JP5391649B2 - Process for producing cystine-keratin hydrolyzate complex - Google Patents

Description

  The present invention relates to adding a new property to a keratin hydrolyzate. More specifically, the present invention relates to a cystine-keratin hydrolyzate that adds a cystine that is a sulfur-containing amino acid to the keratin hydrolyzate and forms a precipitate in a pH-dependent manner. The present invention relates to a cystine-keratin hydrolyzate complex that is a complex and can be used in place of a material that has been conventionally produced using a keratin hydrolyzate as a raw material.

  Keratin is widely recognized by consumers in the health foods and supplements market because it is expected to have a protective effect, an improvement effect, and a growth effect on parts such as hair and nails. Examples of raw materials include wool, feathers, horns and scales. The keratin hydrolyzate is a product obtained by hydrolyzing keratin prepared from wool, feathers, etc. with an enzyme or acid / alkali and purifying it. The keratin hydrolyzate often has a molecular weight of about several thousand, and is an aggregate of peptides having a size of about 300 to 5,000. The final product is provided in the form of dried powders, granules and solutions.

  Keratin, which is the source of keratin hydrolyzate, is a fibrous protein that is contained in the most abundant amounts in the keratinous tissues of multicellular animals including humans. It is a major component of keratinous tissue such as hard hair and nails, and hair and nails are composed of keratin with slightly different amino acid composition called hard keratin and soft skin stratum corneum called soft keratin.

  The functionality of keratin is broadly divided into physical function and physiological function. Examples of physical functions include oxidation / reduction functions and non-biodegradability. Disulfide (-SS-) bond is formed by strong reducibility of cysteine thiol group (-SH) which is sulfur-containing amino acid in keratin, and excellent mechanical properties such as elasticity and toughness of wool and hair are expressed. . Moreover, it is known that wool is a hardly biodegradable fiber compared with cotton, hemp, and silk, and the degradation rate in soil is considerably slow (Non-patent Document 1). The physiological characteristics include a function as a nutrient and a function as a physiological activity / biological regulation function. Nutritional characteristics are composed of about 18 kinds of amino acids as amino acid composition, and are characterized by being rich in 14-18% of Cys (cysteine / cystine) which is hardly contained in other proteins such as silk and fiber. . Further, the ratio of basic amino acids histidine, lysine, and arginine is 1: 3: 10 (Non-patent Document 2). As a physiological activity, it has an anti-oxidative action, an anti-cancer promotion action, and an anti-carcinogenic action (Patent Document 1).

  Cystine is a kind of sulfur-containing amino acid and is known as a main component of protein keratin constituting hair. In vivo, methionine, which is an essential amino acid, is converted into cysteine through five steps, and an amino acid having a structure in which two molecules of cysteine are linked via an SS bond (disulfide bond) generated by oxidation of a thiol group (-SH). It becomes. Disulfide bonds between cysteines in polypeptides are necessary to maintain the secondary structure of proteins, and in hair, disulfide bonds between cysteines in keratin determine the degree of curly hair. Furthermore, hard keratin present in hair is also present in nails, and its amino acid composition is similar to that of hair and contains a large amount of cystine. In hair, it is known that disulfide bonds are formed between cysteine residues in keratin fibers to form cystine, which contributes to improvement in the elasticity and toughness of hair. In addition, when it is metabolized in the body, it generates sulfur and it works to detoxify by reacting with other substances, and it can protect the body from harmful effects such as active oxygen and radiation caused by toxic metals, smoking, drinking, etc. It is said.

  Thus, in the field of hair cosmetics, a cystine-introducing peptide to which the characteristics of cystine have been added has been developed (Patent Document 2), and a hair protecting agent for preventing damaged hair and recovering damaged hair using this technique. (Patent Document 3) and the first agent for permanent wave that can prevent hair damage during permanent wave, can impart excellent wave to hair, and can give gloss and moisture to treated hair (patent Document 4) is provided. In the field of tissue engineering and biotechnology, collagen derivatives that can be cross-linked via disulfide bonds are modified by adding thiol groups for the purpose of application to biomaterials such as implants. 5) has been developed. On the other hand, an organic compound having a thiol group (Patent Document 6) has been developed and provided as a suture, a biological substitute, and an adhesive. As described above, progress has been made in the development of materials having the characteristics of cystine in the fields of hair cosmetics, tissue engineering, and biotechnology, but there is hardly seen in other fields such as food and drink. As mentioned above, cystine is a major component of keratin, and is synthesized from methionine, which is an essential amino acid in vivo. The amount of hair also decreases and hair and nails become brittle. Considering that cystine is consumed with the growth of hair and nails, it is considered effective to take it as a food or drink in order to improve its quality. In addition, by taking cystine in combination with keratin, it is possible to prevent loss of nutritional balance due to single excessive intake of amino acids and to improve absorption efficiency. Thus, materials with the properties of cystine are useful, but their development has not progressed, and there is a need for materials with added properties such as detoxification, hair protection, improvement, and growth. Yes.

