JP6544503B2 - Cosmetic composition and functional food containing collagenase inhibitor and collagenase inhibitor - Google Patents
Cosmetic composition and functional food containing collagenase inhibitor and collagenase inhibitor Download PDFInfo
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Description
本発明は、コラゲナーゼの活性阻害作用を有する新規なコラゲナーゼ阻害剤及びその応用品に関する。 The present invention relates to a novel collagenase inhibitor having a collagenase activity inhibitory action and its application.
皮膚の水分保持性、柔軟性又は弾力性は、真皮に含まれるコラーゲン等の細胞外マトリックス成分に起因しており、コラーゲンの減少又は分解は、皮膚老化の原因である。
真皮線維芽細胞より産生されるコラゲナーゼはコラーゲンの分解する酵素であり、コラゲナーゼの活性を抑制し、皮膚の弾力や張りを維持するコラーゲンの変性を防止することが皮膚老化の防止に効果的である。
また、皮膚組織など美容系のみならず，癌転移においてもコラゲナーゼが活性化することが知られており、コラゲナーゼ阻害による癌転移抑制効果が期待されている。
The moisture retention, flexibility or elasticity of the skin is attributed to extracellular matrix components such as collagen contained in the dermis, and the reduction or degradation of collagen is the cause of skin aging.
Collagenase produced from dermal fibroblasts is an enzyme that degrades collagen, and it is effective in preventing skin aging by suppressing collagenase activity and preventing collagen degeneration that maintains skin elasticity and firmness. .
Moreover, it is known that collagenase is activated not only in cosmetic systems such as skin tissue but also in cancer metastasis, and a cancer metastasis inhibitory effect by collagenase inhibition is expected.
これまでにコラゲナーゼの活性阻害作用を有する成分として、様々な報告がなされており、例えば、アセロラ種子抽出物（特許文献１）、特許文献２ではハマメリス抽出物（特許文献２）、ジャガイモ抽出物（特許文献３）、ウェルウィッチア科植物の抽出物（特許文献４）、レッドクローバー（特許文献５）などが挙げられる。 Until now, various reports have been made as components having a collagenase activity inhibitory action, for example, acerola seed extract (patent document 1), patent document 2 with hamamelis extract (patent document 2), potato extract (patent document 2) Patent Document 3), an extract of a Welwichaceae plant (Patent Document 4), a red clover (Patent Document 5), and the like.
しかしながら、麹菌発酵物を利用したコラゲナーゼ阻害剤についての研究はほとんど進んでいなかった。
かかる状況下、本発明の目的は、従来知られていなかった、新規のコラゲナーゼ阻害剤、及びその応用品を提供することである。
However, little research has been conducted on collagenase inhibitors using gonococci fermentation products.
Under such circumstances, an object of the present invention is to provide a novel collagenase inhibitor which has not hitherto been known, and an application thereof.
本発明者は、上記課題を解決すべく、皮膚外用剤、化粧料、飲食品などの分野に幅広く応用が可能なコラゲナーゼ阻害作用を有する物質を得るために、天然由来の種々の物質について検討を行った。その結果、焼酎の製造に使用される白麹菌(Aspergillus kawachii)や黒麹菌（Aspergillus awamori）を繁殖させた米麹の溶媒抽出物が、優れたコラゲナーゼ阻害作用を有することを見出し、本発明に至った。 In order to solve the above-mentioned problems, the present inventor has studied various substances of natural origin in order to obtain substances having collagenase inhibitory action which can be widely applied to the fields of external skin preparations, cosmetics, food and drink and the like. went. As a result, it has been found that a solvent extract of rice bran grown by Aspergillus niger (Aspergillus kawachii) or Aspergillus niger (Aspergillus awamori), which is used for producing shochu, has an excellent collagenase inhibitory action, leading to the present invention. The
すなわち、本発明は、以下の発明に係るものである。
＜１＞ 白麹菌(Aspergillus kawachii)及び黒麹菌（Aspergillus awamori）から選択される少なくとも１種を繁殖させた米麹の溶媒抽出物を含有するコラゲナーゼ阻害剤。
＜２＞ 前記米麹が、白麹菌(Aspergillus kawachii)を繁殖させた米麹である前記＜１＞に記載のコラゲナーゼ活性阻害剤。
＜３＞ 前記溶媒抽出物が、前記米麹と、水、エタノール及び１，３−ブチレングリコールから選択される１種以上を主溶媒として含有する抽出溶媒とを接触させ、前記米麹に含有される成分を抽出して得られる抽出物である前記＜１＞または＜２＞に記載のコラゲナーゼ阻害剤。
＜４＞ 前記抽出溶媒が、水及び１，３−ブチレングリコールから選択される１種以上である前記＜３＞に記載のコラゲナーゼ阻害剤。
＜５＞ 前記＜１＞から＜４＞のいずれかに記載のコラゲナーゼ阻害剤を配合してなることを特徴とする化粧料組成物。
＜６＞ 前記＜１＞から＜４＞のいずれかに記載のコラゲナーゼ阻害剤を配合してなることを特徴とする機能性食品。
That is, the present invention relates to the following inventions.
<1> A collagenase inhibitor containing a solvent extract of rice bran on which at least one selected from Aspergillus kawachii and Aspergillus awamori is propagated.
<2> The collagenase activity inhibitor according to <1>, wherein the rice bran is a rice bran obtained by breeding Aspergillus kawachii.
<3> The solvent extract is brought into contact with the rice bran and an extraction solvent containing at least one selected from water, ethanol and 1,3-butylene glycol as a main solvent, and the solvent extract is contained in the rice bran The collagenase inhibitor according to <1> or <2>, which is an extract obtained by extracting a component.
<4> The collagenase inhibitor according to <3>, wherein the extraction solvent is one or more selected from water and 1,3-butylene glycol.
<5> A cosmetic composition comprising the collagenase inhibitor according to any one of <1> to <4>.
<6> A functional food comprising the collagenase inhibitor according to any one of <1> to <4>.
本発明によれば、優れたコラゲナーゼ阻害作用を有するコラゲナーゼ阻害剤、及び化粧料組成物及び機能性食品が提供される。 ADVANTAGE OF THE INVENTION According to this invention, the collagenase inhibitor which has the outstanding collagenase inhibitory effect, and a cosmetic composition and a functional food are provided.
