INTRODUCTION:

The use of chemicals enhancers such as surfactants and organic solvent induce irritation, cause damage, and reduce skin barrier function, therefore, it is desirable to deliver the therapeutic agents that maintain the normal skin barrier function without the aid of a chemical enhancer. One such approach is use of vesicular system.

Figure. 1 structure of skin

In the past decade, topical drug delivery by liposomal formulation has evoked considerable interest. Deformation liposomes andtransferosomes were first generation of elastic vesicles introduced by ceve and blume in 1992 and were reported to penetrate intact skin while carrying a therapeutic concentration of drug when applied under non-occluded condition.^[1]

Drug, encapsulated in lipid vesicles, prepared from phospholipids and non-ionic surfactants is known to be transported into across the skin. Vesicles can be used for encapsulating hydrophilic and lipophilic as well as low and high molecular weight drugs.

Figure.2 Structure of ethosomes

Therefore, these lipid rich vesicles are hypothesized to carry a significant quantity of drugs across the skin, thus enhancing the systemic absorption of drugs. Classic liposomes are little or of no value as carrier for drug delivery because they do not penetrate skin deeply, but rather remain confined to upper layer of stratum corneum. It has been investigated and reported that vesicular systems embodying ethanol in relatively high concentration called ethosomes, are very efficient at enhancing the skin permeation of a number of drugs.^[1,2]

Ethosomes:

Figure. 3 Structure of drug entrapment in lipid core in ethosomes

Ethosomes are ethanolic liposomes ^[4]

Ethosomes were developed by Touitou, 1997 as additional novel lipid carriers composed of ethanol, phospholipids and water. They are reported to improve the skin delivery of various drugs. ^[1,7]

Alcohol (ethanol and isopropyl alcohol) is efficient permeation enhancer that is believed to act by affecting the intercellular region of the stratum corneum. Ethosomes are soft, malleable vesicles composed of mainly phospholipids, water, ethanol (relatively at higher concentration).^[11]The “soft vesicles” represent novel vesicles carrier for enhanced delivery through the skin.^[3]The vesicles have been well known for their importance in cellular communication particle transportation from many years.Vesicles would also allow controlling the release rate of drug over an extended time, keeping the drug shielded from immune response or other removal systems and thus be able to release just the right amount of drug and keep that concentration constant for longer periods of time.^[1]

Ethosomes are non-invasive delivery carrier that enable drugs to reach the deep skin layers and/or systemic circulation. Size of ethosomes vesicles can be modulated from tens of nanometer to microns(30nm to few microns). High concentration of ethanol makes ethosomes unique, as ethanol is known for its disturbance of skin lipid bilayer organization. Therefore when integrated into a vesicles the ability to penetrate the stratum corneum. Because of high concentration, lipid membrane is less tightly packed than the conventional vesicles, although it has equivalent stability and improves the drug distribution ability in the stratum corneum lipids.^[3]

Ethosomes permeate through the skin layers more rapid and possess significantly higher transdermal flux in comparison to conventional liposomes.Ethosomes have also been prepared by adding penetration enhancer propylene glycol and shows the penetration efficiency. Penetration enhancers, however, the main mode of action is a perturbation of the lipid organization in the stratum corneum, thereby increasing the transport rate across the skin.The presence of edge-activator agent (i.e ethanol and sodium cholate) in the lipid bilayer noticeably improves the carrier penetration through the stratum corneum allowing an efficacious local and systemic delivery of both hydrophilic and hydrophobic compounds.^[1]

Ethosome composition:

Ethosomes are vesicular carrier comprise of hydroalcoholic orhydro/alcoholic/glycolic phospholipid in which the concentration of alcohols or their combination is relatively high.^[13]Typically, Ethosomes may contain phospholipids with various chemical structures like phosphatidylcholine (PC), hydrogenated PC, phosphatidic acid (PA), phosphatidylserine (PS), Phosphatidylethanolamine (PE), phosphatidylglycerol (PPG), phosphatidylinositol (PI), hydrogenated PC, alcohol (ethanol or isopropyl alcohol), water and propylene glycol (or other glycols). Such a composition enables delivery of highconcentration of active ingredients through skin. ^[4,5]

Drug delivery can be modulated by altering alcohol: water or alcohol-polyol: water ratio. Some preferred phospholipids are soya phospholipids such as Phospholipon 90 (PL-90). It is usually employed in a range of 0.5-10% w/w. Cholesterol at concentrations ranging between 0.1 1% can also be added to the preparation of alcohols, which can be used, including Ethanol and Isopropyl Alcohol.^[12]

