Academic Sciences

International Journal of Pharmacy and Pharmaceutical Sciences

ISSN- 0975-1491 Vol 4, Issue 3, 2012

ResearchArticle

STUDYOFANALGINATE/HPMCBASEDINSITUGELLINGOPHTHALMICDELIVERYSYSTEM FORLEVOFLOXACINHYDROCHLORIDE
MAHESHNS*,MANJULABP
DepartmentofPharmaceutics,GovernmentcollegeofPharmacy#02,P.KalingaRaoRoad,SubbiahCircle,Bangalore560027India. Email:mahesh.satya@yahoo.com Received:19Mar2012,RevisedandAccepted:02May2012 ABSTRACT Conventionaleyedropscanresultinpoordrugbioavailabilityduetouniqueocularanatomyandphysiology1.Ionactivated in situgellingsystems areabletocrosslinkwithcationspresentinthetearfluid,thereforeformingagelontheocularsurface,whichresultsprolongedcornealcontact time. The present work describes the formulation and evaluation of an antibacterial agent levofloxacin hydrochloride, based on the concept of ion activatedinsitugelation.Sodium alginatewas usedasthegellingagentincombinationwithHPMCE15(HydroxyPropylMethylCellulose)that acted as a viscosity enhancing agent. The rheological behaviour of all the formulations were not affected by the incorporation of levofloxacin hydrochloride,bothinvitroandinvivoprecornealretention studiesindicatedthatthealginateorHPMCE15solutionsretainedthedrugbetter thanthealginateorHPMCE15alone.Theformulationsweretherapeuticallyefficacious,sterile,stableandprovidedsustainedreleaseofthedrug over a period of time. These results demonstrate that the developed system is economical, patient compliant and an alternative to conventional ophthalmicdrops,patientcompliance,industriallyorientedandeconomical. Keywords:Ophthalmicdeliverysystems,Sodiumalginate,Levofloxacin. INTRODUCTION Ophthalmic drug delivery is one of the most interesting and challenging areas facing the pharmaceutical scientist. The unique anatomy and physiology of the eye make it difficult to achieve an effectivedrugconcentrationatthetargetsite.Thechallengeremains to circumvent the protective barriers of the eye without causing significant tissue damage or increasing the risk of systemic side effects.Variousdeliverysystemshavebeeninvestigatedduringpast decades, pursuing two main strategies: to increase the corneal permeabilityandtoprolongthecontacttimeontheocularsurface2. While former is mainly achieved by the use of prodrugs3, penetration enhancers4, 5 and colloidal delivery systems such as nanoparticles6, 7, 8, liposomes9 and niosomes10, latter relies on rheological and mucoadhesive properties of the ophthalmic formulation. Polymeric eye formulations can be subdivided into three groups: viscosity enhancing polymers, which simply increase the formulationsviscosity,resultingindecreasedlacrimaldrainageand enhanced bioavailability, muco adhesive polymers, which interact withocularmucin,thereforeincreasingthecontacttimewithocular tissues,and in situgellingpolymers,whichundergosoltogelphase transitionuponexposuretothephysiologicalconditionspresentin theeye.Thelatterarehighlyadvantageousoverthepreformedgels, whichdonotallowforaccurateandreproducibleadministrationofa drugandafteradministration,oftenproduceblurredvision,crusting ofeyelidsandlacrimation.Insitugellingsystems,ontheotherhand, canbeeasilyandaccuratelyinstilledinliquidform,andarecapable ofprolongingtheformulationsresidencetimeonthesurfaceofthe eyeduetogelling11. Keeping the physiology of the ocular surface in mind, three parameters (temperature, pH and ionic strength) can be generally exploited. A number of studies have already been performed on thermosetting gels based on pluronic12, 13 and pH sensitive formulations of chitosan and Carbopol14, 15, but besides a few carbomer based artificial tear products none of these formulations has made it into market. The majority of studies have been conductedonionactivated in situ gellingsystemswhichareableto crosslinkwiththecationspresentinthetearfluid,formingagelon theocularsurface.TheycanbeformulatedatoptimalpHforocular deliveryusingbuffers,canbeeasilyandaccuratelyinstilledatroom temperature and may therefore be less irritating to the ocular tissuesthan in situgellingsystemsdependingonachangeinpHor temperature. Gellan gum (gelrite) based systems have been evaluated for many drugs such as Timolol maleate 16, ciprofloxacin hydrochloride17, indomethacin18 and gatifloxacin19 and have shown prolonged corneal residence time and increased ocular bioavailability of the drug. However, only a limited number of studies have been performed on xanthan gum and carrageenan systems20 withasubstantiallackofstudiescomparingthevarious in situ gellingpolymerformulations.Theobjectiveofthepresentstudy was to develop an ion activated in situ gelling for Levofloxacin, a fluoroquinolone derivative used to treat external infections of the eye such as acute and subacute bacterial conjunctivitis,keratitis, keratoconjuctivitis22andcornealulcerswhichcanpreventfrequent drug administration and enhance patient compliance. Sodium alginatewasusedasthegellingagentincombinationwithhydroxy propylmethylcelluloseE15(HPMCE15)astheviscosityenhancer for the formulation of Levofloxacin eye drops (0.5% w/v), which undergo gelation when instilled into the culdesac of the eye and providesustainedreleaseofthedrug. MATERIALSANDMETHODS Materials LevofloxacinhydrochloridewasobtainedasgiftsamplefromCadila health care ltd., Ahmedabad, India. Sodium alginate (HiMedia labs India)andHPMCE15fromApotexpharma,allotherreagentswere ofanalyticalgrade. Preparationofformulations Table I shows the composition of all the formulations. The alginate/HPMCE15 hydrogel was prepared by dispersing the required amount in 0.9% W/V sodium chloride with constant stirringusing benzalkoniumchloride(0.02% W/V)aspreservative. Levofloxacinhydrochloride(0.5%W/V)wasdissolvedinwaterand added to polymer solution under constant stirring to obtain a uniform solution. Distilled water was then added to make the volumeupto100ml. EvaluationStudies The general appearance of the formulations was observed which included colour & clarity of solution. The pH of the prepared formulationswascheckedusingapHmeter.Thegellingcapacityof the formulations was evaluated for gelling property in order to identify the formulations, suitable for use as insitu gelling system.