以下、本発明について例示物等を示して詳細に説明するが、本発明は以下の例示物等に限定されるものではなく、本発明の要旨を逸脱しない範囲において任意に変更して実施できる。なお、本明細書において、「〜」という表現を用いた場合、その前後の数値または物理値を含む意味で用いることとする。 Hereinafter, the present invention will be described in detail with reference to examples and the like, but the present invention is not limited to the following examples and the like, and can be implemented with various modifications without departing from the scope of the present invention. In addition, in this specification, when the expression "-" is used, it shall be used by the meaning including the numerical value or physical value before and behind that.
＜コラゲナーゼ阻害剤＞
本発明は、白麹菌(Aspergillus kawachii) 及び黒麹菌（Aspergillus awamori）から選択される少なくとも１種を繁殖させた米麹の溶媒抽出物を含有するコラゲナーゼ阻害剤（以下、「本発明の阻害剤」と記載する場合がある。）である。
<Collagenase inhibitor>
The present invention relates to a collagenase inhibitor (hereinafter referred to as "the inhibitor of the present invention") which contains a solvent extract of rice bran on which at least one selected from Aspergillus kawachii and Aspergillus awamori is propagated. And may be written as
本発明の阻害剤は、米に麹菌を繁殖させた米麹のうち、麹菌として白麹菌(Aspergillus kawachii)及び黒麹菌（Aspergillus awamori）から選択される少なくとも１種を繁殖させた米麹を使用していることに特徴があり、優れたコラゲナーゼ阻害作用を発現する。
以下、本発明の阻害剤について詳細に説明する。
The inhibitor according to the present invention is a rice bran obtained by breeding at least one selected from Aspergillus kawachii and Aspergillus awamori as bacilli among rice bran in which Bacilli were propagated in rice. And have excellent collagenase inhibitory action.
Hereinafter, the inhibitor of the present invention will be described in detail.
（米麹）
本発明の阻害剤に含有される米麹は、麹菌として白麹菌(Aspergillus kawachii)を繁殖させた米麹（以下、「米麹（白麹菌）」と記載する場合がある。）または、黒麹菌（Aspergillus awamori）を繁殖させた米麹（以下、「米麹（黒麹菌）」と記載する場合がある。）、また、白麹菌と黒麹菌の両方を繁殖させた米麹である。なお、以下、これらを総称して「本発明の米麹」と記載する場合がある。本発明の米麹のうち、色素の観点からより好ましくは米麹（白麹菌）である。
(Malted rice)
The rice bran contained in the inhibitor of the present invention is a rice bran (hereinafter sometimes referred to as “rice bran (white bacillus)”), which is obtained by propagating Aspergillus kawachii as bacilli (hereinafter referred to as “rice bran (white bacillus)”) or black bacillus. It is a rice bran (hereinafter sometimes referred to as "rice bran (E. coli")) which is produced by breeding (Aspergillus awamori), and also a rice bran having both B. anthracis and E. coli bred. Hereinafter, these may be collectively referred to as "rice bran of the present invention". Among the rice bran according to the present invention, more preferred is rice bran (white bacillus) from the viewpoint of pigment.
白麹菌(Aspergillus kawachii)は、黒麹菌（Aspergillus awamori）の突然変異株であり、焼酎の製造に使用されている麹菌である。
米麹（白麹菌）や米麹（黒麹菌）の溶媒抽出物が優れたコラゲナーゼ阻害作用を発現することについての作用機序の詳細は現段階では詳細は完全に明らかではないが、米麹（白麹菌）や米麹（黒麹菌）の抽出物を加熱して、含まれる酵素を失活させても、あるいはろ過によって抽出物から白麹菌を除去してもコラゲナーゼ阻害活性が認められるため、麹の白麹菌や黒麹菌による低分子産生物が実質的な有効成分である可能性がある。
Aspergillus kawachii is a mutant of Aspergillus awamori and is a bacillus used in the manufacture of shochu.
Although the details of the mechanism of action of the solvent extract of rice bran (Blinter Blight) and rice bran (Bletilla bacillus) express excellent collagenase inhibitory action, the details are not completely clear at this stage. The collagenase inhibitory activity is recognized even if the extract contained in E. coli) or rice bran (E. bacillus) is heated to inactivate the enzymes contained therein, or even if E. coli is removed from the extract by filtration. And low molecular weight products produced by Aspergillus oryzae may be a substantial active ingredient.
一方、白麹菌および黒麹菌以外の麹菌を繁殖させた米麹の抽出物、例えば、清酒製造に使用される黄麹菌（Aspergillus oryzae）を繁殖させた米麹の水抽出物では、コラゲナーゼ阻害作用が認められない、あるいはコラゲナーゼ阻害作用が米麹（白麹菌）の抽出物や米麹（黒麹菌）の場合と比較して極めて小さい。 On the other hand, the extract of rice bran on which Aspergillus oryzae other than B. anthracis and B. anthracis is propagated, for example, the aqueous extract of rice bran on which Aspergillus oryzae used for producing sake is effective in inhibiting collagenase It is not recognized or collagenase inhibitory action is extremely small compared with the case of the extract of rice bran (White mold fungus) or rice bran (black mold fungus).
米麹の原料である米類としては、玄米、ウルチ米、精白米等が挙げられる。これらは必要に応じて、適当な大きさに粉砕して使用してもよい。
米類は、固めに炊く、あるいは蒸したのちに、白麹菌や黒麹菌を繁殖させて本発明に係る米麹とする。
好適な一例を挙げると、精白米を水で精米した後、冷水に浸漬する。その後、余計な水切りを行い、精白米を蒸しあげる。蒸しあがった精白米の水分を飛ばし、麹菌（白麹菌、黒麹菌）を混ぜ込んでいく。次に麹菌を混ぜた精白米を３５〜４０℃で１８時間保温し、その後、麹菌が付着した精白米を均一に広げ３４℃で６時間保温する。これを２回繰り返し、麹を製造する。
Examples of rice which is a raw material of rice bran include brown rice, uruchi rice, polished rice and the like. These may be used after being crushed to an appropriate size, if necessary.
After rice is cooked or steamed, white bacilli and black bacillus are propagated to make a rice bran according to the present invention.
