JP2013166732A - Method for producing endothelin activity inhibitor and method for producing usnea longissima extract - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method for producing an endothelin activity inhibitor containing an active ingredient of the endothelin activity inhibitor in a high concentration and excellent in an endothelin activity inhibitory effect, and a method for producing Usnea longissima extract containing the active ingredient of the endothelin activity inhibitor in a high concentration.SOLUTION: A method for producing Usnea longissima extract highly containing an evernic acid is provided by comprising a process of extracting Usnea longissima at ≥0°C and <20°C temperature by using an alcohol or alcohol solution having ≤10 vol% water content, and a method for producing an endothelin activity inhibitor is provided by comprising a process of extracting Usnea longissima at ≥0°C and <20°C temperature by using the alcohol or alcohol solution having ≤10 vol% water content.

Description

  The present invention relates to a method for producing an endothelin action inhibitor and a method for producing a Nagasaga Ogase extract.

Endothelin is a peptide hormone derived from endothelial cells and acts on various cells and tissues through receptors. For example, it is known to cause an increase in intracellular calcium concentration in vascular smooth muscle cells and the like (Non-Patent Document 1).
In recent years, endothelin has promoted an increase in intracellular calcium concentration in epidermal melanocytes (melanocytes), promoted cell proliferation via intracellular signal transduction system, and enhanced the activity of tyrosinase, the rate-limiting enzyme of melanin synthesis. Has been reported (see Non-Patent Document 2). In addition, endothelin is one of the melanocyte activators produced by epidermal keratinocytes (keratinocytes) (Non-patent Document 3), and is an important factor in ultraviolet-induced pigmentation and senile pigment spot formation. It has been reported (Non-Patent Documents 4 and 5).
A substance capable of suppressing the action of endothelin due to the biological action of endothelin suppresses melanin production and pigmentation and is useful as a whitening agent.

On the other hand, it is known that an extract of Usnea longissima, which is a kind of lichen belonging to the genus Parmaliaceae ( Usnea ), has an inhibitory action on melanin production (Patent Documents 1 and 2, and Non-patent document 6). On the other hand, it is known that evelic acid contained in the Nagasaru Ogase extract has an anti-carcinogenic promotion effect (see Non-Patent Document 7). However, it has not been known that evernic acid has an inhibitory action on melanin production, and it is only used in a method for treating neurodegenerative diseases, an Epstein-Barr virus activation inhibitor, a deodorant composition, etc. Yes (for example, see Patent Documents 3 to 5).

JP-A-4-282319 JP-A-4-282321 International Publication No. 2008/109521 JP 7-69882 A German Patent Application No. 2351864

Hirata Y. et al., Biochem. Biophys. Res. Commun., 1988, vol. 154, p. 868-875 Yada Y. et al., J. et al. Biol. Chem., 1991, vol. 266, p. 18352-18357 Imokawa G. et al., J. et al. Biol. Chem., 1992, vol. 267, p. 24675-24680 Imokawa G. et al., J. et al. Invest. Dermatol., 1995, vol. 105, p. 32-37 Kadono S. et al., J. et al. Invest. Dermatol., 2001, vol. 116, p. 571-577 Moo-Sung Kim et al. Microbiology, 2007, vol. 45, p. 578-582 Yasuhiro Kinoshita, “Pharmacology of lichens”, Nippon Paint periodic technical information magazine, 1997, Vol. 11, p. 25-29

  This invention makes it a subject to provide the manufacturing method of the endothelin action inhibitor which contains the active ingredient of an endothelin action inhibitor in high concentration, and is excellent in the endothelin action inhibitory effect. Moreover, this invention makes it a subject to provide the manufacturing method of the Nagasaga Ogase extract which contains the active ingredient of an endothelin action inhibitor in high concentration.

  In view of the above problems, the present inventors have intensively studied to search for a novel substance that suppresses the action of endothelin. As a result, it has been found that everonic acid represented by the following formula has an inhibitory effect on the increase in calcium (ion) concentration in melanocytes caused by the action of endothelin.

