International Journal of Pharmacy and Pharmaceutical Sciences

ISSN- 0975-1491 Vol 2, Suppl 2, 2010

ResearchArticle

STUDIESONREADYMIXSUSPENSIONOFAMPICILLINTRIHYDRATE:DEVELOPMENT, CHARECTERIZATIONANDINVITROEVALUATION
JAFARM.*,AEJAZA.
DepartmentofPharmaceutics,LuqmanCollegeofpharmacy,Gulbarga585102,Karnataka,India.Email:jafar_31957@yahoo.com Received:26Jan2010,RevisedandAccepted:14Feb2010 ABSTRACT Ampicillin trihydrate is used as an antibacterial agent, with an oral dose of 250500 mg three to four times a day. Reconstitutable Ampicillin trihydratedrysyrupiscurrentlyavailableinthemarket,wherereconstitutionoftheproducthastobedonebytheconsumer,whichmayleadto handling errors. In addition, the shelf life of reconstitutable dry syrup is only for about a week after reconstitution. Ampicillin trihydrate was attemptedtoformulateintoreadymixoralsuspensionwithimprovedstabilityandshelflife.Inthefirstapproachofpreparation,waterwasusedas suspendingmediumandpHoftheformulationswaschosenisintherangeof5to65.Inthesecondapproach,oilslikefractionatedcoconutoiland refinedsunfloweroilwereusedassuspendingmedia.Thecontentuniformityofthepreparedformulationswasanalyzedandfoundtobewithinthe limits. Physical characteristics like sedimentation volume, ease of redispersability and viscosity were evaluated. Particle size determination revealed that majority of the particles was in the size range of 15 75 m. In vitro dissolution studies were carried out and all the formulations showed100%dissolutionat50thminute.Stabilitystudieswerecarriedoutat250C/60%RHand300C/60%RHfor90days.Thedrugcontentwas analyzedon7th,14th.90thdayonanintervalof7days.Sedimentationvolume,viscosity,easeofredispersability,particlesizedistributionandin vitrodissolutionwereanalysedon1stand90thday.FormulationFIandFIVshowed Considerable amount of drug degradation. All other formulations did not show appreciable changes when evaluated. Ampicillin trihydrate degradation during the accelerated stability studies was carried out for 30th day sample using TLC method. It was found that the Rf value of Ampicillin trihydrate in both standard solution as well as formulation was found to be same. This confirmed that there was no degradation of Ampicillin. Hence it was concluded that Ampicillin trihydrate could be formulated into ready mix oral suspension with improved stability and optimumdissolutioncharacteristics. Keywords:Ampicillintrihydrate,Reconstitutable,Oralsuspension,Stabilitystudies INTRODUCTION Asubstantialnumberofdrugsformulatedintheformofreadymix oralsuspensionhavebeenintroducedintomarket. Oralrouteofdrugadministrationhasbeenusedfordecades,which ispreferredtobemostconvenientandeasy.Henceitismostwidely usedamong allroutesof drugadministration. Suspensionsthough havetoundergodissolutionarestilladvantageousoversoliddosage forms as disintegration step is absent and the drug is ready for solubility in the gastro intestinal medium. Because of this suspensionsarewidelyusedfororalrouteofadministration 1 Ampicillin trihydrate is a semi synthetic penicillin derivative, and havinganantibacterialspectrumbroaderthanthatofpenicillinG hasbeenattained.Itisactiveagainstgrampositiveorganismsthat aresusceptibletootherpenicillinsanditismoreactiveagainstsome gramnegativebacteriaandenterococcalinfections