  In addition, as a material for pharmaceuticals, quasi-drugs, and foods and drinks using the characteristics of keratin, for example, beauty and health excellent in antioxidant action, anti-cancer promotion action and anti-carcinogenic action focusing on the physiological activity of keratin Bioactive composition useful for maintenance (patent document 1), food and drink for beautiful nails (patent document 7), and nutritional supplement for hair growth / hair growth (patent document 8) are provided. Many are blends of keratin hydrolyzate and other protein degradation products, plant extracts, etc., and the method of combining keratin hydrolyzate with other functional materials to achieve some synergistic effect is the field of food and drink In not seen. In the field of cosmetics and textiles, materials that combine cystine and keratin have been developed, and its applicability to cosmetics has been shown (Patent Document 2), but high adsorption to hair and skin due to disulfide bonds of cystine. Keratin with added properties is produced and provided as a cystine-introducing peptide having an effect of imparting luster and moisture to hair. However, the use in the field of food and drink is not considered, and it is manufactured through complex chemical synthesis for use in cosmetics. Therefore, a material having both characteristics of cystine and keratin that can be used in the field of food and drink, and a technique for producing it by a simple method are awaited.

JP-A-11-139985 Japanese Patent Laid-Open No. 10-287797 Japanese Patent Laid-Open No. 11-269045 Japanese Patent Laid-Open No. 11-269047 Japanese Patent Laid-Open No. 6-80935 JP-T 9-503490 JP 2001-321125 A JP 2007-236383 A Textile and Industry Vol. 62, no. 11 (2006) "Function of keratin fibers-Role of disulfide bonds-" Ajico News Food and Health Information Magazine Series “Amino Acids” 13 “Hair and amino acids” issued by Ajinomoto Co., Inc.

An object of the present invention, cystine having both the effect of the action and keratin cystine - is to provide a manufacturing how keratin hydrolyzate complex.

  As a result of intensive research in view of the above circumstances, the present inventors have found that an ingestible cystine-keratin complex can be produced without complicated chemical synthesis, and the protection and improvement of hair and nails The present invention has been completed by providing it as a material having both functions.

That is, the gist of the present invention is as follows.
(1) A method for producing a cystine-keratin hydrolyzate complex comprising a keratin hydrolyzate having a molecular weight of 300 to 5000 and cystine,
Dissolving cystine in an alkaline solution having a pH of 10 or higher;
Method for producing a keratin hydrolyzate complex, - the cystine solution and keratin hydrolyzate solution and mixing and stirring Resid such a step to form a complex in a solution of less than pH10 by cystine
(2) Production of cystine-keratin hydrolyzate complex according to (1), wherein the cystine-keratin hydrolyzate complex contains up to twice the amount of cystine relative to a keratin hydrolyzate having a molecular weight of 300 to 5000 Method,
About.

  By adding and ingesting the cystine-keratin hydrolyzate complex of the present invention to food and drink, etc., the whitening effect by the appearance prevention effect of melanin possessed by cystine and the enhancing action of hair and nails possessed by keratin hydrolyzate Synergistic effects can be expected in both cases, and as a result, damage to hair, nails and skin can be prevented, damaged hair, nails and skin can be recovered, and gloss and smoothness can be imparted more than ever. The effect can be expected.

  The cystine-keratin hydrolyzate complex of the present invention is characterized by containing a keratin hydrolyzate and cystine. By having such characteristics, a complex having both the action of cystine and the action of keratin can be obtained.

