JPH0859438A - Hair treating agent - Google Patents

Abstract

PURPOSE: To obtain a hair treating agent capable of increasing the binding amount of anti-hair anti-bodies with hair and imparting the smoothness and flexibility to the hair. CONSTITUTION: This hair treating agent comprises containing a carrier binding a proteolytic enzyme. Trypsin, chymotrypsin, pepsin, papain, cathepsin B1, etc., are cited as the proteolytic enzyme, above all a thiol protease capable of being activated in the presence of a reducing agent such as papain, cathepsin B1, etc., are prefergble, and especially papain is preferred. An agarose gel, a polystyrene, a carboxy-modified polystyrene latex, a liposome, a clextran, etc., are cited as the carrier. The amount of the blended proteolytic enzyme is determined so that 0.01-1 unit thereof based on 50mg hair may be treated, and, e.g. 0.5-20 units based on 1g hair treating agent is preferable. This treating agent does not damage cuticles on the surface of the hair, decomposes keratinized protein present in the surface layer of the cuticles and markedly increases the binding of anti-hair antibodies to the surface of the hair.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a hair treatment agent which increases the amount of anti-hair antibody bound to hair.

[0002]

2. Description of the Related Art In recent years, research on hair cosmetics using anti-hair antibodies obtained by immunizing hair with animals has become active (JP-A-4-29912 and JP-A-4-414).
No. 13, etc.). However, these anti-hair antibodies did not have sufficient binding strength when actually applied to hair. On the other hand, papain has been reported to be a thiol protease protein present in papaya milk with a molecular weight of 23,406 that decomposes the cuticle interstitium of hair to exfoliate cuticle cells (Textile. Res. J. 35.
Vol. 1014, 1965). A composition in which this papain is used in combination with citric acid is disclosed in Japanese Examined Patent Publication No. 6-15462, and it is shown that this composition has an effect of giving gloss to the hair. However, in this composition for hair, papain penetrates deeply into the interstitium and reacts without sufficiently degrading the keratinized protein present in the cuticle surface layer, which causes hair damage such as cuticle cell exfoliation. There was a problem that I could not escape.

As a result of intensive studies to improve the above-mentioned drawbacks, the present inventors have found that proteolytic enzymes such as papain
It has been found that, by using in combination with a reducing agent or by being bound to a carrier, not only the binding property of an antibody to hair is improved, but also smoothness and suppleness are given to the hair, and the present invention has been completed. It was

[0004]

SUMMARY OF THE INVENTION Therefore, an object of the present invention is to provide a hair treatment agent which can increase the amount of anti-hair antibody bound to hair and which imparts smoothness and suppleness to hair. is there.

[0005]

Means for Solving the Problems The above objects include a hair treatment agent containing a protease-binding carrier, a hair treatment agent in which the protease is a thiol protease, a thiol protease or a thiol protease-binding carrier and a reducing agent. The hair treatment agent is achieved by the hair treatment agent wherein the thiol protease is papain.

The present invention will be described in detail below.

The proteolytic enzyme used in the present invention includes trypsin, chymotrypsin, pepsin, papain, cathepsin B1 and the like. Particularly, thiol proteases such as papain and cathepsin B1 which are activated in the presence of a reducing agent are used. Papain is particularly preferable. Examples of the proteolytic enzyme used in the present invention include natural extracts and commercial products.

Examples of the reducing agent used in the present invention include dithiothreitol, 2-mercaptoethanol, cysteine and sodium sulfite.

The hair treatment method used in the present invention includes, for example, to activate proteolytic enzymes and at the same time to effectively cleave disulfide bonds which are considered to be present in keratinized proteins present in the cuticle surface layer. In addition, it is preferable to use a protease together with a reducing agent.

In the present invention, the carrier used as the proteolytic enzyme-binding carrier includes agarose gel, high molecular polymers such as polystyrene, latex such as carboxy-modified polystyrene latex, insoluble carriers such as liposomes and amylose, dextran, gelatin, collagen, Examples thereof include water-soluble polymers such as polylysine.

To bind these carriers to the proteolytic enzyme, known methods may be used, including physical and chemical methods.

When the proteolytic enzyme is used by being bound to a carrier, it is possible to enhance the reaction specificity for the keratinized protein existing on the cuticle surface layer without allowing the proteolytic enzyme to penetrate into the carrier, and the cuticle is keratinized. Since only a part can be decomposed, the action on the cuticle is mild, which is preferable. Therefore, the size of the carrier depends on the width of the cuticle cell stroma 180 Å (Textil
e. Res. J. 35, 1014, 1965).

