JP2021181987A - Method for evaluating moisture content of skin stratum corneum - Google Patents
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- 210000000434 stratum corneum Anatomy 0.000 title claims abstract description 69
- 238000000034 method Methods 0.000 title claims abstract description 43
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 76
- 238000000862 absorption spectrum Methods 0.000 claims abstract description 72
- 238000005259 measurement Methods 0.000 claims abstract description 66
- 239000000523 sample Substances 0.000 claims abstract description 50
- 239000013074 reference sample Substances 0.000 claims abstract description 34
- 210000003491 skin Anatomy 0.000 claims abstract description 26
- 238000001035 drying Methods 0.000 claims abstract description 9
- 239000010410 layer Substances 0.000 claims description 22
- 239000012790 adhesive layer Substances 0.000 claims description 11
- 238000001228 spectrum Methods 0.000 claims description 11
- 229920001296 polysiloxane Polymers 0.000 claims description 8
- 239000000463 material Substances 0.000 claims description 6
- 102000011782 Keratins Human genes 0.000 claims description 5
- 108010076876 Keratins Proteins 0.000 claims description 5
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 6
- 239000000853 adhesive Substances 0.000 description 6
- 230000001070 adhesive effect Effects 0.000 description 6
- 239000013464 silicone adhesive Substances 0.000 description 5
- 238000005033 Fourier transform infrared spectroscopy Methods 0.000 description 4
- 210000004027 cell Anatomy 0.000 description 4
- 210000000245 forearm Anatomy 0.000 description 4
- 238000004566 IR spectroscopy Methods 0.000 description 3
- 239000002390 adhesive tape Substances 0.000 description 3
- XAGFODPZIPBFFR-UHFFFAOYSA-N aluminium Chemical compound [Al] XAGFODPZIPBFFR-UHFFFAOYSA-N 0.000 description 3
- 229910052782 aluminium Inorganic materials 0.000 description 3
- 229910052757 nitrogen Inorganic materials 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 229920002799 BoPET Polymers 0.000 description 2
- 239000004809 Teflon Substances 0.000 description 2
- 229920006362 Teflon® Polymers 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 239000002537 cosmetic Substances 0.000 description 2
- 229910003460 diamond Inorganic materials 0.000 description 2
- 239000010432 diamond Substances 0.000 description 2
- 238000011156 evaluation Methods 0.000 description 2
- 241000282412 Homo Species 0.000 description 1
- 238000007476 Maximum Likelihood Methods 0.000 description 1
- 229910021607 Silver chloride Inorganic materials 0.000 description 1
- 230000002238 attenuated effect Effects 0.000 description 1
- 238000000794 confocal Raman spectroscopy Methods 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 239000002274 desiccant Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000001704 evaporation Methods 0.000 description 1
- 230000008020 evaporation Effects 0.000 description 1
- 238000004299 exfoliation Methods 0.000 description 1
- 229910052732 germanium Inorganic materials 0.000 description 1
- GNPVGFCGXDBREM-UHFFFAOYSA-N germanium atom Chemical compound [Ge] GNPVGFCGXDBREM-UHFFFAOYSA-N 0.000 description 1
- 238000002129 infrared reflectance spectroscopy Methods 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 230000010354 integration Effects 0.000 description 1
- 210000002510 keratinocyte Anatomy 0.000 description 1
- 238000000691 measurement method Methods 0.000 description 1
- 239000004745 nonwoven fabric Substances 0.000 description 1
- 239000013307 optical fiber Substances 0.000 description 1
- 230000035699 permeability Effects 0.000 description 1
- 229920000139 polyethylene terephthalate Polymers 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- SBIBMFFZSBJNJF-UHFFFAOYSA-N selenium;zinc Chemical compound [Se]=[Zn] SBIBMFFZSBJNJF-UHFFFAOYSA-N 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- HKZLPVFGJNLROG-UHFFFAOYSA-M silver monochloride Chemical compound [Cl-].[Ag+] HKZLPVFGJNLROG-UHFFFAOYSA-M 0.000 description 1
- 230000003595 spectral effect Effects 0.000 description 1
- 239000002344 surface layer Substances 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
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Abstract
Description
本発明は、皮膚角層の水分量を評価する方法に関する。 The present invention relates to a method for evaluating the water content of the skin stratum corneum.
皮膚の最表層を構成する組織を角層と呼ぶ。肌荒れや乾燥等の肌状態の違いによって、角層を構成する水分量等が異なることが知られている。また角層は多くのスキンケア剤や化粧料の主たる作用部位である。したがって、角層の水分量やその変化を把握することは、肌状態の把握や、スキンケア剤や化粧料のスキンケア効果の検証等に有用である。
従来、皮膚の水分量を測定する方法として、スキコンやコルネオメーターを始めとする電気的な手法が広く用いられてきている。これらの手法は水分量の変動を鋭敏に且つ簡便にとらえることができる長所を有する一方、測定深度が不明瞭なこと、水分量を絶対量として検出できないこと、水以外の因子の影響を受けやすいこと等の短所も指摘されている。
このような水分測定法の課題を解決すべく、共焦点ラマン分光法を用いる方法（特許文献１）や、近赤外反射分光法を用いる方法（特許文献２）が検討されている。
しかしながら、これらの方法は、装置自体が大掛かりであるため場所の制限もあり、簡便に水分量を測定することはできなかった。このため、簡便に、皮膚の水分量を直接評価する方法が望まれていた。
The tissues that make up the outermost layer of the skin are called the stratum corneum. It is known that the amount of water constituting the stratum corneum differs depending on the difference in skin condition such as rough skin and dryness. The stratum corneum is also the main site of action for many skin care agents and cosmetics. Therefore, grasping the water content of the stratum corneum and its change is useful for grasping the skin condition and verifying the skin care effect of skin care agents and cosmetics.
