Drug delivery system based on cyclodextrin-naproxen inclusion complex incorporated in electrospun polycaprolactone nanofibers

(1)

ContentslistsavailableatScienceDirect

Colloids

and

Surfaces

B:

Biointerfaces

jou rn a l h om ep ag e :w w w . e l s e v i e r . c o m / l o c a t e / c o l s u r f b

Drug

delivery

system

based

on

cyclodextrin-naproxen

inclusion

complex

incorporated

in

electrospun

polycaprolactone

nanoﬁbers

M. Fatih

Canbolat

a,c,∗∗

_,

_Asli

_Celebioglu

a,b

_,

_Tamer

_Uyar

a,b,∗ a_{UNAM-National}_{Nanotechnology}_Research_Center,_Bilkent_University,_Ankara_06800,_Turkey b_Institute_of_Materials_Science_&_{Nanotechnology,}_Bilkent_University,_Ankara_06800,_Turkey c_Suleyman_Demirel_University,_Engineering_Faculty,_Textile_Engineering,_Isparta_32260,_Turkey

a

r

t

i

c

l

e

i

n

f

o

Articlehistory: Received4July2013

Receivedinrevisedform9November2013 Accepted11November2013

Availableonline19November2013 Keywords: Nanoﬁbers Drug Cyclodextrin Inclusioncomplex Naproxen Release

a

b

s

t

r

a

c

t

Inthisstudy,we selectnaproxen(NAP) asareferencedrugand electrospunpoly(␧-caprolactone)

(PCL)nanoﬁbersasaﬁbrousmatrix forourdrug-deliverysystem.NAPwas complexedwith

beta-cyclodextrin(␤CD)toforminclusioncomplex(NAP-␤CD-IC)andthenNAP-␤CD-ICwasincorporated

intoPCLnanoﬁbersviaelectrospinning.TheincorporationofNAPwithoutCD-ICintoelectrospunPCL

wasalsocarriedoutforacomparativestudy.OuraimistoanalyzethereleaseproﬁlesofNAPfrom

PCL/NAPandPCL/NAP-␤CD-ICnanoﬁbersandweinvestigatetheeffectofCD-IContherelease

behav-iorofNAPfromthenanoﬁbrousPCLmatrix.ThecharacterizationofNAP-␤CD-ICandthepresenceof

CD-ICinPCL/NAP-␤CD-ICnanoﬁberswerestudiedbyFTIR,XRD,TGA,NMRandSEM.TheSEMimaging

oftheelectrospunPCL/NAPandPCL/NAP-␤CD-ICnanoﬁbersrevealthattheaverageﬁberdiameterof

thesenanoﬁbersisaround300nm,inaddition,theaggregatesofCD-ICinPCL/NAP-␤CD-ICnanoﬁbersis

observed.ThereleasestudyofNAPinbuffersolutionelucidatethatthePCL/NAP-␤CD-ICnanoﬁbershave

higherreleaseamountofNAPthanthePCL/NAPnanoﬁbersduetothesolubilityenhancementofNAPby

CD-IC.

1. Introduction

Themainfunctionofdrugdeliverysystemsistotransport vari-ousdrugstothetargetsitesinthebodyinasecurewayandadjust thereleasemechanismsbycontrollingtheamountofdrugsand treatmenttime[1,2].Thereareseveralcarriersandformulations usedfordrugdeliverypurposessuchaspolymericmatrices,gels, cyclodextrins,liposomes,microspheres,foams,ﬁlmsandsome oth-ers[3–8].Indrugdelivery,itisexpectedfromacarriermaterialto haveatleastfollowingproperties;biocompatibility,non-toxicity, lackof immunogenicity, acceptablebiodegradationtime, repro-ducibility,andcontinuousactivationtillarrivaltothetarget[9,10]. Nanostructures and cyclodextrins (CD) present signiﬁcant opportunities in drug delivery systems with their unique and promising characteristic features. Nanostructures improve the releasebehaviorandstabilityofthedrugsbymaintainingthedrug

∗ Correspondingauthorat:UNAM-NationalNanotechnologyResearchCenter, BilkentUniversity,Ankara06800,Turkey.Tel.:+903122903571;

fax:+903122664365.

∗∗ Correspondingauthorat:SuleymanDemirelUniversity,EngineeringFaculty, TextileEngineering,Isparta32260,Turkey.Tel.:+902462111188;

fax:+902462111180.

E-mailaddresses:fatihcanbolat@sdu.edu.tr(M.F.Canbolat), tamer@unam.bilkent.edu.tr(T.Uyar).

concentrationwithinatherapeuticwindowandovercomingthe biologicalbarriersforcellularuptake[11,12].Ontheotherhand, CDsinduceimprovementindrugreleaseproﬁlesandenhancement indrugsolubilizationandstabilizationbyforminginclusion com-plexes(ICs)withdrugs[13,14].However,thereisnostandardor idealstructureavailablefordrugdeliverypurposeandmany stud-iesarereportedondevelopingmuchbetterstructuresforthesite speciﬁcdrugtargeting[15–17].

Naproxen(NAP)isapoorlywatersoluble,non-steroidal anti-inflammatory drug (NSAIDs) that is used to relieve pain or inflammation [18,19]. Yet, enhanced solubilityachievements of naproxen by formingcyclodextrin inclusion complexes (CD-IC) were reported [20,21]. Apart from CD, the use of electrospun nanofibersasanincorporatingmatrixalsoenhancedtherelease behaviorofNAPincomparisonwithcastfilms[22].However,the main functionofelectrospunnanofibersin drugdelivery appli-cationscanbedefinedastheircontrolledandsustainedrelease behaviors[23,24].Indrugdeliveryfield,whilethere arestudies aboutincorporationofCDsintopolymericstructuressuchas hydro-gelsandfilms[25,26],averylimitednumberofreportsareavailable relatedtoincorporationofCD-ICofactiveagents suchasdrugs [27],antibacterials[28],essentialoils[29]andflavors/fragrances [30–33]intoelectrospunnanofibersfordeliveryandstabilization purposes.Forinstance,usingcyclodextrinasastabilizingand solu-bilizingagentandelectrospunnanofibermatsasacarriermatrix 0927-7765/$–seefrontmatter©2013ElsevierB.V.Allrightsreserved.

