Effect of hydrophobic nanoparticle cores on the ... · Y. Sirithananchai et al. physiological...
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Effect of hydrophobic nanoparticle cores on the characteristics of poly(ε-caprolactone)-co-d-α-tocopheryl polyethylene glycol 1000 succinate nanoparticles loaded with camptothecin Y. Sirithananchai 1 , VB. Junyaprasert 1,2 , K. Sakchaisri 3 , J. Suksiriworapong 1,2,* 1 Department of Pharmacy, Faculty of Pharmacy, Mahidol University, Bangkok 10400, Thailand 2 Center of Excellence in Innovative Drug Delivery and Nanomedicine, Faculty of Pharmacy, Mahidol University, Bangkok 10400, Thailand 3 Department of Pharmacology, Faculty of Pharmacy, Mahidol University, Bangkok 10400, Thailand Abstract The clinical efficacy of camptothecin (CPT), a potent antitumor drug, is restricted by its insolubility in aqueous and organic solvents and its instability in physiological fluid. The incorporation of drug in nanoparticles can minimize these problems. Since CPT exhibits hydrophobic properties, the nanoparticle core with different hydrophobicity may affect the characteristics of nanoparticles. This study aimed to investigate the effect of hydrophobicity of the nanoparticle core on the characteristics of poly(ε-caprolactone)-co-d-α-tocopheryl polyethylene glycol 1000 succinate (PCL- TPGS) nanoparticles for the entrapment of CPT. The result found that the increasing hydrophobicity of the nanoparticle core significantly enlarged the particle size of blank nanoparticles from 40 nm to 164 nm while it decreased the polydispersity index (PdI) from 0.518 to 0.242 and the zeta potential (ZP) from -1.79 mV to -22.9 mV. After drug loading, the particle size increased but the ZP remained almost unchanged as compared to the blank nanoparticles. The initial fed amount of CPT at 0.1 mg yielded the highest %entrapment efficiency (%EE). The increasing amount of CPT dramatically decreased %EE from 94.03-96.73% to 18.96-22.53% but slightly increased %drug loading from 0.073-0.077% to 0.085-0.092%. The results suggested that the hydrophobicity of the PCL-TPGS nanoparticle core affected the characteristics of nanoparticles. From these results, the 3:1 PCL- TPGS nanoparticles loading with 0.1 mg of CPT showed the desirable size (smaller than 200 nm), narrow PdI, and highest %EE. Therefore, this formulation will be chosen for the development of CPT delivery system. Keyword: Camptothecin, Poly(ε-caprolactone), d-α-Tocopheryl polyethylene glycol 1000 succinate, Nanoparticles, Hydrophobic 1. INTRODUCTION Camptothecin (CPT) is a plant alkaloid isolated from an oriental tree, Camptotheca acuminata. CPT shows the significant antitumor activity against various cancers, including lung, ovarian, breast, pancreas, gliomas, and stomach. It inhibits the activity of DNA topoisomerase I, which is required for replication and transcription of cell cycle. 1, 2 However, the clinical efficacy of CPT is significantly restricted by its insolubility in water and most organic solvents. Moreover, CPT is spontaneously and rapidly inactivated due to hydrolysis of its active lactone form resulting in an inactive carboxylate form (Figure 1). Although this reaction is reversible, its equilibrium favors the carboxylate form at the physiological condition. Several strategies have been proposed primarily to increase the solubility of CPT as well as to improve the stability of CPT in *Corresponding author: E-mail: [email protected] Original Article Mahidol Univ J Pharm Sci 2015; 42 (4), 169-177
Transcript of Effect of hydrophobic nanoparticle cores on the ... · Y. Sirithananchai et al. physiological...
