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A Satishchandra., et al. / International Journal of Advances in Pharmaceutical ResearchIJAPRAvailable Online throughwww.ijapronline.orgResearch PaperISSN: 2230 – 7583ETHOSOMES: A NOVAL VASICULAR CARRIER FORENHANCED DERMALDELIVERY OF AZIDOTHYMIDINEA Satishchandra * , Vedha Hari, AnnapurnaanushaKLR Pharmacy College, Palvoncha, A.P, India.Received on 22 – 11 - 2011 Revised on 25 – 12- 2011 Accepted on 03 – 01 – 201ABSTRACTConsiderable first pass metabolism and a high clearance, thus frequent administration of large dose isrequired toAzidothymidine (AZT) is widely used for treatment of Acquired Immuno deficiency syndrome andrelatedconditions,either alone or in combination with other antiviral agents. AZT has low oral bioavailability(60%) due tomaintain therapeutic drug levels.Thus ethosomal microencapsulation of AZT provides the prolongedrelease of singedose there by minimizing the frequent administration, there by reducing the side effects.Ethosomesare ethanolicphospholipid vesicles which are mainly used for transdermal delivery of drugs. Ethosomes havehigher penetrationrate through the skin as compared to liposomes hence these can be widely used in place ofliposomes. The increasedpermeation of ethosomes is probably due to its ethanolic content. Ethanol increases thecell membrane lipid fluiditywhich results in increased skin penetrability of ethosomes. These ethosomes permeatesinside the skin and fuse withcell membrane lipids and release the drug. Hot and cold methods are used forformulation of ethosomes.Azidothymidine (AZT) loaded ethosomal carriers were prepared, optimized andcharacterized for vesicular shape andsurface morphology,(SEM) scanning electronic microscopy, drug content,entrapment efficiency, in vitro releasestudy. The formulation (Etho 1) having 52% ethonolic content showedgreater entrapment efficiency(85%).Key Words: Azidothymidine (AZT); Ethosomes; phospholipids; Transdermal; Microencapsulation.INTRODUCTIONEthosomes are interesting and innovative vesicularsystems that have appeared in the field opharmaceutical technology and drug delivery in recentyears. This carrier presents interesting featurescorrelated with its ability to permeate intact throughthe human skin due to its high deformability.Ethosomes are soft, malleable vesicles tailored forenhanced delivery of active agents. It has been shownthat the physicochemical characteristics of ethosomesallow this vesicular carrier to transport activesubstances more efficaciously through the stratumcorneum into the deeper layers of the skin thanconventional liposomes . This aspect is of greatimportance for the design of carriers to be appliedboth topically and systemic drug administration. Theskin acts as a major target as well as a principlebarrier for topical/transdermal (TT) drug delivery.Thestratum corneum plays a crucial role in barrierfunction for TT drug delivery. Despite major researchandevelopment efforts in TT systems and theadvantages of these routes, low stratum corneumpermeability limits the usefulness of topical drugdelivery. To overcome this, methods have beenassessed to increase permeation. One controversialmethod is the use of vesicular systems, such asliposomes, ethosomes and niosomes, whoseeffectiveness depends on their physicochemicalproperties.Touitou discovered lipid vesicular systemsethosomes embodying ethanol in relatively highconcentration. Ethosomes contain phospholipids,alcohol (ethanol and isopropyl alcohol) in relativelyhigh concentration and water. Unlike classicalliposomes, ethosomes were shown to permeatethrough the stratum corneum barrier andwerereported to possess significantly highertransdermal flux in comparison to liposomes. Theexact mechanism for better permeation into deeperskin layers from ethosomes is still not clear. However,synergistic effects of combination of phospholipidsand high concentration of ethanol in vesicularIJAPR / Feb. 2012/ Vol. 3 / Issue. 2 / 746 - 752 746

