Agents and methods for maintaining or improving brain function using crude drugs

Abstract

As a new means for maintaining or improving cognitive function, provided is an agent that is for maintaining or improving cerebral function and that contains a herbal medicine selected from a stem part of Japanese sweet flag and a jujube seed. Here, said jujube seed is chosen from a crushed material of the same or an extraction residue of the crushed material, and said stem part of Japanese sweet flag is chosen from a crushed material of the same, an extract of the crushed material, or an extraction residue of the crushed material.

Description

Technical Field

[0001] The present invention relates to agents and methods for maintaining or improving brain function, promoting nerve cell repair or inducing neurogenesis, removing dementia-causing proteins accumulated in the brain, or treating or preventing neurodegenerative diseases using crude drugs, as well as foods and pharmaceuticals containing such agents.

Background Art

[0002] With the aging of society and the westernization of lifestyle and eating habits, the number of dementia patients is increasing rapidly worldwide. The social costs required for their medical care and nursing care and the economic losses due to the decline in the labor force of patients and their families are enormous, posing a major social problem.

[0003] Typical dementias include Alzheimer's disease, frontotemporal dementia, Lewy body dementia, etc. In these diseases, specific proteins aggregate and accumulate in the nervous system, which is thought to cause dementia by leading to nerve cell death. Specifically, Aβ and tau accumulate in the brain in Alzheimer's disease, tau and TDP-43 accumulate in the brain in frontotemporal dementia, and α-synuclein accumulates in the brain in Lewy body dementia (Non-Patent Document 1: Spires-Jones et al., Acta Neuropathol., (2017), 134[2]:187-205).

[0004] As anti-dementia drugs, the development of drugs that suppress the production of these causative proteins or remove the causative proteins from the brain is underway. As candidates for such anti-dementia drugs, for Alzheimer's disease, Aβ production enzyme (β-secretase and γ-secretase) inhibitors (Non-Patent Document 2: Luo et al., Cell & Biosci., (2022), 12:2), Aβ vaccines (Non-Patent Document 3: Valiukas et al., Vaccines, (2022), 10[9]:1527), Aβ antibodies (Non-Patent Document 4: Song et al., Transl. Neurodegener., (2022), 11:18), etc. For frontotemporal dementia, tau vaccines (Non-Patent Document 5: Medina, Int. J. Mol. Sci., (2018), 19[4]:1160), tau antibodies (Non-Patent Document 6: Ji et al., Drugs, (2021), 81

[10] :1135-1152), etc. For Lewy body dementia, α-synuclein inhibitors and α-synuclein antibodies (Non-Patent Document 7: Alzforum website, FBRI LLC, search results for alpha-synuclein Target, https: / / www.alzforum.org / therapeutics / search?fda_statuses=&target_types%5B%5D=33416&therapy_types=&conditions=&keywords-entry=&keywords=, searched in December 2022), etc. have been studied respectively.

[0005] However, most of the anti-dementia drug candidates developed so far have not confirmed the drug efficacy expected in clinical trials for dementia patients and have ended in failure (Non-Patent Document 8: Asher et al., Life Sciences, (2022), 306:120861).

[0006] Ziziphus jujuba var. spinosa is said to have the effect of "stabilizing the mind". Ziziphus jujuba var. spinosa powder is known as a medicinal diet ingredient. Ziziphus jujuba var. spinosa extract is known to have an effect of improving cognitive function (Non-Patent Document 9: Chinese Traditional and Herbal Drugs, (2001), 32[3]:246-247, Non-Patent Document 10: Journal of Guangxi Traditional Chinese Medical University, (2002), 5[3]:11-13, Non-Patent Document 11: Journal of Xi'an Jiaotong University (Medical Sciences), (2010), 31[6]:673-707). Regarding jujuboside and spinosin, which are the main components of Ziziphus jujuba var. spinosa, an anti-dementia effect has been reported (Non-Patent Document 12: Liu et al., Eur. J. Pharmacol., (2014), 738:206-213; Non-Patent Document 13: Zhang et al., Theranostics, (2018), 8:4262-4278; Non-Patent Document 14: Tabassum et al., Sci. Rep., (2019), 9:4512; Non-Patent Document 15: Ko et al., Biomol. Ther., (2015), 23:156-164; Non-Patent Document 16: Lee et al., Pharmacol. Biochem. Behav., (2016), 145:9-16; Non-Patent Document 17: Xu et al., Biomol. Ther., (2019), 27:71-77; Non-Patent Document 18: Cai et al., Biomol. Ther., (2020), 28:131-136; and Non-Patent Document 19: Zhang et al., Biomol. Ther., (2020), 28:259-266).

[0007] Acorus tatarinowii (Sekishoubu) is said to be effective against amnesia because its root part (which may be referred to as "Sekishougen" as appropriate) "improves blood flow and clarifies consciousness" and has been commonly used in traditional Chinese medicine and Kampo medicine. Regarding asarone and eugenol, which are the main components of Sekishougen, there are reports on their anti-dementia effects (Non-Patent Document 20: Geng et al., Biol. Pharm. Bull., (2010), 33:836-843; Non-Patent Document 21: Liu et al., Yakugaku Zasshi, (2010),130[5]:737-746; Non-Patent Document 22: Wei et al., J. Alzheimers Dis., (2013),33:863-880; Non-Patent Document 23: Yang et al., Cell Mol. Neurobiol., (2016), 36:121-130; Non-Patent Document 24: Xue et al., Eur. J. Pharmacol., (2014), 741:195-204; Non-Patent Document 25: Chen et al., Brain Res., (2014), 1552:41-54; and Non-Patent Document 26: Mao et al., Aging Cell, (2015), 14:784-796). However, the efficacy of the stem part of Acorus tatarinowii has not been clearly claimed, and it is hardly used in traditional Chinese medicine and Kampo medicine.

[0008] Moreover, regarding neither Ziziphus jujuba seeds nor Acorus tatarinowii, the effects such as promoting the repair of nerve cells or inducing neurogenesis, removing the dementia-causing proteins accumulated in the brain, treating or preventing neurodegenerative diseases, etc. are not known.

Prior Art Documents

Non-Patent Documents

[0009]

Non-Patent Document 1

Non-Patent Document 2

Non-Patent Document 3

Non-Patent Document 4

Non-Patent Document 5

Non-Patent Document 6

[10] :1135 - 1152

Non-Patent Document 7

Non-Patent Document 8

Non-Patent Document 9

Non-Patent Document 10

Non-Patent Document 11

Non-Patent Document 12

Non-Patent Document 25

Non-Patent Document 26

Non-Patent Document 27

Non-Patent Document 28

Non-Patent Document 29

Non-Patent Document 30

Non-Patent Document 31

Non-Patent Document 32

[14] :4845-4856

Non-Patent Document 33

Non-Patent Document 34

Non-Patent Document 35

Non-Patent Document 36

Non-Patent Document 37

Summary of the Invention

Problems to be Solved by the Invention

[0010] The problem to be solved by the present invention is to provide new means for maintaining or improving brain function, promoting nerve cell repair or inducing neurogenesis, removing dementia-causing proteins accumulated in the brain, or treating or preventing neurodegenerative diseases.

Means for Solving the Problems

[0011] As a result of intensive studies, the present inventors have found that the stems of Ziziphus jujuba var. spinosa and Acorus tatarinowii have various effects such as maintaining or improving brain function, promoting nerve cell repair or inducing neurogenesis, removing dementia-causing proteins accumulated in the brain, or treating or preventing neurodegenerative diseases in each predetermined form, and have completed the present invention.

[0012] That is, the gist of the present invention relates to, for example, the following. [Item 1] An agent for maintaining or improving brain function, containing a crude drug selected from the stems of Ziziphus jujuba var. spinosa (Sansou'nin) and Acorus tatarinowii (Sekishoubu), wherein the Ziziphus jujuba var. spinosa is selected from crushed materials and extraction residues of crushed materials, and the stem part of the Acorus tatarinowii is selected from crushed materials, extracts of crushed materials, and extraction residues of crushed materials. [Item 2] The agent for maintaining or improving brain function according to Item 1, wherein the brain function is a cognitive function. [Item 3] An agent for promoting nerve cell repair or inducing neurogenesis, containing a crude drug selected from the stems of Ziziphus jujuba var. spinosa and Acorus tatarinowii, wherein the Ziziphus jujuba var. spinosa and the stem part of the Acorus tatarinowii are each independently selected from crushed materials, extracts of crushed materials, and extraction residues of crushed materials. [Item 4] An agent for removing causative proteins of dementia that accumulate in the brain, containing crude drugs selected from jujube seeds and the stem part of Acorus tatarinowii, wherein the jujube seeds and the stem part of Acorus tatarinowii are each independently selected from crushed materials, extracts of crushed materials, and extraction residues of crushed materials. [Item 5] The agent according to Item 4, wherein the causative protein of dementia is one or more proteins selected from amyloid β (Aβ), tau, α-synuclein, TDP-43, FUS / TLS, polyglutamine, proteins by RAN (repeat-associated non-ATG) translation, prions, and SOD-1. [Item 6] An agent for treating or preventing neurodegenerative diseases, containing crude drugs selected from jujube seeds and the stem part of Acorus tatarinowii, wherein the jujube seeds and the stem part of Acorus tatarinowii are each independently selected from crushed materials, extracts of crushed materials, and extraction residues of crushed materials. [Item 7] The agent according to Item 6, wherein the neurodegenerative disease is degenerative dementia. [Item 8] The agent according to Item 7, wherein the degenerative dementia is one or more dementias selected from Alzheimer's disease, frontotemporal dementia, and Lewy body dementia. [Item 9] The agent according to any one of Items 1 to 8, wherein the crude drug is administered to the subject in an amount of 0.05 mg to 10 g per day. [Item 10] A food containing the agent according to any one of Items 1 to 8. [Item 11] A medicine containing the agent according to any one of Items 1 to 8. [Item 12] A method for maintaining or improving the brain function of a subject, comprising administering a crude drug selected from jujube seeds and the stem part of Acorus tatarinowii to a subject in need thereof, wherein the jujube seeds are selected from crushed materials and extraction residues of crushed materials, and the stem part of Acorus tatarinowii is selected from crushed materials, extracts of crushed materials, and extraction residues of crushed materials. [Item 13] The method according to Item 12, wherein the brain function is cognitive function. [Item 14] A method for promoting the repair of target nerve cells or inducing neurogenesis, comprising administering a crude drug selected from the seeds of Ziziphus jujuba var. spinosa and the stem part of Acorus tatarinowii to a subject in need thereof, wherein each of the seeds of Ziziphus jujuba var. spinosa and the stem part of Acorus tatarinowii is independently selected from a crushed product, an extract of the crushed product, and an extraction residue of the crushed product. [Item 15] A method for removing a dementia-causing protein accumulated in the brain of a subject, comprising administering a crude drug selected from the seeds of Ziziphus jujuba var. spinosa and the stem part of Acorus tatarinowii to a subject in need thereof, wherein each of the seeds of Ziziphus jujuba var. spinosa and the stem part of Acorus tatarinowii is independently selected from a crushed product, an extract of the crushed product, and an extraction residue of the crushed product. [Item 16] The method according to Item 15, wherein the dementia-causing protein is one or more proteins selected from amyloid-β (Aβ), tau, α-synuclein, TDP-43, FUS / TLS, polyglutamine, a protein by RAN (repeat-associated non-ATG) translation, prion, and SOD-1. [Item 17] A method for treating or preventing a neurodegenerative disease of a subject, comprising administering a crude drug selected from the seeds of Ziziphus jujuba var. spinosa and the stem part of Acorus tatarinowii to a subject in need thereof, wherein each of the seeds of Ziziphus jujuba var. spinosa and the stem part of Acorus tatarinowii is independently selected from a crushed product, an extract of the crushed product, and an extraction residue of the crushed product. [Item 18] The method according to Item 17, wherein the neurodegenerative disease is degenerative dementia. [Item 19] The method according to Item 18, wherein the degenerative dementia is one or more dementias selected from Alzheimer's disease, frontotemporal dementia, and Lewy body dementia. [Item 20] The method according to any one of Items 12 to 19, wherein the crude drug is administered to the subject in an amount of 0.05 mgmg to 10 g / day. [Item 21] A crushed powder of a crude drug selected from the seeds of Ziziphus jujuba var. spinosa and the stem part of Acorus tatarinowii, wherein the proportion of the crushed powder having a particle size of 100 μm or less in the crushed powder is 50% or more. [Item 22] The crushed powder according to Item 21, having a water activity of less than 0.7. [Effect of the Invention]

[0013] According to the present invention, by using the stems of Ziziphus jujuba var. spinosa and Acorus tatarinowii in respective predetermined forms, new means for maintaining or improving brain function, promoting the repair of nerve cells or inducing neurogenesis, removing the dementia-causing proteins accumulated in the brain, or treating or preventing neurodegenerative diseases are provided.

Brief Description of the Drawings

[0014]

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[0015] Hereinafter, the present invention will be described in detail in accordance with specific embodiments. However, the present invention is not limited to the following embodiments, and can be implemented in any form without departing from the spirit of the present invention.

[0016] [Summary] As described above, it is considered that the reason why most of the previously studied anti-dementia drugs have ended in failure, apart from the problem of side effects, is that the administration time of the drugs is too late and the wrong molecules are targeted. It has been shown that the accumulation of Aβ in the brain starts more than 20 years before the onset of Alzheimer's disease, and the accumulation of tau also starts about 10 years ago. Aβ accumulates, tau accumulates, nerve cells begin to die, and then dementia finally develops. That is, by the time dementia develops, many nerve cells have already died. If one wants to remove Aβ or tau, it makes no sense unless it is before nerve cells begin to die. That is to say, the role of drugs targeting Aβ or tau is in prevention rather than treatment. Also, previously, it was thought that the disease developed when insoluble aggregates of proteins such as senile plaques and neurofibrillary changes (accumulations of aggregated tau) killed nerve cells. However, recently, it is thought that soluble oligomers formed at an earlier stage impair the function of nerve cells and cause dementia. Therefore, it is necessary to remove the oligomers of the causative proteins in the prevention of dementia.