Effect of hydrophobic nanoparticle cores on the characteristics of poly(ε-caprolactone)-co-d-α-tocopheryl polyethylene glycol 1000 succinate nanoparticles loaded with camptothecin Y. Sirithananchai1, VB. Junyaprasert1,2, K. Sakchaisri3, J. Suksiriworapong1,2,*
1 Department of Pharmacy, Faculty of Pharmacy, Mahidol University, Bangkok 10400, Thailand2 Center of Excellence in Innovative Drug Delivery and Nanomedicine, Faculty of Pharmacy, Mahidol University, Bangkok 10400, Thailand 3 Department of Pharmacology, Faculty of Pharmacy, Mahidol University, Bangkok 10400, Thailand Abstract Theclinical efficacyof camptothecin (CPT), apotent antitumordrug, is restrictedby itsinsolubilityinaqueousandorganicsolventsanditsinstabilityinphysiologicalfluid.Theincorporationofdruginnanoparticlescanminimizetheseproblems.SinceCPTexhibitshydrophobicproperties,thenanoparticlecorewithdifferenthydrophobicitymayaffectthecharacteristicsofnanoparticles.This study aimed to investigate the effect of hydrophobicity of the nanoparticle core on thecharacteristicsofpoly(ε-caprolactone)-co-d-α-tocopherylpolyethyleneglycol1000succinate(PCL-TPGS)nanoparticlesfortheentrapmentofCPT.Theresultfoundthattheincreasinghydrophobicityofthenanoparticlecoresignificantlyenlargedtheparticlesizeofblanknanoparticlesfrom40nmto164nmwhileitdecreasedthepolydispersityindex(PdI)from0.518to0.242andthezetapotential(ZP)from-1.79mVto-22.9mV.Afterdrugloading,theparticlesizeincreasedbuttheZPremainedalmostunchangedascomparedtotheblanknanoparticles.TheinitialfedamountofCPTat0.1mgyielded the highest%entrapment efficiency (%EE).The increasing amount ofCPT dramaticallydecreased%EE from94.03-96.73% to 18.96-22.53%but slightly increased%drug loading from0.073-0.077% to0.085-0.092%.The results suggested that thehydrophobicityof thePCL-TPGSnanoparticle core affected the characteristics of nanoparticles. From these results, the 3:1 PCL-TPGSnanoparticlesloadingwith0.1mgofCPTshowedthedesirablesize(smallerthan200nm),narrowPdI,andhighest%EE.Therefore,thisformulationwillbechosenforthedevelopmentofCPTdeliverysystem.
Keyword: Camptothecin,Poly(ε-caprolactone),d-α-Tocopherylpolyethyleneglycol1000succinate,Nanoparticles,Hydrophobic
1. INTRODUCTION Camptothecin(CPT)isaplantalkaloidisolated from an oriental tree, Camptotheca acuminata.CPTshowsthesignificantantitumoractivityagainstvariouscancers,includinglung,ovarian,breast,pancreas,gliomas,andstomach.ItinhibitstheactivityofDNAtopoisomeraseI,whichisrequiredforreplicationandtranscription ofcellcycle.1,2However,theclinicalefficacyofCPTissignificantlyrestrictedbyitsinsolubility
inwaterandmostorganic solvents.Moreover,CPTisspontaneouslyandrapidlyinactivateddue to hydrolysis of its active lactone formresulting in an inactive carboxylate form(Figure1).Althoughthisreactionisreversible,itsequilibriumfavorsthecarboxylateformatthephysiologicalcondition. Severalstrategieshavebeenproposedprimarily to increase the solubility of CPTaswellas toimprovethestabilityofCPTin
*Corresponding author: E-mail:[email protected]
Original Article Mahidol Univ J Pharm Sci 2015; 42 (4), 169-177