As a suitable example, after milling polished rice with water, it is immersed in cold water. After that, extra draining and steaming the polished rice. Remove the moisture from the steamed polished rice and mix in the koji molds (white mold and black mold bacteria). Next, the milled rice mixed with Aspergillus oryzae is kept at 35 to 40 ° C. for 18 hours, and then the milled rice to which the fungus is attached is spread uniformly and kept for 6 hours at 34 ° C. Repeat this twice to make a cocoon.
本発明の阻害剤において、本発明の米麹の溶媒抽出物（以下、単に「抽出物」と記載する場合がある。）とは、抽出対象となる本発明の米麹を、溶媒に接触させることで、米麹中の酵素による米麹成分の分解ならびに抽出などして、有効成分の含有量を高めた形態のものを総括した概念である。本発明の阻害剤における抽出物は、具体的には、本発明の米麹を抽出原料として得られる抽出液、該抽出液の希釈液若しくは濃縮液、又はこれらの粗精製物若しくは精製物のいずれもが含まれる。また、液状の抽出物を乾燥して得られる乾燥物も、本発明における抽出物に該当するものとする。 In the inhibitor of the present invention, the solvent extract of the rice bran of the present invention (hereinafter sometimes referred to simply as "extract") brings the rice bran of the present invention to be extracted into contact with a solvent It is the concept which summarized the thing of the form which raised content of the active ingredient by the decomposition | disassembly of the rice bran component by the enzyme in rice bran, extraction, etc. Specifically, the extract of the inhibitor of the present invention is any of an extract obtained using the rice bran of the present invention as an extraction raw material, a diluted solution or concentrate of the extract, or any of these crude products or purified products. Also included. In addition, the dried product obtained by drying the liquid extract also corresponds to the extract in the present invention.
抽出物は、乾燥物を除くいずれの場合においても、必要に応じて、不溶分を除去するために、透析、限外ろ過、ろ過、カラムクロマトグラフィー等による前処理を行ってもよい。ろ過による除去の場合には、必要に応じて、不純物を除去するために活性炭、ベントナイト、セライト等の吸着剤やろ過助剤を添加してもよい。特に抽出液の状態で用いる場合には、メンブレンフィルター等による除菌ろ過を併せて行うことが好ましい。 The extract may be pretreated by dialysis, ultrafiltration, filtration, column chromatography or the like to remove insolubles, if necessary, in any case except for the dry matter. In the case of removal by filtration, if necessary, an adsorbent such as activated carbon, bentonite, celite, or a filter aid may be added to remove impurities. In particular, when used in the state of an extract, it is preferable to carry out sterilization filtration with a membrane filter etc. in combination.
抽出物は、必要に応じて任意の公知の方法で精製してもよく、必要に応じて、医薬および食品分野において許容される塩、誘導体または溶媒和物等に変換してもよい。 The extract may be optionally purified by any known method, and may be converted into a salt, a derivative, a solvate, etc. acceptable in the pharmaceutical and food field as necessary.
抽出物を溶媒抽出で得る場合、抽出溶媒としては、本発明の米麹に含有されるコラゲナーゼ阻害作用を発現する有効成分を抽出できるものであればよい。具体例としては、水、メタノール、エタノール、プロピルアルコール、イソプロピルアルコール等の炭素数１〜５の低級アルコール；アセトン、メチルエチルケトン等の低級脂肪族ケトン；１，３−ブチレングリコール、プロピレングリコール、グリセリン等の炭素数２〜５の多価アルコールなどが挙げられる。なお、抽出溶媒として使用し得る水には、精製水、イオン交換水、生理食塩水、リン酸緩衝液、リン酸緩衝生理食塩水等も含まれる。
これらの抽出溶媒は単独又はこれら２種以上の混合物として使用することができ、２種以上の溶媒の混合液を抽出溶媒として使用する場合、その混合比は適宜調整することができる。
When the extract is obtained by solvent extraction, any extraction solvent may be used as long as it can extract the active ingredient that exhibits the collagenase inhibitory action contained in the rice bran of the present invention. Specific examples thereof include lower alcohols having 1 to 5 carbon atoms such as water, methanol, ethanol, propyl alcohol and isopropyl alcohol; lower aliphatic ketones such as acetone and methyl ethyl ketone; and 1,3-butylene glycol, propylene glycol, glycerin and the like A C2-C5 polyhydric alcohol etc. are mentioned. Water which can be used as an extraction solvent also includes purified water, ion-exchanged water, physiological saline, phosphate buffer, phosphate buffered saline and the like.
These extraction solvents can be used alone or as a mixture of two or more of these, and when a mixture of two or more solvents is used as an extraction solvent, the mixing ratio can be appropriately adjusted.
なお、抽出溶媒には、本発明の効果を損なわない範囲で添加剤を含んでいてもよい。例えば、ｐＨ調整剤、粘度調整剤などが挙げられる。 In addition, the extraction solvent may contain an additive as long as the effect of the present invention is not impaired. For example, pH adjusters, viscosity adjusters, etc. may be mentioned.
本発明に係る抽出物において、本発明の米麹と、抽出溶媒とを接触させ、前記米麹に含有される成分を抽出して得られる抽出物であることが好ましい。
抽出溶媒としては、水、エタノール及び１，３−ブチレングリコールから選択される１種以上を主溶媒として含有する抽出溶媒であることが好ましい。ここで、「主溶媒」とは溶媒全量の５０体積％以上（１００体積％含む）を占める溶媒を意味する。
The extract according to the present invention is preferably an extract obtained by bringing the rice bran of the present invention into contact with an extraction solvent and extracting the components contained in the rice bran.
The extraction solvent is preferably an extraction solvent containing one or more selected from water, ethanol and 1,3-butylene glycol as a main solvent. Here, the "main solvent" means a solvent that occupies 50% by volume or more (including 100% by volume) of the total amount of the solvent.
抽出溶媒としては、特に水及び１，３−ブチレングリコールから選択される１種以上であることが好ましい。水と１，３−ブチレングリコールとの割合は任意であるが、好適には、１，３−ブチレングリコール基準濃度で、０重量％〜５０重量％である。
このような抽出溶媒であれば、得られる抽出物は、他の溶媒で抽出した場合と比較して、より優れたコラゲナーゼに対する活性阻害作用を有する。
The extraction solvent is preferably at least one selected from water and 1,3-butylene glycol. Although the ratio of water to 1,3-butylene glycol is optional, it is preferably 0% by weight to 50% by weight based on 1,3-butylene glycol.
If it is such an extraction solvent, the extract obtained has the activity inhibitory effect with respect to a collagenase more excellent compared with the case where it extracts with another solvent.