Furthermore, the present inventors have studied the extraction conditions for Nagasaru Ogase, from which an extract containing a high content of evernic acid can be obtained efficiently. In Non-Patent Document 6, Nagasagaogase is extracted at a high temperature of 70 to 80 ° C. using methanol as an extraction solvent, and the concentration after filtration is performed at 50 ° C. However, it has been found that when Nagasaru Ogase is treated under such high temperature conditions, alcohol degradation of eberic acid occurs, and the yield of eberic acid is greatly reduced. Further, Patent Document 1 and Patent Document 2 describe that Nagasaruogase is extracted at a low temperature using an alcohol such as methanol as an extraction solvent. However, when an alcohol solution having a high water content is used as an extraction solvent, it has been found that the yield of everonic acid is greatly reduced by hydrolysis of everonic acid. From these results, it is possible to effectively increase the calcium (ion) concentration in melanocytes caused by the action of endothelin by extracting Nagasaruogase at a low temperature using an alcohol or alcohol solution having a low water content as an extraction solvent. It was found that a Nagasaga Ogase extract containing a high concentration of inhibitory evernic acid was obtained.
The present invention has been completed based on these findings.

That is, the present invention provides a method for extracting Nagasaru Ogase extract with a high content of evelic acid, which comprises a step of extracting Nagasal Ogase using an alcohol or an alcohol solution having a water content of 10% by volume or less at a temperature of 0 ° C. or more and less than 20 ° C. Regarding the method.
Moreover, this invention relates to the manufacturing method of an endothelin action inhibitor including the process of extracting Nagasaruogase using the alcohol or alcohol solution whose water content is 10 volume% or less at the temperature of 0 degreeC or more and less than 20 degreeC.
Furthermore, this invention relates to the whitening agent which uses the Nagasaruogase extract obtained by the said manufacturing method or an endothelin action inhibitor as an active ingredient.

  The method for producing an endothelin action inhibitor of the present invention can produce an endothelin action inhibitor that is highly effective in endothelin action suppression by containing eberic acid, which is an active ingredient of an endothelin action inhibitor, at a high concentration. Moreover, the manufacturing method of the Nagasaga Ogase extract of this invention can manufacture the Nagasal Ogase extract which contains the evelic acid which is an active ingredient of an endothelin action inhibitor in high concentration.

  In the production method of the present invention, Nagasagaogase is extracted at a temperature of 0 ° C. or more and less than 20 ° C. using an alcohol or an alcohol solution having a water content of 10% by volume or less as an extraction solvent. By this method, compared to the case of using an alcohol solution having a high water content or the extraction at a temperature of 20 ° C. or higher, Nagasagaogase extract and endothelin action inhibition containing eberic acid having an endothelin action inhibitory effect at a high concentration An agent is obtained.

In the present specification, the "Eberic acid high-content Nagasaga Ogase extract" refers to an extraction defined by the present invention in which the Ebernic acid content in the Nagasal Ogase extract obtained by extraction under the extraction conditions defined in the present invention is defined by the present invention. It means that there is more content of everonic acid in the extract obtained by extracting under conditions other than conditions. For example, as shown in the examples described later, when the extraction is performed under the extraction temperature condition (temperature of 0 ° C. or more and less than 20 ° C.) in the present invention, an alcohol or an alcohol solution having a water content of 10% by volume or less is used as the extraction solvent. The Nagasal Ogase extract obtained by using has a higher evernic acid content than the Nagasal Ogase extract obtained by using an alcohol solution having a water content higher than 10% by volume. Similarly, when extraction is performed using the extraction solvent (alcohol or alcohol solution having a water content of 10% by volume or less) used in the present invention, Nagasagaogase extract obtained when the extraction temperature is set to 0 ° C. or higher and lower than 20 ° C. Increases the content of eberic acid as compared to the Nagasaga Ogase extract obtained when the extraction temperature is 20 ° C. or higher.
Thus, the inclusion of everonic acid in the Nagasaga Ogase extract obtained when either one of the extraction temperature and the extraction solvent is a condition defined in the present invention and the other condition is also a condition defined in the present invention. As long as the amount of the other condition is high compared to the condition other than that defined in the present invention, the Nagasagaogase extract obtained under the extraction conditions defined in the present invention is referred to as “ It shall be included in “Everic acid high content Nagasaru Ogase extract”.