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physiologicalfluid.CPThasbeenincorporatedtoseveralkindsofdrugcarriersystemstoimprove drugsolubilityandstability.3Untilnow,variouskindsofnanocarriersforCPThavebeenstudiedforexample,polymericmicelles,dendrimers,lipid-basednanoparticles,andmicrospheres.2,4-6 Amongthesecarriers,thepolymericnanoparticleshavebeenextensivelyinvestigatedduetomanyadvantages.Theycanenhancedrugsolubilityandstabilityinthebiologicalenvironmentby entrappinghydrophobicdrugsinthecompatiblehydrophobiccore.2,7Meanwhile,thecoverageof these particles by the hydrophilic part ofpolymer chain can avoid the mononuclearphagocytesystem(MPS)leadingtotheprolongedcirculationtime.Inaddition,thenanoparticlescanalsoenhancetheabsorptionofdrugsintothetumortissuesbytheenhancedpermeabilityandretention(EPR)effect.8 Varioustypesofcopolymershavebeenusedforthefabricationofnanoparticles.Amongthemostcommonlyusedbiodegradablesyntheticpolymers, the aliphatic polyesters includingpoly(ε-caprolactone) (PCL) exhibit attractivepropertiessuchasbiocompatibility,nontoxicity,andnonimmunogenicity.9-11Inaddition,thesepolymers havebeen approvedby theUnitedStates Food and Drug Administration (U.S.FDA) for pharmaceutical and biomedicalapplications.Overthedecades,PCLhasbeenextensivelystudiedasbiomaterials.However,theuseofPCLisrestrictedduetothehighdegreeofcrystallinityandslowbiodegradationrate.Additionally,thehighhydrophobicityofPCLacceleratestheremovalofPCLparticlesfrombloodcirculation.11,12Therefore,themodification
ofPCLwithhydrophilicpolymerisnecessarytosolvetheseissues. D-α-Tocopherylpolyethyleneglycol 1000 succinate (TPGS) is awater-soluble derivativeofvitaminE.Itsstructureiscomposed ofhydrophobicalkylchainofvitaminEandhydrophilicpartofPEG.13Ithasbeenapprovedby U.S.FDAaswater-solublevitaminEnutritional supplementanddrugdeliveryvehicle.14DuetomanyattractivecharacteristicsofTPGSincludinganti-cancerproperty,P-glycoproteininhibitoryactivity,andsolubilizingeffectforbothwater-solubleandwater-insolublecompounds, the additionofTPGStothePCLchainmaymodifythepropertiesofPCL.TheentrapmentofCPTbyPCL-TPGSnanoparticlesmaythusimprovethesolubilityandstabilityofCPT.However,onemajorfactoraffectingtheloadingcapacityofnanoparticles is thecompatibilitybetweennanoparticlecoreanddrug.Ithasbeenreportedthattheentrapmentofhydrophobicdrugscouldbeenhancedbyincreasingthehydrophobicnano-particlecore.15Therefore,itisofourinterest to investigate thedifferenthydrophobicPCLchains of PCL-TPGS copolymers on the characteristics and loading capacity of thenanoparticlesforCPT.TheratiosofhydrophobictohydrophilicsegmentsofPCL-TPGScopolymerswerevariedat1:1,2:1,3:1,and4:1andthesecopolymerswereemployed toprepare the nanoparticles. In addition, the initial amountofdrugfedwasvariedtoobservetheeffectontheloadingcapacityofthesystem.
2. MATERIALS AND METHODS2.1 Materials
CPTwaspurchasedfromXi’anLyphar
Figure 1. Reversible transformation of active lactone camptothecin to inactive carboxylate camptothecin.
171Effect of hydrophobic nanoparticle cores on the characteristics of poly(ε-caprolactone)-co-d-α-tocopheryl polyethylene glycol 1000 succinate nanoparticles loaded with camptothecin
BiotechCo.,Ltd.(Shaanxi,China).TPGSandpoloxamer407(P407)weregiftedfromBASF(Ludwigshafen,Germany).Acetonitrile(ACN),methanol, and tetrahydrofuran (THF)wereof high-performance liquid chromatography(HPLC) grade (Burdick & Jackson, Ulsan,Korea).GlacialaceticacidwaspurchasedfromRCILabscanCo.,Ltd. (Bangkok,Thailand).Sodiumacetate trihydrate (CH3COONa)wasobtained from VWR international BVBA(Leuven,Belgium).Sterilewaterforinjectionwas purchased from Thai Nakorn Patana(Bangkok, Thailand). Triethylamine (TEA,(C2H5)3N) was purchased from Carlo Erbareagents(ValdeReui,France).