Conventionally, as a method for measuring the water content of the skin, an electric method such as a skicon or a cornometer has been widely used. While these methods have the advantage of being able to detect fluctuations in water content sensitively and easily, the measurement depth is unclear, the water content cannot be detected as an absolute amount, and they are susceptible to factors other than water. Disadvantages such as things have also been pointed out.
In order to solve such a problem of the water content measurement method, a method using confocal Raman spectroscopy (Patent Document 1) and a method using near-infrared reflection spectroscopy (Patent Document 2) are being studied.
However, in these methods, since the device itself is large-scale, there are restrictions on the location, and it is not possible to easily measure the water content. Therefore, a simple method for directly evaluating the water content of the skin has been desired.
一方、赤外線分光法の一種である全反射減衰赤外分光法（Attenuated Total Reflection-IR、ATR-IR法）用いれば、皮膚を内部反射エレメントに押し付けるだけで角質細胞の状態を直接評価することができるが、やはり測定できる場所や環境、及び被験者に無理な態勢を取らせることに伴う制限がある。そこで測定に、サンプリングした皮膚角層を使用することが考えられる。被験者への負担が軽微かつ簡便なサンプリング方法として、テープ剥離により角層を採取し、ATR-IR法に基づく赤外分光法で、その組成等を解析する方法が提案されている。 On the other hand, if total reflection attenuation infrared spectroscopy (Attenuated Total Reflection-IR, ATR-IR method), which is a type of infrared spectroscopy, is used, the condition of keratinocytes can be directly evaluated by simply pressing the skin against the internal reflection element. However, there are still restrictions due to the place and environment where measurement can be performed, and the unreasonable posture of the subject. Therefore, it is conceivable to use the sampled skin horny layer for the measurement. As a simple sampling method with a light burden on the subject, a method has been proposed in which the stratum corneum is sampled by tape peeling and its composition is analyzed by infrared spectroscopy based on the ATR-IR method.
一例として、シリコーン系粘着層を有する粘着テープで皮膚表面の角層細胞を剥離し、該粘着面の赤外吸収スペクトルを測定する方法がある（特許文献３）。この技術は、角層細胞剥離後の該粘着テープの粘着面の赤外吸収スペクトルより、角層細胞剥離前の該粘着テープの粘着面の赤外吸収スペクトルを、係数を乗じたのちに差し引き、剥離角層に相当するスペクトル成分（以下、「剥離角層赤外吸収スペクトル」とも称す。）を算出することで、角層表層における角層構成成分及び角層付着物に関する化学情報を取得するものである。この方法を用いれば、被験者の測定場所にとらわれず、評価サンプルを取得することができる。しかしながら、このように、テープ剥離により採取した角層には水がほとんど含まれていないと考えられ、この方法では角層中の水分量を評価することはできないと考えられていた。 As an example, there is a method of peeling the stratum corneum cells on the skin surface with an adhesive tape having a silicone-based adhesive layer and measuring the infrared absorption spectrum of the adhesive surface (Patent Document 3). In this technique, the infrared absorption spectrum of the adhesive surface of the adhesive tape before the detachment of the stratum corneum cells is subtracted from the infrared absorption spectrum of the adhesive surface of the adhesive tape after the exfoliation of the stratum corneum cells after multiplying by a coefficient. By calculating the spectral component corresponding to the peeled horny layer (hereinafter, also referred to as "peeled horny layer infrared absorption spectrum"), chemical information on the horny layer constituent component and the horny layer deposit on the surface layer of the horny layer is acquired. Is. By using this method, it is possible to obtain an evaluation sample regardless of the measurement location of the subject. However, as described above, it was considered that the stratum corneum collected by tape peeling contained almost no water, and it was considered that the water content in the stratum corneum could not be evaluated by this method.
本発明が解決する課題としては、被験者への負担が軽微かつ簡便な方法を用いて、測定場所の制限なしに角層中の水分量を直接評価する方法を提供することである。 An object to be solved by the present invention is to provide a method for directly evaluating the water content in the stratum corneum without limiting the measurement location by using a simple method with a light burden on the subject.
本発明者らは、テープ剥離角層の赤外吸収スペクトルを解析した結果、意外にも水がほとんど含まれていないと考えられていたテープ剥離角層中にも少量ながら水分が含まれていることを見出し、その特定波長範囲における赤外吸収スペクトルのピークテープに基づいて、剥離した直後の角層中に含まれる水分量を評価できることを見出した。さらに、剥離したテープを適切に保管することによって、剥離した直後の角層と同様に水分量を評価できることを見出し、測定場所の制限なしに角層中の水分量を直接評価する方法を創出した。 As a result of analyzing the infrared absorption spectrum of the tape peeling horny layer, the present inventors unexpectedly contained a small amount of water in the tape peeling horny layer, which was thought to contain almost no water. It was found that the amount of water contained in the stratum corneum immediately after peeling can be evaluated based on the peak tape of the infrared absorption spectrum in the specific wavelength range. Furthermore, we found that by properly storing the peeled tape, we could evaluate the water content in the same way as the stratum corneum immediately after peeling, and created a method to directly evaluate the water content in the stratum corneum without restrictions on the measurement location. ..
