Evaluation method and design method for ultrafine bubble-containing compositions

Abstract

To provide a novel evaluation method for ultrafine bubble-containing compositions. [Solution] A method for evaluating an ultrafine bubble-containing composition, comprising: a contact step of bringing the ultrafine bubble-containing composition to be evaluated into contact with human skin or a substitute for human skin; an observation step of observing a section of the human skin or substitute for human skin after the contact step; and an evaluation step of evaluating the properties of the ultrafine bubble-containing composition based on the observation results.

Description

Technical Field

[0001] The present invention relates to a method for evaluating and designing an ultrafine bubble-containing composition.

Background Art

[0002] Due to their small size and large surface area characteristics, ultrafine bubbles are expected to be applied in many industries. In particular, their use in water treatment, agriculture, medicine, the cosmetics industry, etc. has been studied, and it is required to maximize their effects. However, existing evaluation methods may not fully reflect the characteristics of ultrafine bubbles, and there are problems with the accuracy, especially when evaluating biocompatibility and safety.

[0003] As a conventional evaluation method for ultrafine bubbles, a method of measuring physical properties such as the size, distribution, and duration of bubbles has been mainstream (Patent Document 1). However, with these methods, it is difficult to comprehensively evaluate the actual effects of ultrafine bubble-containing compositions on the human body and the effects as cosmetics, etc., in terms of application performance.

Prior Art Documents

Patent Documents

[0004]

Patent Document 1

Summary of the Invention

Problems to be Solved by the Invention

[0005] Under such circumstances, a new evaluation method for ultrafine bubble-containing compositions has been demanded, and in particular, the development of a technology that can directly and comprehensively evaluate the characteristics of ultrafine bubble-containing compositions has been demanded. Therefore, an object of the present invention is to provide a new evaluation method for ultrafine bubble-containing compositions.

Means for Solving the Problems

[0006] The present invention, which solves the above problems, is as follows [1] to [7].

[0007] [1] A method for evaluating an ultrafine bubble-containing composition, comprising: a contact step of bringing the ultrafine bubble-containing composition to be evaluated into contact with human skin or a substitute for human skin; an observation step of observing a section of the human skin or a substitute for human skin after the contact step; and an evaluation step of evaluating the properties of the ultrafine bubble-containing composition based on the observation results.

[0008] [2] The method for evaluating an ultrafine bubble-containing composition according to [1], wherein in the evaluation step, the less the structural change of the human skin or human skin substitute after the contact step, the better the ultrafine bubble-containing composition is evaluated to be.

[0009] [3] A method for evaluating an ultrafine bubble-containing composition according to [1] or [2], wherein the above-mentioned properties are one or more selected from non-invasiveness, retention, and permeability.

[0010] [4] A method for evaluating an ultrafine bubble-containing composition according to any one of [1] to [3], wherein in the observation step, the section is observed using a confocal laser scanning microscope.

[0011] [5] An evaluation method for an ultrafine bubble-containing composition according to any one of [1] to [4], comprising an image acquisition step after the observation step for acquiring an image of the section, wherein the evaluation step evaluates the properties of the ultrafine bubble-containing composition based on the image.

[0012] [6] A method for evaluating an ultrafine bubble-containing composition according to any one of [1] to [5], wherein in the contact step, the ultrafine bubble-containing composition to be evaluated is in contact with human skin or a substitute for human skin for 4 hours or more.

[0013] [7] The method for evaluating an ultrafine bubble-containing composition according to any one of [1] to [6], wherein the contact step is a step of bringing the ultrafine bubble-containing composition to be evaluated and a positive control or a negative control into contact with human skin or a human skin substitute, and the evaluation step is a step of evaluating the properties of the ultrafine bubble-containing composition based on a comparison of the observation results of the human skin or human skin substitute that has been in contact with the ultrafine bubble-containing composition and the human skin or human skin substitute that has been in contact with the positive control or negative control.