[0017] From the perspective of prevention, it is desirable that a drug can act on the oligomers of various causative proteins with a single agent, rather than being specific to Aβ, tau, or α-synuclein. Furthermore, an action to repair nerve cells damaged by oligomers and restore brain function is also necessary. Also, since the prevention of dementia is a long-term process, it is desirable that the preventive drug is safe, inexpensive, and can be taken non-invasively by oneself without the help of a doctor if possible. Since there are many requirements for a preventive drug for dementia like this, it is difficult to achieve this with a pharmaceutical product consisting of a single component. Also, if all middle-aged and elderly people have to take it for a long time for the prevention of dementia, there is a risk that the medical economy will collapse with pharmaceuticals in the end.

[0018] To solve such problems, the inventors focused on crude drugs used in traditional Chinese medicine and Kampo medicine, which have a long history. If there are crude drugs effective in improving cognitive function, middle-aged and elderly people can obtain them at their own discretion without seeing a doctor, and by taking them in addition to diet, etc., they can strive to prevent dementia in their daily lives. As a result of intensive studies, the inventors have found that sour jujube seeds and the stem part of Acorus tatarinowii (Acorus stem) each have various effects in a predetermined form, such as maintaining or improving brain function, promoting the repair of nerve cells or inducing neurogenesis, removing dementia-causing proteins accumulated in the brain, or treating or preventing neurodegenerative diseases. The present invention is based on such findings.

[0019] That is, according to one aspect of the present invention, there is provided an agent for maintaining or improving brain function, an agent for promoting the repair of nerve cells or inducing neurogenesis, an agent for removing dementia-causing proteins accumulated in the brain, or an agent for treating or preventing neurodegenerative diseases (these may be collectively referred to as "the agent of the present invention" as appropriate), which contains a crude drug selected from sour jujube seeds and the stem part of Acorus tatarinowii.

[0020] Also, according to one aspect of the present invention, there is provided a food containing the agent of the present invention (this may be referred to as "the food of the present invention" as appropriate).

[0021] Also, according to one aspect of the present invention, there is provided a medicine containing the agent of the present invention (this may be referred to as "the medicine of the present invention" as appropriate).

[0022] Also, according to one aspect of the present invention, there is provided a method for maintaining or improving brain function, a method for promoting the repair of nerve cells or inducing neurogenesis, a method for removing dementia-causing proteins accumulated in the brain, or a method for treating or preventing neurodegenerative diseases (these may be collectively referred to as "the method of the present invention" as appropriate), which includes administering one or more selected from a crude drug selected from sour jujube seeds and the stem part of Acorus tatarinowii, the agent of the present invention, the food of the present invention, and the medicine of the present invention.

[0023] [Ziziphus jujuba Mill. Spinosae Semen] According to one embodiment, the agent of the present invention contains sour jujube seed (Ziziphus jujuba Mill. var. spinosa Hu) as a crude drug. Sour jujube seed is said to have the effect of "stabilizing the mind" and is formulated with polygala root, longan aril, etc. in traditional Chinese medicine and is prescribed for insomnia and amnesia. In the previous notice from the Pharmaceutical Affairs Bureau of the Ministry of Health, Labour and Welfare, sour jujube seed is also treated as a non-pharmaceutical product and can be used as a food.

[0024] The present inventors prepared a powder obtained by drying and crushing sour jujube seed (Cru-ZSS), an extract prepared by extracting the dried powder with hot water or the like (Ext-ZSS), and an extraction residue remaining after extraction (Res-ZSS), and orally administered them to model mice with frontotemporal dementia showing tau pathology (Tau784 mice: Examples A1, A3, and A4), model mice with frontotemporal dementia showing TDP-43 pathology (C9-500 mice: Example C1), and model mice with Alzheimer's disease showing Aβ pathology (APP23 mice: Example A2) for one month to evaluate the cognitive function improvement effect and brain pathology improvement effect. Furthermore, the powder obtained by drying and crushing sour jujube seed (Cru-ZSS) was orally administered to model mice with Lewy body dementia showing α-synuclein pathology (Huα-Syn(A53T) mice: Example A6) for one month to evaluate the brain pathology improvement effect and neurogenesis effect. In addition, the nerve repair effect of sour jujube seed was evaluated based on the induction of the expression of brain-derived neurotrophic factor (BDNF) and the induction of neurogenesis. As a result, it was found that sour jujube seed has various activities such as improvement of cognitive function in model mice, removal of oligomers of Aβ, tau, and α-synuclein, recovery of synapses, induction of the expression of brain-derived neurotrophic factor (BDNF), and induction of neurogenesis not only in the extract (Ext-ZSS) obtained by hot water extraction, which is a common treatment in traditional Chinese medicine, but also in the extraction residue (Res-ZSS) that is usually discarded and even in the powder (Cru-ZSS) simply crushed. Moreover, it is particularly noteworthy that the activity was the strongest in the crushed powder of sour jujube seed, and the cognitive function of the mice given the crushed powder of sour jujube seed became better than that of wild-type mice of the same age.

[0025] The main components of Ziziphus jujuba var. spinosa include spinosin, jujuboside A, and jujuboside B. It is already known that these components have an Aβ removal effect, an effect of inducing BDNF expression, and a neurogenic effect (Non-Patent Document 12 mentioned above: Liu et al., Eur. J. Pharmacol., (2014), 738:206-213; Non-Patent Document 13: Zhang et al., Theranostics, (2018), 8:4262-4278; Non-Patent Document 14: Tabassum et al., Sci. Rep., (2019), 9:4512; Non-Patent Document 15: Ko et al., Biomol. Ther., (2015), 23:156-164; Non-Patent Document 16: Lee et al., Pharmacol. Biochem. Behav., (2016), 145:9-16; Non-Patent Document 17: Xu et al., Biomol. Ther., (2019), 27:71-77; Non-Patent Document 18: Cai et al., Biomol. Ther., (2020), 28:131-136; and Non-Patent Document 19: Zhang et al., Biomol. Ther., (2020), 28:259-266). When the present inventors compared the contents of these components between the powder (Cru-ZSS) obtained by drying and crushing Ziziphus jujuba var. spinosa and the extract (Ext-ZSS) prepared by extracting the dried powder with hot water, although the content of these components was higher in the extract (Ext-ZSS) (Example A5), the cognitive function improving effect was stronger in the crushed powder (Cru-ZSS) (Example A4). Further, when a mixture of the same amounts of spinosin, jujuboside A, and jujuboside B contained in the extract (Ext-ZSS) was administered to mice, the cognitive function improving effect was weaker than when the extract (Ext-ZSS) was administered (Example A5). These results suggest that Ziziphus jujuba var. spinosa contains components effective for improving cognitive function in addition to spinosin, jujuboside A, and jujuboside B, and furthermore, the active components are most abundantly contained in the simply crushed powder.

[0026] [Acori Tatarinowii Rhizoma] According to one embodiment, the agent of the present invention contains the stem part of Acorus gramineus Soland. (this may be appropriately referred to as "Acorus stem" in some cases). Acorus gramineus Soland. is said to be effective for amnesia because its root part (this may be appropriately referred to as "Acorus root" in some cases) "improves blood flow and clarifies consciousness", and has been commonly used in traditional Chinese medicine and Kampo medicine. In the Notification of the Director of the Pharmaceutical Affairs Bureau of the Ministry of Health, Labour and Welfare "Regarding the Guidance and Control of Unapproved and Unauthorized Pharmaceuticals" (Appendix "Standards Regarding the Scope of Pharmaceuticals"), Appendix 3 "List of Substances (Raw Materials) That Are Not Considered Pharmaceuticals Unless They Claim Pharmaceutical Efficacy and Effects" (this may be appropriately referred to as the "Notification of the Director of the Pharmaceutical Affairs Bureau of the Ministry of Health, Labour and Welfare" in some cases), Acorus root is treated as a pharmaceutical. However, the efficacy of Acorus stem has not been clearly claimed, and it is rarely used in traditional Chinese medicine and Kampo medicine. In the aforementioned Notification of the Director of the Pharmaceutical Affairs Bureau of the Ministry of Health, Labour and Welfare, Acorus stem is treated as a non-pharmaceutical and can be used as a food.

[0027] The present inventors prepared powder obtained by drying and crushing Acorus gramineus stems (Cru-AGS), an extract (Ext-AGS) prepared by extracting the dried powder with hot water or the like, and an extraction residue (Res-AGS) remaining after extraction, and orally administered them to model mice of frontotemporal dementia showing tau pathology (Tau784 mice: Examples B1 and B3), model mice of Alzheimer's disease showing Aβ pathology (APP23 mice: Example B2), and model mice of frontotemporal dementia showing TDP-43 pathology (C9-500 mice: Example C1) for one month to evaluate the cognitive function improving effect and brain pathology improving effect. Further, powder obtained by drying and crushing Acorus gramineus stems (Cru-AGS) was orally administered to model mice of Lewy body dementia showing α-synuclein pathology (Huα-Syn(A53T) mice: Example B4) for one month to evaluate the brain pathology improving effect and neurogenesis effect. In addition, the nerve repair effect of Acorus gramineus stems was evaluated based on the induction of the expression of brain-derived neurotrophic factor (BDNF) and the induction of neurogenesis. As a result, Acorus gramineus stems were found to have various activities such as improving the cognitive function of model mice, removing oligomers of Aβ, tau, and α-synuclein, restoring synapses, inducing the expression of brain-derived neurotrophic factor (BDNF), and inducing neurogenesis not only in the extract (Ext-AGS) obtained by hot water extraction, which is a common treatment in traditional Chinese medicine, but also in the extraction residue (Res-AGS) that is usually discarded and even in the powder (Res-AGS) simply crushed. Moreover, it is particularly noteworthy that the activity was the strongest in the crushed powder (Res-AGS) of Acorus gramineus stems.

[0028] The main active ingredients of Acorus tatarinowii root used in traditional Chinese medicine and Kampo medicine are α-asarone, β-asarone, eugenol, etc. It is already known that these ingredients have the effect of removing proteins causing dementia such as amyloid-β, tau, α-synuclein, etc., and the effect of inducing the expression of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), glial cell-derived neurotrophic factor (GDNF), etc. (Non-Patent Document 20 mentioned above: Geng et al., Biol. Pharm. Bull., (2010), 33:836-843; Non-Patent Document 21: Liu et al., Yakugaku Zasshi, (2010), 130:737-746; Non-Patent Document 22: Wei et al., J. Alzheimers Dis., (2013), 33:863-880; Non-Patent Document 23: Yang et al., Cell Mol. Neurobiol., (2016), 36:121-130; Non-Patent Document 24: Xue et al., Eur. J. Pharmacol., (2014), 741:195-204; Non-Patent Document 25: Chen et al., Brain Res., (2014), 1552:41-54; and Non-Patent Document 26: Mao et al., Aging Cell, (2015), 14:784-796). However, it is a finding that has been clarified for the first time by the study of the present inventors that Acorus tatarinowii stem, which contains relatively few of these ingredients and is not commonly used, has a strong cognitive function improving effect on Acorus tatarinowii root containing a large amount of these ingredients.

[0029] [Morphology of crude drugs] According to one embodiment, the semen ziziphi spinosae can be used in any form. Examples of such forms include, but are not limited to, crushed semen ziziphi spinosae, extracts of the crushed product, and extraction residues of the crushed product. Among them, when used as the agent of the present invention for maintaining or improving brain function, the semen ziziphi spinosae is usually used in the form of crushed product or extraction residue of the crushed product, but it is preferably used in the form of crushed semen ziziphi spinosae. On the other hand, when used as the agent of the present invention for promoting nerve cell repair or inducing neurogenesis, removing the causative protein of dementia accumulated in the brain, or treating or preventing neurodegenerative diseases, the semen ziziphi spinosae may be in any form of crushed product, extract of the crushed product, and extraction residue of the crushed product, but it is preferably used in the form of crushed semen ziziphi spinosae.

[0030] According to one embodiment, the acorus tatarinowii rhizome can be used in any form. Examples of such forms include, but are not limited to, crushed acorus tatarinowii rhizome, extracts of the crushed product, and extraction residues of the crushed product. Among them, it is preferably used in the form of crushed acorus tatarinowii rhizome.

[0031] In traditional Chinese medicine and Kampo medicine, it is common to process dried crude drugs with hot water (i.e., decoct them) and take the extracts obtained therefrom, and the remaining residues are discarded. However, according to the findings of the present inventors, when crude drugs selected from semen ziziphi spinosae and acorus tatarinowii rhizome are dried and then used in the form of crushed products such as simply crushed powders, various cognitive function improving effects are most strongly exerted. This can be said to be a surprising finding that cannot be predicted from the common sense of traditional Chinese medicine and Kampo medicine.