本発明は、赤外吸収スペクトルのピークから皮膚角層の水分量を評価する方法であって、
（ｉ）テープ剥離により取得した角層を乾燥処理することで水分を除去し、水分を含まない乾燥角層の基準サンプルを調製する段階、
（ii）基準サンプルの吸収スペクトルＡを取得する段階、
（iii）測定したい皮膚の角層をテープで剥離し、測定サンプルを得る段階、
（iv）測定サンプルの吸収スペクトルを取得する段階、
（ｖ′）吸収スペクトルＡと水単体の吸収スペクトルＢのピークを基に、測定サンプル中に含まれる水分量の指標を決定する段階
を含む方法に関する。
The present invention is a method for evaluating the water content of the skin stratum corneum from the peak of the infrared absorption spectrum.
(I) The stage of preparing a reference sample of a dry stratum corneum that does not contain moisture by removing moisture by drying the stratum corneum obtained by peeling the tape.
(Ii) At the stage of acquiring the absorption spectrum A of the reference sample,
(Iii) The stage where the stratum corneum of the skin to be measured is peeled off with tape to obtain a measurement sample.
(Iv) The stage of acquiring the absorption spectrum of the measurement sample,
(V') The present invention relates to a method comprising a step of determining an index of the amount of water contained in a measurement sample based on the peaks of the absorption spectrum A and the absorption spectrum B of water alone.
また、本発明は、赤外吸収スペクトルの３８００〜３０００ｃｍ-1のピークから皮膚角層の水分量を評価する方法であって、
（ｉ）テープ剥離により取得した角層を乾燥処理することで水分を除去し、水分を含まない乾燥角層の基準サンプルを調製する段階、
（ii）基準サンプルの吸収スペクトルＡを取得する段階、
（iii）測定したい皮膚の角層をテープで剥離し、測定サンプルを得る段階、
（iv）測定サンプルの吸収スペクトルを取得する段階、
（ｖ）測定サンプルの３８００〜３０００ｃｍ-1のピークが、吸収スペクトルＡと水単体の吸収スペクトルＢの足し合わせに一致するよう、（式１）により、ＡとＢのスペクトルにかかる係数ａ、ｂを決定する段階、
ＳSC（λ）≒ａ（Ｓdry-SC（λ））＋ｂ（Ｓwater（λ）） （式１）
ＳSC（λ）：測定サンプルのスペクトル
Ｓdry-SC（λ）：乾燥角質（基準サンプル）の吸収スペクトル（Ａ）
Ｓwater（λ）：水単体の吸収スペクトル（Ｂ）
（vi）ｂ／ａの値を、測定サンプル中に含まれる水分量の指標とする段階
を含む方法に関する。
Further, the present invention is a method for evaluating the water content of the skin stratum corneum from the peak of 3800 to 3000 cm -1 in the infrared absorption spectrum.
(I) The stage of preparing a reference sample of a dry stratum corneum that does not contain moisture by removing moisture by drying the stratum corneum obtained by peeling the tape.
(Ii) At the stage of acquiring the absorption spectrum A of the reference sample,
(Iii) The stage where the stratum corneum of the skin to be measured is peeled off with tape to obtain a measurement sample.
(Iv) The stage of acquiring the absorption spectrum of the measurement sample,
(V) Coefficients a and b on the spectra of A and B according to (Equation 1) so that the peak of 3800 to 3000 cm -1 of the measurement sample matches the sum of the absorption spectrum A and the absorption spectrum B of water alone. Stage to decide,
S SC (λ) ≒ a (S dry-SC (λ)) + b (S water (λ)) (Equation 1)
S SC (λ): Spectrum of measurement sample S dry-SC (λ): Absorption spectrum of dry keratin (reference sample) (A)
S water (λ): Absorption spectrum of water alone (B)
(Vi) The present invention relates to a method including a step of using the value of b / a as an index of the amount of water contained in the measurement sample.
本発明によれば、テープで剥離した皮膚の角層の赤外吸収スペクトルを測定することにより、被験者負担が軽微で、測定する場所や環境によらず、短時間で皮膚角層の水分量を評価することができる。また、角層を剥離したテープを適切に保管することによって、剥離した直後の角層と同様に水分量を評価できる。 According to the present invention, by measuring the infrared absorption spectrum of the skin stratum corneum peeled off with tape, the burden on the subject is light, and the water content of the skin stratum corneum can be measured in a short time regardless of the measurement location or environment. Can be evaluated. In addition, by properly storing the tape from which the stratum corneum has been peeled off, the water content can be evaluated in the same manner as the stratum corneum immediately after peeling off.
本発明において、赤外吸収スペクトルは、ＡＴＲ−ＩＲ分光装置を用いて取得することができる。ＡＴＲ−ＩＲ分光装置としては、通常のＦＴ−ＩＲと、それに接続できるＡＴＲユニットを用いればよい。またＡＴＲユニットは光ファイバー型プローブでも良い。またＡＴＲユニットとＦＴ−ＩＲが一体化したシステムを用いても良い。また内部反射エレメントにも特に制限はないが、例えばゲルマニウム、ダイヤモンド、ＺｎＳｅ、Ｓｉ，ＰＩＲ（塩化銀製）を用いることができる。反射回数や入射角についても、全反射条件が維持できる限り、特に制限は無い。波数分解能は０．５〜３２ｃｍ-1の範囲で測定することが必要だが、感度とＳ／Ｎが両立できる２〜８ｃｍ-1の範囲にて測定することが好ましい。 In the present invention, the infrared absorption spectrum can be obtained by using an ATR-IR spectroscope. As the ATR-IR spectroscope, a normal FT-IR and an ATR unit that can be connected to the normal FT-IR may be used. The ATR unit may be an optical fiber type probe. Further, a system in which the ATR unit and the FT-IR are integrated may be used. The internal reflection element is not particularly limited, but for example, germanium, diamond, ZnSe, Si, and PIR (made of silver chloride) can be used. The number of reflections and the angle of incidence are not particularly limited as long as the total reflection conditions can be maintained. It is necessary to measure the wavenumber resolution in the range of 0.5 to 32 cm -1 , but it is preferable to measure it in the range of 2 to 8 cm -1 where both sensitivity and S / N can be achieved.