[0014] [8] A method for designing an ultrafine bubble-containing composition, comprising: a contact step of bringing the ultrafine bubble-containing composition into contact with human skin or a substitute for human skin; an observation step of observing a section of the human skin or a substitute for human skin after the contact step; an evaluation step of evaluating the properties of the ultrafine bubble-containing composition based on the observation results; and a selection step of selecting a formulation of the components of the ultrafine bubble-containing composition based on the evaluation results. [Effects of the Invention]

[0015] According to the present invention, the properties of an ultrafine bubble-containing composition can be evaluated. [Brief explanation of the drawing]

[0016] [Figure 1] This flowchart shows a method for evaluating an ultrafine bubble-containing composition according to the present invention. [Figure 2] This is a flowchart showing the design method for an ultrafine bubble-containing composition according to the present invention. [Figure 3](a) A figure showing a bright-field only stained image of a tissue section of a three-dimensional cultured epidermal model after treating polyglyceryl-10 laurate (containing sodium calcein), an aqueous solution of sodium calcein, and an aqueous solution of sodium calcein containing ultrafine bubbles for 4 hours. (b) A figure showing a stained image of only sodium calcein of a tissue section of a three-dimensional cultured epidermal model after treating polyglyceryl-10 laurate (containing sodium calcein), an aqueous solution of sodium calcein, and an aqueous solution of sodium calcein containing ultrafine bubbles for 4 hours. [Figure 4] A figure showing a tissue section of a three-dimensional cultured epidermal model after treating polyglyceryl-10 laurate, ultrapure water, and water containing ultrafine bubbles for 4 hours.

Embodiments for Carrying Out the Invention

[0017] <Method for Evaluating an Ultrafine Bubble-Containing Composition> Figure 1 is a flowchart showing a method for evaluating an ultrafine bubble-containing composition according to the present invention. Referring to this, the method for evaluating an ultrafine bubble-containing composition of the present invention will be described in detail. Hereinafter, ultrafine bubbles are also denoted as UFB.

[0018] First, in the contact step, the UFB-containing composition to be evaluated is brought into contact with human skin or a human skin substitute.

[0019] The present invention can be an object of evaluation without particular limitation as long as it is a composition containing UFB. Ultrafine bubbles (UFB) mean bubbles having a diameter of 1 μm or less. Examples of the UFB-containing composition include water containing UFB, cosmetics containing UFB, and quasi-drugs containing UFB.

[0020] Examples of the cosmetic material include, for example, skin cosmetics, hair cosmetics, detergents, shaving foams, finger disinfectants, and the like. Examples of the above skin cosmetics include cosmetics, lotions, milks, beauty essences, creams, face packs, foundations, essences, eye creams, and the like. Examples of the above hair cosmetics include hair rinses, conditioners, treatments, hair colors, hair color treatments, hair growth agents, hair styling agents, hair lotions, and the like. Examples of the above detergents include facial cleansers, cleansing agents, hair shampoos, body cleansers, and the like.

[0021] Any component of the UFB-containing composition in the present invention is not particularly limited and includes components usually used in cosmetics. Examples of the components include surfactants, oils, powders (pigments, dyes, resins), thickeners, resins, fragrances, ultraviolet absorbers (including organic and inorganic types, and may correspond to either UV-A or B), natural plant extract components, seaweed extract components, crude drug components, amino acids, whitening agents, blood circulation promoters, anti-inflammatory agents, bactericides, cooling agents, antiperspirants, salts, antioxidants, skin activators, pigments, vitamins, neutralizing agents, pH adjusters, and the like. Note that each of these agents is not limited to its use as such an agent and can be diverted for other uses according to the purpose. For example, vitamins can be used as antioxidants, or combined with other uses. For example, a surfactant can be used as an agent having an effect as a stabilizer for microbubbles. Furthermore, it is not limited to the above examples, and all that are commonly used by those skilled in the art can be used.

[0022] Here, the UFB-containing composition to be evaluated containing a cosmetic may be brought into contact with the human skin or human skin substitute, or before or after applying the cosmetic to the human skin or human skin substitute, the UFB-containing composition to be evaluated may be brought into contact with the human skin or human skin substitute.

