Unlocking the potential of cosmetic dermal delivery with ethosomes: A comprehensive review

In a world where hair loss, acne, and skin whitening are common concerns, ethosomes emerge as a captivating breakthrough in cosmetic drug delivery.


| INTRODUC TI ON
The skin is the largest organ in the human body and can be utilized to administer drugs for both localized and systemic effects.
However, the stratum corneum, the outermost layer of the skin, acts as a barrier that restricts the penetration of hydrophilic drugs and those with a high molecular weight. 1 Ethosomes have gained attention as a type of vesicular drug delivery system that can enhance drug penetration by increasing the fluidity and permeability of the cell membrane.They are composed of a monolayer of phospholipids and ethanol, 2 and can carry both hydrophilic and hydrophobic compounds, making them a versatile and effective alternative to traditional transdermal drug delivery systems. 3hosomes can be classified into several types depending on their composition and preparation method, including lipid-based ethosomes, 4 protein-based ethosomes, 5 polymer-based ethosomes, 6 microemulsion-based ethosomes, 7 liposome-based ethosomes, 8 and dendrimer-based ethosomes. 9e use of ethosomes in drug delivery has shown promise in several preclinical and clinical studies.Ethosomes have been successfully used to improve the delivery of drugs such as diclofenac, 10 terbinafine, 11 and tamoxifen, 12 through the skin, as well as through other routes of administration such as the oral and nasal routes. 13,14Moreover, the use of ethosomes has been associated with increased bioavailability, improved drug efficacy, and reduced toxicity. 15,16ven their potential in transdermal drug delivery and their increasing popularity in the pharmaceutical industry, there is a growing need for a comprehensive review of the current state of research on ethosomes.Such a review can provide an overview of the various types of ethosomes, their method of preparation, physicochemical properties, drug loading and release mechanisms, as well as their applications in cosmetic drug delivery.It can also highlight the challenges and opportunities associated with the use of ethosomes and identify future directions for research.

| PREPAR ATI ON S OF E THOSOME S
There are several methods for preparing ethosomes, and may vary depending on the specific conditions and requirements of the ethosome preparation.Their production is relatively simple and costeffective, making them a suitable alternative to other drug delivery systems. 17Table 1 highlighted the most important methods that can be used to prepare ethosomes.

| Modified hot method path
In this method, two dispersions were heated separately to 40°C, the first mixture is composed of phospholipid dispersion and the second one is composed of propylene glycol and ethanol.The two dispersions were mixed after dissolving the drug in water or ethanol. 18e foreseeable size of vesicles can be fulfilled by sonication, and extrusion. 19

| Modified cold method path
This method is a commonly used technique for preparing ethosomes. 20It consists of two phases; the organic and the aqueous phases.The organic phase is made by blending a phospholipid dispersion in ethanol at room temperature, followed by heating in a water bath to 30°C.Then, the organic phase is gradually added to 30°C water and stirred for 5 min in a covered container.The mixture is then cooled to room temperature.The desired size of the vesicles can be achieved by using sonication or extrusion. 21

| Vortex/sonication method edge
This method is a simple and effective common technique used for ethosome preparation.In this method, phospholipids and edge activators are mixed in a phosphate buffer and subjected to forceful shaking and vortexing to ensure an even distribution of the components.The suspension is then sonicated using a vortex or sonicator to create vesicles.The size of the vesicles can be controlled by adjusting the sonication time and intensity.Finally, the suspension is extruded through membranes of various sizes to obtain vesicles of the desired size.This method is versatile, scalable, and can be used to prepare a wide range of drugs, both hydrophilic and hydrophobic.
It is also cost-effective and can be easily scaled up for large-scale production. 22

| Rotary film evaporation
Rotary film evaporation is a useful technique for producing ethosomes for drug delivery applications. 23Phospholipids are dispersed in an organic solvent within a spherical bottom flask.The organic solvent is evaporated using a rotary evaporator, which leaves a thin film of lipids around the inner walls of the flask.The lipid film is then hydrated using an aqueous medium containing the drug that leads to formation of lipid bilayer vesicles containing the drug of interest.The size of the ethosomes is controlled using sonication and extrusion. 24