本発明の米麹に対する抽出溶媒の量は特に制限はないが、通常、米麹（白麹菌）に対して重量比で３〜１００倍量程度である。抽出溶媒が水の場合には、米麹（白麹菌）に対して重量比で５〜２０倍量であることが好ましく、１，３−ブチレングリコールである場合には５〜２０倍量であることが好ましい。米麹（黒麹菌）の場合も米麹（白麹菌）の場合と同様である。 The amount of the extraction solvent for rice bran according to the present invention is not particularly limited, but is usually about 3 to 100 times by weight the amount of rice bran (Blinter Bacilli). When the extraction solvent is water, the amount is preferably 5 to 20 times by weight ratio to rice bran (White bacillus), and if it is 1,3-butylene glycol it is 5 to 20 times Is preferred. The case of rice bran (black bacillus) is similar to the case of rice bran (white bacillus).
本発明の米麹から溶媒抽出物を抽出する方法は特に限定されず、常法に従って行なれる。例えば、米麹（白麹菌）と、抽出溶媒とを互いに充分に攪拌し混合後、米麹（白麹菌）から溶媒中に、有効成分が十分に抽出されるまで一定期間静置する。抽出時間は、抽出溶媒の種類や、米麹（白麹菌）と抽出溶媒との割合を考慮し適宜選択されるが、抽出溶媒が水の場合は１〜３時間であり、１，３−ブチレングリコールの場合は１〜３時間である。米麹（黒麹菌）の場合も米麹（白麹菌）の場合と同様である。
また、抽出物に含まれる残渣を取り除くため、濾過や遠心分離を行ってもよい。また、得られた抽出液はそのまま利用してもよいが、常法に従って希釈、濃縮、乾燥、精製等の処理を施してもよい。
なお、この抽出操作は品質劣化を避けるために常温で行うのが好ましいが、抽出効率を上げるために加温状態にして行うことも可能である。加温により米麹のデンプンやタンパク質などが分解され豊富な栄養成分（アミノ酸・糖など）を抽出液中に含有させることができる。また、必要に応じて抽出物の割合を高めるため、減圧濃縮や凍結乾燥により溶媒除去してもよい。
The method for extracting a solvent extract from rice bran according to the present invention is not particularly limited, and can be carried out according to a conventional method. For example, after the rice bran (white bacillus) and the extraction solvent are sufficiently stirred and mixed with each other, they are allowed to stand from the rice bran (white bacillus) in the solvent for a certain period until the active ingredient is sufficiently extracted. The extraction time is appropriately selected in consideration of the type of the extraction solvent and the ratio of rice bran (white mold) to the extraction solvent, but is 1 to 3 hours when the extraction solvent is water, and 1,3-butylene In the case of glycol, it is 1 to 3 hours. The case of rice bran (black bacillus) is similar to the case of rice bran (white bacillus).
Moreover, in order to remove the residue contained in the extract, filtration or centrifugation may be performed. The obtained extract may be used as it is, but may be subjected to treatments such as dilution, concentration, drying, purification and the like according to a conventional method.
Although it is preferable to carry out this extraction operation at normal temperature in order to avoid quality deterioration, it is also possible to carry out the heating operation in order to increase the extraction efficiency. By heating, starch, protein and the like of rice bran are decomposed and abundant nutrient components (amino acid, sugar and the like) can be contained in the extract. In addition, in order to increase the proportion of the extract as necessary, the solvent may be removed by concentration under reduced pressure or lyophilization.
本発明に係る抽出物として、米麹（白麹菌）や米麹（黒麹菌）を水の混合物をろ過したもの、米麹（白麹菌）や米麹（黒麹菌）と水の混合物を加温後濾過したものが好適な一例として挙げられる。 As an extract according to the present invention, one obtained by filtering a mixture of rice bran (white mold) or rice bran (black mold), and heating a mixture of rice bran (white mold) or rice bran (black mold) and water What was post-filtered is mentioned as a suitable example.
本発明に係る抽出物は、コラゲナーゼ阻害作用を有すると共に、人体に対する毒性や刺激性が少ない。そのため、各種外用剤、服用剤、化粧料組成物、機能性食品等の用途に使用することができる。 The extract according to the present invention has collagenase inhibitory action and is less toxic and irritating to the human body. Therefore, it can be used for applications such as various external preparations, medications, cosmetic compositions, functional foods and the like.
本発明の阻害剤として、米麹（白麹菌）や米麹（黒麹菌）の抽出物を、原液をそのまま用いても、濃縮して濃縮液として用いてもよく、原液あるいは濃縮液を希釈溶媒に溶解して使用してもよい。この希釈溶媒としては、コラゲナーゼ阻害作用を損なわないものが選択され、水、エチルアルコール、イソプロピルアルコール、グリセリン、プロピレングリコール、１，３−ブチレングリコール等を例示することができる。 As the inhibitor of the present invention, the extract of rice bran (White bacillus) or rice bran (black bacillus) may be used as a stock solution as it is or may be concentrated as a concentrate, and the stock solution or concentrate may be used as a dilution solvent It may be dissolved in As this dilution solvent, one which does not impair the collagenase inhibitory action is selected, and water, ethyl alcohol, isopropyl alcohol, glycerin, propylene glycol, 1,3-butylene glycol and the like can be exemplified.
本発明の阻害剤によるコラゲナーゼ阻害作用は、当該阻害剤が含有する米麹（白麹菌）や米麹（黒麹菌）の抽出物の量に依存し、コラゲナーゼ阻害率が３０％以上（好適には４０％以上）となる範囲で決定される。
本発明の阻害剤は、特にコラゲナーゼ阻害作用が高いため、比較的低濃度（例えば、２〜１０重量％）でもコラーゲンの分解抑制効果が期待できる。
The collagenase inhibitory action by the inhibitor of the present invention depends on the amount of the rice bran (white mold) and the rice bran (black mold) extract contained in the inhibitor, and the collagenase inhibitory rate is 30% or more (preferably) It is decided in the range which becomes 40% or more).
The inhibitor of the present invention is particularly high in collagenase inhibitory activity, and thus can suppress collagen degradation even at relatively low concentrations (for example, 2 to 10% by weight).
