JP6746109B2 - Measurement method of skin stratum corneum water content using terahertz wave - Google Patents

Description

The present invention relates to a method for measuring the amount of water in the skin stratum corneum.

The outermost layer of the skin, the stratum corneum, is the main protective barrier against water evaporation and microorganisms, poisons and the like. In the stratum corneum, water is known to control physical properties such as substance permeability and plasticity of the stratum corneum, and the activity of an enzyme that has an important effect on desquamation of stratum corneum cells of the skin surface layer (non-patent document) Reference 1). Therefore, it is important to know the water content in the stratum corneum and its change in detail in order to evaluate the stratum corneum function.

Various methods have been devised so far for measuring the amount of water in the stratum corneum and its change. For example, paying attention to the fact that the electrical properties such as the electrical conductivity, capacitance, and dielectric constant of the stratum corneum change depending on the amount of water in the stratum corneum. An apparatus that tries to know the water content has been proposed (Patent Document 1, Non-Patent Document 2, Non-Patent Document 3).

These devices were useful in that the amount of water in the stratum corneum can be quickly and easily determined without damaging the skin. However, on the skin to which external preparations for skin such as cosmetics and its raw materials have been applied, the measured value may be affected by the applied material hindering or promoting the flow of electricity. There is a problem in that it is not possible to obtain a measured value that reflects only the water content of the.

In addition, focusing on the fact that the absorption of near-infrared rays changes depending on the amount of water in the stratum corneum, it is possible to measure near-infrared rays by using diffuse reflectance spectroscopy or attenuated total reflection spectroscopy (ATR method) using near-infrared rays. A method for measuring the layer water content has been proposed (Non-Patent Document 4).

These methods are also useful in that the amount of water in the stratum corneum can be known without damaging the skin. However, it is difficult to separate the near-infrared absorption spectrum of water from the absorption spectrum of substances contained in cosmetics and skin external preparations, or substances in the skin, and multiple scattering in skin tissue and absorption by melanin pigments. There is a problem that it is easily affected by substances other than water in the stratum corneum, and it cannot be said that the technique is sufficient to easily and accurately evaluate the stratum corneum function.

Furthermore, a method of observing the distribution of water content in the skin from a signal obtained by nuclear magnetic resonance of hydrogen atoms constituting water has also been proposed (Non-Patent Document 5). With the method using nuclear magnetic resonance, the water content distribution can be acquired as an image without damaging the skin, so it is possible to acquire data according to the desire to obtain information on the water content in which part of the skin. There is an advantage that can be done. However, there is a possibility that an accident such as a burn may occur depending on the fact that very expensive equipment and facilities are required, that the sample needs to be placed in a very strong magnetic field, and the composition of the external skin preparation applied to the skin. There was such a problem.

In addition, focusing on the fact that the intensity of Raman scattering changes depending on the amount of water in the skin, a method for measuring the distribution of the amount of water in the skin by using a confocal laser scanning Raman spectroscope has been proposed (non- Patent Document 6). In this way, the stratum corneum was assumed to consist of water and protein, the CH ₃ stretching vibration derived from the CH ₃ group constituting the Raman scattering intensity and protein based on OH stretching vibration derived from the OH group constituting the water By correcting the change in the detected intensity due to the depth from the sample surface from the ratio of the Raman scattering intensities based on it, it becomes possible to express the water content at each depth point in weight percent. However, in the case of cosmetics or skin to which an external preparation for skin has been applied, the substance that has CH ₃ stretching vibration is almost always contained in the applied product, so the Raman scattering intensity by CH ₃ stretching vibration was used. The problem was that the correction would not give the correct moisture weight percent.

Electromagnetic waves of 0.1 to 10 terahertz called terahertz waves are located in the intermediate region between light waves and radio waves. Terahertz waves have good material permeability, can be safely used by the human body, have small scattering in living tissues, and have extremely large absorption by water. Focusing on these characteristics, in recent years, attempts have been made to image the distribution of water and perform spectroscopic measurement using terahertz waves. For example, using terahertz wave time-domain spectroscopy, echoes of terahertz waves generated on the discontinuity surface of the group refractive index caused by the difference in the group refractive index of the stratum corneum, the epidermis, the dermis, and the stratum corneum of the subcutaneous tissue that make up the skin tissue. A method of measuring the amount of water based on pulse intensity information has been proposed (Non-Patent Document 7).