| Classic mechanical-dispersion method
In this method, a lipid mixture consisting of phospholipids and ethanol is first prepared.The drug or active ingredient is then added to the lipid mixture and the mixture is heated to form a clear solution.
The clear solution is then cooled and subjected to mechanical dispersion using a high-shear homogenizer or ultrasonicator to form small vesicles, typically in the range of 100-300 nm in diameter.This process involves subjecting the mixture to high shear stress to break up the lipid bilayers and promote the formation of ethosomes.
Finally, the resulting ethosome suspension is filtered through a membrane to remove any large particles or aggregates, and the size and shape of the ethosomes are confirmed using techniques such as dynamic light scattering (DLS) or transmission electron microscopy (TEM). 25,26

| Transmembrane pH-gradient method
In this method nonmedicated binary ethosomes is prepared initially followed by active loading of the drug.The phospholipid (e.g., phosphatidylcholine) is dissolved in an alcoholic phase consisting of ethanol and propylene glycol. 27A citrate buffer solution is TA B L E 1 Overview of ethosome preparation methods and their key characteristic.

Manufacturing scale
Hot method • Easy and quick method.
• High encapsulation efficiency.gradually added to the solution with continuous stirring at 700 rpm and 30 ± 1°C.The system is cooled to room temperature and the binary ethosomes are ready.Then the drug is actively loaded into the ethosomes and the system is continuously agitated at 700 rpm to effectively disperse and dissolve the drug.A pH gradient can be established between the outer (alkaline) phase and inner (acidic) phase of the ethosomal system by adding sodium hydroxide (NaOH) solution (0.5 M) to adjust the outer pH.The system is then incubated at an appropriate time and temperature to allow the unionized drugs to actively pass through the lipid bilayer of ethosomes and become entrapped in the vesicles. 28

| CHAR A C TE RIZ ATION OF E THOSOME S
Ethosomes can be visualized and evaluated to optimize their efficacy.Visualization of ethosomes can be achieved using TEM and scanning electron microscopy (SEM). 29These techniques allow the study of the microstructure of materials at high magnification.While the size and zeta potential of ethosomes can be measured using DLS, photon correlation spectroscopy (PCS) 30 and Zeta meter.The measurement of the zeta potential is crucial in determining the aggregation of vesicles and the stability of the vesicular system.Additionally, the entrapment efficiency of ethosomes can be determined using ultracentrifugation, a technique that separates particles based on their size, shape, and density.Different types of ultracentrifugation exist, including isopycnic centrifugation, Rate-zonal centrifugation, and density gradient centrifugation, each suited to different types of analysis. 31,32reover, the surface tension activity of ethosomes can be measured using the Du Nouy ring tensiometer, which measures surface and interfacial tension.This device uses the ring method, which involves immersing a small ring-shaped sample in the liquid at the interface and measuring the force required to pull the ring from the interface.The force is converted into a surface tension value by calibration.The ring is usually made of platinum, gold, or other metals. 33,34This method is useful for measuring the surface tension of liquids and the interfacial tension between two immiscible liquids, or liquid-gas or liquid-solid interface.
Furthermore, differential scanning calorimetry (DSC) can be used to determine the transition temperature of ethosomes in drug delivery. 35DSC measures the heat absorbed or released by a sample as a function of temperature or time.By measuring the heat absorbed or released at various temperatures, DSC provides information about the thermodynamic properties of the sample, such as the melting point, crystallization behavior, and phase transitions.In the case of ethosomes, DSC can be used to determine the transition temperature of the lipids used in the formulation, providing insight into the stability and performance of the ethosomes as a drug delivery system. 36timately, confocal laser scanning microscopy (CLSM) can be used to evaluate the depth of penetration of ethosomes into the skin or other tissue. 37This technology helps in understanding the efficacy and mechanism of the drug delivery system by analyzing the distribution of the vesicles.
Finally, to assess the stability of ethosomes, the preparations can be stored at different temperatures (25 ± 2°C, 37 ± 2°C and 45 ± 2°C) for different periods of time. 38Changes in the size and shape of the vesicles can be measured using DLS and TEM to evaluate the ability of ethosomes to sustain the drug over time and under different conditions. 39e flexibility of ethosomal formulations can be evaluated using the extrusion method, while the clarity of the preparation can be measured using a nephelometer.These methods help to determine the degree of deformability and turbidity of the ethosomal preparation, respectively, and evaluate the performance of the formulation. 40

