KR100406125B1 - A compositions for skin external application containing the torilin - Google Patents

Abstract

The present invention relates to an external composition for skin containing torilin represented by the following formula (1) as an active ingredient for hair growth, and more specifically, tolopecia, including tolylin, which has been separated and purified from a bee box, as an active ingredient. To inhibit the activity of the 5 alpha-reductase (5α-reductase) associated with, to prevent hair loss and provides a composition for external application for skin having a very good hair growth promoting effect.

Description

A composition for skin external application containing the torilin}

The present invention relates to a topical skin composition containing torilin represented by the following formula (1). More specifically, it is an external preparation that contains tolylin extracted from the bee scab box as an active ingredient to inhibit the activity of 5 alpha-reductase related to alopecia and impart very good hair growth effect. It can be mix | blended with external skin preparations, such as a cosmetics.

[Formula 1]

The human hair is about 100,000 to 150,000 pieces, each hair has a different cycle and grow and drop out through the anagen, catagen, and telogen. This cycle is repeated over three to six years, with the result that on average 50-100 hairs fall out normally. Generally, the term alopecia refers to the increase in the number of hairs that fall abnormally due to shortening of the hair growth rate in the growth phase and more hair in the degenerative or resting phase.

On the other hand, androgenetic alopecia is directly dependent on the amount of male hormones because it is dependent on male hormones, and thus, many studies have recently been reported for the prevention and treatment of hair loss through inhibition of male hormone activity. Describing the mechanism of hair loss caused by male hormones, 5 alpha-reductase is present in testosterone-reactive tissues such as sebaceous glands, hair follicles, prostate, and epididymis and is one of the male hormones. dihydrotestosterone) is an enzyme involved in metabolism, and its conversion requires NADPH. In addition, testosterone is involved in male sexual impulses, skeletal muscle increase, male external genitalia, scrotum growth, spermatogenesis, etc., and dihydrotestosterone (dihydrotestosterone) is involved in tissues such as acne, sebum increase, hair loss, and prostatic hypertrophy ( Diane et al. JID 1995). Therefore, studies are being actively conducted to develop sebum secretion inhibitors and anti-hair loss agents using inhibitors of this enzyme. Excessive secretion of male hormones after puberty causes acne and hair loss, and studies are attempting to develop hair restorers or hair loss inhibitors using 5-alpha-reductase inhibitors to prevent excessive production of dihydrotestosterone in hair follicle cells of hair follicles. This is going on.

Therefore, a female hormone such as estrogen has been conventionally used for the treatment of androgenetic alopecia due to this testosterone suppression. However, such female hormones, as well as the effect, there are reports of the occurrence of side effects due to skin inflammation, hormone administration, etc. Currently, the use of a very small amount is discontinued. Recently, Merck has been commercially available as a hair restorer to synthesize finasteride, an inhibitor of 5 alpha-reductase, but it has been limited to use due to problems such as expensive side and side effects. There is a situation.

In order to solve the above problems, the present inventors studied materials that are safe for human body while preventing hair loss, and torilin, especially, tolylin extracted from a bee box was 5 alpha-reducta. The present invention has been accomplished by discovering that it inhibits azease and exhibits very good hair growth effects.

Accordingly, it is an object of the present invention to provide an external preparation for skin of 5-alpha-reductase inhibition having excellent hair loss preventing effect and hair growth effect.

Figure 1 shows the ¹ H-NMR spectrum of torilin.

Figure 2 shows the ¹³ C-NMR spectrum of torilin.

3 shows the EI-MS spectrum of torilin.

Figure 4 is a graph showing the effect on the 5 alpha-reductase activity when the amount of extract of the bee box.

5 is a graph showing the degree to which the bee box fractions inhibit 5 alpha-reductase.

Figure 6 is a graph showing the inhibitory ability of each of the tollin (torilin) for 5 alpha-reductase.

In order to achieve the above object, the present invention provides a topical skin composition containing tolyrin represented by the formula (1).

[Formula 1]

The external preparation composition for skin is characterized by inhibiting the activity of 5-alpha-reductase by containing tolyrin. The external preparation composition which concerns on this invention can be used especially as an external preparation for hair growth. Preferably, the tolyrin is used to extract from the bee box.