This method is to estimate the water content of the stratum corneum using the intensity of the echo pulse obtained from the air-stratum corneum interface, but echoes are obtained from the stratum corneum-skin interface where no clear interface is formed. It is difficult to obtain, and due to the thinness of the stratum corneum with respect to the pulse width of the terahertz wave, it remains difficult to obtain information on the water content inside the stratum corneum with high accuracy.

In addition, by measuring the attenuation of the terahertz wave due to the interaction between the surface wave (evanescent wave) in the terahertz band and the sample generated by injecting the terahertz wave under the condition of total reflection, the complex refractive index of the sample or Attempts have been made to obtain a complex dielectric constant (Patent Document 2, Non-Patent Document 8). In Patent Document 2, by measuring two spectra with different incident angle conditions, the complex refractive index of the sample is obtained without requiring a liquid layer having an extremely thin thickness unlike the transmission method. In Non-Patent Document 8, in order to eliminate the influence of water laminated on an ultrathin thin film sample such as a single cell layer, a so-called two-interface model that considers reflection at the interface between the thin film sample and water is used to measure the thin film sample. The complex permittivity is calculated.

However, both documents merely show that the complex refractive index or complex permittivity was measured. Further, the stratum corneum of the skin is not a single cell layer but a complex stack of dead keratinocytes, and it was not known whether it could be used for a method for measuring the amount of water in the stratum corneum.

Japanese Patent Publication No. 63-19016 JP, 2008-304444, A

Rawlings A et al., The effect of glycerol and humidity on desmosome degradation in stratum corneum., Arch Dermatol Res, 287, 457-64, 1995. Tagami H et al., Evaluation of the skin surface hydration in vivo by electrical measurement., J Invest Dermatol, 75, 500-7, 1980. Kumiko Hashimoto et al., Comparison of high-frequency conductivity measuring device and corneometer for measuring water content in the stratum corneum., Journal of Kosho, 11, 252-8, 1987. Potts RO et al., A noninvasive in vivo technique to quantitatively measure water concentration of the stratum corneum using attenuated total-reflectance infrared spectroscopy., Arch Dermatol Res, 277, 489-95, 1985. Mirrashed F et al., In vivo quantitative analysis of the effect of hydration (immersion and Vaseline treatment) in skin layers using high-resolution MRI and magnetization transfer contrast. Skin Research and Technology, 10, 14-22, 2004. Caspers PJ et al., In vivo confocal Raman microspectroscopy of the skin: Noninvasive determination of molecular concentration profiles., J Invest Dermatol, 116, 434-42, 2001. Takeshi Yasui et al., Development of non-contact and local skin water content measurement using terahertz electromagnetic pulse, Biomedical engineering, 42, 390-4, 2004. K. Shiraga et al., Determination of the complex dielectric constant of an epithelial cell monolayer in the terahertz region., Applied Physics Letters, 102, 053702, 2013.

By the way, when the measurement is performed using the ATR method, the terahertz wave needs to be totally reflected on the surface of the skin. The condition for total reflection is uniquely determined by the refractive index of the medium on the incident side, the refractive index of the medium on the emitting side, and the incident angle. That is, the ATR measurement condition depends on the incident angle of the terahertz wave, the refractive index of the prism (medium on the output side) that is in contact with the skin, and the refractive index of the stratum corneum (medium on the incident side) that is in contact with the prism.

However, the skin has a layered structure consisting of the stratum corneum, epidermis layer, dermis layer, and subcutaneous tissue layer, and it may be affected by moisture in the epidermis layer below the stratum corneum, so far. However, there is no known method for accurately measuring the refractive index of the stratum corneum, and the reality is that the refractive index of the stratum corneum is unknown. Therefore, it is not known whether the incident terahertz wave is totally reflected under the arbitrarily set conditions, and it is not possible to accurately grasp the condition for total reflection in the stratum corneum.