When Ampicillin is given orally, it is absorbed from intestinal tract to produce peak blood level concentration in about two hours. Ampicillin trihydrate acts on microorganism by interfering with development of bacterial cell wall. Specifically, they inhibit biosynthesis of dipeptidoglycon that is needed to provide strength and rigidity to bacterial cell wall2 .Seham A.Elkheshen, Sabry, S.Budawi and AlinA.et al., have done work on optimization of a reconstitutablesuspension ofRifampicin.They havederivedavery easyandasimplemethodforestimatingtheeaseofredispersibility3. J.M.Hempenstall,IrwinW.J,Wanpo,Ali,andAndrewA.H.havedone studyonAntibioticgranulesforreconstitutionassyrupwherethe drugschosenwerePhenoxymethylpenicillinandAmpicillin.They havereportedthatthestabilityofthesuspensiondependsmainlyon the suspending medium used4.M.R.Vora, Patel M.M, Gohel M.C and Chauhan G.M. have done study on Formulation of Tinidazole suspensions. They have reported that the excipients used in the preparation of a suspension play a very vital role in its normal stability,aswellonthephotostabilityofthesuspension5.Ampicillin trihydrate has been attempted to formulate in ready mix oral suspension. The existing Ampicillin dry syrup has to be reconstituted before use. The direction given for dry syrup is to reconstitute by adding purified water up to the mark given on the label,whichistobedonebytheuseronly.Becauseoffaultylabelit mayaffectthedosageregimen.Toavoidthisproblemanattemptis made in the present investigation to prepare ready mix oral suspensionofAmpicillinwithimprovedshelflife. 109 EXPERIMENTAL MATERIALSANDMETHODS Ampicillin trihydrate is obtained as a gift sample from K.A.P.L Peenya, Bangalore, Carboxy methylcellulose sodium (Loba chemie Pvt. Ltd.)Aerosil (SmithKline Beecham, Mysore) Tartrazine Colour (Himedia,India.)Sodiumbenzoate(Ranbaxylabltd.)Sodiumacetate (Ranbaxy fine chemicals Ltd).Ninhydrin reagent (Loba chemie Pvt. Ltd.)PineappleFlavour(GenuinechemicalsCo,Mumbai).Citricacid (Ranbaxy fine chemicals Ltd). Sodium hydroxide (Ranbaxy fine chemicalsLtd)FractionatedCoconutOil(TransWorldOilsPvt. Ltd, Kerala.)RefindSunflowerOil(SunpurePvt.Ltd.,Mysore.) Methodofpreparationofsuspension Trial and error method was followed to reach the optimum formulation using different quantities of excipients. The various formulationsthatwerepreparedarelistedin(Table1). All the ingredients were added in geometric proportions. Preparationsthentransferredtohomogeniserandhomogenizedfor 15minutes.FinallyvolumeandpHwereadjustedwhereverrequired 6. Assayfordrugcontent Exactly1mlofsuspensionwastransferredto100mlvolumetricflask and volume was made to 100ml with 5NSodium hydroxide, from this1mlwaswithdrawn,transferredtoa10mlvolumetricflaskand volumewasmadeto10mlwith5NSodiumhydroxide.Theamount ofdrugpresentintheabovesolutionwasanalyzedbymeasuringthe absorbanceat272nm7. Sedimentationvolume8 Sedimentation volume of the formulations was determined using followingformula. Vs=Hu/H0 Vs= Sedimentation volume, Hu= Ultimate settled height of suspension,H0=Originalheightofthesuspensionbeforesettling Easeofredispersibility3 The suspension was allowed to settle in a measuring cylinder. The mouthofthecylinderwasclosedandwasinvertedthrough1800and IntJPharmacyPharmSci

