Bioresorbable Stereochemically Defined Polymers for Tissue Engineering and Wireless Bio-Integrated Electronic Device Applications

Yen-Hao Hsu

BIORESORBABLE STEREOCHEMICALLY DEFINED POLYMERS FOR TISSUE ENGINEERING AND WIRELESS BIO-INTEGRATED ELECTRONIC DEVICE APPLICATIONS

A Dissertation

Presented to

The Graduate Faculty of The University of Akron

In Partial Fulfillment

of the Requirements for the Degree

Doctor of Philosophy

Yen-Hao Hsu

March, 2021

BIORESORBABLE STEREOCHEMICALLY DEFINED POLYMERS FOR TISSUE ENGINEERING AND WIRELESS BIO-INTEGRATED ELECTRONIC DEVICE APPLICATIONS

Yen-Hao Hsu

Dissertation

Approved: Accepted:

______Advisor Interim Director of SPSPE Dr. Matthew L. Becker Dr. Ali Dhinojwala

______Committee Member Interim Dean of the College Dr. Yu Zhu Dr. Craig Menzemer

______Committee Member Interim Director, Graduate School Dr. Chrys Wesdemiotis Dr. Marnie Saunders

______Committee Member Date Dr. Xiong Gong

______Committee Member Dr. Kevin A. Cavicchi

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ABSTRACT

In most synthetic bioresorbable polymers, changing the physical properties such as elasticity and toughness by monomers results in a change to the crystallinity of the material, which manifests through alteration of its mechanical performance. “Thiol-yne” click chemistry has been discovered as an efficient methodology for step-growth polymerization between thiols and activated alkynes. Variation of the solvent polarity and base strength results in a series of elastomers possessing a wide variation of cis stereochemistry and crystallinity resulting in wide range of elasticity and stiffness. These materials are noteworthy that they do not rely on hard block-soft block interactions for their elastic properties nor are they crosslinked, which facilitates their degradation and use in regenerative medicine applications. Significantly, the crystalline domains that form between the cis alkene units and degradable segments to provide unexpected water barrier properties while retaining the ability to be resorbed. Thus, this innovation opens the new route for developing bioresorbable elastomers that can be widely applied into tissue engineering and wireless medical bio-electronics.

DEDICATION

I would like to dedicate this work to my wife, Yu-Chia Lai, who has sacrificed her past five years to allow me to pursue my education. You are my loyal listener, most trustworthy supporter, and my best friend.

ACKNOWLEDGEMENTS

The completion of this work could not have been completed without the aid and assistance of many individuals. First, I would like to thank my advisor, Dr. Matthew Becker, for his support, guidance, and enthusiasm during my graduate studies. I especially am thankful for the room and the respect that he granted me during my graduate work. I would also like to thank my committee members, Dr. Yu Zhu, Dr. Xiong Gong, Dr. Chrys

Wesdemiotis, and Dr. Kevin Cavicchi for spending their precious time giving me some suggestions and inputs.

This work of this dissertation would not have been possible without the help and assistance of my fellow group members. I would like to especially acknowledge: Dr. Jiayi

Yu, Shantanu Nikam, Yongjun Shin, Dr. Derek Luong, Dr. Jason Nettleton, Karissa

Nettleton, Dr. Darya Asheghali, Dr. Alex Kleinfehn, Dr. Zach Zander, Dr. Nathan Dreger, Dr.

Garrett Bass, and Peiru Chen. I will always cherish the memories and keep Becker lab championship attitude in my mind.

Last, but not least, I would like to thank my family and friends. Their selfless love and continuous support have kept me focused and positive all the time. Each of you has helped me obtain my goals.

TABLE OF CONTENTS

Page BIORESORBABLE STEREOCHEMICALLY DEFINED POLYMERS FOR TISSUE ENGINEERING AND WIRELESS BIO-INTEGRATED ELECTRONIC DEVICE APPLICATIONS ...... iii LIST OF TABLES ...... ix LIST OF FIGURES ...... x LIST OF SCHEMES ...... xxi CHAPTER1 I. INTRODUCTION ...... 1 1.1. Background for Synthetic Biodegradable Polymers ...... 1

1.2. Design and Synthesis of Biodegradable Ester-based Polymers ...... 2

1.3. Synthetic Biodegradable Polymers via Thiol-yne Step-growth Polymerization .. 7

1.4. Biodegradable Elastomers as Encapsulation in Wireless Bioresorbable Medical

Electronics ...... 10

II. MATERIALS AND INSTRUMENTATION ...... 12 2.1. Materials ...... 12

2.2. Instrumentation ...... 13

III. CROSSLINKED INTERNAL ALKYNE-BASED STEREOELASTOMERS: POLYMERS WITH TUNABLE MECHANICAL PROPERTIES ...... 17 3.1. Abstract ...... 17

3.2. Introduction...... 18

3.3. Experimental ...... 20

3.4. Results and Discussion ...... 30

3.5. Conclusion ...... 41

3.6. Acknowledgement ...... 42

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IV. SHAPE MEMORY BEHAVIOR OF BIOCOMPATIBLE POLYURETHANE ELASTOMERS SYNTHESIZED VIA THIOL-YNE MICHAEL ADDITION ...... 43 4.1. Abstract ...... 43

4.2. Introduction...... 44

4.3. Experimental ...... 47

4.4. Results and Discussion ...... 55

4.5. Conclusion ...... 65

4.6. Acknowledgement ...... 66

V. BIORESORBABLE ELASTOMERS WITH TUNABLE CRYSTALLINITY AS ENCAPSULATION LAYER IN WIRELESS BIO-ELECTRONICS TO ENHANCE WATER BARRIER ...... 67 5.1. Abstract ...... 67

5.2. Introduction...... 68

5.3. Experimental ...... 70

5.4. Results and Discussion ...... 78

5.5. Conclusion ...... 85

5.6. Acknowledgement ...... 86

VI. CONCLUSION ...... 87 REFERENCES ...... 91 APPENDIX A-SUPPORTING FIGURES ...... 112 APPENDIX B-SUPPORTING SCHEMES ...... 164

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LIST OF TABLES

Table Page

Table 3.1. Molecular masses, thermal and mechanical properties of internal alkyne-based (co)polymers, end-capped functionalized polymer, and crosslinked polymers...... 39

Table 4.1. Stereochemistry and molecular masses of thiol-yne polymers (U6T6) were obtained using different polymerization conditions from 5,14-dioxo-4,15-dioxa-6,13- diazaoctadecane-1,18-diyl dipropiolate (U6) and 1,6-hexanedithiol (T6) precursors and thermal properties were obtained by DSC...... 58

Table 4.2. Thermal and mechanical properties for five different %cis U6T6 polymers. ... 60

Table 5.1. Molecular masses, thermal and properties of (co)polymers with different molar fraction of bis(3-mercaptopropyl) succinate (CSS) incorporation...... 82

Table 5.2. Mechanical properties of (co)polymers with different molar fraction of bis(3- mercaptopropyl) succinate (CSS) incorporation...... 84

LIST OF FIGURES

Figure Page

Figure 1.1. A) Chemical structures of moieties susceptible to hydrolysis. B) Three different mechanisms of hydrolysis. C) Well-known biodegradable (co)polyesters...... 3

Figure 1.2. Polymerization methodologies for polyesters. A) Preparation of polyesters via condensation polymerization of either from I) diacids and diols or II) hydroxyl acids. B) Ring-opening polymerization (ROP) using either I) anionic living polymerization or II) metal-catalyzed initiated polymerization...... 5

Figure 1.3. Schematic illustration of thiol-yne step-growth polymerizations modified from previously reported literatures. A) Stereocontrolled step-growth polymerization via a nucleophilic thiol-yne addition demonstrates that low and high % cis can be achieved by altering the organic basicity and solvent polarity. B) A new series of biodegradable elastomer-like polymers reported that incorporates degradable succinate-based monomer units. The degradation rates can be widely tuned by incorporating different molar fraction of succinate-based monomer or changing % cis content while maintaining same amount of succinate-based monomer incorporation. C) A schematic depiction of how the stereochemistry contributes to the mechanical properties...... 9

Figure 3.1. A) A based-directed thiol-yne step-growth (co)polymerizations with x% of 2- butyne-1,4-diyl dipropiolate (I) under CHCl3 with DBU for high cis content. B) The stacked 1H NMR spectra of the (co)polymers with high cis- content via step-growth polymerization revealed the stoichiometry and %cis content is determined easily from the ratio of integration of resonance a to resonance g...... 33

1 three different wt% crosslinkers (1 wt%, 3 wt% and 5 wt%) in CHCl3. B) The H NMR spectra of end-capped polymer EI100 demonstrates phenyl ring resonance a can only be observed between  = 7.20-7.35 ppm. This confirms that benzyl mercaptan reacted with 1 I100 successfully. C) The H NMR spectra of crosslinked polymer X3EI100 demonstrates resonance d at  = 5.30 ppm which corresponds to the methylene resonance from 6 (see Figure 7.10). Resonance d also confirms the formation of triazoles between polymer chains by RuAAC click reaction...... 35

Figure 3.3. A) Differential scanning calorimetry (DSC) for internal alkyne-based polymers (I100, I70A30, I30A70, I10A90). The stacked thermograms display the glass transition temperatures (Tg) have a positive correlation with % 2-butyne-1,4-diyl dipropiolate (I) incorporation. B) DSC thermogram for end-capped polymer (EI100), clicked polymers (X1EI100 and X3EI100) revealed an increase of Tg with increasing %crosslinker...... 37

X1EI100 (red) and X3EI100 (blue) after click reaction at 37 C in the non-linear region displayed tunable mechanical properties with different wt% loading of crosslinker. All data were collected using three samples for the same measurement to illustrate the reproducibility...... 38

Figure 3.5. The hysteresis performances that can be utilized to determine shape recovery behavior by load-unloaded cyclic stress vs. strain curves at 37 C (end-capped functionalization polymer EI100: solid red line, 1 wt% loading crosslinked polymer X1EI100: dash blue line, 3 wt% loading crosslinked polymer X3EI100: dash green line). The crosslinked polymers by RuAAC reaction reduced significantly Young’s modulus (E0) due to disrupted chain packing resulting in the more elastic property that provided remarkable shape recovery behavior and tunable mechanical properties by feeding different wt% crosslinker...... 41

Figure 4.1. Characterization of stereocontrolled polyurethanes. A) A base-directed thiol- yne step-growth polymerization for different %cis content polymers was carried out with 5,14-dioxo-4,15-dioxa-6,13-diazaoctadecane-1,18-diyl dipropiolate (U6) and the distilled 1 1,6-hexanedithiol (T6) in different solvents and catalytic bases. B) Stacked H NMR spectra in CDCl3 demonstrated two clear vinyl proton doublets at δ = 7.1 ppm (cis, 9 Hz) and δ = 7.7 ppm (trans, 15 Hz), respectively. Significantly, the ratio of the cis to trans is xi determined by the resonance H1 to reveal the stereochemistry can be controlled under thiol-yne step-growth polymerization with specific solvent and catalytic base. C) SEC chromatograms determined by polystyrene (PS) standards in CHCl3 for polyurethanes with tunable stereochemistry...... 56