本発明のコラゲナーゼ阻害剤は、皮膚の老化の一因であるコラゲナーゼによるコラーゲンの分解を抑制することができるので、抗老化剤として使用できる可能性がある。ここで、「抗老化作用」とは、皮膚のしわ、たるみなどを有効に防止・改善などを含む概念である。 The collagenase inhibitor of the present invention can be used as an anti-aging agent because it can suppress the degradation of collagen by collagenase, which is a cause of skin aging. Here, the "anti-aging action" is a concept including effectively preventing or improving wrinkles, sagging and the like of the skin.
本発明のコラゲナーゼ阻害剤の形態としては特に限定はないが、一般に皮膚に塗布する形の皮膚外用剤として用いられる場合には、液状やクリーム状である。この場合、コラゲナーゼ阻害剤は、必要に応じて、通常医薬品、医薬部外品に配合される、油性成分、可溶化剤、保湿剤、色素、乳化剤、増粘剤、香料等の任意の成分を含有することができる。また、本発明の効果を損なわない範囲で、他の薬効植物抽出物を添加してもよい。 The form of the collagenase inhibitor of the present invention is not particularly limited, but generally it is in the form of liquid or cream when it is used as an external preparation for skin application on the skin. In this case, the collagenase inhibitor is an optional ingredient such as an oily component, a solubilizer, a moisturizer, a pigment, an emulsifier, a thickener, a flavor and the like which is usually added to pharmaceuticals and quasi drugs as needed. It can be contained. In addition, other medicinal plant extracts may be added as long as the effects of the present invention are not impaired.
また、本発明のコラゲナーゼ阻害剤は、入浴剤、ボディーソープ、シャンプー等の入浴用組成物に配合して用いてもよい。剤型としては、一般に用いられる、水溶液、Ｗ／Ｏ型又はＯ／Ｗ型エマルション、適当な賦形剤等を用いて顆粒剤その他の粉末、錠剤等としてもよい。また、本発明の効果を損なわない範囲で、他の薬効植物抽出物を添加してもよい。 In addition, the collagenase inhibitor of the present invention may be used by blending in a bathing composition such as a bath agent, a body soap, a shampoo and the like. As a dosage form, an aqueous solution, a W / O type or an O / W type emulsion generally used, granules and other powders, tablets and the like using appropriate excipients and the like may be used. In addition, other medicinal plant extracts may be added as long as the effects of the present invention are not impaired.
また、本発明のコラゲナーゼ阻害剤は経口用組成物とすることもできる。この場合、経口投与に利用される剤形としては、具体的には、固形製剤として、粉末剤、顆粒剤、錠剤、カプセル剤、トローチ等が挙げられる。また、液状製剤として内用液剤、外用液剤、懸濁剤、乳剤、シロップ剤等が例示され、これら剤形やその他の剤形が目的に応じて適宜選択される。また、必要に応じて、医薬品・医薬部外品・食品などに配合される任意の成分を含有することができる。また、本発明の効果を損なわない範囲で、他の薬効植物抽出物を添加してもよい。 Moreover, the collagenase inhibitor of this invention can also be used as the composition for oral. In this case, as a dosage form used for oral administration, specifically, as a solid preparation, powder, granules, tablets, capsules, troches and the like can be mentioned. Further, as liquid preparations, internal use liquids, external use liquids, suspensions, emulsions, syrups and the like are exemplified, and these dosage forms and other dosage forms are appropriately selected according to the purpose. In addition, it can contain optional components to be blended into medicines, quasi-drugs, foods, etc., as needed. In addition, other medicinal plant extracts may be added as long as the effects of the present invention are not impaired.
＜化粧料組成物＞
本発明の化粧料組成物は、上記本発明のコラゲナーゼ阻害剤を配合してなることを特徴とする。上述の通り、本発明のコラゲナーゼ阻害剤に含まれる米麹（白麹菌）や米麹（黒麹菌）の抽出物は、コラゲナーゼ阻害作用を有し、かつ、各種化粧料基材及び化粧料添加物に対する安定性も高いため、化粧料組成物に好適に使用することができる。
<Cosmetic composition>
The cosmetic composition of the present invention comprises the collagenase inhibitor of the present invention. As described above, the extract of rice bran (Branium bacillus) and rice bran (Ebony bacillus) contained in the collagenase inhibitor of the present invention has a collagenase inhibitory action, and various cosmetic bases and cosmetic additives It is also highly suitable for cosmetic compositions because it has high stability.
化粧料組成物への本発明の阻害剤の配合割合は任意であるが、米麹（白麹菌）や米麹（黒麹菌）の抽出物の含有量として、化粧料組成物が抗老化作用を十分に発揮する範囲で適宜選択される。米麹（白麹菌）や米麹（黒麹菌）の抽出物の化粧料組成物に対する割合は、抽出物における溶媒の割合や溶媒の種類等にもよるが、例えば、化粧料組成物の形態がジェルの場合は２〜５０重量％程度、形態が乳液の場合は２〜５０重量％程度、形態がクリームの場合は２〜５０重量％程度である。 Although the blending ratio of the inhibitor of the present invention to the cosmetic composition is arbitrary, the cosmetic composition has an anti-aging effect as the content of the extract of rice bran (white mold) or rice bran (black mold) It is suitably selected in the range which exhibits sufficiently. Although the ratio of the extract of rice bran (white mold) or rice bran (black mold) to the cosmetic composition depends on the ratio of the solvent in the extract, the type of solvent, etc., for example, the form of the cosmetic composition is It is about 2 to 50% by weight in the case of gel, about 2 to 50% by weight in the case of emulsion, and about 2 to 50% by weight in the case of cream.
本発明の化粧料組成物は、慣用の化粧料基材を適宜配合し、所望の剤型とすることができる。その形態は特に制限はないが、ジェル、乳液、クリーム等の形態が挙げられる。 The cosmetic composition of the present invention can be formulated into a desired formulation by appropriately blending a conventional cosmetic base. The form is not particularly limited, and examples include gel, emulsion, cream and the like.