Hereinafter, the present invention will be described in detail.

In general, the bee- eater is also called the Toriolis japonica Decandolle , and the fruit of the herb is a biennial with a ridge growing in the grass. In oriental medicine, it is used as astringent anti-inflammatory agent, and it is useful for genital pruritus, umjong, and postpartum oro. In addition, folk remedies are known to improve the symptoms of women's itching and secretion from cold caused by boiled bees box boiled with water or bathed in bath water.

On the other hand, the extract of beolsa box of the present invention is extracted with lower alcohol (alcohol), ethyl acetate (ethylacetate), hexane (hexane) or diethylether, or anhydrous or hydrous ethanol (methanol) or methanol (methanol) extract Hexane (hexane), ethyl acetate (ethylacetate), can be used to extract the fraction of the water layer.

Skin external preparations containing tolylin, which is an extract of the bee sac box of the present invention, include softening cream, nutrient cream, massage cream, essence, pack and hair tonic, hair lotion and the like. More preferably, the formulation is prepared in the form of an ointment and applied to the scalp.

Tolylin, which is an extract of the bee venom, may be contained in an amount of 0.0001 to 5% by weight, preferably 0.001 to 1% by weight, based on the dry weight of the skin external preparation composition.

Hereinafter, the present invention will be described in more detail.

After purchasing commercially available bee-box herb medicine, it is applied to organic solvents such as ethanol, methanol, etc., 3 ~ 5 times the dry weight alone, and 50 ~ 100 in the extractor equipped with cooling capacitor. Extract by heating to 1-10 hours. Alternatively, the active ingredient may be extracted by dipping at 0 to 40 ° C. for 1 to 30 days in the solvent used above. The bee box extract extracted by the above method is completely dried using a reduced pressure evaporator.

The present invention will be described in more detail with reference to Examples and Test Examples of the present invention, but the present invention is not limited only to these examples.

As used in the present invention, beolsa box extract means that the active ingredient is extracted and concentrated to dry by the method of Example 1-1, 1-2, 1-3.

Example 1 Preparation of a bee box extract

1-1. Preparation of bee box extract

250 g of 70% ethanol was added to 100 g of the bee box, and the extract was heated and extracted at 78 ° C., which is the boiling point of ethanol, in an extractor equipped with a cooling condenser for 2 hours. The extract was concentrated under reduced pressure at 60 ° C. using a distillation apparatus equipped with a cooling capacitor to obtain a dry residue 2g.

1-2. Bee box extract manufacture

200 g of chloroform was added to 100 g of the bee box, and the extract was heated and extracted at 61 ° C., which is a boiling point of chloroform, in an extractor equipped with a cooling capacitor for 2 hours. The extract was concentrated under reduced pressure at 60 ° C. using a distillation apparatus equipped with a cooling capacitor to obtain a dry residue of 1.5 g.

1-3. Preparation of bee box extract

200 ml of ethylacetate (ethylacetate) was added to 100 g of the bee box, and the extract was heated at 77 ° C., the boiling point of ethyl acetate in an extractor equipped with a cooling capacitor, extracted for 2 hours, filtered to remove the residue, and a bee box extract was prepared. The extract was concentrated under reduced pressure at 50 ° C. using a distillation apparatus equipped with a cooling capacitor to obtain 2.1 g of dry residue.

Example 2 Purification of Toriiline

10 g of a bee-box ethylacetate extract obtained in Examples 1 to 3 was dissolved in ethylacetate, adsorbed onto a minimum amount of silica gel, and 150 g of silica gel was suspended in hexane to fill the tube. Loaded on. Hexane (hexane: ethylacetate) mixed solution (10: 1, 5: 1, 4: 1, 3: 1, 2; 1, 1; 1) through the 15 fractions in 100ml order Was collected and 5alpha-reductase inhibitory activity was investigated. In the fourth fraction (0.13 g), 5 alpha-reductase inhibitory activity was observed, which was again preparative silica gel TLC under the condition of hexane-ethyl acetate (3: 1) to give 25.1 mg of torilin.