[0032] Specifically, when the crude drug selected from Ziziphus jujuba Mill. var. spinosa (Bunge) Hu ex H. F. Chow and Acorus tatarinowii Schott is in the form of a crushed product such as crushed powder, the processing conditions are not particularly limited, but are as follows, for example. First, the crude drug is dried. The conditions for the drying treatment are not limited, and various known conditions may be used. For example, it can be dried at a temperature of 0 to 100 °C in the form of natural drying or heat drying. Also, roasting may be performed. Next, the dried or roasted crude drug is subjected to a crushing treatment. The means for the crushing treatment is not limited, and various known means may be used. Examples include a crushing method by handwork and crushing using a crusher such as a wet pulverization method or a dry pulverization method. Examples of the dry pulverization method include a hammer mill crushing method and a pneumatic flow pulverization method. In the pneumatic flow pulverization method, a swirling airflow is generated in the pulverization chamber by a rotating rotor and blades, and the input sample is pulverized by repeatedly colliding with each other by the swirling airflow. The finely pulverized sample moves to a recovery chamber called a classification chamber, but the insufficiently pulverized sample returns to the pulverization chamber and the pulverization progresses. The obtained crushed product of the crude drug may be used as it is, or a powder having a particle size controlled to a predetermined value or less may be obtained by passing it through a sieve. For example, a sieve with a mesh size of 0.02 mm to 20 mm can be used. More specifically, for example, the Ziziphus jujuba Mill. var. spinosa (Bunge) Hu ex H. F. Chow powder can be sieved through a sieve with a mesh size of 2 to 3 mm and then a 500 μm sieve, and the Acorus tatarinowii Schott stem powder can be sieved through a sieve with a mesh size of 2 to 3 mm and then a 500 μm sieve to adjust the particle size.

[0033] From the viewpoint of usability, it is preferable to adjust the particle size of the crushed powder so that the crushed product having a particle size below a predetermined value accounts for a predetermined ratio or more among the crushed products of the crude drug. Specifically, among the crushed products of the crude drug, it is preferable that the ratio of the crushed product having a particle size of 100 μm or less is 50% or more, and more preferably 75% or more. Also, from the viewpoint of storage stability, the finally obtained crushed powder preferably has a water activity of less than 0.7, more preferably less than 0.64, and even more preferably less than 0.6. Note that the crushed product of the crude drug in which the crushed product having a particle size below a predetermined value accounts for a predetermined ratio or more is also included in the scope of the present invention.

[0034] In addition, when the crude drug selected from Ziziphus jujuba and Acorus tatarinowii is in the form of a crushed extract or extraction residue, the treatment conditions are not particularly limited. For example, an extraction solvent may be added to the dried and crushed crude drug for extraction. Specifically, for 1 part by mass of the dried and crushed crude drug, for example, 1 to 100 parts by mass of the extraction solvent may be added and held for, for example, 0.1 to 100 hours to perform extraction. The extraction solvent is also not particularly limited, and various known solvents can be used. Examples include water and hydrophilic organic solvents such as ethanol. The extraction using the extraction solvent is carried out at room temperature or with heating. For example, when heating with water, it can be heated at 30 to 100 °C with hot water added to the crushed material. In addition, if necessary, stirring treatment, ultrasonic treatment, heating reflux treatment, etc. can be used in combination to enhance the extraction efficiency. After completion of the extraction, the liquid component and the solid component are separated by means such as manual operation or filtration. The separated liquid component can be used as the extract of the crushed material, and the solid component can be used as the extraction residue of the crushed material. In either case, the solvent may be evaporated and concentrated before use if necessary.

[0035] [Action of agents] According to one embodiment, the agent of the present invention is an agent for maintaining or improving brain function. In the present invention, "maintaining or improving brain function" means, but is not limited to, maintaining or improving cognitive function and / or motor function.

[0036] In the present invention, "maintaining or improving cognitive function" means, but is not limited to, preventing the occurrence of mild memory loss, memory impairment, disorientation, impairment of judgment and comprehension, executive function impairment, apraxia, agnosia, aphasia, etc. seen in dementia, or improving these abnormalities.

[0037] In the present invention, the maintenance or improvement of "motor function" means, but is not limited to, preventing the occurrence of abnormalities such as tremors, muscle rigidity, postural reflex disorders, parkinsonism such as akinesia and hypokinesia seen in Parkinson's disease, involuntary movements seen in Huntington's disease, coordination disorders seen in spinocerebellar degeneration, and muscle weakness seen in amyotrophic lateral sclerosis, or improving these abnormalities.

[0038] The effect of maintaining or improving brain function by the agent of the present invention can be evaluated, for example, as shown in the following examples, by administering the agent to a model animal that develops brain pathology and then subjecting it to cognitive function tests such as the Morris water maze test and motor function tests such as the rotarod test, or by staining and observing brain sections.

[0039] According to one embodiment, the agent of the present invention is an agent for promoting nerve cell repair or inducing neurogenesis. In the present invention, "promoting nerve cell repair" includes, but is not limited to, restoring the number and function of decreased synapses, restoring the function of decreased nerve cells, promoting the maintenance and recovery of the functions of synapses and nerve cells, and enhancing the expression of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), etc. that protect nerve cells from various stresses. In the present invention, "inducing neurogenesis" means, but is not limited to, inducing the new appearance of immature nerve cells that actively perform DNA synthesis. Indicators of DNA synthesis include actively incorporating nucleic acid analogs such as BrdU administered from the outside, and expressing doublecortin, etc. as markers of immature nerve cells. The effect of promoting nerve cell repair or inducing neurogenesis by the agent of the present invention can be evaluated, for example, as shown in the following examples, by observing the induction of the expression of brain-derived neurotrophic factor (BDNF) after administering the agent to a model animal that develops dementia or brain pathology, or by staining and observing brain sections.

[0040] According to one embodiment, the agent of the present invention is an agent for removing causative proteins of dementia that accumulate in the brain. In the present invention, the "causative protein of dementia" means a protein that has been identified or presumed as a causative substance of dementia. Examples include, but are not limited to, one or more proteins selected from amyloid-β (Aβ), tau, α-synuclein, TDP-43, FUS / TLS, polyglutamine, proteins by RAN (repeat-associated non-ATG) translation, prions, SOD-1, etc. The action of maintaining or improving brain function by the agent of the present invention can be evaluated, for example, as shown in the examples described later, by administering the agent to a model animal that develops dementia and then staining and observing brain sections.

[0041] According to one embodiment, the agent of the present invention is a therapeutic or prophylactic agent for neurodegenerative diseases. In the present invention, the "neurodegenerative disease" means, but is not limited to, a neurodegenerative disease in which any of the aforementioned causative proteins of dementia accumulates in the brain. Examples of neurodegenerative diseases include, but are not limited to, Alzheimer's disease (Aβ, tau), frontotemporal lobar degeneration (tau, TDP-43, FUS / TLS), Lewy body dementia (α-synuclein), Parkinson's disease (α-synuclein), multiple system atrophy (α-synuclein), Huntington's disease (polyglutamine), amyotrophic lateral sclerosis (TDP-43, FUS / TLS, RAN protein, SOD-1), spinocerebellar degeneration (RAN protein), Creutzfeldt-Jakob disease (prion), etc. (Examples of causative proteins of dementia that accumulate in each neurodegenerative disease are shown in parentheses.). Among them, the therapeutic or prophylactic agent for neurodegenerative diseases of the present invention is preferably a therapeutic or prophylactic agent for degenerative dementia. Examples of degenerative dementia include, but are not limited to, Alzheimer's disease, frontotemporal dementia, Lewy body dementia, etc. Among them, if frontotemporal dementia in which tau accumulates is further classified in detail, Pick's disease, corticobasal degeneration, progressive supranuclear palsy, etc. can be mentioned. The therapeutic or prophylactic effect of the agent of the present invention on neurodegenerative diseases can be evaluated, for example, as shown in the examples described later, by administering the agent to a model animal that develops a neurodegenerative disease (for example, degenerative dementia) and then subjecting it to a cognitive function test such as the Morris water maze test or a motor function test such as the rotarod test, or by staining and observing brain sections.

[0042] [Dosage form, usage and dosage of agents] The dosage form of the agent of the present invention is not particularly limited. For example, crude drugs selected from Ziziphus jujuba var. spinosa and Acorus tatarinowii can be used as they are in any form such as crushed materials, extracts of crushed materials, extraction residues of crushed materials, etc., or formulated together with other components such as desired excipients and / or carriers. Further, when used in the form of a food (the food of the present invention) or a medicine (the medicine of the present invention) containing the agent of the present invention, Ziziphus jujuba var. spinosa and / or Acorus tatarinowii, which are the active ingredients of the agent of the present invention, can be mixed and used together with other components according to the form of the food or medicine, respectively. Details will be described later.

[0043] The usage of the agent of the present invention is not particularly limited either, but it is usually administered orally. In particular, the agent of the present invention is preferably used in the form of a food (the food of the present invention) or an oral medicine (the medicine of the present invention). Details will be described later.

[0044] The dosage of the agent of the present invention is not particularly limited either. For example, the agent of the present invention can be administered to a subject such that Ziziphus jujuba var. spinosa and / or Acorus tatarinowii, which are its active ingredients, are in an amount of usually 0.05 mg / day or more, particularly 0.5 mg / day or more, more particularly 1.0 mg / day or more, and usually 10 g / day or less, particularly 5 g / day or less, more particularly 1 g / day or less.

[0045] [Food] According to one aspect of the present invention, a food (the food of the present invention) containing the agent of the present invention is provided. The food of the present invention is characterized by containing any one of the agents of the present invention and is used for the use of the agent.

[0046] The food of the present invention can be in any form that can be orally ingested, such as a solution, suspension, emulsion, powder, solid molded product, etc. Further, in the same manner as the medicine of the present invention described later, it can be formed into dosage forms such as capsules, lozenges, syrups, granules, etc.

[0047] The food of the present invention can be manufactured as beverages such as tea, black tea, coffee, soft drinks, alcoholic beverages, carbonated beverages, milk beverages, fruit juice beverages, nutritional drinks, concentrated beverages, powdered beverages (such as powdered juice, powdered soup, etc.); supplements; confectioneries such as candies, gummies, gums, chocolates, cookies, biscuits, etc.; frozen desserts such as ice cream; dairy products such as yogurt, processed milk, etc.; wheat flour products such as cereals, bread, cake mixes, etc.; noodles such as buckwheat noodles; oil processed products such as mayonnaise, whipped cream, dressings, etc.; fishery processed products; livestock processed products; and agricultural processed products. By adding and containing the agent of the present invention during the manufacture of these foods, the food of the present invention can be manufactured.

[0048] In addition to other food materials, the food of the present invention may contain, as necessary, sweeteners, colorants, preservatives, thickeners, stabilizers, gelling agents or sizing agents, antioxidants such as ascorbic acid, color developers, bleaching agents, fungicides or insect repellents, yeast food, gum base, potassium carbonate, bitter agents, enzymes, brighteners, fragrances, acidulants, chewing gum softeners, seasonings, coagulants for tofu, emulsifiers, pH adjusters, swelling agents, vitamins, minerals, nutritional fortifiers such as amino acids, and additives such as manufacturing agents.

[0049] The food of the present invention may be provided as a food indicating the action, effect, function, or use of the agent of the present invention under the systems of various countries. For example, in Japan, the food of the present invention can also be manufactured as a health functional food (specific health functional food, functionally labeled food, nutritional functional food).

[0050] The content of Ziziphus jujuba and / or Acorus tatarinowii, which are the active ingredients of the agent of the present invention in the food of the present invention, can be appropriately set within the range in which the effects of the present invention can be obtained in the same manner as the agent of the present invention. Specifically, the content may be adjusted so that Ziziphus jujuba and / or Acorus tatarinowii, which are the active ingredients of the agent of the present invention, are usually 0.1 mg / day or more, particularly 0.5 mg / day or more, more preferably 1.0 mg / day or more, and usually 10 g / day or less, particularly 5 g / day or less, more preferably 1 g / day or less.

[0051] The number of times and frequency of taking the food of the present invention are arbitrary, and can be appropriately set as 1 to several times a day, daily, every other day, every two days, or any number of times and frequency from 1 to 7 days a week. According to the desired number of times and frequency of taking, by blending an amount of Ziziphus jujuba seeds and / or Acorus tatarinowii rhizome necessary to obtain the effect of the agent of the present invention into the food, a food of the present invention capable of expecting a desired effect according to the type of desired action effect can be provided.

[0052] Since the food of the present invention is characterized by containing the agent of the present invention, it shares the advantages of various agents of the present invention and is extremely useful. In addition, since it is a food, it can be safely and easily used not only by patients suffering from specific diseases but also by healthy people.

[0053] In addition, the food of the present invention can be used not only for humans but also for animals other than humans to which the use of the agent of the present invention can be applied.

[0054] [Medicine] According to one aspect of the present invention, a medicament (the medicament of the present invention) containing the agent of the present invention is provided. The medicament of the present invention is characterized by containing any one of the agents of the present invention and is used for the use of the agent.

[0055] The administration route of the medicament of the present invention is not limited, but it is usually an oral medicine (oral agent) administered orally.

[0056] The medicament of the present invention can be manufactured as an arbitrary dosage form by adding the agent of the present invention as an active ingredient, such as capsule agents such as tablets, soft capsules, and hard capsules, solid preparations such as powders, granules, drops, and pills, semi-solid preparations such as jelly, and liquid preparations such as syrups, suspensions, and internal use liquids. In this case, the medicament of the present invention can be formulated as a pharmaceutical composition combining the agent of the present invention and other additives usually used in the manufacture of oral agents by a method for manufacturing pharmaceuticals known to those skilled in the art.

[0057] Examples of additives used in the production of the medicament of the present invention include excipients, disintegrants, binders, lubricants, coating agents, dispersants, fluidizing agents, stabilizers, preservatives, buffers, flavoring agents, suspending agents, emulsifiers, flavoring agents, solubilizing agents, coloring agents, thickening agents, and the like. Further, by further combining a pharmaceutically acceptable carrier, the medicament of the present invention having enhanced action and effect of the agent of the present invention can be provided.

[0058] The medicament of the present invention includes products conforming to pharmaceuticals under the systems of various countries. Examples of this include quasi-drugs in Japan.

