Synthesis and Characterization of Porphyrin Nanoparticles to model Heme Protein Iron Coordination Graham Beaton , Samuel Pazicni gct5@wildcats.unh.edu; Parsons Hall, 23 Academic Way, Durham NH 03824 Monomer Characterization Introduction Nanoparticle Collapse Heme proteins serve many vital functions in biological systems, including oxygen delivery, electron transport and redox catalysis. Their function depends largely on the reactivity of the iron metal center, which is not only influenced by covalent interactions of the Heme prosthetic group but also by interactions of the secondary coordination sphere. A Porphyrin cored nanoparticle was synthesized to model heme cofactor iron coordination and the effects of the Heme’s secondary coordination sphere on the iron center’s reactivity were explored. Several methacrylate monomers were synthesized and subjected to RAFT polymerization with porphyrin cored CTA to create a functionalized porphyrin cored polymer. An Iron insertion incorporated the metal center into the porphyrin cored polymer and upon photodimerization with 350 nm light, it was collapsed into a nanoparticle where functionalized chains allowed for a tunable microenvironment around the metal center. Figure 1: 400 MHz 1H NMR of AMMA Figure 2: 400 MHz 1H NMR of PFPMA = = = H2O Figure 6: Photodimerization of AMMA units with 350 nm light facilitating nanoparticle collapse Nanoparticle collapse was monitored by UV-Vis (5 min. increments) and Gel Permeation Chromatography (10 min. increments) Hypochromic shift in Anthracene bands and increased retention time are indicative of successful collapse Polymer Synthesis and Characterization Monomer Synthesis Figure 7: UV-Vis of PCNP collapse Figure 8: GPC of PCNP collapse (5) Fe(III) center was reduced to Fe(III) with Dithionite and binding studies were run with Imidazole and Carbon monoxide as axial bound ligands. This binding was monitored with UV-Vis and the soret shifts were consistent with reduced and small molecule bound species. Monomers were chosen based on their susceptibility to RAFT polymerization, ability to facilitate nanoparticle collapsing and functionality to afford realistic protein microenvironments. Acrylamide System Incorporation of PFPMA and AMMA units allow for photodimerization collapse mechanism as well as post polymerization modifications via amine activated ester functionality Figure 9: UV-Vis spectroscopy of reduced and ligated PCNP species (1) C Future Work With the methacrylate polymer system made and conditions understood further work can be done to probe the reactivity of the metal center. The activated ester functionality of the PFPMA units afford a polymer susceptible to many post polymerization modifications, this will allow the secondary coordination sphere to be tuned towards hydrophobic, hydrophilic or hydrogen bond rich microenvironments. This will also be accomplished with further development of the acrylamide based polymer system. B B (2) A C A Methacrylate System Polymer Backbone (B) Figure 1: 400 MHz 1H NMR of Por (PFPMA Co AMMA) at 12% AMMA incorporation Iron Insertion Acknowledgements (3) Deprotonation of the pyrrole Nitrogens with 2,4,6-colidine afford the free base porphyrin core, upon metalation with Iron Bromide, the iron cored star polymer is synthesized and the reaction is monitored via UV-Vis spectroscopy. Dr. Erik Berda Dr. Kyle J. Rodriguez Dr. Roy Planalp Hamel Center SURF Fellowship UNH Instrumentation Center Dr. Patricia Wilkinson Metalated star polymer References (4) Rodriguez, K. J.; Hanlon, A. M.; Lyon, C. K.; Cole, J. P.; Tuten, B. T.; Tooley, C. A.; Berda, E. B.; Pazicni, S., Inorg. Chem. 2016, 55 (19), 9493-9496. (3) Lyon, C. K.; Prasher, A.; Hanlon, A. M.; Tuten, B. T.; Tooley, C. A.; Frank, P. G.; Berda, E. B., Polym. Chem. 2015, 6 (2), 181-197. (4) Das, A.; Theato, P., Chem Rev 2016, 116 (3), 1434-95. (5) Levere, M. E.; Ho, H. T.; Pascual, S.; Fontaine, L., Polym. Chem. 2011, 2 (12), 2878-2887. (6) Beil, J. B.; Zimmerman, S. C., Macromolecules 2004, 37 (3), 778-787. (7) Dixon, D. W.; Barbush, M.; Shirazi, A., Inorg. Chem. 1985, 24 (7), 1081-1087. (8) Nakamura, M.; Ikeue, T.; Fujii, H.; Yoshimura, T., J. Am. Chem. Soc. 1997, 119 (27), 6284-6291. Free-base star polymer Figure 3: UV-Vis spectrum of PCSP pre and post iron insertion Acrylamide monomers (1),(2) Purified via Liquid-Liquid extraction, Organic phase washed with H2O and Brine, dried over MgSO4 and isolated via column chromatography 2:1 Hexanes , Ethyl Acetate (3) Organic phase washed with H2O, Brine, Sodium Bicarbonate and 1M HC (4) Vaccum filtered to remove DCU, isolated via column chromatography 5:1 Toluene , Ethyl Acetate Hyperchromic shift of alpha, Beta Q bands and the hypochromic shift of soret band are consistent with metalation of porphyrin with Fe(II)