The present invention has been made in view of such background art, and an object of the present invention is to provide a method for measuring the water content of the stratum corneum using terahertz waves.

The measuring method of the present invention is a method for measuring the amount of water in the stratum corneum of the skin by total reflection attenuation spectroscopy using a terahertz wave, and irradiates a terahertz wave by bringing the skin surface into contact with the prism surface that is the terahertz wave emitting surface. And a step of obtaining an absorption coefficient.

According to the present invention, the amount of water in the stratum corneum of the skin can be easily measured using terahertz waves.

FIG. 1 is a schematic diagram showing the measurement concept of the present invention. FIG. 2 is a schematic diagram showing the measurement concept of the two-interface model. FIG. 3 is a diagram showing an absorption spectrum of water. FIG. 4 is a diagram showing an absorption spectrum of an aqueous sucrose solution. FIG. 5 is a diagram showing the relationship between the mass moisture content of the sucrose aqueous solution and the absorption coefficient, where (a) shows the measurement results at 0.5 THz and (b) shows the measurement results at 1.0 THz. ● indicates experimental value and ○ indicates literature value. FIG. 6 is a diagram showing an absorption spectrum of the stratum corneum model. FIG. 7 is a diagram showing the relationship between the mass moisture content and the absorption coefficient using the stratum corneum model. (a) shows the measurement result at 0.25 THz, (b) shows the measurement result at 0.5 THz, (C) is a figure which shows the measurement result in 1.0 THz. FIG. 8 is a diagram showing an absorption spectrum of pig skin (dried for 48 hours). FIG. 9 is a diagram showing the relationship between the mass moisture content of pig skin and the number of times of stripping. (a) shows the measurement result at 0.25 THz, (b) shows the measurement result at 0.5 THz, (c) ) Is a diagram showing a measurement result at 1.0 THz. FIG. 10 is a diagram showing an absorption spectrum of porcine skin (dry for 48 hours) after applying the two-interface model. FIG. 11 is a diagram showing the relationship between the absorption coefficient of pig skin and the number of strippings after the application of the two-interface model. (a) shows the measurement results at 0.25 THz, and (b) shows the results at 0.5 THz. (C) is a figure which shows the measurement result at 1.0 THz. FIG. 12 is an example of a diagram showing the complex refractive index of the pig stratum corneum obtained using the two-interface model, where (a) shows its real part and (b) shows its imaginary part. FIG. 13 is a diagram showing changes in the absorption coefficient before and after the application of the two-interface model, showing the rate of change before the application. (A) is a figure which shows the measurement result in 0.25 THz, (b) shows the measurement result in 0.5 THz, (c) is a figure which shows the measurement result in 1.0 THz. FIG. 14 is a diagram showing an absorption spectrum of an albumin aqueous solution. FIG. 15 is a diagram showing the relationship between the mass water content of an aqueous albumin solution and the absorption coefficient. (A) is a figure which shows the measurement result in 0.25 THz, (b) shows the measurement result in 0.5 THz, (c) is a figure which shows the measurement result in 1.0 THz. FIG. 16 is a diagram showing the relationship (0.5 THz) between the mass water content of pig skin and the number of strippings when an albumin aqueous solution is used as a stratum corneum model.

The measuring method of the present invention is a method for measuring the amount of water in the stratum corneum of the skin by total reflection attenuation spectroscopy using a terahertz wave, and irradiates a terahertz wave by bringing the skin surface into contact with the prism surface that is the terahertz wave emitting surface. Then, there is a step of obtaining a skin absorption coefficient.