[0059] The content of zizyphi spinosae semen and / or acori graminei rhizoma, which are the active ingredients of the agent of the present invention in the medicament of the present invention, can be appropriately set within the range in which the effects of the present invention can be obtained in the same manner as the agent of the present invention. Specifically, the content of zizyphi spinosae semen and / or acori graminei rhizoma, which are the active ingredients of the agent of the present invention, is usually 0.1 mg / day or more, particularly 0.5 mg / day or more, more preferably 1.0 mg / day or more, and usually 10 g / day or less, particularly 5 g / day or less, more preferably 1 g / day or less.

[0060] The number of administrations and the frequency of the medicament of the present invention are arbitrary and can be appropriately set at 1 to several times a day, daily, every other day, every two days, or at any number and frequency of 1 to 7 days per week. Depending on the desired number of administrations and frequency, by formulating an amount of zizyphi spinosae semen and / or acori graminei rhizoma necessary to obtain the effect of the agent of the present invention, an administration of the present invention that can expect a desired effect according to the type of desired action effect can be provided.

[0061] Since the medicament of the present invention is characterized by containing the agent of the present invention, it shares the advantages of the agent and is extremely useful.

[0062] In addition to humans, the medicament of the present invention can also be used for animals other than humans to which the use of the agent of the present invention can be applied.

[0063] [Method] According to one aspect of the present invention, there is provided a method for maintaining or improving brain function, promoting nerve cell repair or inducing neurogenesis, removing dementia-causing proteins accumulated in the brain, or treating or preventing neurodegenerative diseases (the method of the present invention), which includes administering to a subject one or more selected from crude drugs selected from Semen Ziziphi Spinosae and the stem part of Acori Tatarinowii Rhizoma, the agent of the present invention, the food of the present invention, and the medicine of the present invention.

[0064] For the method of the present invention, Semen Ziziphi Spinosae and / or the stem of Acori Tatarinowii Rhizoma, which are the active ingredients of the agent of the present invention, may be administered to the subject in the form of the agent of the present invention, the food of the present invention, and / or the medicine of the present invention. Details thereof are as described in detail in the above description regarding the agent of the present invention, the food of the present invention, and the medicine of the present invention.

Examples

[0065] Hereinafter, the present invention will be described in more detail with reference to examples. However, these examples are merely examples shown for convenience of explanation and the present invention is not limited to these examples in any sense.

[0066] [Materials and methods] · Preparation of hot water extracts, ethanol extracts, extraction residues, and crushed materials of Ziziphus jujuba Mill. Spinosae Semen (ZSS) and Acori Tatarinowii Rhizoma (AGS), and hot water extract of Acori Tatarinowii Radix (AGR) The preparation of the hot water extract (Ext-ZSS) of Semen Ziziphi Spinosae in Example Group A was carried out as follows. 1 kg of dried Semen Ziziphi Spinosae (ZSS) (origin: Hebei Province, China) was crushed, 10 kg of water was added and boiled for 1 hour for hot water extraction. Then it was filtered, the filtrate was concentrated and heat sterilized, 40 g of dextrin was added, and dried to obtain 200 g of Semen Ziziphi Spinosae extract.

[0067] The preparation of the ethanol extract (EtOH-ext-Z) of Ziziphus jujuba seeds in Example Group A was carried out as follows. To crushed Ziziphus jujuba seeds (200 g), 50% ethanol water (900 g) was added and hot water extraction was performed at 90 °C for 1 hour, followed by filtration to obtain a filtrate. To the remaining residue, 50% ethanol water (900 kg) was added and hot water extraction and filtration were performed again under the same conditions. The filtrates were combined, concentrated, heat sterilized (80 °C, 30 minutes), passed through a 30-mesh filter, and then freeze-dried to obtain the uncrushed ethanol water extract powder of Ziziphus jujuba seeds (24.5 g).

[0068] The preparation of the extraction residue (Res-ZSS) of Ziziphus jujuba seeds in Example A3 and the crushed powder (Cru-ZSS) of Ziziphus jujuba seeds in Examples A3, A5, and A6 was carried out according to the following procedure. 5.3 kg of dry Ziziphus jujuba seeds obtained from Auropure Life Science Co, Ltd. (Zhuzhou, Hunan, China) were sterilized (115 °C, 1.5 hours), finely pulverized using a hammer mill, and passed through a 3-mm screen to obtain 2.1 kg of coarsely crushed powder. The obtained coarsely crushed powder (2.1 kg) was sieved through a vibrating sieve with an opening of 500 μm to obtain 1.3 kg of dry crushed powder. To the above-mentioned dry crushed powder of Ziziphus jujuba seeds (10 g), water (140 ml) was added, and hot water extraction was performed at 90 - 95 °C for 3 hours. Then, it was filtered and the residue was dried under reduced pressure at 40 °C overnight to obtain the extraction residue (8.29 g).

[0069] The preparation of the hot water extract (Ext-AGR) of Acorus tatarinowii roots and the hot water extract (Ext-AGS) of Acorus tatarinowii stems in Example B1 was carried out as follows. 500 g of dry roots and 400 g of dry stems of Acorus gramineus Solander (origin: Hubei Province, China) were cut into small pieces and added to 8 times and 30 times the amount of water, respectively. Each mixture was heated at 90 °C for 1 hour and passed through a 32-mesh and then a 200-mesh filter. The filtrates were collected, and the residues were extracted by heating two more times. The filtrates extracted three times each were combined, concentrated, and freeze-dried to obtain a viscous liquid extract. The yield of the extract of the hot water extract (Ext-AGS) of Acorus tatarinowii stems was 130 g, and the solid content was 50.0% by mass. The yield of the hot water extract (Ext-AGR) of Acorus tatarinowii roots was 98 g, and the solid content was 52.2% by mass.

[0070] The preparation of the extraction residue (Res-AGS) and crushed powder (Cru-AGS) of Acorus tatarinowii rhizome in Example B3 was carried out according to the following procedure. The dried Acorus tatarinowii rhizome (5.3 kg) was coarsely crushed using a cutter mill, passed through a 2-mm screen, and 5.1 kg of coarsely crushed product was obtained. The coarsely crushed product was sterilized (170 °C, 3 s), finely crushed with a ball mill, and then screened through a vibrating sieve with an opening of 500 μm to obtain 4.0 kg of dried crushed powder. To the above-mentioned dried crushed powder of Acorus tatarinowii rhizome (50 g), water (700 ml) was added, and hot water extraction was carried out at 90 - 95 °C for 3 hours. Then, it was filtered, and the residue was dried under reduced pressure at 40 °C overnight to obtain the extraction residue (43.41 g).

[0071] · Component analysis of Ziziphus jujuba Mill. Spinosae Semen (ZSS) The component analysis of Ziziphus jujuba seed (ZSS) was entrusted to the Japan Food Analysis Center (Tokyo, Japan). Ziziphus jujuba seed (ZSS) is known to contain jujuboside A, jujuboside B, and spinosin as the main components (Non-Patent Document 27: Liu et al., Talanta, (2007), 71:668 - 675, Non-Patent Document 28: Wang et al., J. Pharm. Anal., (2019), 9[6]:406 - 413). 0.2 g of the crushed powder of Ziziphus jujuba seed (ZSS) was suspended in 30 mL of 50% ethanol and shaken for 10 minutes. After centrifugation, the supernatant was collected, and the pellet was extracted with ethanol two more times. The supernatants obtained from the three extractions were combined to obtain a total of 100 mL of the extract of Ziziphus jujuba seed (ZSS).

[0072] The detection of jujuboside A and B was carried out by diluting the extract of Ziziphus jujuba seed (ZSS) 10-fold and then separating it by high-performance liquid chromatography (HPLC). As the column, a reverse-phase InertSustain C18 column (GL Sciences, Tokyo, Japan) was used, and as the mobile phase, a mixed solvent of 0.1% formic acid and acetonitrile (63:37) was used. Each fraction was sequentially analyzed by electrospray ionization mass spectrometry (ESI-MS) using Xevo TQ MS (Waters Corporation, Milford, MA).

[0073] The detection of spinosin was performed by separating the extract of Ziziphus jujuba seeds (ZSS) using high-performance liquid chromatography (HPLC). As the column, a reversed-phase Unison UK-C18 column (Imtakt USA, Portland, Oregon, USA) was used, and as the mobile phase, a mixed solvent of 0.1% formic acid and methanol (65:35) was used. The absorbance at 270 nm of each fraction was measured.

[0074] [Mouse] Six different mouse models of neurodegenerative dementia were used.

[0075] APP23 mice are an AD model that expresses human APP with the Swedish (KM670 / 671NL) mutation (Non-Patent Document 29: Stalder et al., Am. J. Pathol., (1999), 154[6]:1673-1684). These mice show memory impairment at 3 months of age. According to the past research of the present inventors, these mice show the accumulation of Aβ oligomers, synaptic loss, and amyloid deposition at 15 months of age.

[0076] Tau784 mice are an FTD model that expresses both 3-repeat and 4-repeat human tau at adult age due to the presence of the tau intron mutation (in which 4-repeat human tau is dominantly expressed) (Non-Patent Document 30: Umeda et al., Am. J. Clin. Pathol., (2013), 183[1]:211-225). According to the past research of the present inventors, these mice show hyperphosphorylation of tau, tau oligomer formation, synaptic loss, and memory impairment at 6 months of age due to the unbalanced expression of tau isoforms, show microglial activation at 12 months of age, and show the formation of neurofibrillary changes and neuron loss at 15 months of age.

[0077] The Huα-Syn(A53T) system G2-3 mice were originally created as a PD model that expresses human α-synuclein with the A53T mutation (Non-Patent Document 31: Rota et al., Transl. Neurodegener., (2019), 8:5). According to the past research of the present inventors, these mice show the accumulation of α-synuclein oligomers from 4 months after birth, and show cognitive impairment at 6 months after birth and motor dysfunction at 9 months after birth. Therefore, it can be regarded as a model of DLB until 9 months after birth.

[0078] APPOSK mice are transgenic (Tg) mice that express Osaka mutant human APP and were created as an AD model that does not form amyloid plaques but induces amyloid-β oligomers (Non-Patent Document 32: Tomiyama et al., J. Neurosci., (2010), 30

[14] :4845-4856).

[0079] OSK-K1 mice are knock-in mice in which the Osaka mutation was introduced into the mouse APP gene and were created as an AD model (Non-Patent Document 33: Umeda et al., Acta Neuropathol. Commun., (2017), 5:59).

[0080] The C9-500 mouse is an FTD / ALS model mouse in which the full-length sequence of C9orf72 was introduced into intron 1a (Non-Patent Document 36: Liu et al., Neuron., (2016), 90(3): 521-34). According to the past research of the present inventors, accumulation of phosphorylated TDP-43 was observed at 3 months after birth, and synaptic loss, neuronal dropout, and activation of microglia were observed after 6 months after birth (Non-Patent Document 37: Hatanaka Y et al., Biomedicines., (2022), 10(5):1080). Cognitive function begins to decline at 4.5 months after birth, but motor function remains normal until 12 months after birth. Therefore, the model mouse can be regarded as an FTD-TDP model until 12 months after birth. Transgenic mice (Tg) were mated with wild-type FVB / N Jc1 mice to maintain heterozygotes of the transgene.

[0081] All transgenic (Tg) mice were maintained and used as heterozygous animals. All animal experiments were approved by the Ethics Committee of Osaka Metropolitan University (Osaka, Japan) and were conducted in accordance with the Osaka Metropolitan University Animal Experiment Guide.

[0082] [Treatment of mice] To compare the effects of the hot water extract of Acorus calamus rhizome (Ext-AGS) and the hot water extract of Acorus calamus root (Ext-AGR), suspensions were prepared by diluting each extract with a solid content of about 50% by mass to 2.5 mg / mL with water. 400 μL of each suspension (i.e., 0.5 mg of solid content) was orally administered to male and female Tau784 mice 5 days a week (Monday to Friday) for 1 month. As a control, the same amount of water was administered to age-matched Tg and non-Tg littermates.

[0083] To examine the effects of Ziziphus jujuba seed (ZSS) and Acorus gramineus stolon (AGS), the hot water extract (Ext-ZSS), crushed powder (Cru-ZSS), and extraction residue (Res-ZSS) of Ziziphus jujuba seed, as well as the hot water extract (Ext-AGS), crushed powder (Cru-AGS), and extraction residue (Res-AGS) of Acorus gramineus stolon, were suspended in water by ultrasonic treatment to a concentration of 0.33 or 1.65 mg / mL. 300 μL of each suspension (i.e., 0.1 mg or 0.5 mg of powder) was orally administered to male and female Tau784, APP23, C9-500, and Huα-Syn (A53T) mice 5 days a week for 1 month.

[0084] To test the mixture of the main components of Ziziphus jujuba seed (ZSS), jujuboside A, jujuboside B, and spinosin (all from Biosynth, Compton, Berkshire, UK) were dissolved in water to concentrations of 1.52 g / mL, 0.667 g / mL, and 2.33 g / mL, respectively, to prepare a mixed solution. 300 μL of the resulting mixed solution was administered to Tau784 mice for 1 month. The dosage of each component corresponded to the amount of each component in 0.5 mg of the hot water extract of Ziziphus jujuba seed (Ext-ZSS), i.e., 0.455 g of jujuboside A, 0.2 g of jujuboside B, and 0.7 g of spinosin.

[0085] To evaluate neurogenesis, the thymidine analog 5-bromo-2'-deoxyuridine (BrdU; Sigma-Aldrich), which is selectively incorporated into the DNA of proliferating cells, was dissolved in Tris-buffered saline (pH 7.6) to a concentration of 5 mg / mL. 300 μL of the BrdU solution (i.e., 1.5 mg) was intraperitoneally injected into Huα-Syn (A53T) mice during the last 5 days of the treatment with the crushed powder.