Among Glycols, Propylene Glycol and Transcutol are generally used. In addition, non-ionicsurfactants (PEG-alkyl ethers) can be combined with the phospholipids in these preparations. Cationic lipids like cocoamide, POE alkyl amines, dodecylamine, cetrimide etc can be added too. The concentration of alcohol in the final product may range from 20 to 50%. The concentration of the non-aqueous phase (alcohol and glycol combination) may range between 22 to 70%.^[6]

Table-1 Different Additive Employed In Formulation of Ethosomes:

Class	Example	Uses
Phospholipids	Soya phosphatidyl choline Egg phosphatidyl choline Dipalmitylphosphatidyl choline Distearylphosphatidyl choline	Vesicles forming component
Polyglycol	Propylene glycol	As a skin penetration enhancer
Alcohol	Ethanol Isopropyl alcohol	For providing the softness for vesicle membrane As a penetration enhancer
Cholesterol	Cholesterol	For providing the stability to vesicle membrane
Dye	Rhodamine-123 Rhodamine red FluoresceneIsothiocynate(FITC) 6- Carboxy fluorescence	For characterization study
Vehicle	CarbopolD934, HPMC	As a gel former

Ethosomes carrier for dermal and transdermal drug delivery:

These were reported to be effective at delivering molecules to and through the skin to the systemic circulation. Ethosomal 5% acyclovir system vs 5% acyclovir cream (zovirax, zc) for topical treatment for herpetic infection. Observation was made by using fluorescent probe D-289 to study skin penetration. Authors concluded that the classic liposomes did not facilitate probe penetration into skin rather resulted in small reservoir in the upper layers of skin. Using hydroethanolic solutions, a relatively deep penetration, but of relatively very low fluorescent activity was observed. Use of ethosomal system resulted in increase in both depth and fluorescent activity. Ethosomes have also been reported to improve invivo and invitro skin delivery of many drugs under occlusive and non-occlusive condition.^[1]

Mechanism of action:

It is thought that first part of the mechanism is due to the ethanol effect, where ethanol interacts with the lipid molecules in the polar head group region resulting in a reduction in the transition temperature of the lipids in the stratum corneum, increasing their fluidity and decreasing the density of the lipid multilayer. This is followed by the ‘ethosomal effect’, which includes lipid penetration and permeation by the opening of new pathways, due to a malleability and fusion of ethosomes with skin lipids, resulting in the release of a drug into the deep layers of the skin.Ethanol may also provide vesicles with soft flexible characteristics, which allow them to penetrate more easily into the deeper layer of the skin.^[4,8]

Stability:

Liposomes apparently having the cholesterol effect which cause liposomes to be less permeable membrane by filling holes or disruption. Niosomes are similar to that of liposomes but niosomes are preferred over liposomes due to chemical stability and economy. Pharmacosomes are preferred over liposomes andniosomes as a unique advantage having potential alternative to convectional vesicles.^[10]However, due to poor skin permeability, liposomes and niosomes could not be successfully used for systemic drug delivery and their use was limited for topical use. To overcome problems of poor skin permeability Ceve et al. and Touitou et al. recently introduced two new vesicular carrier systems transfersomes and ethosomes, respectively for non-invasive delivery of drugs into or across the skin.^[14]Transfersomes and ethosomes incorporated edge activators (surfactants) and penetration enhancers (alcohols and polyols), respectively, to influence the properties of vesicles and stratum corneum.^[1]

Liposome fuse and forms bigger vesicles and drugs leakage from the vesicles and there was a breakage in the liposomes during storage which leads to instability. Ethosomes causes a modification of net charge of the system and confers some degree of steric stabilization leading to increase stability of dispersion against agglomeration. Increase in ethanol to 15-45% increases entrapment efficiency owing to increase n fluidity of membrane. Further increase in ethanol >45% makes a membrane more leaky thus there is a decrease in entrapment causes destabilization. Ethosomes bearing melatonin offered a suitable approach for transdermal delivery when compared to liposomes andhydroethanolic solution Increase in thermodynamic activity due to evaporation of ethanol, increases penetration of drug molecule due to reduction in barrier property of stratum corneum by ethanol.^[1,7]

Advantages:^[^3,5]

1. Ethosomes enhance permeation of the drug through skin transdermal and dermal delivery.

2. Ethosomes are platforms for the delivery of large and diverse groups of drugs (peptides, protein molecules)

3. Ethosomal systems are much more efficient at delivering a fluorescent probe (quantum dots) to the skin in terms of quantity and depth.

