, lubricants, wetting agents, fragrances, emulsifiers and preservatives.

 <109>

 Formulations for parenteral administration may be formulated in the form of injections, creams, lotions, external ointments, oils, moisturizers, gels, aerosols and nasal inhalants in the art. These formulations are statements that are commonly known prescriptions for all pharmaceutical chemistries.

¾ (Remington's Pharmaceutical Sciences, 19th ed., Mack Publishing Company, East on, PA, 1995).

<ιιι> The total effective amount of the pharmaceutical composition of the present invention may be administered to a patient in a single dose, and by a long-term fractional treatment protocol in multiple doses. May be administered. The pharmaceutical composition of the present invention may vary the content of the active ingredient depending on the extent of the disease. Preferably the preferred total dose of the composition of the present invention may be about 0.01 to 500 mg, most preferably 0.1 // g to 100 mg per kg body weight per day. However, the dose of the composition may be determined by considering various factors such as the age, weight, health condition, sex, severity of disease, diet and excretion rate, as well as the route of administration and the frequency of treatment of the pharmaceutical composition. In view of this, a person of ordinary skill in the art would be able to determine an appropriate effective dosage for the particular use of the composition as a prophylactic or therapeutic agent for dementia or cognitive dysfunction. will be. The pharmaceutical composition according to the present invention is not particularly limited to its formulation, route of administration and method of administration as long as the effect of the present invention is shown.

 <112>

 On the other hand, the present invention provides a use for the production of a preventive or therapeutic agent for dementia of Lactobacillus helvetticus or its culture.

The present invention also provides a use for the production of a preventive or therapeutic agent for cognitive dysfunction of Lactobacillus helvetticus or its culture.

 <115>

 The present invention also provides a method for preventing or treating dementia, characterized by administering Lactobacillus helvetica or its culture in an effective amount to a subject in need thereof.

 The present invention also provides a method for preventing or treating cognitive dysfunction, characterized by administering Lactobacillus helvetticus or its culture in an effective amount to a subject in need thereof.

 <118>

 The lactobacillus helvetica or the culture thereof of the present invention may be administered in an effective amount via various routes including oral, transdermal, subcutaneous, intravenous or intramuscular. From above

An effective amount is an amount that, when administered to a patient, represents the therapeutic effect of dementia or cognitive dysfunction.

 The subject may be an animal, preferably a mammal, particularly an animal including a human, or may be a cell, tissue, organ or the like derived from the animal. The subject may be a patient in need of treatment.

 The lactobacillus helvetticus or the culture thereof of the present invention may be administered as it is or prepared in various formulations as described above, and preferably, a desired effect, ie dementia or cognitive function. It can be administered until the therapeutic effect of the disorder is derived.

 <122>

The present invention also provides a food composition for enhancing cognitive function containing Lactobacillus helvetica or its culture as an active ingredient. The Lactobacillus helveticus may be any of the same types of lactic acid bacteria that are effective in the symptoms of cognitive dysfunction, and the range includes all subspecies or variants of the Lactobacillus helveticus species. Preferably it may be Lactobacillus helveticus IDCC 3801 of accession number KCTC-11332BP.

 <125>

 <126> The food composition is a functional food (functional food), nutritional supplements

 It includes all forms such as nutritional supplements, health foods and food additives. Food compositions of this type can be prepared in various forms according to conventional methods known in the art.

 <127>

 For example, the health food may be prepared by drinking the fermented lactic acid bacteria of the present invention in the form of tea, juice and drink, or may be ingested by granulation, encapsulation and powdering. In addition, the lactic acid bacterium fermentation product of the present invention may be mixed with other active ingredients to prepare a composition.

 Functional foods also include beverages (including alcoholic beverages), fruits and processed foods (eg canned fruit, canned food, jams, marmalade, etc.), fish, meat and processed foods (eg ham, Sausages, corned beef, etc., breads and noodles (e.g. udon noodles, soba noodles, ramen noodles, spaghetti macaroni, etc.), fruit juices, various drinks, cookies, candy, dairy products (e.g. butter, cheese), edible water It may be prepared by adding the fermented lactic acid bacteria of the present invention to margarine, vegetable protein, retort food, frozen food, various seasonings (eg, miso, soy sauce, sauce, etc.).

In addition, in order to use the lactic acid bacteria fermented product of the present invention in the form of a food additive, it can be prepared in powder or concentrate form.

 <131>

 The preferred content of the present invention and lactic acid bacteria fermentation product in the food composition of the present invention may include about 0.01 to 10% by weight based on the total weight of the food.