[0086] [Behavioral test] The spatial reference memory of mice was evaluated using the Morris water maze test (Non-Patent Document 34: Kelley Bromley-Brits et al., J. Vis. Exp., (2011), 53e2920). A platform with a diameter of 10 cm was placed at a depth of 1 cm below the water surface in a circular pool with a diameter of 1 m in a location where it could not be seen by the mice. The mice were placed in the pool and allowed to swim for 60 seconds, and the time taken to reach the platform was measured (expressed as Escape latency). If they did not reach it within 60 seconds, the experimenter placed the mice on the platform and allowed them to rest for a while. This trial was conducted 5 times a day at 5-minute intervals for 4 consecutive days for each mouse. Thereby, the ability to acquire memory was measured.

[0087] The motor function of mice was evaluated using the rotarod test with an MK-610 mouse rotarod treadmill (Muromachi Kikai, Tokyo). The mice were placed on a rod (rotation axis) that rotated 5 times per minute, and trained for 180 seconds to keep walking without falling. Next, training was conducted twice under the condition of increasing the rotation speed from 4 rotations per minute to 40 rotations per minute over 240 seconds. The next day, the accelerating rotarod test was conducted twice at 1-hour intervals under the same conditions. The time when the mice fell off the rod was measured, and the average value of the two times was calculated.

[0088] [Histological analysis of neuropathology] After the behavioral tests, the mice in each group were divided into two, one for histological analysis and the other for subsequent biochemical analysis. Brain sections were prepared according to the previously reported method (Non-Patent Document 32: Tomiyama et al., J. Neurosci., (2010), 30

[14] :4845 - 4856). Phosphorylated tau and tau oligomers were stained with mouse monoclonal AT8 antibody (Thermo Scientific, Waltham, Massachusetts, USA) and rabbit polyclonal T22 antibody (Sigma-Aldrich), respectively. Aβ oligomers and amyloid deposits were stained with mouse monoclonal 11A1 antibody (IBL, Fujinomiya, Japan) and rabbit polyclonal β001 antibody (Non-Patent Document 35: Lippa et al., Arch Neurol., (1999), 56:1111 - 1118), respectively. Phosphorylated α-synuclein and α-synuclein oligomers were stained with rabbit monoclonal anti-α-synuclein (phospho S129) antibody (EP1536Y; Abcam, Cambridge, UK) and rabbit polyclonal Syn33 antibody (Sigma-Aldrich), respectively. Synaptophysin was stained with a mouse monoclonal antibody against synaptophysin (SVP-38; Sigma-Aldrich). TDP-43 was stained with a mouse monoclonal antibody against phosphorylated TDP-43 (pSer409 / 410-TDP-43; Cosmo Bio). The staining intensity or positive area of a certain brain region was quantified using NIH ImageJ software.

[0089] [Histological analysis of BDNF expression and neurogenesis] BDNF expression was evaluated in Tau784 mice administered with each crushed powder of Ziziphus jujuba seeds and Acorus gramineus rhizomes (Cru-ZSS and Cru-AGS). Brain sections were stained with a mouse monoclonal anti-BDNF antibody (#9; DSHB, Iowa City, Iowa). The staining intensity of a certain brain region was quantified using NIH ImageJ software.

[0090] Neurogenesis was evaluated in Huα-Syn(A53T) mice administered with crushed powders of Ziziphi Spinosae Semen and Acori Tatarinowii Rhizoma (Cru-ZSS and Cru-AGS). Brain sections were double-stained with mouse monoclonal anti-BrdU (IBL) and rabbit polyclonal anti-doublecortin antibody (Abcam). Cells positive for both BrdU and doublecortin in the stationary brain regions were counted as newly generated neurons.

[0091] [Statistical analysis] For comparison of the means among three or more groups, ANOVA or two-way repeated measures ANOVA (for behavioral tests) was used, followed by Fisher's PLSD test. A p-value < 0.05 was considered a significant difference.

[0092] [Example group A: Examination of Ziziphus jujuba Mill. Spinosae Semen (ZSS)] · Example A1: Examination of the effect of hot water extract (Ext-ZSS) of Ziziphus jujuba Mill. Spinosae Semen on Tau784 mice The effect of Ziziphi Spinosae Semen (ZSS) on Tau784 mice was examined. A hot water extract of Ziziphi Spinosae Semen (Ext-ZSS) was orally administered to 14-month-old Tau784 mice (average body weight 35.1 g) at 0.1 and 0.5 mg / day for 1 month.

[0093] Figure 1 is a graph showing the results of the Morris water maze test of Tau784 mice administered with a hot water extract of Ziziphi Spinosae Semen (Ext-ZSS) at 0.1 mg / day and 0.5 mg / day, compared with Tau784 mice administered with water and non-transgenic mice. The memory of the mice was improved in a dose-dependent manner. That is, at the high dose, it completely recovered to the level similar to that of non-Tg littermates, while at the low dose, only a moderate effect was shown.

[0094] Figure 2 shows the tau pathology in the CA2 / 3 region of the hippocampus of Tau784 mice administered with the hot water extract of Ziziphus jujuba var. spinosa (Ext-ZSS) at 0.1 mg / day and 0.5 mg / day, compared with Tau784 mice administered with water and non-transgenic mice. Figure 2A is a photograph showing the staining results of phosphorylated tau and tau oligomers, and Figure 2B is a graph showing the results of quantifying the staining intensity of each photograph. According to the evaluation results of tau pathology in the hippocampus using immunohistochemistry, the levels of phosphorylated tau and tau oligomers were significantly decreased in a dose-dependent manner.

[0095] Figure 3 shows the synaptopodin pathology in the mossy fibers of the CA2 / 3 region of the hippocampus of Tau784 mice administered with the hot water extract of Ziziphus jujuba var. spinosa (Ext-ZSS) at 0.1 mg / day and 0.5 mg / day, compared with Tau784 mice administered with water and non-transgenic mice. Figure 3A is a photograph showing the staining results of synaptopodin, and Figure 3B is a graph showing the results of quantifying the staining intensity of each photograph. The synaptopodin levels in the CA2 / 3 region of the hippocampus also significantly recovered in a dose-dependent manner.

[0096] · Example A2: Examination of the effect of hot water extract (Ext-ZSS) of Ziziphus jujuba Mill. Spinosae Semen on APP23 mice The effect of Ziziphus jujuba var. spinosa (ZSS) on APP23 mice was investigated. The hot water extract of Ziziphus jujuba var. spinosa (Ext-ZSS) was orally administered to 13- to 15-month-old APP23 mice (average body weight 28.9 g) at 0.1 mg / day for 1 month. In addition, the hot water extract of Ziziphus jujuba var. spinosa (Ext-ZSS) was orally administered to 15- to 16-month-old APP23 mice (average body weight 28.9 g) at 0.5 mg / day for 1 month.

[0097] Figure 4 is a graph showing the results of the Morris water maze test of APP23 mice administered with the hot water extract of Ziziphus jujuba seeds (Ext-ZSS), compared with APP23 mice administered with water and non-transgenic mice. Figure 4A is a graph showing the results of administration at 0.1 mg / day, and Figure 4B is a graph showing the results of administration at 0.5 mg / day. The hot water extract of Ziziphus jujuba seeds (Ext-ZSS) improved the memory of mice at any dose. In particular, the hot water extract of Ziziphus jujuba seeds (Ext-ZSS) at 0.5 mg / day significantly improved the memory of mice to the same level as that of non-Tg littermates.

[0098] Figure 5 is a diagram showing the amyloid-β oligomer pathology in the cerebral cortex (CC) and hippocampus (HC) of APP23 mice administered with the hot water extract of Ziziphus jujuba seeds (Ext-ZSS) at 0.1 mg / day, compared with APP23 mice administered with water. Figure 5A is a photograph showing the staining results of amyloid-β oligomers, and Figure 5B is a graph showing the results of quantifying the staining intensity of each photograph. The hot water extract of Ziziphus jujuba seeds (Ext-ZSS) significantly reduced the levels of Aβ oligomers in the cerebral cortex and hippocampus.

[0099] Figure 6 is a diagram showing the amyloid deposition pathology in the cerebral cortex (CC) and hippocampus (HC) of APP23 mice administered with the hot water extract of Ziziphus jujuba seeds (Ext-ZSS) at 0.1 mg / day, compared with APP23 mice administered with water. Figure 6A is a photograph showing the staining results of amyloid deposition, and Figure 6B is a graph showing the results of quantifying the staining intensity of each photograph. The hot water extract of Ziziphus jujuba seeds (Ext-ZSS) significantly decreased amyloid deposition in both the cerebral cortex (CC) and hippocampus (HC).

[0100] Figure 7 shows the synaptopodin pathology in the hippocampal CA2 / 3 region of APP23 mice administered with the hot water extract of Ziziphus jujuba seeds (Ext-ZSS) at 0.1 mg / day, compared with APP23 mice administered with water and non-transgenic mice. Figure 7A is a photograph showing the staining results of synaptopodin, and Figure 7B is a graph showing the results of quantifying the staining intensity of each photograph. The hot water extract of Ziziphus jujuba seeds (Ext-ZSS) significantly decreased the synaptopodin level in the hippocampal CA2 / 3 region.

[0101] · Example A3: Examination of the effects of hot water extract (Ext-ZSS), crushed powder (Cru-ZSS), and extraction residue (Res-ZSS) of Ziziphus jujuba Mill. Spinosae Semen on Tau784 mice In many cases of traditional Chinese herbal medicines, herbs are used as decoctions, and their extraction residues are usually discarded. However, in hot water extraction, heat-sensitive components may be decomposed, and volatile substances may be lost by evaporation. Therefore, in order to examine whether there are differences in effects in a given form, the effects of the hot water extract of Ziziphus jujuba seeds (Ext-ZSS), crushed powder (Cru-ZSS), and extraction residue (Res-ZSS) on Tau784 mice were examined.

[0102] Specifically, to compare the hot water extract of Ziziphus jujuba seeds (Ext-ZSS) with the crushed powder (Cru-ZSS), Tau784 mice aged 8 - 12 months (average body weight 29.3 g) were administered at 0.1 mg / day for 1 month. Also, to compare the crushed powder of Ziziphus jujuba seeds (Cru-ZSS) with the extraction residue (Res-ZSS), Tau784 mice aged 8 - 11 months (average body weight 31.2 g) were administered at 0.1 mg / day for 1 month.

[0103] Figure 8 is a graph showing the results of the Morris water maze test of Tau784 mice administered with the hot water extract (Ext-ZSS), crushed powder (Cru-ZSS), and extraction residue (Res-ZSS) of Ziziphus jujuba seed at 0.1 mg / day, in comparison with Tau784 mice administered with water as a control and non-transgenic mice. Figure 8A is a graph showing the results of the test comparing the hot water extract (Ext-ZSS) and the crushed powder (Cru-ZSS). The hot water extract (Ext-ZSS) improved the memory of the mice, but the effect was incomplete. In contrast, the crushed powder (Cru-ZSS) significantly enhanced the memory of the mice to a level even higher than that of non-Tg littermates. Figure 8B is a graph showing the results of the test comparing the extraction residue (Res-ZSS) and the crushed powder (Cru-ZSS). Also in this evaluation, the crushed powder (Cru-ZSS) showed an extremely excellent memory improvement effect. On the other hand, in the extraction residue (Res-ZSS), similar to the hot water extract (Ext-ZSS), the memory was improved, but the effect was incomplete. These results support the speculation of the inventors that when medicinal plants are made into decoctions, their functional components may be lost.

[0104] Figure 9 is a figure showing tau pathology in the hippocampal CA2 / 3 region of Tau784 mice administered with the hot water extract (Ext-ZSS), crushed powder (Cru-ZSS), and extraction residue (Res-ZSS) of Ziziphus jujuba seed at 0.1 mg / day, in comparison with Tau784 mice administered with water as a control and non-transgenic mice. Figure 9A is a photograph showing the staining results of phosphorylated tau and tau oligomers, and Figure 9B is a graph showing the results of quantifying the staining intensity of each photograph. The levels of phosphorylated tau and tau oligomers were significantly decreased by all of the hot water extract (Ext-ZSS), crushed powder (Cru-ZSS), and extraction residue (Res-ZSS), but the crushed powder (Cru-ZSS) showed the strongest effect.

[0105] Figure 10 shows the synaptophysin and BDNF pathology in the hippocampal CA2 / 3 region of Tau784 mice administered with the hot water extract (Ext-ZSS), crushed powder (Cru-ZSS), and extraction residue (Res-ZSS) of Ziziphus jujuba var. spinosa at 0.1 mg / day, in comparison with Tau784 mice administered with water and non-transgenic mice. Figure 10A is a photograph showing the staining results of synaptophysin and BDNF, and Figure 10B is a graph showing the results of quantifying the staining intensity of each photograph. The synaptophysin level in the hippocampal CA2 / 3 region was only slightly affected by the hot water extract (Ext-ZSS) and extraction residue (Res-ZSS), while the crushed powder (Cru-ZSS) significantly recovered to a level equivalent to that of non-Tg littermates. The expression level of BDNF in the hippocampus was also only slightly affected by the hot water extract (Ext-ZSS) and extraction residue (Res-ZSS), while the crushed powder (Cru-ZSS) significantly increased to a level even higher than that of non-Tg littermates.

[0106] · Example A4: Examination of the effects of the main components jujuboside A, jujuboside B, and spinosin of Ziziphus jujuba Mill. Spinosae Semen (ZSS) on Tau784 mice Jujuboside A, jujuboside B, and spinosin, which are the main components of Ziziphus jujuba var. spinosa (ZSS), have all been reported to have anti-dementia effects. Therefore, it was speculated that the reason why the crushed powder (Cru-ZSS) of Ziziphus jujuba var. spinosa showed a stronger effect on tau than the hot water extract (Ext-ZSS) was that the crushed powder (Cru-ZSS) contained more of these components than the hot water extract (Ext-ZSS).