Figure 4.2. Thermomechanical characterization of polyurethane stereoelastomers. A) DSC nd thermograms of the 2 heating cycle presented for U6T6 with five different %cis. Higher cis content revealed higher glass transition temperature (Tg) to show more stiffer property. B) DSC thermograms of the 2nd heating cycle with three different ramp speeds presented for 82% cis content. Polymer chains formed better chain packing by hydrogen bonding interaction resulting in remarkable thermal-induced crystallinity (Tc) at 78 °C under slow ramp speed. C) Representative stress vs. strain curves for U6T6 with the various %cis content at 10 mm/ min under room temperature. D) Storage modulus, loss modulus, and tan  were presented by a DMA temperature ramp-frequency sweep for the 82% cis content stereoelastomer...... 59

Figure 4.3. Shape Memory characterization of 82% cis content. A) Representative curves of three shape memory cycles for 82% cis content stereoelastomer are shown. B) Visual demonstration of shape memory is observed in the 3rd cycle for the 82% cis content stereoelastomer by heating to 50 °C and the shape is shown to recover up to 90%...... 62 Figure 4.4. A) Mass loss over time with different %cis elastomers (black: 82%; red: 71%; blue: 62%; pink: 46%; olive: 32%, respectively) shows stereochemistry dependent surface erosion behavior. B) Enlarged scale at degradation region. C) SEM analysis of test coupons from 62% cis content exposed to accelerated degradation conditions indicates uniform degradation from surface erosion processes. Cell viability of 82% cis content. D) LIVE/DEAD imaging at 20× magnification of L929 cells on a 82% cis stereoelastomer disc and after exposure to 70% EtOH for 1 h. E) Cell viability as measured by XTT assay...... 64

Figure 5.1. Stereocontrolled synthesis and characterization of a series of bioresorbable elastomers. A) A based-directed thiol-yne step-growth (co)polymerizations with x% of bis(3-mercaptopropyl) succinate (CSS) under CHCl3 with DBU for 80-82% cis content. B) SEC chromatograms determined by polystyrene (PS) standards in HPLC grade CHCl3. C) 1 Stacked H NMR spectra in CDCl3 demonstrated two clear vinyl proton doublets at δ = 7.1 ppm (cis, 10 Hz) and δ = 7.6 ppm (trans, 15 Hz) respectively and the ratio of the cis to trans is determined to reveal the controlled stereochemistry...... 80

Figure 5.2. Thermal and mechanical properties of stereocontrolled polymers with different % of bis(3-mercaptopropyl) succinate (CSS) incorporation. Stacked thermogram xii of differential scanning calorimetry (DSC) at A) second heating cycle shows that the glass transition temperatures (Tg) and melting temperature (Tm) have a positive correlation with %CSS incorporation and B) second cooling cycle displays the fraction of crystallinity have the same trend. C) Overlapped data of thermogravimetric analysis (TGA) were performed to determine the degradation profile for each species. D) Increasing the amount of (Css) which is a longer, bulkier comonomer relatively reduced the UTS and the modulus of the resulting elastomers based on the uniaxial tensile tests. Mechanical property is highly tunable depending on the amount of Css content...... 83

Figure 5.3. A) Mass loss over time with different %CSS elastomers (black: 20%; red: 30%; blue: 40%; pink: 50%; olive: 60%, respectively) shows CSS dependent degradation rate profiles. B) A schematic illustration of polymer thin film demonstrates how the semicrystalline elastomers delayed the water penetration, then eventually degraded and fully resorbed...... 85

Figure 7.1. The 1H NMR spectrum of compound 2 shows only resonance a, which confirms full conversion and high purity (CD3OD, 300 MHz)...... 112

Figure 7.2. The 13C NMR spectrum of compound 2 shows quantitative resonances, which confirm full conversion and high purity (CD3OD, 75 MHz)...... 113

Figure 7.3. The 1H NMR spectrum of compound 4 shows appearance of resonances a, b and c, coming from the methyl (CH3) and phenol ring (aromatic) of tosyl groups (CDCl3, 300 MHz)...... 114

Figure 7.4. The 13C NMR spectrum of compound 4 shows appearance of resonances corresponding to the formation of disubstituted tosylation (CDCl3, 75 MHz)...... 115

Figure 7.5. The 1H NMR spectrum of 2-butyne-1,4-diyl dipropiolate (I) demonstrates two singlet resonances a and b corresponding to terminal alkyne and methylene group, respectively. Two singlet resonances also confirm I was synthesized from nucleophilic substitution (SN2) for both groups (CDCl3, 300 MHz)...... 116

Figure 7.6. The 13C NMR spectrum of 2-butyne-1,4-diyl dipropiolate (I) demonstrates resonance 3 corresponding to the carbonyl signal that confirms the formation of I via nucleophilic substitution (CDCl3, 75 MHz)...... 117

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Figure 7.7. The 1H NMR spectrum of compound 5 demonstrates resonance b come from methylene (-CH2-) group, which is adjacent to sulfur atom and resonance d confirms the formation of the thiol-yne reaction. The integration of resonance d illustrates 75% cis content compound 5 in CHCl3 with 1 mol% DBU (CDCl3, 300 MHz)...... 118

Figure 7.8. The 1H NMR spectrum of 1,6-diazidohexane displays a triplet splitting pattern a corresponding to methylene group (CDCl3, 300 MHz)...... 119

Figure 7.9. The 13C NMR spectrum of 1,6-diazidohexane displays only three resonances that confirm the disubstituted formation of the diazido compound (CDCl3, 75 MHz). . 120

Figure 7.10. The 1H NMR spectrum of compound 6 demonstrates resonances f, g, and h come from the aliphatic chain. An integration ratio of 1:1:1 also confirms the disubstituted formation of the triazole structure; resonance e is attributed to the methylene (-CH2-) protons adjacent to the ester group shifts downfield (~5.29 ppm) that affords the evidence of the triazole formation of the compound 6 (CDCl3, 300 MHz)...... 121

1 Figure 7.11. The H NMR spectrum of thiol-yne polymer I100 demonstrates the high cis- content for the thiol-yne step-growth polymer. % of Cis- can be calculated by the J coupling constants (Jcis = 9 Hz for and Jtrans = 15 Hz) of the respective resonances. Resonance d, e and f are from the dithiol monomer and resonance a is from the dialkyne monomer. The ratio of a, d, e, and f displays a 1:1:1:1 integration to afford pure thiol-yne polymer I100, revealing %cis/ %trans = 78 %: 22 % in CHCl3 with 1 mol% DBU (CDCl3, 300 MHz)...... 122

Figure 7.12. SEC chromatogram of I100 of thiol-yne step-growth polymer with 100% incorporation of I; Mn = 48.5 kDa, Mw = 79.9 kDa, Ð M = 1.7 (SEC DMF with 0.1 M LiBr, based on PS standards)...... 123

1 Figure 7.13. The H NMR spectrum of thiol-yne copolymer I70A30 demonstrates the high cis content for the thiol-yne step-growth copolymer. The ratio of resonance a to resonance g displays 70%: 30% that affords polymer I70A30 with 70% incorporation of I (CDCl3, 300 MHz). %cis = 80%...... 124

Figure 7.14. SEC chromatogram of I70A30 of thiol-yne step-growth polymer with 70% incorporation of I; Mn = 49.4 kDa, Mw = 87.6 kDa, Ð M = 1.8 (SEC DMF with 0.1 M LiBr, based on PS standards)...... 125 xiv

1 Figure 7.15. The H NMR spectrum of thiol-yne copolymer I30A70 demonstrates the high cis content for the thiol-yne step-growth copolymer. The ratio of resonance a to resonance g displays 30%: 70% that affords polymer I30A70 with 30% incorporation of I (CDCl3, 300 MHz). %cis = 80%...... 126

Figure 7.16. SEC chromatogram of I30A70 of thiol-yne step-growth polymer with 30% incorporation of I; Mn = 53.8 kDa, Mw = 108.2 kDa, Ð M = 2.0 (SEC DMF with 0.1 M LiBr, based on PS standards)...... 127

1 Figure 7.17. The H NMR spectrum of thiol-yne copolymer I10A90 demonstrates the high cis content for the thiol-yne step-growth copolymer. The ratio of resonance a to resonance g displays 10%: 90% that affords polymer I10A90 with 10% incorporation of I (CDCl3, 300 MHz). %cis = 80%...... 128

Figure 7.18. SEC chromatogram of I10A90 of thiol-yne step-growth polymer with 10% incorporation of I; Mn = 31.4 kDa, Mw = 105.3 kDa, Ð M = 3.4 (SEC DMF with 0.1 M LiBr, based on PS standards)...... 129

1 Figure 7.19. The H NMR spectrum of end-capped functionalization polymer EI100 demonstrates phenyl ring resonance a can only be observed at the region between 7.20- 7.35 ppm. This confirms that benzyl mercaptan was reacted with I100 successfully (CDCl3, 300 MHz)...... 130

1 Figure 7.20. The H NMR spectrum of crosslinked polymer X3wt_EI100 demonstrates resonance d at ~ 5.30 ppm which corresponds to the methylene signal from molecule 6 (see Figure S10). Resonance d also confirms the formation of triazoles between polymer chains by RuAAC click reaction (CDCl3, 300 MHz)...... 131

Figure 7.21. The overlapped thermogravimetric analysis (TGA) data were performed to determine the degradation profile for each species...... 132

Figure 7.22. Exemplar of stress vs strain curves for I100 (100% internal alkyne) at A) ambient temperature (25 °C ); B) 37 °C were tested at 10 mm/ min in the non-linear region. Data for three samples are shown to illustrate the reproducibility...... 132

Figure 7.23. Exemplar of stress vs strain curves for different % internal alkyne-based copolymers A) copolymer I70A30 (70% internal alkyne); B) copolymer I30A70 (30% internal alkyne); C) copolymer I10A90 (10% internal alkyne) were tested at 10 mm/ min at ambient temperature in the non-linear region. Data for three samples are shown to illustrate the reproducibility...... 133

Figure 7.24. Exemplar of stress vs. strain curves for A) end-capped functionalization polymer EI100 (before crosslinked); B) crosslinked polymer X1EI100 by loading 1 wt% crosslinker; C) crosslinked polymer X3EI100 by loading 3 wt% crosslinker were tested at 10 mm/ min at 37 °C in the non-linear region. Data for three samples are shown to illustrate the reproducibility...... 133

Figure 7.25. The hysteresis was performed by load-unloaded cyclic stress vs. strain curves stretching up to 300% with 4 cycles at 10 mm/min strain rate at 37 oC with 4 cycles for A) end-capped functionalization polymer EI100; B) crosslinked polymer X1EI100; C) crosslinked polymer X3EI100...... 134

Figure 7.26. The 1H NMR spectrum of compound 2 shows broad resonance d which confirms NCO functional group converted to urethane functional group and appearance of resonances a, b and c, all coming from 3-bromo-1-propanol (CDCl3, 300 MHz)...... 134

Figure 7.27. The 13C NMR spectrum of compound 2 displayed resonance 4 at 156.47 ppm corresponding to carbonyl group (C=O) of urethane (CDCl3, 75 MHz)...... 135

Figure 7.28. The 1H NMR spectrum of 5,14-dioxo-4,15-dioxa-6,13-diazaoctadecane-1,18- diyl dipropiolate (U6) demonstrates one singlet resonance a corresponding to terminal alkyne and resonance b, methylene group (CH2), shifted from 3.47 ppm to 4.27 ppm compared with Figure S1 shows the methylene (CH2) is adjacent to ester functional group. These two resonances confirm U6 was synthesized from nucleophilic substitution (SN2) of sodium propiolate for both groups (CDCl3, 300 MHz)...... 136