角層を剥離するために用いるテープは、ベースフィルムに粘着層を有するもので、ベースフィルムは、水を吸収しない素材であれば特に制限されず、粘着層は、水及び角層由来のピークに重ならない点から、シリコーン系粘着層であるのが好ましい。テープとしては、シリコーン粘着層を有するテープが好ましい。
シリコーン粘着層を有するテープは、一般には耐熱性や耐薬品性が求められる用途に市販されているため、ベースフィルムにテフロン（登録商標）やアルミが使用されているテープが入手し易い。これらのベースフィルムは強度もあるため、皮膚への貼り付け・取り外しが容易であり、本発明に適している。しかしテフロン（登録商標）やアルミ以外のベースフィルムでも問題は無い。
具体的には、シリコーン系粘着層を有するテープは、実施例に記載のアズワン株式会社ほか、３Ｍ社、日東電工株式会社、株式会社オカド、株式会社寺岡製作所等々より購入することができる。また、東レ・ダウコーニング株式会社、信越化学工業株式会社、株式会社レヂテックス等々より市販のシリコーン系粘着剤を各種フィルムや不織布等に塗工することにより、シリコーン系粘着層を有するテープを調製することができる。
The tape used to peel off the stratum corneum has an adhesive layer on the base film, and the base film is not particularly limited as long as it is a material that does not absorb water, and the adhesive layer has peaks derived from water and the stratum corneum. A silicone-based adhesive layer is preferable because it does not overlap. As the tape, a tape having a silicone adhesive layer is preferable.
Since the tape having a silicone adhesive layer is generally marketed for applications requiring heat resistance and chemical resistance, it is easy to obtain a tape using Teflon (registered trademark) or aluminum for the base film. Since these base films are also strong, they can be easily attached to and removed from the skin, and are suitable for the present invention. However, there is no problem with base films other than Teflon (registered trademark) and aluminum.
Specifically, the tape having a silicone-based adhesive layer can be purchased from AS ONE Corporation described in Examples, 3M Company, Nitto Denko Corporation, Okado Corporation, Teraoka Seisakusho Co., Ltd., and the like. In addition, a tape having a silicone-based adhesive layer may be prepared by applying a silicone-based adhesive commercially available from Toray Dow Corning Co., Ltd., Shin-Etsu Chemical Co., Ltd., Reditex Co., Ltd., etc. to various films and non-woven fabrics. Can be done.
シリコーン粘着面への角層の付着方法としては、該シリコーン粘着面を皮膚表面に直接押し付けて角層を付着させてもよいし、粘着層を有する別のテープを皮膚表面に押し付けて採取した角層を、該シリコーン粘着面に転写させてもよい。 As a method of adhering the horny layer to the silicone adhesive surface, the horny layer may be adhered by directly pressing the silicone adhesive surface against the skin surface, or another tape having an adhesive layer is pressed against the skin surface to collect the horns. The layer may be transferred to the silicone adhesive surface.
以下、本発明の皮膚の表面（角層）水分量の評価方法の手順について説明する。
（ｉ）テープ剥離により取得した角層を乾燥処理することで水分を除去し、水分を含まない乾燥角層の基準サンプルを調製する：
ここで、テープ剥離により取得する角層は、基準サンプルとなるものである。後に測定するサンプル（測定サンプル）と同一の種類（例えば、ヒト、ブタ等）であれば、その部位は制限されないが、同じ部位であるのが好ましい。例えば、頬の水分量を評価しようとする場合であっても、腕等の角層を基準サンプルとしても良い。また、測定する人の基準サンプルは必ずしも必要ではなく、例えば、ヒトであれば、基準サンプルの吸収スペクトルは、ほぼ同様なものが得られるため、他のヒトの基準サンプルを用いても良い。
Hereinafter, the procedure of the method for evaluating the water content on the surface (horny layer) of the skin of the present invention will be described.
(I) Moisture is removed by drying the stratum corneum obtained by peeling the tape, and a reference sample of the dry stratum corneum containing no moisture is prepared:
Here, the stratum corneum obtained by peeling the tape serves as a reference sample. If it is the same type (for example, human, pig, etc.) as the sample to be measured later (measurement sample), the site is not limited, but it is preferably the same site. For example, even when the water content of the cheek is to be evaluated, the stratum corneum such as the arm may be used as a reference sample. Further, the reference sample of the person to be measured is not always necessary. For example, in the case of humans, since the absorption spectra of the reference samples are almost the same, other human reference samples may be used.