 <133>

 Advantageous Effects

Therefore, the pharmaceutical composition containing Lactobacillus helvetticus of the present invention or a culture thereof as an active ingredient, the use thereof, and a method for preventing or treating dementia or cognitive dysfunction may be used for the prevention or treatment of dementia or cognitive dysfunction. effective. In particular, the composition of the present invention is excellent in promoting the production of ΑΡΡα with a neuroprotective effect and suppression of the production of beta amyloid, preventing the dementia or cognitive dysfunction such as Alzheimer's disease Or has excellent efficacy in treatment. In addition, the culture production method of the present invention is effective for the production of a culture having an excellent effect in the prevention or treatment of dementia or cognitive dysfunction of the present invention using the strain of the present invention. Therefore, the pharmaceutical composition containing Lactobacillus ¾Beticus or a culture thereof as an active ingredient, the use thereof, a method for preventing or treating dementia or cognitive dysfunction, or a method for preparing the culture may cause dementia or cognitive dysfunction. It can be used in various fields such as the manufacture of prophylactic or therapeutic medicines for people of age or prone age.

 <135>

 [Brief Description of Drawings]

 1 is a result of confirming the β-secretase inhibitory effect of IDCC 3801 lactic acid bacteria culture by Western blot (Media: ΑΡΡα / β in the culture cultured BA-3 cell line using only culture medium without administration of precipitate IDCC 3801: band of ΑΡΡα / β in the culture cultured ΒΑ-3 cell line by administering the acetone precipitate of the culture of IDCC 38 to the culture medium).

 FIG. 2 is a result of comparing the active ingredient precipitation according to the volume ratio of the culture medium and the precipitation organic solvent by Western blot (Media: Culture cultured BA-3 cell line using only culture medium without administration of precipitate) Band of ΑΡΡα / β in cell lysate, IDCC 3801: culture in which BA-3 cell line was cultured by administering ethanol sediment of the culture of IDCC 3801 to the culture medium / band of ΑΡΡα / β in cell lysate).

 <! 38> FIG. 3 is a graph showing a comparison of spontaneous alternating behavior (%) in order to determine the effect of improving memory in dementia model mice of IDCC 3801 lactobacillus cultures through the labyrinth maze test of dementia mice. : Mice administered SG medium instead of IDCC 3801 precipitate; IDCC 3801: Mice administered IDCC 3801 precipitate; Initial: Measurements when IDCC 3801 culture was administered to dementia model mice by oral administration; Measured at the end of the 8-week oral administration period.

 4 is a graph showing the change in the amount of beta amyloid in the brain according to the administration of IDCC 3801 lactic acid bacteria culture in the dementia model mice using ELISA ¾ (medium: mice administered SG medium instead of IDCC 3801 precipitate). IDCC 3801: mice administered IDCC 3801 precipitate).

 FIG. 5 shows the underwater maze test of scopolamine induced memory injury model mice.

IDCC3801 This is a graph comparing the time taken to find the platform for the memory improvement effect of the lactic acid bacteria culture (Escape latency time (s): Time taken to take (sec) Saline + saline: treated with saline instead of drug for 2 weeks and then treated with saline instead of scopolamine; Saline + scopolamine: scopolamine treated group after 2 weeks of physiological saline instead of drug; Donepezil + Scopolamine: Scopolamine treated group after administration of Donepezil for 2 weeks; Medium + scopolamine: Scopleamine treated group after administration of medium alcohol precipitate for 2 weeks; Lactic acid bacteria + scopolamine: scopolamine treated group after administration of lactic acid bacteria alcohol precipitate for 2 weeks). <i4i>. FIG. 6 is a graph showing the results of measuring the time taken to enter the dark room to examine the memory improvement effect of the IDCC3801 lactic acid bacteria culture in the passive avoidance test of scopolamine induced memory damage model mice (Retention Latency time (s) : Time taken to enter dark room (seconds); Sal + Sal: Group treated with physiological saline instead of scopolamine for 2 weeks after saline instead of drug; Sal + Sco: Scopolamine treated with physiological saline for 2 weeks instead of drug Group; Do + Sco: Scopolamine treated group after administration of Donepezil for 2 weeks; M + Sco: Scopolamine treated group after administration of medium spirit precipitate for 2 weeks; 3801 + Sco: Scopolamine after administration of lactic acid bacteria alcohol precipitate for 2 weeks Treatment group).

 <142>

 [Form for implementation of invention]

 Hereinafter, the present invention will be described in detail by way of examples.

 However, the following examples are merely to illustrate the invention's content is not limited to the following examples.