2.2 Nanoparticle preparation
Thenanoparticleswerepreparedby nanoprecipitationmethod.16,17Briefly,PCL-TPGS(54mg)withanincreasinghydrophobictohydrophilicratio(1:1,2:1,3:1,and4:1)andTPGS(6mg)weredissolvedin10mLofTHF.The solution was added to 10 mL of watercontainingP4070.6%w/v.THFwasevaporatedat roomtemperatureand the resultantwas centrifugedat1,370×gtoeliminateaggregates.ForCPT-loadednanoparticles,variousamountsofCPT(0.1,0.3,and0.5mg)weredissolvedwith PCL-TPGS polymer in THF and thenanoparticles were prepared as previouslydescribed. The nanoparticles were kept as adispersionformforfurtheranalysis.
2.3 Particle size analysis
Theparticlesizeandsizedistribution(PdI) of nanoparticles were determined byphoton correlation spectroscopy (PCS)usingZetasizer Nano ZS (Malvern Instruments,Malvern,UK).Thesamplesweremeasuredafterdilutionwithdeionizedwatertoobtainasuitablescatteringintensityandperformedatanangleof173°,25°C.Allmeanparticlesizeand PdI values were attained by averagingvaluesof10measurements.
2.4 Zeta potential (ZP) analysis
The surface charge of nanoparticles,definedas thevalueofelectricalpotential at
the shear plane of particles,was determinedbyZetasizerNanoZS(MalvernInstruments,Malvern,UK).Allmeasurementswereperformed at25°C.Themeasurementofeachsamplewasrepeatedthreetimesandanaveragevaluewasreported.
2.5 Determination of drug loading efficiency
Theloadingefficiencyofnanoparticleswasevaluatedintermsofpercentdrugloading(%DL)andpercententrapmentefficiency(%EE).The measurement of %DL and %EE wasperformedbydirectandindirectmethods.Fortheindirectmethod,thenanoparticledispersionwascentrifugallyfilteredat14,811×gfor15minthroughMicroconUltracelYM-30tube(MWcut-off 3000 Da, Millipore, Schwalbach, Germany).Theamountofdruginfiltratewasanalyzed by HPLC. For the direct method,thelyophilizedsamplewasdissolvedinACNandacetatebuffer.Theprecipitatewasfilteredthrough 0.22 μm nylon syringe filter. TheclearsolutionwasanalyzedbyHPLC.The amount of drug present in the nanoparticleswas calculated by the different amount ofdrug found in thefiltrateand the lyophilizedsample.The%DLand%EEwerecalculatedaccordingtotheequationsbelow.
2.6 Percent yield (%yield)
Theyieldofnanoparticleswasdetermined by recording thedryweightofnanoparticlesafterlyophilization.Afterpreparation,3mLofnanoparticleswastransferredintotheaccurately weighedvialandthenfrozenat-80°Covernight.Subsequently, the frozen nanoparticleswerelyophilized for 24 h. Finally, the weight ofdriedproductwasrecorded.The%yieldwascalculatedbythefollowingequation.
(1)
(2)
(3)
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2.7 HPLC Analysis
TheanalysisofCPTwasperformedby HPLCmethod18usingShimadzuHPLCmachine (Japan)equippedwithDGU-20A5Degasser,LC-20ADPump,SIL-10AFAutosampler,andSPD-20AUV/VISdetector.Areversed-phaseC18 column (Phenomenex® Gemini-NX 5µC18110Ả,150×4.6mm,withaguardcolumn)wasusedasastationaryphase.ThemixtureofACNandacetatebufferpH5.5with1%v/vTEA at the ratio of 21:79v/vwas utilized as a mobilephase.CPTwaselutedthroughthe gradient mode of mobile phase which wasvariedfrom21to45%for5min.Eluentwaspumpedthroughthecolumnataflowrateof1mL/min.The injection volumewas 20µLand the detection wavelength was 370 nm.CalibrationcurvesoflactoneandcarboxylateformsofCPTwereconstructedatconcentrations of0.025,0.1,1,5,10and20µg/mLwithR2ofatleast0.9995.Therelativestandarddeviationof interday and intraday precisions was lessthan 2%.The%recovery ofCPTwas in theacceptablerangeof98.5-101.5%.19
2.8 Statistical analysis
Theresultsareexpressedasthemean ± standard deviation (SD) from at least 3measurements. The one way ANOVA orstudent’s t-test was determined using SPSSprogram for multiple or pair comparisons,respectively. The statistical difference isconsideredattheprobabilitylevelof0.05.