  In the Nagasaga Ogase extract, evernic acid may be present in the form of a salt. Although it does not specifically limit as a salt of evernic acid, For example, Alkali metal salts, such as lithium salt, sodium salt, potassium salt, Alkaline earth metal salts, such as calcium salt and magnesium salt, Alkylamine salts, such as trimethylamine and triethylamine, and 4 Examples thereof include quaternary ammonium salts, alkanolamine salts such as triethanolamine, diethanolamine and monoethanolamine, and amino acid salts such as lysine, histidine and arginine. Alkali metal salts, alkanolamine salts and amino acid salts are preferred.

The Nagasarogase used in the present invention is a kind of lichen belonging to the genus Parnealiaceae ( Usnea ). In the present invention, naturally-occurring Nagasarogose may be collected and used, or a commercially available product may be obtained and used.
In the present invention, any arbitrary part (overall, whole grass, lichen, branch, leaf, fruiting body, etc.) of Nagasaru Ogase can be used, and a plurality of parts may be used in combination.
The Nagasaru Ogase used in the present invention may be in a naturally existing state, but since the extraction temperature in the present invention is relatively low, in order to increase the extraction efficiency, such as drying, freeze drying, chopping, pulverization, etc. It is preferable to use one that has been treated.

  As shown in the examples described later, when Nagasaruogase is extracted at a temperature of 20 ° C. or higher, the alcoholysis of eberic acid occurs, and the yield of eberic acid is greatly reduced. Therefore, in this invention, Nagasaru Ogase is extracted at a temperature of 0 ° C. or more and less than 20 ° C. The extraction temperature is preferably 0 to 15 ° C. Although there is no restriction | limiting in particular in extraction time, 3 hours-30 days are preferable, and 8 hours-10 days are more preferable. In order to increase extraction efficiency, stirring may be performed during extraction, or homogenization treatment may be performed in a solvent. In addition, although there is no restriction | limiting in particular in the usage-amount of an extraction solvent, it is preferable that it is 1-50 times amount by mass ratio with respect to Nagasaru Ogase of an extraction raw material, and it is more preferable that it is 5-30 times amount.

  Examples of the alcohol used as the extraction solvent in the present invention include lower alcohols such as methanol, ethanol, isopropyl alcohol, and butanol (preferably lower alcohols having 1 to 4 carbon atoms), ethylene glycol, propylene glycol, 1,3-butylene. Examples thereof include polyhydric alcohols such as glycol. In the present invention, these solvents may be used alone or in combination of two or more. In the present invention, lower alcohols such as methanol, ethanol and isopropyl alcohol are preferably used, and methanol and ethanol are more preferably used.

  In the present invention, the water content of the alcohol or alcohol solution used as the extraction solvent is 10% by volume or less. As will be shown in Examples described later, when an alcohol solution having a high water content is used as an extraction solvent, the yield of everonic acid is greatly reduced due to hydrolysis of everonic acid. The water content of the extraction solvent is preferably 5% by volume or less.

  After the extraction step, a fraction having a high activity may be fractionated by a suitable separation means such as gel filtration, chromatography, precision distillation or the like, if necessary. The extract obtained by the production method of the present invention includes the various extracts thus obtained, a diluted solution thereof, a concentrated solution thereof, a purified product thereof or a dry powder thereof.

  As shown in the below-mentioned Examples, an ebernic acid-rich Nagasaruogase extract can be obtained by the extraction step. Therefore, since eberic acid can suppress the production of melanin by suppressing the action of endothelin on melanocytes, the Nagasaga Ogase extract obtained by the production method of the present invention has an excellent endothelin action suppressing effect and whitening effect. In the present specification, “whitening (action)” refers to the suppression of melanin formation and pigmentation to return to an original transparent skin color without excess melanin, or pigment such as skin blackening or stain / sobacas It means preventing and suppressing deposition.

  The Nagasaga Ogase extract obtained by the present invention may be used as it is as an endothelin inhibitor or whitening agent. Moreover, you may mix and use the Nagasaroogase extract obtained by this invention, and an appropriate liquid or solid excipient | filler or extender, such as a titanium oxide, a calcium carbonate, distilled water, lactose, starch, for example. Furthermore, other endothelin action inhibitors, other whitening agents, moisturizers, antioxidants, UV absorbers, surfactants, thickeners, colorants, oily substances, polymer compounds, preservatives, powders , Pigments, fragrances, emulsion stabilizers, pH adjusters, and the like, may be used in admixture with components used in ordinary whitening agents.