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Gellingwasassessedbymixingtheformulationwithsimulatedtear fluidintheproportion25:7andthegelationwasassessedbyvisual examination 21.Thetimetakenforgeltoformandthetimetakenfor ittodissolvewasnoted.Thedrugcontentwasdeterminedbytaking 1ml of the formulation and diluting it to 100ml with phosphate bufferpH6.8.Aliquotof1mlwaswithdrawnandfurtherdilutedto 25mlwithphosphatebufferpH6.8.Levofloxacinconcentrationwas

determined at 288.5nm by using UVVisible spectrophotometer (ElicoSL159).Viscosityoftheinstilledformulationisanimportant factor in determining residence time of drug in the eye. The prepared solutions were allowed to gel in the simulated tear fluid and then the viscosity determination were carried out by using Brookefield DVII+ pro extra viscometer with spindle LV3with angularvelocityrunfrom10to100rpm17.

Table1:FormulationDesignofLevofloxacinHydrochloridecontaininginsituhydrogels. S.No. 1 2 3 4 5 6

Ingredients LevofloxacinHCl Sodiumalginate HPMCE15 Sodioumchloride Benzalkoniumchloride Distilledwater

ConcentrationIn%W/V F1 F2 0.5 0.5 0.25 0.5 1 1 0.9 0.9 0.02 0.02 100ml 100ml

F3 0.5 0.75 1 0.9 0.02 100ml

F4 0.5 1.0 1 0.9 0.02 100ml

In vitro release studies of levofloxacin from formulations was studied through a fabricated apparatus consisting of 1ml of formulation placed in donor compartment over a cellophane membrane which was rinsed and soaked for 24hours in diffusion medium. The donor compartment was immersed in the receptor compartment containing 50ml of STF (Simulated Tear Fluid) maintained at 371C with constant stirring at 22 rpm using magnetic stirrer. At specified intervals of time, 1ml of the sample solution was withdrawn from the receptor compartment and replaced with the fresh STF. The samples were analyzed after necessary dilutions by UVVisible spectrophotometer at 288.5nm. The in vitro release studies were also carried out for the marketed conventionalophthalmicdropsoflevofloxacinhydrochloride(0.5% W/V) in order to compare the release profile of the drug with the preparedinsitugellingsystem. Sterilization of the selected formulations was carried out by using autoclave at 121C, 15lbs pressure for 20mins and sterility testing was performed26. The ethical committee of the institution had permitted the ocular irritation studies. Four male albino rabbits of each weighing 1.52kg were instilled with selected sterile

formulations twice a day for a period of 21 days and the rabbits wereobservedperiodicallyforredness,swellingandwateringofthe eyes. The selected formulations were stored at ambient humidity conditionsbetween5C,25C2C/65%RH5%and40C2C/75% RH5%foraperiodofonemonth.Thesampleswerewithdrawnat frequent intervals and evaluated for the parameters viz. pH, appearance,gelationstudies,drugcontentandinvitrodrugrelease23. RESULTSANDDISCUSSION The formulations were light yellow in color and the clarity was found to be satisfactory. The pH of all the formulations was within theacceptablerangeandhencewouldnotcauseanyirritationupon administration. The drug content of all the formulations was in range.TheevaluationresultsarealsomentionedintableII.Thetwo main prerequisites of gelling system are viscosity and gelling capacity.ExceptfortheformulationsF1andF2,alltheformulations gelledinstantaneouslywitha translucent matrix on addition tothe STF, which may due to ionic cross linking of the alginate chains by thedivalentcationandextendedforfewhours.

