DESIGN AND SYNTHESIS OF MULTIFUNCTIONAL POLYESTERS WITH 'PEPTIDE-LIKE' PENDANT GROUPS A Dissertation Presented to The Graduate Faculty of The University of Akron In Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy Ying Xu May, 2016 DESIGN AND SYNTHESIS OF MULTIFUNCTIONAL POLYESTERS WITH 'PEPTIDE-LIKE' PENDANT GROUPS Ying Xu Dissertation Approved: Accepted: ______________________ _______________________ Advisor Department Chair Dr. Abraham Joy Dr. Coleen Pugh ______________________ _______________________ Committee Member Dean of the College Dr. Matthew L. Becker Dr. Eric J. Amis ______________________ _______________________ Committee Member Dean of the Graduate School Dr. Coleen Pugh Dr. Chand Midha ______________________ _______________________ Committee Member Date Dr. Ali Dhinojwala ______________________ Committee Member Dr. Younjin Min ii ABSTRACT Current biomaterials including polylactic acid have good mechanical and biodegradable properties.1 But they are devoid of functional groups that enable integration with the cellular environment. We have designed a platform of modular multifunctional polyesters with pendant functional groups that address the lack of functional cues in current biomaterials.2 The polyesters were synthesized at room temperature by carbodiimide- mediated polymerization of pendant functionalized diols and succinic acid.3 The pendant groups were designed to mimic the side chains of peptides. It was shown that the physical properties of the polyesters can be modulated over a wide range by the selection of pendant groups. Orthogonal functionalization of the pendant groups with ligands such as fluorophores, poly (ethylene glycol) (PEG) or Arg-Gly-Asp (RGD) was shown. One specific application of functional polyester was the design of mussel inspired adhesives by incorporation of catechol groups into side chain of such polyesters. The first generation adhesive polyester showed the effect of 3,4-dihydroxyphenylalanine (DOPA) groups, but the adhesion strength on aluminum substrate decreased in wet conditions. The second generation adhesive polyester was a copolymer with soybean oil based monomer, coumarin and DOPA monomer. It showed good adhesion under both dry and wet conditions. Adhesion tests on porcine skin were also performed and the results demonstrated that our polymer had higher adhesion strength than the commercial fibrin glue. iii A second application was the fabrication of nanofiber mats through electrospinnin for extended dual release of model drugs. Two types of electrospun fiber mats were made. For one of them, two dyes (Rhodamine B and coumarin dye) were non-covalently encapsulated within the polymer fibers. For the other one, rhodamine B was covalently attached to the fibers, while coumarin dye was physically entrapped. For the fibers with non-covalently encapsulated dyes, the release of dyes over 90 days showed that the coumarin dye had a faster release profile compared to the rhodamine B dye. For the fibers where coumarin dye was encapsulated and rhodamine B was tethered, the release of coumarin dye was similar to the first one. The oxime bond of the covalently tethered rhodamine B was stable over 90 days, and there was no release of rhodamine B in 1×phosphate buffer saline(PBS) (pH = 7.4) which was similar to the studies by Raines and coworkers.4 Another project was to study the differentiation of stem cell into osteoblasts. Three polymers with 40% of carboxylic (COOH), amine (NH2), or hydroxyl (OH) pendant groups, were synthesized. The three polymers were used for examining the differentiation of mouse pre-osteoblast cell lines (MC3T3) into osteoblasts. Alkaline Phosphatase (ALP) staining and ALP acitivity of MC3T3 differentiated for 14 days were performed. From the ALP staining images, it was seen that the ALP production increased as COOH > blank > NH2 > TCPS (Tissue Culture treated Polystyrene) > OH. Alizarin Red staining and von Kossa staining of the MC3T3 differentiated for 21 days were performed. From Alizarin Red staining, polymer with COOH group demonstrated the largest influence on differentiation, it was seen that the polymers with COOH or NH2 provided the most differentiation. iv ACKNOWLEDGEMENTS I would like to express my sincere gratitude to my advisor, Professor Abraham Joy, for all his support, encouragement, and guidance during the past few years. I have benefited tremendously from his passion for science, profound knowledge and creative ideas. I would like to thank Professor Ali Dhinojwala for the collaboration and his valuable discussions and suggestions. I would like to take this opportunity to thank my dissertation committee members: Professor Coleen Pugh, Professor Matthew Becker, Professor Ali Dhinojwala and Professor Younjin Min for their valuable time and suggestions. I would like to thank Dharamdeep Jain for the collaboration. I appreciate the help from students in College of Polymer Science and Engineering, in particular Erin Childers, Dr. Shiwang Chen, Dr. Hao Sun, Qianhui Liu, Dr. Shuangyi Sun, Dr. Sachin Gokhale, John Swanson, Chao Peng and all my group members. Their kind friendship, support and encouragement have helped me to go through this special journey in my life. Finally, I would like to give my special thanks to my husband Jinjun Zhou, my sister and my parents for their sincere love and understanding, and support during my Ph.D. studies. v TABLE OF CONTENTS Page LIST OF TABLES .............................................................................................................. x LIST OF FIGURES ........................................................................................................... xi LIST OF SCHEMES ....................................................................................................... xvii CHAPTER I. INTRODUCTION ........................................................................................................... 1 1.1 Evolution of Biomaterials ......................................................................................... 1 1.2 Synthesis of Functional Aliphatic Polyesters ............................................................ 3 1.2.1 Functional aliphatic polyesters synthesized via condensation ........................... 4 1.2.2 Functional aliphatic polyesters made by ROP .................................................... 5 II. A LIBRARY OF MULTIFUNCTIONAL POLYESTERS WITH ‘PEPTIDE-LIKE’ PENDANT FUNCTIONAL GROUPS ............................................................................. 17 2.1 Abstract ................................................................................................................... 17 2.2 Introduction ............................................................................................................. 17 2.3 Experimental Section .............................................................................................. 19 2.3.1 Materials and instrumentation .......................................................................... 19 2.3.2 Synthesis of functional diols with different pendant Groups ........................... 20 2.3.3 Synthesis of functional polyesters .................................................................... 33 2.3.4 1H NMR of polyesters ...................................................................................... 34 2.3.5 Conjugation of dyes to p(mLys0.5-co-Propargyl0.5) ........................................ 39 vi t 2.3.6 Functionalization of p(mAla0.4-co-mAsp Bu0.4-co-Propargyl0.2) with PEG and RGD ........................................................................................................................... 40 2.4 Results and Discussion ........................................................................................... 43 2.4.1 Synthesis of functional polyesters and modulation of physical properties ...... 43 2.4.2 Conjugations of two dyes to functional polyester ............................................ 49 2.4.3 Conjugation of RGD and PEG to functional polyester and cell adhesion study ................................................................................................................................... 51 2.5 Conclusion .............................................................................................................. 54 III. MUSSEL INSPIRED ADHESIVES BASED ON ‘PEPTIDE-LIKE’ FUNCTIONAL POLYESTER .................................................................................................................... 55 3.1 Abstract ................................................................................................................... 55 3.2 Introduction ............................................................................................................. 56 3.3 Experimental Section .............................................................................................. 59 3.3.1 Materials and Instrumentation .......................................................................... 59 3.3.2 Synthesis and characterization of monomers ................................................... 60 3.3.3 Synthesis of polymers ....................................................................................... 64 3.3.4 Water contact angle measurement .................................................................... 68 3.3.5 Lap shear adhesion test of catechol containing polymers and control polymers ..................................................................................................................................