Starting from today, we will begin our first specialized science popularization: Methods for evaluating the efficacy of cosmetics. We know that whether effective cosmetics are good or not, no matter how flashy the marketing is, ultimately it depends on evidence and data, and the first thing to consider is whether your product can be absorbed and utilized by the skin. Therefore, today's first science popularization article will elaborate on transdermal absorption testing methods.
With the rapid development of the cosmetics industry and the increasing awareness of scientific skincare among consumers, the efficacy of cosmetics is no longer a vague marketing slogan, but a scientific conclusion that needs to be verified through rigorous experiments and accurate data. This article will be based on professional research results, comprehensively dissecting the underlying logic of the effectiveness of active ingredients in cosmetics from the physiological basis, influencing factors, and detection methods of transdermal absorption.
Transdermal absorption: the core prerequisite for the efficacy of cosmetics
（1） The core definition and industry significance of transdermal absorption
Transdermal absorption of cosmetics refers to the process in which active ingredients in cosmetics act on specific parts of the skin according to their claimed efficacy, and accumulate and stabilize their effects in that area. This process is fundamentally different from transdermal drug delivery: non surface drugs need to enter the systemic circulation through the skin to achieve systemic treatment, while cosmetic active ingredients cannot only stay in the stratum corneum or enter the systemic circulation, and need to accurately reach the targeted layer of the skin. From this, it can be seen that transdermal drug delivery technology for cosmetics is a technical activity, not necessarily the smaller the better, but rather a choice based on demand and application.
Transdermal absorption is the first hurdle for cosmetics to achieve efficacy, and it is also the core link in efficacy evaluation. No matter how many high-end active ingredients are added to a cosmetic, if it cannot break through the skin barrier and reach the targeted area, all claims of efficacy are just empty talk. Meanwhile, the safety of transdermal absorption is equally crucial - if ingredients such as sunscreen and preservatives excessively penetrate deep into the skin or even enter the bloodstream, they may cause skin irritation, allergies, and even systemic safety risks. Therefore, studying the transdermal absorption mechanism is not only the core technology of cosmetics research and development, but also the basis for ensuring consumer safety and effectiveness.
（2） The physiological basis of transdermal absorption: the "offensive and defensive battle" of the skin barrier
The skin is the largest organ in the human body, a natural barrier between the body and the outside world, and the biggest challenge for the transdermal absorption of active ingredients in cosmetics. The skin is divided into stratum corneum, epidermis, dermis, and subcutaneous tissue from the outside to the inside. Each layer forms different resistance to the penetration of ingredients and is also a targeted site for different functional ingredients.
1. Three major pathways for transdermal absorption of active ingredients
There are only three "channels" through which cosmetic ingredients can enter the skin, and the applicable ingredients and penetration efficiency of each channel are completely different:
① Intercellular pathway (main pathway): Through the intercellular spaces of the stratum corneum, it directly enters the dermis or subcutaneous tissue from the epidermis. This is the main permeation pathway for lipophilic components, and the lipid bilayer in the stratum corneum provides a "similar and soluble" permeation channel for lipophilic components, which is also the core absorption method for most cosmetic ingredients.
② Skin accessory pathway (secondary pathway): It enters the deep part of the skin through skin accessories such as hair follicles, sweat glands, sebaceous glands, etc. This channel does not pass through the stratum corneum barrier and is suitable for the penetration of large molecular components such as collagen and peptides. However, the proportion of skin appendages on the skin surface is extremely low, only 1% -2%, and the overall absorption efficiency is limited.
③ Intracellular pathway (auxiliary pathway): A small amount of substances are absorbed through the interior of the stratum corneum cells, mainly suitable for small molecule water-soluble components, relying on the hydration of keratinocytes to achieve permeation, with a relatively low overall permeation amount.
2. Targeted levels of different functional ingredients
The efficacy of cosmetics varies greatly, and the corresponding targeted action levels of ingredients are also completely different. Accurate "delivery" is the key to the effectiveness of ingredients:
① Skin surface: Sunscreen, closed moisturizer (such as some oils), only need to stay on the skin surface to form a protective film, without penetration, to avoid skin irritation caused by penetration.
② Skin Accessories: General anti acne, antiperspirant, and hair growth ingredients that need to act on hair follicles and sweat glands, targeting acne, excessive sweating, and hair loss problems.
③ Dermis layer: whitening agents, anti allergic, anti-aging ingredients that need to penetrate to the basal layer of the epidermis or superficial dermis. Whitening ingredients need to inhibit the activity of melanocytes in the basal layer, while anti allergic ingredients need to alleviate inflammation in the dermis layer. Anti aging ingredients should reach the dermis to act on fibroblasts, promote the synthesis of collagen and elastin, and achieve anti wrinkle and firming effects.