Figure 7.29. The 13C NMR spectrum of 5,14-dioxo-4,15-dioxa-6,13-diazaoctadecane-1,18- diyl dipropiolate (U6) displays two carbonyl signals, resonance 7 from urethane matched with Figure S2 and resonance 3 corresponding to the carbonyl signal from sodium propiolate that confirms the formation of U6 via nucleophilic substitution (CDCl3, 75 MHz)...... 137

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Figure 7.30. The 1H NMR spectrum shows the 82% cis content for the thiol-yne step- growth polymer. Cis content can be calculated by the J coupling constants (Jcis = 9 Hz and Jtrans = 15 Hz) of the respective resonances. Resonance j, k and l are from the dithiol monomer and resonance a, b, c, d, e, f and g are from urethane-based monomer (U6). The polymer reveals %cis/ %trans = 82 %: 18 % in CHCl3 with 1 mol% DBU (CDCl3, 300 MHz)...... 138

Figure 7.31. SEC chromatogram of 82% cis content of thiol-yne step-growth polymer; Mn = 52.9 kDa, Mw = 94.4 kDa, Ð M = 1.8 (SEC DMF with 0.1 M LiBr, based on PS standards)...... 139

Figure 7.32. The 1H NMR spectrum of 71% cis content for the thiol-yne step-growth polymer. Cis content can be calculated by the J coupling constants (Jcis = 9 Hz and Jtrans = 15 Hz) of the respective resonances. Resonance j, k and l are from the dithiol monomer and resonance a, b, c, d, e, f and g are from urethane-based monomer (U6). The polymer reveals %cis/ %trans = 71 %: 29 % in DMSO/ CHCl3 (1/ 3) with 1 mol% Et3N (CDCl3, 300 MHz)...... 140

Figure 7.33. SEC chromatogram of 71% cis content of thiol-yne step-growth polymer; Mn = 51.3 kDa, Mw = 98.2 kDa, Ð M = 1.9 (SEC DMF with 0.1 M LiBr, based on PS standards)...... 141

Figure 7.34. The 1H NMR spectrum of 62% cis content for the thiol-yne step-growth polymer. Cis content can be calculated by the J coupling constants (Jcis = 9 Hz and Jtrans = 15 Hz) of the respective resonances. of the respective resonances. Resonance j, k and l are from the dithiol monomer and resonance a, b, c, d, e, f and g are from the urethane- based monomer (U6). The polymer reveals %cis/ %trans = 62 %: 38 % in DMSO/ CHCl3 (1/ 4) with 1 mol% Et3N (CDCl3, 300 MHz)...... 142

Figure 7.35. SEC chromatogram of 62% cis content of thiol-yne step-growth polymer; Mn = 56.0 kDa, Mw = 93.5 kDa, Ð M = 1.7 (SEC DMF with 0.1 M LiBr, based on PS standards)...... 143

Figure 7.36. The 1H NMR spectrum of 46% cis content for the thiol-yne step-growth polymer. Cis content can be calculated by the J coupling constants (Jcis = 9 Hz and Jtrans = 15 Hz) of the respective resonances. of the respective resonances. Resonance j, k and l are from the dithiol monomer and resonance a, b, c, d, e, f and g are from the urethane-

xvii based monomer (U6). The polymer reveals %cis/ %trans = 46 %: 54 % in DMSO/ CHCl3 (1/ 5) with 1 mol% Et3N (CDCl3, 300 MHz)...... 144

Figure 7.37. SEC chromatogram of 46% cis content of thiol-yne step-growth polymer; Mn = 45.8 kDa, Mw = 100.6 kDa, Ð M = 2.2 (SEC DMF with 0.1 M LiBr, based on PS standards)...... 145

Figure 7.38. The 1H NMR spectrum of 32% cis content for the thiol-yne step-growth polymer. Cis content can be calculated by the J coupling constants (Jcis = 9 Hz and Jtrans = 15 Hz) of the respective resonances. of the respective resonances. Resonance j, k and l are from the dithiol monomer and resonance a, b, c, d, e, f and g are from the urethane- based monomer (U6). The polymer reveals %cis/ %trans = 32 %: 68 % in DMSO/ CHCl3 (1/ 6) with 1 mol% Et3N (CDCl3, 300 MHz)...... 146

Figure 7.39. SEC chromatogram of 32% cis content of thiol-yne step-growth polymer; Mn = 34.5 kDa, Mw = 74.0 kDa, Ð M = 2.1 (SEC DMF with 0.1 M LiBr, based on PS standards)...... 147

Figure 7.40. The overlapped TGA data of U6T6 polymers (82%, 71%, 62%, 46%, and 32% cis content respectively) were performed to determine the degradation profile for each species. The decomposition temperature (Td) was determined by the point at 5% weight loss...... 148

Figure 7.41. Overlapped FT-IR spectroscopy for 82% cis content U6T6 with before (black line) and after (red line) stretching shows the peaks at 3334 cm-1 and 1686 cm-1 are associated with hydrogen bonded N-H and C=O, respectively to support urethane linkages forming physical cross-linked network based hydrogen bonding.117 ...... 149

Figure 7.42. The hysteresis tests from 82% cis content were performed by load-unloaded cyclic stress vs. strain curves stretching up to different elongation (A: 20%; B: 50%, and C: 100% respectively) with 5 cycles at 10 mm/min strain rate...... 150

1 Figure 7.43. The H NMR spectrum of P1 (with 0% CSS) shows the 81% cis content for the thiol-yne step-growth polymer. Cis- content can be calculated by the J coupling constants of the respective resonances. Resonance f, g, h, i, and j are from the dithiol monomer (C10S) and resonance a, b, and c are from the 1,3-propane dipropiolate monomer (C3A).

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The polymer P1 reveals cis %/ trans % = 81 %: 19 % in CHCl3 with 1 mol% DBU (CDCl3, 500 MHz)...... 151

Figure 7.44. SEC chromatogram of P1 (81% Cis) of thiol-yne step-growth polymer; Mn = 32.5 kDa, Mw = 72.8 kDa, Ð M = 2.24 (SEC CHCl3, based on PS standards)...... 152

1 Figure 7.45. The H NMR spectrum of P2 (with 20% CSS) shows the 80% cis content for the thiol-yne step-growth copolymer with 20% incorporation of CSS in CHCl3 with 1 mol% DBU. The ratio of resonance n (from CSS) to resonance h (from C3A) displays 20%: 80% that affords polymer P2 with 20% incorporation of CSS (CDCl3, 500 MHz)...... 153

Figure 7.46. SEC chromatogram of P2 (80% Cis) of thiol-yne step-growth polymer with 20% incorporation of CSS; Mn = 32.6 kDa, Mw = 68.6 kDa, Ð M = 2.10 (SEC CHCl3, based on PS standards)...... 154

1 Figure 7.47. The H NMR spectrum of P3 (with 30% CSS) shows the 80% cis content for the thiol-yne step-growth copolymer with 30% incorporation of CSS in CHCl3 with 1 mol% DBU. The ratio of resonance n (from CSS) to resonance h (from C3A) displays 30%: 70% that affords polymer P3 with 30% incorporation of CSS (CDCl3, 500 MHz)...... 155

Figure 7.48. SEC chromatogram of P3 (80% Cis) of thiol-yne step-growth polymer with 30% incorporation of CSS; Mn = 29.1 kDa, Mw = 60.4 kDa, Ð M = 2.08 (SEC CHCl3, based on PS standards)...... 156

1 Figure 7.49. The H NMR spectrum of P4 (with 40% CSS) shows the 80% cis content for the thiol-yne step-growth copolymer with 40% incorporation of CSS in CHCl3 with 1 mol% DBU. The ratio of resonance n (from CSS) to resonance h (from C3A) displays 40%: 60% that affords polymer P4 with 40% incorporation of CSS (CDCl3, 500 MHz)...... 157

Figure 7.50. SEC chromatogram of P4 (80% Cis) of thiol-yne step-growth polymer with 40% incorporation of CSS; Mn = 26.7 kDa, Mw = 57.0 kDa, Ð M = 2.13 (SEC CHCl3, based on PS standards)...... 158

1 Figure 7.51. The H NMR spectrum of P5 (with 50% CSS) shows the 80% cis content for the thiol-yne step-growth copolymer with 50% incorporation of CSS in CHCl3 with 1 mol% DBU.

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The ratio of resonance n (from CSS) to resonance h (from C3A) displays 49%: 51% that affords polymer P5 with 49% incorporation of CSS (CDCl3, 500 MHz)...... 159

Figure 7.52. SEC chromatogram of P5 (80% Cis) of thiol-yne step-growth polymer with 50% incorporation of CSS; Mn = 45.4 kDa, Mw = 94.9 kDa, Ð M = 2.09 (SEC CHCl3, based on PS standards)...... 160

1 Figure 7.53. The H NMR spectrum of P6 (with 60% CSS) shows the 80% cis content for the thiol-yne step-growth copolymer with 60% incorporation of CSS in CHCl3 with 1 mol% DBU. The ratio of resonance n (from CSS) to resonance h (from C3A) displays 60%: 40% that affords polymer P6 with 60% incorporation of CSS (CDCl3, 500 MHz)...... 161

Figure 7.54. SEC chromatogram of P6 (80% Cis) of thiol-yne step-growth polymer with 60% incorporation of CSS; Mn = 29.0 kDa, Mw = 71.7 kDa, Ð M = 2.47 (SEC CHCl3, based on PS standards)...... 162

Figure 7.55. Exemplar stress vs. strain curves for (co)polymers (0%, 20%, 30%, 40%, 50%, 60% CSS) tested at 10 mm/min. Data for 3 samples are shown to illustrate the reproducibility...... 163

Figure 7.56. Illustration of in vitro medical device fabrication...... 163

LIST OF SCHEMES

Scheme Page

Scheme 7.1. The synthetic route demonstrates end-capped modification molecule (5) synthesized from thiol-yne reaction in CHCl3 with benzyl mercaptan and 1 mol% DBU. Internal clicked molecule (6) was synthesized by RuAAC in DCM, as a typical azide-alkyne cycloaddition...... 164

Scheme 7.2. The synthetic route for urethane-based dipropiolate monomer (U6)...... 164

Scheme 7.3. The general thiol-yne step-growth polymerization for U6T6 polymers ..... 165

Scheme 7.4. The synthesis of 1,3-propane diyl dipropiolate (C3A)...... 165

Scheme 7.5. The synthesis of Bis(3-mercaptopropyl) succinate (CSS)...... 165

xxi

CHAPTER I

INTRODUCTION

1.1. Background for Synthetic Biodegradable Polymers

Synthetic biodegradable and bioresorbable polymers are attractive because of remaining comprehensive studies for biological and biomedical applications in soft-tissue engineering (vascular,1-3 hernia,4, 5 skin6), orthopaedic repair7-11, and drug delivery

(degradable template,12, 13 encapsulation14, 15). Architecture of chemistry enables polymer scientists better handle of the chemical structures, molecular masses, and physical properties, making them a versatile branch of biomaterials with tunable degradation and mechanical properties.16-18 Certain features should be taken into consideration when evaluating the biodegradable polymer for tissue engineering: i) the polymer must allow for cell adhesion and growth, then induce differentiation;19, 20 ii) mechanical properties of the polymer must be similar to the native human tissue, degrade into non-toxic, resorbable monomers or oligomers under physiological conditions, and have the degradation rate that aligns with the rate of tissue regeneration.21, 22

Thus, synthetic polymeric biomaterials containing degradable moieties including ester,22 anhydride,23 ortho-ester,24 amide,25 urethane,26, 27 urea,28 and carbonate29 group have been extensively studied as for resorbable sutures and/ or supporters, drug delivery,

1 and bone screws due to molecular chain scission of polymers.21, 22 The degradation process of these polymers can be initiated either by passively acid- or base-catalyzed hydrolysis or enzyme-catalyzed process16, 30 (see Figure 1.1.A and 1.1.B). Notably, the polyester-based biomaterials are most commonly and relatively feasible to be designed, synthesized, and controlled to reach targeting physical properties and degradation rates such as poly(lactic acid) (PLA),17 poly(glycolic acid) (PGA),31 poly(lactic acid-co-glycolic acid)

(PLGA),32 poly(ε-caprolactone) (PCL),33 poly(butylene succinate) (PBS),34 and amino acid- based poly(ester urea)s (PEUs)9, 35 (see Figure 1.1.C).