角層の乾燥処理は、角層細胞を破壊せずに処理できる方法であればいずれでも良い。
例えば、乾燥窒素をフローさせた容器内に一定時間（１時間程度）放置する、乾燥した空気を送風する、乾燥材などを用いて低湿度にした容器内に放置する、自然乾燥する、容器内の気圧を減圧させて乾燥するなどの方法により、テープ剥離により取得した角層の水分を除去することができる。
これにより、水分を含まない乾燥角層の「基準サンプル」を得ることができる。
なお、乾燥角層の「基準サンプル」は、「（iv）測定サンプルの吸収スペクトルを取得する：」で用いた「測定サンプル」を乾燥処理して得ることもできる。
The drying treatment of the stratum corneum may be any method as long as it can be treated without destroying the stratum corneum cells.
For example, leave it in a container in which dry nitrogen has flowed for a certain period of time (about 1 hour), blow dry air, leave it in a container with low humidity using a desiccant, etc., allow it to dry naturally, in the container. Moisture in the stratum corneum obtained by peeling the tape can be removed by a method such as depressurizing the air pressure and drying.
This makes it possible to obtain a "reference sample" of the dry stratum corneum that does not contain water.
The "reference sample" of the dried horny layer can also be obtained by drying the "measurement sample" used in "(iv) Obtaining the absorption spectrum of the measurement sample:".
（ii）基準サンプルの吸収スペクトルＡを取得する：
前記により得られた基準サンプルと、必要に応じて、水単体について、それぞれの赤外吸収スペクトルを取得する。
赤外吸収スペクトルは、水由来のピークを取得できることから、少なくとも３８００〜３０００ｃｍ-1の範囲を測定するのが好ましい。
得られた赤外吸収スペクトルは、基準サンプルは、吸収スペクトルＡ，水単体は、吸収スペクトルＢとする。
なお、水単体の赤外吸収スペクトルである吸収スペクトルＢは、必ずしも毎回測定する必要はなく、測定装置の校正の際に行うなど、必要に応じた測定で足りる。
(Ii) Obtain the absorption spectrum A of the reference sample:
Infrared absorption spectra of the reference sample obtained as described above and, if necessary, water alone are obtained.
Since the infrared absorption spectrum can acquire a peak derived from water, it is preferable to measure at least in the range of 3800 to 3000 cm -1.
The obtained infrared absorption spectrum has an absorption spectrum A for the reference sample and an absorption spectrum B for water alone.
It should be noted that the absorption spectrum B, which is the infrared absorption spectrum of water alone, does not necessarily have to be measured every time, and measurement as needed, such as when calibrating the measuring device, is sufficient.
（iii）測定したい皮膚の角層をテープで剥離し、測定サンプルを得る：
次に、測定したい皮膚の角層をテープで剥離し、「測定サンプル」とする。
角層をテープで剥離するには、基準サンプルと同様に行えばよい。
得られた測定サンプルは、取得後直ちに吸収スペクトルを測定するか、それが困難な場合、角層の状態や水分量が変わらない環境で保存することにより、その後いつどこで測定しても良い。例えば、測定サンプルは、水が蒸散しない素材の、言い換えると水蒸気透過性がないか、あっても極めて小さいシートに、角層が付着したテープごと貼り付けて保存することができる。水が蒸散しない素材のシートは、材質としては、ＰＥＴなどを用いることができ、例えば、市販のＯＨＰシート（ＯＨＰフィルム 品番：VF-1420N（KOKUYO）、材質：リサイクルＰＥＴフィルム）等が挙げられる。さらに、角層が付着したテープを張り付けたＯＨＰシートを、別の粘着ラミネートフィルムで上下を挟むことで、角層中の水分の蒸散を防ぐことができ、1週間程度保存することができる。粘着ラミネートフィルムは水が蒸散しない素材であれば良く、例えばＰＥＴフィルムやアルミコート層を含むラミネートフィルムなどが挙げられる。
このように保存することにより、測定装置がない場所で測定サンプルを取得した後、測定装置のある場所まで運び、測定することが可能となる。例えば、測定したい皮膚の角層について、自宅でテープ剥離して得た測定サンプルをシートに貼り付け、検査機関に郵送などして、水分量の評価を依頼することも可能である。
(Iii) Peel off the stratum corneum of the skin to be measured with tape to obtain a measurement sample:
Next, the stratum corneum of the skin to be measured is peeled off with tape to obtain a "measurement sample".
To peel off the stratum corneum with tape, it may be done in the same manner as the reference sample.
The obtained measurement sample may be measured at any time and anywhere thereafter by measuring the absorption spectrum immediately after acquisition or, if it is difficult, storing it in an environment where the state of the stratum corneum and the water content do not change. For example, the measurement sample can be stored by attaching the tape with the horny layer to a sheet made of a material that does not evaporate water, in other words, a sheet having no water vapor permeability or even if it is extremely small. As a sheet made of a material that does not evaporate water, PET or the like can be used, and examples thereof include commercially available OHP sheets (OHP film product number: VF-1420N (KOKUYO), material: recycled PET film) and the like. Further, by sandwiching the transparency to which the tape to which the stratum corneum is attached is sandwiched between the upper and lower sides with another adhesive laminated film, the evaporation of water in the stratum corneum can be prevented and the OHP sheet can be stored for about one week. The adhesive laminated film may be any material as long as it does not evaporate water, and examples thereof include a PET film and a laminated film containing an aluminum coat layer.
By storing in this way, it becomes possible to acquire a measurement sample in a place where there is no measuring device and then carry it to a place where the measuring device is present for measurement. For example, it is possible to request an evaluation of the water content of the stratum corneum of the skin to be measured by attaching a measurement sample obtained by peeling the tape at home to a sheet and mailing it to an inspection institution.