 <145>

 <146> <Example 1>

 <147> 3-secretase activity inhibited lactic acid bacteria screening

 <148>

 <1-1> Preparation method of fermented product sample of lactic acid bacteria

 <150> Collect feces from breastfeeding infants within 2 weeks of birth at a postpartum cooking center in Gyeonggi-do and smear them on an LBS flat medium (containing pH 4.0, 0.3 bile, LBS agar, Beet on, Dickson and Company, USA). A total of nine kinds of lactic acid bacteria were isolated and fermented products cultured with these strains were examined for their ΑΡΡβ inhibitory activity.

 <151>

 <152> <1-2> Sample Preparation of Lactic Acid Bacteria Fermented Products

The lactic acid strain isolated in Example <1-1> was subcultured or precultured in MRS medium (de Man-Rogosa-Sharpe media). The preculture was inoculated with SG (10% skim milk powder, 0.5% glucose, pH 6.8) medium or milk (3% (v / v)) medium containing skim milk powder at 37 ^° C. The culture was obtained by the method of standing culture for 24 hours. After cultivation in SG medium or milk, the culture state showed curd formation like lactic acid bacteria fermented milk and separated from whey. The medium to which 3% MRS medium was added was maintained under the same conditions and used as a control. Each culture was centrifuged at 15,000 rpm for 10 minutes to recover the supernatant, which was first filtered with a 0.22 πι syringe filer. Next, ultrafiltration was performed with a filter membrane for filtering molecular weight of 5,000 or less. Then, acetone or alcohol was added to the ultrafiltrate first, and then mixed 1 times, followed by centrifugation at 15,000 rpm for 10 minutes to obtain a 1x precipitate, and to the remaining supernatant, 1x volume of the ultrafiltrate was added again. A double precipitate was obtained by the method. A four-fold precipitate was also secured in the same way. Each precipitate was recovered, dried, and weighed, and then suspended in an appropriate concentration in 0.5% TFACtrifluoroacetic acid) to be used for the test.

 <154>

 <155> <1-3> Screening of lactic acid bacteria inhibiting β-secretase activity

 Acetone precipitates of nine strain cultures obtained in Example <1-2> (vs. ultrafiltration

 Lactobacillus having the inhibitory effect of β-secretase activity was selected using the precipitate obtained by adding twice the acelon). As a control, only a precipitate of acetone precipitated in the same manner as the SG medium culture medium not inoculated with lactic acid bacteria was used.

 In order to investigate the effects of lactic acid bacteria fermentation product samples on the metabolism of amyloid precursor protein (hereinafter called APP), the β-secretase and wild-type ΑΡΡ695 genes were injected to express the overexpressed ΗΕΚ293 cell line (hereinafter, ΒΑ-3, Yeon et al, Peptides, 2007, 28: 838-844).

 <i58> BA-3 Overexpression of β-Secretase Produces ΑΡΡβ and Increases β-Secretase

 This is useful for searching for active ingredients that directly or indirectly affect activity. ΒΑ-3 was incubated for 1 day by dispensing 6-well plate using DMEM containing 10% FBS at a density of 80% or more. After removing the medium and adding 0.9 ml of fetal bovine serum (FBS) free of fetal bovine serum (FBS) warmed to 37 ° C., lOOug / ml of the sample of acetone or alcohol precipitate of 0.1 ml free DM l suspended in Example <1-2>. Treatment was carried out at concentration and incubated for 24 hours. 0.2 ml of each medium treated with each sample was subjected to electrophoresis on 8% SDS—PAGE, followed by Western blot using MAB1560 (monoclonal, CHEMICON, USA) and Rb53 (polyclonal, Cephalon, USA) antibodies, antibodies of ΑΡΡα and ΑΡΡβ. ΑΡΡα and β in the culture were measured.

In order to identify ΑΡΡα and ΑΡΡβ in the cells, the cells are collected, suspended in phosphate saturated saline (PBS), centrifuged to remove supernatant, and protease inhibitors are included. Lysis buffer (lysis buffer; 50 mM Tris-HCl, pH7.6, 180 mM NaCl, 2 mM

EDTA, 1% Triton X-100) was added and left on ice for 30 minutes. Sonication was briefly performed, and the supernatant was recovered by centrifugation at 15,000 rpm for 10 minutes at 4 ° C., the protein was quantified, and Western blot was performed using MAB1560 and Rb53 antibodies in the same manner as described above.

 <160>

 <i6i> As a result, the presence of lactic acid bacteria inhibiting the production of ΑΡΡβ than the control group

 IDCC 3801 was shown to inhibit the production of ΑΡΡβ.