1.2. Design and Synthesis of Biodegradable Ester-based Polymers

The majority of biodegradable polyesters can be achieved by two well-defined polymerization methodologies, which are acid-catalyzed polycondensation36, 37 and ring- opening polymerization (ROP).38, 39 The former can be successfully accomplished either by using two specific functional group at both end chains (dicarboxylic acids and diols) or hydroxyl acids with catalytic acid and both methods are needed to be carried out at high temperature to further remove water that allows reaction to follow Le Chatelier’s principle forward to obtaining high molecular mass polymer instead of oligomer 40(see

Figure 1.2.A). The latter can be accomplished by utilizing cyclic carboxylic ester monomers such lactones with specific catalyst and initiator to undergo ring-opening polymerization 2 to provide polymers with targeting molecular mass by controlling the feed ration between initiator to monomer 38(see Figure 1.2.B).

Figure 1.1. A) Chemical structures of moieties susceptible to hydrolysis. B) Three different mechanisms of hydrolysis. C) Well-known biodegradable (co)polyesters. 3

Certainly, both polymerizations are offsetting mutual lacks such as conversion, reactivity, controlled molar mass, polydispersity (PDI), chemical structure feasibility, and polymer architecture.38, 41 The conversion of monomers, the polycondensation is faster than ROP at early stage, yet the targeting molecular mass or repeating unit is difficult to be handled. Based on Carothers’ theory, the narrowest PDI for polycondensation is 2 at infinite degree of polymerization (DP)42; however, the PDI of ROP could be controlled down to 1.1 or less and present more consistency of DP.38 Some chemical structures are difficult to have cyclic form such aromatic moieties, then the polycondensation turns out be an ideal choice. Interestingly, the polymer architecture regarding the copolymers such as block, gradient, and alternating building blocks can be accomplished by ROP which means polycondensation usually provides “random” copolymers instead of block copolymers or gradient copolymers. Thus, the polymerization methodologies based on the proposed applications provide differences in morphology, thermal, mechanical properties, especially in degradation rates.43

Unfortunately, few shortages have been still remained to limit these materials made from abovementioned methods for soft-tissue regeneration.21, 22 The unsatisfied elastic properties and degradation rates related to natural human tissues, limiting the application of commercial biodegradable polymers. Although their mechanical properties can be achieved by modulating the molecular mass or using copolymers such as PLGA,

PCL, poly(glycerol sebacate) (PGS), and polyurethanes (PUs), they still have had difficulty in matching natural tissues perfectly.22, 44, 45 To PCL, it provides better mechanical property to be great candidate for surgery suture, yet does not replicate the elastic behavior of native tissues.22 While the tunable degradation rate of PLGA by controlling the stoichiometry of lactic acid and glycolic acid, it turns out to be shown semicrystalline and poor elasticity that allows only as a blending function to enhance mechanical properties.46 The chemically crosslinked polymer, PGS, can relatively produce elastic property; however, difficult to control the crosslinking density by polycondensation resulting in thermosetting that cannot be thermally processed after crosslinked.21, 45

Polyurethanes, widely applied into catheter medical devices, provide tunable elasticity by altering monomer selections from isocyanates, polyols, and short chain diols; unfortunately; struggling to control degradation rate between hard segment (isocyanates) and soft segment (polyols and diols) and generating urea linkage formation resulting from water contamination that are matched the time frame of the degradation.44, 47 6

1.3. Synthetic Biodegradable Polymers via Thiol-yne Step-growth Polymerization

Efforts to change the chemical structure to vary the mechanical and thermal

properties present the key view in biodegradable polymers that have made it hard to

decouple the effects of chemistry and mechanical properties on degradability and tissue

regeneration.21, 22 Particularly, no synthetic elastomeric or elastomer-like polymer system

has provided independent control of mechanical properties and degradation before

2016.22 After that, Dove and our group co-reported the first metal-free, stereocontrolled

step-growth polymerization via a nucleophilic thiol-yne addition which yielded a series of

thermally processable elastomer-like polymers in which the mechanical properties were

controlled by the double bond stereochemistry of the backbone (see Figure 1.3.A).48 The

double bond stereochemistry (% cis) in each polymer from thiol-yne step-growth

polymerization was well-controlled that based on solvent polarity and organic base which

are able to preferentially directs the thiol addition to the cis stereochemistry. Significantly,

Truong et al. have shown that low and high % cis can be achieved by changing the base

from Et3N (pKa = 10.75) to DBU (pKa = 13.5) while maintaining chloroform (CDCl3) as the

49 solvent. However, moderately high % cis subunits can be achieved with Et3N base when

a more polar solvent such as DMSO or DMF is used. All high % cis polymers were formed

using DBU/ CHCl3 but lower % cis contents were formed by using Et3N and varying

compositions of DMF and CHCl3 (17:3, 7:3, and 100% DMF). Nevertheless, in this initial 7 report, the materials were non-degradable and display no significant mass loss over years in 5 M KOH (aq) solution as the in vitro accelerated degradation condition, most likely a result of resistance to ester hydrolysis due to conjugation.

In order to translate these elastomer-like systems into regenerative medicine applications, a new series of biodegradable elastomer-like polymers have been developed and recently reported that incorporate degradable succinate-based monomer units (see

Figure 1.3.B).50 By changing the stoichiometry of succinate moiety incorporation, the degradation rate of the material can be tuned precisely while retaining control over the mechanical properties by maintaining the cis/trans stereochemistry of the double bond.

This structural control enables the independent tuning of mechanical and degradation properties and thus overcomes a major hurdle in biomaterials (see Figure 1.3.C).

1.4. Biodegradable Elastomers as Encapsulation in Wireless Bioresorbable Medical

Electronics

In past two decades, the ability to measure pressure inside the human body at the

targeting areas has become a main topic in modern clinical medicine. Up to now, there

are many procedures that still highly rely on the wire electronic implants in monitoring

and treating conditions that range from acute to traumatic injuries such as intracranial,

intra-abdominal, intravascular, and intraocular hypertension.51-53 While the traditional

and long-term implantable medical sensors provide the stability, accuracy for measuring

pressures, the procedure of surgical removal after recovery of patient could potentially

cause infections arising from biofilm formation from bacteria along with transdermal

wires that migrate within the wound area or the body.54-57

To solve these concerns, the sensors that can be fully resorbable by the

metabolism in the human body avoid the demand for further surgeries that have been

reported recently.58-62 A range of materials such as metals and their derivatives,63

biopolymers,64 and inorganic molecules60, 65 form the foundations of technologies that

suitable designs allow resorption and dissolution of the device eventually without

cytotoxicity and material residues. Certainly, eradicating the need for extraction surgeries

after a controlled period is desirable and there are few reported bioresorbable materials

including inorganic materials as encapsulation functions of the sensors. Unfortunately, 10 these materials are less stretchable and flexible that limit the development as encapsulation layer of the sensors. Thus, the bioresorbable elastomers with tunable crystallinity, elasticity, and degradation rates by utilizing thiol-yne step-growth polymerization have emerged to provide a great opportunity for solving issues in such these wireless medical devices.

CHAPTER II

MATERIALS AND INSTRUMENTATION

2.1. Materials

All commercial reagents and solvents were used as received without further purification except 1,6-hexanedithiol and 1,10-decandedithiol. The chloroform-d (CDCl3) and methanol-d4 (CD3OD) were purchased from Cambridge Isotopes Laboratories, Inc

(Tewksbury, MA). Chloroform (CHCl3) was purchased from VWR Chemicals (99% with amylene as inhibitor) and Fischer Scientific (HPLC grade, 99.5+% with amylene as

i inhibitor). Diethyl ether (Et2O) and isopropyl alcohol ( PrOH) were purchased from EMD

Millipore (Burlington, MA). Anhydrous methylene chloride (CH2Cl2), ethyl acetate (EtOAc),

N,N-dimethylformamide (DMF), dimethyl sulfoxide (DMSO), methanol (MeOH), sodium hydroxide (NaOH), potassium hydroxide (KOH), sodium sulfate (Na2SO4), sodium bicarbonate (NaHCO3), ammonium chloride (NH4Cl), propiolic acid, 3-bromo-1-propanol, hexamethylene diisocyanate (HDI), dibutyltin dilaurate, p-toluenesulfonyl chloride (TsCl),

2-butyne-1,4-diol, 1,3-propanediol, 1,6-dibromohexane, benzyl mercaptan, 1,8- diazabicyclo[5.4.0]undec-7-ene (DBU), sodium azide (NaN3), sulfuric acid (H2SO4), hexanes, 1,6-hexanedithiol, butylated hydroxytoluene (BHT), chloro(pentamethylcyclopentadienyl)(cyclooctadiene)ruthenium(II) (CpRuCl(COD)) were

12 purchased from Sigma-Aldrich (St. Louis, MO). 3-mercapto-1-propanol (>97%), 1,10- decanedithiol (>98%), were purchased from Tokyo Chemical Industry Ltd (Philadelphia,

PA).

2.2. Instrumentation

1H NMR (300 MHz) and 13C NMR (75 MHz) spectra were obtained using a Varian

Mercury NMR spectrometer operated at 303 K. All chemical shifts are reported in ppm (δ)

1 and referenced to the chemical shifts of residual solvent resonances (CDCl3 H: δ = 7.26

13 1 13 ppm, C: δ = 77.16 ppm; CD3OD H: δ = 4.87, 3.31 ppm, C: δ = 49.00 ppm).

Differential scanning calorimetry (DSC) was performed using a TA Instruments

Q200 DSC (TA Instruments – Waters L.L.C., New Castle, DE) on sample sizes between 5 –

10 mg using temperature ramps for heating of 10 °C∙min-1 and a cooling rate of 10 °C∙min-

1 . from -30 °C to 150 °C. The glass transition temperature (Tg) and melting temperature

(Tm) were determined from the second heating cycle of DSC. The crystalline temperature

(Tc) was determined in the second cooling cycle of DSC.

