JPH064537B2 - Stress dysfunction improver - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION (Field of Industrial Use) The present invention relates to a stress dysfunction-improving agent, and in particular, contains a phenylpropanoid glycoside as an active ingredient.
This relates to stress dysfunction improving agents.

(Prior Art) Phenylpropanoid glycosides have the formula (In the formula, R ¹ and R ² represent a hydrogen atom, a hydroxy group or a methoxy group, R ³ represents a hydrogen atom, a β-D-glucopyranosyl group or a β-D-apiofuranosyl group, and R ⁴ represents hydrogen. (Meaning an atom or an acetyl group). As phenylpropanoid glycosides known in the past, acteosides (R ¹ , R ² isolated from lily of the Asteraceae plant, rodent moth and Asteraceae, as well as the nambangiceel of the Nematode family plant, and the thorn of the Spodoptera punctaceae plant, etc. = Hydroxy group, R ³ , R ⁴ = hydrogen atom) and the following compounds are examples.

Echinakosaido (R ^1, R ^2: hydroxy group, R ^3: β-D- glucopyranosyl group, R ^4: a hydrogen atom), 2'-acetyl accession Theo Side (R ^1, R ^2: hydroxy groups, R ^3: a hydrogen atom , R ^4: acetyl group), Osman scan side ^{B (R 1, R 2,} R 4, R 4: water water atom), fault perilla side B (R ^1, R ^2: hydroxy group, R ^3: β-D - Apio furanosyl group, R ^4: for pharmacological or physiological action of the hydrogen atoms) these phenylpropanoid glycosides have been reported that the Echinakosaido and fault perilla side B both have antimicrobial activity (A. Stoll et al. “Helv. Chim. Acta.” P. 238, 1950 and Endo et al. “Abstracts of the 28th Annual Meeting of the Japanese Society of Biopharmacy” p. 20, 1981), and acteoside has anti-parkinsonism and β-blocking. It has been reported to have an action [West German Patent Application Publication No. 2609533] It has been reported that it has an antitumor effect (Numata et al., "Abstracts of the 33rd Annual Meeting of the Kinki Branch of the Pharmaceutical Society of Japan", page 76, 1983).

On the other hand, flesh roe, which is a parasitic plant of the family Nematodeaceae, is included in the oldest book of herbs, "Shinno Honzo Sutra," and has been used as a tonic since ancient times, but it is a pharmacological substance that supports the tonic action. The actual situation is that the physiological action has not been proved, and only a slight hypotensive action and salivary secretion promoting action have been reported ("Summary of the 1956 Thesis Report of the Chinese Academy of Medical Sciences, II", p. 76, 1956 and "Summary of Chuanther research literature", p. 259, 1975). For this reason, the inventors of the present invention understand that the traditional efficacy of beef broth, that is, "five to seven wounds" refers to the healing of various symptoms caused by the functional deterioration of the vital organs due to stress and mental fatigue. As a result of repeated studies on beef roe focusing on this point, it was found that beef roe contains the following novel phenylpropanoid glycosides in addition to various known phenylpropanoid glycosides. , And reported a patent application for the isolation method (Kobayashi et al., "Chem. Pharm. Bull." Volume 32, 3009 and 3880).
Pp. 1984 and Japanese Patent Application No. 60-29335).

Cystannoside A (R ¹ : hydroxy group, R ² : methoxyl group, R ³ : β-D-glucopyranosyl group, R ⁴ : hydrogen atom) Cystannoside B (R ¹ , R ² : methoxyl group, R ³ : β-D- Glucopyranosyl group, R ⁴ : hydrogen atom) Cistanoside C (R ¹ : hydroxy group, R ² : methoxyl group, R ³ , R ⁴ : hydrogen atom) and cystanoside D (R ¹ , R ² : methoxyl group, R ³ , R) ⁽⁴ : Hydrogen atom) These cystanosides were presumed to have anti-stress effects as a result of preliminary tests, but they have not been sufficiently clarified.

An animal test method for demonstrating the efficacy of crude drugs classified as supplements in Oriental medicine was established by Saito et al.
66, 1983), recovery of dysfunction caused by stress is the eleuthero extract (JP 59-53426), syringin (JP 59-33221), syringaresinol diglucoside (JP Sho 59-33221). 59-116220) and iridoid, its glycosides or salts (JP-A-59-164717).

