pubs.acs.org/Macromolecules Published on Web 03/26/2010 r 2010 American Chemical Society Macromolecules 2010, 43, 3801–3808 3801 DOI: 10.1021/ma100292w Fabrication of “Clickable” Hydrogels via Dendron-Polymer Conjugates Huseyin Altin, Irem Kosif, and Rana Sanyal* Department of Chemistry, Bogazici University, Bebek, 34342, Istanbul, Turkey Received February 5, 2010; Revised Manuscript Received March 16, 2010 ABSTRACT: Functionalizable hydrogels are of great interest as three-dimensional (3D) scaffolds for cell growth and tissue engineering. The ability to covalently immobilize biologically relevant molecules with accurate control of their density within the hydrogel matrix is highly desirable. Dendron-polymer conjugates prepared via Huisgen type “click” reaction provides a unique precursor for reactive hydrogels. A family of dendron-polymer conjugates were prepared by coupling second- and third-generation alkyne appended polyester dendrons with linear poly(ethylene glycol) diazides, PEG2K and PEG6K. Controlled cross-linking of alkyne-functionalized dendron-polymer-dendron conjugates with a hydrophilic diazide provides hydrogels with gelation efficiencies greater than 80%. Excess leftover alkynes can be used to functionalize these hydrogels as desired. Fine tuning of degree of cross-linking and functionalization is demonstrated by immobilization of streptavidin. Introduction Recent years have witnessed remarkable advances in the area of synthesis of well-defined hydrogels. Interest in developing novel synthetic approaches in this area of research is fueled by the hydrogel-based emerging technologies ranging from off-the-shelf consumer products such as contact lenses and wound dressings to medical applications such as scaffolds for tissue engineering. The impact of synthetic methodologies in macromolecular engi- neering is rapidly shaping the field of hydrogel synthesis. 1-10 Advances in this area have seen developments in new cross- linking methodologies and synthesis of designer hydrogels that can be precisely functionalized. 11-20 Traditional approaches for incorporating entities such as small molecules, peptides, and large biomacromolecules like enzymes or growth factors into hydrogel matrix have relied upon encapsulation and physiabsorption. 21-29 Covalent immobilization techniques have been generally limited to attachment of the molecule of interest into a polymerizable macromonomer. 32-35 Postfunctionalization of hydrogels has been evaluated as an attractive alternative in recent years. 36-39 This approach relies on the presence of reactive functional groups in the hydrogel matrix that can undergo efficient functionalization under mild reaction conditions. Advent of “click” reactions 40-47 has dramatically influenced postpolymerization functional group transformations due to their near-quantitative conversions under mild reaction conditions. Indeed, recent works from Hilborn 48 and Hawker 49 groups have reported efficient synthesis of hydro- gels using Huisgen type cross-linking of telechelic PEGs with multivalent azide based cross-linkers. Since then, many other groups have reported synthesis of polymers 50-60 and hydrogels using the Huisgen click cycloaddition-based strategy. 61-75 One strategy in hydrogel preparation has been the use of triblock copolymers. 76-87 A triblock copolymer consisting of a hydrophilic block in between two hydrophobic blocks forms a network micellar structure under aqueous conditions. Polymeri- zable groups such as acrylate double bonds at the end of the hydrophobic blocks allow photo-cross-linking of the structure to provide chemically cross-linked hydrogels. In recent years, dendron-polymer-dendron-based triblock copolymers 88-91 have emerged as an attractive building block for synthesis of hydrogels. 92-98 Grinstaff and co-workers reported the synth- esis of biodegradable photo-cross-linkable hybrid dendritic- linear triblock copolymers and applied them to seal corneal lacerations. 99-101 We envisioned that the multivalent nature of the dendrons would allow one to utilize some of the reactive end groups for cross-linking to afford the gel, whereas the residual reactive groups would allow covalent postfunctionalization of the hydrogels with molecules of interest. In our design, we utilized the Huisgen-type copper-catalyzed click reaction 102,103 between biodegradable polyester dendrons 104-109 and biocompatible hy- drophilic linear PEG polymers to access dendron-polymer- dendron conjugates necessary for the hydrogel formation. Func- tionalization of the dendron periphery with alkyne groups affords reactive hydrogel precursors. While some of these alkyne groups are cross-linked using a bisazide to fabricate the hydrogel (second Huisgen type “click” reaction), the residual alkynes allow efficient covalent functionalization of the hydrogel matrix with molecules of interest via the third consecutive “click” reaction (Scheme 1). Experimental Section Materials. 2,2-Bis(hydroxymethyl)propionic acid (BMPA), Dowex X50WX2, propargyl alcohol, and 4-pentynoic acid were purchased from Alfa Aesar. All poly(ethylene glycol) were obtained from Fluka. All solvents were purchased from Merck and used as obtained without further purification unless other- wise noted. Azide-functionalized PEGs were synthesized ac- cording to literature procedures. 110 Syntheses of dendrons 1 and 2 are given in the Supporting Information. Methods. The monomer and copolymer characterizations involved 1 H NMR spectroscopy (Varian 400 MHz) and Fourier transform infrared (ATR-FTIR) spectroscopy (Thermo Fisher Scientific Inc. Nicolet 380). The molecular weights were esti- mated by gel permeation chromatography (GPC) analysis using a Viscotek GPCmax VE-2001 analysis system. PLgel (length/i.d. 300 mm  7.5 mm, 5 μm particle size) Mixed-C column was caliberated with polystyrene standards (1K-150K), using a refractive index detector. THF was used as eluent at a flow rate of 1 mL/min at 30 °C. Elemental analyses were obtained from Thermo Electron SpA FlashEA 1112 elemental analyzer *Corresponding author: e-mail rana.sanyal@boun.edu.tr; Tel þ90(212)3597613; Fax þ90(212)2872467.