A Satishchandra., et al. / International Journal of Advances in Pharmaceutical Researchformulations have been suggested to be responsiblefor deeper distribution and penetration in the skinlipid bilayer. The use of lipid vesicles as drug deliverysystems for skin treatment has attracted increasingattention in recent years. However, it is generallyaccepted that conventional ethosomes are of littlevalue for this purpose. Ethosomes remain confine tothe upper layer of stratum corneum (SC) and hence,are suitable for topical drug delivery. Only speciallydesigned vesicles were shown to deliver drugs acrossthe skin layers [1]. Hence the aim of present studywas to develop sustained topical ethosomal gel ofAzidothymidine and to evaluate with respect tovarious in vitro evaluation tests.MATERIAL AND METHODAzidothymidine, Soya phosphatidyl choline, ethanol,water.Method Of PreparationAzidothymidine ethosome was prepared as describedby Touitou et al .ePhospholipid and drug were takenin required quantity and dissolved in ethanol. Thismixture was heated to 30 degree centigrade in a waterbath. Double distilled water heated to 30 degrees wasslowly added as a fine stream to lipid mixture withconstant stirring at 700rpm in closed vessel. Mixingwas continued for additional 5minutes.The resultingvesicles were sonicated for 3 cycles of 5 minutes with5 minutes rest between the cycles.Characterization Of Ethosomal PreparationOptical Microscope ObservationThe ethosomal dispersion was spread on theglass slide using a glass rod. Formation ofmultilamellar vesicles was confirmed byexamining the ethosomal suspension under anoptical microscope with the magnificationpower of 100 x.Vesicular shape and surface morphologyScanning electronic microscopy (SEM)was conducted to characterize the surfacemorphology of the ethosome vesicles were analyzedby scanning electron microscopy (SEM). Prior toanalysis, the ethosome were mounted onto doublesidedtape that has previously been secured oncopper stubs and coated with platinum, thenanalyzed at different magnifications.Particle SizeParticle size of the prepared ethosomalformulations were determined by particle sizeanalyzer (Microtract S3500, USA). 1ml of theethosomal samples was poured in to the samplinghollow chamber and the mean particle size wasdetected using software system.Drug ContentThe % drug content of ethosomalpreparation was determined by using followingformula (1)%drug content =Sample absorbanceStandard absorbanceEntrapment efficiencyThe entrapment capacity of azidothymidineethosomes was measured by the ultracentrifugemethod .Vesicular preparations containingAzidothymidine was kept overnight at 4°C andcentrifuged in a ultracentrifuge at 4°C, at 30 000 rpmfor 2 h. Azidothymidine was assayed both in thesediment and in the supernatant. The entrapmentefficiency was calculated using following formula (2):%entrapment efficiency = Amount of drugadded- amount of drug non encapsulated ×100Amount of drug loadedIn Vitro Permeation StudiesThe in-vitro permeation ofAzidothymidine from ethosomal formulationwas studied using locally fabricated diffusioncell. The in-vitro diffusion of the drug throughdialysis membrane was performed. Themembrane was soaked in a buffer for 6-8hours. Required quantity of ethosomalpreparation was filled in to dialysis membraneand both the ends of dialysis membrane aresealed without leakage with thread. This actedas donor compartment. 100ml of 0.1N Hcl wastaken in a beaker which was used as a receptorcompartment. The donor compartment waskept in contact with the receptor compartmentand the temperature was maintained at 37 ±0.1 ºC. The solutions of the receptor side werestirred by externally driven Teflon-coatedmagnetic bars. At predetermined timeintervals, sample was withdrawn and replacedby 5ml of 0.1N Hcl . The drug concentrationsin the aliquot were determined at 267 nmagainst appropriate blank. This experiment wasdone in triplicate and average value wasreported. In- vitro skin permeation studieswere conducted for different formulation andeffect of variation in composition of ethanol onpermeation rate was observed.IJAPR / Feb. 2012/ Vol. 3 / Issue. 2 / 746 - 752 747

A Satishchandra., et al. / International Journal of Advances in Pharmaceutical ResearchGParticle SizeThe particle size of ethosomal preparation were examined by using Particle Size Analyser and resultswere discussed in the (Fig. 2A and 2B). 2A)H2B)Drug ContentThe %drug content of ethosomal formulations was calculated according to formula (1) and was foundto be 83.4% for etho1, 118.38% for etho2 and 111.08%etho3.Entrapment EfficiencyThe entrapment efficiency of ethosomal formulation was calculated according to formula (2) and was foundto be 85% for etho1, 79% for etho2 and 77% for etho3.In Vitro Permeation StudyThe composition of formulation was as follows: 1) Azidothymidine 2) soya lecithin 3) ethanol4) water. Release study of different formulations was taken in a suitable apparatus. The drug incorporated was 0.12g; lecithinconcentration is 0.6g and ethanol concentration was varied. In etho1 the ethanol concentration was 52% which showed 96.98% ofdrug releaseIJAPR / Feb. 2012/ Vol. 3 / Issue. 2 / 746 - 752 749