Thermogravimetric (TGA) analysis was performed using a TA Instruments TGA Q50

(TA Instruments – Waters L.L.C, New Castle, DE) on sample sizes of ca. 10 mg using a

-1 heating ramp of 20 °C∙min from r.t. to 700 °C. The decomposition temperature (Td) was determined at 5% mass loss.

Size exclusion chromatography (SEC) was performed on some samples using an

EcoSEC HLC-8320 GPC (Tosoh Bioscience LLC, King of Prussia, PA) equipped with a TSKgel

GMHHR-M mixed bed column and refractive index (RI) detector. Molecular masses were calculated using a calibration curve determined from polystyrene standards (PStQuick

MP-M standards, Tosoh Bioscience, LLC) with DMF with 0.1 M LiBr as eluent flowing at

1.0 mL∙min-1 at 323K, and a sample concentration of 3 mg∙mL-1. Some samples of SEC data were performed by using an EcoSEC HLC-8320GPC (Tosoh Bioscience LLC, King of Prussia,

PA) equipped with a TSKgel GMHHR-M mixed bed column and refractive index (RI) detector. Molecular masses were calculated using a calibration curve determined from polystyrene standards (PStQuick C and D standards, Tosoh Bioscience, LLC, King of Prussia,

PA) as eluent flowing at 0.5 mL∙min-1 at 313K, and a sample concentration of 3 mg∙mL-1

HPLC grade CHCl3.

The vacuum compression machine from TMP Technical Machine Products Corp. was used to fabricate thin films of each polymer. The machine was preheated to 150 °C.

Polymer was then added into the 50 × 50 × 0.5 mm mold and put into the compression machine with vacuum on. After 15 minutes of melting, the system was degassed three times. Next, 5 lbs*1000, 10 lbs*1000, 20 lbs*1000 of pressure were applied for 5 minutes, respectively. Following compression, the mold was cooled while maintaining 20 lbs*1000 of pressure to prevent wrinkle formation on the film’s surface. The press process 14 mentioned above was repeated twice to enable removal of air bubbles. Following the press, the mold was cooled before removed. The films were visually inspected to ensure that no bubbles were present. Dumbbell-shaped samples were cut using a custom ASTM

Die D-638 Type V.

Uniaxial tensile tests were performed on a custom ASTM Die D-638 Type V dumbbell-shaped samples using an Instron 4204 Universal Testing Machine at 99 °F (37

°C) and an Instron 5567 Universal Testing Machine at 75 °F (25 °C). The gauge length was set as 7 mm and the neck dimensions of the specimens were 7.11 mm in length, 1.70 mm in width and 0.50 mm in thickness.

Tensile tests were carried out using an Instron 4204 Universal Testing Machine at

99 °F (37 °C) and Instron 5567 Universal Testing Machine at 75 °F (25 °C). The gauge length was set as 7 mm and the crosshead speed was set as 10 mm/min. The dimensions of the neck of the specimens were 7.11 mm in length, 1.70 mm in width and 0.50 mm in thickness. The modulus was obtained from the slope of the initial linear region (1-5% strain). The reported results are average values from three individual measurements.

Load-unloaded cyclic tests were carried out using Instron instruments Instron 4204

Universal Testing Machine at 99 °F (37 °C). The gauge length was set as 7 mm and the crosshead load-unloaded speed was set as 10 mm/min constantly. The dimensions of the

15 neck of the specimens were 7.11 mm in length, 1.70 mm in width and 0.50 mm in thickness.

Dynamic Mechanical Analysis (DMA) was perform using a DMA Q800 instrument to obtain temperature sweep data. The rectangular DMA specimens (25 x 5 x 0.5 mm) were prepared by compression molding. Single frequency (1 Hz), strain-based (15 µm amplitude), temperature sweep (-50 to 120 °C at a rate of 3 °C∙min-1) experiments were conducted on three independent samples. The cyclic thermomechanical testing was completed in controlled force mode with heating and cooling rates of 10 °C∙min-1. The fixity and recovery parameters for shape memory were calculated using the standard shape memory equations shown in previous literature.66

CHAPTER III

CROSSLINKED INTERNAL ALKYNE-BASED STEREOELASTOMERS: POLYMERS WITH

TUNABLE MECHANICAL PROPERTIES

This work has been submitted to Macromolecules. Yen-Hao Hsu, Andrew P. Dove, Matthew L. Becker

3.1. Abstract

New methods to introduce and control polymer network crosslinking and improve mechanical properties of the resulting materials have been investigated extensively.

Methods to enhance mechanical properties of polymers and elastomers for industrial applications include “vulcanization” by which polymer chains are crosslinked through chemical bonds. In this work, we outline a new method to crosslink well-defined, synthetic elastomers using “click” reactions. Specifically, 2-butyne-1,4-diyl dipropiolate which possesses an internal alkyne, was synthesized as a functional monomer and further copolymerized to yield a series of elastomeric materials possessing variations in cis stereochemistry. Notably, the glass transition temperature and mechanical properties of the resulting copolymers can be tuned by changing the stereochemistry of the thiol-yne addition. The alkyne functionalities at the polymer chain ends and within the backbone allow for post-polymerization end-group functionalization and interchain crosslinks which 17 form polymer networks using a ruthenium-catalyzed azide-alkyne cycloaddition (RuAAC).

Hysteresis tests have shown that tensile modulus and recovery can be controlled by the density of the crosslinking within the network.

3.2. Introduction

Thiol-yne click chemistry is a powerful and efficient tool for the synthesis of polymeric materials67-70 and surface functionalization71-73 due to the fast reaction rates, improved yield over other condensation reactions and a high tolerance for a wide range of functional groups. Fairbanks et al. showed that terminal alkynes in thiol-yne click reactions typically react faster than terminal alkenes in thiol-ene type reactions.74 From a chemistry standpoint, thiol-yne click reactions can be utilized widely as they allow the consecutive addition of functional materials containing thiol groups to the terminal alkyne group available on the surface of a polymer.68, 74 Thiol-yne chemistry has also been applied to synthesize hyperbranched polymers and dendrimers via radical-mediated mechanisms under UV irradiation.75, 76 This method affords alternatives to large-scale synthesis and high-density branching networks.

Despite current advantages, thiol-yne click chemistry still poses some challenges that must be overcome when photoinitiated radical polymerization methods are used.

The major challenge is to obtain linear polymers using this technique, where multiple additions can lead to branching and covalent network formation. To address these issues, conditions for a single addition, nucleophilic thiol-yne Michael addition polymerization in

18 the presence of an organic catalyst were reported recently by our team48, 77, 78 and Tang et al.79 Significantly, the nucleophilic polyaddition is a typical Michael addition but is also a way to form linear polymers via anti-Markovnikov addition79 by utilizing alkyne functionality with electron-withdrawing groups (e.g., carbonyl group) to form conjugated systems. The interaction of the organobase with the activated alkyne is dependent on the solvent and when bound via hydrogen bonding is able to direct the stereochemistry of the addition product. This discovery has shown that the synthesis of cis- and trans- isomers can be finely controlled by the choice of solvents and organic bases producing contents ranging from high trans- to high cis- content structures.48, 49

Elastomeric materials with tunable mechanical properties have been studied by organocatalytic thiol-yne step-growth polymerization. Polymer networks can be formed by vulcanization that can dramatically enhance the ability of recovery after enforced deformation.48 Unfortunately, a loss in mechanical properties for these materials is typically observed with decreasing tensile stress after vulcanization which is attributed to the loss of alkenes in the backbone and reduced crystallinity. Significantly, the crosslinked polymer by vulcanization process has become the thermosetting property instead of thermoplastic that cannot allow us to undergo any thermal processes. Herein, we demonstrate a thiol-yne step-growth polymerization followed by ruthenium-catalyzed click reactions between internal alkynes that can be used as an alternative to vulcanization for control of crosslinking density while maintaining the alkene stereochemistry that allows for tunable mechanical properties. Additionally, propane-

1,3-dipropiolate (A) was chosen as a soft segment monomer copolymerized along with the internal alkyne-based monomer (I) to yield a series of elastomeric copolymers with tunable glass transition temperature (Tg) and mechanical properties. Further polymer networks can be controlled using a ruthenium-catalyzed azide-alkyne cycloaddition

(RuAAC) that makes polymers after crosslinked still maintain their thermoplastic property.

3.3. Experimental

Materials. All commercial reagents and solvents were used as received without further purification except 1,6-hexanedithiol. The chloroform-d (CDCl3) and methanol-d4

(CD3OD) were purchased from Cambridge Isotopes Laboratories, Inc (Tewksbury, MA).

Chloroform was purchased from VWR chemicals (ACS Grade, stabilized with amylene).

i Diethyl ether (Et2O) and isopropyl alcohol ( PrOH) were purchased from EMD Millipore

(Burlington, MA). Anhydrous methylene chloride (CH2Cl2), ethyl acetate (EtOAc), N,N- dimethylformamide (DMF), methanol (MeOH), sodium hydroxide (NaOH), potassium hydroxide (KOH), sodium sulfate (Na2SO4), sodium bicarbonate (NaHCO3), ammonium chloride (NH4Cl), propiolic acid, p-toluenesulfonyl chloride (TsCl), 2-butyne-1,4-diol, 1,8- diazabicyclo[5.4.0]undec-7-ene (DBU), sodium azide (NaN3), sulfuric acid (H2SO4), 1,3- propanediol, 1,6-dibromohexane, hexanes, butylated hydroxytoluene (BHT), benzyl mercaptan, chloro(pentamethylcyclopentadienyl)(cyclooctadiene)ruthenium(II)

(CpRuCl(COD)) were purchased from Sigma-Aldrich (St. Louis, MO).

Characterization. 1H NMR (300 MHz) and 13C NMR (75 MHz) spectra were obtained using a Varian Mercury NMR spectrometer operated at 303 K. All chemical shifts are reported in ppm (δ) and referenced to the chemical shifts of residual solvent resonances

1 13 1 13 (CDCl3 H: δ = 7.26 ppm, C: δ = 77.16 ppm; CD3OD H: δ = 4.87, 3.31 ppm, C: δ = 49.00 ppm).

Differential scanning calorimetry (DSC) was performed using a TA Instruments

Q200 DSC (TA Instruments – Waters L.L.C., New Castle, DE) on sample sizes between 5 –

10 mg using temperature ramps for heating of 10 °C∙min-1 and a cooling rate of 10 °C∙min-

1 . from -30 °C to 150 °C. The glass transition temperature (Tg) was determined from the midpoint in the second heating cycle of DSC.

Thermogravimetric (TGA) analysis was performed using a TA Instruments TGA Q50

(TA Instruments – Waters L.L.C, New Castle, DE) on sample sizes of ca. 10 mg using a

-1 heating ramp of 20 °C∙min from r.t. to 700 °C . The decomposition temperature (Td) was determined at 5% mass loss.

Size exclusion chromatography (SEC) was performed on all samples using an

EcoSEC HLC-8320 GPC (Tosoh Bioscience LLC, King of Prussia, PA) equipped with a TSKgel

GMHHR-M mixed bed column and refractive index (RI) detector. Molecular masses were calculated using a calibration curve determined from polystyrene standards (PStQuick

MP-M standards, Tosoh Bioscience, LLC) with DMF with 0.1 M LiBr as eluent flowing at

1.0 mL∙min-1 at 323K, and a sample concentration of 3 mg∙mL-1.