3. RESULTS AND DISCUSSION3.1 Blank nanoparticles
The PCL-TPGS nanoparticles werepreparedusingTPGSincombinationwithP407asstabilizers.ThehydrophobictohydrophilicratioofPCL-TPGScopolymersandtheinitiallyfed amount of CPT were investigated sincethesemayaffectthepropertiesofthenanopar-ticles. It was postulated that the PCL-TPGSnanoparticlesformedbyaggregatingPCLandvitamin E segments inside the nanoparticlecoresurroundedbythehydrophilicPEGsegment. Withoutanystabilizer,thenanoparticlestended
toaggregateresultinginmicron-sizedparticles.Furthermore,wealsofoundthattheemulsifica-tionefficiencyofsoleTPGSwasinadequateto formthestablenanoparticles(datanotshown).Therefore,thecombinationofTPGSandP407wereusedasstabilizers.Uponthepolymeraggregation,TPGSactingasanemulsifierinsertedvitaminEparttothecoreofnanoparticlesandtetheredthePEGchaintotheaqueousenviron-ment.9MeanwhileP407physicallyadsorbedon thesurfaceofnanoparticlesandformedthestericstabilizinglayersurroundingthenanoparticlespreventingtheaggregationtendencyofparticles.The combination of TPGS and P407 couldsynergisticallyfacilitatetheformationofPCL-TPGSnanoparticles.TheresultsofblankPCL-TPGSnanoparticlesaresummarizedinFigure2. Varioushydrophobic to hydrophilic ratios ofPCL-TPGScopolymersat1:1,2:1,3:1,and4:1resultedinthe%hydrophobicchainof27.43%,52.73%, 65.54%, 72.20%, respectively, ascalculated by nuclear magnetic resonancespectroscopy. It was discovered that the smallestparticleswithhydrodynamicdiameterof40±5nm wasobtainedfrom1:1PCL-TPGSformulation. However,thisformulationhadthewidestsizedistribution(0.518±0.009).AsshowninFigure 2Aand2B, the increasinghydrophobic tohydrophilicratiofrom1:1to4:1ledtothesignificant increment of particle size from40±5nmto164±1nm(p-value<0.05)andthenarrowPdIfrom0.518±0.009to0.242±0.020,respectively.All polymersused in this studycontained the sameTPGSpart.Hence the differenthydrophobicityofPCL-TPGSstemmedfrom the different chain length of PCL.Theincreasinghydrophobictohydrophilicratio from1:1to4:1increasedthePCLchainlengthandthusincreasedthehydrophobicityofpolymer. The longer hydrophobicPCLchain enlargedthe size of nanoparticleswhich agreedwellwith the previous report.20 The 1:1 and 2:1PCL-TPGSnanoparticleshadhigherPdIvaluethan the 3:1 and 4:1 nanoparticles probablyduetothefactthatthesmallparticlestendedtoaggregateuponthenanoparticleformationinordertoreducethefreeenergyofparticles.21 Thelongerhydrophobictohydrophilicratio
173Effect of hydrophobic nanoparticle cores on the characteristics of poly(ε-caprolactone)-co-d-α-tocopheryl polyethylene glycol 1000 succinate nanoparticles loaded with camptothecin
ofPCL-TPGScopolymerstendedtoformthe morecompactedhydrophobiccoreofnanoparticlesandhadthegreaterthermodynamicstabilitythanthoseformedbyshorterhydrophobicchainof PCL-TPGS.22 Therefore, the 3:1 and 4:1nanoparticleshadlessfreeenergythanthe1:1 and 2:1 PCL-TPGS nanoparticles resultinginthelowerPdIvalue.Theseresultswereinconsistentwiththepreviousreport.15Itwasestablishedthat theparticleformationwas primarilyaffectedbythenatureandthelengthofhydrophobicblockwhereasthehydrophilicblockhadonlyaslighteffect.TheZPofallpolymer ratiosshowedthenegativelychargedsurface.TheZP decreased from -1.8 ± 1.0mV to -
22.9±2.0mVwhenincreasingthehydrophobicityfrom1:1to4:1.ItwasobviousthatthelongerhydrophobicPCLsegmentprovidedthehighernegativelysurfacecharge.ThiswasprobablyduetothepresenceofcarbonylgroupsofPCLsegmentsonthenanoparticlesurface.23At1:1and2:1hydrophobictohydrophilicratios,theZPwasalmostzeroduetothePEGsegmentenablingtocoveralmostcompletelythesurfaceofnanoparticles.Theobtaineddataagreedwellwiththeliteraturesreportingthattheloweringof absolute ZP value was possibly due totheadsorptionofpolymersonthesurfaceofnanoparticles.24,25The%yieldof all blank formulationswasalmost100%(Table1).