| MECHANIS MS OF E TH SOME S S K IN DELIVERY
Ethosomes provide alternative pathways for drugs to reach the underlying tissues.The unique composition of ethosomes can enhance the penetration of drugs through the skin, using various pathways leading to improved efficacy. 41The pathway used depends on the composition and formulation of the ethosomes, as well as the properties of the drug being delivered.Ethosomes can interact with the skin's natural lipids, 42 or fuse (vesicle fusion) with the skin cells, 43 or penetrate transfollicular through the hair follicles, 44,45 or pass transcellular across the cells. 46Additionally, the presence of ethanol and phospholipids in ethosomes can increase the fluidity and permeability of the stratum corneum based on the interfacial activity 47,48 ; Ethanol effect is the first step for ethosomes penetration, in which ethanol changes the conformation of stratum corneum lipid which are extremely compacted.Ethanol interacts with the hydrophilic (polar) heads of the lipid layers which results in increasing lipid fluidity and decrease the consistency of the lipid layers, thus providing the vesicles with flexibility and enabling them to penetrate to deeper layers. 49Ethosomes can penetrate the skin via both the percutaneous pathway of the hair follicle and stratum corneum, and the breaking up of the vesicles can allow the drug to continue to permeate into the deeper layers of the skin.The superior percutaneous penetration of ethosomes is attributed to the synergistic effects of their ingredients, including ethanol and phospholipids. 49

| E THOSOME S APPLI C ATI ON S IN COS ME TI C DELIVERY
Ethosomes have gained considerable attention as a potential drug delivery system for various cosmetic applications, including hair loss, acnes, and skin whitening.

| Ethosomes for hair loss
Hair loss could be contributed to various potential causes.This includes genetic factors, hormonal changes, certain medical conditions, and diseases like thyroid disorders, autoimmune conditions such as alopecia areata, scalp conditions (such as psoriasis or seborrheic dermatitis).Additionally, some medications like chemotherapy drugs, antidepressants, and blood thinners may cause hair loss or treatments like radiation therapy or some cosmetic procedures may cause temporary or permanent hair loss in the treated areas.Moreover, physical, emotional, nutritional deficiencies or aging may lead to hair loss.
Ethosomes have been investigated as a potential treatment for hair loss by delivering drugs such as minoxidil and finasteride directly to the hair follicles.The enhanced penetration of the skin by ethosomes can improve the efficacy of these drugs and promote hair growth. 50The drug molecules can penetrate the hair shaft and reach the target site more efficiently, promoting hair growth.Studies have shown that the use of minoxidil and finasteride in ethosomes formulations significantly improves the efficacy of these drugs compared to traditional creams and solutions.However, more research is needed to confirm the safety and efficacy of ethosomes as a hair loss treatment.
Minoxidil is the only FDA approved medication to treat androgenic alopecia.It is available as a topical solution and sold under different brand names like Rogaine® and many others. 51It works by increasing blood flow to the hair follicles, which in turn stimulates hair growth.However, minoxidil is known to have poor skin penetration, which limits its effectiveness.By incorporating minoxidil into ethosomes, the drug can be delivered directly to the hair follicles, leading to improved efficacy and increased hair growth.Ethosomes enhance the penetration of minoxidil to reach pilo sebaceous follicle. 52Minoxidil ethosomes is not a drug that is currently available in the market. 53However, more research is needed to confirm the safety and efficacy of minoxidil ethosomes, and to establish its ability to get FDA approval.
Finasteride is a drug that is commonly used to treat male pattern baldness.It is an oral medication that is approved by FDA for the treatment of hair loss, and it is sold under the brand name Propecia.
It works by inhibiting the action of an enzyme called 5-alpha reductase, which is responsible for converting testosterone into dihydrotestosterone which is the hormone that causes hair loss. 54By incorporating finasteride into ethosomes, the drug can be delivered directly to the hair follicles, leading to improved efficacy and decreased hair loss.Currently, finasteride ethosomes is not a drug that is available in the market.There have been some studies that have investigated the potential of finasteride ethosomes as a treatment for hair loss, but these studies have been done in laboratory and preclinical level, and it is not yet FDA approved. 55ltidrug ethosomes are a promising area of research in drug delivery systems for hair loss treatments.Wang et al. (2022) 56 demonstrated the potential benefits of combining two or more drugs in a single vesicular formulation.By targeting multiple mechanisms of hair loss, multidrug ethosomes could improve the efficacy of hair loss treatments.However, the development of drug formulations and their effectiveness in treating hair loss is an ongoing and complex area of research.Therefore, further studies are needed to assess the safety and efficacy of multidrug ethosomes in hair loss treatment.