Examples of the raw material of the keratin hydrolyzate used in the present invention include animal hair, feathers, nails, horns, and hooves, and it is particularly preferable to use proteins prepared from wool, feathers, and the like. As a method for producing the keratin hydrolyzate, the keratin raw material can be produced by alkaline decomposition, oxidative decomposition with an inorganic acid / organic acid, reductive decomposition with a reducing agent, enzymatic decomposition with an enzyme, or decomposition by a bioreactor method. The conditions for alkaline decomposition, oxidative decomposition, reductive decomposition, enzymatic decomposition, and bioreactor decomposition are not particularly limited, and may be known conditions.
The keratin hydrolyzate may have a molecular weight of about 300 to 5000.

  Cystine used in the present invention has a structure in which two cysteine molecules, which are sulfur-containing amino acids, are connected via an S—S bond (disulfide bond), and a naturally occurring L form is used. Although it is possible to produce a complex using a mixture of D-forms or D-form alone, the present invention is not limited to this.

As a method for producing the cystine-keratin hydrolyzate complex of the present invention,
A step of dissolving cystine in an alkaline solution having a pH of 10 or more (first step);
A step of forming a complex in a solution having a pH of less than 10 by mixing and stirring the cystine solution and the keratin hydrolyzate solution (second step)
It is characterized by including.

In the first step, when cystine is dissolved in an alkaline solution having a pH of 10 or more, cystine and an alkaline solution having a pH of 10 or more are mixed while adjusting the pH to 10 or more, or cystine is mixed in a strong alkaline aqueous solution, Various methods such as adding a pH adjusting agent to adjust the pH to around 10 can be performed.
The alkali used in the present invention can be used as an aqueous solution having a pH of 10 or more. For example, potassium carbonate, potassium hydroxide, sodium carbonate, sodium hydroxide can be used, but from the viewpoint of manufacturing foods and drinks and cosmetics. It is preferable to use potassium carbonate / sodium carbonate. The solvent may be water. The amount of alkali in the alkaline solution is not particularly limited as long as the pH is 10 or more. In addition, if it is less than pH10, it cannot fully melt | dissolve and it is not preferable to use for this invention.
When mixing the cystine and the alkaline solution, the alkaline solution is preferably heated, so that it can be dissolved quickly.

Next, in the second step, the cystine solution obtained in the first step and the keratin hydrolyzate solution are mixed and stirred.
The keratin hydrolyzate solution may be an aqueous solution of keratin hydrolyzate. It is prepared by mixing keratin hydrolyzate in water or warm water. The content of cystine in the resulting cystine solution is preferably 2% by weight or more. The pH of the cystine solution is 10 or higher.

  The cystine solution and the keratin hydrolyzate solution may be mixed and stirred at room temperature. A complex is formed by adjusting the pH of the mixed solution to 10 or less. Formation of the complex can be confirmed by the cloudiness of the mixed solution. In the present invention, the cystine solution and the keratin hydrolyzate solution may be mixed at a time, but it is preferable to mix any of them little by little. Moreover, the stirring speed should just be suitable.

The mixed solution in which white turbidity has occurred can be promoted to form a complex by allowing to stand at room temperature.
The liquid mixture that has been allowed to stand may be added with a pH adjuster to reduce the pH to around 7. The standing time is not particularly limited, but is preferably 30 to 60 minutes.

The cystine-keratin hydrolyzate complex produced in the mixed solution in a suspended state in which white turbidity is produced can be obtained by separating it from the solvent by known means such as centrifugation and filtration.
The resulting cystine-keratin hydrolyzate complex can be washed with a solvent such as water, a buffer solution, or the like, and further dried to obtain a solid.

  The cystine-keratin hydrolyzate complex obtained as described above can be obtained by removing reagents used in the synthesis process without going through the conventional complicated chemical synthesis as in, for example, Patent Document 2 above. Since it does not need to be purified, it can be easily added to foods, cosmetics, and other products that require attention to safety, and it can be easily used in forms other than solutions by collecting it as a precipitate. It is a cystine-keratin hydrolyzate complex.

  Therefore, the present invention is obtained by dissolving cystine in an alkaline solution having a pH of 10 or more, mixing the obtained cystine solution and a keratin hydrolyzate solution, and adjusting the pH to less than 10. It relates to a keratin hydrolyzate complex.

  In the cystine-keratin hydrolyzate complex of the present invention, it is obtained by the production method as described above, and cystine and keratin or a hydrolyzate form a complex in the examples described later. It can confirm from the result of description. Although the structure of the complex is not clear, the cystine-keratin hydrolyzate of the present invention does not form a strong bond such as a covalent bond between the keratin hydrolyzate and cystine. It is easy to think that the properties of keratin and cystine are easily played together.