Mechanical Property Measurements. Tensile Tests at Different Strain Rates: Thin films of each polymer were fabricated using a vacuum compression machine (TMP

Technical Machine Products Corp.). The machine was preheated to 150 °C. Polymer was then added into the 50 × 50 × 0.5 mm mold and put into the compression machine with vacuum on. After 15 minutes of melting, the system was degassed three times. Next, 5 lbs*1000, 10 lbs*1000, 20 lbs*1000 of pressure were applied for 5 minutes, respectively.

Following compression, the mold was cooled while maintaining 20 lbs*1000 of pressure to prevent wrinkle formation on the film’s surface. The films were visually inspected to ensure that no bubbles were present. Dumbbell-shaped samples were cut using a custom

ASTM Die D-638 Type V. Rates tested were 0.1, 1, 5, 10, 20 mm/min. A rate of 10 mm/min was determined to be appropriate. Tensile tests at different stretching velocities were carried out using an Instron 4204 Universal Testing Machine at 99 °F (37 °C) and an Instron

5567 Universal Testing Machine at 75 °F (25 °C). The gauge length was set as 7 mm and the neck dimensions of the specimens were 7.11 mm in length, 1.70 mm in width and

0.50 mm in thickness.

Tensile Tests at 10 mm/min: Dumbbell-shaped samples were prepared using the method noted above. Tensile tests were carried out using an Instron 4204 Universal

Testing Machine at 99 °F (37 °C) and Instron 5567 Universal Testing Machine at 75 °F (25

°C). The gauge length was set as 7 mm and the crosshead speed was set as 10 mm/min.

The dimensions of the neck of the specimens were 7.11 mm in length, 1.70 mm in width and 0.50 mm in thickness. The modulus was obtained from the slope of the initial linear

22 region (1-5% strain). The reported results are average values from three individual measurements.

Hysteresis: Dumbbell-shaped samples were prepared using the same method as stated previously. Load-unloaded cyclic tests were carried out using Instron instruments

Instron 4204 Universal Testing Machine at 99 °F (37 °C). The gauge length was set as 7 mm and the crosshead load-unloaded speed was set as 10 mm/min constantly. The dimensions of the neck of the specimens were 7.11 mm in length, 1.70 mm in width and

0.50 mm in thickness.

Syntheses for Monomers and Polymers. Sodium propiolate (2): This compound was synthesized according to the procedure described by Bonnesen et al.80 In a dark hood, sodium hydroxide (0.645 g, 0.016 mol) was dissolved in 50 mL methanol in a 250 mL round-bottom flask. The solution was cooled to 0 °C for 10 min. Then propionic acid (1.000 mL, 0.016 mol) was added with stirring. The solution was returned to ambient temperature and stirred for an additional 2 h. The solvent was then removed by rotary evaporation. A white solid product was formed and dried under high vacuum to yield the white solid product (1.44 g, 97%). The product was stored in the dark due to light

1 13 sensitivities. H NMR (300 MHz, CD3OD) δ 2.95 (s, 1H). C NMR (75 MHz, CD3OD) δ 160.64,

81.83, 69.12.

But-2-yne-1,4-diyl bis(4-methylbenzenesulfonate) (4): But-2-yne-1,4-diyl bis(4- methylbenzenesulfonate) was synthesized according to the procedure described by

Maisonial et al.81 Briefly, p-toluenesulfonyl chloride (24.000 g, 0.126 mol) and

23 commercially available 2-butyne-1,4-diol (4.000 g, 0.046 mol) were dissolved in Et2O (300 mL). The mixture was cooled to -15 C for 15 min before potassium hydroxide (16.000 g,

0.285 mol) was added slowly. The resulting solution was stirred at 0 C for 3 h and then poured into a 1 L flask with ice water (300 mL), the solution was extracted with DCM (200 mL × 3) and the organic layer was collected, dried by anhydrous Na2SO4, filtered and evaporated to remove DCM. The solid was washed by Et2O (100 mL × 3) and dried under

1 vacuum 24 h to yield the white solid product (14.66 g, 80%). H NMR (300 MHz, CDCl3) δ

7.77 (d, J = 8.8 Hz, 4H), 7.34 (d, J = 8.8 Hz, 4H), 4.58 (s, 4H), 2.46 (s, 6H). 13C NMR (75 MHz,

CDCl3) δ 145.54, 132.80, 130.01 (× 2), 128.16 (× 2), 81.04, 57.21, 21.76.

2-Butyne-1,4-diyl dipropiolate (I): In the dark hood, compound 2 (7.600 g, 0.083 mol) and compound 4 (12.000 g, 0.030 mol) were dissolved in DMF (120 mL), then the mixture was heated to 50 C and stirred for additional 24 h. After the reaction was cooled down to room temperature, a saturated solution of NH4Cl (200 mL) was added to the mixture and stirred for 10 min. The mixture was extracted with DCM (150 mL × 3) and the organic extracts were combined. The extracted organic solution was extracted further with a saturated solution of NaHCO3 (150 mL× 3) to remove excess acid. The organic layer was combined and dried over anhydrous Na2SO4, filtered, and concentrated. The residue was purified by flash column chromatography on silica gel (EtOAc/hexanes 1:3; Rf = 0.30).

After removal of the solvent, the final product was further purified by distillation under

1 high vacuum at 110 °C to yield a colorless oil (3.76 g, 65%). H NMR (300 MHz, CDCl3) δ

13 4.81 (s, 4H), 2.97 (s, 2H). C NMR (75 MHz, CDCl3) δ 151.78, 80.60, 76.34, 73.82, 53.28.

+ + ESI-MS for C10H6O4Na, m/z theoretical: [M+Na] = 213.02 Da, observed: [M+Na] = 213.0

Da.

Procedure of thiol-yne step-growth polymerization for poly(bis(4-(propioloyloxy) but-2-yn-1-yl)-3,3'-(hexane-1,6-diylbis(sulfanediyl))) (I100): 2-butyne-1,4-diyl dipropiolate

(5.500 g, 0.029 mol) and 1,6-hexanedithiol (4.300 g, 0.028 mol) were added to a 500 mL round bottom flask with 200 mL CHCl3. The solution was then cooled to -15 C with stirring for 20 min before DBU (44 μL, 29 mmol) was added in one portion. Notably, the addition of DBU caused the solvent to reflux due to an exothermic reaction. After 10 min, the reaction was allowed to warm to room temperature. After 1 h, several drops of 2-butyne-

1,4-diyl dipropiolate (I) was added into reaction solution with CHCl3 (5 mL). After another

0.5 h, the solution was diluted with CHCl3 (50 mL) and butylated hydroxytoluene (BHT)

(0.480 g, 0.002 mol) was added. The polymer solution was then precipitated into diethyl ether (1.5 L) and collected by decanting the supernatant. The polymer was then dissolved in CHCl3 (150 mL) and reprecipitated into diethyl ether (1.5 L), collected by decanting the supernatant, and dried by high vacuum system at room temperature for 24 h to obtain

1 the pale-yellow I100 polymer (8.30 g, 85%). H NMR (CDCl3, 300 MHz) % cis: % trans = 78

%: 22 %, and cis content was determined by vinyl proton resonances. δ 7.74 (d, Jtrans = 15

Hz, 0.22H), 7.15 (d, Jcis, 9 Hz, 0.78H), 5.90-5.72 (m, 1H), 4.79 (s, 2H), 2.82-2.75 (m, 2H),

1.74-1.65 (m, 2H), 1.46-1.41 (m, 2H) (see Figure 7.11). SEC (DMF + 0.1M LiBr, based on PS standards): Mn = 48.5 kDa, Mw = 79.9 kDa, Ð M = 1.7. DSC: Tg = 24 C. TGA: Td = 279 C.

Tensile tests: E0 = 31.0 ± 6.7 MPa, break = 754 ± 50 %, UTS = 55.0 ± 7.0 MPa.

I70A30 random copolymer as example for general procedure of thiol-yne step- growth copolymerization: 2-butyne-1,4-diyl dipropiolate (2.66 g, 0.014 mol), propane-1,3- diyl dipropiolate (1.080 g, 0.006 mol) 48, and 1,6-hexanedithiol (3.050 g, 0.020 mol) were added to a 500 mL round bottom flask with 200 mL CHCl3. The solution was then cooled to -15 C with stirring for 20 min before DBU (44 μL, 29 mmol) was added in one portion.

Notably, the addition of DBU caused the solvent to bubble due to an exothermic reaction.

After 10 min, the reaction was allowed to warm to room temperature. After 1 h, several drops of 2-butyne-1,4-diyl dipropiolate was added into reaction solution with CHCl3 (5 mL). After another 0.5 h, the solution was diluted with CHCl3 (50 mL) and BHT (0.480 g,

0.002 mol) was added. The polymer solution was then precipitated into diethyl ether (1.5

L) and collected by decanting the supernatant. The polymer was then dissolved in CHCl3

(150 mL) and reprecipitated into diethyl ether (1.5 L), collected by decanting the supernatant, and dried by high vacuum system at room temperature for 24 h to obtain

1 the pale yellow I70A30 polymer (6.32 g, 92.3%). H NMR (CDCl3, 300 MHz) % cis ~ 79-80 with 30% propane-1,3-diyl dipropiolate incorporation. Cis content was determined by vinyl proton resonances, and % incorporation of propane-1,3-diyl dipropiolate was determined by ratio of integration of resonance a to resonance g δ 7.77-7.65 (m, 0.18H),

7.17-7.06 (m, 0.72H), 5.89-5.69 (m, 0.72H), 4.79 (s. 1.38H), 4.27-4.21 (m, 0.59H), 2.82-

2.75 (m, 2H), 2.07-1.99 (m, 0.30H), 1.73-1.64 (m, 2H), 1.47-1.41 (m, 2H) (see Figure 7.13).

SEC (DMF + 0.1M LiBr, based on PS standards): Mn = 49.4 kDa, Mw = 87.6 kDa, Ð M = 1.8.

DSC: Tg = 16 C. TGA: Td = 336 C. Tensile tests: E0 = 2.7 ± 0.1 MPa, break = 1560 ± 40 %,

UTS = 42.2 ± 4.5 MPa.

Copolymerization of I30A70: The polymer was prepared by the general procedure

1 described above. H NMR (CDCl3, 300 MHz) % cis ~ 79-80 with 70% propane-1,3-diyl dipropiolate incorporation. Cis content was determined by vinyl proton resonances, and

% incorporation of propane-1,3-diyl dipropiolate was determined by ratio of integration of resonance a to resonance g. δ 7.77-7.65 (m, 0.20H), 7.17-7.06 (m, 0.78H), 5.89-5.69 (m,

1H), 4.79 (s. 0.60H), 4.27-4.21 (m, 1.41H), 2.82-2.75 (m, 2H), 2.07-1.99 (m, 0.70H), 1.73-

1.64 (m, 2H), 1.47-1.41 (m, 2H) (see Figure 7.15). SEC (DMF + 0.1M LiBr, based on PS standards): Mn = 53.8 kDa, Mw = 108.2 kDa, Ð M = 2.0. DSC: Tg = 5 C. TGA: Td = 359 C.

Tensile tests: E0 = 9.3 ± 0.5 MPa, break = 1930 ± 60 %, UTS = 26.2 ± 2.9 MPa.