(2)

16 M.F.Canbolatetal./ColloidsandSurfacesB:Biointerfaces115(2014)15–21 forsustainedreleasemayopenupnewpathwaysfordrugdelivery

purposebasedonimprovedreleaseperformanceofthedrug. CDs(␣-,␤-,and ␥-types)are cyclic oligosaccharides which areenzymaticallyconvergedfromstarchthatcomprise glucopy-ranosideunitslinkedwith␣(1–4)bonds[34].CDsarehollowand truncatedconeshapedmoleculesthathavehydrophobiccavityand hydrophilicoutershellwhichenablethemtoconstitute noncova-lenthost–guestinclusioncomplexationwithvarietyofmolecules invariousforms[34,35].Ontheotherhand,electrospunnanofibers areotherpromisingnanostructuresindrugdeliveryapplications [36–38].Therearemanyreasonsfornanofiberstructurestobe pre-ferredandusedinbiomedicalapplicationareas,i.e.,smallfibersize, highporosity,interconnectedporousstructure,andcapabilityto embedvariousfunctionaladditivesintothem.

In this study, naproxen (NAP) and inclusion complex of naproxenwithbeta-cyclodextrin(NAP-␤CD-IC)wasincorporated intopolycaprolactone(PCL)electrospunnanofibersforour drug-deliverysystem.Wecompared therelease profilesofNAPfrom PCL/NAPand PCL/NAP-␤CD-ICnanofibersandwe examinedthe effectofinclusion complexationonthereleasebehavior ofNAP fromthenanofibrousPCLmatrix.Inthisregard,wefirsttestedthe effectof_␤CDonthesolubilityofNAPbyforminginclusion complex-ation.NAPshowshighersolubilityinNAP-␤CD-ICwhencompared tofreeNAPwhichisconsistentwiththeliterature[39,40].Parallel tothisresult,improvedreleaseprofileofNAPfromPCL/NAP- ␤CD-ICnanofibersisobserved,aswell.

2. Experimental 2.1. Materials

Naproxen (NAP) was commercially purchased from Abdi Ibrahim Pharmaceutical Company (Turkey). Polycaprolactone (PCL)(Mw:80,000,SigmaAldrich),N,N-dimethylformamide(DMF)

(Riedel,Pestanal),dicholoromethane (DCM)(Sigma,ExtraPure), and potassium dihydrogen phosphate (VWR, Chromanorm for HPLC)wereobtainedcommerciallyfromvarioussuppliers. Beta-cyclodextrin (␤CD) was obtained from Wacker Chemie AG (Germany)andthede-ionizedwaterwasobtainedfromthe Mil-liporeMilli-QUltrapure WaterSystem.Allmaterialswereused withoutanypuriﬁcation.

2.2. ThepreparationofsolidˇCD-NAPinclusioncomplex (NAP-ˇCD-IC)

FortheNAP-␤CD-ICformation,1gof␤CDwasdissolvedin18ml waterand250mgNAPwasdispersedin2ml water,separately. Then,theNAPsolutionwasaddedintoCDsolutionslowly. Ulti-matesolutionwasstirredover-nightandaturbiddispersionwas obtained.Itwaskeptat−80◦_C_and_freeze-dried _to_obtain

NAP-␤CD-ICpowder.

2.3. Thepreparationofelectrospinningsolutions

TheNAP-_␤CD-ICincludingPCLsolutionwasobtainedby dis-persingtheNAP-␤CD-ICpowderinclearandhomogenousPCL(15% (w/v),withrespecttosolvent)DMF/DCM(3:1,v/v)solution,atthe 20%(w/w)polymerconcentration.Forcomparison,thepurePCL (15%,w/v)andonlyNAPincluding(4%(w/w)withrespectto poly-merconcentration)PCL(15%,w/v)solutionswerealsopreparedin DMF/DCM(3/1,v/v)blendsystem.

2.4. Electrospinning

ThePCL,PCL/NAPandPCL/NAP-␤CD-ICsolutionswereplaced ina 3mlsyringe ﬁttedwitha metallic needle of 0.6mm inner

diameter. The syringe was fixed horizontally on the syringe pump (model SP 101IZ, WPI, USA). The positive electrode of thehigh-voltagepowersupply (MatsusadaPrecision,AUSeries, Japan)wasclampedtothemetalneedle tip,and thecylindrical aluminum collectorwasgrounded.The parameters ofthe elec-trospinningwereadjustedas;feedrateofsolutions=1ml/h,the appliedvoltage=15kV,andthetip-to-collector distance=10cm. Electrospunnanofibersweredepositedonagroundedstationary cylindricalmetalcollectorcoveredwithapieceofaluminumfoil. TheelectrospinningapparatuswasenclosedinaPlexiglasbox,and electrospinningwascarried outat25◦C,20% relativehumidity. Thecollectednanofibersweredriedatroomtemperatureunder thefumehoodovernight.