Table2:EvaluationresultsofpH,DrugContentandGellingCapacity. Parameters pH DrugContent(%) GellingCapacity F1 7.1 103.5 + F2 7.1 101.6 + F3 7.1 98.0 ++ F4 7.1 103.0 +++

Note:+gelsslowlyanddissolves;++gelationimmediateandremainsforafewhours;+++gelationimmediateandremainsforanextendedperiod.

Generally viscosity values in the range of 1550cps significantly improvethecontacttimeintheeye.Theviscosityoftheformulations F1 to F4 ranged from 1448cps with increase in the alginate concentration within the system. All the formulations exhibited pseudoplasticrheology,asshownbyshearthinningandadecreasein theviscositywithincreasedangularvelocityfigI.Theadministration

ofophthalmic preparationsshould haveaslittle effectaspossible on thepseudoplasticcharacteroftheprecornealfilm24.Sincetheocular shear rate is very high, ranging from 0.03 s1 during interblinking periodsto425028500s1duringblinking25,viscoelasticfluidswitha viscositythat ishigh underlow shear rate conditions and low under thehighshearrateconditionsareoftenpreferred.

Fig.I:(SPINDLENOLV03) 656

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TheInvitroreleasestudiesfigIIindicatedthatamongstalltheformulations,F4sustaineddrugreleasefor8hours,whichmaybeduetothehigher concentrationofsodiumalginatealongwithHPMC.

Fig.2:PlotofinvitroreleaseoflevofloxacinasafunctionoftimeforformulationsF1F4frominsitugellingsystems Note:%CDR=Cumulative%DrugReleased The higher regression coefficient values tableIII for each formulation suggested that the formulations F1 to F4 behaved matrix type of drug release. Table3:Regressioncoefficientanalysisandbestfitanalysisfortheformulations Formulations F1 F2 F3 F4

Zeroorder 0.7935 0.8608 0.8082 0.7663

Firstorder 0.7941 0.8609 0.8083 0.7664

Matrix 0.9957 0.9965 0.9961 0.9952

Peppas 0.9885 0.9960 0.9916 0.7664

HixonCrowell 0.9420 0.8609 0.8083 0.7664

The comparative in vitro drug release profile fig III between the marketed conventional ophthalmic drops and the formulation F4 showed41.69%and33.98%afterinitial60min.Attheendof180
[

min.thedrugreleasewasfoundtobe99.35%and51.16%fromthe marketed product and F4 indicating that the drug release was significantlyprolongedbyusingtheinsitugellingsystems.

Fig.3:ComparisonofdrugreleaseprofileofmarketedconventionaleyedropwithformulationF4

The formulation F4passed the sterility test as there was no appearanceofturbidityandhencenoevidenceofmicrobialgrowth whenincubatedfornotlessthan14daysat3035Cincaseoffluid

thioglycollatemediumandat2025Cinthecaseofsoyabeancasein digestmedium.Theresultsoftheocularirritationstudiesindicated that the formulation F4 is nonirritant with excellent ocular 657