  The endothelin action inhibitor and whitening agent obtained by the method of the present invention can be applied to the skin and the like as skin cosmetics, external medicines, quasi-drugs, etc., and their dosage forms are also aqueous solutions, solubilization systems, It can take a wide variety of forms such as an emulsifying system, a powder system, a gel system, an ointment system, a cream, a water-oil two-layer system, and a water-oil-powder three-layer system. Examples thereof include face wash, lotion, milky lotion, cream, lotion, gel, essence (beauty serum), pack, mask, foundation, ointment, sheet-like product and the like.

There is no restriction | limiting in particular in content of the Nagasaru Ogase extract in the endothelin action inhibitor obtained by the method of this invention, and a whitening agent, and it can set suitably depending on the eberic acid density | concentration in Nagasaga Ogase extract.
Moreover, there is no restriction | limiting in particular in the usage-amount of the endothelin action inhibitor obtained by the method of this invention, and it changes with content of an active ingredient, For example, in the case of cream form and ointment form, it is 0.00 per 1 cm < ² > of skin surfaces. 1 to 5 μg is preferable, and in the case of a liquid preparation, 0.1 to 10 μg per 1 cm ^{2 of} the skin surface is preferable.

  EXAMPLES Hereinafter, although this invention is demonstrated further in detail based on an Example, this invention is not limited to these.

Test Example 1 Verification of endothelin action inhibitory action of eberic acid (production of eberic acid)
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 50 g and 50 L% ethanol aqueous solution 1 L were mixed, extracted at 20 ° C. for 14 days and filtered to obtain 940 mL of Nagasaru Ogase extract (solid content 1.76 g). The solid content of 55.3 mg in this extract was fractionated by HPLC to obtain 4.4 mg of everonic acid (yield 7.96%).
The structural analysis of the isolated component was performed by NMR. It was isolated because the NMR spectral data of the isolated component was almost identical to the spectral data of eberic acid reported in the literature (Phytochemistry, Vol. 48, No. 5, pp. 815-822, 1998) The component was identified as everonic acid. The results of structural analysis by NMR are shown in Table 1. In Table 1 below, Me represents a methyl group.

(Preparation of German chamomile extract)
German chamomile extract (Japanese name: chamomile, obtained from Shinwa Bussan Co., Ltd.) and 40 mL of 50 vol% ethanol-containing aqueous solution are mixed, extracted at room temperature for 14 days, filtered, and German chamomile extract (221 mL) (Evaporation residue 2.63%) was obtained.

  About the evelic acid and the German chamomile extract, the endothelin action inhibitory effect was verified using the following evaluation system. Endothelin is known to act on melanocytes to increase calcium concentration and increase melanin production (Yada Y. et al., J. Biol. Chem., 1991, vol. 266, p. 18352). -18357; Imokawa G. et al., J. Biol. Chem., 1992, vol. 267, p. 24675-24680, etc.), substances that suppress the action of endothelin are useful as whitening ingredients.

Normal human neonatal epidermis-derived melanocytes (NHEMs; manufactured by Kurabo Industries) are seeded in 96-well plates for fluorescence detection at 3 × 10 ⁴ cells / well (200 μL / well) and cultured at 37 ° C. under 5% CO _2. did. Medium 254 containing PMA (-) growth additive (HMGS) was used as the medium.
After culturing for 3 days, the medium was removed from the culture plate by decantation, replaced with assay buffer containing intracellular Ca ^{2 ++} measurement reagent Fluo4-AM, and incubated at 37 ° C. for 1 hour. Next, 20 seconds after the start of measurement in an FDSS (Functional Drug Screening System) instrument, pretreatment with a predetermined concentration of eberic acid or German chamomile extract is performed for 1 minute, followed by addition of the ligand endothelin (ET-1, final concentration 100 nM). Fluo4-AM fluorescence was measured using Ex. 480 nm / Em. Detection was performed over time from the start of measurement at a measurement wavelength of 540 nm until 5 minutes later.
Relative value (%) with Fluo4-AM fluorescence increase ratio (Max.ratio-Min.ratio) in control as 100, increase in intracellular calcium (calcium ion) concentration by endothelin stimulation in each compound or extract The rate (%) was calculated, and the endothelin action inhibitory effect was evaluated. In addition, what added ethanol was used as control.
The results are shown in Table 2.

