JP4878748B2 - Acetylcholinesterase inhibitor - Google Patents

Description

  The present invention relates to an acetylcholinesterase (hereinafter also referred to as “AchE”) inhibitor containing tea tree oil as an active ingredient, and uses thereof.

  In recent years, an aging society has progressed, and the number of senile dementia patients has increased, and among them, Alzheimer-type dementia patients have increased. The cause of this Alzheimer-type dementia is unclear, but a decrease in the level of acetylcholine, a neurotransmitter in the patient's brain, is observed, which is caused by a decline in cholinergic nerve function. (Non-patent Document 1). Therefore, for Alzheimer-type dementia, treatment methods aiming at increasing the concentration of acetylcholine to prevent the function of cholinergic nerves from becoming mainstream have become mainstream.

  Acetylcholine is synthesized by acetylcholine transferase and maintained at a constant concentration by being decomposed by acetylcholinesterase. Therefore, it is a method of suppressing the reduction of acetylcholine by inhibiting the activity of acetylcholinesterase to suppress the degradation of acetylcholine. Although this method does not completely treat the symptoms, it has been found that it can improve the symptoms or slow the progression of the symptoms.

  By the way, it is known that the scent of essential oils of plants exhibits antibacterial action, psychological action, and biological rhythm regulating action, and aromatherapy (aroma therapy) that heals the mind and body by the scent of plants has attracted attention.

  In recent years, it has been reported that aroma components of plants such as mint, lavender or grapefruit essential oils exhibit not only the aroma but also AchE inhibitory activity (Non-patent Document 2). Ginkgo biloba extract is also said to have an effect of improving symptoms of Alzheimer's dementia.

  It is of great interest that such naturally derived ingredients have been confirmed to have AchE inhibitory activity associated with the treatment of Alzheimer's dementia.

By the way, Australian tea tree oil (essential oil) has biological activity such as antiseptic, antibacterial activity, and deodorant activity (Non-patent Documents 3 and 4), and is used as a perfume and cosmetics. The importance is increasing. However, there has been no report to date that tea tree oil has AchE inhibitory activity.
Weinstock M .: Neurodegeneration, 4, pp.349-356, 1995 Miyazawa M .: Nat. Prod. Lett., 15, pp.205-210, 2001 Laakso PV Proc. 25 th Congr. Pharm. Sci. 1965; pp485-492 Altman PM Aust, J. Pharm. Sci. 1988; pp276-278

  An object of the present invention is to provide a novel AChE inhibitor containing plant essential oil as an active ingredient and use thereof.

  The present inventors have found that an essential oil component of a specific plant (for example, tea tree oil) exhibits a strong AChE inhibitory activity, and further studies on this result complete the present invention. It was.

  That is, the present invention provides the following acetylcholinesterase inhibition and the like.

  Item 1. An acetylcholinesterase inhibitor containing tea tree oil as an active ingredient.

  Item 2. An acetylcholinesterase inhibitor containing terpinen-4-ol, γ-terpinene, α-terpinene, p-cymene and 1,8-cineol as active ingredients.

  Item 3. Contains 30-40 parts by weight of terpinen-4-ol, 15-25 parts by weight of γ-terpinene, 4-12 parts by weight of α-terpinene, 3-11 parts by weight of p-cymene and 2-6 parts by weight of 1,8-cineol Item 3. The acetylcholinesterase inhibitor according to Item 2.

  Item 4.1 An acetylcholinesterase inhibitor comprising, as an active ingredient, a bicyclomonoterpene having a 1-methylcyclohexene skeleton and containing no heteroatom.

  Item 5. Item 5. The acetylcholinesterase inhibitor according to Item 4, comprising (+)-2-carene or (+)-3-carene as an active ingredient.

  Item 6. An acetylcholinesterase inhibitor containing, as active ingredients, tea tree oil and a bicyclomonoterpene having a 1-methylcyclohexene skeleton and no heteroatoms.

  Item 7. Item 7. A preventive or therapeutic agent for senile dementia, comprising the acetylcholinesterase inhibitor according to any one of Items 1 to 6.

  Item 8. Item 7. A preventive or therapeutic agent for Alzheimer's dementia, comprising the acetylcholinesterase inhibitor according to any one of Items 1 to 6.

  Item 9. Item 7. Use of the acetylcholinesterase inhibitor according to any one of Items 1 to 6 for the manufacture of a prophylactic or therapeutic agent for senile dementia.

  Item 10. A method for preventing or treating senile dementia, comprising administering a pharmaceutically effective amount of the acetylcholinesterase inhibitor according to any one of Items 1 to 6 to a patient with senile dementia.

  Item 11. Item 7. A fragrance containing the acetylcholinesterase inhibitor according to any one of Items 1 to 6.