the number of inversions necessary to restore a homogeneous suspension was determined. If the homogeneity of the suspension was attained in oneinversion,then thesuspension was considered 100%easilyredispersable.Everyadditionalinversiondecreasesthe percentageofeaseofredispersibilityby5%.

Viscositydetermination TheviscosityofallformulationswasdeterminedbyusingBrookfield digital viscometer. The measurements were carried out using spindle number3 (disc type) rotating at 10, 20, and 100 rpm. The temperaturewasmaintainedat30oC. IV 6.25% 1.83% 0.75% q.s 0.45% 90% q.s Waterq.s 5.5 q.s 6.25% 1.83% 0.75% q.s 0.45% 90% q.s Waterq.s 6.5 q.s V 6.25% 1.83% 0.75% q.s 0.45% 90% q.s Fractionated coconutoilq.s q.s VI 6.25% 1.83% 0.75% q.s 0.45% 90% q.s Fractionated coconutoilq.s q.s

Table1:CompositionofAmpicillintrihydratereadymixoralsuspensions Formulation I Ampicillin 6.25% trihydrate Carboxy methyl 1.83% cellulosesodium Aerosil 0.75% Tartrazine q.s (color)* SodiumBenzoate 0.45% Sugar 90% Pharmaceutical grade Pineappleflavor* q.s Suspending Waterq.s medium* pH 5.0 Citricacid q.s *Quantitysufficient Particlesizedistribution9,10: Using optical microscope particle size distribution studies were carriedout. 1. 2. 3. Eye piece micrometer was calibrated using stage micrometer, Samplewasuniformlysuspendedinparaffinoil. A slide of above suspension was prepared, placed under microscopeandmeasuredthesizeoftheparticles. II 6.25% 1.83% 0.75% q.s 0.45% 90% q.s Waterq.s 6.0 q.s III

At the end of the 90th day samples were analyzed for viscosity, redispersibility, sedimentation volume, drug content and in vitro dissolutionprofile TLCStudies TLCplateswerepreparedbyusingsilicagelGasastationaryphase and3% w/vsolutionofsodiumacetateinwaterasmobilephase, developing the plates in a saturated chamber. Spraying 5% w/v solutionofninhydrinidentifiedthespots.Ampicillintrihydratepure drug was used as standard. The Rf values were calculated for standardandsample. RESULTSANDDISCUSSION The prepared readymix formulations of ampicillin trihydrate were foundtopocessanexcellentredispersibilitypropertywithoptimum particle size distribution. Sedimentation studies showed that the sedimentationvolumeofallformulationsisbelow1,whichindicates thattheformulationswereoptimumandacceptable.Theviscosityof alltheformulationswassuchthatitwouldbeeasilypourable from the container and also showed a shear thinning effect. The percentagedrugcontentofthepreparedsuspensionwaswithinthe standard limits of the pharmacopoeia. Results of the Comparative evaluation of Ampicillin trihydrate suspension formulations are showninTable2.

Invitrotestofdissolution Prepared suspension formulations were subjected for dissolution usingaUSP(XXII)rotatingpaddledissolutionapparatus(apparatus II). The dosage forms were placed in 900ml of distilled water as a mediumat370(10 C).Themediawasagitatedbypaddlerotatingat 1002rpm.Aliquotsof10mlofdissolutionmediumweredrawnat intervalsof10th,20th,30th,40th,50thand60thminutes.Anequivalent volume of fresh dissolution medium was added in the dissolution vessel after each sample withdrawing. The percentage of drug dissolvedwasdeterminedbymeasuringtheabsorbanceat320nm11, 12. Acceleratedstabilitystudies

Dissolution of the prepared formulations proved that ampicillin The prepared formulations were stored at 250C/60%RH and trihydratereleasefromalltheformulationswasalmostsimilarwith 300C/60%RH.Samplesfromthestoredpreparationweretakenand analyzed after every 7th day for the period of 90 days for drug 100%dissolutionwithin50minutes(Fig1&2) contentuniformitycalculations. Table2:ComparativeevaluationofAmpicillintrihydratesuspensionformulations Parameters Evaluated Appearance Taste Viscosity (mps100rpm) Sedimentation Volume(After24hrs) Redispersibility(%) Particlesizerange(m) Drugcontent(%) Invitro%drug release(After50mins) RfValue FI Paleyellow Sweet 373 0.86 95 15250 1040.12 100.55 0.95 FII Paleyellow Sweet 315 0.84 95 15250 102.10.08 100.49 0.95 FIII Paleyellow Sweet 320 0.84 95 15250 1040.09 100.76 0.95 FIV Paleyellow Sweet 310 0.81 95 15250 104.10.04 100.96 0.95 FV Paleyellow Sweet 2850 0.77 90 15250 104.20.04 100.67 0.95 FVI Paleyellow Sweet 2901 0.76 90 15250 102.20.09 100.25 0.95

110

IntJPharmacyPharmSci

 Fig.1:PercentageofdrugdissolvedVstimeforformulationFI,FII&FIII

Fig.2:PercentageofdrugdissolvedVstimeforformulationFIV,FV&FVI AcceleratedStabilityStudiesat400C/75%RH percentage drug content, viscosity, ease of redispersibility, particle size distribution and drug dissolution after and during accelerated stability studies. Formulations FI and FIV showed a significant decrease in percentage drug content after 70th (95.7%0.12 & 96.2%0.04)and77th(94.6%0.12&94.120.16)dayrespectively. But it was observed that the physical and dissolution properties of theseformulationsremainedunaltered.TLCresultsshowedthatthe Rf valueofAmpicillintrihydrateinbothstandardsolutionaswellas formulation was found to be same. This confirmed that there is no degradation of Ampicillin. Based on above observations, it can be concludedthatAmpicillintrihydratecanbeformulatedasreadymix oralsuspensionwithimprovedshelflife.