German chamomile extract is known to suppress the increase in Ca concentration of melanocytes by the action of endothelin and suppress melanin production (for example, Document Pigment Cell Res., 1997, vol. 10, p. 218-228). reference). As is clear from Table 2, in the system to which the German chamomile extract was added, a concentration-dependent calcium concentration increase inhibitory action was confirmed. In particular, in a system with a concentration of 1 v / v%, an increase in calcium concentration was suppressed by 20% or more, and an excellent endothelin inhibitory action was confirmed.
On the other hand, as is clear from Table 2, in the system added with eberic acid, the increase in calcium concentration in melanocytes by endothelin stimulation was suppressed in a concentration-dependent manner. That is, it can be seen that evelic acid has an excellent endothelin inhibitory action equivalent to or higher than that of a German chamomile extract, which is a known whitening component, and is useful as a whitening component.

Comparative Example 1
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 10 volume% ethanol solution 200 mL were mixed, extracted at 20 ° C. for 8 days, filtered, and Nagasal Ogase extract 173.0 g (evaporation residue 0.342 (w / v)%).

Comparative Example 2
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 10 volume% ethanol solution 200 mL were mixed and extracted at 30 ° C. for 8 days followed by filtration. Nagasaru Ogase extract 169.6 g (evaporation residue 0.372 (w / v)%).

Comparative Example 3
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 10 volume% ethanol solution 200 mL were mixed, extracted at 40 ° C. for 8 days, filtered, and Nagasaru Ogase extract 168.6 g (evaporation residue 0.422 (w / v)%).

Comparative Example 4
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 50 g and 50 L% ethanol solution 1 L were mixed and extracted at 15 ° C. for 8 days, followed by filtration. Nagasaru Ogase extract 863.0 g (evaporation residue 0.334 (w / v)%).

Comparative Example 5
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 50 g and 50 volume% ethanol solution mL were mixed and extracted at 20 ° C. for 8 days followed by filtration. Nagasaga Ogase extract 858.5 g (evaporation residue 0.374 (w / v)%).

Comparative Example 6
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 50 g and 50 L% ethanol solution 1 L were mixed, extracted at 30 ° C. for 8 days, filtered, and Nagasal Ogase extract 853.5 g (evaporation residue 0.420 (w / v)%).

Example 1
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 95 volume% ethanol solution 200 mL were mixed and extracted at 15 ° C. for 8 days followed by filtration. Nagasaru Ogase extract 149.0 g (evaporation residue 0.276 (w / v)%).

Comparative Example 7
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 95 volume% ethanol solution 200 mL were mixed, extracted at 20 ° C. for 8 days, filtered, and 148.5 g of Nagasal Ogase extract (evaporation residue 0.260 (w / v)%).

Comparative Example 8
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 95 volume% ethanol solution 200 mL were mixed and extracted at 30 ° C. for 8 days followed by filtration. Nagasaru Ogase extract 147.3 g (evaporation residue 0.344 (w / v)%).

Comparative Example 9
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 95 volume% ethanol solution 200 mL were mixed and extracted at 40 ° C. for 8 days followed by filtration. Nagasaru Ogase extract 145.5 g (evaporation residue 0.414 (w / v)%).

Test Example 2 Measurement of Everonic Acid Concentration The Everonic acid concentration in each Nagasaruogase extract obtained in Example 1 and Comparative Examples 1 to 10 was analyzed under the following conditions using HPLC. The apparatus and measurement conditions used for the measurement are as follows.

Measuring instruments (HPLC): Agilent Technologies Open LAB CDS LC02-1260, Autosampler G1329B 1260 ALS, Column Oven G1316A 1260 TCC, Detector G1315D 1260 DAD VL, Pump G1312B 1260 Bin pump, G1379B 1260 μ-Degasser

<HPLC analysis conditions>
Flow rate: 0.75 mL / min
Column used: Inertsil ODS-3 5 μm (GL Science), 3.0 ID × 150 mm
Detection wavelength: 254 nm
Column temperature: 40 ° C
Mobile phase: (0.1% trifluoroacetic acid-containing aqueous solution): (0.1% trifluoroacetic acid-containing methanol) = 30: 70 (retention time of everonic acid: 8.900 minutes)

The results are shown in Table 3.