  Item 12. Item 7. Essential oil for aromatherapy containing the acetylcholinesterase inhibitor according to any one of Items 1 to 6.

  Hereinafter, the present invention will be described in detail.

  The AchE inhibitor of the present invention contains an essential oil (tea tree oil) that is an extract from tea tree as an active ingredient.

  The tea tree oil used in the present invention is obtained from Melaleuca alternifolia Cheel belonging to the genus Melaleuca, and its composition is mainly composed of a mixture of terpenoids.

  The tea tree oil of the present invention means a component extracted from the tea tree, and may be extracted from any part such as roots, stems, leaves, flowers, fruits, seeds and the like. Among these, those extracted from the above-ground parts (stems, leaves, flowers, berries, seeds, etc.) of the yellow cornflower are preferable. The tea tree can be used for extraction as it is, but may be used after being dried and pulverized.

  As a method for extracting tea tree oil, known methods such as a steam distillation method, a pressing method, a solvent extraction method and the like are used because the essential oil component has a low boiling point. Of these, the steam distillation method is preferably employed.

  After the essential oil is obtained from the tea tree by the method described above, in the present invention, the obtained extract can be used as it is as an AchE inhibitor. Alternatively, if necessary, the essential oil can be obtained by solid-liquid separation from the residue by a conventional method such as filtration or centrifugation.

  Furthermore, if necessary, one or more suitable separation and purification means such as alumina column chromatography, silica gel chromatography, gel filtration chromatography, ion exchange chromatography, hydrophobic chromatography, and high performance liquid chromatography may be combined. A fraction or compound having AchE inhibitory activity can be taken out and used as an AchE inhibitor. A known elution solvent may be used, but a nonpolar organic solvent having a low boiling point is preferably used as a main component. Thereby, the outstanding activity can be exhibited with a small amount of essential oil.

  Tea tree oil itself has strong AchE inhibitory activity. The tea tree oil contains individual terpenoids such as terpinen-4-ol, γ-terpinene, α-terpinene, p-cymene, 1,8-cineole as main components. Although it is slightly lower than oil, it has strong AchE inhibitory activity.

  These terpenoids exhibit high AchE inhibitory activity almost equal to that of tea tree oil when mixed at a predetermined mixing ratio. The above-mentioned predetermined mixing ratio is the content ratio of each terpenoid contained in tea tree oil (terpinen-4-ol, γ-terpinene, α-terpinene, p-cymene, 1,8-cineole = 36: 20: 8 : 7: 4, weight ratio), terpinen-4-ol 30 to 40 parts by weight, γ-terpinene 15 to 25 parts by weight, α-terpinene 4 to 12 parts by weight, p-cymene 3 to A mixture containing 11 parts by weight and 1,8-cineole 2-6 parts by weight shows strong AchE inhibitory activity, in particular 34-38 parts by weight terpinen-4-ol, 18-22 parts by weight γ-terpinene, α- Mixtures containing 6 to 10 parts by weight of terpinene, 5 to 9 parts by weight of p-cymene and 3 to 5 parts by weight of 1,8-cineol show stronger activity.

  This is because the AchE inhibitory action of the individual terpenes constituting the tea tree oil does not necessarily reach the tea tree oil, but when these are mixed at the above mixing ratio, the AchE inhibitory action equivalent to that of the tea tree oil is exhibited. . This is not expressed by a single strong active ingredient contained in tea tree oil, but is considered to be due to a synergistic action between essential oil components. Such knowledge was found for the first time by the present inventors and can be said to be a very interesting behavior. This point can be easily understood from the description of the first embodiment, for example.

  Since terpenoid mixture mixed with tea tree oil and a predetermined mixing ratio has strong AchE inhibitory activity, it is widely used as an AchE inhibitor.

  Further, as a result of screening AchE inhibitory action for various terpenoids, it was revealed that bicyclomonoterpenes having a 1-methylcyclohexene skeleton and not containing a heteroatom have extremely high AchE inhibitory action. Specific compounds include (+)-α-pinene, (−)-α-pinene, (+)-2-carene, and (+)-3-carene. Especially, at least 1 sort (s) chosen from the group which consists of (+)-2-carene and (+)-3-carene from an active point is preferable, and the thing containing (+)-3-carene is especially preferable.

  Bicyclomonoterpenes having a 1-methylcyclohexene skeleton and not containing a heteroatom are widely used as AchE inhibitors because of their strong AchE inhibitory activity.

  The above-mentioned tea tree oil, terpenoid mixture mixed at a predetermined mixing ratio, and bicyclomonoterpene having a 1-methylcyclohexene skeleton and not containing a heteroatom may be mixed as necessary and used as an AchE inhibitor. it can. The mixing ratio of each component in the case of mixing is not particularly limited, but bicyclomonoterpene (particularly (+)-3-carene) having a 1-methylcyclohexene skeleton and not containing a heteroatom is preferably 1 in the mixture. It is preferable to contain about ~ 95 wt% (especially 50 to 95 wt%).