(Problems to be solved by the invention) In the present age where scientific and technological innovation is rapidly advancing, the social mechanism is complicated, and the people are forced to live a lot of stress with it. As a result, psychosomatic disorders and neuroses tend to occur frequently. Psychosomatic disorders appear as disorders in the respiratory system, digestive system, circulatory system or urogenital system, and neurosis appears as insomnia, emotional instability, amnesia and the like. Therefore, in order to carry out a healthy social life under the modern living environment, a drug that improves physical and neurological dysfunction caused by stress is extremely significant.

Therefore, a problem to be solved by the present invention is to provide an anti-stress agent containing an active ingredient of a drug of this type, in particular, a crude drug, thereby improving functional disorder due to stress. .

(Means and Actions for Solving Problems) As described above, it has been reported that phenylpropanoid glycosides have a few pharmacological or physiological actions, but an anti-stress action. Was not previously known.

The present inventors have been using tonics as a tonic since ancient times.
It is a herbal medicine that is effective for forgetfulness, low back and knee cold pain, enuresis, infertility, obita, constipation, etc. We have been conducting research on beef roe used in medicinal liquor together with the fact that as a result, we have already announced that beef roe contains various phenylpropanoid glycosides. ,
After further research, the fractions at each stage obtained in the extraction of beef roe, and the pharmacological and physiological actions of the constituents obtained by further purifying these fractions were examined by Saito et al. According to the method, it was found that the sexual behavior disorder and the decrease in learning and memory ability due to stress were improved by the administration of phenylpropanoid glycosides, and the present invention was completed.

Therefore, according to the invention, the formula (Wherein R ¹ , R ² , R ³ and R ⁴ have the above-mentioned meanings), and contains at least one phenylpropanoid glycoside as an active ingredient.
An agent for improving dysfunction caused by stress is provided.

As the active ingredient of the agent according to the present invention, isolated and purified acteoside, etinacoside, cystanoside A, cystanoside C and the like are advantageous, but it is also possible to use them without such isolation and purification.

(Dosage Form and Dosage) There is no particular limitation on the dosage form of the agent according to the present invention, and therefore, at least one compound of phenylpropanoid glycosides can be administered as a syrup or as a formulation. In preparation, oral preparations such as powders, fine granules, granules, capsules, liquids and syrups, or parenteral preparations such as injections and enteral injections You can also

The dose depends on the patient's age, weight, symptoms, dosage form, etc., but when orally administered to adults, 0.05-5 g / day of phenylpropanoid glycoside should be taken in 1-3 divided doses. Is appropriate.

(Effects of the Invention) The present invention is based on the discovery and confirmation of a new pharmacological or physiological action related to phenylpropanoid glycosides, that is, an anti-stress action. When the agent according to the present invention is administered, various effects are caused by stress. It is possible to reduce or cure functional disorders, particularly sexual dysfunction and amnesia, and to prevent or treat psychosomatic disorders.

The toxicity of phenylpropanoid glycosides is extremely low,
Therefore, the agent according to the present invention has excellent use safety.

(Formulation Examples, etc.) Next, the present invention will be described more specifically with reference to production examples of phenylpropanoid glycosides, pharmacological test examples and formulation examples.

Examples of production of phenylpropanoid glycosides Phenylpropanoid glycosides are used for lilacs, rodent moths, antler, privets, etc. of the family Asteraceae, for smoky tobacco, etc. It is contained in the celestial herbs, etc., and in the periwinkle of the family Scutellariae, and therefore can be obtained by extracting these plants as raw materials. In consideration of the large number of phenylpropanoid glycosides contained in these various raw materials, we adopted meat roe as a raw material, and this was adopted as Kobayashi et al. "Chem.Pharm.Bull."
Volume 32, Pages 3009 and 3880, 1984 and Japanese Patent Application No. 60-293
A phenylpropanoid glycoside was prepared by treatment according to the method described in No. 35.