FIG. 1 is a schematic diagram showing the measurement concept of the present invention. In the total reflection attenuation spectroscopy, the terahertz wave is incident on the prism under the condition of total reflection at the interface between the ATR prism (hereinafter referred to as “prism”) surface and the skin surface, and reflected at the interface between the prism surface and the skin surface. The terahertz wave is observed by a detector (not shown). At this time, the electric field intensity E to _out of the reflected terahertz wave is represented by the product of the electric field intensity E _in of the incident terahertz wave and the Fresnel reflection coefficient r to ₁₂ at the interface between the prism and the skin surface, as shown in Equation 1. analysis relationship holds and to the _{E ~ out = r ~ 12 ×} E in is performed. The Fresnel reflection coefficient r~ ₁₂ is obtained by the following mathematical formula 2. Here, in Equation 2, n ₁ is the refractive index of the prism, n ₂ is the complex refractive index of the skin, and θ is the incident angle of the terahertz wave.

(Equation 1)
E~ _out = r~ ₁₂ x E _in

Here, in the case of a substance having absorption of terahertz waves, assuming that the thickness of the sample on the prism surface is t and the absorption coefficient of the sample is α, the absorption intensity A(t) of the ATR spectrum observed from the reflected terahertz wave is When the thickness t of the sample is sufficiently larger than the seeping depth d _{p of the} evanescent wave (t→∞), the absorption intensity A(∞) of the ATR spectrum is expressed by the following Equation 3 The absorption intensity A(∞) is proportional to the seeping depth d _p of the evanescent wave. The seepage depth d _p of the evanescent wave is the depth at which the electric field strength of the incident wave is attenuated to 1/e as shown in Equation 5, the wavelength λ of the incident terahertz wave, the refractive index n _{1 of the} prism, It is uniquely determined by the refractive index n _{2 of the} sample and the incident angle θ with respect to the prism surface.

Further, since the absorption intensity A(t) of the ATR spectrum depends on the seeping depth d _p of the evanescent wave, that is, the wavelength λ of the incident terahertz wave, the absorption intensity A(t) depends on the wavelength λ by the ATR correction for dividing the ATR spectrum by the wavelength λ. The peak intensity is made uniform.

On the other hand, the propagation of an electromagnetic wave in a substance having absorption is represented by a complex refractive index n~. The complex index of refraction n~ is expressed by Equation 6 using the extinction coefficient κ of the substance, and it is known that the extinction coefficient κ and the absorption coefficient α of the substance have a proportional relationship shown in Equation 7. In Expression 6, n is the refractive index of the substance, and i is the imaginary unit.

That is, when the thickness t of the sample is sufficiently larger than the seeping depth d _{p of the} evanescent wave, the ATR spectrum A(∞) is proportional to the absorption coefficient α (equation 4), and the absorption coefficient α is extinction. There is a relationship that is proportional to the coefficient κ (Equation 7). Therefore, the absorption coefficient α can be obtained by directly measuring the absorption intensity A(t) of the ATR spectrum or by obtaining the extinction coefficient κ of the substance.

The ATR spectrum after reflection can be observed by terahertz time domain spectroscopy. In this method, a reflection spectrum and a phase difference spectrum are obtained by Fourier transforming the acquired time waveform of the terahertz wave. When the thickness t of the sample is sufficiently larger than the seepage depth d _{p of the} evanescent wave, the reflection spectrum R(ω) and the phase difference spectrum Φ(ω) are represented by the following Equations 8 and 9, respectively. .. In addition, the Fresnel reflection coefficient r _{12 at} the interface between the prism and the sample is expressed by Equation 2. Therefore, the complex index of refraction of the sample is obtained by obtaining the solution of the simultaneous equations of Equations 2, 8 and 9 from the reflection spectrum R(ω) and the phase difference spectrum Φ(ω) obtained by the terahertz time domain spectroscopy. n ₂ ~ is required. Note that r _ref in the equations 8 and 9 is the Fresnel reflection coefficient when the sample is not placed. Then, the absorption coefficient α of the substance is obtained from the imaginary part (extinction coefficient κ) of the obtained complex refractive index n ₂ ~ and the mathematical expression 7. That is, the measurement method of the present invention is a step of measuring the reflection spectrum and the phase difference spectrum obtained by irradiating the terahertz wave while contacting the skin with the prism surface, which is the terahertz wave emission surface, and the obtained reflection spectrum and the phase difference. Calculating the absorption coefficient of the stratum corneum from the spectrum.