Example 3 Structure Determination of Tolyrin

Instrumental analysis was performed to identify the structure of torilin obtained in Example 2.

¹ H-NMR and ¹³ C-NMR spectra (Jeol spectrometer), IR spectrum (Bio-Rad FTS-40 spectrometer), and mass spectrum (JMS-700 Mass spectrometer (Jeol)) were measured and The results are shown in Table 1 and FIGS.

A characteristic light absorption band of an ester group was found at IR? Max cm ⁻¹ (KBr): 1725, 1259 cm ⁻¹ , and a characteristic light absorption band of an alpha, beta unsaturated ester group was observed at 1695, 1639 cm ⁻¹ .

Spectroscopy of torilin (CDCl ₃ ) Carbon number ¹³ C shift DEPT 1H shift One 51.14 CH 2.44m 2 41.22 CH ₂ 2.07dd (18.3, 3.0) 2.59dd (18.3, 6.0) 3 208.26 C 4 135.02 C 5 173.68 C 6 25.74 CH ₂ 2.50 br d (13.5) 2.87d (13.5) 7 46.35 CH 2.41dd (10.2, 3.6) 8 70.77 CH 5.45 ddd (7.5, 7.5, 3.6) 9 40.55 CH ₂ 1.63m 2.26ddd (14.5, 7.5, 1.2) 10 33.45 CH 1.49m 11 84.47 C 12 23.98 CH ₃ 1.51 s 13 24.57 CH ₃ 1.53 s 14 22.71 CH ₃ 1.02d (6.9) 15 8.06 CH ₃ 1.72d (1.5) One' 15.83 CH ₃ 2.01dq (1.5, 7.2) 2' 138.34 CH 6.08qq (1.5, 7.2) 3 ' 127.79 C 4' 20.63 CH ₃ 1.90m 5 ' 166.86 C COCH ₃ 22.70 CH ₃ 1.98 s 170.40 C

Hereinafter, a formulation example is demonstrated.

Example 4 Nutrients

ingredient Content (% by weight) Bee box extract 0.5 Beeswax 4.0 Polysorbate 60 1.5 Solbitan Sesquiolade 0.7 Liquid paraffin 5.0 Squalane 5.0 Caprylic / Capric Triglycerides 5.0 glycerin 3.0 Butylene glycol 3.0 Propylene glycol 3.0 Carboxy Vinyl Polymer 0.1 Triethanolamine 0.2 antiseptic Quantity Pigment Quantity Spices Quantity Purified water To 100

Example 5 Hair Tonic

ingredient Content (% by weight) ethanol 55.0 Castor Oil 5.0 glycerin 3.0 Pyrotonamine 0.1 Bee box extract 1.0 Spices Quantity Purified water To 100

Example 6 Hair Lotion

ingredient Content (% by weight) Cetostearyl alcohol 2.0 Stearyl Triethyl Ammonium Chloride 2.0 Hydroxyethyl cellulose 0.5 Pyrotonolamine 0.1 Bee box extract 1.0 Spices 0.5 Purified water To 100

[Test Example 1] 5 alpha-reductase (5α-reductase) inhibitory effect test

Mature Sprague-Dawley male rats (7-8 weeks old) were killed with diethyl ether to remove abdominal prostate and remove connective tissue. A buffer solution (0.32M sucrose, 0.1mM dithiothreitol, 20mM sodium acetate) was added thereto, finely chopped with a grinder and suspended. The suspension was filtered with gauze, centrifuged at 1500 x g, and the pellets were taken and suspended in the buffer at a concentration of 20-30 mg / ml.

After taking a portion of the suspension was placed in a 0.02 molar (M) phosphate buffer, the substrate with the isotopes ³ H is attached, by reacting with testosterone (testosterone) was measured for production of DHT (dihydrotestosterone) in the product.

At this time, the reaction solution is dithiothreitol (dithiothreitol), 20mM of phosphate salt of 1mM, pH 6.5, in 50mM NADPH, [1,2,6,7- ³ H] testosterone 1pM / testosterone (1 × 10

^-9 M), 565 mL of 36 mg enzyme suspension.