 <162>

 In addition to lactic acid bacteria, similar results were observed in Bacillus subtilis and E. coli cultures, but they did not inhibit ΑΡΡβ and production.

<164>

<165> [Table 1] ^'

 <i66> Inhibitory Effect on the ΑΡΡβ Production of SG Cultured Lactic Acid Bacteria.

 <167>

 <I68> <l-4> Identification of Selected Lactic Acid Bacteria

 In order to identify the IDCC 3801 strain, the lactic acid bacterium having the highest inhibitory effect on the ΑΡΡβ production, a phylogenetic analysis was performed by comparing 16S rRNA gene sequences.

<170>^'blast similarity research programs (Blast similarity search program:

 The nucleotide sequence of 16S rRNA of IDCC 3801 strain and 16CC rRNA of ATCC 15009, Lactobacillus helveticus standard strain, were compared using the National Institute of Biotechnology Informat ion. As a result, it was confirmed that the homology with 98.9%.

 <i7i> Also 9 genus and 84 Lactobacillus species registered on GenBank

(Species) and multiple sequence alignments with 16S rRNA gene sequences of 87 strains of Ashdophyllus strains were performed to determine the position in the phylogenic tree. Multiple sequence alignment program program) uses ClustalX. The 16S rRNA gene of strain IDCC 3801. on the phylogenetic tree was located closest to Lactobacillus helvetikus. In addition, homology with 10-week 16S rRNA gene sequences registered with Lactobacillus helveticus, such as AY369116, AM113779, DQ 123580, AB362629, EF536362, EU377824, FJ 172346, EU420177, FJ459815, GQ131286, showed a high identity of 99.1% on average. It was. As a result of molecular biological identification of the above and the split IDCC 3801 strain, the strain was classified as Lactobacillus helvetticus. Therefore, the strain with excellent probiotic properties such as acid resistance and bile acid resistance and high inhibitory effect of ΑΡΡβ production was named as Lactobacillus helveticus IDCC 38, and the Korean Microbial Conservation Center (www.kccm.or.kr) Deposited on 8 May (Accession Number: KCTC-11332BP).

 <173>

 <Example 2>

 Inhibitory Effect of Lactobacillus Beticus IDCC38 () 1 on B-Secretase Activity and α-Scretase Activity

 <176

 The inhibitory effect of ΑΡΡβ production on Lactobacillus helvetticus IDCC 3801 strain cultures was tested in acetone precipitated samples. It was confirmed that the acetone precipitated sample of the ultrafiltrate of the Lactobacillus Helveticus IDCC 3801 culture filtrate obtained by culturing in SG also showed the inhibitory effect of ΑΡΡβ production even at 25ug / ml (see FIG. 1).

 <178>

 Acetone precipitates in SG culture filtrates were difficult to be applied as pharmaceuticals due to solvents, so LTC was tested for Lactobacillus helvetticus IDCC 3801 strains.

 <180> Treat the ethanol precipitate of the Lactobacillus helvetticus IDCC3801 SG culture filtrate obtained in Example <1-2 at a concentration of lOOug / ml to inhibit the production of ΑΡΡβ by the method of Example <1-3> ， ΑΡΡα It was investigated whether to promote the production of. As a control, only precipitated SG medium culture medium without the inoculation of lactic acid bacteria was used in the same manner.

<ΐ8ΐ> As a result, when treating the ethanol sediment of Lactobacillus Helveticus IDCC 3801 strain culture solution, the sediment precipitated at 4 times the alcoholic beverage compared with the culture filtrate. Unlike 100% inhibitory effect was confirmed. Thus, twice the alcoholic precipitates had high inhibitory activity. What is shown means that the active ingredient in the culture product of Lactobacillus helvetticus IDCC 3801 strain is specifically precipitated and concentrated by two-fold alcohol treatment. In addition, it was confirmed that the ethanol sediment samples of Lactobacillus helvetticus IDCC 3801 strain increased the accumulation of ΑΡΡα in cells. (See Figure 2).

 <182>

 <183> In conclusion, it was confirmed that the active ingredient of Lactobacillus helvetticus IDCC 3801 alcohol sediment selectively inhibits the production of ΑΡΡβ but has the property of inducing the accumulation of ΑΡΡα in cells.

 <184>

 <185> <Example 3>

 <186> Toxicity Test of Alcohol Sediment in Lactobacillus ¾Beticus IDCC 3801 SG Culture Solution

<187>

 A single dose toxicity test was performed to determine the toxicity induced by oral administration of Lactobacillus helvetticus IDCC 3801 SG culture. At 6 weeks of age, five male ICR mice were observed orally for 7 days after a single oral dose of 3,000 mg / kg.