In the measurement method of the present invention, since the skin is the measurement target, the absorption coefficient α obtained above represents the absorption coefficient of the skin, but the terahertz wave has a very large absorption by water, and skin tissue and body hair. The absorption coefficient α obtained above can be said to represent absorption by skin moisture, because it is less susceptible to absorption by proteins, lipids, blood components, and the like that make up.

In the present invention, the terahertz wave is emitted under the condition that the terahertz wave is totally reflected at the interface between the prism and the skin surface. Here, since the incident angle of the terahertz wave is larger than 0° and smaller than 90°, the condition for total reflection is that the refractive index of the prism is at least equal to or higher than the complex refractive index of the stratum corneum. Will be required. As will be described below, according to the actual measurement by the present inventor, it has been found that the real part of the complex refractive index of the stratum corneum of pig skin is about 2.5 or less when the terahertz wave of 0.1 THz or more is irradiated. This was confirmed by calculation using the so-called two-interface model described below, which takes into account the reflection coefficient occurring at the interface between the epidermis layer below the stratum corneum and the stratum corneum. That is, the complex refractive index of the stratum corneum can be calculated by applying the two-interface model using the complex refractive index of the skin layer and the thickness of the stratum corneum. The complex refractive index of the epidermal layer used for the calculation can be obtained, for example, by measuring the epidermal layer obtained by removing the keratin by stripping by the ATR method. The thickness of the stratum corneum may be measured, but the number of times of stripping may be used as the thickness, or an estimated value may be used.

Pig skin is very similar in composition and structure to human skin, and it can be said that the complex refractive index of the horny layer of human skin and that of the horny layer of porcine skin are almost the same. Therefore, the refractive index of the prism varies depending on the frequency of the terahertz wave used for measurement, but at a frequency lower than about 1 THz, at least 2.0 or more, preferably 2.5 or more, and preferably 3.0 or more is desired. In addition, a prism having a refractive index of 2.0 or more is used in a frequency band higher than about 1 THz. That is, a prism having a refractive index of 3.0 or higher can be measured using a terahertz wave of 0.1 THz or higher. As long as the prism has such a refractive index, its material is not limited and may be appropriately selected. For example, the material may be silicon (n=3.4) and MgO (n=3.1). The incident angle also depends on the refractive index of the prism and the frequency of the terahertz wave, and is set to an angle that satisfies the condition of total reflection.

The terahertz wave that can be irradiated in the present invention may be any electromagnetic wave in the terahertz wave region, and the lower limit of the frequency band is, for example, 0.1 THz, 0.25 THz, 0.5 THz, 1.0 THz. obtain. Further, the upper limit thereof may be 10.0 THz, 5.0 THz, 3.0 THz, and 2.0 THz, for example. As described below, the higher the frequency of the terahertz wave, the higher the absorption coefficient α of water and the better the sensitivity. However, the higher the frequency, the more the exudation depth d _p becomes too small and the amount of water in the stratum corneum is sufficient. Tend not to be reflected in. On the other hand, the bleeding depth d _p of the evanescent wave is proportional to the wavelength λ of the terahertz wave and is inversely proportional to the frequency thereof, as shown in Formula 5, so that the bleeding depth d _p is large on the low frequency side. Become. Therefore, on the low frequency side, the absorption in the epidermis layer below the stratum corneum is the main component, and it cannot be said that the absorption coefficient α of the stratum corneum is reflected. From this point of view, the terahertz wave of 0.1 THz or more and 3.0 THz is preferable.