2.5. Measurementsandcharacterization

TheexactmolarratiobetweenNAP:␤CDintheinclusion com-plexwasdeterminedbyusingprotonnuclearmagneticresonance (1_H_NMR,_Bruker_D_PX-400)_system._The_NAP-_␤CD-IC_powder_was

dissolvedind6-DMSO(in20g/Lconcentration).Thespectrawere recordedat 400MHz and at16 total scan. A rheometer(Anton PaarPhysicaCR301)equippedwithacone/plateaccessory (spin-dletypeCP40-2)wasusedtomeasuretherheologicalbehaviorof PCL,PCL/NAPandPCL/NAP-␤CD-ICsolutionintherangeof0.1to 1001/sshearrate.Thescanningelectronmicroscope(SEM)(FEI Quanta200FEG)wasusedforthemorphologicalanalysisofthe electrospunnanofibers.Samplesweresputteredwith5nmAu/Pd priortoSEMimaging.Theaveragefiberdiameter(AFD)was deter-minedfromtheSEMimages,andaround100fiberswereanalyzed. The crystallinestructure determination of the NAP, ␤CD, NAP-␤CD-ICpowderandPCL,PCL/NAPandPCL/NAP-␤CD-ICnanofibers wereinvestigated byusingX-raydiffraction(XRD) (PANalytical X’Pertpowderdiffractometer)havingCuK␣radiationinarangeof 2=5–30◦.Thethermalpropertiesofelectrospunnanofiberswere studiedbythermogravimetricanalysis(TGA)(TAQ500)andthe measurementswerecarriedout from25to500◦C at20◦C/min heatingrate,andN2wasusedasapurgegas.Theinfraredspectraof

thenanoﬁberswereobtainedbyusingaFouriertransforminfrared spectrometer(FTIR)(Bruker-VERTEX70).For measurement,the samplesweremixedwithpotassiumbromide(KBr)andpressed aspellets.Thescans(64scans)wererecordedbetween4000cm−1 and400cm−1ataresolutionof4cm−1.UV–vis-spectroscopy (Var-ianCary5000) wasused todeterminethesolubility difference betweenpureNAPandNAP-␤CD-IC.Forthispurpose,5×10−4M NAPpowderandNAP-␤CD-ICthatincludesthesameamountof NAPweredissolvedinwater.After24hstirring,thesolutionswere ﬁlteredandtheUVabsorbanceofsampleswasmeasuredinthe 250–370nmrange.

2.6. TheNAPreleaseprofilefromelectrospunPCLnanofibers TheHPLCsystem(Agilent1200Series)wasusedtoinvestigate thereleaseprofilesofPCL/NAP andPCL/NAP-␤CD-ICnanofibers. TheseparationofNAPwasperformedwithZorbaxEclipseXDB-C18 column(150mm×4.6mm,5␮mparticlesize)anditwasdetected at 230nm wavelength. Acetonitrile (100%) was used asmobile phaseataflowrateof1ml/minandtheinjectionvolumewaskept at10␮l.Forthetest,30mgweightedPCL/NAPandPCL/NAP- ␤CD-ICnanofiberswereimmersedinto30mlbuffersolutionsandthey werekeptinthatbuffertodeterminethereleasedamountofNAP attheprogressingtimeintervals.Theexperimentswererepeated threetimesforbothcompositenanofibers.Thecalibrationcurve of NAP was prepared by using stock solutions in 7 different concentrations;20ppm, 10ppm, 5ppm, 2ppm, 1ppm, 0.5ppm and0.2ppm.ItshowedlinearityandacceptabilitywithR2_≥_0.99.

(3)

Fig.1.1_H_NMR_spectrum_of_NAP-␤CD-IC_dissolved_in_d6-DMSO.

Themeasurementresultswereadaptedtothiscalibrationcurve intermsofpeakareaundercurves.

3. Resultsanddiscussion

Naproxen(NAP)waschosenasareferencedruginthisstudy based on its well-known inclusion complex formation ability withbeta-cyclodextrin(␤CD)[19,41,42].Thereleasebehaviorof NAPafterdirectincorporationintoPCLnanofibers(PCL/NAP)and afterNAP-␤CD-ICformationandincorporationintoPCLnanofibers (PCL/NAP-␤CD-IC)wereexaminedinthepresentstudy.Themain objectiveofthisstudyistogetbetterreleaseprofileofNAPfrom PCL/NAP-␤CD-ICnanofibersduetohighersolubilityenhancement ofNAPby␤CD-ICformation.

In the first step, NAP-␤CD-IC was formed by freeze-drying method.Themixingratiosof1:4(w/w)waschosenforNAP-␤CD to provide molar ratio as 1:1.2 for the proper inclusion com-plex formation. In further steps, several characterization tests were performed to prove theinclusion complex formation. To testthesolubilityofNAP by␤CD-ICformation,UV–visanalysis wasperformed.Then,releaseprofilesofNAPfromPCL/NAPand PCL/NAP-␤CD-ICnanofibrousmatsinbuffersolutionswere ana-lyzedbyHPLCmethod.

3.1. Inclusioncomplexcharacterization

1_H_NMR_study_was_performed_to_ﬁgure_out_the_molar_ratio_of

NAP-␤CDandamountofNAPintheinclusioncomplex.Fig.1 indi-catesthe1_H_NMR_spectrum_of_the_NAP-_␤CD-IC_powder._The_molar

ratiowascalculatedbytakingtheintegralofNAPpeakatabout 1.4ppm[43]and␤CD’scharacteristicpeakatabout4.8ppm[44]in d6-DMSOsystem.Itwasfoundthat,NAP-␤CD-IChave1:1.2molar ratiowhentheintegralsofmentionedpeakswereproportionedto eachotherandthisratioisquitewellagreewithourinitialmixing ratio.