 As a result, the toxicity test mice did not die and no long-term abnormalities were found, indicating no toxicity (results not shown).

 <190>

 <191> <Example 4>

 <i92> Chide Improvement Efficacy Test of Alcohol Sediment in In vivo Lactobacillus Beticus IDCC 3801 SG Culture Solution

 <193>

 <194> <4-1> Y-Maze Test

 <| 95> Algheimer's animal model Tg2576 (Swedish mutant APP overexpression mouse, 4,

 9 months, female) was evaluated for the efficacy of ultrafiltration of Lactobacillus helvetticus IDCC 3801 SG culture ultrafiltration liquor using medium ultrafiltration liquor sediment as a control. For administration, each sample was suspended in 1% carboxymethyl cellulose (CMC), orally administered at a dose of 500 mg / kg 3 times a week for 8 weeks.

 <196>

The test extended three arms to form the letter Y and each branch The length was 25c [n, height 14cm, width 5cm and Y-shaped maze measuring device located at the same angle. At the end of a passage in the Y-shaped labyrinth, the head of the animal was pointed freely for eight minutes. It was recognized as an arm entry by deciding the entry into the passage to the hind paw of dementia rats, and the actual alteration when passing through three passages consecutively. One point was given and the spontaneous alteration behavior (%) was calculated by calculating the actual number of crossovers as a percentage of the maximum possible crossovers (ie, the total number of crossovers minus two).

 <198>

 Y—The maze was performed once before sample processing and once after sample processing to compare the effects of pre- and post-test memory improvement. Y—The maze test result was approximately 44 after sample treatment compared to before sample treatment. % Showed improvement in memory (see FIG. 3). The medium treated group, which was a control group, showed no significant difference from before the test.

 <200>

 <4i> Aβ-inhibitory Effect of Alcohol Sediment from IDCC 3801 SG Culture in Alzheimer's Model Rats

 After the test of Example <4-1>, the brain tissues of the mice were extracted and quantitatively compared to Aβ40 to determine the Aβ inhibitory effect of alcoholic sediment in the IDCC 3801 culture.

After completion of the test, the blood in the brain tissue was removed by perfusion with PBS, and the brain was extracted and weighed. Solution B (PBS, 5% BSA, 0.03% Tween-20, protease), which was maintained at room temperature for 3 hours by adding 8 times solution A (5M guanidine HCl, 50 mM Tris HCl, pH 8.0) by weight Inhibitor solution) was added 20 times relative to volume. The supernatant was recovered by centrifugation at 15,000 rpm for 20 minutes at 4 ° C. to quantify the protein and quantitatively compare Aβ40 in the extract using an ELISA kit (the GENETICS Company, Switzerland).

 As a result, the Aβ40 value of the brain tissue volume was 302.1 30 2.1pg / ml in the control group, whereas the alcoholic sediment administered group of Lactobacillus helvetticus IDCC 3801 culture was 245.2 土 3.2pg / ml in the control group. Aβ40 decreased by about% (see FIG. 4).

 <205>

 <206> Lactobacillus Helveticus IDCC Results

Alcohol sediment in 3801 SG culture filtrate ultrafiltrate was treated in animal model of Alzheimer's disease. It was confirmed that every improvement effect.

 <207>

<208>^'<Example5>^' _.

 <209> Efficacy test of cognitive function of alcohol sediment in in vivo IDCC 3801 SG culture

<2io> Nine-week-old dominant ICR mice were used for each treatment group for cognitive improvement. The Lactobacillus helvetticus IDCC 3801 culture and the ethanol sediment of the present invention were suspended in 1% carboxymethylsalose to prepare a sample for administration, and the control drug donepezil was used in physiological saline. 500 mg / kg of Lactobacillus Helveticus IDCC 3801 culture and medium alcohol infiltrate and 2 mg / kg of donepezil were administered orally twice a week for 2 weeks and lmg of scopolamine 60 minutes before the start of the test. After oral administration of / kg induces memory damage, water maze and passive avoidance tests were performed. As a vehicle, a group treated with scopolamine and a group treated only with physiological saline after administration of saline for two weeks instead of drugs.

 <211>

 <212> <5-1> Water Labyrinth Test

 During the maze test, the reference memory test was performed after adapting by allowing the swimmer to swim freely in a tank without a flat for 60 seconds one day before the test. The test was performed by measuring the time to find the platform submerged 4-5 times daily for 5 days (escape latency, in seconds), and the maximum allowable time was limited to 60 seconds. By the second day of the test, if the platform could not be located, it was led to find the platform within the 60-second time limit.