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tolerance.Nooculardamageorabnormalclinicalsignstothecornea, irisorconjunctivawerevisible. The stability studies indicated that the formulation F4 was physicallyandchemicallystablewithnosignificantchangeinanyof the parameters evaluated when stored at the ambient humidity conditionsbetween5C,25C2C/65%RH5%and40C2C/75% RH5%.Fromstabilitystudiesitwasobservedthatthe insitugelling systemoflevofloxacinwasstableatselectedstorageconditions. CONCLUSION Levofloxacin, a broad spectrum antibacterial agent used in the treatmentofocularinfectionswassuccessfullyformulatedasanion activated in situ gel forming ophthalmic solution using sodium alginate in combination with HPMCE15 as a viscosity enhancer which sustained the drug release over a period of 8 hours. The polymersusedareinexpensiveandeasilyavailable.Theformulation also promises to reduce thefrequency of drugadministration, thus improvingpatientcompliance.Astheconceptinvolvedisnovel and the methodology used for the preparation is simple as that of conventional ophthalmic liquid dosage form, it is industrially orientedandeconomical. REFERENCES Manish K, Kulkarni GT. Recent advances in ophthalmic drug deliverysystem.IntJPharmPharmaSci2012;4(1):38794. 2. Jarvinen K, Jarvinen T, Urtti A. Ocular absorption following topicaldelivery. 3. AdvDrugDelivRev1995;16:319. 4. LeeVHL,LiVHK.Prodrugsforimprovedoculardrugdelivery. 5. AdvDrugDelivRev1989;3:138. 6. Kaur IP, Smitha R. Penetration enhancers and ocular bioadhesives: two newavenues for ophthalmic drug delivery. DrugDevIndPharm2002;28:35369. 7. Saettone MF, Chetoni P, Cerbai R, Mazzanti G,Braghiroli L. Evaluationof ocular permeation enhancers: in vitro effects on corneal transport of fourbetablockers, and in vitro/in vivo toxicactivity.IntJPharm1996;142:10313. 8. Diebold Y, Calonge M. Applications of nanoparticles in ophthalmology.ProgRetinEyeRes2010;29:596609. 9. Nagarwal RC, Kant S, Singh PN, Maiti P, Pandit JK. Polymeric nanoparticulate system: a potential approach for ocular drug delivery.JControlRel2009;136:213. 10. Zimmer A, Kreuter J.Microspheres and nanoparticles used in oculardeliverysystems.AdvDrugDelivRev1995;16:6173. 11. Meisner D, Mezei M. Liposome ocular delivery systems. Adv DrugDelivRev1995;16:7593. 12. Kaur IP, Garg A, Singla AK, Aggarwal D. Vesicular systems in oculardrugdelivery:anoverview.IntJPharm2004;269:114. 1. 13. KraulandAH,LeitnerVM,BernkopSA.Improvementinthein situ gelling properties of deacetylatedgellan gum by the immobilization of thiol groups. J PharmSci 2003; 92: 1234 41. 14. ElKamelAH.InvitroandinvivoevaluationofPluronicF127 based oculardelivery system for timolol maleate. Int J Pharm 2002;241:4755. 15. QiH,ChenW,HuangC,LiL,ChenC,LiW,WuC.Development ofa poloxamer analogs/Carbopolbased in situ gelling and mucoadhesive ophthalmicdelivery system for puerarin. Int J Pharm2007;337:17887. 16. Gupta H, Velpandian T, Jain S. Ion and pHactivated novel in situgelsystemforsustainedoculardrugdelivery.JDrugTarget 2010;18:499505. 17. SultanaY,AqilM,AliA,ZafarS.Evaluationofcarbopolmethyl cellulose based sustainedrelease ocular delivery system for pefloxacinmesylate using rabbit eye model. Pharm DevTechnol2006;11:31319. 18. RozierA,MazuelC,GroveJ,PlazonnetB.Gelrite(R):anovel,ion activated,insitugellingpolymerforophthalmicvehicles.Effect onbioavailabilityoftimolol.IntJPharm1989;57:16368. 19. Balasubramaniam J, Pandit JK. Ionactivated in situ gelling systems for sustained ophthalmic delivery of ciprofloxacin hydrochloride.DrugDeliv2003;10:18591. 20. Balasubramaniam J, Kant S, Pandit JK. In vitro and in vivo evaluation of the Gelritegellan gumbased ocular delivery systemforindomethacin.ActaPharm2003;53:25161. 21. KalamMA,SultanaY,SamadA,AliA,AqilM,SharmaM,Mishra AK.Gelritebased in vitro gelation ophthalmic drug delivery systemofgatifloxacin.JDispersionSciTechnol2008;29:8996. 22. CeulemansJ,VinckierI,LudwigA.Theuseofxanthanguminan ophthalmicliquiddosageform:rheological characterizationof theinteractionwithmucin.JPharmSci2002;91:111727 23. Carlfors J, Edsman K, Peterson R , Jornving K.Rheological evaluationofGelritein situ gelsforophthalmicuse.EurJPharm Sci.1998;6:11319. 24. Vijay DW, Dipak UA, Sanjay JS. Drug delivery and Pharmacotherapy for dry eye disease. Int J Pharm Pharma Sci 2012;4(2):426. 25. KulkarniGT, GowthamarajanK, Suresh B. Stability testing of Pharmaceutical products: An overview. Indian J Pharm Edu2004;38:19498. 26. Bothner H, Waaler T, Wik O. Rheological Characterization of tearsubstitutes.DrugDevIndPharm1990;16:75568. 27. Kumar SR, Himmestein KJ. Modification of in situ gelling behavior of carbopol solutions by hydroxypropyl methylcellulose.JPharmSci1995;84:34448. 28. Indian Pharmacopeia (1996). The Controller ofPublications, Delhi,India,Testforsterility.Vol.II,A117.

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