化粧料基材としては、目的とする剤型に合わせて、水、水溶性有機溶媒、油脂、ロウ、高級脂肪酸、高級アルコール、エステル類等の従来公知の化粧料基材が適宜選択される。
また、本発明の化粧料組成物には、本発明の効果を損なわない範囲で通常化粧料や皮膚外用医薬で使用される任意の成分を添加することができる。かかる任意成分の具体例としては、増粘・ゲル化剤、酸化防止剤、紫外線吸収剤、色素剤、金属封鎖剤、防腐剤、ｐＨ調整剤、香料、ミツロウ等が挙げられる。これら任意成分の配合割合は、その目的に応じて適宜選択して決定することができる。また、本発明の効果を損なわない範囲で、他の薬効植物抽出物を添加してもよい。
As cosmetic base materials, conventionally known cosmetic base materials such as water, water-soluble organic solvents, fats and oils, waxes, higher fatty acids, higher alcohols, esters and the like are appropriately selected according to the intended formulation.
In addition, to the cosmetic composition of the present invention can be added any component that is usually used in cosmetics and external skin preparations as long as the effects of the present invention are not impaired. Specific examples of such optional components include thickening / gelling agents, antioxidants, ultraviolet light absorbers, coloring agents, sequestering agents, preservatives, pH adjusters, perfumes, beeswax and the like. The blending ratio of these optional components can be appropriately selected and determined according to the purpose. In addition, other medicinal plant extracts may be added as long as the effects of the present invention are not impaired.
＜機能性食品＞
一方、日常的に飲食することで、本発明のコラゲナーゼ阻害剤を摂取したい場合には、該食品、飲料に含有させて機能性食品としてもよい。
ここでいう「機能性食品」とは、一般食品に加えて、健康食品、栄養補助食品、栄養機能食品等、健康の維持の目的で摂取する食品および／又は飲料を意味している。なお、機能性食品として製品化する場合には、食品に用いられる様々な添加剤、具体的には、着色料、保存料、増粘安定剤、酸化防止剤漂白剤、防菌防黴剤、酸味料、調味料、乳化剤、強化剤、製造用剤、香料等を添加していてもよい。
<Functional food>
On the other hand, when it is desired to ingest the collagenase inhibitor of the present invention by eating and drinking on a daily basis, the food or beverage may be incorporated as a functional food.
The term "functional food" as used herein means, in addition to general foods, foods and / or beverages to be ingested for the purpose of maintaining health, such as health foods, nutraceuticals, and nutritious functional foods. In the case of commercialization as functional food, various additives used for food, specifically, colorants, preservatives, thickening stabilizers, antioxidant bleaches, antibacterial fungicides, An acidulant, a seasoning, an emulsifier, a fortifying agent, a manufacturing agent, a flavor and the like may be added.
本発明の機能性食品の対象となる、食品、飲料は特に限定されるものではない。例えば、食品として、ソーセージ、ハム、魚介加工品、ゼリー、キャンディー、チューインガムなどの食品類が挙げられる。また、飲料としては、各種の茶類、清涼飲料水、酒類、栄養ドリンクなどが挙げられる。この中でも、茶、ゼリーであることが特に好ましい。
本発明のコラゲナーゼ阻害剤は、このような食品、飲料に添加することにより、簡易に経口摂取することができる。
Foods and beverages to which the functional food of the present invention is applied are not particularly limited. For example, foods include sausages, ham, processed fish products, jellies, candies, chewing gum and other foods. Moreover, as a drink, various teas, soft drinks, liquors, a nutrient drink, etc. are mentioned. Among these, tea and jelly are particularly preferable.
The collagenase inhibitor of the present invention can be easily taken orally by adding it to such foods and beverages.
以下、実施例により本発明を更に詳細に説明するが、本発明は、その要旨を変更しない限り以下の実施例に限定されるものではない。なお、以下において、「１，３−ブチレングリコール」を「ＢＧ」と記載する場合がある。 Hereinafter, the present invention will be described in more detail by way of examples, but the present invention is not limited to the following examples as long as the gist thereof is not changed. In addition, "1, 3- butylene glycol" may be described as "BG" below.
＜米麹（白麹菌）＞
白麹菌(Aspergillus kawachii)を繁殖させた米麹は、株式会社ビオックから入手した白麹Ｋ型菌を用いて製造した。
＜米麹（黒麹菌）＞
黒麹菌（Aspergillus awamori）を繁殖させた米麹は、株式会社ビオックから入手した泡盛用黒麹種菌を用いて製造した。
<Rice bran (white bacillus)>
The rice bran on which Aspergillus niger (Aspergillus kawachii) was propagated was produced using Shirasagi K type bacteria obtained from Bioc Corporation.
<Rice bran (black blight fungus)>
The rice bran on which Aspergillus niger (Aspergillus awamori) was bred was produced using the awamori black mold species obtained from Bioc, Inc.
＜コラゲナーゼ阻害率の評価方法＞
〔試薬の調製〕
基質溶液：
Ｐｚペプチド(BACHEM社製)３．９ｍｇを、０．１Ｍトリス塩酸緩衝液(ｐＨ７．１、含２０ｍＭ塩化カルシウム)１０ｍＬに溶解して使用した(０．５ｍＭに相当)。
酵素溶液：コラゲナーゼ(TYPE IV、SIGMA-ALDRICH社製)５ｍｇを緩衝液１ｍＬに溶解させ２０μＬずつ分注し、−８０℃で保管する。使用時に緩衝液で５０倍に希釈して反応に用いた。
<Method of evaluating collagenase inhibition rate>
Preparation of Reagents
Substrate solution:
3.9 mg of Pz peptide (manufactured by BACHEM) was used by dissolving it in 10 mL of 0.1 M Tris-HCl buffer solution (pH 7.1, containing 20 mM calcium chloride) (corresponding to 0.5 mM).
Enzyme solution: 5 mg of collagenase (TYPE IV, manufactured by SIGMA-ALDRICH) is dissolved in 1 mL of buffer solution, aliquoted in 20 μL portions, and stored at -80 ° C. At the time of use, it was diluted 50 times with buffer and used for the reaction.