  Moreover, the cystine-keratin hydrolyzate complex of the present invention can be added to a food or drink or cosmetic as a raw material to prepare a food or drink or cosmetic that has both a cystine-specific action and a keratin-specific action.

  Next, the present invention will be described more specifically with reference to examples. However, this invention is not limited only to those Examples. In the following examples and the like, “%” indicating the concentration of a solution or the like is “% by weight” unless otherwise indicated.

Example 1
The cystine-keratin hydrolyzate complex was prepared by dissolving L-cystine in a potassium carbonate solution having a pH of 10 or higher at 90 ° C. to prepare a 2% cystine solution. This was returned to room temperature (25 ° C.), and a 50% aqueous solution of keratin hydrolyzate was gradually added thereto and stirred. As a keratin hydrolyzate solution to be used, a 33% aqueous solution of a keratin hydrolyzate having a molecular weight of 300 to 5000 (manufactured by Ichimaru Falcos) was prepared and added to the 2% cystine solution. At room temperature (25 ° C), 10 g of 2% cystine solution and 30 g of 33% solution of the above keratin hydrolyzate (manufactured by Ichimaru Falcos) were gradually mixed and stirred, and when the pH was lowered to around 7, white turbidity was produced. It was. Subsequently, the reaction was continued for 60 minutes at room temperature. After the reaction was completed, 7.5 g of 1M phosphate buffer was added to adjust the pH to 7. The reaction mixture was centrifuged at 49000 xg (20000 rpm) to remove the supernatant, the remaining supernatant and precipitate were washed with 0.1 M phosphate buffer, and the suspension was centrifuged at 49000 xg (20000 rpm). The supernatant was removed and dried to obtain a precipitate of cystine-keratin hydrolyzate complex.

Comparative Example As a control for Example 1, at room temperature (25 ° C.), 5 g of 2% cystine-2% potassium carbonate solution was mixed and stirred with 2.0 g of 1M sodium dihydrogen phosphate solution to lower the pH. As a result, white turbidity occurred. Subsequently, the mixture was allowed to react for 30 minutes at room temperature. After the reaction was completed, 5.5 g of 1M phosphate buffer was added to adjust the pH to 7, and 30 g of a 33% solution of keratin hydrolyzate having a molecular weight of 300 to 5000 was obtained. Were gradually mixed and stirred. The reaction mixture was centrifuged at 49000 xg (20000 rpm) to remove the supernatant, the remaining supernatant and precipitate were washed with 0.1 M phosphate buffer, and the suspension was centrifuged at 49000 xg (20000 rpm). The supernatant was removed and dried to obtain a cystine precipitate.

Test example 1
A portion of the precipitate obtained in Example 1 was placed in a Durham tube, degassed and acid hydrolyzed at 150 ° C. for 1 hour, and then amino acid composition analysis was performed using an amino acid analyzer. The total amount of radix was detected as 33.7% as half cystine. Furthermore, aspartic acid, glutamic acid, serine, glycine, and proline, which are amino acids that characterize keratin SH powder (manufactured by Ichimaru Falcos) used as a keratin hydrolyzate in the present invention, are 12%, 17%, 19%, and 13%, respectively. -11% was detected. A part of the precipitate obtained in the comparative example as a control was taken in a Durham tube, deaerated and acid-hydrolyzed at 150 ° C. for 1 hour, and then amino acid composition analysis was performed with an amino acid analyzer.・ Aspartic acid, glycine, and proline were hardly detected.

  The amino acid composition of the peptide in the precipitate by reaction with cystine, excluding free amino acids, calculated from the results obtained in Example 1 and the comparative example, is the amino acid composition of human hair keratin / wool keratin (Non-patent Document) Along with 2), the amino acid content is shown in FIG. The ratio of amino acids such as cystine and methionine in the raw material keratin hydrolyzate is not shown because it hardly contains.

  As apparent from FIG. 1, in the precipitate obtained in the cystine-keratin hydrolyzate reaction experiment, amino acids such as aspartic acid, glutamic acid, serine, glycine, and proline derived from the keratin hydrolyzate were detected in addition to cystine. Similarity was confirmed when compared with the amino acid composition of human keratin and wool keratin. As described above, it was revealed that the keratin hydrolyzate detected in the precipitate formed by the reaction accompanied by the change in pH forms a complex with cystine.