Since the obtained absorption coefficient α is a value that reflects the water content of the stratum corneum, the relative water content of the stratum corneum can be known from the calculated absorption coefficient α. Further, since the absorption coefficient α is proportional to the molar concentration of the substance, the molar concentration of the substance can be obtained from the absorption coefficient α. That is, the mass water content (water content) of the stratum corneum is obtained from the obtained absorption coefficient α using the known relationship between the mass water content and the absorption coefficient α. When the mass water content of the stratum corneum is obtained, for example, it can be obtained by comparison with the absorption coefficient α measured using a stratum corneum model whose mass water content is known. An arbitrary model is used as the stratum corneum model. Since the stratum corneum is a layered structure of dead keratinocytes, for example, a sheet model obtained by compressing a powder of protein component collected from the skin to obtain moisture, and a stratum corneum containing protein and water. An example is a model consisting of an aqueous solution of albumin protein that is assumed to be composed of

Further, on the low frequency side, the seeping depth d _p of the evanescent wave becomes larger than the thickness d of the stratum corneum, which is considered to be affected by the moisture of the epidermis layer. Therefore, the absorption coefficient α can be obtained by applying the two-interface model described in Non-Patent Document 8 to obtain the corrected reflection coefficients r to ₁₂₃ . The two-interface model is a thin-layered sample and a laminated-layered sample in which the reflection coefficient r to ₁₂ at the interface between the prism and the thin-layered sample is changed when the laminated-layered sample is stacked on the thin-film sample as shown in FIG. It means a model using the total reflection coefficient (corrected reflection coefficient) r to ₁₂₃ in consideration of the reflection coefficients r to ₂₃ at the interface with and. That is, the interface between the thin film sample and the laminated sample is treated as having attenuation proportional to the Fresnel reflection coefficient r to ₂₃ at the interface, and the attenuation in the thin film sample is taken out and analyzed by removing this attenuation. Since the skin has a structure in which the epidermis layer is laminated on the extremely thin stratum corneum, it is considered that there is an interface between the stratum corneum and the epidermis layer. Therefore, if the reflection coefficients r to ₂₃ at this interface are taken into consideration, the error when the evanescent wave seeps out from the thickness of the stratum corneum can be reduced.

The reflection coefficient r~ _{23 at} the interface between the stratum corneum and the skin layer is calculated by the mathematical expression 10, and the corrected reflection coefficient r~ ₁₂₃ is _calculated by the following mathematical expression 11. Further, as the absorption coefficient α, the corrected reflection coefficients r to ₁₂₃ are substituted in place of the reflection coefficients r to _{12 in} the above formulas 8 and 9, and the simultaneous formulas (2), (10), (11) and (8) are used. It can be obtained by finding the solution of the number of steps. The thickness d of the stratum corneum and the complex refractive index n ₃ of the skin layer necessary for this correction are actually measured values, estimated values, and may be literature values.

Further, when the two-interface model is applied, if the seeping depth d _{p of} the evanescent wave is smaller than the thickness d of the stratum corneum, an error occurs due to the correction, and thus the seeping depth d _{p of the} evanescent wave is Frequency bands are used that are greater than the layer thickness d. For example, it can be a terahertz wave of preferably 3 THz or less, 2 THz or less, more preferably 1.5 THz or less, and desirably 1 THz or less.

As described above, by using the measuring method of the present invention, an extremely thin water content of the stratum corneum such as several tens of μm can be easily measured without damaging the skin.

Hereinafter, the present invention will be described more specifically based on the following examples, but the present invention is not limited to the following examples.

[Measurement of absorption coefficient of water]
In measuring the water content of the stratum corneum, the absorption coefficient α of water was first measured. The terahertz total reflection type attenuation spectrophotometer used for the measurement is equipped with a Si (n=3.42) prism as an ATR module and has an incident angle of 57° (hereinafter the same). Water was dropped on the prism surface for measurement. The reference was air. The result is shown in FIG.

Next, sucrose aqueous solutions having different water contents were measured, and the relationship between the absorption coefficient α and the mass water content (sucrose concentration) of the sucrose aqueous solution was investigated. The measurement was performed by dropping 1 mL of an aqueous sucrose solution on the prism surface. The results are shown in FIGS. 4 and 5. From this result, it was confirmed that the mass water content of the sucrose aqueous solution and the absorption coefficient α are proportional (FIG. 5). The coefficient of determination R ^{2 at} this time is, for example, 0.99 or more in the 0.5 THz and 1.0 THz bands, respectively, and the absorption coefficient at 0.5 THz is a literature value (Keiichiro Shiraga et al., Food Chemistry, 140, 315-320). , 2013).