[0023] The human skin may be from any part of the body, but is preferably the skin of the whole body, scalp, face, neck, arms, hands, and feet. When selecting the area to be evaluated, it is preferable to choose skin that is suitable for the expected usage environment.

[0024] The human skin substitute is not particularly limited as long as it can evaluate the properties of the UFB-containing composition, but includes, for example, porcine skin, a three-dimensional cultured skin model, or artificial skin. In particular, the human skin substitute is preferably a three-dimensional cultured skin model. These human skin substitutes can simulate the reactions of human skin to cosmetics and external stimuli. In particular, three-dimensional cultured skin models offer a high degree of fidelity to the dermis and epidermis, allowing for accurate evaluation of the penetration depth and retention volume of UFB-containing compositions. Furthermore, these human skin substitutes are resistant to environmental conditions such as temperature and humidity, enabling the simulation of various usage environments and yielding highly reliable results.

[0025] In the aforementioned contact step, the conditions for contacting the UFB-containing composition to be evaluated with human skin or a human skin substitute can be set in various ways depending on the purpose of evaluation and the characteristics of the composition.

[0026] The temperature conditions during the contact process are preferably adjusted to a range of 32°C to 37°C, which is close to normal body temperature. The lower limit of the temperature conditions in the contact process is preferably 33°C or higher, more preferably 34°C or higher, and even more preferably 35°C or higher. Depending on the specific evaluation purpose, lower temperatures of 4°C to 20°C or higher temperatures of 40°C or higher can also be used as temperature conditions.

[0027] The contact time during which the UFB-containing composition to be evaluated is brought into contact with human skin or a human skin substitute is preferably adjusted according to the properties of the UFB-containing composition being evaluated.

[0028] For example, when evaluating the short-term effects of a UFB-containing composition, it is preferable that the contact time between the UFB-containing composition being evaluated and human skin or a human skin substitute is short. The short contact time is preferably 1 second or more, more preferably 5 seconds or more, even more preferably 10 seconds or more, and even more preferably 1 minute or more. Furthermore, the short contact time is preferably 5 minutes or less, more preferably 4 minutes or less, even more preferably 3 minutes or less, and even more preferably 2 minutes or less.

[0029] Furthermore, when evaluating the long-term effects of a UFB-containing composition, it is preferable that the contact time between the UFB-containing composition being evaluated and human skin or a human skin substitute is long. The long contact time is preferably 1 hour or more, more preferably 2 hours or more, even more preferably 3 hours or more, and even more preferably 4 hours or more. Furthermore, the long-term contact time is preferably 24 hours or less, more preferably 20 hours or less, even more preferably 18 hours or less, and even more preferably 12 hours or less.

[0030] When evaluating the immediate effect of a UFB-containing composition, contact should be made for at least 1 second, preferably 5 seconds or more, more preferably 10 seconds or more, and even more preferably 1 minute or more. Furthermore, when evaluating the immediate effect of the UFB-containing composition, contact should be performed for 5 minutes or less, preferably 4 minutes or less, more preferably 3 minutes or less, and even more preferably 2 minutes or less.

[0031] Furthermore, when evaluating the storage capacity of the UFB-containing composition, contact should be performed for at least one hour, preferably for at least two hours, more preferably for at least three hours, and even more preferably for at least four hours. In particular, it is preferable to continuously supply the UFB-containing composition using an infinite capacity system.

[0032] Since the concentration of the UFB-containing composition also affects the evaluation, the UFB-containing composition may be appropriately diluted before use. The dilution ratio is 1 to 1000 times. When a UFB-containing composition is used at a high concentration, its effects can be evaluated in a concentration-dependent manner.

[0033] Methods for contacting the UFB-containing composition to be evaluated with human skin or a human skin substitute include directly applying the UFB-containing composition to the human skin or human skin substitute, immersing it, spraying it, or adding the UFB-containing composition to a culture cup containing human skin or a human skin substitute. The method of contact can be selected according to the purpose of the test. In addition to evaluation in a static environment, evaluation in a dynamic environment is also possible, and the properties of the UFB-containing composition can be evaluated in a dynamic fluid environment using a flow system or the like.