（iv）測定サンプルの吸収スペクトルを取得する：
測定サンプルの吸収スペクトルを、前記と同様にして測定する。
吸収スペクトルは、測定サンプル取得後直ちに測定しても良く、前記のように、保存後に測定しても良い。
(Iv) Obtain the absorption spectrum of the measurement sample:
The absorption spectrum of the measurement sample is measured in the same manner as described above.
The absorption spectrum may be measured immediately after obtaining the measurement sample, or may be measured after storage as described above.
（ｖ）測定サンプルの３８００〜３０００ｃｍ-1のピークが、吸収スペクトルＡと水単体の吸収スペクトルＢの足し合わせに一致するよう、（式１）により、ＡとＢのスペクトルにかかる係数ａ、ｂを決定する：
ＳSC（λ）≒ａ（Ｓdry-SC（λ））＋ｂ（Ｓwater（λ）） （式１）
ＳSC（λ）：測定サンプルのスペクトル
Ｓdry-SC（λ）：乾燥角質（基準サンプル）の吸収スペクトル（Ａ）
Ｓwater（λ）：水単体の吸収スペクトル（Ｂ）
(V) Coefficients a and b on the spectra of A and B according to (Equation 1) so that the peak of 3800 to 3000 cm -1 of the measurement sample matches the sum of the absorption spectrum A and the absorption spectrum B of water alone. To decide:
S SC (λ) ≒ a (S dry-SC (λ)) + b (S water (λ)) (Equation 1)
S SC (λ): Spectrum of measurement sample S dry-SC (λ): Absorption spectrum of dry keratin (reference sample) (A)
S water (λ): Absorption spectrum of water alone (B)
すなわち、（式１）で示されるように、測定サンプルのスペクトル（ＳSC（λ））は、乾燥角質（基準サンプル）の吸収スペクトルＡ（Ｓdry-SC（λ））と、水単体の吸収スペクトルＢ（Ｓwater（λ））との足し合わせで、上記（式１）のように近似的に表現することができる（ａ及びｂは係数、λは波数）。
３８００〜３０００ｃｍ-1の吸収帯のピークに対して上記式を適用して、係数ａ、ｂを算出する。（式１）を最も良く再現する係数ａ、ｂの算出には、最小二乗法や最尤法を用いることができる（図１参照）。
That is, as shown in (Equation 1), the spectrum of the measuring sample (S SC (lambda)), the absorption spectrum of the dry keratin (reference sample) A (S dry-SC ( λ)) and the absorption of water alone By adding to the spectrum B (S water (λ)), it can be expressed approximately as in the above (Equation 1) (a and b are coefficients, λ is a wave number).
The above equations are applied to the peaks of the absorption band of 3800 to 3000 cm -1 to calculate the coefficients a and b. A least squares method or a maximum likelihood method can be used to calculate the coefficients a and b that best reproduce (Equation 1) (see FIG. 1).
（vi）ｂ／ａの値を、測定サンプル中に含まれる水分量の指標とする：
得られたａ及びｂより、ｂ／ａの値を、測定サンプル中に含まれる水分量の指標とする。
ｂ／ａは、測定サンプル中に含まれる水分量を相対的に示す値である。この値が大きいほど水分量が多いことを示し、皮膚表面（角層）の水分量を相対的に評価することができる。なお、ｂ／ａの値について、他のｂ／ａの値と比較する場合は、同じ乾燥角層（基準サンプル）の吸収スペクトル（Ａ）を用いることとする。
(Vi) The value of b / a is used as an index of the amount of water contained in the measurement sample:
From the obtained a and b, the value of b / a is used as an index of the amount of water contained in the measurement sample.
b / a is a value that relatively indicates the amount of water contained in the measurement sample. The larger this value is, the larger the water content is, and the water content of the skin surface (horny layer) can be relatively evaluated. When comparing the value of b / a with the value of other b / a, the absorption spectrum (A) of the same dry stratum corneum (reference sample) is used.
実施例で用いた装置、測定条件は、以下のとおりである。
（１）ＦＴ−ＩＲ装置：
Ｆｒｏｎｔｉｅｒ（Ｐｅｒｋｉｎ−Ｅｌｍｅｒ社、ＴＧＳ検出器）に、ユニバーサルＡＴＲユニット（ダイアモンド、４５°）またはＮｉｃｏｌｅ ｉＳ５（Ｔｈｅｒｍｏ Ｆｉｓｈｅｒ Ｓｃｉｅｎｔｉｆｉｃ社）にＡＴＲユニット（ｉＤ５ ＡＴＲ）を取り付けたものを用いた。
The apparatus and measurement conditions used in the examples are as follows.
(1) FT-IR device:
A Universal ATR unit (diamond, 45 °) or Nicole iS5 (Thermo Fisher Scientific) with an ATR unit (iD5 ATR) attached to a Frontier (Perkin-Elmer, TGS detector) was used.