| Ethosomes for acnes
Acne is a common skin condition that occurs when hair follicles become clogged with oil and dead skin cells.The development of acne is a complex process that involves several factors, including hormones, bacteria, inflammation, and genetics.Other factors such as stress, diet, and certain medications can also contribute to the development of acne.Bacteria play a role in the development of acne.
Propionibacterium acnes (P.acnes) is a type of bacteria that lives on the skin and is normally harmless.However, when hair follicles become clogged with oil, P. acnes can multiply and cause inflammation of sebaceous follicle, leading to the formation of pimples. 57,58It was found that by loading 1%-3% of bacitracin (Gram positive bactericidal agent) in ethosome using 25% ethanol, acne caused by P.acnes was cured.It was found that bacitracin loaded in ethosomes was effectively delivered to the site of the infection, curing acne and preventing it from occurring. 59 addition, ethosomes have been explored as a potential treatment for acnes by delivering drugs such as retinoids and benzoyl peroxide directly to the affected area. 60The enhanced penetration of the skin by ethosomes can improve the efficacy of these drugs and reduce the formation of new acnes.
Tretinoin is a medication that is approved by FDA for the treatment of acne and fine wrinkles, sun-damaged skin, and dark spots.It is a retinoid, which means that it is a derivative of vitamin A. It works by increasing cell turnover and reducing the formation of comedones (clogged pores) that reduce the formation of blackheads and whiteheads.Tretinoin is usually used in topical form under the brand name Retin-A. 61However, tretinoin can be irritating to the skin, which limits its effectiveness.By incorporating tretinoin into ethosomes, the drug can be delivered directly to the sebaceous glands, where it can target the underlying causes of acne, leading to improved efficacy and reduced side effects.Currently, tretinoin ethosomes is not a drug that is available in the market.While some studies have investigated the potential of tretinoin ethosomes as a treatment for various skin conditions such as acne and photoaging, more research is needed to confirm the safety and efficacy of Tretinoin ethosomes and to establish its ability to get FDA approval. 62reover, benzoyl peroxide is an antimicrobial agent that is commonly used to treat acne.Benzoyl peroxide is usually used in topical form and it is available over the counter under different brand names like Clearasil®, Oxy® and many others. 63It works by killing the P.
acnes bacteria.By incorporating benzoyl peroxide into ethosomes, the drug can be delivered directly to the sebaceous glands, leading to improved efficacy and reduced side effects.While some studies have investigated the potential of benzoyl peroxide ethosomes as a treatment for acne, more research is needed to confirm their safety and efficacy. 64rthermore, multidrug ethosomes can potentially enhance the efficacy of acne treatments and may be especially useful for patients with moderate to severe acne. 65For example, a combination of tretinoin and benzoyl peroxide ethosomes can address multiple aspects of the acne pathogenesis. 66Tretinoin helps to regulate the abnormal shedding of skin cells that can clog pores and contribute to acne formation, while benzoyl peroxide helps to kill the bacteria that contribute to the development of acne lesions.Additionally, the use of ethosomes can help to increase the delivery of these medications into the skin, thereby enhancing their efficacy.However, the development of drug formulations and their efficacy in treating acne is a complex and ongoing area of research.Further studies are needed to determine the safety and efficacy of multidrug ethosomes specifically in acne treatment, and it is important to consult with a healthcare professional before starting any new treatment.