[0034] The pH and osmotic pressure of a composition affect its impact on skin and skin substitutes; therefore, the pH may be adjusted within the range of acidic, neutral, and alkaline, and the properties of the UFB-containing composition are evaluated based on this. Furthermore, if the UFB-containing composition contains cosmetic ingredients, pharmaceuticals, moisturizing ingredients, etc., it is possible to comprehensively evaluate the interactions and effects of these ingredients on human skin and human skin substitutes.

[0035] In the evaluation process described later, when evaluating the properties of the ultrafine bubble-containing composition based on a comparison of observation results between the human skin or human skin substitute that has been in contact with the UFB-containing composition and the human skin or human skin substitute that has been in contact with the positive control or negative control, it is preferable that the contact step is a step in which the ultrafine bubble-containing composition to be evaluated and the positive control or negative control are brought into contact with human skin or a human skin substitute, respectively.

[0036] For the positive control, general cosmetics and quasi-drugs can be used. For the negative control, water or water containing ultrafine bubbles can be used.

[0037] Next, in the observation step, the section of human skin or human skin substitute after the contact step is observed.

[0038] The aforementioned observations can be performed using various microscopes, such as confocal laser scanning microscopes, optical coherence tomography (FF-OCT, LC-OCT, SS-OCT, SD-OCT) systems, multiphoton microscopes, second harmonic generation light microscopes, photoacoustic microscopes, photoacoustic microscopes, confocal Raman microscopes, optical interference microscopes (OCM), and scanning electron microscopes. In particular, the use of a confocal laser scanning microscope is preferred.

[0039] In the present invention, the section of the human skin or human skin substitute after the stained contact step may be observed. The dyeing method is not particularly limited, and any known method can be used.

[0040] The present invention preferably includes an image acquisition step, after the observation step, in which an image of the section is acquired.

[0041] In the image acquisition step, an image of the section of human skin or human skin substitute after the contact step is acquired. The aforementioned images can be captured using the various microscopes described above.

[0042] Finally, in the evaluation process, the properties of the UFB-containing composition are evaluated. The properties of the UFB-containing composition are evaluated based on the observation results of the observation step. Preferably, the properties of the UFB-containing composition are also evaluated based on the image.

[0043] The aforementioned properties include non-invasiveness, retention, permeability, antibacterial properties, stability, rapid action, cleansing action, moisturizing action, and anti-inflammatory action. Preferably, the aforementioned properties include non-invasiveness, retention, or permeability.

[0044] In the evaluation step, preferably, the less structural change there is in the human skin or human skin substitute after the contact step, the more highly the UFB-containing composition is evaluated as being non-invasive.

[0045] Preferably, in the evaluation step, the properties of the ultrafine bubble-containing composition are evaluated based on a comparison of the observation results of the human skin or human skin substitute that has been in contact with the UFB-containing composition and the human skin or human skin substitute that has been in contact with the positive control or negative control.

[0046] More preferably, the observation results of the human skin or human skin substitute that has been in contact with the UFB-containing composition are compared with those of the human skin or human skin substitute that has been in contact with the positive control or negative control. The less the structural change in the human skin or human skin substitute that has been in contact with the UFB-containing composition, the more non-invasive the UFB-containing composition is evaluated to be.

[0047] In the present invention, "minimal structural change" means that when the UFB-containing composition comes into contact with human skin or a human skin substitute, there is little physical and / or chemical damage or change to the tissue structure of the human skin or human skin substitute.

[0048] Preferably, if, after contacting the UFB-containing composition with human skin or a human skin substitute, the tissue structure of the human skin or human skin substitute is maintained without disintegration, the UFB-containing composition can be evaluated as having excellent non-invasive properties. More preferably, if the tissue structure is maintained stably overall, the UFB-containing composition can be evaluated as having excellent non-invasive properties. This invention makes it possible to evaluate whether a UFB-containing composition is suitable for sensitive skin or delicate tissues.