[Water absorption coefficient in the stratum corneum model]
The absorption coefficient of water contained in the stratum corneum model was measured using the stratum corneum model. A sheet obtained by pressing a powder of interstitial components (hide powder non-chromated, manufactured by SIGMA) collected from the skin dermis was used as a stratum corneum model (sheet model). A sheet was prepared by repeatedly applying a pressure of 294 Pa or 589 Pa to 0.12 g of powder for 2 seconds three times. 200 to 500 μL of distilled water was added dropwise to the sheet, and the sheet was allowed to stand still in a container having a humidity of 100% for 48 hours so that the sheet was evenly spread with water. The water content was calculated from the wet mass immediately after the measurement and the dry mass after drying at 70° C. for 24 hours.

The obtained sheet was placed on a prism, and weights were placed on the prism for measurement. The results are shown in FIGS. 6 and 7. As shown in FIG. 7, it was shown that the mass water content and the absorption coefficient are proportional, and the mass water content can be calculated from the absorption coefficient.

[Measurement of water content of pig skin]
The water content of the stratum corneum of the pig was measured using the skin of the pig, and then the change in the water content due to stripping was measured. Frozen pig skin (2 cm×2 cm) was thawed at room temperature (about 20° C.), and then left to dry at room temperature (about 20° C.) for 48 hours with the side of the skin covered with adhesive tape. Then, a paper waste cloth (trade name "Kimwipe", manufactured by Nippon Paper Clare Co., Ltd.) containing 2 mL of distilled water was placed on the dried skin for 150 seconds and left to stand. After that, the surface of the skin was lightly wiped with a paper waste to obtain water-containing skin. In addition, after the measurement, the measurement was performed while removing the stratum corneum by stripping using cellophane tape, and the measurement was repeated until stripping was performed 25 times. In this stripping, it is considered that one layer of the stratum corneum (thickness of about 1 μm) was peeled off by one stripping, and the stratum corneum was almost removed by the stripping of 25 times.

The pig skin was placed on the prism, and a weight was placed on the prism for measurement. The water content was calculated from the measured absorption coefficient using a graph showing the relationship between the absorption coefficient and the mass water content obtained in Example 2. The results are shown in FIGS. 8 and 9. Similar to water and sucrose aqueous solution, the skin absorption coefficient tended to increase on the high frequency side. The water content of pig skin decreased with increasing stripping, but it tended to increase thereafter. As a result of water being absorbed into the skin after drying for an extremely short time, water absorption occurred in the outermost layer, but the water content was not sufficiently absorbed in the subsequent middle layer part, and the water content was low, and the deep skin part It is considered that the water content remained high as a result of not being dried sufficiently. Under this measurement condition, the evanescent wave seepage depth d _p is calculated to be about 70 to 10 μm, and it is considered that the water content of the horny layer of pig skin is measured. These measurement results not only reflect the water content of the pig skin, but also report that there is a water gradient in the stratum corneum (Wu J et al., Confocal Raman microspectroscopy of stratum corneum: apre-clinical validation. study, International Journal of Cosmetic Science, 30, 1, 47-56, 2008.: Egawa M et al., In vivo Estimation of Stratum Corneum Thickness from Water Concentration Profiles Obtained with Raman Spectroscopy, Acta Dermato Venereologica, 87, 4-8 , 2007), and it can be said that the method according to the present invention is a valid method. As described above, in the present invention, it is possible to accurately grasp the amount of water or the change in the amount of water in an extremely thin thin film sample such as the stratum corneum.

