Table Of ContentBestMasters
Springer awards „BestMasters“ to the best master’s theses which have been com-
pleted at renowned universities in Germany, Austria, and Switzerland.
Th e studies received highest marks and were recommended for publication by
supervisors. Th ey address current issues from various fi elds of research in natural
sciences, psychology, technology, and economics.
Th e series addresses practitioners as well as scientists and, in particular, off ers guid-
ance for early stage researchers.
Tamara Bernadette Aigner
Photopolymerizable
Porous Polyorgano-
phosphazenes
Degradable Matrices for
Tissue Engineering
Tamara Bernadette Aigner
Bayreuth, Germany
BestMasters
ISBN 978-3-658-09319-8 ISBN 978-3-658-09320-4 (eBook)
DOI 10.1007/978-3-658-09320-4
Library of Congress Control Number: 2015933839
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Institutsprofil
Das Institut fu¨r Chemie der Polymere (ICP) an der Johannes Kepler Universita¨t (JKU)
Linz, dasichvon2007anneuaufbauendurfte, fokussiertseineForschungaufdieEntwick-
lungfunktionalerPolymerefu¨rdenEinsatzinDrugDeliveryundControlledRelease,imBe-
reichTissueEngineering,inHautpflegemittelnundWaschmitteln,alsTra¨gervonKatalysatoren
sowiealsFestphaseninChromatographieundExtraktion. Vorallemwerdenfu¨reinigedieser
ZweckebioabbaubarePolyphosphazenemaßgeschneidert.BionikundBiomimetikspieleneine
RollebeieinemProjekt,beidemesumneuartigeDNA-Analogageht,alsoPolymere,diespa¨ter
einmalinderAntisense-Therapieeingesetztwerdenko¨nnten. Polymereausnachwachsenden
RohstoffenkommenauchzumEinsatz. AußerdemarbeitetdasICPaneinigenProjekten,bei
denenesumindustrielleAspektevonPolymerengeht.Sosollenz.B.Kunststoffefu¨rzuku¨nftige
thermischeSolarkollektorenu¨berchemischeKopplungenvonStabilisatorenandiePolymerket-
tenderartmodifiziertwerden,dassdieKunststoff-Kollektorendeutlichla¨ngereHaltbarkeiten
aufweisen.
DasICPbefindetsichmittlerweileineinerbesondersfruchtbarenUmgebung,miteinem
Polymerforschungsschwerpunkt an der JKU, mit neu hinzugekommenen Kunststofftechnik-
Instituten,miteinerstarkenKunststoff-Industrieinna¨chsterNa¨heundmiteinerneuerrichteten
Medizinfakulta¨timJahr2014.
Univ.-Prof.Dr.OliverBru¨ggemann
Institutsvorstand
Institutfu¨rChemiederPolymere(ICP)
V
Acknowledgement
Thismasterthesiswouldnothavecomeoffwithoutthecontributionofmanypeople.First
I would like to thank my supervisor Univ.-Prof. Dr. Oliver Bru¨ggemann for giving me the
chancetoworkonthisgreatproject. FurthermoreIthankmy”TissueEngineeringTeam”:Dr.
IanTeasdaleforhisexcellentguidanceandhisirrepressibleoptimismnomatterwhathappened,
Aitziber Iturmendi for her enthusiasm and spreading good cheer throughout the laboratory,
Sandra Wilfert, PhD for her valuable ideas and lending me her ear everytime and Dipl.Ing.
MartinaPrambauerforintroducingmesonicelytothelaboratory.Ialsowanttothanktheother
membersoftheinstitutefortheirhelpandthegreattime.
MoreoverIthankourprojectpartnerswithoutthemtheprojectwouldnothavebeenpossi-
ble.MyspecialthanksgotoDr.FlorianHildnerandMariaRigaufromRedCrossandGbenga
Olawale,MScfromBioMedzetfortheirgreatbiologicalcontributionfundedbyRegio13and
toTransfercenterfu¨rKunststofftechnikfortheirfinancialsupport.
ThankstoDr. WolfgangScho¨ffbergerforthesolidstatemeasurement. TheNMRexperi-
mentswereperformedattheUpperAustrian-SouthBohemianResearchInfrastructureCenter
inLinz,co-financedbytheEuropeanUnioninthecontextoftheproject”RERI-uasb”,EFRE
RU2-EU-124/100-2010(ETCAustria-CzechRepublic2007-2013,projectM00146).
Iwouldliketothankmyparents,mybrotherandmyfriendsfortheirconstantsupportand
understanding.LastbutnotleastIthankThomasforbeingatmysideallthetime.
