J. Pharm. Pharmacol. 1999, 51: 1123±1134 # 1999 J. Pharm. Pharmacol.
Skin Delivery of Oestradiol from Deformable and Traditional
Liposomes: Mechanistic Studies
GAMAL M. M. EL MAGHRABY, ADRIAN C. WILLIAMS AND BRIAN W. BARRY
Drug Delivery Group, Postgraduate Studies in Pharmaceutical Technology, The School of Pharmacy,
University of Bradford, Bradford, BD7 1DP, UK
The drug finds it difficult to cross the barrier of skin provided by stratum corneum. The lipid vesicles have to cross this barrier which it finds difficult to overcome. Hence, most of the effect of these vesicles 1.e Liposomes is found in outer skin layers like epidermis and dermis. A new type of vesicle called transferosome is found to penetrate the intact skin, thus, giving transdermal effect. In vitro skin studies using both conventional liposomes and transferosomes is investigated in this study. Oestradiol was used as a model drug to investigate the mechanism to optimise its delivery via skin. Five mechanisms of these vesicles were investigated which are free drug mechanism, penetration enhancement, drug uptake by skin, intact vesicle permeability and . in drug free mechanism, the amount of free drug permeating the skin is calculated. Pretreating the skin with empty vesicles helped to study the penetration enhancement. Dipping stratum corneum in different formulations of liposome helped to determine the drug uptake. It was seen that vesicles of the size 200-300 nm can penetrate through the skin. The vesicles bigger than 500 is believed not to permeate through the skin. Different edge activators or surfactants like tween, span and sodium cholate are used to prepare transferosomes. Cholesterol is mixed in conventional liposomes as it is found to stabilise the vesicles
The vesicle were prepared using hand- shaking method. In this the lipid mixture was dissolved in ethanol or 2:1 ethanol : chloroform ration which were the organic solvents. These organic solvents dissolved the cholesterol and lipid. Oestradiol, the model drug was then added to this mixture which was previously radiolabelled. The mixture was then kept in rotary evaporatot. After complete evaporation of the organic phase, the thin film was hydrated with water and ethanol in 7% v/v. the vesicles were then then allowed to get swollen.
Smaller vesicles were prepared using bath sonicator at room temperaturefor 30mins. All the vesicle sizes were homogenized using 10 times extrusion using 200 and 100nm polycarbonate membranes.
Determination of entrapment efficiency
The entrapped and unentrapped drug were separated using mini column centrifuge. Sephadex gel was formed by soking Sephadex for 5h in wter. Whattman pads were used tas filter in bottom of the barrels. Centrifugation was done at 3000 rev per min for 3 minutes. Liposomes were recovered from the first and second stage of centrifugation.
Determination of drug release
The amount of free drug released by mini- column centrifugation was calculated indirectly from the amount of drug entrapped. The amont of drug entrapped helps to calculate the unentrapped amount. Unentrapped drug was calculated by considering the amount at zero time as initial amount.
Determination of vesicle size
Sizing was done using zeta sizer (photon correlation spectroscopy). Here the samples prepared in distilled water was filtered through 2µm membranes
Preparation of human skin membranes
Mid-line Caucasian skin samples were used for this purpose. These samples were stored at -20°C in vacuum. Heat separation technique was used to prepare epidermal membrane. The skin was heated at 60°C for 45 minutes in water bath which was then peeled off the underlying dermis. Stratum corneum was also prepared.
Results and discussion
This is expressed as percent entrapment of entrapped drug. The lipophilic drug estrodiol showed high entrapment efficiency. Phospholipid concentration, cholesterol and surfactant affected this entrapment efficiency. 99% was the maximum entrapment efficiency achieved.
The release of progesterone and steroid was found to be negligible for about 60h in DPPC liposomes. Steroid release was checked for diffusion lipid composition. It was found that in the formulation’s stable period negligible release was noted. In conventional liposome, 6% drug was released in 48 hours. Stabiliser like cholesterol was found to decrease drug release.
Use of deformable vesicles using surfactant showed sigmoidal shapes. The leaks were found to be greater due to leaky membranes. It was concluded that the lipid composition determines the entrapment of oestrodiol. Cholesterol decreased drug release whereas surfactants increased the drug release.
The average size of unilamellar vesicles was found to be 127 to 146nm irrespective of the liposome formulation.. the result is after sonication or manual extrusion. The sizes of multilamellar vesicles were dependant on the formulation. Vesicles with cholesterol showed greater size while with surfactants showed smaller size.
Delivery through skin
Deformable vesicles were found to be giving optimal delivery via skin. Data analysis were plotted of amount permeated against time. Deformable vesicles with cholate delivered the maximum amount of drug. The traditional liposomes only showed superficial drug delivery. deformable vesicles increased the skin permeation more than the partitioning of the compound.
In this experiment it was concluded that oestradiol delivery through vesicle is better than saturated aqueous solution. Deformable vesicles are found to be more efficient fir drug delivery via skin. It provided deeper drug penetration through stratum corneum which traditional liposomes failed to do.