[0049] Furthermore, preferably, the UFB-containing composition can be evaluated as having excellent non-invasive properties if the tissue structure of the human skin or human skin substitute is maintained when the UFB-containing composition is brought into contact with the human skin or human skin substitute. The degree to which the tissue structure of human skin or a human skin substitute is maintained can also be evaluated, for example, based on pre-established criteria. Specifically, the observation results of the tissue structure of human skin or a human skin substitute can be evaluated in stages such as "no peeling or collapse of the tissue structure of human skin or a human skin substitute (Stage 1)", "part of the tissue structure of human skin or a human skin substitute is peeling (Stage 2)", "the entire stratum corneum is collapsing (Stage 3)", and "the tissue beneath the tissue structure of human skin or a human skin substitute is also collapsing (Stage 4)". The degree of non-invasiveness of the UFB-containing composition can then be evaluated based on predetermined criteria for the degree to which the tissue structure of human skin or a human skin substitute is maintained.

[0050] Furthermore, in the evaluation step, preferably, the more the UFB-containing composition penetrates in the depth direction of the human skin or human skin substitute after the contact step, the more the UFB-containing composition is evaluated as having excellent permeability.

[0051] The penetration depth of the UFB-containing composition can be confirmed by examining tissue sections under a microscope or other means. This invention makes it possible to evaluate the ability of a UFB-containing composition to deliver its components deep into the skin.

[0052] Preferably, when the UFB-containing composition is brought into contact with the human skin or human skin substitute, and the UFB-containing composition penetrates to the depth of the human skin or human skin substitute after the contact step, compared to the tissue structure of the human skin or human skin substitute to which the negative control has been brought into contact, the UFB-containing composition can be evaluated as having excellent permeability.

[0053] Furthermore, preferably, a UFB-containing composition can be evaluated as having excellent permeability if the degree to which the UFB-containing composition penetrates in the depth direction of the human skin or human skin substitute after contact with the human skin or human skin substitute is 1 or more than the degree to which the UFB-containing composition penetrates in the depth direction of the human skin or human skin substitute after contact with the positive control or negative control. A UFB-containing composition can be evaluated as having excellent permeability if the degree of penetration is 1.1 times or more, more preferably 1.2 times or more, even more preferably 1.5 times or more, and even more preferably 2 times or more.

[0054] Furthermore, in the evaluation step, preferably, the more the UFB-containing composition is stored in the depth direction of the human skin or human skin substitute after the contact step, the better the UFB-containing composition is evaluated to have excellent storage properties.

[0055] In this invention, "storage ability" refers to the ability of the UFB-containing composition to remain in the skin for a long period of time after it has penetrated the skin.

[0056] For evaluating retention, observation of sections of human skin or human skin substitute after the contact process, after leaving them for an extended period, is suitable. This allows evaluation of the extent to which the penetrated components are retained and the duration of that retention. Furthermore, retention can be confirmed by taking samples of the sections and quantitatively measuring the residual amount over time.

[0057] Preferably, if 50% or more of the UFB-containing composition remains in the human skin or human skin substitute after the contact step, 24 hours after contact, the UFB-containing composition is evaluated as having excellent retention properties. More preferably, in the human skin or human skin substitute after the contact step, when the UFB-containing composition remains in the skin at 60% or more, more preferably 70% or more, even more preferably 80% or more, even more preferably 90% or more, and most preferably 100% at 24 hours after contact, it is evaluated as a UFB-containing composition having excellent retention.

[0058] Also, preferably, in the human skin or human skin substitute contacted with the positive control or negative control, when the degree of retention of the UFB-containing composition in the human skin or human skin substitute after the contact step is 1 time or more compared to the degree of retention of the positive control or negative control, it can be evaluated as a UFB-containing composition having excellent permeability. More preferably, when the degree of retention is 1.1 times or more, more preferably 1.2 times or more, more preferably 1.5 times or more, and even more preferably 2 times or more, it can be evaluated as a UFB-containing composition having excellent retention.