Then,weusedFTIRspectroscopytoobservethespectralchanges and therepresentative bands of thespectrafor the substances beforeandafterICformation.TheFTIRspectraofpureNAP,pure ␤CDandNAP-␤CD-ICaredepictedinFig.2a.IntheFTIRspectrum ofNAP,distinctabsorptionbandat1029cm−1correspondstoC O stretching,at1228cm−1 correspondsto O stretching,and at 1395cm−1 correspondstoCH3 bending.Peaksat1685cm−1 and

1729cm−1correspondtoanti-symmetricalandsymmetricalC O stretchingvibrations[45,46].Incaseof␤CDspectra,characteristic peaksareappearedat1029cm−1,at1080cm−1and1157cm−1due toC Ostretch,at1638cm−1duetoH OHbending,at2927cm−1 duetoC Haliphaticstretch,andat3401cm−1duetoO H stretch-ing[45–47].TheFTIRspectrumofNAP-␤CD-ICshowsabandat about1730cm−1whichcomesfromNAPwithaslightshiftwhich isinanagreementwiththeliterature[41,42,46].Alsoitwasseen thattypicalpeaksoftheNAPweresuppressedintheNAP-␤CD-IC spectrawhichsuggestedthesuccessfulICformation.

Fig.2. FTIRspectraof(a)NAP,␤-CDandNAP-␤CD-IC,and(b)PCL,PCL/NAPand PCL/NAP-␤CD-ICnanoﬁbers.

(4)

18 M.F.Canbolatetal./ColloidsandSurfacesB:Biointerfaces115(2014)15–21

Fig.3.XRDdiffractionpatternsof(a)NAP,␤-CDandNAP-␤CD-IC,and(b)PCL, PCL/NAPandPCL/NAP-␤CD-ICnanoﬁbers.

TheXRDpatternsofthepureNAP,pure␤CDandNAP- ␤CD-IC were recorded toinvestigate thepossible differences in the crystallinityofthestructures.AsitcanbeseenfromFig.3a,for theNAP-␤CD-IC,distinctdiffractionpeaksforthecrystallineNAP weredetectedintheXRDpatternsindicatingthatsomefreeNAP presentintheNAP-␤CD-ICpowder.Alsothecharacteristicpeaks ofchannel-typepackingstructureat2∼12◦,18◦,and19◦of␤CD wereobservedfortheNAP-␤CD-ICwhichshowsthesuccessfulIC formationofNAPwith␤CD[48].

TGAthermogramsofpureNAP,pure_␤CDandNAP-_␤CD-ICare shown inFig. 4a. TGAthermogramsshow weightlosses below 100◦Cforboth␤CDandNAP-␤CD-ICduetowaterlossandmain degradation wasobserved for NAP at 268◦C and at 350◦C for ␤CD.Thewaterlosswasabout11%for␤CDwhileitwasaround 8%for NAP-␤CD-IC.The 3%difference mightbe attributableto theexistenceofNAP insteadofwater inthe␤CD cavity.Other thantheinitialweightloss,therearetwoweightlossesseenfor NAP-␤CD-IC.Firstoneisbetween150◦C and250◦Candsecond oneisbetween300◦Cand350◦CwhichcorrespondstofreeNAP and IC weight losses that merged with theCD decomposition, respectively.Therelativelessdecomposedamountofuncomplexed free NAP than the initial amount and the higher temperature shift in the NAP–␤CD part prove the successful formation of NAP-␤CD-IC.

TheUV–visspectroscopymeasurementsofNAPandNAP- ␤CD-ICsolutionsweredisplayedinFig.5.Asitcanbeseenfromthe spectra,theabsorptionintensityofNAP-␤CD-ICsolutionishigher thanNAP powder for thesameamount ofNAP (5×10−4M).It ismainlybecause,theinclusioncomplexationofNAPwith␤CD enhancethesolubilityofinsolubledrug,NAP,in watermedium andleadstohigherintensityoccurrenceinUV–vis-spectra.Thus, theinclusioncomplexationandthesolubilityenhancementarealso provedbytheUV–vismeasurements.

Fig.4.TGAthermogramsof(a)NAP,␤-CDandNAP-␤CD-IC,and(b)PCL,PCL/NAP andPCL/NAP-␤CD-ICnanoﬁbers.

3.2. Characterizationofelectrospunnanoﬁbers

Following the characterization studies of NAP-␤CD-IC, PCL polymer matrix was chosen for the production of electrospun nanofibers.SincePCLiswaterinsolubleandbiodegradable poly-mer[49,50],itisthoughtthattheuseofPCLnanofibersasadrug deliverysystemmightexhibitconvenientreleaseprofileforNAP. FollowingconcentrationadjustmentsofthePCL/NAPand PCL/NAP-␤CD-ICblends,electrospinningandcharacterizationstudieswere performed.

Weinvestigatedthemorphologyofelectrospunnanofibersof purePCL,PCL/NAPandPCL/NAP-_␤CD-ICbySEMimaging.The rep-resentativeimagesofSEMmicrographsweregiveninFig.6and fiberdiameterdistributiondataweresummarizedinTable1. Uni-form,beadfreenanofiberswithdiametervariationwereobtained fromallthreePCLbasednanofibroussamples.Itis clearlyseen intheFig.6cthatthereareICcrystalaggregatesaccumulatedin

Fig.5.SolubilityanalysisofNAPbyUV–visspectroscopy;solubilityofpureNAPand solubilityofNAPfromNAP-␤CD-IC.

(5)

Table1

Polymersolutionparametersandaveragefiberdiametervaluesofelectrospunnanofibers(thefibersizeisreportedastheaverage±standarddeviation;foreachcase100 fiberswereanalyzed).