〔コラゲナーゼ阻害作用の測定法〕
これらの試料溶液５０μＬ、コラゲナーゼ溶液５０μＬ及び基質溶液４００μＬを混合し、３７℃で３０分間反応させた。次いで、２５ｍＭクエン酸溶液１ｍＬで反応を停止し、酢酸エチル５ｍＬで抽出した。遠心分離(１６００ｇ、１０分間)後、酢酸エチルを対照として、酢酸エチル層の波長３２０ｎｍにおける吸光度を測定した。対照には、試料溶液の代わりに各抽出溶媒を用い、また、それぞれのブランクとして、酵素溶液の代わりに超純水を加えて同様の操作を行った。これらの値からコラゲナーゼ阻害率を次式により算出した。
コラゲナーゼ阻害率(％)＝〔１−（Ａ−Ｂ）／（Ｃ−Ｄ）〕×１００
但し、Ａ：試料溶液の３２０ｎｍにおける吸光度、
Ｂ：試料溶液ブランクの３２０ｎｍにおける吸光度、
Ｃ：対照溶液の３２０ｎｍにおける吸光度、
Ｄ：対照溶液ブランクの３２０ｎｍにおける吸光度である。
[Measuring method of collagenase inhibitory action]
50 μL of these sample solutions, 50 μL of collagenase solution and 400 μL of substrate solution were mixed, and reacted at 37 ° C. for 30 minutes. The reaction was then quenched with 1 mL of 25 mM citric acid solution and extracted with 5 mL of ethyl acetate. After centrifugation (1600 g, 10 minutes), the absorbance at a wavelength of 320 nm of the ethyl acetate layer was measured using ethyl acetate as a control. As a control, each extraction solvent was used instead of the sample solution, and as a blank, ultrapure water was added instead of the enzyme solution to perform the same operation. From these values, the collagenase inhibition rate was calculated by the following equation.
Collagenase inhibition rate (%) = [1- (A-B) / (C-D)] × 100
Where A: absorbance of the sample solution at 320 nm,
B: Absorbance at 320 nm of sample solution blank,
C: Absorbance at 320 nm of control solution,
D: Absorbance at 320 nm of a control solution blank.
＜１．米麹（白麹菌）（抽出溶媒：水）のコラゲナーゼ阻害作用の評価＞
（実験例１）
抽出溶媒としての水（５０ｍＬ）に、米麹（白麹菌）（５ｇ）を浸漬し、室温で３時間静置した。遠心分離後、上澄みを濾過して、実験例１の抽出物を得た。
<1. Evaluation of the collagenase inhibitory action of rice bran (white bacillus) (extraction solvent: water)>
(Experimental example 1)
Rice bran (Blinter Bacilli) (5 g) was immersed in water (50 mL) as an extraction solvent and allowed to stand at room temperature for 3 hours. After centrifugation, the supernatant was filtered to obtain the extract of Experimental Example 1.
（実験例２）
抽出溶媒としての水（３０ｍＬ）に、麹（白麹菌）（１０ｇ）を浸漬し、５５℃で５時間静置した。遠心分離後、上澄みを濾過して、実験例２の抽出物を得た。
(Experimental example 2)
Aspergillus niger (B. koji) (10 g) was immersed in water (30 mL) as an extraction solvent and allowed to stand at 55 ° C. for 5 hours. After centrifugation, the supernatant was filtered to obtain the extract of Experimental Example 2.
（評価）
実験例１，２の抽出物について、上述の方法でコラゲナーゼの阻害率を評価した。結果を図１に示す。
(Evaluation)
The extracts of Experimental Examples 1 and 2 were evaluated for the collagenase inhibition rate by the method described above. The results are shown in FIG.
＜２．米麹（白麹菌）（抽出溶媒：水-ＢＧ混合溶媒）のコラゲナーゼ阻害作用の評価＞
（実験例３）
米麹（白麹菌）５ｇを、５０重量％ＢＧ/水５０ｍＬに浸漬し、３時間攪拌した。遠心分離後、上澄みをろ過して得られた抽出液を実験例３の抽出物とした。
<2. Evaluation of the collagenase inhibitory action of rice bran (white bacillus) (extraction solvent: water-BG mixed solvent)>
(Experimental example 3)
5 g of rice bran (white bacillus) was immersed in 50 mL of 50% by weight BG / water and stirred for 3 hours. The extract obtained by filtering the supernatant after centrifugation was used as the extract of Experimental Example 3.
（実験例４）
抽出溶媒としての５０重量％ＢＧ/水（３０ｍＬ）に、麹（白麹菌）（１０ｇ）を浸漬し、５５℃で５時間静置した。遠心分離後、上澄みを濾過して、実験例４の抽出物を得た。
(Experimental example 4)
Aspergillus (Bacillus bacillus) (10 g) was immersed in 50% by weight BG / water (30 mL) as an extraction solvent, and allowed to stand at 55 ° C. for 5 hours. After centrifugation, the supernatant was filtered to obtain the extract of Experimental Example 4.
（評価）
実験例３，４の抽出物について、上述の方法でコラゲナーゼの阻害率を評価した。結果を図２に示す。
(Evaluation)
The extracts of Experimental Examples 3 and 4 were evaluated for the collagenase inhibition rate by the method described above. The results are shown in FIG.
＜３．米麹（黒麹菌）のコラゲナーゼ阻害作用の評価＞
（実験例５）
抽出溶媒としての水（５０ｍＬ）に、米麹（黒麹菌）（５ｇ）を浸漬し、室温で３時間静置した。遠心分離後、上澄みを濾過して、実験例５の抽出物を得た。
<3. Evaluation of the collagenase inhibitory action of rice bran (E. bacillus)>
(Experimental example 5)
The rice bran (black bacillus) (5 g) was immersed in water (50 mL) as an extraction solvent and allowed to stand at room temperature for 3 hours. After centrifugation, the supernatant was filtered to obtain the extract of Experimental Example 5.
（実験例６）
米麹（黒麹菌）５ｇを、５０重量％ＢＧ/水５０ｍＬに浸漬し、３時間攪拌した。遠心分離後、上澄みをろ過して得られた抽出液を実験例６の抽出物とした。
(Experimental example 6)
5 g of rice bran (E. coli) was immersed in 50 mL of 50% by weight BG / water and stirred for 3 hours. The extract obtained by filtering the supernatant after centrifugation was used as the extract of Experimental Example 6.
（実験例７）
抽出溶媒としての５０重量％ＢＧ/水（３０ｍＬ）に、麹（黒麹菌）（１０ｇ）を浸漬し、５５℃で５時間静置した。遠心分離後、上澄みを濾過して、実験例７の抽出物を得た。
(Experimental example 7)
Aspergillus niger (10 g) was immersed in 50% by weight BG / water (30 mL) as an extraction solvent and allowed to stand at 55 ° C. for 5 hours. After centrifugation, the supernatant was filtered to obtain the extract of Experimental Example 7.