[0059] <Method for designing UFB-containing composition> The present invention also relates to a method for designing a UFB-containing composition. FIG. 2 is a diagram showing a method for designing a UFB-containing composition using the above-described evaluation method.

[0060] First, an evaluation of the UFB-containing composition is performed by a contact step of contacting the UFB-containing composition with human skin or a human skin substitute, an observation step of observing a section of the human skin or human skin substitute after the contact step, and an evaluation step of evaluating the characteristics of the UFB-containing composition based on the observation results. Here, the UFB-containing composition to be evaluated may be any one, such as an existing UFB-containing composition with a clear formulation of its components.

[0061] Next, in the selection process, the formulation of the components of the UFB-containing composition is selected based on the evaluation results. If the design is not completed at this stage, the formulation of the components of the UFB-containing composition that were evaluated is selected.

[0062] Then, the selected UFB-containing composition is evaluated again. Note that a comparison may be made between the UFB-containing composition before and after the change in formulation.

[0063] As described above, the design of the UFB-containing composition is completed when it is decided to proceed with the evaluation of the properties of the UFB-containing composition and the selection of component formulations, and an excellent component formulation for the UFB-containing composition is obtained.

[0064] The present invention will be described in detail below with reference to examples, but the present invention is not limited to these examples. [Examples]

[0065] <1> Preparation of UFB-containing composition First, as a base, an aqueous solution of sodium calcein salt (solvent: ultrapure water) was prepared to a final concentration of 0.1%. Ultrapure water was also prepared as a base.

[0066] Furthermore, as UFB-containing compositions, ultrapure water containing UFB (final UFB concentration of 218.9 million particles / mL), and an ultrapure aqueous solution containing 0.1% sodium calcein salt and UFB were prepared (final UFB concentration of approximately 137.1 million particles / mL). The number concentration of UFBs in the solution was measured using a nanoparticle analyzer (NanoSight NS300) manufactured by Malvern Panalytical.

[0067] <2> Evaluation of UFB-containing compositions The effects of a polyglyceryl-10 laurate and UFB-containing composition on three-dimensional cultured epidermis were observed. To demonstrate the penetration of the components, samples containing sodium calcein salt (Figure 3) and samples without it (Figure 4) were prepared. The specific procedure is described below.

[0068] <Sample Processing> (1) Culture cups containing a three-dimensional cultured skin model (LabCyte EPI-MODEL24 (J-TEC, 401124)) were transferred to a 24-well plate containing assay medium, and pre-cultured overnight in a CO2 incubator. (2) The following day, the culture cup was transferred to a sample vial containing PBS that had been stored in a 32°C constant temperature bath, 200 μL of PBS was added and hydrated for 30 minutes to 1 hour, and then the PBS was removed with a micropipette. (3) 500 μL of each test sample was added to a hydrated three-dimensional cultured skin model and treated for 4 hours.

[0069] <Frozen Section Protocol> (1) After adding each sample and treating the culture cups for 4 hours, they were washed with PBS and fixed by immersion in 10% neutral buffered formalin (Wako, 062-01661). (2) The fixed specimens were washed three times with Dulbecco's Phosphate-Buffered Saline (DPBS, MatTek, TC-PBS), then immersed in a 10% sucrose solution and left to stand at 4°C for 2 hours. (3) After that, the samples were immersed in a 20% sucrose solution and left to stand again at 4°C for 2 hours. (4) Using a disposable scalpel, the tissue was cut out from the culture cup and embedded in OCT compound. (5) After the embedded specimens were cooled to -30°C using a cryostat, sections were prepared. (6) Subsequently, the sections were nuclear-stained and mounted on slides using DAPI-Fluoromount-G (registered trademark, SBA, 0100-20) for novel nuclear staining.

[0070] <2> Tissue section observation results Each stained section was observed using a confocal laser scanning microscope (KEYENCE BZ-X800) (magnification 10x and 20x). The results are shown in Figures 3 and 4.