[Measurement of Water Content of Pig Skin (Corneum) Using Two-Interface Model]
From the measurement data obtained in Example 3, the absorption coefficient and the mass water content of the horny layer of pig skin were calculated using a two-interface model. The results are shown in FIGS. 10 and 11. In addition, an example of a diagram showing the relationship between the refractive index n (real part) and extinction coefficient κ (imaginary part) of the complex refractive index n to ₂ of the stratum corneum derived to calculate the absorption coefficient of the stratum corneum Is shown in FIG. The horny layer thickness d of porcine skin required for calculation of the reflection coefficients r to ₁₂₃ in the two-interface model was assumed to be 30 μm. Further, the complex refractive index n to ₃ of the skin layer is the complex refractive index after 25 strippings in Example 3.

As shown in FIG. 12, the real part of the complex refractive index n~ ₂ of the stratum corneum was measured to be 2.5 or less. Further, the skin absorption coefficient and the relationship between the stripping frequency and the absorption coefficient showed the same tendency as in Example 3. On the other hand, considering whether the two-interface model is applied or not, as shown in FIG. 13, when the two-interface model is applied, the skin absorption coefficient tends to be small as a whole, and the low-frequency side has a tendency to be smaller than the high-frequency side. Became smaller. It can be said that this is a result of the influence of the epidermis being eliminated because the evanescent wave seepage depth becomes larger on the low frequency side. In addition, the change in absorption coefficient depending on the application was smaller on the high frequency side than on the low frequency side.

[Corneum model using water-soluble albumin solution]
It was feared that the stratum corneum model used in Example 2 could not measure a water content having a mass water content of 30% or less due to the presence of voids. Therefore, the same measurement was performed using an aqueous solution of water-soluble albumin with a known mass concentration (w/w%) as a model in which voids do not exist. FIG. 14 shows the relationship between the absorption coefficient and the frequency. Further, FIG. 15 shows the relationship between the absorption coefficient and the mass water content. Further, FIG. 16 shows the relationship between the mass moisture content obtained from the measurement data obtained in Example 3 using the two-interface model and the number of strippings. The relationship between the mass moisture content and the absorption coefficient in FIG. 15 was derived from the following Equation 12. In Formula 12, M _W is the mass of water, M _D is the mass of albumin, V is the volume of the aqueous solution, and W _M is the mass concentration of water. In the terahertz band, the magnitude of the absorption coefficient is considered to have a linear relationship with the molar concentration of water molecules, but in an aqueous albumin solution, not only the mass of water molecules but also the mass of albumin should be considered when calculating the water content by mass. And the mass water content is not proportional to the molar concentration. On the other hand, when the albumin concentration increases (mass water content decreases), the decrease range of the water absorption coefficient is considered to be proportional to the increase amount of albumin (absorption is small enough to be ignored). It is considered that the coefficient also has a linear relationship. Then, the concentration of water and the concentration of albumin can be expressed by Equations 13 and 14 with A, B, C, and D as constants, respectively. Therefore, A, B, C, and D are obtained by approximating the measured absorption coefficient and the mass concentration of water, and the measured absorption coefficient and the mass concentration of albumin by a linear function, respectively, and the relationship shown in FIG. ) Got.

By using the albumin aqueous solution as a stratum corneum model, the influence of voids was reduced, and the moisture content of the stratum corneum could be measured even when the mass water content was 30% or less. Further, when the relationship between the mass water content and the stripping frequency was determined from this calibration curve, the same tendency as in Example 4 was obtained.

The present invention provides a new measurement method for measuring the water content of the stratum corneum.

Claims (4)

A method of measuring the water content of the stratum corneum of the skin by total reflection attenuation spectroscopy using terahertz waves, where the skin surface is brought into contact with the prism surface, which is the terahertz wave emission surface, and the frequency is 0.1 THz or more and 1.5 THz or less. A method for measuring the amount of water in the stratum corneum of the skin, comprising the step of irradiating the terahertz wave to determine the absorption coefficient. The measuring method according to claim 1 , wherein the refractive index of the prism is 2.0 or more. The measurement method according to claim 1 , wherein the refractive index of the prism is 3.0 or more. The method according to any one of claims 1 to 3 , further comprising a step of obtaining a water content of the stratum corneum from a comparison between the obtained absorption coefficient and an absorption coefficient measured using a skin model having a known water content. The measurement method described.