4. Low risk profile- the technology has no large scale drug development risk, as the toxicological profiles of the ethosomes components are well documented in the scientific literature.

5. High patient compliance- the ethosomes drugs are administered in a semisolid form (gel/cream), producing high patient compliance. In contrast, iontophoresisandphonophoresis are relatively complicated to use, which will affect patient compliance.

6. High market attractiveness for products with proprietary technology. Relatively simple to manufacture with no complicated technical investments required for the production of ethosomes.

7. The ethosomes system is passive, non-passive and available for immediate commercialization.

Future perspective:^[^1,2]

1. Introduction of ethosomes has intiated new area in transdermal drug delivery

2. Further research in this area will allow better control over drug release in vivo allowing physicians to make therapy more efficient.

3. It offers good opportunity for non-invasive delivery of small-, medium- and large-sized drug molecules.

4. Special emphasis is given to skin delivery of proteins and other macromolecules and for transcutaneous immunization.

Table 2 Characterization techniques:

Parameters	Methods
Vesicle shape (morphology)	Transmission electron microscopy Scanning electron microscopy
Entrapment efficiency	Mini column centrifugation method Fluorescence spectrophotometry
Vesicle size and size distribution	Dynamic light scattering method
Vesicle Skin interaction study	Confocal laser scanning microscopy Fluorescence microscopy Transmission electron microscopy Eosin-Hematoxylin staining
Phospholipid-ethanol interaction	31P NMR Differential scanning calorimeter
Degree of deformability	Extrusion method
Zeta potential	Zeta meter
Turbidity	Nephalometer
In vitro drug release study	Franz diffusion cell with artificial or biological membrane, Dialysis bag diffusion
Drug deposition study	Franz diffusion cell
Stability study	Dynamic light scattering method Transmission electron microscopy

Application of Ethosomes:

Therapeutic application:^[^1,5]

a. Ethosomes for transcutaneous immunization, and antigen-loaded ethosomes for transcutaneous immunization against hepatitis B were prepared and which shows the greater entrapment efficiency compared to conventional liposomes.Ethosomes are also used in pilosabeceous targeting.

b. Transcutaneous delivery potential of antigen-loaded antigen system, demonstrated much higher skin permeation of the antigen in comparison to the conventional liposomes.

c. Oral administration of hormones is associated with completion like first pass metabolism, low bioavailability and several dose dependent side effects. Testosterone ethosomes across rabbit pinna skn with marketed transdermal patch of testosterone shows 30 times higher skin permeation compared to other formulation.

d. Encapsulation of CBD (cannabidiol) in ethosomes significantly increased its skin permeation, accumulation and hence its biological activity.

e. Psychoactive drug THP compared its delivery with that of classical liposomal formulation for the treatment of Parkinson’s disease observed that 4.5% times higher in action than that of liposomes.

f. Cyclosporine Aethosomal formulation for treatment of inflammatory skin disease like psoriasis, atopic dermatitis and disease of hair follicle like alopecia areata etc.

g. Ethosomes formulation of antibiotic could be high efficient and would overcome the problem with convectional therapy.

Cosmetic application:^[^1]

Above of apply ethosomes in cosmoceuticals is not only to increase stability of cosmetic chemicals but also to decrease skin irritation. Composition and size are main factors to be considered to obtain this advantage of elastic vesicles for cosmoceuticals.

Topical administration of many antioxidants leads to diminish oxidative injury in the skin so ‘anti-oxidant ethosomes’ for topical delivery utilizing the synergistic properties of vitamin A, vitamin E, vitamin C.Vitamin A and vitamin E in the lipid bilayer and vitamin C in the aqueous core complete protection of ethosome formulation.

Table: 3 Application of Ethosomes as a Drug Carrier:

Drug

Results

NSAIDS (Diclofenac)

Selective delivery of drug to desired side for prolong period of time

Acyclovir

Increase skin permeation, Improved in biological activity two to three times,

Improved in Pharmacodynamic profile

Insulin

Significant decrease in blood glucose level,

Provide control release

TrihexyphenidylHydrochloride

Improved transdermal flux,

Provide controlled release

Improved patient compliance,

Biologically active at dose several times lower than the currently used formulation.