（3） Six key factors affecting transdermal penetration
Whether active ingredients can successfully break through the skin barrier is determined by the characteristics of the ingredients themselves and the physiological state of the skin, with six major factors directly determining the penetration efficiency:
1. Molecular size: the "hard threshold" for permeation
Molecular size is the "primary factor" affecting transdermal absorption, usually measured by molecular weight (Da) or molecular volume, with a linear relationship between the two. The industry recognized absorption threshold is 500Da - small molecular components with a molecular weight less than 500Da (such as vitamin C, niacinamide) are easily absorbed by the skin; Large molecular components larger than 500Da, such as hyaluronic acid, some peptides, and collagen, are difficult to penetrate the stratum corneum barrier. However, with the development of encapsulation technologies such as liposomes and microcapsules, a few large molecular components can also achieve efficient permeation.
2. Distribution coefficient: the balance art of lipophilic and hydrophilic
The distribution coefficient (logP) represents the lipophilicity and hydrophilicity of the components. The stratum corneum is rich in lipophilic substances, and lipophilic components are easier to distribute and penetrate, but not necessarily the stronger the lipophilicity, the better. If the fat solubility of the component is too strong, it will be difficult to enter the hydrophilic dermal tissue and instead form a permeation barrier; If the hydrophilicity is too strong, it cannot penetrate the lipid barrier of the stratum corneum. Only ingredients with a balanced lipophilic hydrophilic ratio can achieve optimal transdermal absorption.
3. Skin hydration: opening the "temporary channel" for penetration
Hydration refers to the absorption of water by the stratum corneum, which undergoes hydration and is the "gentlest way" to enhance the penetration of ingredients. Keratinocytes contain hydrophilic fibrin, and as the water content increases, the pore size and intercellular space of the cell membrane passively expand, greatly improving the efficiency of component permeation. Data shows that when the moisture content of the stratum corneum exceeds 10%, it enters a hydrated state; When the moisture content exceeds 50%, the permeability of the active ingredient can increase by 5-10 times. Hydration has a more significant increase in the permeability of lipophilic molecules and a smaller impact on hydrophilic molecules. This is also the core reason why the skin absorption effect is better after applying the facial mask - the sealing of the facial mask quickly improves the hydration of the stratum corneum.
4. Barrier integrity: a direct reflection of TEWL
Elevated TEWL directly reflects damage to the epidermal barrier. When the stratum corneum barrier is intact, the penetration of components is slow and controllable; When the barrier is damaged due to excessive cleaning, UV radiation, or skin diseases, TEWL and transdermal absorption rate increase synchronously, and external components are easily infiltrated in large quantities, which may enhance efficacy but also increase the risk of allergies and irritation.
5. Thickness of stratum corneum: "absorption differences" between location and population
The thickness and density of the stratum corneum in different parts of the skin vary greatly, with varying permeability. The order of absorption efficiency is: behind the ear>forehead>scalp>abdomen>arms>legs>sole. Meanwhile, age and gender can also affect the thickness of the stratum corneum: infants have a thinner stratum corneum and a much stronger absorption capacity than adults; The density of the stratum corneum on female skin is slightly lower than that on male skin, and the absorption efficiency is relatively higher. Due to the scarcity of human skin samples, pig, mouse, and rat skin are commonly used as substitutes in scientific research. Among them, pig skin has the structure and thickness closest to human skin, making it the most ideal animal substitute model.
6. Temperature and pH: The "invisible regulation" of absorption by the environment
When the skin temperature rises, blood circulation accelerates, the hydration of the stratum corneum increases, and the permeability of ingredients significantly improves. At the same time, the normal pH range of the human epidermal stratum corneum is 5.2-5.6, which is a weakly acidic environment. Within this pH range, the skin has the best ability to absorb ingredients.
Transdermal absorption detection method:
Scientific verification from in vivo to in vitro
With the implementation of the EU ban on animal testing for cosmetics, in vitro alternative testing has become mainstream in the industry, and transdermal absorption testing has formed a complete system of "in vivo testing+in vitro testing+new technologies", each method has its own applicable scenarios, advantages, and limitations.
（1） Live detection method: closest to real usage scenarios
Live detection uses human or animal subjects as experimental subjects, directly measuring the content of ingredients in the skin. The data is closest to the actual use effect, but there are issues such as invasiveness and ethical limitations.