Solvents Concentrations Viscosity(Pas) Averageﬁberdiameter(nm)

PCL DMF/DCM(3:1,v/v) 15%(w/v) 0.465 336±100

PCL/NAP DMF/DCM(3:1,v/v) 15%(w/v)/4%(w/w) 0.675 361±152

PCL/NAP-␤CD-IC DMF/DCM(3:1,v/v) 15%(w/v)/20%(w/w) 0.745 389±167

theﬁbermatrixwhichprovesthesuccessfulincorporationof

NAP-␤CD-ICintoelectrospunPCLnanoﬁbers.Itwasfoundoutthatﬁber

diameterdistributions have good correlationwiththeviscosity

measurementresultsascanbeseeninTable1.Theincorporation

ofNAPandNAP-␤CD-ICintoPCLmatrixcausedviscosityincrease inthepolymersolutionswhichﬁnallytriggeredtheformationof nanoﬁberswithlargerdiameters.

WeperformedFTIRstudiesfornanoﬁbersofpurePCL,PCL/NAP andPCL/NAP-␤CD-ICsamples(Fig.2b).TypicalpeaksforthePCLare observedat2949cm−1and2865cm−1duetoCH2stretching

vibra-tions,at1731cm−1duetoC Ostretchingvibrations,at1471cm−1, 1397cm−1,and 1365cm−1 due toCH2 bendingvibrations.Also

C OandC Cstretchingvibrationsat1293cm−1,C O C stretch-ingvibrationsat1240cm−1,1169cm−1,1108cm−1,and1048cm−1 wedetected[51–53].InFTIRspectraofPCL/NAPnanofibers,the characteristicpeaksofPCLmostlyexistwhilenoneofthe character-isticpeaksofNAPwasobservedwhichismostprobablyduetothe dominanteffectofthePCLpeaksandtherelativelylow concentra-tionofNAPinthepolymermatrix(∼4%,w/w).Forinstance,almost allthepeaksinthefingerprintregionarequitewellfittedwith PCLcharacteristicpeaks.However,thereisagoodindicationabout theexistenceofNAPduetosomeshiftedpeaksat2869cm−1and 2952cm−1whiletheyarelocatedat2865cm−1and2949cm−1in thePCLspectra.TheFTIRspectrumofPCL/NAP-␤CD-ICnanofibers hasverysimilarcharacteristicfeatureswithPCL/NAPnanofibers. Again,PCLpeakswereoversaturatedinthespectrawhile␤CD-IC peaksweresuppressed,evenoneofthemostdistinctpeakof␤CD at1029cm−1isnotvisible.AlthoughtheFTIRspectradidnotdepict

Fig.6. SEMmicrographsof(a)PCL,(b)PCL/NAPand(c)PCL/NAP-␤CD-ICnanoﬁbers.

anyinteractionbetweenPCLand NAP-␤CD-ICin terms ofpeak shifts,wecannotrule-outsuchcase.Itisduetothefactthatthe signalfromtheFTIRspectraoriginatesfromtwospecies,namely, interactionandnon-interactionofPCLwithNAP-␤CD-IC.Alsowe haveonly4%ofNAPinPCLandhencethesignalispredominantly comingfromPCL.

X-raydiffractionpatternsofpurePCL,PCL/NAPand PCL/NAP-␤CD-ICnanofibrousmatsaregiveninFig.3b.NAPandNAP-␤CD-IC arecrystallinematerials,however,theXRDpatternsofPCL/NAP andPCL/NAP-_␤CD-ICnanofibrousmatsrevealedthat both NAP-␤CD-ICandNAPtransformedintoamorphousphasefollowingthe incorporationintothePCLnanofibrousmatrix.

In TGA thermograms,main degradation of NAP in PCL/NAP nanofiberswasfoundoutbetween200–250◦Cwhilefor PCL/NAP-␤CD-ICnanofibers, it wasfound outbetween 300–375◦C from theTGAanalysis.TheTGAthermograminFig.4brevealsthatPCL nanofibersshowmaindegradationatabout420◦C.BylookingTGA thermograms,itispossibletoclaimtheexistenceofNAPmolecules andNAP-␤CD-ICinthePCL/NAP-␤CD-ICnanofibers.Inbothcases, forPCL/NAP andPCL/NAP-␤CD-ICnanofibers,two weightlosses wereseenafter100◦Cwhichareindependentfromwaterlosses.

We studied the release profiles of NAP from PCL/NAP and PCL/NAP-_␤CD-ICnanofibersinbuffersolutionforabout20htime period by HPLC. The amount of PCL/NAP and PCL/NAP-␤CD-IC nanofibersusedforthereleasestudywasadjustedaccordinglyin ordertohavethesameamountofNAPinthesesamples.Forboth samples,afterslightburstingofthedrug,slowreleasebehavior wasobservedfor12hperiod.Then,NAPshowedsustainedrelease profilesduetothebalancedconditionssuchashavingsame diffu-sionresistancefordifferenttimeintervals[54].Thereleaseprofiles revealedthatPCL/NAP-␤CD-ICnanofibroussystemhasmorethan twotimeshigherreleaseratethanPCL/NAPnanofibroussystem whichisverypromisingresultforthedrugdeliverypurpose(Fig.7). ItalsorevealedthattheformationofNAP-␤CD-IChelpedNAPto releasefromnanofibrousmatmucheasierwhichisquitevitalin drugdeliveryapplications.Asitwasprovenbyoursolubilitytest, CD-ICformationhelpsNAPtodissolveinwatermuchbetterwhich showsitseffectonthereleasebehaviorofNAPafterincorporated intonanofibrousmat.Theeasyandhigherreleaseofdrugis impor-tantfor somespecifictargetsindrugdelivery.It iswell-known

Fig.7.ReleaseproﬁleofNAPfromPCL/NAPandPCL/NAP-␤CD-ICnanoﬁbrousmats byHPLCwithstandarddeviations(eachanalysisrepeated3times,n=3).