（２）測定条件：
波数分解能４ｃｍ-1、積算４回
(2) Measurement conditions:
Wavenumber resolution 4 cm -1 , integration 4 times
実施例１
ヒト前腕内側部にシリコーン系粘着層を有するテープ（アズフロンテープＰＴＷ１３−００８、アズワン株式会社）を貼り付け、剥がすことにより角層を採取した。このテープ剥離角層を、乾燥窒素をフローさせた容器内に１時間放置した後、容器から取り出し、基準サンプルとした。容器から取り出した基準サンプルは、速やかにＩＲ測定を行い、基準サンプルの吸収スペクトルＡを取得した。また、同様に、水単体の吸収スペクトルＢを取得した。
低湿度環境下（２０℃、１５％）で１時間馴化した後、前記と同様のテープ（アズフロンテープＰＴＷ１３−００８、アズワン株式会社）を前腕内側部に貼り付け、剥がすことにより角層を採取した（測定サンプル）。この角層が付着したテープを、剥離後すぐにＯＨＰシート（ＯＨＰフィルム 品番：VF-1420N（KOKUYO））に貼りつけた。その後剥離テープをＯＨＰシートからはがしてすぐにＩＲ測定を行い、測定サンプルの吸収スペクトルを取得した。
測定サンプルの３８００〜３０００ｃｍ-1のピークが、基準サンプルの吸収スペクトルＡと水単体の吸収スペクトルＢの足し合わせに一致するよう、前記（式１）において、ＡとＢのスペクトルにかかる係数ａ、ｂを最小二乗法によって決定した。
また、高湿度環境下（２０℃、９０％）でも同様の測定を行った。
各湿度について、５点ずつ測定し、ｂ／ａの平均値を求めた。
その結果、低湿度環境下では、ｂ／ａ＝０．１１９であった。
また、高湿度環境下では、ｂ／ａ＝０．１７４であった。
以上の結果を図２に示す。これらより、ｂ／ａの値の平均値を比較すると、高湿度環境下では低湿度環境下よりｂ／ａが大きく、水分量が多いことを示す結果であった。
Example 1
A tape having a silicone-based adhesive layer (Asflon Tape PTW13-008, AS ONE Corporation) was attached to the inner part of the human forearm, and the stratum corneum was collected by peeling it off. This tape peeling horny layer was left in a container in which dry nitrogen was flowed for 1 hour, and then taken out from the container and used as a reference sample. The reference sample taken out of the container was immediately subjected to IR measurement, and the absorption spectrum A of the reference sample was obtained. Similarly, the absorption spectrum B of water alone was acquired.
After acclimatization in a low humidity environment (20 ° C, 15%) for 1 hour, the same tape as above (AS ONE Tape PTW13-008, AS ONE Corporation) is attached to the inner part of the forearm and peeled off to collect the stratum corneum. (Measurement sample). The tape to which this stratum corneum was attached was attached to an OHP sheet (OHP film product number: VF-1420N (KOKUYO)) immediately after peeling. After that, the release tape was peeled off from the OHP sheet, IR measurement was performed immediately, and the absorption spectrum of the measurement sample was obtained.
In the above (Equation 1), the coefficients a, which are applied to the spectra of A and B, so that the peak of 3800 to 3000 cm -1 of the measurement sample coincides with the sum of the absorption spectrum A of the reference sample and the absorption spectrum B of water alone. b was determined by the least squares method.
Further, the same measurement was performed even in a high humidity environment (20 ° C., 90%).
Each humidity was measured at 5 points, and the average value of b / a was calculated.
As a result, it was b / a = 0.119 in a low humidity environment.
Further, in a high humidity environment, b / a = 0.174.
The above results are shown in FIG. From these results, when the average values of b / a were compared, it was shown that the b / a was larger and the water content was higher in the high humidity environment than in the low humidity environment.
実施例２
ヒト前腕内側部にシリコーン系粘着層を有するテープ（アズフロンテープＰＴＷ１３−００８、アズワン株式会社）を貼り付け、剥がすことにより角層を採取した。このテープ剥離角層を、乾燥窒素をフローさせた容器内に１時間放置した後、容器から取り出し、基準サンプルとした。容器から取り出した基準サンプルは、速やかにＩＲ測定を行い、基準サンプルの吸収スペクトルＡを取得した。また、同様に、水単体の吸収スペクトルＢを取得した。
前記と同様のテープ（アズフロンテープＰＴＷ１３−００８、アズワン株式会社）を前腕内側部に貼り付け、剥がすことにより角層を採取した（測定サンプル）。この角層が付着したテープについて、剥離後すぐにＩＲ測定を行い、剥離直後の測定サンプルの吸収スペクトルｘを取得した。剥離テープ（測定サンプル）は、ＩＲ測定後すぐにＯＨＰシート（ＯＨＰフィルム 品番：VF-1420N（KOKUYO））に貼り付け、室温にて約３時間保存した。
室温にて約３時間保存した後、剥離テープ（測定サンプル）をＯＨＰシートからはがしてすぐにＩＲ測定を行い、保存後の測定サンプルの吸収スペクトルｙを取得した。
剥離直後の吸収スペクトルｘ、保存後の吸収スペクトルｙそれぞれについて、測定サンプルの３８００〜３０００ｃｍ-1のピークが、基準サンプルの吸収スペクトルＡと水単体の吸収スペクトルＢの足し合わせに一致するよう、前記（式１）において、ＡとＢのスペクトルにかかる係数ａ、ｂを最小二乗法によって決定した。
これらの測定を３サンプル（(1)、(2)、(3)）について行い、それぞれ、ｂ／ａの値を求めた。
その結果、ｂ／ａの値は、剥離直後において、(1)０．１１１、(2)０．１６４、(3)０．１５７であり、保存後において、(1)０．１１１、(2)０．１６３、(3)０．１５６であった。これらより、保存後も剥離直後と同様に、皮膚角層の水分量を評価できることが確認された。
Example 2
A tape having a silicone-based adhesive layer (Asflon Tape PTW13-008, AS ONE Corporation) was attached to the inner part of the human forearm, and the stratum corneum was collected by peeling it off. This tape peeling horny layer was left in a container in which dry nitrogen was flowed for 1 hour, and then taken out from the container and used as a reference sample. The reference sample taken out of the container was immediately subjected to IR measurement, and the absorption spectrum A of the reference sample was obtained. Similarly, the absorption spectrum B of water alone was acquired.
