Thesis (MSc (Chemistry and Polymer Science))--University of Stellenbosch, 2009. / Thesis submitted in partial fulfilment for the degree of Master of Science (polymer science) at Stellenbosch University / ENGLISH ABSTRACT: The goal of this study was to synthesize hybrid hydrogels via a chemical crosslinking mechanism through use of chain end functional poly(N-vinyl pyrrolidone)(PVP) with various topologies. The crosslinking chemistries should be benign in nature i.e. at physiological pH ranges and at 37 °C. The degradation products should be biologically tolerable and renal clearance should be possible (< 30 000 g/mol PVP0. PVP of various topologies, controlled molar mass and quantitative chain end functionality was obtained via Reversible Addition Fragmentation chain Transfer (RAFT) mediated polymerization (PDI = 1.1- 1.4). The synthesized polymers were chain end functionalized to introduce thiol or aldehyde moieties. Thiol chain ends were obtained through post polymerization modification of xanthate functional PVP with either aminolysis or reduction. The aldehyde moiety was obtained by post polymerization modification of xanthate end functional PVP with sequential hydrolysis and thermolysis. Thiol functional four arm star PVP was reacted with acrylate difunctional poly(ethylene glycol) (DIAC PEG) crosslinker under standard Michael addition conditions. In order to obtain thioether crosslinked hydrogels from tetra functional star PVP molecules it was found that a minimum thiol functionalization of 30% and a molar ratio of acrylate:thiol of 1:1.1 is required. The Schiff base reaction was used to synthesize imine or secondary amine (after reduction) crosslinks with the lysine residues on either lysozyme or bovine serum albumin (BSA) or the primary amines of bis-(2-amino ethyl)amine). Hydrogels were obtained from aldehyde functionalized PVP molecules with a fraction of functional aldehyde chain ends of 0.88 for difunctional molecules and 0.50 for tetra functional star PVP molecules with lysozyme or BSA crosslinkers. The reaction rate was favoured by lowered pH (<6.0) and an optimum molar ratio of amine : aldehyde of 1:0.8. Hydrogels were analyzed by equilibrium swelling calculations to determine the molar mass between crosslinks and the estimated pore size. In both crosslinking systems the properties of the formed hydrogels were seen to be affected by molar ratio used and by the topology of the crosslinking agent. PVP BSA and PVP PEG hydrogels were tested for 24 h and 48 h cell viability by using H9C2 myoblast cells. A concentration range of 0.25 x 10(2) to 0.01 g/mL was studied. Cell mortality was tested by Trypan blue staining and results were verified with MTT assay. A very low cell death precentage (<37%)was observed. Cells even appeared to experience a stimulatory effect after 48 h of exposure at low concentrations of PVP PEG hydrogel treatments. The properties of the formed hydrogel could be tuned by the molar mass ratios of PVP and crosslinker. The functionality of the crosslinker directly affects the molar mass between crosslinks and thus indirectly the degradation profile. It was concluded that PVP molecules with various topologies, well-defined molar masses and chain end functionality could be obtained via RAFT mediated polymerization. Obtained polymers were successfully modified and crosslinked to obtain hydrogels with stoichiometrically tuneable properties i.e. initial swelling ratio, degradation time, molar mass between crosslinks. The hydrogels had very positive cell viability results that would definitely justify further research into these materials as “tissue-mimetic” materials. / AFRIKAANSE OPSOMMING: Die doel van die studie is om poli(N-viniel pirollidoon) (PVP) gebaseerde hibried hidrogelle te sintetiseer deur middel van kovalente kruisbindings met toepaslike kruisverbinder molekules. Die chemiese reaksies betrokke in die vorming van hierdie kovalente kruisbindings moet gematig van aard wees, by fisiologiese pH en by 37 °C plaasvind. Die degradasieprodukte van die hibried-hidrogel moet biologies verdraagsaam en ook uitskeibaar deur die endokrinologiese sisteem wees. PVP van verskillende topologieë, beheerde molêre massa en kwantitatiewe kettingendfunksionaliteit is berei deur ‘n omkeerbare addisie-fragmentasiekettingoordrag (OAFO)-beheerde polimerisasieproses (PDI = 1.1-1.4). Xantaat-kettingend-PVP is aangepas na thiol of aldehied kettingendfunksies. Thiolendfunksies is verkry deur middel van ‘n aminolisasie-reaksie. Xantaat kettingend-PVP is stapsgewys gemodifiseer deur hidroliese en verhittingstappe om die aldehied ketting-endfunksionaliteit te bekom. Thiol ketting-endfunksionele vier-armige ster-PVP is kovalent gebind aan difunksionele poly(etileen glikol) deur middel van die Michael-addisiereaksie. PVP PEG hidrogelle het slegs gevorm met vier-armige ster-PVP molekules wat oor ‘n minimum van 30 % thiol-funksionaliteit beskik het en ‘n optimale molêre massa verhouding van 1:1.1 vir ankrilaat to thiol. Die Schiff-basisreaksie is gebruik om hidrogel te sintetiseer wat met imiene of amiene (na redusering) kovalente bindings gekruisbind is. In hierdie sisteme het hidrogel slegs gevorm as die aldehied-PVP molekules oor ‘n fraksie funksionele kettingend-waarde van 0.88 vir dialdehied-PVP molekules en 0.5 vir vier armige ster-PVP molekules beskik het. Die reaksie snelheid van die Schiff-basis kovalente bindings is bevoordeel deur die pH te verlaag (≤ 6.0) en ‘n gunstige molêre massa verhouding van 1:0.8 vir die nukleofiel teen oor die akseptor molekule is waargeneem. Ewewigswel berekeninge is gebruik om die molêre massa tussen kruisbindings en die gemiddelde benaderde porieë binne die drie-dimensionele interne struktuur van die hydrogel te bepaal. Die seltoksisiteit van PVP-BSA en PVP-PEG hidrogelle is oor 24 h en 48 h in die teenwoordigheid van H9c2 mioblast-selle getoets. Die hydrogel behandelings is uitgevoer in ‘n konsentrasie reeks van 0.25 x 10(2) tot 0.01 g/mL. Selmortaliteit is getoets deur ‘n Trypan-blou verkleuringstudie. Hierdie resultate is ondersteun deur MTT sel-lewensvatbaarheidstoetse. ‘n Lae selmortaliteit (≤ 37 %) is waargeneem en, opspraakwekkend, het van die selle na 48 h verhoogde vitaliteit getoon in die teenwoordigheid van lae konsentrasies PVP-PEG hidrogelle. Dit is bevind dat hidrogel eienskappe deur stoichiometriese molêre massa verhoudings asook die keuse in die topologie van kruisverbinder beïnvloed word. Hierdie eienskappe het ‘n direkte effek op die degradasieprofiel van die gevormde hidrogel. Samevattend dus is PVP molekules met ‘n variasie van topologieë, spesifieke molêre massas en kettingfunksionalitete deur middel van OAFO-gemedieerde polimerisasies gesintetiseer. Xantaatkettingendfunksionele PVP-molekules kon suksesvol omgeskakel word na die kettingendfunksionaliteit van ons keuse om ‘n hibriedhidrogel met stoichiometries-manupileerbare eienskappe te sintetiseer. Die positiewe sel-lewensvatbaarbheidstudie resultate staaf verdere ondersoeke in hierdie PVPgebaseerde hibried hidrogelmateriaal as ‘n weefsel nabootsingsmateriaal.

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