Examples of the form of the hair treatment agent of the present invention include a proteolytic enzyme, a reducing agent, a proteolytic enzyme-binding carrier and the like mixed with a general buffer solution such as a sodium phosphate buffer solution (pH 6.7). However, it may be in the form of usual treatment, conditioner, shampoo, hair cream, hair lotion and the like.

The reaction conditions of the hair treatment agent of the present invention are preferably carried out in the vicinity of the optimum pH of the enzyme to be used. For example, the reaction of papain and the reducing agent on the hair is performed at 25 ° C. with a buffer such as a phosphate buffer. It is better to react in liquid (pH 6.7).

The proteolytic enzyme of the present invention may be added to the hair treatment agent in an amount of 0.01 to 1 unit per 50 mg of hair. For example, it may be 0.5 per 1 g of hair treatment agent.
~ 20 units are preferred. However, in order to increase the amount of anti-hair antibody bound to hair, 0.01 to 50 mg of hair is used.
0.5 g per 1 g of hair treatment agent so that 0.1 unit is treated
It is preferable to set it to 2 units. One unit here is the amount of enzyme that hydrolyzes 1 μmol of benzoyl-L-arginine ethyl ester at 25 ° C. and pH 6.2 per minute.

In the present invention, the amount of the reducing agent added to the hair treatment agent is preferably 0.1 to 2% by weight based on the total amount of the hair treatment agent.

In the present invention, the amount of the proteolytic enzyme bound to the carrier in the hair treatment agent is 50 mg of hair.
It may be determined so that 0.1 to 10 units are treated, preferably 1.0 to 20 units per 1 g of the hair treatment agent. However,
In order to increase the binding amount of the anti-hair antibody to the hair, it is preferable that the amount is 1 to 200 units per 1 g of the hair treatment agent so that 0.1 to 10 units are treated for 50 mg of hair.

The reaction conditions for the hair treating agent of the papain-binding carrier of the present invention to the hair are, for example, reaction at 25 ° C. in a buffer solution (pH 6.7) such as a phosphate buffer solution.

The anti-hair antibody used in the present invention is an antibody against a protein which is a constituent of hair, such as an anti-keratin antibody and an anti-matrix antibody. These antibodies are disclosed in
It can be manufactured according to the methods described in JP-A-41413 and JP-A-5-279231.

The hair treatment agent of the present invention is present on the cuticle surface layer without damaging the cuticle on the hair surface by using a proteolytic enzyme and a reducing agent in combination, or by using a proteolytic enzyme bound to a carrier. By degrading the keratinized protein, the binding of anti-hair antibody to the hair surface can be significantly increased.

[0021]

The present invention will be described in more detail with reference to the following examples.

Prior to the examples, a method for preparing an antibody latex, a hair treatment of an antibody latex used for evaluating a hair treatment agent, a hair dyeing evaluation and an electron microscope observation method will be described.

Method for producing antibody latex 1. Preparation of antibody According to the method described in Japanese Patent Laid-Open No. 5-155783,
The anti-keratin antibody (anti-hair antibody) was prepared as follows. After washing 10 g of healthy hair, 0.2 M Tris-HCl buffer containing 8 M urea and 0.2 M 2-mercaptoethanol (pH 9.
2) The extract obtained by repeating the extraction operation in 2.5 l at 50 ° C. for 1 hour was centrifuged. Add 400 g of supernatant to 760 ml of Tris solution
200 g of a monoiodoacetic acid solution was added, and the mixture was stirred and reacted at room temperature for 1 hour. The reaction was stopped by adding 7 ml of 2-mercaptoethanol, and after dialyzing against a sufficient amount of water, a permeate was obtained through a 2 μm filter. 0.4 volume of this permeate is 0.
One part by volume of 5M sodium acetate buffer (pH 4.2) was added, the pH was adjusted to 4.2 with acetic acid, and hair keratin was subjected to isoelectric focusing. The precipitate obtained by centrifugation was dissolved in physiological saline, sterilized by passing through a 2 μm filter, and then concentrated with an ultrafiltration membrane to obtain a hair keratin antigen.

The hair keratin antigen was dissolved in physiological saline (protein concentration 20 mg / ml) and mixed with Freund's complete adjuvant to prepare a water-in-oil emulsion. 5.0 ml of this emulsion was subcutaneously administered to the neck of a pregnant Holstein cow two months before birth. At an interval of 10 days thereafter, an emulsion containing the same amount of antigen as the initial immunization prepared with Freund's incomplete adjuvant was administered subcutaneously for the 1st to 3rd booster immunizations and the 4th and 5th intramuscular injections. did.