A certain amount of Example 1 was dissolved in 10% dimethyl sulfoxide (DMSO) and added to 10 ml per reaction, and the same volume of solvent was used as a control experiment, and riboflavin was used as a positive control experiment. In addition, the reaction proceeded at 37 ° C. for 30 minutes by adding an enzyme suspension, and then 600 ml of ethylacetate was added to stop the reaction and the product was dried through nitrogen gas. In this 50㎖ of ethyl acetate (ethylacetate) the total amount of plastic sheets silica gel 60 F ₂₅₄ kayijel was dissolved in _{(Silica plastic sheet kieselgel 60 F 254} ) was spotted on a developing solvent of ethyl acetate (ethylacetate) - cyclohexane (cyclohexane) Developed using (1: 1).

Next, after drying the silica plastic sheet in air, the Bas system was used to measure the amount of isotopes, and the dried plastic sheet and the exposed film were put together in a Bas cassette and stored for one week. The isotope amounts of testosterone and dihydrotestosterone produced in the film were then quantified using the exposed intensity. The experiment was used repeatedly for two times, and the inhibition rate was calculated by the following calculation method based on the measurement results.

* T: Amount of ³ H radioactivity in testosterone (5a-reductase) region

DHT: The amount of ³ H radioactivity in the dihydrotestosterone region

* Conversion rate: Radiation amount / total radiation amount in DHT area

Inhibition rate: 100 x (control conversion-sample conversion) / control conversion

5 alpha-reductase inhibitory effect was observed by varying the amount of the bee box extract, the inhibition rate was calculated by the above calculation method, and the results are shown in FIG. 4.

In addition, 5 alpha-reductase inhibitory activity of the fractions obtained by performing silica gel tube chromatography as described in Example 2 with respect to the bee box extract, and the results are shown in FIG. In addition, 5 alpha-reductase inhibitory activity was measured by varying the amount of purely separated tolyrin as in Example 2, and is shown in FIG. 6.

As can be seen in the accompanying FIG. 4, it can be seen that tolylin, which is a bee-box extract of the present invention, has an inhibitory effect of 5 alpha-reductase. In addition, as can be seen in Figure 5, it was confirmed that 5 alpha-reductase inhibitory activity is strong in 4 of 15 fractions, IC from 50 to inhibit 5 alpha-reductase of tolyrin from Figure 6 _{The 50} value was calculated to be 0.4 µg / ml.

Test Example 2 Hair Growth Effect Test

A 30% hydrous ethanol solution containing 2% of the plant extract dry extract prepared in Examples 1 to 3 was prepared and used for the test. The hair growth effect test was performed using a mouse (ICR), 47-53 days of age, to remove the back hairs, and the back area was selected to be clean. Each ml was applied.

The length of hair and the degree of hair growth over time were compared by adding scores according to the degree of restoration after hair removal. In order to compare the degree of hair growth, as a control, a 30% alcohol solution was applied to each individual to observe the growth state. The results are shown in Table 2 below.

Elapsed days 10 12 15 18 Examples 1-3 0.49 1.02 2.64 ± 0.45 2.95 Control 0.11 0.41 1.03 ± 0.33 1.75 ± 0.40

As can be seen in Table 2, it was confirmed that the plant extract containing tolyrin according to Examples 1 to 3 has a hair growth promoting effect.

As described above, tolylin (torinlin), a bee-box extract provided by the present invention has an excellent effect of inhibiting 5 alpha-reductase and promotes hair growth at a concentration that does not affect the safety of the product. It is effective and can be used in effective concentrations for external skin preparations that exhibit hair loss prevention and hair growth effects.

Claims (4)

A composition for external application for skin, containing tolyrin represented by the following formula (1) as an active ingredient. [Formula 1] The composition for external application for skin according to claim 1, wherein the tolyrin inhibits the activity of 5-alpha-reductase. The external preparation composition for skin according to claim 1, wherein the external preparation composition for skin can promote hair growth. The composition for external application for skin according to claim 1, wherein the tolyrin is a bee box extract.