(6)

20 M.F.Canbolatetal./ColloidsandSurfacesB:Biointerfaces115(2014)15–21 phenomenathatnanoﬁberscanenhancethereleasebehaviorof

drugswiththeirhighsurfaceareatovolumeratio[55,56].Inthis study,thepositiveeffectofinclusioncomplexformationonthe releaseproﬁlesofNAPinadditiontonanoﬁberincorporationhas beendemonstratedandthismaybeapromisingresultfordesigning noveldrugdeliverysystems.

4. Conclusions

Themainideabehindthisstudywastocomparetheefficiencies oftwodifferentdeliverysystemsfortheNAP;direct incorpora-tionofNAPandNAPincludedcomplexincorporationfollowingIC formationwith␤CDintoelectrospunPCLnanofibers.Initially,the formationofNAP-␤CD-ICwasstudiedandthentheincorporation ofNAPandNAP-_␤CD-ICintoPCLnanofiberswasperformedvia electrospinning.ThereleaseperformanceoftheNAPwasincreased morethantwotimesincaseofPCL/NAP-␤CD-ICnanofiberswhen comparedwithPCL/NAP nanofibers.Thus, it is understoodthat incorporationofNAP-␤CD-ICinapolymericnanofibroussystem stillpreservetheimprovedsolubilityeffectofCD-IContherelease rateofNAPandprovidesastableenvironmentforit.

Acknowledgment

StatePlanningOrganization(DPT)ofTurkeyisacknowledged for the support of UNAM-National Nanotechnology Research Center, BilkentUniversity. Dr. T. Uyar acknowledges TUBITAK-The Scientiﬁc and Technological Research Council of Turkey (project#111M459)andEUFP7-PEOPLE-2009RGMarieCurie-IRG (NANOWEB,PIRG06-GA-2009-256428)andTheTurkishAcademy ofSciences-OutstandingYoungScientistsAwardProgram (TUBA-GEBIP)for funding the research. A. Celebioglu acknowledges TUBITAK-BIDEBforthenationalPh.D.studyscholarship.

References

[1]T.M.Allen,P.R.Cullis,Drugdeliverysystems:enteringthemainstream,Science 303(2004)1818–1822.

[2]J.L. Frandsen, H. Ghandehari, Recombinant protein-based polymers for advanceddrugdelivery,Chem.Soc.Rev.41(2012)2696–2706.

[3]J.H.Jung,J.H.Lee,J.R.Silverman,G.John,Coordinationpolymergelswith impor-tantenvironmentalandbiologicalapplications,Chem.Soc.Rev.42(2013) 924–936.

[4]Y.Zhang,J.Zhang,T.Jiang,S.Wang,Inclusionofthepoorlywater-soluble drugsimvastatininmesocellularfoamnanoparticles:drugloadingandrelease properties,Int.J.Pharm.410(2011)118–124.

[5]R.Machin,J.R.Isasi,I.Vélaz,␤-Cyclodextrinhydrogelsaspotentialdrugdelivery systems,Carbohydr.Polym.87(2012)2024–2030.

[6]J.Kowapradit,A.Apirakaramwong,T.Ngawhirunpat,T.Rojanarata,W. Sajom-sang, P.Opanasopit, MethylatedN-(4-N,N-dimethylaminobenzyl)chitosan coatedliposomesfororalproteindrugdelivery,Eur.J.Pharm.Sci.47(2012) 359–366.

[7]E.D‘Aurizio,P.Sozio,L.S.Cerasa,M.Vacca,L.Brunetti,G.Orlando,A.Chiavaroli, R.J.Kok,W.E.Hennink,A.D.Stefano,Biodegradablemicrospheresloadedwith ananti-parkinsonprodrug:aninvivopharmacokineticstudy,Mol.Pharm.8 (2011)2408–2415.

[8]J.B.Wolinsky,Y.L.Colson,M.W.Grinstaff,Localdrugdeliverystrategiesfor can-certreatment:gels,nanoparticles,polymericﬁlms,rods,andwafers,J.Control. Release159(2012)14–26.

[9]S.D.Koker,R.Hoogenboom,B.G.D.Geest,Polymericmultilayercapsulesfor drugdelivery,Chem.Soc.Rev.41(2012)2867–2884.

[10]J.H.Kang,D.H.Oh,Y.-K.Oh,C.S.Yong,H.-G.Choi,Effectsofsolidcarriersonthe crystallineproperties,dissolutionandbioavailabilityofﬂurbiprofeninsolid self-nanoemulsifyingdrugdeliverysystem(solidSNEDDS),Eur.J.Pharm. Bio-pharm.80(2012)289–297.

[11]J.M.Rabanel,V.Aoun,I.Elkin,M.Mokhtar,P.Hildgen,Drug-loaded nanocarri-ers:passivetargetingandcrossingofbiologicalbarriers,Curr.Med.Chem.19 (2012)3070–3102.

[12]L.H.Reddy,J.L.Arias,J.Nicolas,P.Couvreur,Magneticnanoparticles:design characterizationtoxicitybiocompatibilitypharmaceuticalbiomedical applica-tions,Chem.Rev.112(2012)5818–5878.

[13]T.Loftsson,M.E.Brewster,Cyclodextrinsasfunctionalexcipients:methodsto enhancecomplexationefﬁciency,J.Pharm.Sci.101(2012)3019–3032.