 24 hours after the end of the reference memory test, a probe test was performed by removing the flats in the tank and allowing them to swim freely for 60 seconds to measure the time of staying at the flats.

 As a result, in the reference memory test, the physiological saline treatment group treated with scopolamine did not improve the time to find the flats during the test period, and the medium alcohol sediment treatment group had a weak learning effect. In contrast, the donepezil treated group used as a reference drug was tested.

From day 2, the time to find the flats was markedly reduced, showing a similar improvement as the scopolamine non-administered group. The alcoholic sediment treatment group of Lactobacillus helvetticus IDCC 3801 cultures was found to have a shorter time to find the flats as the test period elapsed, and after the physiological saline administration on the 5th day of the test, it was 30.5 6.2 seconds compared to 50.0 5 3.3 seconds of the scopolamine treatment group. U There was a significant difference (p <0.05). 3 days (medium alcohol sediment treated group 51.0 土 4.0 sec, Lactobacillus helvetticus IDCC 3801 culture alcoholic sediment 35.3 土 7.1 sec. P <0.05) and 4 days (medium alcohol sediment treated group 49.0 Lactobacillus Helveticus IDCC 3801 cultured alcoholic sediment showed significant difference in 4.2 sec, Lactobacillus Helveticus IDCC 3801 cultured alcoholic sediment 36.9 4.6 seconds, p <0.05). See FIG. 5). In the probe, Lactobacillus Helveticus IDCC 3801 cultured liquor sediment treatment group recovered 1.6 土 0.6 (36.5%) from the place where the platform was located, and 0.8 土 0.3 (17.3%) of physiological saline treatment group. Statistically significant, 2.1 times higher (p <0.05).

 <216>

 <217> <5-2> Manual evasion test

 The passive evasion test was performed using a shuttle box divided into two compartments as a guillotine door. One compartment was illuminated to lighten the other, and the other compartment was unlit and wrapped in black cloth and darkened. On the first day, place the dementia rat in a bright room, give 30 seconds of search time, open the partition door to enter the dark room, measure the acquisition latency time into the dark room, and enter the partition door when entering the dark room. The rats were remembered for electrical stimulation by applying an electric current of 0.5 mA through the grid bottom for 3 seconds. After 24 hours, the demented rats were placed in a bright room, and after 30 seconds of searching time, the partition door was opened to measure the retention latency time up to 300 seconds for all the feet of the rats to enter the dark room. The larger the retention latency time, the better the memory of passive avoidance through learning.

<219> As a result of measuring the retention latency time of each treatment group after learning electrical stimulation, the scopolamine treatment group was 123.7 土 11.0 seconds after the physiological saline solution administration, and the media alcohol sediment treatment group was 80.3 士 35.9. Second, the alcoholic sediment treatment group of Lactobacillus helvetus IDCC 3801 culture was 276.1 sul 21.5 seconds, and the alcoholic sediment treatment group of Lactobacillus helvetus IDCC 3801 culture was significantly more than 232% compared to the physiological saline treatment group (ρθ.01) , Compared with the medium alcohol sediment treatment group showed more than 343% (p <0.01) memory improvement effect. In addition, the ethanol sediment treated group of the Lactobacillus helvetticus IDCC 3801 culture showed a better effect than the donepezil treated group under the current test conditions (see FIG. 6). As a result, it was confirmed that the composition of the present invention has excellent efficacy in improving memory and cognitive function by dementia.

 <221>

 Examples of the formulation of a pharmaceutical composition comprising a compound of the present invention will be described, but the present invention is not intended to be limited thereto but only to be described in detail.

 <223>

 Preparation Example 1 Preparation of Powder

 <225> 20 mg of alcohol sediment in Lactobacillus helvetticus culture

 <226> Lactose 100 mg

 <227> Talc 10 mg

 The above ingredients are mixed and filled in an airtight cloth to prepare a powder.

<229>

 Preparation Example 2 Preparation of Tablet

 <231> alcoholic sediment 10 rag of Lactobacillus helvetus culture

 <232> corn starch 100 mg

 <233> Lactose 100 mg

 <234> magnesium stearate 2 mg

 After mixing the above components and tableting according to the conventional manufacturing method of the tablet to prepare a tablet.

 <236>

 <237> <Example 3> Preparation of a voxel agent

 <238> Alcoholic sediment 10 rag of Lactobacillus helvetus culture

 <239> crystalline cellulose 3 m

 <240> Lactose 14.8 mg

 <24i> Magnesium Stearate 0.2 rag

 According to a conventional capsule preparation method, the above ingredients are mixed and filled into gelatin capsules to prepare capsules.