[0107] To verify this possibility, the amounts of three components in the hot water extract (Ext-ZSS) and crushed powder (Cru-ZSS) of Ziziphus jujuba var. spinosa were analyzed. The results are shown in Table 1 below. Surprisingly, despite the strong dementia-suppressing effect of the crushed powder (Cru-ZSS), its content of these components was lower than that of the hot water extract (Ext-ZSS). These results suggest that the effect of the crushed powder (Cru-ZSS) of Ziziphus jujuba var. spinosa is due to components other than these.

[0108]

Table 1

[0109] Therefore, to evaluate the contributions of jujuboside A, jujuboside B, and spinosin to the improvement of mouse cognitive function, these compounds were dissolved in water to prepare a mixed solution containing the final contents corresponding to 0.5 mg of the hot water extract of Ziziphus jujuba seeds (Ext-ZSS), namely, 0.455 g of jujuboside A, 0.2 g of jujuboside B, and 0.7 g of spinosin in 300 μL, and this was administered to Tau784 mice (average body weight 31.9 g) aged 13 - 16 months for 1 month.

[0110] Figure 11 is a graph showing the results of the Morris water maze test of Tau784 mice administered a mixed solution of jujuboside A, jujuboside B, and spinosin, which are the three main components of Ziziphus jujuba seeds, in comparison with Tau784 mice administered water as a control and non - genetically modified mice. The daily doses of jujuboside A, jujuboside B, and spinosin in the mixed solution are 0.455 g, 0.2 g, and 0.7 g, respectively, and these correspond to the amounts of each component in 0.5 mg of the hot water extract of Ziziphus jujuba seeds (Ext-ZSS). The effect of the mixed solution of jujuboside A, jujuboside B, and spinosin on the memory of mice was much weaker than the effect of 0.5 mg of the hot water extract of Ziziphus jujuba seeds (Ext-ZSS) (Figure 1). These results suggest that Ziziphus jujuba seeds (ZSS) contain active substances other than jujuboside A, jujuboside B, and spinosin, and most of them may be lost during hot water extraction.

[0111] · Example A5: Examination of the effect of crushed powder (Cru-ZSS) of Ziziphus jujuba Mill. Spinosae Semen on Huα-Syn(A53T) mice The effects of Ziziphus jujuba seeds (ZSS) in an α-synucleinopathy model were investigated. Eight-month-old Huα-Syn (A53T) mice (average body weight, 29.8 g for ZSS) were orally administered crushed Ziziphus jujuba seeds (Cru-ZSS) at 0.1 mg / day for 1 month. Immunohistochemistry was used to evaluate α-synuclein pathology in the hippocampus. Also, since it was suggested that crushed Ziziphus jujuba seeds (Cru-ZSS) have a trophic effect on neurons (Figs. 8-10), the levels of neurogenesis in the dentate gyrus and substantia nigra were evaluated. The latter brain regions are particularly vulnerable to α-synuclein-induced neurodegeneration and cause motor dysfunction in PD.

[0112] Figure 12 shows the comparison of α-synuclein pathology in the CA2 / 3 region of the hippocampus of Huα-Syn (A53T) mice administered crushed Ziziphus jujuba seeds (Cru-ZSS) at 0.1 mg / day with Tau784 mice and non-transgenic mice administered water as controls. Figure 12A is a photograph showing the staining results of phosphorylated α-synuclein and α-synuclein oligomers, and Figure 12B is a graph showing the results of quantifying the staining intensity of each photograph. The levels of both phosphorylated α-synuclein and α-synuclein oligomers were significantly decreased by crushed Ziziphus jujuba seeds (Cru-ZSS).

[0113] Figure 13 shows the comparison of the levels of neurogenesis in the dentate gyrus (DG) and substantia nigra (SN) of Huα-Syn (A53T) mice administered crushed Ziziphus jujuba seeds (Cru-ZSS) at 0.1 mg / day with Huα-Syn (A53T) mice and non-transgenic mice administered water as controls. Figure 13A is a photograph showing the immunofluorescence staining results of BrdU (red) and doublecortin (DCX, green), and Figure 13B is a graph showing the results of counting double-positive cells (yellow) in each photograph as newly generated neurons. Crushed Ziziphus jujuba seeds (Cru-ZSS) significantly increased the number of double-positive cells of BrdU and doublecortin to a level higher than that of non-Tg littermates in both brain regions.

[0114] · Example A6: Examination of the effect of crushed powder (Cru-ZSS) of Ziziphus jujuba Mill. Spinosae Semen on Huα-Syn(A53T) mice The effects of Ziziphus jujuba seed (ZSS) on the motor function of mice were investigated. Crushing powder of Ziziphus jujuba seed (Cru-ZSS) was orally administered to 8-month-old Huα-Syn (A53T) mice (average body weight, 29.8 g for ZSS) at a dose of 0.1 mg / day for 1 month. Subsequently, the motor function was evaluated by the rotarod test.

[0115] Figure 14 is a graph showing the results of the rotarod test of Huα-Syn (A53T) mice administered with crushing powder of Ziziphus jujuba seed (Cru-ZSS) at a dose of 0.1 mg / day in Example A6, compared with Huα-Syn (A53T) mice administered with water and non-transgenic mice. The crushing powder of Ziziphus jujuba seed (Cru-ZSS) significantly increased the motor function of Huα-Syn (A53T) mice to a level close to that of non-Tg littermates.

[0116] · Example A7: Comparison of the effects of hot water extract (HOT water ext-Z) and ethanol extract (EtOH ext-Z) of Ziziphus jujuba Mill. Spinosae Semen on APPOSK mice The effects of hot water extraction and ethanol extraction of crushed Ziziphus jujuba seed on APP / PS1 mice were investigated. A group of 10 APP / PS1 mice at 12 - 16 months of age was divided into three groups of 10 mice each. One group was administered with the hot water extract of crushed Ziziphus jujuba seed at a dose of 0.1 mg / day, one group was administered with the ethanol extract of crushed Ziziphus jujuba seed at a dose of 0.1 mg / day, and the remaining one group was orally administered with 300 μL of water 5 days a week for a total of 1 month. Non-Tg mice of the same age were orally administered with the same amount of water.

[0117] Figure 15 is a graph showing the results of the Morris water maze test of APP / PS1 mice administered with the hot water extract (HOT water ext-Z) or ethanol extract (EtOH ext-Z) of Ziziphus jujuba seed, compared with APP / PS1 mice administered with water and non-transgenic mice. As long as crushed Ziziphus jujuba seed was used, there was no difference in the drug efficacy between the two, and both were equally effective.

[0118] · Example A8: Comparison of crushed and uncrushed Ziziphus jujuba Mill. Spinosae Semen OSK-KI mice at 14 to 16 months of age were divided into 4 groups of 9 mice each. One group was administered the hot water extract of crushed wild jujube seeds (Crushed-HOT water ext-Z) at 0.1 mg / day, one group was administered the hot water extract of intact wild jujube (Intact-HOT water ext-Z) at 0.1 mg / day, another group was administered the ethanol extract of intact wild jujube seeds (Intact-EtOH ext-Z) at 0.1 mg / day, and the remaining group was orally administered water, 300 μL each, 5 days a week for a total of 1 month. Non-KI mice of the same age were orally administered the same amount of water.

[0119] Figure 16 is a graph showing the results of the Morris water maze test of OSK-KI mice administered the hot water extract of crushed wild jujube seeds (Crushed-HOT water ext-Z), the hot water extract of intact wild jujube (Intact-HOT water ext-Z), or the ethanol extract of intact wild jujube seeds (Intact-EtOH ext-Z), in comparison with OSK-KI mice and non-genetically modified mice administered water as a control. Compared with the hot water extract of crushed wild jujube seeds, the hot water extract and ethanol extract of intact wild jujube were both slightly less effective. There was no difference between the hot water extract and ethanol extract of intact wild jujube seeds.

[0120] [Example group B: Examination of Acori Tatarinowii Rhizoma (AGS)] · Example B1: Examination of the effects of hot water extracts (Ext-AGS) and hot water extract (Ext-AGR) of Acori Tatarinowii Rhizoma on Tau784 mice The effects of Acorus gramineus stems (AGS) and Acorus gramineus roots (AGR) on Tau784 mice were compared. The former is used in traditional Chinese medicine, while the latter is usually discarded. Hot water extracts of Acorus gramineus stems (Ext-AGS) and Acorus gramineus roots (Ext-AGR) containing 0.5 mg of solid content were orally administered to 17-month-old Tau784 mice (average body weight 36.1 g) for 1 month.

[0121] Figure 17 is a graph showing the results of the Morris water maze test of Tau784 mice administered with the hot water extract of Acorus gramineus Soland. stems (Ext-AGS) and the hot water extract of Acorus gramineus Soland. roots (Ext-AGR) at 0.5 mg / day, in comparison with Tau784 mice administered with water as a control and non-transgenic mice. In the water maze test, both extracts significantly improved the memory of the mice. Unexpectedly, however, AGS showed a stronger effect than AGR.

[0122] Figure 18 is a diagram showing tau pathology in the hippocampal CA2 / 3 region of Tau784 mice administered with the hot water extract of Acorus gramineus Soland. stems (Ext-AGS) and the hot water extract of Acorus gramineus Soland. roots (Ext-AGR) at 0.5 mg / day, in comparison with Tau784 mice administered with water as a control and non-transgenic mice. Figure 18A is a photograph showing the staining results of phosphorylated tau and tau oligomers, and Figure 18B is a graph showing the results of quantifying the staining intensity of each photograph. According to the evaluation results of tau pathology in the olfactory cortex using immunohistochemistry, both extracts significantly reduced the level of phosphorylated tau. In addition, both extracts decreased the level of tau oligomers, but AGS showed a stronger effect than AGR.

[0123] Figure 19 is a diagram showing synaptopodin pathology in the mossy fibers of the hippocampal CA2 / 3 region of Tau784 mice administered with the hot water extract of Acorus gramineus Soland. stems (Ext-AGS) and the hot water extract of Acorus gramineus Soland. roots (Ext-AGR) at 0.5 mg / day, in comparison with Tau784 mice administered with water as a control and non-transgenic mice. Figure 19A is a photograph showing the staining results of synaptopodin, and Figure 19B is a graph showing the results of quantifying the staining intensity of each photograph. In both extracts, the level of synaptopodin in the hippocampal CA3 region recovered to the same level as that of non-Tg littermates. As far as the inventors know, these results are the first evidence indicating that AGS has an effect of improving cognitive function equal to or greater than that of AGR.

[0124] · Example B2: Examination of the effect of hot water extract (Ext-AGS) of Acori Tatarinowii Rhizoma on APP23 mice The effect of Acorus gramineus Soland. (AGS) on APP23 mice was investigated. APP23 mice at 13 - 15 months of age (average body weight 28.9 g) were orally administered a hot water extract of Acorus gramineus Soland. (Ext-AGS) at a dose of 0.1 mg / day for 1 month.

[0125] Figure 20 is a graph showing the results of the Morris water maze test of APP23 mice administered a hot water extract of Acorus gramineus Soland. (Ext-AGS) at a dose of 0.1 mg / day, compared with APP23 mice administered water as a control and non-transgenic mice. The hot water extract of Acorus gramineus Soland. (Ext-AGS) improved the memory of mice at a dose of 0.1 mg / day.

[0126] Figure 21 is a figure showing the amyloid-β oligomer pathology in the cerebral cortex (CC) and hippocampus (HC) of APP23 mice administered a hot water extract of Acorus gramineus Soland. (Ext-AGS) at a dose of 0.1 mg / day, compared with APP23 mice administered water as a control. Figure 21A is a photograph showing the staining results of amyloid-β oligomers, and Figure 21B is a graph showing the results of quantifying the staining intensity of each photograph. The hot water extract of Acorus gramineus Soland. (Ext-AGS) significantly reduced the levels of Aβ oligomers in the cerebral cortex and hippocampus.

[0127] Figure 22 is a figure showing the amyloid deposition pathology in the cerebral cortex (CC) and hippocampus (HC) of APP23 mice administered a hot water extract of Acorus gramineus Soland. (Ext-AGS) at a dose of 0.1 mg / day, compared with APP23 mice administered water as a control. Figure 22A is a photograph showing the staining results of amyloid deposition, and Figure 22B is a graph showing the results of quantifying the staining intensity of each photograph. The hot water extract of Acorus gramineus Soland. (Ext-AGS) significantly decreased amyloid deposition in both the cerebral cortex (CC) and hippocampus (HC).

[0128] Figure 23 shows the synaptophysin pathology in the hippocampal CA2 / 3 region of APP23 mice administered with the hot water extract of Acorus tatarinowii Schott (Ext-AGS) at 0.1 mg / day, in comparison with APP23 mice administered with water and non-transgenic mice. Figure 23A is a photograph showing the staining results of synaptophysin, and Figure 23B is a graph showing the results of quantifying the staining intensity of each photograph. The hot water extract of Acorus tatarinowii Schott (Ext-AGS) significantly decreased the synaptophysin level in the hippocampal CA2 / 3 region.

[0129] · Example B3: Examination of the effects of hot water extract (Ext-AGS), crushed powder (Cru-AGS), and extraction residue (Res-AGS) of Acori Tatarinowii Rhizoma on Tau784 mice In many cases of traditional Chinese herbal medicines, herbs are used as decoctions, and their extraction residues are usually discarded. However, in hot water extraction, heat-sensitive components may be decomposed, and volatile substances may be lost by evaporation. Therefore, in order to examine whether there are differences in effects in a given form, the effects of the hot water extract of Acorus tatarinowii Schott (Ext-AGS), crushed powder (Cru-AGS), and extraction residue (Res-AGS) on Tau784 mice were examined.