Test example 2
Collagen containing cystine-keratin hydrolyzate complex for 10 monitors for the functionality (effect on skin, hair, nails) of cystine-keratin hydrolyzate complex obtained in Example 1 A peptide drink (containing 30 mg equivalent of the cystine-keratin hydrolyzate complex) was distributed, and one tasting per day was continued for 2 weeks. A five-step evaluation was made to answer questions (improvement items) regarding whether or not the effect on skin, hair, and nails was felt at the 3rd, 7th, 10th, and 14th days of 2 weeks. The efficacy was quantified. The numerical value indicates that 5 is the maximum and the effect is the highest, and 1 is the minimum and the effect is the lowest. The results are shown in FIGS. The control in FIGS. 2 to 4 means an experiment in which a beverage containing a mixture of cystine and keratin is ingested. The placebo in FIGS. 2 to 4 means an experiment in which a beverage containing only a collagen peptide is ingested. The evaluation in the figure is an average value. From the results of FIGS. 2 to 4, the cystine-keratin hydrolyzate complex according to the present invention has a whitening action which is a function peculiar to cystine and a protective and improving action for hair and nails which is a peculiar function to keratin. It is found that both can be realized and that the effect is realized more than when a mixture of cystine and keratin hydrolyzate is ingested.
The mixture of cystine and keratin used in the control was prepared by mixing cystine with keratin without dissolving it in an alkaline solution.

Example 2
Each component was mixed according to the following formulation to prepare a beverage.

Example 3
Each component was mixed according to the following formulation to prepare a cosmetic (cream).

Example 4
Each component was mixed according to the following formulation to prepare a cosmetic (hair restoration / hair growth cream).

  In addition, when the criticality of the cystine content in the cystine-keratin hydrolyzate complex was tested, a complex obtained by adding and mixing 10 g of an excessive amount of keratin hydrolyzate (molecular weight of about 300 to 5000) to 200 mg of cystine. When calculated based on the results of amino acid composition analysis of the body, cystine and keratin hydrolyzate form a complex at a ratio of cystine: keratin hydrolyzate = 2: 1 (weight ratio). Thus, in the production method of the present invention, although the amount of cystine and keratin hydrolyzate contained in the cystine-keratin hydrolyzate complex can be changed depending on the mixing ratio of cystine and keratin hydrolyzate, a certain molecular weight It is believed that at most twice the amount of cystine is adsorbed to the distribution of keratin hydrolysates.

  Since the cystine-keratin hydrolyzate complex of the present invention is excellent in functionality provided with the functions of cystine and keratin hydrolyzate as described above, skin rust prevention, water retention, enhancement of hair and nails, It can be suitably used as an active ingredient in foods and drinks for the purpose of inhibiting production / extraction of melanin causing freckles and freckles, antioxidant action, inactivation of active oxygen, and improvement of liver function.

FIG. 1 shows the amino acid composition in the keratin hydrolyzate precipitated against cystine obtained in Example 1 and the comparative example, together with the amino acid composition of human hair keratin and wool keratin, as the content of amino acid residues. It is a graph. FIG. 2 is a graph showing the results of evaluating the effects of the cystine-keratin hydrolyzate complex obtained in Example 1 on the hair of the control and placebo. FIG. 3 is a graph showing the results of evaluating the effect of the cystine-keratin hydrolyzate complex obtained in Example 1 on the nails of the control and placebo. FIG. 4 is a graph showing the results of evaluating the skin effect of the cystine-keratin hydrolyzate complex obtained in Example 1 and the control and placebo.

Claims (2)

A method for producing a cystine-keratin hydrolyzate complex comprising a keratin hydrolyzate having a molecular weight of 300 to 5000 and cystine,
Dissolving cystine in an alkaline solution having a pH of 10 or higher;
The cystine solution and keratin hydrolyzate solution and mixing and stirring Resid such a step to form a complex in a solution of less than pH10 by cystine - method of manufacturing a keratin hydrolyzate complex. The method for producing a cystine-keratin hydrolyzate complex according to claim 1, wherein the cystine-keratin hydrolyzate complex contains up to twice the amount of cystine relative to a keratin hydrolyzate having a molecular weight of 300 to 5,000 .

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