Table 1. The %yield of blank and CPT-loaded PCL-TPGS nanoparticles (mean ± SD, n = 3)
Formulation
Hydrophobictohydrophilicratio
1:1 2:1 3:1 4:1
Blanknanoparticles 99.67±0.64 100.00±0.21 99.98±0.12 100.10±0.09 CPT-loadednanoparticles(0.1mg) 99.30±1.22 98.29±0.85 99.86±0.36 99.90±0.63 CPT-loadednanoparticles(0.3mg) 99.82±0.81 100.37±1.08 99.21±2.01 98.75±2.68 CPT-loadednanoparticles(0.5mg) 99.72±1.39 99.14±1.45 99.24±1.98 99.79±0.37
3.2 CPT-loaded nanoparticles
Theparticlesize,PdIandZPofCPT-loadednanoparticlesareillustratedinFigure2.Afterdrug loading,almostallCPT-loadednanoparticles exhibited larger size than theblank nanoparticles. TheZP values of drug-loaded1:1and2:1PCL-TPGSnanoparticlesbecame more negative possibly due to thepresenceofdrugineitherlactoneorcarboxylateformonthesurfaceofnanoparticles.Theoverall%EE and %DL were ranged from 18.96 to96.73%and0.073to0.092%,respectively,asshowninFigure3.The%yieldofCPT-loadednanoparticleswasfoundtobeintherangeof98.29-100.37%(Table1).
Effect of hydrophobic to hydrophilic ratio of polymers
AtthesameamountofCPT,theparticle
size increased with the longer hydrophobicPCLsegment(p-value<0.05).Theparticlesize of CPT-loaded nanoparticles enlarged from33±5nmto343±15nmwiththeincreasing hydrophobicityofnanoparticlecore.Thisresultwasinagreementwiththeblanknanoparticlesand the previous report20 as aforementioned.The particle sizes of CPT-loaded 1:1, 2:1,3:1,and4:1PCL-TPGSnanoparticleswereintherangeof33-54nm,80-85nm,166-199nm,and287-343nm, respectively.Similar to theblanknanoparticles,thePdIvaluesofallCPT-loadednanoparticles tended todecreasewithincreasinghydrophobictohydrophilicratioofnanoparticlecorefrom0.742±0.079to0.172± 0.003 (p-value > 0.05). However, the 3:1and4:1nanoparticlesshowedthenarrowsizedistributionofaround0.200probablydueto theaggregationtendencyofmorehydrophobic polymersresultingintheuniformityofparticle
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size as mentioned previously.22 The surfacechargeofmostnanoparticlesremainedalmostunchangedascomparedtotheblanknanoparticlessuggestingthatallCPTmoleculeswereentrapped
inside thenanoparticles.The incrementof hydrophobictohydrophilicratioofPCL-TPGSpolymerinsignificantlyaffected%EEand%DL (p-value>0.05)asshowedinFigure3.