Copolymerization of I10A90: The polymer was prepared by the general procedure

1 described above. H NMR (CDCl3, 300 MHz) % cis ~ 79-80 with 90% propane-1,3-diyl dipropiolate incorporation. Cis content was determined by vinyl proton resonances, and

% incorporation of propane-1,3-diyl dipropiolate was determined by ratio of integration of resonance a to resonance g. δ 7.77-7.65 (m, 0.19H), 7.17-7.06 (m, 0.78H), 5.89-5.69 (m,

1H), 4.79 (s. 0.19H), 4.27-4.21 (m, 1.80H), 2.82-2.75 (m, 2H), 2.07-1.99 (m, 0.92H), 1.73-

1.64 (m, 2H), 1.47-1.41 (m, 2H) (see Figure 7.17). SEC (DMF + 0.1M LiBr, based on PS standards): Mn = 31.4 kDa, Mw = 105.3 kDa, Ð M = 3.4. DSC: Tg = 3 C. TGA: Td = 359 C.

Tensile tests: E0 = 48.3 ± 4.8 MPa, break = 1987 ± 51 %, UTS = 33.9 ± 0.8 MPa.

Synthesis of thiol-yne reaction for But-2-yne-1,4-diyl bis(3-(benzylthio) acrylate) (5):

Benzyl mercaptan (0.380 g, 3 mmol) was added into a 100 mL round bottom flask with 2- butyne-1,4-diyl dipropiolate (M1) (0.285 g, 1.5 mmol) in CHCl3 (20 mL) added slowly to the 100 mL round bottom flask. The solution was then cooled to -15 C with stirring for

20 min before DBU (2.4 μL, 0.015 mmol) was added in one portion. After stirring for 10 min, the reaction was allowed to warm to room temperature with continuous stirring.

After 2 h, the solvent was removed and the crude was directly purified by flash column chromatography on silica gel (pure DCM; Rf = 0.30) to afford a white solid product (0.567

1 g, 85%). H NMR (300 MHz, CDCl3) δ 7.77 (d, Jtrans = 15 Hz), 7.34-7.27 (m, 10H), 7.14 (d, Jcis

= 9 Hz), 5.86-5.79 (m, 2H), 4.76 (s, 4H), 4.03-3.97 (m, 4H). ESI-MS for C24H22O4S2Na, m/z theoretical: [M+Na]+ = 461.09 Da, observed: [M+Na]+ = 461.1 Da.

1,6-Diazidohexane: The diazides with less than six carbon atoms should be avoided due to explosion dangers. This compound was synthesized according to the procedure

82 described by Thomas et al. NaN3(1.90 g, 29.2 mmol) was added to a solution of the 1,6- dibromohexane (2.38 g, 9.8 mmol) with DMF (25.0 mL) in a 100 mL round bottom flask.

The mixture was stirred at 60 °C for 12 h. Then water (100 mL) was added and the product was extracted with ether (50 mL× 3). The organic layer was collected and dried over anhydrous Na2SO4, filtered, and concentrated. The concentrated product was concentrated under high vacuum overnight to yield a pure colorless oil product (1.50 g,

1 91%). H NMR (300 MHz, CDCl3) δ 3.27 (t, J = 6 Hz, 4H), 1.63-1.56 (m, 4H), 1.42-1.37 (m,

13 4H). C NMR (75 MHz, CDCl3) δ 51.40, 28.82, 26.39.

Model reaction molecule (Hexane-1,6-diylbis(1H-1,2,3-triazole-1,4,5-triyl)) tetrakis(methylene) tetrakis(3-(benzylthio) acrylate) (6): Cp*RuCl(COD) (9 mg, 0.024 mmol) and compound 5 (300 mg, 0.68 mmol) were added to a 100 mL two-neck round bottom flask. The round bottom flask was evacuated and purged with N2 three times before anhydrous DCM (40 mL) was added. 1,6-diazidohexane (56 mg, 0.33 mmol) was added into the reaction solution by a syringe and the reaction was stirred at room temperature for 12 h. After reaction solution was removed, then the crude product was directly purified by flash column chromatography on silica gel (DCM+3 wt% MeOH; Rf =

1 0.25) to afford a dark brown oil product (273 mg, 80%). H NMR (300 MHz, CDCl3) δ 7.73-

7.66 (m, 1H), 7.29-7.23 (m, 10H), 7.19-7.02 (m, 3H), 5.79-5.71 (m, 4H), 5.29 (s, 8H), 4.27

(br, 4H), 3.98-3.91 (m, 8H), 1.83 (br, 4H), 1.31 (br, 4H). ESI-MS for C54H56N6O8S4Na, m/z theoretical: [M+Na]+ = 1067.29 Da, observed: [M+Na]+ = 1067.3 Da.

Synthesis of end-capped functionalization polymer (EI100): The poly(bis(4-

(propioloyloxy) but-2-yn-1-yl)-3,3'-(hexane-1,6-diylbis(sulfanediyl))) (I100A0, 0.500 g, Mn =

23.6 kDa, Mw = 35.1 kDa, Ð M = 1.5.) was dissolved CHCl3 (50 mL) followed by addition of several drops of benzyl mercaptan into the polymer solution. The solution was cooled to

-15 C for 10 min before catalytic DBU (1 μL ) was added in a one portion to stir for 10 min. Then the polymer solution was allowed to warm to room temperature for 2 h. The end-capped functionalization polymer EI100 was precipitated into Et2O (400 mL), collected, and concentrated under high vacuum system for 24 h. SEC (DMF+ 0.1M LiBr, based on PS

29 standards) Mn = 24.7 kDa, Mw = 35.4 Da, Ð M = 1.5. DSC: Tg = 22 C. TGA: Td = 287 C. Tensile tests (at 37 C): E0 = 22.6 ± 4.3 MPa, break = 847 ± 30 %, UTS = 16.1 ± 1.1 MPa.

Crosslinked polymer X1EI100 as example for general procedure of crosslinking reaction. 1.000 g of end-capped functionalization polymer EI100 (Mn = 24.7 kDa, Mw = 35.4 kDa, Ð M = 1.5) and Cp*RuCl(COD) (0.001 g, 0.003 mmol) were added to a 250 mL two neck round bottom flask then the round bottom flask was evacuated and purged with N2 three times before dried CHCl3 (60 mL) was added. Then the 1 wt% of 1,6-diazidohexane (0.010 g, 0.059 mmol) as crosslinker was added into the reaction solution by syringe and allowed to stir for 12 h, respectively. The solution of crosslinked polymer X1EI100 was precipitated into Et2O (500 mL), collected, and concentrated using a high vacuum system for 24 h. DSC:

Tg = 30 C. TGA: Td = 251 C. Tensile tests (at 37 C): E0 = 5.9 ± 1.5 MPa, break = 526 ± 17

%, UTS = 25.0 ± 0.3 MPa.

Crosslinked polymer X3EI100: The crosslinked polymer was prepared by the general procedure described above using 3 wt% of 1,6-diazidohexane (0.030 g, 0.177 mmol) as crosslinker to yield X3EI100 polymer. DSC: Tg = 34 C. TGA: Td = 247 C. Tensile tests (at 37

C): E0 = 6.3 ± 0.2 MPa, break = 403 ± 17 %, UTS = 28.3 ± 3.0 MPa.

3.4. Results and Discussion

Monomer and Polymer Syntheses

In this work, an internal alkyne-based dipropiolate monomer, 2-butyne-1,4-diyl dipropiolate (I), was designed and synthesized (see Figure 3.1.A) to serve as a functional

30 monomer for a stereocontrolled thiol-yne step-growth polymerization with 1,6- hexanedithiol to yield the elastomeric polymer I100 (Mn =48.5 kDa, Mw = 79.9 kDa, Ð M =

1.7) with high cis content (~78 %, identified by J coupling of resonances of vinyl protons in 1H NMR spectrum shown in Figure 7.11) by using DBU as a catalyst in chloroform based on our conditions developed previously.48

In order to investigate the potential of these elastomeric systems, a copolymerization strategy that incorporates propane-1,3-dipropiolate (A) (30%, 70%, and

90% feed ratios) with the internal alkyne-based dipropiolate (I) and 1,6-hexanedithiol was employed to afford a series of high cis content copolymers (see Figure 3.1.A). The % incorporation of propane-1,3-dipropiolate (A) was calculated quantitatively by the integration ratio of singlet methine protons a  = 4.78 ppm (from monomer I) to triplet methine protons g  = 4.23 ppm (from monomer A) based on the 1H NMR spectra of the

(co)polymers in solution (CDCl3) (see Figure 3.1.B). The resulting stoichiometry-controlled copolymers each maintain high cis content (80-82 % cis).

Noticeably, the resulting polymer (I100) possesses the functional alkyne within each repeating unit of the backbone as well as at the chain ends. The end-group functionalization could ensure that further click reaction using internal alkyne can be performed via ruthenium-catalyzed azide-alkyne cycloaddition (RuAAC) reaction.83, 84

Especially, RuAAC reactions are classical click reactions that give complementary access to 1,5-disubstituted triazoles rather than 1,4-disubstituted triazoles from CuAAC.85, 86 Also,

31 modified ruthenium catalysts for RuAAC click reactions allows fully substituted triazoles via cycloaddition of internal alkynes with organic azides.84, 87

As a proof of concept, two consecutive click reactions (base-directed thiol-yne and

RuAAC) were initially utilized to synthesize a prototypical molecule that we designed as a model reaction before end-group modification and post-internal click reaction with I100 polymer. In the synthetic route, benzyl mercaptan (C6H5CH2SH) was chosen as an end- capping agent and treated with 2-butyne-1,4-diyl dipropiolate (I) in chloroform with 1 mol% DBU to undergo thiol-yne click reaction. Furthermore, the end-capped monomer was allowed to react with 1,6-diazidohexane acting as a crosslinker in dichloromethane

(DCM) with 2.5 mol% ruthenium catalyst at room temperature for 12 hours affording the model molecule (see Scheme 7.1).

Benzyl mercaptan end-capped polymer (EI100) was prepared from I100 polymer and benzyl mercaptan via thiol-yne click reaction, and 1H NMR analysis of the purified polymer also revealed aromatic proton resonances around  = 7.20-7.35 ppm. This confirms that benzyl mercaptan reacted with I100 successfully (see Figure 3.2.B and Figure 7.19). We also demonstrated that [Ru]-catalyzed click reactions with internal alkynes can be used as an alternative crosslinking method for vulcanization. The control over crosslink density while maintaining the stereochemistry of cis/ trans alkene bonds within the elastomeric polymers allows for the tuning of thermal and mechanical properties. Three different wt%

(1 wt%, 3 wt%, and 5 wt%, respectively) of 1,6-diazidohexane were used to crosslink I100 in the presence of catalytic amount of CpRuCl(COD) in CHCl3 at room temperature for 12 hours. The polymer network was then precipitated in diethyl ether (Et2O) to afford slightly crosslinked polymer networks (X1EI100 and X3EI100) (see Figure 3.2.C and Figure 7.20).

Unfortunately, 5 wt% crosslinker loading polymers (X5EI100) could not be isolated using this methodology due to high interchain crosslinking which caused the formation of insoluble polymer gel-like system precluding further characterization. However, the gel- like polymer revealed that the crosslinking process can undergo with the crosslinker less than 5 wt%, and the resulting polymers that remain below the gel point can be thermally processable, and remain soluble in common solvents for further processing and characterization.