[0130] Specifically, in order to compare the effects of the hot water extract of Acorus tatarinowii Schott (Ext-AGS), crushed powder (Cru-AGS), and extraction residue (Res-AGS), 10-month-old Tau784 mice (average body weight 29.3 g) were administered at 0.1 mg / day for 1 month.

[0131] Figure 24 is a graph showing the results of the Morris water maze test of Tau784 mice administered with the hot water extract (Ext-AGS), crushed powder (Cru-AGS), and extraction residue (Res-AGS) of Acorus tatarinowii rhizome at 0.1 mg / day, in comparison with Tau784 mice administered with water and non-transgenic mice. Figure 24A is a graph showing the results of the hot water extract (Ext-AGS) and the crushed powder (Cru-AGS). The hot water extract (Ext-AGS) improved the memory of the mice, but the effect was incomplete. In contrast, the crushed powder (Cru-AGS) enhanced the memory of the mice to the same level as that of non-Tg littermates. Figure 24B is a graph showing the results of the extraction residue (Res-AGS) and the crushed powder (Cru-AGS). The crushed powder (Cru-AGS) achieved complete recovery, while the extraction residue (Res-AGS) showed only a moderate effect. These results support the speculation of the present inventors that when medicinal plants are made into decoctions, their functional components may be lost.

[0132] Figure 25 is a figure showing tau pathology in the hippocampal CA2 / 3 region of Tau784 mice administered with the hot water extract (Ext-AGS), crushed powder (Cru-AGS), and extraction residue (Res-AGS) of Acorus tatarinowii rhizome at 0.1 mg / day, in comparison with Tau784 mice administered with water and non-transgenic mice. Figure 25A is a photograph showing the staining results of phosphorylated tau and tau oligomers, and Figure 25B is a graph showing the results of quantifying the staining intensity of each photograph. The level of phosphorylated tau decreased significantly in the crushed powder (Cru-AGS), while only a slight decrease was observed in the hot water extract (Ext-AGS) or the extraction residue (Res-AGS). The level of tau oligomers decreased significantly by all three preparations, but the crushed powder (Cru-AGS) showed the strongest effect.

[0133] Figure 26 shows the synaptophysin and BDNF pathology in the hippocampal CA2 / 3 region of Tau784 mice administered with the hot water extract (Ext-AGS), crushed powder (Cru-AGS), and extraction residue (Res-AGS) of Acorus tatarinowii rhizome at 0.1 mg / day, in comparison with Tau784 mice administered with water and non-transgenic mice. Figure 26A is a photograph showing the staining results of synaptophysin and BDNF, and Figure 26B is a graph showing the results of quantifying the staining intensity of each photograph. The synaptophysin level in the hippocampal CA2 / 3 region was also significantly decreased by all three preparations, but only the crushed powder (Cru-AGS) recovered to a level similar to that of non-Tg littermates (Figure 5D). The BDNF level in the hippocampus increased to some extent in the crushed powder (Cru-AGS), and also slightly increased in the hot water extract (Ext-AGS) and extraction residue (Res-AGS), although none of them reached the level of non-Tg littermates.

[0134] · Example B4: Examination of the effect of crushed powder of Acorus tatarinowii Schott (Cru-AGS) on Huα-Syn (A53T) mice The effects of Acorus tatarinowii rhizome (AGS) in an α-synucleinopathy model were investigated. Eight-month-old Huα-Syn(A53T) mice (average body weight, AGS 29.0 g) were orally administered with the crushed powder of Acorus tatarinowii rhizome (Cru-AGS) at 0.1 mg / day for 1 month. The pathology of α-synuclein in the hippocampus was evaluated by immunohistochemistry. Also, since it was suggested that the crushed powder of Acorus tatarinowii rhizome (Cru-AGS) had a trophic effect on neurons (Figures 24 - 26), the levels of neurogenesis in the dentate gyrus and substantia nigra were evaluated. The latter brain region is particularly vulnerable to neurodegeneration induced by α-synuclein and causes motor dysfunction in PD.

[0135] Figure 27 shows the α-synuclein pathology in the hippocampal CA2 / 3 region of Huα-Syn(A53T) mice administered with crushed powder of Acorus tatarinowii rhizome (Cru-AGS) at 0.1 mg / day, in comparison with Tau784 mice and non-transgenic mice administered with water as a control. Figure 27A is a photograph showing the staining results of phosphorylated α-synuclein and α-synuclein oligomers, and Figure 27B is a graph showing the results of quantifying the staining intensity of each photograph. The levels of phosphorylated α-synuclein and α-synuclein oligomers were both significantly decreased by the crushed powder of Acorus tatarinowii rhizome (Cru-AGS).

[0136] Figure 28 shows the levels of neurogenesis in the dentate gyrus (DG) and substantia nigra (SN) of Huα-Syn(A53T) mice administered with crushed powder of Acorus tatarinowii rhizome (Cru-AGS) at 0.1 mg / day, in comparison with Huα-Syn(A53T) mice and non-transgenic mice administered with water as a control. Figure 28A is a photograph showing the immunofluorescence staining results of BrdU (red) and doublecortin (DCX, green), and Figure 28B is a graph showing the results of counting double-positive cells (yellow) in each photograph as newly generated neurons. The levels of neurogenesis in the dentate gyrus and substantia nigra were restored to levels similar to those of non-Tg littermates by the crushed powder of Acorus tatarinowii rhizome (Cru-AGS).

[0137] [Example Group C: Examination regarding the effect comparison between Acorus tatarinowii Schott (AGS) and Ziziphus jujuba Mill. var. spinosa (ZSS)] The effects of Acorus tatarinowii rhizome (AGS) and Ziziphus jujuba seed (ZSS) on C9-500 mice (Jackson Laboratory) were compared. A suspension in water (0.33 mg / mL) containing 0.1 mg of the crushed product of Acorus tatarinowii rhizome (Cru-AGS) and Ziziphus jujuba seed (Cru-ZSS) was orally administered to 7-month-old C9-500 mice (average body weight 31 g) for 1 month.

[0138] · Example C1: Comparative examination of the effects of Acorus tatarinowii Schott (AGS) and Ziziphus jujuba Mill. var. spinosa (ZSS) on C9-500 mice Figure 29 is a graph showing the results of the Morris water maze test on C9-500 mice administered with crushed products of Acorus tatarinowii (Cru-AGS) and Ziziphus jujuba var. spinosa (Cru-ZSS) at 0.1 mg / day. In the water maze test, both extracts significantly improved the memory of the mice. Unexpectedly, however, Acorus tatarinowii (AGS) showed a stronger effect than Ziziphus jujuba var. spinosa (ZSS).

[0139] Figure 30 is a diagram showing the comparison of TDP-43 pathology in the CA2 / 3 region of the hippocampus of C9-500 mice administered with crushed products of Acorus tatarinowii (Cru-AGS) and Ziziphus jujuba var. spinosa (Cru-ZSS) at 0.1 mg / day with C9-500 mice administered with water and non-transgenic mice as controls. Figure 30A is a photograph showing the staining results of phosphorylated TDP-43, and Figure 30B is a graph showing the results of quantifying the puncta in each photograph. Both crushed products decreased the level of TDP-43, but Acorus tatarinowii (AGS) showed a stronger effect than Ziziphus jujuba var. spinosa (ZSS).

[0140] Figure 31 is a diagram showing the comparison of synaptopodin pathology in the mossy fibers of the CA2 / 3 region of the hippocampus of C9-500 mice administered with crushed products of Acorus tatarinowii (Cru-AGS) and Ziziphus jujuba var. spinosa (Cru-ZSS) at 0.1 mg / day with C9-500 mice administered with water and non-transgenic mice as controls. Figure 31A is a photograph showing the staining results of synaptopodin, and Figure 31B is a graph showing the results of quantifying the staining intensity in each photograph. The level of synaptopodin in the CA3 region of the hippocampus significantly increased with the crushed product of Acorus tatarinowii (AGS).

[0141] · Example C2: Comparative examination of the effects of Acorus tatarinowii Schott (AGS) and Ziziphus jujuba Mill. var. spinosa (ZSS) on HuαSyn mice The effect of Acorus tatarinowii (AGS) in the α-synucleinopathy model was investigated. Six- to seven-month-old Huα-Syn(A53T) mice (average body weight, AGS 28.0 g) were orally administered with crushed powder of Acorus tatarinowii (Cru-AGS) or crushed powder of Ziziphus jujuba var. spinosa (ZSS) at 0.1 mg / day for 1 month, and the Morris water maze test was performed. The results are shown in Figure 32. In the water maze test, both extracts significantly improved the memory of the mice. Unexpectedly, however, Acorus tatarinowii (AGS) showed a stronger effect than Ziziphus jujuba var. spinosa (ZSS).

[0142] [Example Group D: Examination regarding the crushing method of Ziziphus jujuba Mill. var. spinosa (ZSS)] · Example D1: Examination of the crushing preparation method of Ziziphus jujuba Mill. var. spinosa (ZSS) 8.6 kg obtained by flash roasting the dried seeds of Ziziphus jujuba var. spinosa obtained from Auropure Life Science Co., Ltd. (Zhuzhou, Hunan, China) at 260 °C for 57 seconds was processed with a superfine grinder (Mascoloider, manufactured by Masuko Sangyo Co., Ltd., #MKZA10 - 10JLC `αH), and a coarse ground product (8.49 kg) was obtained by sieving with a sieve having an opening of 2.38 mm. Of the obtained coarse ground product (8.49 kg), 100 g was subjected to airflow pulverization (manufactured by Micro Powdertec Co., Ltd., #MP10 - 550), and then a whole grain powder (64 g) was obtained by sieving with a sieve having an opening of 0.5 mm.

[0143] The particle size distributions of the crushed powders of Ziziphus jujuba var. spinosa (ZSS) obtained by the airflow pulverization method and the crushed powders of Ziziphus jujuba var. spinosa (ZSS) obtained by the hammer mill pulverization method (used in Examples A3, A5, and A6) were analyzed. For the particle size analysis, a laser diffraction / scattering particle size distribution analyzer (LMS - 3000, manufactured by Malvern) was used. While circulating 150 mL of isopropanol, a micro spatula full of the circulating Ziziphus jujuba var. spinosa powder was prepared and added to the dispersion tank. While performing ultrasonic treatment, the light diffraction and scattering data of the sample particles were monitored, and the particle size distribution was measured when the dispersion state reached uniformity. The measurement range was set to 0.01 - 3500.00 μm. The results are shown in Fig. 33. The volume (%) on the vertical axis of Fig. 33 is the value obtained by dividing (the scattering intensity of each particle size) by (the sum of the scattering intensities of all particle sizes). As shown in Fig. 33, as a result, in the hammer mill pulverization method, peak tops were detected at 1 μm, 6 μm, and 300 - 400 μm, and in the airflow pulverization method, peak tops were detected at 1 μm, 6 μm, 20 - 30 μm, and 100 μm. Also, as shown in the cumulative volume ratio, in the hammer mill pulverization method, those with a particle size of 100 μm or less were 50% or less, whereas in the airflow pulverization method, those with a particle size of 100 μm or less were 75% or more. Since the crushed powder obtained by the airflow pulverization method has a finer particle size as a whole, it does not form lumps and has a smooth texture.

[0144] Furthermore, the physical properties of the crushed powders of Ziziphus jujuba seeds (ZSS) obtained by the hammer mill crushing method and the airflow crushing method were analyzed respectively. The results are shown in Table 2. [Table 2]

[0145] (Angle of repose) The granules were deposited from the orifice (funnel) onto a horizontal plane to form a cone. The angle formed by the generatrix of the cone and the bottom surface was defined as the angle of repose α, and was calculated from the radius r of the bottom surface and the height h of the cone by the following formula.

[0146] [Equation]

[0147] (Bulk density) Measured by a common method using a graduated cylinder. The sample (100 g) was passed through a sieve with an opening of 1.0 mm or more and placed in a 250 mL graduated cylinder without being compacted. The loose bulk volume V0 was measured, and the bulk density (g / mL) was calculated.

[0148] (Water activity) The sample (1.0 - 1.1 g) was placed on an aluminum dish with a diameter of 20 mm (the mass of which was W0 (g)), and the sample and a saturated solution were placed in a Conway unit. The aluminum dish with the sample was placed in the inner chamber installed in the Conway unit, and a standard saturated solution (selected from Table 3 below; the mass of which was W1 (g)) was placed in the outer chamber. After sealing, it was incubated at 25 °C for 2 hours. After the treatment, the aluminum dish was weighed (the mass of which was W2 (g)). The increase or decrease per 1 g of the sample was plotted on the Y-axis and the water activity of the saturated solution was plotted on the X-axis according to the following formula, and the intersection with the X-axis was calculated to obtain the water activity of the sample.

[0149] [Equation]

[0150] [Table 3]

[0151] (Acid value) To 15 g of the sample, 200 mL of diethyl ether was added to extract the oil and fat components. Subsequently, dehydration filtration and solvent distillation were performed to obtain the extracted oil. This was dissolved in 60 mL of a 1:1 mixture of ethanol and diethyl ether, the glass electrode GE-101B was connected to COM-1760 manufactured by Hiranuma Sangyo Co., Ltd., and a 0.05 mol / L potassium hydroxide standard solution was titrated to measure the potential difference.

[0152] (Peroxide value) To 15 g of the sample, 200 mL of diethyl ether was added to extract the oil and fat components. Subsequently, dehydration filtration and solvent distillation were performed to obtain the extracted oil. This was dissolved in 50 mL of a 2:3 mixture of isooctane and acetic acid, nitrogen substitution was carried out, and then 0.1 mL of saturated potassium iodide was added. After reacting for 1 minute, 30 mL of water was added and infiltrated. Using a 1% starch solution as an indicator, the resulting solution was titrated with a 0.01 mol / L sodium thiosulfate standard solution.