Figure 2. The hydrodynamic diameter (A), polydispersty index (PdI, B), and zeta potential (ZP, C) of blank and CPT-loaded PCL-TPGS nanoparticles. An error bar indicates the standard deviation from at least three measurements. *Significant difference comparing between blank and CPT-loaded nanoparticles at the same hydrophobic to hydrophilic ratio of polymer and the same amount of CPT. **Significant difference comparing among all amounts of CPT at the same hydrophobic to hydrophilic ratio of polymer. ***Significant difference comparing among various hydrophobic to hydrophilic ratios of polymer at the same amount of CPT.
175Effect of hydrophobic nanoparticle cores on the characteristics of poly(ε-caprolactone)-co-d-α-tocopheryl polyethylene glycol 1000 succinate nanoparticles loaded with camptothecin
Effect of initially fed amount of CPT
In this study, theamountofpolymerwassetconstantsotheincreasingamountofCPT from 0.1 to 0.5mg increased the drugto polymer ratio. In Figure 2, the increasingamount of CPT insignificantly affected thehydrodynamic diameter, PdI, and ZP valuesofthenanoparticles.However,theincreasingdrugtopolymerratiohadmucheffectontheloadingefficiency.Thehighest%EEwasobtainedwhenCPTwasinitiallyfedat0.1mg.TheincreasingCPTamount to0.3and0.5mg dramaticallyreduced%EEfrom94.03-96.73%to29.42-30.90%and18.96-22.53%,respectively(p-value<0.05).However,itincreased%DLfrom 0.073-0.077% to 0.080-0.084% and0.085-0.092%, respectively (p-value<0.05).
The%yieldofallnanoparticlesremainedalmostconstant when increasing amount of CPT(Table1).TheinitialfedamountofCPTat0.1mg yielded the highest %EE and providedthe%DL approaching the theoretical value. This result indicated that these PCL-TPGSnanoparticlesreachedthemaximumcapacityfortheentrapmentofCPTat0.1mgofinitiallyfedamountofdrug.Therefore,0.1mgofCPTwasselectedforthedevelopmentofCPTdeliverysystem. Fromtheseresults,itcanbeconcludedthat the hydrophobicity of PCL-TPGS nanoparticles had impacts on the particle size, size distribution, and surface charge ofthe nanoparticles while the amount of CPTloadingaffectedtheloadingefficiencyofthesystem.
Figure 3. The percentages of entrapment efficiency (%EE) (A) and drug loading (%DL) (B) of CPT-loaded PCL-TPGS nanoparticles. An error bar indicates the standard deviation from at least three measurements. *Statistically significant difference comparing among various amounts of CPT at the same hydrophobic to hydrophilic ratio of polymer.
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4. CONCLUSION Theresultssuggestedthatthehydro-phobicityofPCL-TPGSnanoparticle core affected the characteristics of nanoparticles.Theincreasinghydrophobictohydrophilicratio of PCL-TPGS polymer enlarged the particlesizesofblankandCPT-loadednanoparticles.Inthemeantime,itreducedthesizedistributionandresultedinmorenegativesurfacecharge.Nevertheless, it slightly affected the loadingefficiency for theentrapmentofCPT.The increasingamountofCPTlowered%EEand%DL.The loadingefficiencywas limitedbythe fed amount of CPT. From these results,the3:1PCL-TPGSnanoparticlesloadingwith0.1mgofCPTshowedthedesirablesize(lessthan200nm),narrowPdI,andhighest%EE.Therefore,thisformulationwillbechosenforfurtherdevelopmentofCPTdeliverysystem.
5. ACKNOWLEDGEMENTS Wewould like to acknowledge thefinancialsupportfromtheThailandResearchFund, the Office of the Higher EducationCommission andMahidol University (GrantNo.MRG5680013),fromthe60thYearSupremeReignofhisMajestyKingBhumibolAdulyadejScholarshipandfromtheOfficeoftheHigherEducationCommissionandMahidolUniversityunder the National Research UniversitiesInitiative.
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