| Ethosomes for skin whitening
Hyperpigmentation is a condition in which areas of the skin become darker than the surrounding skin.This can be caused by an increase in the production of melanin, the pigment that gives color to our skin, hair, and eyes.The mechanism behind hyperpigmentation involves the activation of melanocytes, which are specialized skin cells that produce melanin.Melanocytes can be stimulated by various factors such as exposure to sunlight, inflammation, hormonal changes, and certain medications.When stimulated, melanocytes produce more melanin, which is then transferred to surrounding skin cells to provide protection against further damage from UV radiation or other harmful stimuli.Hyperpigmentation can also be caused by the accumulation of melanin in the upper layers of the skin.This can occur due to impaired shedding of dead skin cells, which can lead to the buildup of pigmented cells in the skin.While hyperpigmentation is usually harmless, it can have a significant impact on a person's appearance and self-esteem.
Ethosomes have been investigated as a potential treatment for skin hyperpigmentation by delivering drugs directly to the affected area.The enhanced penetration of the skin by ethosomes can improve the efficacy of these drugs and reduce the appearance of dark spots and uneven skin tone. 67Different drugs were delivered by ethosomes applications in skin whitening including hydroquinone, 68,69 kojic acid 70,71 and vitamin C. 72 Hydroquinone is a skin-lightening agent that is commonly used to treat hyperpigmentation.It works by inhibiting the production of melanin, the pigment that gives color to the skin.Hydroquinone is a medication that is approved by FDA for the treatment of hyperpigmentation (dark spots) caused by sun damage, acne, or other skin conditions.Hydroquinone is usually used in topical form and it is sold under different brand names like Melamin®, Nuquin® and many others. 68By incorporating hydroquinone into ethosomes, the drug can be delivered directly to the melanocytes, the cells responsible for producing melanin, leading to improved efficacy and reduced side effects.Hydroquinone ethosomes is not a drug that is available in the market.While some studies have investigated the potential of hydroquinone ethosomes as a treatment for skin whitening, more research is needed to confirm the safety and efficacy of hydroquinone ethosomes. 69 addition, Kojic acid is a naturally occurring skin-lightening agent that is commonly used to treat hyperpigmentation.It is commonly used in topical form as a skin lightening agent in many over-the-counter and prescription products.It is sold under different brand names like Kojic acid cream, serum, and many others. 70jic acid is derived from various fungus, including Aspergillus parasiticus, Aspergillus oryzae, Asperagillus falvus, and Asperagillus tamari.
However, the most used fungal strain for commercial production of kojic is Aspergillus oryzae.This fungus is a widely used as well in the food and beverage industry for fermentation of soybeans, rice, and other products.Kojic works by inhibiting the production of melanin, the pigment that gives color to the skin.It is used to treat hyperpigmentation caused by sun damage, acne, or other skin conditions.By incorporating kojic acid into ethosomes, the drug can be delivered directly to the melanocytes, leading to improved efficacy and reduced side effects.While some studies have investigated the potential of kojic acid ethosomes as a treatment for skin whitening and hyperpigmentation, more research is needed to confirm the safety and efficacy of Kojic acid ethosomes and to establish its ability to get FDA approval. 71reover, vitamin C is an antioxidant that is commonly used to treat hyperpigmentation.Vitamin C is a powerful antioxidant that helps to protect the skin from environmental damage, boost collagen production, and even out skin tone.Vitamin C is also available in different concentrations and different brand names like Obagi®, Skinceuticals®, and many others. 72Vitamin C is commonly used as a skin-lightening agent and in many skincare products like serums, creams, and lotions. 73It is found in different forms like L-ascorbic acid, magnesium ascorbyl phosphate, and ascorbyl palmitate.It works by inhibiting the production of melanin, stimulating the collagen to improve skin texture and protecting the skin from sun damage.By incorporating Vitamin C into ethosomes, the drug can be delivered directly to the skin, leading to improved efficacy and reduced side effects.Vitamin C ethosomes is not a drug that is available in the market, but there are some studies that have investigated the potential of Vitamin C ethosomes in the field of skin care and anti-aging. 74