A Satishchandra., et al. / International Journal of Advances in Pharmaceutical ResearchFig No: 03, In ethanol 2 the ethanol concentration was 60% which showed 99.89% of drug release with in 3hrsFig : 3B, etho3 contains ethanol concentration of 67.5% which showed drug release of 102.29% within3hrs.(Fig:3C)Table No: 01, Composition of prepared ethosomesComposition Etho 1 Etho 2 Etho 3Drug (Azidothymidine) 0.12 g 012g 0.12gPhospholipid (Soya0.6g 0.6g 0.6gLecithin)Ethanol 52% 60% 67.5%Water q.s to 30ml q.s to 30ml q.s to 30ml% Entrapment Efficiency 85% 79% 77%Etho1 0.9811Etho2 0.7437Etho3 0.9811The kinetics parameters were tabulated in table (2) and shown in (Fig 4A, 4B and 4c).Kinetic ModelsZero order First order Higuchi KorsemeyerpeppasHixonR² R² R² R²R².8814 0.9611 .8826 0.95820.774 0.8614 0.9142 0.8560.5414 0.7523 0.8779 0.7932IJAPR / Feb. 2012/ Vol. 3 / Issue. 2 / 746 - 752 750

A Satishchandra., et al. / International Journal of Advances in Pharmaceutical Research4B)IJAPR / Feb. 2012/ Vol. 3 / Issue. 2 / 746 - 752 751

A Satishchandra., et al. / International Journal of Advances in Pharmaceutical ResearchIR Studies: IR studies of azidothymidine and lecithin were determined and the peaks are shown in (Fig5A and5B).5A)68.36560676.60597.35651.61492.4355632.1550%T45403462.343161.633033.762813.152082.831438.791466.231259.801280.17789.071142.87734.30984.56956.381006.28934.76 759.91897.11846.011110.851065.891088.67560.7435301683.1525.64000.0 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 450.0cm-15B)86.0985.885.685.485.22938.192385.2185.084.8528.1584.63567.63722.43%T 84.484.284.083.883.63838.993673.143735.36854.5883.41615.3583.283.01744.041363.761216.6882.804000.0 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 450.0cm-1Fig 5 : IR Spectra of A) Pure drug (AZT) , B) LecithinCONCLUSIONThe aim of current investigation is to evaluate thetransdermal potential of novel vesicular carrierethosomes, bearing Azidothymidine, anti-viral drughaving limited transdermal permeation. Byformulating AZT as an ethosomal formulation, wecan increase the permeation of drug in which ethanolacts as permeation enhancer. Ethosomal systems arecapable of delivering higher amounts of AZT atcontrolled release rate. It provides better remissionfrom the disease and reduces the duration of therapy.REFERENCES1. Vivek dave.,Dhirendra kumar., Shaila Lewis., Sarvesh paliwal.Ethosome For Enhanced Transdermal Drug Delivery OfAceclofenac.International Journal Of Drug Delivery .2010;2:81-92.2.Kundlik Girhepunje., Ranju Pal., HiteshGevariya.,Atanukumar ehera.,Thirumoorthy N.Ethosomes:A Novel Vesicular Carrier For EnhancedDermal Delivery Of Ciclopirox Olamine. Der PharmaciaLettre.2010;2(1):360-367.3.M.K.Das., K.Rama Rao. Microencapsulation OfZidovudine By Double Emulsion Solvent DiffusionTechnique Using Ethylcellulose.Indian .J.Pharm.Sci.2007;69(2):244-250.4.E.Touitou., B.Godin., N.Dayan.,C.Weiss.,A.Piliponsky., F.Levi-Schaffer. Intracellular DeliveryMediated By An EthosomalCarrier.Biomaterials.2001;22:3053-3059.5.Gangwar Satyam., Singh Shivani., GargGarima.Ethosomes: A Novel Tool For Drug DeliveryThrough Skin. Journal PharmacyResearch.2010;3(4):688-691.6.Ehab R. Bendas ., Mina I . Tardos.EnhancedTransdermal Delivery Of Salbutamol Sulfate ViaEthosomes.AAPS PharmSciTech.2007;8(4)Article107:E1-E9.7. M.K.Bhalaria., Sachin Naik., A.N.Misra.Ethosomes:A Novel Delivery System For Antifungal Drugs InTreatment Of Topical Fungal Diseases.INDIAN J EXPBIOL.2009;Vol.47:368-375.IJAPR / Feb. 2012/ Vol. 3 / Issue. 2 / 746 - 752 752