[14]F.V.d.Manakker,T.Vermonden,C.F.V.Nostrum,W.E.Hennink, Cyclodextrin-based polymeric materials: synthesis, properties, and pharmaceutical/ biomedicalapplications,Biomacromolecules10(2009)3157–3175. [15]W.Gu,C.Wu,J.Chen,Y.Xiao,Nanotechnologyinthetargeteddrugdeliveryfor

bonediseasesandboneregeneration,Int.J.Nanomed.8(2013)2305–2317. [16]R. Cheng,F. Meng,C.Deng, H.-A.Klok,Z.Zhong,Dual andmulti-stimuli

responsivepolymericnanoparticlesforprogrammedsite-speciﬁcdrug deliv-ery,Biomaterials34(2013)3647–3657.

[17]M.A.Petersen,M.A.Hillmyer,E.Kokkoli,Bioresorbablepolymersomesfor tar-geteddeliveryofcisplatin,Bioconjug.Chem.24(2013)533–543.

[18]N.Sadlej-Sosnowska,L.Kozerski,E.Bednarek,J.Sitkowski,Fluorometricand NMRstudiesofthenaproxen-cyclodextrininclusioncomplexesinaqueous solutions,J.Incl.Phenom.Macrocycl.Chem.37(2000)383–394.

[19]A.Banik,P.Gogoi,M.D.Saikia,Interactionofnaproxenwith␤-cyclodextrinand itsderivatives/polymer:experimentalandmolecularmodelingstudies,J.Incl. Phenom.Macrocycl.Chem.72(2012)449–458.

[20]M.Cirri,F.Maestrelli,G.Corti,S.Furlanetto,P.Mura,Simultaneouseffectof cyclodextrincomplexation,pH,andhydrophilicpolymersonnaproxen solubi-lization,J.Pharm.Biomed.Anal.4(2)(2006)126–131.

[21]H.L.Ramírez,R.Cao,A.Fragoso,J.J.Torres-Labandeira,A.Dominguez,E.H. Schacht, M.Ba ˜nos, R. Villalonga, Improvedanti-inﬂammatory properties fornaproxenwithcyclodextrin-graftedpolysaccharides,Macromol.Biosci.6 (2006)555–561.

[22]S.Tungprapa,I.Jangchud,P.Supaphol,Releasecharacteristicsoffourmodel drugsfromdrug-loadedelectrospuncelluloseacetateﬁbermats,Polymer48 (2007)5030–5041.

[23]E.-R.Kenawy,F.I.Abdel-Hay,M.H.El-Newehy,G.E.Wnek,Processingof poly-mernanoﬁbersthroughelectrospinningasdrugdeliverysystems,Mater.Chem. Phys.113(2009)296–302.

[24]T.J.Sill,H.A.v.Recum,Electrospinningapplicationsindrugdeliveryandtissue engineering,Biomaterials29(2008)1989–2006.

[25]M.J.Kutyla,L.K.Lambert,N.M.Davies,R.P.McGeary,P.N.Shaw,B.P.Ross, Cyclodextrin-crosslinkedpoly(acrylicacid):synthesis,physicochemical char-acterizationandcontrolledreleaseofdiﬂunisalandﬂuconazolefromhydrogels, Int.J.Pharm.444(2013)175–184.

[26]M.Jug,F.Maestrelli,P.Mura,Nativeandpolymeric␤-cyclodextrinsin per-formanceimprovementofchitosanﬁlmsaimedforbuccaldeliveryofpoorly solubledrugs,J.Incl.Phenom.Macrocycl.Chem.74(2012)87–97.

[27]T. Vigh, T. Horváthová, A. Balogh, P.L. Sóti, G. Drávavölgyi, Z.K. Nagy, G. Marosi,Polymer-freeand polyvinylpyrrolidone-basedelectrospunsolid dosageformsfordrugdissolutionenhancement,Eur.J.Pharm.Sci.49(2013) 595–602.

[28]F.Kayaci,O.C.O.Umu,T.Tekinay,T.Uyar,Antibacterialelectrospun polylac-ticacid(PLA)nanoﬁbrouswebsincorporatingtriclosan/cyclodextrininclusion complexes,J.Agric.FoodChem.61(2013)3901–3908.

[29]F.Kayaci,Y.Ertas,T.Uyar,Enhancedthermalstabilityofeugenolbycyclodextrin inclusioncomplexencapsulatedinelectrospunpolymericnanoﬁbers,J.Agric. FoodChem.61(2013)8156–8165.

[30]F.Kayaci,T.Uyar,Encapsulationofvanillin/cyclodextrininclusioncomplexin electrospunpolyvinylalcohol(PVA)nanowebs:prolongedshelf-lifeandhigh temperaturestabilityofvanillin,FoodChem.133(2012)641–649.

[31]T.Uyar,Y.Nur,J.Hacaloglu,F.Besenbacher,Electrospinningoffunctional poly(methyl methacrylate) (PMMA) nanoﬁbers containing cyclodextrin-mentholinclusioncomplexes,Nanotechnology20(2009)125703.

[32]T.Uyar,J.Hacaloglu,F.Besenbacher,Electrospunpolyethyleneoxide(PEO) nanoﬁberscontainingcyclodextrininclusioncomplex,J.Nanosci.Nanotechnol. 5(2011)3949–3958.

[33]T.Uyar,J.Hacaloglu,F.Besenbacher,Electrospunpolystyreneﬁberscontaining hightemperaturestablevolatilefragrance/ﬂavorfacilitatedbycyclodextrin inclusioncomplexes,React.Funct.Polym.3(2009)145–150.

[34]J.Szejtli,Introductionandgeneraloverviewofcyclodextrinchemistry,Chem. Rev.98(1998)1743–1754.

[35]E.D.Valle,Cyclodextrinsandtheirusesareview,ProcessBiochem.39(2004) 1033–1046.