[0071] In samples treated with polyglyceryl-10 laurate, changes in the structure of the stratum corneum were observed (Figures 3 and 4). In particular, in samples containing sodium calcein salt, significant structural breakdown of the stratum corneum was observed (Figure 3). On the other hand, in samples treated with ultrapure water, UFB-containing water, sodium calcein salt aqueous solution, and UFB-containing sodium calcein salt aqueous solution, almost no structural changes in the stratum corneum were observed, and the tissue was confirmed to be normally maintained. This suggests that treatment with UFB-containing water does not cause stratum corneum destruction and is more non-invasive compared to treatment with polyglyceryl-10 laurate.

[0072] Furthermore, in samples treated with a UFB-containing sodium calcein salt aqueous solution, the stratum corneum structure was preserved, similar to samples treated with sodium calcein salt aqueous solution alone, and no tissue breakdown or abnormal changes were observed. This indicates that adding UFB to a sodium calcein salt aqueous solution does not adversely affect the stratum corneum.

[0073] Furthermore, it was found that in samples treated with a UFB-containing sodium calcein salt aqueous solution, sodium calcein salt penetrated the tissue more easily compared to samples treated with sodium calcein salt aqueous solution alone. This suggests that the permeability is improved by including UFB in the sodium calcein salt aqueous solution.

[0074] From the above results, it is understood that the properties of UFB-containing compositions can be evaluated using the evaluation method of the present invention. [Industrial applicability]

[0075] This invention can be used for evaluation and design in the research and development of ultrafine bubble-containing compositions.

Claims

1. A method for evaluating an ultrafine bubble-containing composition, A contact step involves bringing the ultrafine bubble-containing composition to be evaluated into contact with human skin or a human skin substitute, An observation step of observing the section of human skin or human skin substitute after the contact step, An evaluation step to evaluate the properties of the ultrafine bubble-containing composition based on the observation results, A method for evaluating an ultrafine bubble-containing composition, comprising the following features.

2. The method for evaluating an ultrafine bubble-containing composition according to claim 1, wherein in the evaluation step, the less the structural change of the human skin or human skin substitute after the contact step, the better the ultrafine bubble-containing composition is evaluated to be.

3. A method for evaluating an ultrafine bubble-containing composition according to claim 1 or 2, wherein the aforementioned characteristics are one or more selected from non-invasiveness, retention, and permeability.

4. The method for evaluating an ultrafine bubble-containing composition according to claim 1 or 2, wherein in the observation step, the section is observed using a confocal laser scanning microscope.

5. The observation step is followed by an image acquisition step in which an image of the section is obtained. The method for evaluating an ultrafine bubble-containing composition according to claim 1 or 2, wherein the evaluation step evaluates the properties of the ultrafine bubble-containing composition based on the image.

6. The method for evaluating an ultrafine bubble-containing composition according to claim 1 or 2, wherein in the contact step, the ultrafine bubble-containing composition to be evaluated is brought into contact with human skin or a substitute for human skin for four hours or more.

7. The aforementioned contact step is a step of bringing the ultrafine bubble-containing composition to be evaluated and the positive control or negative control, respectively, into contact with human skin or a human skin substitute. The evaluation step evaluates the properties of the ultrafine bubble-containing composition based on a comparison of observation results between the human skin or human skin substitute that has been in contact with the ultrafine bubble-containing composition and the human skin or human skin substitute that has been in contact with the positive control or negative control. A method for evaluating an ultrafine bubble-containing composition according to claim 1 or 2.

8. A method for designing an ultrafine bubble-containing composition, A contact step involves bringing an ultrafine bubble-containing composition into contact with human skin or a human skin substitute, An observation step of observing the section of human skin or human skin substitute after the contact step, An evaluation step to evaluate the properties of the ultrafine bubble-containing composition based on the observation results, A selection step to select the formulation of the components of the ultrafine bubble-containing composition based on the evaluation results, A method for designing an ultrafine bubble-containing composition.

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