That is, the beef roast is shredded and extracted with an aqueous alcohol solution (water and a highly polar organic solvent can also be used together), and the resulting extract or its n-butanol-soluble part is a styrene-based non-polar adsorption resin such as Using commercially available Diaion HP-20 (manufactured by Mitsubishi Kasei Co., Ltd.) and Amberlite XAD-2 (manufactured by Rohm and Haas), the crude glycoside fraction (hereinafter referred to as “fraction
-1 "). Next, this fraction-1 was treated with a column filled with a polyamide resin, for example, a commercially available polyamide C-200 (manufactured by Wako Pure Chemical Industries, Ltd.) to give a crude iridoid glycoside and a crude lignan glycoside mixed fraction. (Hereinafter referred to as "fraction-2") and a crude phenylpropanoid glycoside fraction (hereinafter referred to as "fraction-3").

The results of thin layer chromatographic analysis of Fraction-2 under the following conditions are shown in Fig. 2. As is clear from the analysis results, this fraction contained 8-epiloganic acid (1), Ajugor (2), Musaenosidic acid (3), Geniposidic acid (4), 6-Deoxycatalpol (6), Gluroside (7), Baltosioside (8), 8-Epideoxyloganic acid (9), Cystanine (12) and other iridoids and their glycosides, lignan glycosides such as syringaresinol diglucoside (5) and syringaresinol monoglucoside (11), and 8-hydroxygeraniol-1-glucoside Compounds such as (10) are included, but phenylpropanoid glycosides are not included.

Thin plate: Silica gel 60F ₂₅₄ (Merck) Developer: Chloroform / methanol / formic acid / water = 70/30
/ 2/1 Color development method: heated at 105 ° C after spraying 20 v / v (%) sulfuric acid On the other hand, the result of thin layer chromatographic analysis of Fraction -3 under the following conditions is shown in Fig. 1. As is clear from this analysis result, in this fraction, phenylpropanoid glycosides cystanosides A to D (2, 4, 5 and 8 in the figure), etinacoside (1), acteoside (3), It contains 2'-acetylacteoside (6) and Osmans side B (7) but no iridoid or lignan related compounds.

Thin layer plate: Silica gel 60F ₂₅₄ (manufactured by Merck) Developer: Chloroform / methanol / water = 6/4/1 Coloring method: 20v / v (%) Heated at 105 ° C after spraying sulfuric acid Contained in fraction-3 In order to isolate each of the various phenylpropanoid glycosides, the fraction-3 is treated with silica gel column chromatogram and a non-polar adsorption resin (eg Pharmacia) using the thin layer chromatogram shown in FIG. 1 as an index.・ Sephadex LH-2 manufactured by Fine Chemicals
0) Perform column chromatogram processing.

Pharmacological and pharmacological effect test example 1 Phenylpropanoid glycosides for sexual behavior disorder and learning and memory disorder caused by stress by the method of Saito et al. ("16th Japanese and Chinese medicine symposium lecture summary", page 66, 1983). The improvement effect by was investigated.

(1) Experimental animals Males of the IV-CS strain with normal sexual behavior over 9 weeks of age (body weight 28
-32g), temperature 23 ± 1 ℃, humidity 55 ± over one week
Preliminary breeding was carried out under the condition of 5%. In this case, "normal sexual behavior" means that male mice bred in a single-chamber cage cohabit with 10 female mice in estrus by daily subcutaneous administration of 10 μg / kg of estradiol for 10 minutes every day. Refers to the case where the intromission is successful at least five times a week.

(2) Stress load A male mouse, which is an experimental animal, is suspended in air and fixed at a height level position where the tip of the nose comes into contact with the water surface. This state is maintained for 30 minutes on the first day of the test, and suspended for the second and subsequent days. On the 15th day of the final day of the test, the stress was gradually extended and maintained in the suspended state for 2 hours to apply stress. using this method
After daily stress for 15 days, no weight loss was observed compared to the control non-stressed group, and there was no motor coordination disorder, decreased muscle tone, or decreased spontaneous or exploratory movements. However, a decrease in sexual behavior and learning and memory behavior was observed.