1. Quantitative techniques
① Tape peeling method: suitable for ingredients targeting the stratum corneum (such as sunscreen). By peeling off the stratum corneum with multiple layers of tape, quantitatively detecting the content of components on the tape, and accurately calculating the total amount of components in the stratum corneum. The advantage is non-invasive and easy to operate, making it a commonly used method for detecting human stratum corneum components; The limitation is that it cannot detect the components of the dermis and deep layers, and the peeling procedure needs to be optimized to avoid component degradation.
② Microdialysis technology: One of the few techniques that can directly quantify the concentration of dermal components. By implanting microdialysis probes into the skin, real-time collection of dermal tissue fluid is performed to determine the concentration and pharmacokinetic characteristics of the components. Can evaluate whether the ingredients have reached therapeutic concentrations, compare the bioavailability of different formulations, without the need for biological fluids, and can establish multiple sampling points simultaneously; The limitation is that it belongs to minimally invasive detection, and probe implantation may cause slight damage to the skin.
③ Bubble suction method: forming blisters on the skin surface through negative pressure, extracting interstitial fluid and serum from the blisters, and quantitatively analyzing the content of components. The concentration time distribution curve of the ingredients in the skin can be obtained, but it belongs to invasive detection and is prone to skin damage, making it difficult to promote on a large scale.
2. In vivo imaging technology
Mainly used for animal experiments, tracking the distribution, retention time, and relative concentration of ingredients in the skin through fluorescent labeling (such as FITC, green fluorescent protein). Suitable for components such as collagen, hyaluronic acid, and polymer materials, allowing for intuitive observation of component permeation pathways; The limitation is that fluorescent labeling is required for the components, which has a limited scope of application. Currently, it is only used for animal experiments and cannot be directly applied to humans.
（2） In vitro testing method: skin model as the main method
In vitro testing does not require living organisms, and simulates the human skin environment through skin models and biomimetic membranes. It is easy to operate, low-cost, and has good reproducibility, and is currently the mainstream method for cosmetics research and efficacy verification.
The most commonly tested in vitro model is the in vitro skin model, which can be divided into animal skin model and artificially reconstructed skin model. Commonly used animal skins include mouse skin, rat skin, and pig skin, with pig skin being the closest to the human body; Artificially reconstructing skin models using human keratinocytes, melanocytes, and fibroblasts as raw materials to construct a three-dimensional model highly similar to the structure of human skin. From early two-dimensional cell monolayer culture to today's three-dimensional complex models, human body chips, and in vitro skin models, the degree of biological simulation continues to improve.
The most commonly used and classic method for in vitro detection of transdermal absorption is the in vitro diffusion cell method: the ex vivo skin, reconstructed skin model, or biomimetic membrane is placed between the diffusion cell and the receiving cell, with the stratum corneum facing the supply cell. After adding the test sample, it is incubated at a constant temperature, and the component content in the receiving liquid is detected regularly to calculate the permeation rate and retention amount. The advantages are simple operation, accurate data, batch detection, and direct reflection of the transdermal absorption capacity of ingredients; The limitation is that the in vitro model lacks complete microcirculation and there are certain individual differences, making it impossible to fully simulate the real human environment.
（3） New instrument detection: non-invasive, precise, and efficient
With the advancement of technology, new detection methods have emerged one after another, promoting transdermal absorption detection into the era of precision:
1. Microscopic radioactive self imaging technology
By labeling components with radioactive isotopes, subcellular level skin imaging can be achieved, which can accurately observe the localization of components in the epidermis, sebaceous glands, hair follicles, dermal papilla and other parts, especially suitable for evaluating the components delivered by hair follicles. The advantage is extremely high sensitivity and precise positioning; Limitations include radioactivity, need for biopsy, difficulty in labeling some components, and limited application.
2. Confocal Raman Microscopy Method
By combining Raman spectroscopy and microscopic analysis, non-invasive, real-time, and three-dimensional monitoring of the penetration of components within the stratum corneum can be achieved, accurately measuring the depth and content of component penetration without the need for labeling, making it fast and intuitive. It is currently the cutting-edge technology for non-invasive detection of transdermal absorption in the human body, widely used in the efficacy verification of high-end cosmetics.
3. Microfluidic chips and mass spectrometry imaging
Microfluidic organ chips can simulate human skin metabolism and physiological status, accurately replicate skin barrier function, and are suitable for component tolerance and penetration detection;
In recent years, there have been frequent controversial news events regarding the absorption and addition of certain components. In summary, a single detection method has limitations, and the joint verification of multiple methods is an industry consensus in order to draw scientifically reliable conclusions.