<244> <Preparation Example> Preparation of injection

<245> 1 mg of alcohol sediment in Lactobacillus helvetus culture

<246> Mannet 180 rag-<247> Sterile distilled water for injection 2793 nig Na ₂ HP0 ₄ 12H ₂ 0 26 mg

 According to the conventional method for preparing an injection, the amount of the above ingredient is prepared per ampoule (2 mi).

Preparation Example 5 Preparation of Liquid

 Alcohol sediment 2 mg from Lactobacillus helvetica culture

 10 g of isomerized sugar

Mantle 5 _g

 Purified water

 According to the conventional method of preparing a liquid solution, each component is added to the purified water to dissolve it, and then the lemon flavor is added, the above components are mixed, the purified water is added, the whole is purified to add the purified water, and then layered into a brown bottle. The solution is prepared by sterilization.

Preparation Example 6 Preparation of Health Food

 Alcohol sediment 500rag from Lactobacillus helvetus culture

 Vitamin Complex Proper

 Vitamin A Acetate 70 iig

 Vitamin E 1.0 mg

 Vitamin B1 0.13 rag

 Vitamin B2 0.15 rag

 Vitamin B6 0.5 rag

 Vitamin B12 0.2 βξ

 Vitamin C 10 mg

 Biotin 10 fig

 Niacinamide 1.7 rag

 Folic acid 50

 Calcium pantothenate 0.5 rag

 Mineral Mixtures Proper

 Ferrous Sulfate 1.75 rag

 Zinc oxide 0.82 rag

Magnesium Carbonate 25.3 rag Crab Potassium Phosphate 15 mg

 <277> Dibasic calcium phosphate 55 rag

 <278> potassium citrate 90 mg

 <279> calcium carbonate 100 mg

 <280> magnesium chloride 24.8 rag

 The composition ratio of the vitamin and mineral mixtures described above is a relatively suitable composition suitable for health foods in a preferred embodiment, but may be modified arbitrarily, the composition ratio of the above ingredients according to the conventional health food manufacturing method After mixing, the granules may be prepared and used to prepare a health food composition according to a conventional method.

 <282>

 <283> <Example 7> Preparation of health drink

 <284> Alcoholic sediment lOOrag of Lactobacillus helvetticus culture

 <285> vitamin C 15 g

 <286> 100 g of vitamin E (powder)

 <287> iron lactate 19.75 g

 <288> zinc oxide 3.5 g

 <289> nicotinic acid amide 3.5 g

 <290> vitamin A 0.2 g

 <29i> vitamin B10.25 g

 <292> vitamin B20.3 g

 <293> water

 After mixing the above components in accordance with a conventional health beverage manufacturing method, and then stirred and heated at 85 ° C. for about 1 hour, the resulting solution is filtered and sealed sterilized in a sterilized 2 I container It is refrigerated and then used to prepare the healthy beverage composition of the present invention.

 Although the composition ratio is a relatively suitable composition for the preferred beverage in a preferred embodiment, the mixing ratio may be arbitrarily modified according to the regional and ethnic preference, such as the demand hierarchy, the demand country, the intended use.

 <296>

 Industrial Applicability

As described above, the pharmaceutical composition containing Lactobacillus helvetus or a culture thereof according to the present invention as an active ingredient, its use and dementia or cognitive dysfunction Prophylactic or alternative methods are effective in preventing or treating dementia or cognitive dysfunction. In particular, the composition of the present invention has an excellent effect of inhibiting the production of beta amyloid and promoting the production of AΡΡα having a neuroprotective effect, and thus has an excellent efficacy in preventing or treating dementia or cognitive dysfunction such as Alzheimer's disease. In addition, the culture production method of the present invention is effective for the production of a culture having excellent efficacy in the prevention or treatment of dementia or cognitive dysfunction of the present invention using the strain of the present invention. Therefore, the pharmaceutical composition, the use thereof, the method of preventing or treating dementia or cognitive dysfunction, or the method for preparing the culture of Lactobacillus helvetticus or a culture thereof according to the present invention may have dementia or cognitive dysfunction. It can be used in various fields, such as the manufacture of prophylactic or therapeutic medicines for people of prone age, which is likely to be used in industry.

ATTESTATION CONCERNING THE LATER INDICATION OR AN AMENDMENT OF THE SCIENTIFIC DESCRIPTION AND / OR PROPOSED TAXONOMIC DESIGNATION

 . issued pursuant to Rule 8.2

 TO: LEE, Kum-Ki

 Ii Dong pharmaceutical Ca, Ltd.