[36]S.T. Yohe,V.L.M.Herrera,Y.L.Colson,M.W.Grinstaff, 3D superhydropho-bicelectrospunmeshesasreinforcementmaterialsforsustainedlocaldrug delivery against colorectal cancer cells, J. Control. Release 162 (2012) 92–101.

[37]X. Li, M.A. Kanjwal, L. Lin, I.S. Chronakis, Electrospun polyvinyl-alcohol nanoﬁbersasoralfast-dissolvingdeliverysystemofcaffeineandriboﬂavin, ColloidsSurf.B103(2013)182–188.

[38]R.Murugan,S.Ramakrishna,Nano-featuredscaffoldsfortissueengineering:a reviewofspinningmethodologies,TissueEng.12(2006)435–447.

[39]T. Miletic, K. Kyriakos, A. Graovac, S. Ibric, Spray-dried voriconazole– cyclodextrincomplexes:solubility,dissolutionrateandchemicalstability, Car-bohydr.Polym.98(2013)122–131.

[40]H.S.Mahajan,M.H.Pingale,K.M.Agrawal,Solubilityanddissolution enhance-ment ofsaquinavirmesylatebyinclusioncomplexationtechnique,J.Incl. Phenom.Macrocycl.Chem.76(2013)467–472.

[41]J.Blanco,J.L.Vila-Jato,F.Otero,S.Anguiano,Inﬂuenceofmethodofpreparation oninclusioncomplexesofnaproxenwithdifferentcyclodextrins,DrugDev.Ind. Pharm.17(1991)943–957.

[42]S.Junco,T.Casimiro,N.Ribeiro,M.N.D.Ponte,H.C.Marques,Acomparative study ofnaproxen–betacyclodextrincomplexespreparedbyconventional methodsandusingsupercriticalcarbondioxide,J.Incl.Phenom.Macrocycl. Chem.44(2002)117–121.

(7)

[43]E.Marco-Urrea,M.Pérez-Trujillo,P.Blánquez,T.Vicent,G.Caminal, Biodegra-dationoftheanalgesicnaproxenbytrametesversicolorandidentiﬁcationof intermediatesusingHPLC-DAD-MSandNMR,Bioresour.Technol.101(2010) 2159–2166.

[44]S.Divakar,M.Maheswaran,Structuralstudiesoninclusioncompoundsof ␤-cyclodextrinwithsomesubstitutedphenols,J.Incl.Phenom.Macrocycl.Chem. 27(1997)113–126.

[45]H.E.Grandelli,B.Stickle,A.Whittington,E.Kiran,Inclusioncomplexformation of␤-cyclodextrinandnaproxen:astudyonexothermiccomplexformationby differentialscanningcalorimetry,J.Incl.Phenom.Macrocycl.Chem.77(2013) 269–277.

[46]M.Valero,B.I.Perez-Revuelta,L.J.Rodriguez,EffectofPVPK-25ontheformation ofthenaproxen:␤-cyclodextrincomplex,Int.J.Pharm.253(2003)97–110. [47]L.-F.Chen,Q.Shen,J.-P.Shen,D.-T.Shi,T.Chen,H.-R.Yu,Studiesandcomparison

oftheliquidadsorptionandsurfacepropertiesof␣-,␤-,and␥-cyclodextrins byFTIRandcapillaryrisemethod,ColloidsSurf.A411(2012)69–73. [48]A.Harada,M.Okada,J.Li,M.Kamachi,Preparationandcharacterizationof

inclu-sioncomplexesofpoly(propyleneglycol)withcyclodextrins,Macromolecules 28(1995)8406–8411.

[49]K.Tapan,T.K.Dash,V.B.Konkimalla,Poly-␧-caprolactonebasedformulations fordrugdeliveryandtissueengineering:areview,J.Control.Release158(2012) 15–33.

[50]C.Wu,T.F.Jim,Z.Gan,Y.Zhao,S.Wang,Aheterogeneouscatalytickineticsfor enzymaticbiodegradationofpoly(␧-caprolactone)nanoparticlesinaqueous solution,Polymer41(2000)3593–3597.

[51]A.Elzubair,C.N.Elias,J.C.M.Suarez,H.P.Lopes,M.V.B.Vieira,Thephysical char-acterizationofathermoplasticpolymerforendodonticobturation,J.Dent.34 (2006)784–789.

[52]S.A.Catledge,W.C.Clem,N.Shrikishen,S.Chowdhury,A.V.Stanishevsky,M. Koopman,Y.K.Vohra,Anelectrospuntriphasicnanoﬁbrousscaffoldforbone tissueengineering,Biomed.Mater.2(2007)142–150.

[53]L.Ghasemi-Mobarakeh,M.P.Prabhakaran,M.Morshed,M.H.Nasr-Esfahani, S.Ramakrishna,Bio-functionalizedPCLnanoﬁbrousscaffoldsfornervetissue engineering,Mater.Sci.Eng.C30(2010)1129–1136.

[54]X.Sheng,L.Fan,C.He,K.Zhang,X.Mo,H.Wang,VitaminE-loadedsilkﬁbroin nanoﬁbrousmatsfabricatedbygreenprocessforskincareapplication,Int.J. Biol.Macromol.56(2013)49–56.

[55]X.-Z.Sun,G.R.Williams,X.-X.Hou,L.-M.Zhu,Electrospuncurcumin-loaded ﬁberswithpotentialbiomedicalapplications,Carbohydr.Polym.94(2013) 147–153.

[56]D.W.Chen,Y.-H.Hsu,J.-Y.Liao,S.-J.Liu,J.-K.Chen,S.W.-N.Ueng, Sustain-ablereleaseofvancomycin,gentamicinandlidocainefromnovelelectrospun sandwich-structuredPLGA/collagennanoﬁbrousmembranes,Int.J.Pharm.430 (2012).