  The AchE inhibitor of the present invention described above can be used for various applications.

  For example, when the AchE inhibitor of the present invention is used as a pharmaceutical, it is used as a preventive or therapeutic agent for senile dementia in mammals (particularly humans), particularly as a preventive or therapeutic agent for Alzheimer-type dementia.

  The AchE inhibitor of the present invention is a tablet, powder, fine granule, granule, coated tablet, capsule, syrup, troche, inhalant, suppository, injection, ointment, ocular by a conventional method. It can be formulated into agents such as ointments, eye drops, nasal drops, ear drops, poultices, lotions and the like.

  Excipients, binders, lubricants, colorants, flavoring agents, and if necessary stabilizers, emulsifiers, absorption promoters, surfactants, pH adjusters, preservatives, Antioxidants and the like can be used. In general, ingredients and blending amounts that are generally used as raw materials for pharmaceutical preparations are appropriately selected to prepare a preparation by a conventional method.

  When administering the pharmaceutical formulation of this invention, the form is not specifically limited, What is necessary is just the method used normally, and oral administration or parenteral administration may be sufficient. The pharmaceutical dosage according to the present invention can be appropriately selected from pharmaceutically effective doses according to the degree of symptoms, age, sex, body weight, dosage form / salt type, specific type of disease, etc. .

  In addition, since the AchE inhibitor of the present invention has strong AchE inhibitory activity, it can be used in addition to products such as bathing agents, soaps, fragrances, aromatherapy essential oils, perfumes, hairdressing agents, and the like.

  The AchE inhibitor of the present invention may be usually contained in an amount of about 0.01 to 100% by weight (preferably about 0.1 to 5% by weight) based on the entire product.

When the above-mentioned product containing the AchE inhibitor of the present invention is used, the relaxation effect and refreshing effect due to the scent of essential oils are exhibited, and the healing effect of the mind and body is exhibited in a modern society with many stresses. By using the above products in an environment where elderly people, sick people, etc. live, the effects of preventing and improving senile dementia are also exhibited. Of course, the antiseptic and antibacterial effects of tea tree oil are also exhibited. In addition, when used as an essential oil for aromatherapy, the AchE inhibitor of the present invention can be used by mixing with other natural plant essential oils. Further, the essential oil for aromatherapy can be used in such a form that it can be spread and diffused in a predetermined place and inhaled. The oil may be distributed and diffused using a diffuser or the like. The distribution / diffusion place can be distributed, for example, in a home room, a hotel room, a conference room, a hospital room, an event hall where people gather, a break plaza, various relaxation facilities, and the like. In particular, it is preferably used in hospitals and facilities where senile dementia patients live.

  In addition, the AchE inhibitor of the present invention can be blended in various foods and beverages such as soft drinks, dairy products (processed milk, yogurt), and confectionery (jelly, chocolate, biscuit, gum, tablet confectionery).

  The AChE inhibitor of the present invention contains tea tree oil, (+)-3-calen, etc., and has a strong AChE inhibitory activity, and is used as a therapeutic or prophylactic agent for senile dementia, particularly Alzheimer-type dementia. Useful. In addition, the AChE inhibitor of the present invention can be contained in a fragrance, a bath agent, an aromatherapy essential oil, or the like, and can exert physical and mental healing effects such as a relaxation effect and a refreshing effect.

  Next, the present invention will be specifically described with reference to examples, but the present invention is not limited thereto.

Example 1
(1) When component search was performed using GC-MS for tea tree oil (manufactured by Yamamoto Fragrance Co., Ltd.) under the conditions shown in Table 1 below, it was found that the monoterpene hydrocarbon compound was the main constituent. (Table 2).

(2) The AChE inhibition test was performed according to the method of G. L. Ellman, K. D. Courtney, V. Andres. Jr., R. M. Featherstone. Biochem. Pharmacol. 7, 88-95 (1961). 0.04 unit / mL AChE solution, 0.1M phosphate buffer, inhibitor (tea tree oil or monoterpene) and color former (5,5'-dithiobis (2-nitrobenzoic acid; DTNB) in a test tube at 25 ° C After 5 minutes pre-incubation, substrate (acetylthiocholine iodide; ATC) was added and incubated for 20 minutes at 25 ° C. After that, the yellow-colored anion (TNB) generated by hydrolysis reaction of the substrate with AchE at 412 nm Absorbance was measured (see Fig. 1), and the value of inhibitory activity was calculated by comparing the value with the value of no added essential oil as a control value. Indicated.