_. 60 Yangae-dong, Seoc o-gu, Seoul 137-733

 Re ublic of Korea

Claims

[Claims]

 [Claim 1]

 Lactobacillus helveticus IDCC 3801 (Accession No .: KCTC-11332BP), which produces a product that inhibits the production of beta amyloid protein.

[Claim 2]

 A pharmaceutical composition for preventing or treating dementia, comprising Lactobacillus helvetikus or a culture thereof as an active ingredient.

[Claim 3]

 3. The method of claim 2, wherein the dementia is Alzheimer's disease dementia, Vascular dementia, Parkinson's disease dementia, Lewy body dementia, Hunt ington. s disease) type dementia, Creutzfeldt-Jacob disease type dementia and Pick's disease type dementia.

[Claim 4]

 Use for the manufacture of a preventive or therapeutic agent for dementia of Lactobacillus helvetica or its culture.

[Claim 5]

 A method for preventing or treating dementia, characterized by administering Lactobacillus helvetticus or its culture to an individual in need thereof in an effective amount.

[Claim 6]

 A pharmaceutical composition for preventing or treating cognitive impairment, comprising Lactobacillus helvetica or its culture as an active ingredient.

[Claim 7]

7. The method of claim 6, wherein the cognitive dysfunction is associated with cognitive deficits associated with schizophrenia, cognitive deficits associated with age-induced memory impairment or psychiatric disorders, cognitive deficits associated with diabetes, cognitive deficiencies associated with post-stroke, hypoxia Memory deficiency, senile dementia and lead Related cognitive and attention deficits, memory problems associated with mild cognitive impairment, impaired cognitive function associated with dementia, impaired cognitive function associated with Alzheimer's disease, impaired cognitive function associated with Parkinson's disease, impaired cognitive function associated with vasculitis, and tumor-related cognition Cognitive impairment selected from the group consisting of problem, Pick's disease, cognitive deficit due to autism, cognitive deficit after electroconvulsion therapy, cognitive deficit associated with traumatic brain injury, memory loss disorder and delirium Characterized in that the composition.

[Claim 8]

 Use for the manufacture of a preventive or therapeutic agent for cognitive dysfunction of Lactobacillus helvetticus or its culture.

[Claim 9]

 A method for preventing or treating cognitive dysfunction, characterized by administering Lactobacillus helvetticus or its culture to an individual in need thereof in an effective amount.

[Claim 10]

 A food composition for enhancing cognitive function, comprising Lactobacillus helvetikus or a culture thereof as an active ingredient.

[Claim 11]

 The composition according to any one of claims 2, 6 and 10, wherein the Lactobacillus helvetikus has an accession number of KCTC-11332BP.

[Claim 12]

 The composition according to any one of claims 2, 6 and 10, wherein the culture is incubated with a culture solution containing dairy material.

[Claim 13]

 The composition of claim 12, wherein the dairy material is one or a combination thereof selected from the group consisting of skim milk, milk, goat oil, camel oil and goat milk.

[Claim 14] The method according to claim 4 or 8, wherein the Lactobacillus helvetticus has a deposit number of KCTC-11332BP.

[Claim 15]

 10. The method according to claim 5 or 9, wherein the Lactobacillus helveticus has an accession number of KCTC-11332BP.

[Claim 16]

 (a) inoculating the medium containing lactose into the culture medium by inoculating Lactobacillus helvetticus;

 (b) removing the lactic acid bacteria from the culture; And

 (c) Lactobacillus helveti having a dementia prophylactic or therapeutic effect comprising the step of precipitating and separating the active ingredient by adding a solvent selected from the group consisting of acetone or an alcohol having 1 to 6 carbon atoms to remove the culture lactic acid bacteria; Method of Making Cous Cultures.

[Claim 17]

 The method of claim 16, wherein the manufacturing method

 (a) inoculating the medium containing lactose into the culture medium by inoculating Lactobacillus helvetticus;

 (b) centrifuging the culture to remove lactic acid bacteria;

 (c) obtaining a substance having a molecular weight of 5,000 or less in the culture in which the lactic acid bacteria have been removed; And

 (d) lactose having a dementia prevention and treatment effect, comprising the step of precipitating and isolating the active ingredient by adding a solvent selected from the group consisting of acetone and an alcohol having 1 to 6 carbon atoms to the obtained material; Method for the preparation of Bacillus helvetticus culture.

[Claim 18]

 18. The method according to claim 16 or 17, wherein the Lactobacillus helveticus has an accession number of KCTC-11332BP.