TamaraBernadetteAigner
VII
Abstract
Tissueengineeringisanemergingfieldcombiningbiology,chemistryandmaterialssci-
encetorepair,restoreorregeneratelivingtissues.Itisbasedonamatrixconstruct,commonly
fromabiodegradablepolymer,whichmustfulfillthefollowingrequirements: mechanicalsta-
bilitytocarrythecellload,interconnectedporousstructurestoenablecellinfiltrationandcom-
munication,cellrecognitionsitestoallowpositivecellinteraction,non-toxicdegradationprod-
uctsandanadapteddegradationratesothatcellscanrebuildtheextracellularmatrix.
Inthisthesisweaimedtodevelopphotopolymerizablematricesbasedonbiodegradable
poly(organophosphazenes).Acrucialelementofallthesepolymerswasanallylgrouptoenable
thiol-ene photochemistry. Thereby thiol-bearing molecules were reacted with the polymers
to functionalize the poly(organophosphazenes) with several functional moieties to tailor the
chemical,physicalandbiologicalproperties. Moreoverthiol-enephotochemistrywasapplied
tocrosslinkthepolymerstoobtainmechanicallystrengthenednetworks.Blendingagentswere
added to the crosslinking media, which influence the performances of the final materials in
termsofmechanicalstability,degradationrateandbiologicalapplicability. Aninterconnected
porousstructurewasintroducedusinganovelphotocrosslinkingparticulate-leachingtechnique.
ThematerialswerecharacterizedthoroughlyusingNMRandFTIRspectroscopy, GPC,
CT,SEMandelementalanalysis.Degradabilityisamainissueintissueengineering,thussev-
eraldegradationstudieswereperformedandthestructureofthepolymerswasadjustedtotailor
degradability. Thisfinallyledtoanaminoacidcontainingsidechaintofitbiologicalsystems.
Inordertoassesstheirsuitabilityasmatricesfortissueengineering,cytotoxicity,adhesionand
proliferationtestswereperformed.Thesebiologicaltestsshowedpromisingresults,pavingthe
wayforpoly(organophosphazenes)intissueengineering.
IX
Contents
1 Aimofproject 1
2 Introduction 3
2.1 Tissueengineering . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
2.1.1 Basicprinciple . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
2.1.2 Polymerscaffolds . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
2.1.3 Vascularizationandsignalingmolecules . . . . . . . . . . . . . . . . . 7
2.2 Poly(organophosphazenes) . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8
2.3 Thiol-enephotochemistry . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
2.4 Poregeneration-matrixformation . . . . . . . . . . . . . . . . . . . . . . . . 10
2.5 Polymerfunctionalization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
3 Experimental 15
3.1 Materialsandmethods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
3.2 Trimersynthesis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16
3.2.1 Trimermodification. . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
3.3 Polymersynthesis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18
3.3.1 One-potpolymerization. . . . . . . . . . . . . . . . . . . . . . . . . . 18
3.3.2 Livingpolymerization . . . . . . . . . . . . . . . . . . . . . . . . . . 19
3.3.3 Macrosubstitution . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
3.4 Thiol-enephotochemistry . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
3.4.1 Functionalization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
3.4.2 Crosslinking. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26
3.5 Poregeneration-matrixformation . . . . . . . . . . . . . . . . . . . . . . . . 31
3.6 Degradationstudies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33
3.6.1 Trimer1c1andPolymer3c1 . . . . . . . . . . . . . . . . . . . . . . . 33
3.6.2 Polymer3p1,3p2and3p3 . . . . . . . . . . . . . . . . . . . . . . . . 33
3.6.3 Polymer5p1,5p3andTestsubstance3 . . . . . . . . . . . . . . . . . 33
4 Resultsanddiscussion 35
4.1 Trimers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35
4.1.1 Crosslinking. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 36
XI
4.1.2 Poregeneration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 38
4.1.3 Degradationstudies . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41
4.2 Polymers. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 42
4.2.1 Crosslinking. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45
4.2.2 Poregeneration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 46
4.2.3 Degradationstudies . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48
4.3 Blends . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 49
4.3.1 Crosslinking. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50
4.3.2 Poregeneration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 50
4.3.3 Degradationstudies . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52
4.3.4 Elementalanalysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54
4.4 Functionalization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54
4.4.1 Thioglycolicacid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55
4.4.2 Ethyl-3-mercaptopropionate . . . . . . . . . . . . . . . . . . . . . . . 55
4.4.3 Glutathione . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 56
4.4.4 Fluorescein . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
4.5 Biomedicaltests . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
4.5.1 Cytotoxicitystudies . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
4.5.2 Celladhesionandproliferationstudies . . . . . . . . . . . . . . . . . . 65
5 Conclusionandfuturework 67
Bibliography 69
XII