（評価）
実験例５〜７の抽出物について、上述の方法でコラゲナーゼの阻害率を評価した。結果を図３に示す。
(Evaluation)
The percentage of inhibition of collagenase was evaluated for the extracts of Experimental Examples 5 to 7 by the method described above. The results are shown in FIG.
＜４．加熱処理によるコラゲナーゼ阻害作用への影響＞
上記実験例１，２の抽出物を、沸騰水中で、１分加熱して、加熱前後の実験例１，２の抽出物によるコラゲナーゼ阻害率を比較した。結果を図４に示す。
本実験は、米麹（白麹菌）の抽出物にみられるコラゲナーゼ阻害活性に、米麹抽出物中に存在する菌体あるいは白麹菌が生産した酵素群が関与するかを調べるために行ったものである。加熱により菌体および酵素を失活させた抽出物においてもコラゲナーゼ阻害活性が認められたため、菌体および酵素は関与しないことが分かった。
<4. Effect of heat treatment on collagenase inhibitory action>
The extracts of Experimental Examples 1 and 2 were heated in boiling water for 1 minute, and collagenase inhibition rates by the extracts of Experimental Examples 1 and 2 before and after heating were compared. The results are shown in FIG.
This experiment was carried out to determine whether the collagenase inhibitory activity found in the extract of rice bran (Blinter Bacilli) involves the cells present in the extract of rice bran or the enzyme group produced by E. coli. It is. The collagenase inhibitory activity was also observed in the extract in which the cells and the enzyme were inactivated by heating, and it was found that the cells and the enzyme were not involved.
＜５．米麹（白麹菌）抽出物のエラスターゼ阻害作用の評価＞
参考例として、上記実験例４の米麹（白麹菌）抽出物を使用して、エラスターゼ阻害作用の評価を行った。
<5. Evaluation of the elastase inhibitory action of rice bran (Blinter Blight) extract>
As a reference example, evaluation of elastase inhibitory action was carried out using the rice bran (Branium tuberculosis) extract of Experimental Example 4 described above.
〔エラスターゼ活性阻害率の評価方法〕
エラスターゼ活性阻害作用は、ブタ膵臓由来エラスターゼ（SIGMA-ALDRICH社製）および合成基質スクシニル-Ｌ-アラニル-Ｌ-アラニル-Ｌ-アラニン-ｐ-ニトロアニリド）（SIGMA-ALDRICH社製）を用いて評価した。基質をジメチルスルホキシドで０．０５Ｍに調整し、使用時に、０.２Ｍ Ｔｒｉｓ-ＨＣｌ緩衝液（ｐＨ８．０）で１０倍希釈した５ｍＭ溶液を使用した。酵素は０.２Ｍ Ｔｒｉｓ-ＨＣｌ緩衝液（ｐＨ８．０）で１０μｇ/ｍＬに調製した。基質５０μＬを酵素１００μＬおよび試料５０μＬとともに３７℃で８分間反応させ、その後、吸光度４０５ｎｍで測定した（Ａ）。さらに酵素を添加せずに緩衝液を加えて同様の操作と測定を行った（Ｂ）。同様の測定を、試料を添加せずに抽出溶媒を加えて実施した（Ｃ、Ｄ）。上記方法で吸光度を求め、次式により、阻害率を算出した。
エラスターゼ活性阻害率（％）＝[１−（Ａ−Ｂ）/（Ｃ−Ｄ）] × １００
但し、Ａ：試料溶液の４０５ｎｍにおける吸光度、
Ｂ：試料溶液ブランクの４０５ｎｍにおける吸光度、
Ｃ：対照溶液の４０５ｎｍにおける吸光度、
Ｄ：対照溶液ブランクの４０５ｎｍにおける吸光度である。
[Method for evaluating elastase activity inhibition rate]
Elastase activity inhibitory activity was evaluated using porcine pancreas-derived elastase (manufactured by SIGMA-ALDRICH) and the synthetic substrate succinyl-L-alanyl-L-alanyl-L-alanine-p-nitroanilide (manufactured by SIGMA-ALDRICH) did. The substrate was adjusted to 0.05 M with dimethyl sulfoxide, and at the time of use, a 5 mM solution diluted 10-fold with 0.2 M Tris-HCl buffer (pH 8.0) was used. The enzyme was adjusted to 10 μg / mL with 0.2 M Tris-HCl buffer (pH 8.0). 50 μL of substrate was reacted with 100 μL of enzyme and 50 μL of sample for 8 minutes at 37 ° C., and then measured at an absorbance of 405 nm (A). Furthermore, the buffer solution was added without adding the enzyme, and the same operation and measurement were performed (B). Similar measurements were performed with the extraction solvent added without the sample added (C, D). The absorbance was determined by the above method, and the inhibition rate was calculated by the following equation.
Elastase activity inhibition rate (%) = [1-(A-B) / (C-D)] x 100
Where A: absorbance of the sample solution at 405 nm,
B: Absorbance at 405 nm of sample solution blank,
C: Absorbance at 405 nm of control solution,
D: Absorbance at 405 nm of a control solution blank.
（評価）
実験例４の抽出物について、上述の方法でエラスターゼ阻害率を評価したところ、エラスターゼ阻害率は１４．５％であった。
(Evaluation)
The elastase inhibition rate of the extract of Experimental Example 4 was evaluated by the above-mentioned method to be 14.5%.
本発明のコラゲナーゼ阻害剤は、優れたコラゲナーゼ活性阻害作用を示すため、各種外用剤、服用剤、化粧料組成物、機能性食品等の用途に使用することができる。 Since the collagenase inhibitor of the present invention exhibits an excellent collagenase activity inhibitory action, it can be used for applications such as various external preparations, medications, cosmetic compositions, functional foods and the like.
Claims (5)
前記溶媒抽出物が、前記米麹と、水または水を含有する混合溶媒からなる抽出溶媒とを接触させ、前記米麹に含有される成分を抽出して得られる抽出物であることを特徴とするコラゲナーゼ阻害剤（但し、抗酸化作用に基づく抗老化用途及び美白用途に用いるためのものを除く。）。 A solvent extract of rice bran on which at least one selected from Aspergillus kawachii and Aspergillus awamori is propagated,
The solvent extract is characterized in that the extract is obtained by bringing the rice bran into contact with an extraction solvent consisting of water or a mixed solvent containing water, and extracting the components contained in the rice bran. Collagenase inhibitors , except for those used for anti-aging and whitening applications based on antioxidant activity .
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