(3) Experimental method and method for determining effect of test drug Stress was applied every day from the specified time (1 pm), and the test drug was orally administered after the stress was applied.
We investigate sexual behavior from time and learning (memory) behavior from 11:00,
This is repeated for 15 days to examine the improving effect of the test drug.

a) Judgment of sexual behavior One male mouse, which is an experimental animal, was allowed to cohabit for 10 minutes in a cage (30x40x20cm) in which 10 female mice each receiving estradiol (10 μg / kg) were placed. , The number of sexual behaviors, including mounting and intromission, the number of behaviors and the time until the behaviors are measured. In this case, one group consisted of 10 animals, the χ ₂ test for the number of animals was used for the significant difference test between groups, and the analysis of variance method was adopted for the others.

b) Judgment of learning (memory) behavior Rubber plug (diameter 5c) on the floor where current (DC36V, 0.1mA) is flowing.
(m, height 5 cm) is placed, and the male mouse, which is an experimental animal, is allowed to learn for 5 minutes so that it will not descend from the rubber stopper. Male mice subjected to this learning treatment were placed on the rubber stoppers, and the number of times they accidentally descended from the rubber stoppers within 5 minutes and the number of mice that accidentally descended in 10 mice per group were measured.
In this case, the analysis of significant differences between groups adopted the analysis of variance method for the number of times of erroneous landings, and the χ ₂ test for the number of erroneous animals.

(4) Results and its analysis a) Effects on sexual behavior i) Results The results of examining the reduction of sexual behavior due to stress load and its recovery by administration of each test drug were as shown in the table below.

ii) Analysis Sexual behavior of male mice follows the course of licking, mounting, intromission, and ejaculation. Therefore, the effect of the treatment can be determined by investigating what kind of change occurs in the sexual behavior of the male mouse that has been treated with the female mouse when it cohabits with the female mouse in the estrus state. That is, it is judged that the more the number of sexual behaviors among 10 animals in a group within a certain time, the more the number of sexual behaviors, and the shorter the time until sexual behaviors occur, the more sexual behaviors are active. You can do things.

As is clear from the above table, the sexual behavior of stress-loaded male mice is generally reduced. When comparing the stressed test group with the non-stressed control group, it was revealed that there was a marked difference in the later sexual behavior rather than the initial sexual behavior of licking. . In addition, referring to the number of animals that cause each sexual behavior, the number of times and the time required to start, the stress load has a relatively small effect on the number of animals that perform sexual behavior, but the number of times each mouse takes sexual behavior, There was a clear difference in the time required to initiate sexual behavior.

Thus, a clear improvement in sexual behavior was observed by administering the test drug to male mice whose sexual behavior was reduced due to stress loading. Regarding the relationship between the type of test drug and its improving effect, Fraction-1 (crude glycoside fraction,
Various phenylpropanoid glycosides and lignans and iridoid-related compounds were also the least effective, and an improvement was observed at a dose of 50mg / kg, and fraction-2 (lignans and iridoids-related compounds) was found. The latter, that is, phenylpropanoid glycosides, is more effective when comparing Fraction-3 (comprising compounds as constituent components) and Fraction-3 (comprising various phenylpropanoid glycosides as constituent components) It was also found that acteoside, etinacoside, cystanoside A and cystanoside C, which were obtained by further purifying Fraction-3, were effective in improving even if a relatively small dose of 10 mg / kg was administered.

These facts show that the phenylpropanoid glycoside has an effect of effectively improving the decline of sexual function due to stress.

b) Effects on learning (memory) behavior i) Results The test results are shown in Fig. 3.

ii) Analysis In the stress-free control group, the number of mistakenly getting off the rubber stopper within the test time of 5 minutes was 0.75 times a day during the 15-day test period, whereas the stress In the loaded test group, the average was 2.6 times a day, and it was revealed from this that the stress load causes a decrease in learning behavior.

Thus, by administering the test drug to male mice whose learning behavior was reduced by stress, the frequency of accidentally getting off the rubber plug was significantly reduced. A similar tendency was observed in the number of mice that made an error.

The action of etinacoside is weaker than that of other phenylpropanoid glycosides, and the action of fraction-2 containing lignan and iridoid-related compounds constitutes various phenylpropanoid glycosides. It was weaker than the action of individual phenylpropanoid glycosides other than the component Fraction-3 and etinacoside.

These facts show that phenylpropanoid glycosides have an effect of improving the decline in memory ability due to stress.

Pharmacological and pharmacological test example 2 (toxicity test) A ddY mouse (body weight 25-30 g) has a temperature of 23 ± 1 ° C. and a humidity of 55 ±.
A healthy individual was preliminarily bred for 1 week under the condition of free feeding and free water supply in a room of 5%, and a healthy group was made into 5 animals. Then
After fasting for 18 hours, Fraction-3, etinacoside, acteoside and cystanoside A are each 1 g / kg
And 500 mg / kg of cystanoside C were orally, intraperitoneally and intravenously administered respectively.