A tape similar to the above (AS Fluoron Tape PTW13-008, AS ONE Corporation) was attached to the inner part of the forearm and peeled off to collect the stratum corneum (measurement sample). The tape to which the stratum corneum was attached was subjected to IR measurement immediately after peeling, and the absorption spectrum x of the measurement sample immediately after peeling was obtained. The release tape (measurement sample) was attached to an OHP sheet (OHP film product number: VF-1420N (KOKUYO)) immediately after IR measurement, and stored at room temperature for about 3 hours.
After storing at room temperature for about 3 hours, the release tape (measurement sample) was peeled off from the OHP sheet, IR measurement was performed immediately, and the absorption spectrum y of the measurement sample after storage was obtained.
For each of the absorption spectrum x immediately after peeling and the absorption spectrum y after storage, the peak of 3800 to 3000 cm -1 of the measurement sample coincides with the sum of the absorption spectrum A of the reference sample and the absorption spectrum B of water alone. In (Equation 1), the coefficients a and b on the spectra of A and B were determined by the minimum square method.
These measurements were performed on 3 samples ((1), (2), (3)), and the value of b / a was obtained for each.
As a result, the values of b / a were (1) 0.111, (2) 0.164, and (3) 0.157 immediately after peeling, and (1) 0.111, (2) after storage. ) 0.163, (3) 0.156. From these, it was confirmed that the water content of the skin stratum corneum can be evaluated even after storage as in the case immediately after peeling.
Claims (5)
（ｉ）テープ剥離により取得した角層を乾燥処理することで水分を除去し、水分を含まない乾燥角層の基準サンプルを調製する段階、
（ii）基準サンプルの吸収スペクトルＡを取得する段階、
（iii）測定したい皮膚の角層をテープで剥離し、測定サンプルを得る段階、
（iv）測定サンプルの吸収スペクトルを取得する段階、
（ｖ′）吸収スペクトルＡと水単体の吸収スペクトルＢのピークを基に、測定サンプル中に含まれる水分量の指標を決定する段階
を含む方法。 It is a method to evaluate the water content of the skin stratum corneum from the peak of the infrared absorption spectrum.
(I) The stage of preparing a reference sample of a dry stratum corneum that does not contain moisture by removing moisture by drying the stratum corneum obtained by peeling the tape.
(Ii) At the stage of acquiring the absorption spectrum A of the reference sample,
(Iii) The stage where the stratum corneum of the skin to be measured is peeled off with tape to obtain a measurement sample.
(Iv) The stage of acquiring the absorption spectrum of the measurement sample,
(V') A method including a step of determining an index of the amount of water contained in a measurement sample based on the peaks of the absorption spectrum A and the absorption spectrum B of water alone.
（ｉ）テープ剥離により取得した角層を乾燥処理することで水分を除去し、水分を含まない乾燥角層の基準サンプルを調製する段階、
（ii）基準サンプルの吸収スペクトルＡを取得する段階、
（iii）測定したい皮膚の角層をテープで剥離し、測定サンプルを得る段階、
（iv）測定サンプルの吸収スペクトルを取得する段階、
（ｖ）測定サンプルの３８００〜３０００ｃｍ-1のピークが、吸収スペクトルＡと水単体の吸収スペクトルＢの足し合わせに一致するよう、（式１）により、ＡとＢのスペクトルにかかる係数ａ、ｂを決定する段階、
ＳSC（λ）≒ａ（Ｓdry-SC（λ））＋ｂ（Ｓwater（λ）） （式１）
ＳSC（λ）：測定サンプルのスペクトル
Ｓdry-SC（λ）：乾燥角質（基準サンプル）の吸収スペクトル（Ａ）
Ｓwater（λ）：水単体の吸収スペクトル（Ｂ）
（vi）ｂ／ａの値を、測定サンプル中に含まれる水分量の指標とする段階
を含む方法。 It is a method of evaluating the water content of the skin stratum corneum from the peak of 3800 to 3000 cm -1 in the infrared absorption spectrum.
(I) The stage of preparing a reference sample of a dry stratum corneum that does not contain moisture by removing moisture by drying the stratum corneum obtained by peeling the tape.
(Ii) At the stage of acquiring the absorption spectrum A of the reference sample,
(Iii) The stage where the stratum corneum of the skin to be measured is peeled off with tape to obtain a measurement sample.
(Iv) The stage of acquiring the absorption spectrum of the measurement sample,
(V) Coefficients a and b on the spectra of A and B according to (Equation 1) so that the peak of 3800 to 3000 cm -1 of the measurement sample matches the sum of the absorption spectrum A and the absorption spectrum B of water alone. Stage to decide,
S SC (λ) ≒ a (S dry-SC (λ)) + b (S water (λ)) (Equation 1)
S SC (λ): Spectrum of measurement sample S dry-SC (λ): Absorption spectrum of dry keratin (reference sample) (A)
S water (λ): Absorption spectrum of water alone (B)
(Vi) A method including a step of using the value of b / a as an index of the amount of water contained in the measurement sample.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2020086606 | 2020-05-18 | ||
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