As is apparent from the results in Table 3, according to the method of the present invention, a Nagasaruogase extract containing a high content of eberic acid can be obtained.
On the other hand, when the water content in the alcohol solution used as the extraction solvent was high, the evernic acid concentration in the Nagasaru Ogase extract was greatly reduced (Comparative Examples 1 to 6). Further, when the extraction temperature is 20 ° C. or higher, even if an alcohol or an alcohol solution having a water content of 10% by volume or less is used, the concentration of everonic acid is greatly reduced as compared with the case of extraction at a low temperature (Comparative Examples 7 to 9). ).

Example 2
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 3.65 g and 100 volume% methanol 73 mL were mixed and extracted at 5 ° C. for 8 days followed by filtration. Nagasaru Ogase extract 48.7 g (evaporation residue 0.330 (w / V)%).

Example 3
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 100 volume% methanol 200 mL were mixed and extracted at 15 ° C. for 8 days followed by filtration. Nagasaru Ogase extract 144.2 g (evaporation residue 0.386 (w / v )%).

Comparative Example 10
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 100 volume% methanol 200 mL were mixed and extracted at 20 ° C. for 8 days followed by filtration. Nagasaru Ogase extract 143.4 g (evaporation residue 0.360 (w / v )%).

Comparative Example 11
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 100 volume% methanol 200 mL were mixed and extracted at 30 ° C. for 8 days followed by filtration. Nagasaru Ogase extract 143.7 g (evaporation residue 0.452 (w / v )%).

Comparative Example 12
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 100 volume% methanol 200 mL were mixed and extracted at 40 ° C. for 8 days followed by filtration. Nagasaru Ogase extract 143.72 g (evaporation residue 0.540 (w / v )%).

Example 4
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 100 volume% methanol 200 mL were mixed, extracted at 5 ° C. for 3 hours, filtered, and 144.2 g of Nagasal Ogase extract (evaporation residue 0.180 (w / v )%).

Example 5
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 100 volume% methanol 200 mL were mixed and extracted at 5 ° C. for 8 hours followed by filtration. Nagasaru Ogase extract 144.0 g (evaporation residue 0.184 (w / v )%).

Example 6
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 100 volume% methanol 200 mL were mixed and extracted at 5 ° C. for 24 hours, followed by filtration. Nagasaru Ogase extract 144.6 g (evaporation residue 0.198 (w / v )%).

Example 7
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd.) 10 g and 100 volume% methanol 200 mL were mixed and extracted at 5 ° C. for 48 hours followed by filtration. Nagasaru Ogase extract 144.4 g (evaporation residue 0.208 (w / v )%).

Example 8
Nagasaru Ogase (obtained from Shinwa Bussan Co., Ltd., China) 10 g and 100 volume% methanol 200 mL were mixed and extracted at 5 ° C. for 72 hours, followed by filtration. Nagasaru Ogase extract 145.0 g (evaporation residue 0. 216 (w / v)%).

Test Example 3 Measurement of Everonic Acid Concentration The Everonic acid concentration in each Nagasagaogase extract obtained in Examples 2-8 and Comparative Examples 11-13 was measured in the same manner as in Test Example 2. The results are shown in Table 4.

As is apparent from the results in Table 4, according to the method of the present invention, the extraction temperature is set to less than 20 ° C., and thus a Nagasaruogase extract containing a high content of eberic acid can be obtained.
On the other hand, when the extraction temperature was 20 ° C. or higher, the concentration of everonic acid was greatly reduced as compared with the case where extraction was performed at a low temperature (Comparative Examples 10 to 12).

  As described above, a Nagasaru Ogase extract containing a high amount of evernic acid is produced by extracting Nagasaga Ogase at a temperature of 0 ° C. or more and less than 20 ° C. using an alcohol or an alcohol solution having a water content of 10% by volume or less. be able to. Since eberic acid has an excellent endothelin inhibitory effect, Nagase Ogase is extracted with alcohol or alcohol or an alcohol solution having a water content of 10% by volume or less at a temperature of 0 ° C. or higher and lower than 20 ° C. An endothelin action inhibitor excellent in effect can be produced.

(Prescription example)
Endothelin action inhibitors or whitening agents in the form of lotions, milky lotions, cosmetic liquids and creams having the composition shown below, each containing the extract obtained in the above example as an active ingredient, were prepared by conventional methods.