As AchE, bovine erythrocyte AchE manufactured by Sigma Co. Ltd. was used. DTNB and ATC were manufactured by Tokyo Chemical Industry Co. Ltd. (TCI). The terpenoid used was Tokyo Chemical Industry Co., Ltd. and Wako Pure Chemical Industries, Ltd.
(3) First, Table 3 and FIG. 2 show the results of enzyme inhibition tests of essential oils and low-polarity hydrocarbon fractions and high-polarity oxygen-containing fractions obtained by fractionating essential oils by column chromatography. Show. The essential oil showed a strong inhibitory activity, and the IC ₅₀ value as a 50% inhibitory concentration was 51.2 μg / mL. Moreover, the inhibitory activity of the hydrocarbon fraction and the oxygen-containing fraction did not reach the inhibitory activity of tea tree oil.

  Next, when the active component of the hydrocarbon fraction was searched, the five components of terpinen-4-ol, γ-terpinene, α-terpinene, p-cymene and 1,8-cineole, which are the main components of tea tree oil, were found. It became clear that the comparatively strong inhibitory activity was shown (refer Table 3 and FIG. 2). However, it was found that the activity value of each component alone excluding 1,8-cineole does not reach the essential oil.

(4) Abundance ratio in essential oil of 5 compounds of terpinen-4-ol, γ-terpinene, α-terpinene, p-cymene and 1,8-cineole, which are main components of tea tree oil, 36: 20: 8 As a result of preparing a mixture similar to: 7: 4 (weight ratio) and conducting an inhibition test, the IC ₅₀ value was 65.5 μg / mL. The inhibitory activity of the mixture was slightly weaker than that of tea tree oil, but was found to be higher than each component alone. (See Table 3 and FIG. 3). This suggested that the AcnE inhibitory activity of tea tree oil was not due to a single strong inhibitory active component, but due to a synergistic effect between essential oil components.
(5) Next, the enzyme inhibition mode of tea tree oil was examined. As the method, the Lineweaver-Burk plot method and the Dixon plot method, which are widely used for kinetic methods, were used. The results are shown in Table 4.

  Tea tree oil showed mixed inhibition of AchE from the intersection of the Lineweaver-Burk plot, and the inhibition constant was 54.7 μg / mL from the intersection of the Dixon plot. In contrast, the main terpenoids showed antagonistic inhibition of AchE, whose inhibitory activity increased with decreasing substrate concentration from the crossing of the Dixon plot. Each terpenoid and mixture antagonized the substrate at the active center of the enzyme.

  As described above, tea tree oil can be suitably used as an AchE inhibitor.

Example 2
(1) AChE inhibition test was carried out on 17 kinds of bicyclic monoterpenoids contained in plant essential oil in the same manner as in Example 1 (2). The results are shown in Table 5.

(2) Further, the inhibition constants of the above four compounds were measured in the same manner as in Example 1 (5). The results are shown in Table 6.

  From the above results, (+)-α-pinene, (−)-α-pinene, (+)-2-carene and (+)-3-carene showed strong AChE inhibitory action, and in particular, (+) − It was found that the activity of 3-carene was high.

BRIEF DESCRIPTION OF THE DRAWINGS It is a figure which shows the outline of the AchE inhibitory activity test used in Example 1. FIG. It is a figure which shows the result of the AchE inhibitory activity test of tea tree oil and its main component in Example 1. 2 is a graph showing the results of AchE inhibitory activity test of a mixture of tea tree oil and 5 compounds in Example 1. FIG.

Claims (6)

Tea tree oil and at least one compound selected from the group consisting of (+)-α-pinene, (−)-α-pinene, (+)-2-carene and (+)-3-carene as active ingredients Contains an acetylcholinesterase inhibitor. A mixture comprising the tea tree oil and at least one compound selected from the group consisting of (+)-α-pinene, (−)-α-pinene, (+)-2-carene and (+)-3-carene inside,
50 to 95% by weight of at least one compound selected from the group consisting of (+)-α-pinene, (−)-α-pinene, (+)-2-carene and (+)-3-carene The acetylcholinesterase inhibitor according to claim 1. At least one compound selected from the group consisting of (+)-α-pinene, (−)-α-pinene, (+)-2-carene and (+)-3-carene is (+)-3. The acetylcholinesterase inhibitor according to claim 1 or 2, which is -carene. A preventive or therapeutic agent for senile dementia, comprising the acetylcholinesterase inhibitor according to any one of claims 1 to 3. A preventive or therapeutic agent for Alzheimer-type dementia, comprising the acetylcholinesterase inhibitor according to any one of claims 1 to 3. Use of the acetylcholinesterase inhibitor according to any one of claims 1 to 3 for the manufacture of a preventive or therapeutic agent for senile dementia.

Images