[0153] Powders prepared by autoclave sterilization and hammer mill crushing, and crushed products treated by airflow pulverization after roasting sterilization, were found to have a particle size of 0.5 mm or less by particle size distribution analysis. They have an angle of repose similar to that of general starches (angle of repose 45 - 58°: Precision Powder Supply Technology, by Chikara Shibata, IP Science Co., Ltd.), soybean flours (51°: Precision Powder Supply Technology, by Chikara Shibata, IP Science Co., Ltd.), wheat flours (53 - 56°: Precision Powder Supply Technology, by Chikara Shibata, IP Science Co., Ltd.), etc., and a bulk density similar to that of general wheat flours (0.56 - 0.72: Precision Powder Supply Technology, by Chikara Shibata, IP Science Co., Ltd.), powdered sugars (0.66: Powder Process Technology Integration - Basic Technology Volume, Industrial Technology Center), soybean flours (0.66: Precision Powder Supply Technology, by Chikara Shibata, IP Science Co., Ltd.). Also, while it was shown that they are hardly oxidized from the peroxide value, it was suggested from the acid value that they contain a large amount of free fatty acids. A particularly notable point is the water activity. Both the hammer mill crushing method and the airflow pulverization method have a water activity of less than 0.7, which is said to have high preservability, and the airflow pulverization method has a water activity of less than 0.5, where microorganisms cannot propagate. From the above, both the hammer mill crushing method and the airflow pulverization method are excellent in preservability from the viewpoint of suppressing bacterial growth, etc., and industrial applications including food processing similar to wheat flour with a water activity of 0.61 - 0.63 (Ministry of Health, Labour and Welfare "Manual for Sanitary Control Incorporating the Concept of HACCP in Wheat Flour Manufacturing") can be expected.

[0154] Using a simple crushing method of crushing 50 g of wild jujube seeds each at 4°C with a mill (Russell Hobbs model #7660JP) without sterilization, simple crushed products of wild jujube seeds were obtained. The nutritional components of the crushed powders of wild jujube seeds obtained by the simple crushing method, the hammer mill pulverization method, and the airflow pulverization method were analyzed. The results are shown in Table 4 below.

[0155]

Table 4

[0156] (Moisture) Place a 3.9 - 4.1 g sample (with its mass denoted as S (g)) into an aluminum weighing tube (with its mass denoted as W1 (g)), and incubate it in a vacuum dryer at 70 °C for 5 hours. After cooling in a silica gel desiccator, measure the total mass of the weighing tube and the sample (denoted as W2 (g)), and calculate the moisture content using the following formula.

[0157]

Number

[0158] (Protein) Take a 0.3 - 0.4 g measurement sample of quartz, and analyze it using a total nitrogen analyzer (manufactured by Shimizu Chemical Analysis Center). Use high - purity helium gas (purity 99.99% or higher) as the carrier gas and high - purity oxygen gas (purity 99.99% or higher) as the combustion support gas. Set the reaction furnace temperature at 870 °C or higher, the reduction furnace temperature at 600 °C, the detector temperature at 100 °C, and the column temperature at 70 °C. The calibration curve was prepared using ethylenediaminetetraacetic acid (EDTA) as the standard. Calculate the protein mass from the obtained nitrogen mass using the following formula. Here, N is the nitrogen mass (mg) calculated from the calibration curve, S is the sample mass (g), and K is the nitrogen - protein conversion factor 6.25.

[0159]

Number

Number

[0160] (Lipid) A measurement sample of 0.5 - 1.4 g (Sg) was prepared, and 2 mL of ethanol was added. Subsequently, hydrochloric acid and pure water were mixed at a ratio of 25 to 11, 10 mL of the mixture was added, and it was decomposed in a constant temperature water bath at a temperature range of 70 - 80 °C for 30 - 40 minutes. The obtained acid decomposition product was placed in a Majonier tube, 10 mL of ethanol was added, then 25 mL of diethyl ether was added, and it was shaken and mixed. Further, 25 mL of petroleum ether was added and shaken and mixed. Among the separated ether mixed liquid phase (referred to as "E1") and the aqueous phase, the aqueous phase was obtained, 40 mL of a diethyl ether - petroleum ether mixed solution (equal amount mixing) was added and shaken and mixed, and among the separated ether mixed liquid phase (referred to as "E2") and the aqueous phase, the aqueous phase was obtained. For the separated aqueous phase, 30 mL of a diethyl ether - petroleum ether mixed solution (equal amount mixing) was added and mixed, the separation of the ether mixed liquid phase (referred to as "E3") and the aqueous phase was confirmed, and the aqueous phase was removed. The obtained ether mixed liquid phases E1, E2, and E3 were mixed, placed in a weighed fat bottle (W1 g), the solvent was removed, and it was dried at 105 °C for 1 hour. After cooling in a silica gel desiccator, it was weighed (referred to as W2 (g)).

[0161] [Number]

[0162] (Ash content) 1.0 - 1.2 g of a measurement sample (its mass is S (g)) was placed in a weighed porcelain crucible (its mass is W1 (g)) and weighed. After preliminary ashing, it was ashed at 550 °C, cooled in a silica gel desiccator, and then weighed (referred to as W2 (g)). The distribution was calculated from the obtained mass by the following formula.

[0163] [Number]

[0164] (Carbohydrates) Based on the Food Labeling Standards (Cabinet Office Ordinance No. 10 of 2015), it was calculated by the following formula.

[0165] [Number]

[0166] (Carbohydrate) Calculated according to the following formula based on the Food Labeling Standards (Cabinet Office Ordinance No. 10 of 2015).

[0167] [Number]

[0168] (Dietary Fiber) Approximately 1 g of the sample was placed in a centrifuge tube and defatted twice with petroleum ether. It was then dispensed in half into two 500 mL Erlenmeyer flasks and adjusted to 50 mL each with 0.08 mL of phosphate buffer (pH 6.0). 0.05 mL of Termamyl (manufactured by Novozymes) was added, and the mixture was incubated for 30 minutes with shaking in a boiling water bath. After cooling, it was adjusted to pH 4.3 ± 0.3 with approximately 10 mL of 0.325 mol / L hydrochloric acid, and then 0.1 mL of amyloglucosidase (manufactured by Sigma-Aldrich) was added, followed by incubation for 30 minutes with shaking under conditions of 60°C. Subsequently, 95 V / V% ethanol at 60°C was diluted 4-fold and allowed to stand at room temperature for 1 hour. Then, it was suction filtered through a 2G2 glass filter (packed with 1 g of celite), and the residue was washed three times with 20 mL of 78 V / V% ethanol, twice with 10 mL of 95 V / V% ethanol, twice with 10 mL of acetone, and once with 50 mL of petroleum ether, and dried overnight at 105°C. After that, its mass was weighed and designated as R1 and R2. One of the two obtained samples was treated at 525°C for 5 hours for ashing and weighed to obtain the ash mass A. Also, the remaining sample was used to determine the protein mass P by the nitrogen determination conversion method (coefficient 6.25). The dietary fiber mass was calculated from the above mass data using the following formula. Here, R is the mass average value of the residue, P is the protein (%) in the residue, A is the ash (%) in the residue, S is the sample mass, r is the mass of the blank residue, p is the protein (%) in the blank residue, and a is the ash (%) in the blank residue.

[0169] [Number]

[0170] (Energy) Based on the Food Labeling Standards (Cabinet Office Ordinance No. 10 of 2015), it was calculated as 4 for protein, 9 for lipids, 4 for carbohydrates, and 2 for dietary fiber as energy conversion counts.

[0171] (Sodium) 1 g of the sample was placed in a quartz beaker, ashed at 500 °C for 10 hours, 2.5 mL of 20% hydrochloric acid was added, and then it was evaporated to dryness. To the obtained sample, 2.5 mL of 20% hydrochloric acid was added, heated and extracted, then filtered (filter paper No. 5A) and made up to 50 mL. Using an atomic absorption spectrophotometer (SpectrAA240FS, manufactured by Agilent Technologies), with a sodium hollow cathode lamp (manufactured by Agilent Technologies) as the light source, the measurement wavelength was 589.0 nm, and the flame was measured with acetylene at 2.00 L / min and air at 13.50 L / min.

[0172] Based on the nutritional component (moisture, protein, lipid, ash, carbohydrate, sugar, dietary fiber) data of the powder prepared by the simple crushing method, the nutritional component data of the powder prepared by the hammer mill crushing method and the airflow pulverization method were normalized and shown in Table 5 below. In the hammer mill crushing method, moisture and sugar decreased, while carbohydrates and dietary fiber tended to increase. In the airflow pulverization method, moisture decreased dramatically, sugar also decreased, while protein, lipid, ash, carbohydrate, and dietary fiber tended to increase. As described later, since the powder prepared by the hammer mill crushing method shows a tendency to more strongly improve the cognitive function of the pathological model mice than the powder prepared by the airflow pulverization method, it is possible that the pharmacological active ingredients are contained in more of the nutritional components. From this study, it was considered that dietary fiber may contain pharmacological active ingredients. Also, as described above, since pharmacological activity remained in both the hot water extract of Ziziphus jujuba and its residue, it is possible that both water-soluble dietary fiber and insoluble dietary fiber contain pharmacological active ingredients.

[0173]

Table 5

[0174] (Sodium equivalent) It was calculated by multiplying the sodium mass by 2.54.

[0175] · Example D2: Examination of the influence on the effect by the crushing preparation method In the same manner as in Example A3, the effect on the effect of the crushing preparation method of the crushed powder of Ziziphus jujuba (ZSS) on Tau784 mice was examined. The results are shown in Fig. 34. It is a graph showing the results of the Morris water maze test of Tau784 mice administered with the crushed powder of Ziziphus jujuba (Cru-ZSS new) prepared based on Example D1 at 1 mg / kg / day or 3 mg / kg / day, in comparison with Tau784 mice administered with water as a control and non-genetically modified mice. Even in the crushed powder of Ziziphus jujuba obtained by the airflow pulverization method, an extremely excellent memory improvement effect was shown. [Industrial Applicability]

[0176] The present invention can be widely applied to the fields of functional foods, pharmaceuticals, etc. where maintenance or improvement of cognitive function, etc. is required, and its utility value is extremely large.

Claims

1. An agent for maintaining or improving cognitive function and / or motor function, containing a crude drug selected from the stem parts of Ziziphus jujuba var. spinosa (Sansou'nin) and Acorus tatarinowii (Sekishoubu), wherein the Ziziphus jujuba var. spinosa is in a crushed form, and the stem part of the Acorus tatarinowii is selected from a crushed product, a hot water extract of the crushed product, and a hot water extraction residue of the crushed product.

2. An agent for promoting nerve cell repair or inducing neurogenesis, containing a crude drug selected from the stem parts of Ziziphus jujuba var. spinosa and Acorus tatarinowii, wherein the Ziziphus jujuba var. spinosa is in a crushed form, and the stem part of the Acorus tatarinowii is selected from a crushed product, a hot water extract of the crushed product, and a hot water extraction residue of the crushed product.

3. An agent for removing dementia-causing proteins that accumulate in the brain, containing a crude drug selected from the stem parts of Ziziphus jujuba var. spinosa and Acorus tatarinowii, wherein the Ziziphus jujuba var. spinosa is in a crushed form, and the stem part of the Acorus tatarinowii is selected from a crushed product, a hot water extract of the crushed product, and a hot water extraction residue of the crushed product.

4. The agent according to claim 3, wherein the dementia-causing protein is one or more proteins selected from amyloid-β (Aβ), tau, α-synuclein, TDP-43, FUS / TLS, polyglutamine, proteins by repeat-associated non-ATG (RAN) translation, prions, and SOD-1.

5. An agent for treating or preventing degenerative dementia and one or more neurodegenerative diseases selected from Parkinson's disease, multiple system atrophy, Huntington's disease, amyotrophic lateral sclerosis, and spinocerebellar degeneration, containing a crude drug selected from the stem parts of Ziziphus jujuba var. spinosa and Acorus tatarinowii, wherein the Ziziphus jujuba var. spinosa is in a crushed form, and the stem part of the Acorus tatarinowii is selected from a crushed product, a hot water extract of the crushed product, and a hot water extraction residue of the crushed product.

6. The agent according to claim 5, wherein the degenerative dementia is one or more dementias selected from Alzheimer's disease, frontotemporal dementia, and Lewy body dementia.

7. The agent according to any one of claims 1 to 6, wherein the crude drug is administered to the subject in an amount of 0.05 mg to 10 g per day.

8. A food containing the agent according to any one of claims 1 to 6, wherein the food is a food for maintaining or improving cognitive function and / or motor function, a food for promoting nerve cell repair or inducing neurogenesis, a food for removing dementia-causing proteins accumulated in the brain, or a food for treating or preventing degenerative dementia and one or more neurodegenerative diseases selected from Parkinson's disease, multiple system atrophy, Huntington's disease, amyotrophic lateral sclerosis, and spinocerebellar degeneration.

9. The crude drug selected from jujube seeds and the stem part of Acorus tatarinowii contained in the food is a crushed powder, and the proportion of the crushed powder with a particle size of 100 μm or less in the crushed powder is 50% or more. The food according to claim 8.

10. The water activity of the crushed powder is less than 0.

7. The food according to claim 9.

11. A pharmaceutical containing the agent according to any one of claims 1 to 6, wherein the pharmaceutical is a pharmaceutical for maintaining or improving cognitive function and / or motor function, a pharmaceutical for promoting nerve cell repair or inducing neurogenesis, a pharmaceutical for removing dementia-causing proteins accumulated in the brain, or a pharmaceutical for treating or preventing degenerative dementia and one or more neurodegenerative diseases selected from Parkinson's disease, multiple system atrophy, Huntington's disease, amyotrophic lateral sclerosis, and spinocerebellar degeneration.

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