| ADVANTAG E S OF E THOSOME S DELIVERY
Ethosomes have several advantages as a drug delivery system.They can be used for both topical and transdermal drug delivery. 75They are biocompatible, biodegradable, 76 and non-toxic making them a safe and effective drug delivery system. 77,78In addition, they can carry a wide range of drugs, including hydrophilic and hydrophobic compounds, making them a versatile drug delivery system. 79,80They have a high drug-loading capacity, which means that they can carry a large amount of drug per vesicle, that improves the overall efficacy of the treatment and reduces the number of applications needed. 81reover, the ethanol present in ethosomes can help stabilize the vesicles and protect them from degradation.This improves the shelf life of the drug and reduces the need for frequent reapplication. 82rthermore, ethosomes increased the drug penetration through the skin by increasing the fluidity and permeability of the cell membrane that allows for a higher amount of drug to reach the underlying tissues and improve the overall efficacy of the treatment. 83timately, ethosomes can be engineered to target specific tissues or cells, improving the overall specificity of the treatment. 84

| LIMITATI ON S OF E THOSOME S DELIVERY
While ethosomes offer several advantages as a drug delivery system, there are also several limitations that must be considered.
Some of the key limitations include complex preparation and timeconsuming, which can make the production process less efficient, 85 limited stability due to sensitivity to temperature and humidity, that can make it difficult to store and transport the vesicles. 86In addition, there are limited in vivo studies that make it difficult to fully understand the potential benefits and risks of using ethosomes as a drug delivery system. 87Moreover, they can cause skin irritation and erythema if they are not optimized, 88 and show some incompatibility with certain drugs. 89The interference may have physical or chemical or therapeutic effects.This may alter the drug solubility, stability, or bioavailability.For example, the encapsulation of a hydrophilic drug within a lipid bilayer may reduce its solubility, making it less available for delivery, or may not be compatible with the lipid-based nature of ethosomes and may exhibit reduced efficacy, or the encapsulation of a drug within an ethosome may alter its stability, leading to the formation of inactive metabolites or toxic byproducts.Furthermore, they are sensitive to pH and temperature changes, 90 and may have potential interactions with other excipients and drugs in the formulation. 91,92However, many of these limitations can be overcome by optimizing the composition and preparation of the ethosomes and using appropriate formulations. 93It is important to note that further research is needed to fully understand the potential benefits and limitations of ethosomes as a drug delivery system.

| FUTURE OF E THOSOME S
The potential use of ethosomes in skin care delivery is not limited to the mentioned examples.They have also been investigated for the delivery of other cosmetic actives, such as hyaluronic acid, collagen, and peptides. 94These ingredients have shown great promise in improving skin hydration, firmness, and elasticity, and their delivery using ethosomes may enhance their efficacy.
The use of ethosomes in skin cosmetic delivery is still in its early stages, and there are several challenges that need to be addressed, such as stability, shelf-life, and skin irritation.However, the potential benefits of using ethosomes as a cosmetic delivery system are significant, and further research is needed to fully explore their potential in this field.

| CON CLUS ION
In conclusion, Ethosomes have shown promising results as drug delivery systems in the pharmaceutical industry, and their potential for skin care applications is increasingly being explored.The unique structure and composition of ethosomes make them suitable for the delivery of active cosmetic ingredients for the treatment of hair loss, acne, and skin whitening.such as vitamins, antioxidants, and skinlightening agents, through the skin.Their ability to enhance the penetration of drugs through the skin makes them a versatile and effective alternative to traditional transdermal drug delivery systems.The use of ethanol in their composition allows for increased stability, enhanced drug-loading capacity, and targeted drug delivery.However, the preparation of ethosomes can be complex and time-consuming and their stability can be affected by temperature and humidity.
However, further research is needed to fully understand the potential benefits and limitations of using ethosomes in these indications, and to optimize the formulations and methods of administration.

AUTH O R CO NTR I B UTI O N S
Rana Abu-Huwaij conceptualization; Abdullah N. Zidan wrote the original draft; Rana Abu-Huwaij edited, and reviewed the manuscript.

CO N FLI C T O F I NTE R E S T S TATE M E NT
The authors declare no conflict of interest.

DATA AVA I L A B I L I T Y S TAT E M E N T
Data sharing not applicable to this article as no datasets were generated or analysed during the current study.

E TH I C S S TATEM ENT
Not applicable.