1. Preparation of lotion (composition) (Formulation: mass%)
1,3-butylene glycol 8.0
Glycerin 5.0
Ethanol 3.0
Nagasaruogase extract obtained in Example 1 3.0
Chamomile extract 3.0
Kyokyo Extract 1.0
Clove extract 1.0
Xanthan gum 0.1
Hyaluronic acid 0.1
Disodium phosphate 0.1
Sodium dihydrogen phosphate 0.1
Ascorbic acid 2-glucoside 2.0
Purified water Remaining fragrance Appropriate amount Preservative Appropriate amount

2. Preparation of milky lotion (composition) (formulation: mass%)
Nagasaruogase extract obtained in Example 1 1.0
Chamomile extract 1.0
Kyokyo Extract 1.0
Altea extract 2.0
Squalane 3.0
Olive oil 3.0
Glycerin 5.0
Polyoxyethylene hydrogenated castor oil (added moles of ethylene oxide: 40) 1.0
Carboxyvinyl polymer 0.2
Potassium hydroxide 0.1
Xanthan gum 0.2
Edetate disodium 0.02
Purified water Remaining antiseptic

3. Preparation of serum (composition) (Composition: mass%)
Nagasaruogase extract obtained in Example 1 1.0
Chamomile extract 1.0
Kyokyo Extract 1.0
Clove extract 1.0
Carboxyvinyl polymer 0.2
Acrylic acid / alkyl methacrylate copolymer 0.2
Potassium hydroxide 0.2
Xanthan gum 0.1
Hyaluronic acid 0.2
Sodium citrate 0.15
Citric acid 0.03
Glycerin 10.0
1,3-butylene glycol 5.0
Edetate disodium 0.05
Purified water Remaining preservative Appropriate amount Perfume Appropriate amount

4). Preparation of serum (composition) (Composition: mass%)
Nagasagaogase extract obtained in Example 3 3.0
Chamomile extract 1.0
Clove extract 1.0
Kyokyo Extract 1.0
Xanthan gum 0.2
Carboxymethylcellulose 0.2
Carboxyvinyl polymer 0.2
Potassium hydroxide 0.1
Citric acid 0.03
Sodium citrate 0.15
Glycerin 5.0
Propylene glycol 3.0
Polyethylene glycol (molecular weight 1500) 1.0
Polyethylene glycol monostearate 0.5
Purified water Remaining antiseptic

5. Preparation (composition) of cream (formulation: mass%)
Nagasagaogase extract obtained in Example 3 3.0
Chamomile extract 2.0
Kyokyo Extract 2.0
Clove extract 2.0
Methylpolysiloxane 3.0
Squalane 2.0
Neopentyl glycol dicaprate 3.0
Stearyl alcohol 1.5
Cetanol 1.0
Polyoxyethylene hydrogenated castor oil (addition moles of ethylene oxide: 60) 0.5
Acrylic acid / alkyl methacrylate copolymer 0.3
Potassium hydroxide 0.15
Xanthan gum 0.1
Edetate disodium 0.05
Purified water Remaining preservative Appropriate amount Perfume Appropriate amount

Claims (7)

A method for producing a Nagasaru Ogase extract with a high content of evelic acid, comprising a step of extracting Nagase Ogase ( Usnea longissima ) at a temperature of 0 ° C. or more and less than 20 ° C. using an alcohol or an alcohol solution having a water content of 10% by volume or less.   The manufacturing method of Claim 1 whose said alcohol or alcohol solution is 100 volume% methanol.   The manufacturing method of Claim 1 whose said alcohol or alcohol solution is an ethanol solution whose water content is 10 volume% or less. The manufacturing method of an endothelin action inhibitor including the process of extracting Nagasaru Ogase ( Usnea longissima ) using the alcohol or alcohol or alcohol solution whose water content is 10 volume% or less at the temperature of 0 degreeC or more and less than 20 degreeC .   The manufacturing method of Claim 4 whose said alcohol or alcohol solution is 100 volume% methanol.   The manufacturing method of Claim 4 whose said alcohol or alcohol solution is an ethanol solution whose water content is 10 volume% or less. A whitening agent comprising, as an active ingredient, a Nagasal Ogase extract or an endothelin action inhibitor obtained by the production method according to claim 1.