Colostrum of the immunized cow was collected for 3 days after delivery. The fat layer was removed from the colostrum using a cream separator to obtain skim milk. Add 0.1N hydrochloric acid to this skim milk to adjust the pH.
It was adjusted to 4.5 and casein was precipitated. After removing the precipitate with a filter cloth, a supernatant was obtained by continuous centrifugation at 2,500 × g. After neutralization, ammonium sulfate was added to reach 33% saturation, and the antibody was salted out. The precipitate was collected by continuous centrifugation at 2,500 xg and dissolved in physiological phosphate buffer. The obtained solution was dialyzed against 10 mM phosphate buffer (pH 7.5) and applied to 2 l of DEAE cellulose column (DE-52, Whatman) equilibrated with the same buffer in 5 times. . After washing off non-adsorbed protein with the same buffer, the antibody was eluted with the same buffer containing 50 mM sodium chloride to obtain 200 g of the antibody. The purity of this antibody was 90% or more.

This antibody was applied to 400 ml of an affinity carrier (Affigel 15, Bio-Rad), to which hair keratin was bound by a conventional method, in 5 batches. The anti-hair antibody bound to the affinity carrier was treated with 0.2 M glycine hydrochloride buffer (pH
After elution in 2.5), the pH was immediately adjusted to around 8 with 3M Tris solution to obtain an anti-hair keratin antibody that specifically binds to the hair keratin antigen. Hereinafter, it is simply referred to as an anti-keratin antibody.

An antibody was purified from colostrum of non-immunized cows (antibody purity of 90% or more) and used as a control antibody.

2. Binding of Antibody to Latex Carboxy-modified polystyrene latex (particle size: 0.19 μm, manufactured by Nippon Synthetic Rubber Co., Ltd.) adsorbing a red dye was dispersed in 1 ml of purified water at a solid content concentration of 1%. To this, 1 mg of anti-keratin antibody or control antibody was dissolved in 1 ml of physiological phosphate buffer, and the mixture was stirred at 4 ° C. overnight. The supernatant was removed by centrifugation, and the suspension was dispersed in a physiological phosphate buffer containing 0.1% bovine serum albumin. Centrifuge again, redisperse in physiological phosphate buffer containing 0.1% bovine serum albumin, and resuspend in physiological phosphate buffer containing 0.1% bovine serum albumin and 0.1% polyoxyethylenesorbitan monolaurate (Tween 20). An antibody latex was obtained.

3. Method for treating hair with antibody latex The pretreatment agent for anti-hair antibody described in Examples below is applied at 25 ° C. for 24 hours.
After treating the hair bundle (50 mg) under water for a period of time and washing with water, the antibody latex dispersed in a physiological phosphate buffer containing 0.1% bovine serum albumin and 0.1% polyoxyethylene sorbitan monolaurate (Tween 20) was applied to the hair. The bundle was added and rotated for 1 hour. Then, it was washed with physiological saline containing 0.05% Tween 20 and dried.

4. Hair dyeing evaluation After carrying out antibody latex treatment, the degree of coloring can be visually inspected.
It was judged according to the following criteria. 4 ... Remarkably dark red. 3: It was dyed in deep red. 2 ... It was dyed red. 1 ... It was dyed in a light red color. 0 ... It didn't stain at all.

5. Electron microscopy Observation of hair after antibody latex treatment with ion sputter (E-102,
After depositing platinum-palladium with Hitachi, Ltd., the binding state of antibody latex and the degree of damage to the cuticle were observed with a scanning electron microscope (S-800, Hitachi).

Examples 1 to 5 (hair treatment agent containing papain and reducing agent) Papain (22.8 units / mg, manufactured by Worthing) 0.01, 0.
03, 0.1, 0.3, 1 unit and 0.2% dithiothreitol were dissolved in 1 ml of 0.1N sodium phosphate buffer (pH 6.7) to prepare a hair treatment composition.

Examples 6 to 10 (Agarose as a papain-binding carrier-hair treatment agent containing papain) Agarose-papain (8.5 units / mg, Elastin Pro
ducts) 0.1, 0.3, 1, 3, 10 units and 0.2%
Add dithiothreitol to 0.1N sodium phosphate buffer (pH
6.7) Dissolved in 1 ml to prepare a hair treatment agent.

Comparative Example 1 (Reducing agent-containing hair treatment agent) 0.2% dithiothreitol was dissolved in 1 ml of 0.1N sodium phosphate buffer (pH 6.7) to prepare a hair treatment agent.