Resultsarecitedinthetable.DatainTable3(a)and3(b)showsthat the drug is not stable at 400C and 75% RH. As it is clear from the table that within 15 days the drug is degraded below the pharmacopoeiallimits.AccordingtotheICHguidelines13ifthereis any significant change like 5% potency loss or degradant exceeds specification limits or other parameters which fail to meet the specificationsorifsignificantchangeoccursat400C/75%RH,then stabilitytestingistobecarriedoutat300C/60%RH.Sotestingwas carried out at 300C /60% RH and an additional study was carried out at 250C /60% RH. Formulations FII, FIII, FV and FVI were stable and no significant change was observed with respect to Table3(a)Resultsof%drugcontentofformulationsFI,FII,andFIII. Timeindays 1stday 7thday 14thday *StandarddeviationN=3 Labelclaim 25mg/ml %Drugcontent(MeanS.D*) FI FII 104.20.08 1040.08 98.10.16 99.10.08 64.10.14 68.10.14

FIII 104.30.04 96.80.08 72.60.12

Table3(b)Resultsof%drugcontentofformulationsFIV,FVandFVI. Timeindays 1stday 7thday 14thday *StandarddeviationN=3 Labelclaim 25mg/ml %Drugcontent(MeanS.D*) FIV FV 102.40.04 102.30.09 97.60.12 96.50.16 59.90.12 68.60.87 FVI 1040.08 97.330.12 76.10.04

111

IntJPharmacyPharmSci

ACKNOWLEDGEMENTS Authors thanks to Karnataka Antibiotics Pvt.Ltd, Bangalore for providing gift sample of Ampicillin, and to the Principal of J.S.S CollegeofPharmacy,Mysoreforprovidinglaboratoryfacilities. REFERENCES 1. 2. 3. 4. . Ansel, H.C., Eds.In: Introduction to Pharmaceutical Dosage Forms,2ndEdn.9598. Deorge, R.D., Eds.In: Wilson and Gisvold Text Book of Organic Medicinal and Pharmaceutical Chemistry, J.B.Lippincoot Co.U.S.A,8thEdn,241. Elkheshen,S.A.,OptimizationofaReconstitutable Suspension ofRifampicinUsing24FactorialDesigns,DrugDev.Ind.Pharm, 1996,22(7),623630. Hempenstall, J.M., Antibiotic Granules for Reconstitution as Syrups: Product Uniformity and Stability depend upon ReconstitutionProcedure,Int.J.Pharm.,1985,23,131146

5. 6. 7. 8. 9. 10. 11. 12. 13.

Vora, M.R., Studies on Formulation of Tinidazole Suspensions, TheEasternPharmacist,January1992,Vol.37(409),189191. Kohli,D.P.S.,Eds.In:DrugFormulationManual,542543. Klaus Florey. Eds.In: Analyfical Profiles of Drug substances, VolII,3834. Banker.G. S., Eds.In: Modern pharmaceutics, Marcel Dekker Inc.,Vol7,347349. Eugene,L.P.,Eds.In:ExperimentalPharmaceutics,4thEdn.21. Martin,A.,Eds.In:PhysicalPharmacy,4thEdn.426430. Wagh, A., Invitro Evaluation of Commercial Ampicillin Capsules,TheEasternPharmacist,October1995,Vol.37(454), 145147. Indian pharmacopoeia. The controller of publication, Delhi, 1996,Vol.I,55. International conference on harmonization of technical requirements for registration of pharmaceuticals for human use,ICH3,chapter9,ICHTechnicalcoordination,London

112

IntJPharmacyPharmSci