DNA

Better expression of genes,

Selective targeting to dermal cells

Antibiotic, Cannabidol, Erythromycin

Improved skin deposition,

Improved biological activity,

Prolonging drug action

Bacitracin

Improved dermal deposition,

Improved intracellular delivery

Increased bioavailability

Anti-HIV agents

Zidovudine

Lamivudine

Improved transdermal flux,

Improved in biological activity two to three times,

Prolonging drug action,

Affected the normal histology of skin

Azelaic acid

Prolong drug release

CONCLUSION:

Ethosomes carrier opens new challenge and opportunities for the development of novel improved therapies. Transdermal route is promising alternative to drug delivery for systemic effect. Ethosomes have initiated a new area in vesicular research for transdermal drug delivery which can provide better skin permeation than liposomes. The main limiting factor of transdermal drug delivery system i.e. epidermal barrier can be overcome by ethosomes to significant extent. Application of ethosomes provides the advantages such as improved permeation through skin and targeting to deeper skin layers for various skin diseases. Ethosomes have been tested to encapsulate hydrophilic drugs, cationic drugs, proteins and peptides. Further, research in this area will allow better control over drug release in vivo and long-term safety data, allowing the therapy more effective. Thus, ethosomal formulations possess promising future in effective dermal/transdermal delivery of bioactive agents.

REFERENCES:

1. Poonam Varma and K. Pathak, Therapeutic and cosmeceutical potential of ethosomes: An overview, Journal of Advanced Pharmaceutical and Research,V-1(3); Jul-Sep, 2010, PMC3255417.

2. Nirali Dave, Sameer Sheaikh. Ethosomes: A Novel Approach for Transdermal Drug Delivery, research article, International Journal of Current Research Vol. 5, Issue, 04, pp.823-827, April, 2013 ISSN: 0975-833X.

3. Hitesh Jain, Jitendra Patel, Kruti Joshi, Parth Patel and U M Upadhyaye. Ethosomes: A novel drug carrier pharmacieglobale, International journal of comprehensive pharmacy, review article, ISSN0976-8157.

4. Abhishek Chandel, Vishal Patil, Rohit Goyal, Hitesh Dhamija and Bharat Parashar. Ethosomes: A Novel Approach towards Transdermal Drug Delivery, International journal of pharmaceutical and chemical sciences, review article, ISSN:22775005.

5. Vivek Dave, Ashutosh Pareek, Sarvesh Paliwal. Ethosome: A Novel Approach of Transdermal Drug Delivery System, International Journal of Advanced Research in Pharmaceutical and Bio sciences, ISSN2277 – 6222.

6. Tarun Parashar, Soniya, Roopesh Sachan, Vishal Singh, Gaurav Singh, Satyanand Tyagi, Chirag Patel, Anil Gupta, Ethosomes: A recent vesicle of transdermal drug delivery system, International Journal of Research and Development in Pharmacy and Life Sciences, review article, February - March, 2013, Vol. 2, No.2, pp 285-292.

7. R Rakesh, KR Anoop, Ethosomes for Transdermal and Topical Drug Delivery, International Journal of Pharmacy and Pharmaceutical Sciences Vol 4, Suppl 3, 2012.

8. D. Akiladevi, Sachinandan Basak, Ethosomes A Noninvasive Approach for Transdermal drug delivery system, review article, International Journal of Current Pharmaceutical Research, Vol 2, Issue 4, 2010.

9. D. Ainbinder, D. Paolino, M. Fresta, and E. Touitou, Drug Delivery Applications with Ethosomes Journal of Biomedical Nanotechnology, Vol. 6, 558–568, 2010.

10. Saurabh Bansal, Chandan Prasad Kashyap, Geeta Aggarwal and SL Harikumar, A comparative review on vesicular drug delivery system and stability isssues, International journal of research in pharmacy and chemistry review article.

11. Anujdhiman, Deepika Singh, Manjubala, Kavita Sharma, potencial phytotherapeutic agents in design of ethosomes: a review, Journal of Pharmaceutical and Scientific Innovation. Sept-Oct 2012, 26-30.

12. Prasanthi. D, P.K. Lakshmi, Vesicles-Mechanism of Transermal permeation: A Review, Asian Journal of Pharmaceutical and Clinical Research ,Vol 5, Issue 1, 2012 Vol. 4, Issue 3.

13. Sunil Kamboj, Vipin Saini, Nancy Magon, Suman Bala1, Vikas Jhawat, Vesicular drug delivery systems: A novel approach for drug targeting review article, International Journal of Drug Delivery 5 (2013) 121-130.

14. Nikalje Anna Pratima, Tiwari Shailee Ethosomes: A Novel Tool for Transdermal Drug Delivery, review article, International Journal of Research in Pharmacy and Science.

Received on 22.02.2015 Accepted on 05.03.2015

Research J. Topical and Cosmetic Sci. 6(1):Jan.–June 2015 page 7-14

DOI: 10.5958/2321-5844.2015.00002.3