Test Example 1 (Hair Dyeing Test of Human White Hair Bundles) Human white hair bundles were treated with the reducing agent-containing hair treating agent obtained in Comparative Example 1, the papain-containing hair treating agent obtained in Examples 1 to 5 and execution. The agarose-papain-containing hair treatment agents obtained in Examples 6 to 10 were reacted at 25 ° C. for 24 hours, washed with 5 liters of purified water, treated with antibody latex, and then evaluated for hair dyeing. Indicated. In the human hair bundle treated with papain, the degree of coloring by the anti-keratin antibody latex was increased as compared with the treatment with only the reducing agent. In addition, a wide range of 0.1 to 10 units of agarose-papain-treated human hair bundles of human hair showed increased coloration by anti-keratin antibody latex.
The control antibody latex did not stain when treated with either papain or agarose-papain.

[0036]

[Table 1]

Test Example 2 (Electron Microscopic Observation of Human Black Hair) Human black hair was treated at 25 ° C. with the reducing agent-containing hair treating agent obtained in Comparative Example 1 and the agarose-papain-containing hair treating agent obtained in Examples 6-10. After reacting for 24 hours, it was washed with 5 liters of purified water and treated with antibody latex. Electron micrographs taken after the treatment are shown in FIGS. As you can see from Figures 1-6,
In Comparative Example 1, the anti-hair antibody was bound only to the cuticle cleft, but in Example, it was evenly bound to the entire hair surface. Also, in the examples, it was possible to significantly increase the binding of the antibody latex to the hair surface without damaging the cuticle on the hair surface.

In any of the methods of Examples 1 to 10, human hair did not damage the cuticle on the hair surface, and remarkably bound the antibody latex to the hair surface by degrading the keratinized protein present on the surface layer of the cuticle. It is clear to increase.

Example 11 (Agarose-papain-containing hair treatment) Agarose-papain and a reducing agent were added to the hair treatment to prepare an agarose-papain-containing hair treatment. The compounding composition is as follows.

[0040]

[Table 2]

Comparative Example 2 (Reducing Agent-Containing Hair Treatment) A reducing agent-containing hair treatment was prepared by incorporating a reducing agent into the hair treatment. The compounding composition is as follows.

[0042]

[Table 3]

The panel 20 persons were divided into two groups, and after the hair treatments of Example 11 and Comparative Example 2 were continuously used for 2 weeks, the degree of improvement of smoothness and suppleness of hair was examined by a questionnaire, 11 is good
9 out of 10 people, 1 was a little better. On the other hand, in Comparative Example 2, 1 out of 10 people answered that it was good, 2 people answered that it was slightly good, and 7 people answered that it did not change. In addition, Example 11 imparted smoothness and suppleness to the hair.

[0044]

Industrial Applicability According to the present invention, it is possible to provide a hair treatment agent capable of increasing the binding of an anti-hair antibody obtained by immunizing hair keratin or the like to hair and imparting smoothness and suppleness to the hair. Is clear.

[Brief description of drawings]

FIG. 1 is a view showing an electron micrograph showing a result of using a reducing agent-containing hair treatment agent of Comparative Example 1 in Test Example 2.
(B) is an enlarged view of a part of (A) (a portion surrounded by □).

FIG. 2 is a diagram showing an electron micrograph of results obtained by using the reducing agent-containing hair treatment agent of Example 6 in Test Example 2.
(B) is an enlarged view of a part of (A) (a portion surrounded by □).

FIG. 3 is a diagram showing an electron micrograph of results obtained by using the reducing agent-containing hair treatment agent of Example 7 in Test Example 2.
(B) is an enlarged view of a part of (A) (a portion surrounded by □).

FIG. 4 is a diagram showing an electron micrograph of results obtained by using the reducing agent-containing hair treatment agent of Example 8 in Test Example 2.
(B) is an enlarged view of a part of (A) (a portion surrounded by □).

FIG. 5 is a view showing an electron micrograph showing the results of using the reducing agent-containing hair treatment agent of Example 9 in Test Example 2.
(B) is an enlarged view of a part of (A) (a portion surrounded by □).

FIG. 6 is a diagram showing an electron micrograph showing the results of using the reducing agent-containing hair treatment agent of Example 10 in Test Example 2.
(B) is an enlarged view of a part of (A) (a portion surrounded by □).

Claims (4)

[Claims] 1. A hair treatment agent comprising a carrier for binding a protease. 2. The hair treatment agent according to claim 1, wherein the proteolytic enzyme is a thiol protease. 3. A hair treatment agent comprising a thiol protease or a thiol protease binding carrier, and a reducing agent. 4. The hair treatment agent according to claim 2, wherein the thiol protease is papain.