A sedative effect and a decrease in body temperature were observed for about 5 hours immediately after the administration of any of the test drugs and by any administration route, but they recovered after about 6 hours, and no death was observed at all. After that, we examined the body temperature and general symptoms for one week, but no difference was observed between the control group and
Moreover, no abnormality was found by autopsy.

This indicates that the toxicity of phenylpropanoid glycosides is extremely low, and suggests that the safety is extremely high when used as an active ingredient of a drug.

Formulation Example 1 (Granule) A crude extract obtained by immersing beef in 50% ethanol aqueous solution was dissolved in water and treated with Diaion HP-20 column chromatogram to obtain a crude glycoside fraction (fraction-1). ) Got. This fraction was treated with a polyamide column chromatogram to obtain a crude phenylpropanoid glycoside fraction (fraction-3).

To 20 g of this fraction, 95 g of starch and 80 g of lactose were added and mixed uniformly, then wet granulated by a conventional method using hydroxypropyl cellulose (5 g) ethanol solution, dried and sized to obtain granules. It was This granule is packaged in 1 g of laminated paper. The dosage of this granule is 3 times a day, 1 each
It is addressed to the parcel.

Formulation Example 2 (Capsule) 100 g of lactose and 85 g of starch were added to 10 g of etinacoside and mixed uniformly, and then hydroxypropyl cellulose (5 g)
Wet granulation was carried out by a conventional method using an ethanol solution, dried, sized, and filled to 200 mg per capsule to obtain a hard capsule. The dose of this capsule is 3 times a day, 2 each
It is addressed to the capsule.

Formulation Example 3 (tablets) 10 g of acteoside, 90 g of starch, 97 g of lactose, 1 g of hydroxypropyl cellulose and 2 magnesium stearate
g was added and uniformly mixed, and wet sizing was carried out by a conventional method, followed by drying, sizing, and tableting to obtain a tablet of 200 mg / tablet. The dose of this tablet is 3 times a day, each for 3 tablets.

Formulation Example 4 (internal drug solution) 10 g of cystanoside A, 100 g of glucose, and an appropriate amount of fragrance were dissolved in a 14% ethanol solution to make a total amount of 1000 ml to obtain an internal drug solution. The dose of this solution is once a day,
It is 10 ml.

[Brief description of drawings]

Fig. 1 is a thin-layer chromatogram of crude phenylpropanoid glycosides (fraction-3) extracted from beef, and Fig. 2 is an iridoid- and lignan-related compound extracted from beef (fraction- Fig. 3 is a thin-layer chromatogram of 2). Fig. 3 shows the effects of stress on learning and memory behavior in mice, and the reduction of learning and memory behavior by fraction-2 and fraction-3 and phenylpropanoid glycosides. It is a graph which shows the improving effect which it exerts.

Claims (5)

[Claims] 1. A formula (In the formula, R ¹ and R ² represent a hydrogen atom, a hydroxy group or a methoxy group, R ³ represents a hydrogen atom, a β-D-glucopyranosyl group or a β-D-apiofuranosyl group, and R ⁴ represents hydrogen. (Meaning an atom or an acetyl group) is contained as an active ingredient in at least one of phenylpropanoid glycosides represented by
An agent for improving functional disorders caused by stress. 2. An ethinacoside, wherein R ¹ and R ² represent a hydroxy group, R ³ represents a β-D-glucopyranosyl group, and R ⁴ represents a hydrogen atom, as an active ingredient. The functional disorder improving agent according to claim 1. 3. The method according to claim ^1, wherein R ¹ and R ² are hydroxy groups, and R ³ and R ⁴ are acteosides, which are hydrogen atoms. Functional disorder improving agent. 4. A cystanoside A in which R ¹ represents a hydroxy group, R ² represents a methoxyl group, R ³ represents a β-D-glucopyranosyl group, and R ⁴ represents a hydrogen atom as an active ingredient. The dysfunction-improving agent according to claim 1, wherein 5. A cystanoside C having R ¹ as a hydroxy group, R ² as a methoxyl group, and R ³ and R ⁴ as a hydrogen atom as an active ingredient. The dysfunction-improving agent according to item 1.