Figure 3.2. A) The synthetic route demonstrates benzyl mercaptan end-capped functionalization polymer (EI100) synthesized from benzyl mercaptan and I100 polymer in CHCl3 with catalytic amounts of DBU. Subsequent crosslinked polymers (X1EI100, X3EI100, X5EI100) were synthesized by ruthenium-catalyzed click reaction with EI100 polymer and 1 three different wt% crosslinkers (1 wt%, 3 wt% and 5 wt%) in CHCl3. B) The H NMR spectra of end-capped polymer EI100 demonstrates phenyl ring resonance a can only be observed between  = 7.20-7.35 ppm. This confirms that benzyl mercaptan reacted with 1 I100 successfully. C) The H NMR spectra of crosslinked polymer X3EI100 demonstrates resonance d at  = 5.30 ppm which corresponds to the methylene resonance from 6 (see Figure 7.10). Resonance d also confirms the formation of triazoles between polymer chains by RuAAC click reaction.

Thermal and Mechanical Properties

The thermal properties of the materials were measured using differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA). Each of the polymers showed glass transition temperature (Tg), but no melting temperature (Tm) or crystalline temperature (Tc) were observed in heating/cooling cycles at 10 C/ min ramp speed. I100 polymer (with 100% incorporation of internal alkyne-based monomer I) possessed the highest glass transition temperature (Tg = 24 °C) while I10A90 (with 10 % incorporation of

I) had the lowest glass transition (Tg = 3 °C) (see Figure 3.3.A). Notably, incorporation of higher mole fractions of 2-butyne-1,4-diyl dipropiolate (I) resulted in higher glass transition temperature. Compared with propane-1,3-dipropiolate (A) described in previous work,9 the internal alkyne-based monomer (I) is relatively hard/less flexible segment in this copolymer system. Interestingly, the Tg of crosslinked polymers increased gradually from 22 C to 34 C revealing the decrease of chain mobility due to the increase of crosslinking density which resulted in more rigidity/ less flexibility (see Figure 3.3.B).

Decomposition temperature (Td) are also listed in Table 3.1 and show that higher percentages of triazole crosslinking by [Ru]-catalyzed click reactions decreased the decomposition temperature down to 251 C (X1EI100) and 247 C (X3EI100), respectively.

(see Figure 7.21).

The mechanical properties for (co)polymers with various % of 2-butyne-1,4-diyl dipropiolate (I) were further probed by tensile testing. The polymer I100 (100% incorporation of internal alkyne monomer) displayed stiff behavior due to glass transition

(Tg) around room temperature. In order to explore possibility of biomaterial applications, the polymer was also examined at 37 C. In comparison to room temperature, the elongation of polymer was extended to 1200 % and exhibited more elastic behavior (see

Figure 3.4.A). The Young’s modulus (Ε) of the copolymers ranged from 2.7 ± 0.1 MPa

(I70A30) to 48.3 ± 4.8 MPa (I10A90) likely due to strong interchain interactions. The results displayed less plastic behavior for each of the polymers with stiffer regime in the linear region of the stress vs strain curve, and suggested the possibility of developing elastomeric materials without a yield point. The I100 polymer with the highest amount of internal alkyne incorporation possessed the highest ultimate tensile strength (UTS) of 55 ± 7.0 37

MPa, and the lowest strain at break (beak) 754 ± 50% revealing an inverse relationship.

(see Figure 3.4.B)

Figure 3.4. Stress vs. strain curves. A) I100 polymer at ambient temperature and 37 C. B) Copolymers revealed the tunable mechanical properties with different % incorporation of 2-but-yne-1,4-diyl dipropiolate (I) at ambient temperature. C) Polymers for benzyl mercaptan end-capped functionalization polymer (EI100) prior to click reaction (black), X1EI100 (red) and X3EI100 (blue) after click reaction at 37 C in the non-linear region displayed tunable mechanical properties with different wt% loading of crosslinker. All data were collected using three samples for the same measurement to illustrate the reproducibility.

The mechanical performance of the end-capped polymer (EI100) was relatively elastic with an elongation at break (beak) of 847 ± 30 % and an ultimate tensile strength

(UTS) of 16.1 ± 1.1 MPa (see Figure 3.4.C). While X1EI100 and X3EI100 showed that UTS increases up to 25.0 ± 0.3 MPa and 28.3 ± 3.0 MPa for 1 wt% and 3 wt% crosslinker, respectively. Additionally, elongation at break (beak) decreased from 847 ± 30 % (see black in Figure 3.4.C) to 526 ± 17 % (see red in Figure 3.4.C) with 1 wt% crosslinker and 403 ±

17 % with 3 wt% (see blue in Figure 3.4.C). Strain hardening was noticeable in 1 wt% and

3 wt% crosslinked materials compared to the polymer in the absence of any crosslinker.

The increase in strain hardening also trends with wt% crosslinker as postulated. These data confirmed that modulus of elastomeric polymers can be controlled by altering wt% crosslinker with end-capped polymer EI100. (All data are listed in Table 3.1)

Table 3.1. Molecular masses, thermal and mechanical properties of internal alkyne-based (co)polymers, end-capped functionalized polymer, and crosslinked polymers.

% of Mw ÐM Tg Td E0 break UTS I[a] (kDa) (C) [b] (C)[c] (MPa) (%) (MPa) 100 79.9 1.7 24 279 31.0 ± 6.7 754 ± 50 55.0 ± 7.0 70 87.6 1.8 16 336 2.7 ± 0.1 1560 ± 40 42.2 ± 4.5 30 108.2 2.0 5 359 9.3 ± 0.5 1930 ± 60 26.2 ± 2.9 10 105.3 3.4 3 359 48.3 ± 4.8 1987 ± 51 33.9 ± 0.8 100 35.4 1.5 22 287 22.6 ± 4.3 847 ± 30[d] 16.1 ± 1.1[d] 100 - - 30 251 5.9 ± 1.5 526 ± 17[d] 25.0 ± 0.3[d] 100 - - 34 247 6.3 ± 0.2 403 ± 17[d] 28.3 ± 3.0[d] [a] % of 2-butyne-1,4-diyl dipropiolate (I) was determined by 1H NMR spectrometry. [b] Glass transition temperature (Tg) of all materials were determined from the second thermal cycle of differential scanning calorimetry. [c] Decomposition temperature (Td) was calculated at 5 % mass loss. [d] The mechanical properties were obtained based on stress vs strain curves at 37 C.

Elastic Hysteresis Properties

Shape recovery and energy loss of elastomeric materials can be determined by cyclic loading and unloading manipulation. Thus, benzyl mercaptan end-capped functionalized polymer, poly(bis(4-(propioloyloxy) but-2-yn-1-yl)-3,3'-(hexane-1,6- diylbis(sulfanediyl))) (EI100), and both crosslinked polymers (X1EI100 and X3EI100) were subjected to cyclic loading and unloading up to 300 % extension based on the stress vs

39 strain behavior shown in Figure 3.4.C. While the end-capped sample did not recover well

(280 %) due to weak interchain backbone interactions and permanent deformation during elongation. However, both crosslinked samples (X1EI100 and X3EI100) showed 164 % and

203 % recovery of their original length, respectively. The recovery enhancement after crosslinking revealed a modulus suppression (X1EI100 and X3EI100) attributed to − stacking and disruption of chain packing domains from formation of the triazole moieties between chains (see Figure 3.5). Additionally, crosslinked materials X1EI100 (with 1 wt% crosslinker) and X3EI100 (with 3 wt% crosslinker) yielded noteworthy changes to intrinsic mechanical properties resulting in strain hardening behavior. Conspicuously, the modulus in X3EI100 (3 wt%) was slightly higher than X1EI100 (1 wt%). Enhanced tensile stress and relatively low recovery in X3EI100 potentially resulted from more significant strain-induced crystallization during the extension process and increased difficulty recovering from crystallization to amorphous domains during unloading. Most notably, X1EI100 (1wt% crosslinker) in cyclic loading and unloading reveals enhanced elastic properties and providing compelling evidence for a covalently crosslinked network via RuAAC reaction.

Additionally, varying wt% crosslinker correlated to crosslinking density and mechanical properties in a tunable and controlled fashion.

3.5. Conclusion

We demonstrated the successful synthesis of internal alkyne-based elastomeric polymers via thiol-yne step-growth polymerization using a specific organic base and solvent to afford high cis content of alkenes. Further investigations of the thermal and mechanical properties of the copolymers revealed that these properties can be widely tunable with different % incorporation of 2-butyne-1,4-diyl dipropiolate. The end-capped polymer can be obtained by post polymerization thiol-yne modification. Consequently,

41 internal click reaction by RuAAC can be applied to connect polymer backbones resulting in crosslinked materials with tunable thermal and mechanical properties. The benefits of incorporating internal alkyne units are observable in future studies which could include post functionalization of dyes, peptides, and drugs for bioimaging, bioactive performance, and drug delivery. Especially, the crosslinkers can be potentially selected from degradable units such as ester moiety. Combination of thiol-yne and RuAAC click chemistry may be geared to another avenue en route to various biomedical applications.

3.6. Acknowledgement

The authors gratefully acknowledge financial support from the W. Gerald Austen

Endowed Chair in Polymer Science and Polymer Engineering. Mass spectrometry was performed by Selim Gerislioglu in the lab of Dr. Chrys Wesdemiotis at The University of

Akron Mass Spectrometry Center.

CHAPTER IV

SHAPE MEMORY BEHAVIOR OF BIOCOMPATIBLE POLYURETHANE ELASTOMERS

SYNTHESIZED VIA THIOL-YNE MICHAEL ADDITION

This work has not yet been submitted for publication. Yen-Hao Hsu, Derek Luong, Darya Asheghali, Andrew P. Dove, Matthew L. Becker

4.1. Abstract

Biodegradable shape memory elastomers have the potential for use in soft tissue

engineering, drug delivery and device fabrication applications. Unfortunately, few

materials are able to meet the targeted degradation and mechanical properties needed

for long-term implantable devices. In order to overcome these limitations, we have

designed and synthesized a series of unsaturated polyurethanes that are elastic,

degradable and non-toxic to cells in vitro. The polymerization included a nucleophilic

thiol-yne Michael addition between a urethane-based dipropiolate and a dithiol to yield

an α,β-unsaturated carbonyl moiety along the polymer backbone. The alkene

stereochemistry of materials was tuned between 32-82% cis content using the

combination of an organic base and solvent polarity which collectively direct the

nucleophilic addition. The bulk properties such as tensile strength, modulus, and glass

transition temperature can also be tuned broadly, and the hydrogen bonding imparted

by the urethane moiety allows for these materials to elicit cyclic shape memory behavior.

We also demonstrated the in vitro degradation properties are highly dependent on the

alkene stereochemistry.

4.2. Introduction

Polyurethanes (PUs)47, 88 are used widely in commercial products such as foams,89-

91 coatings,88, 92, 93 adhesives,88, 94 and medical devices95-99 due to their unique and tunable

thermoplastic and thermoset properties. Polyurethanes are also used in biomedical

applications including short-term implants such as surgical sutures,100, 101 catheters,98, 102-

104 and wound dressings.105-107 Polyurethanes have also been used in shape memory

applications.26, 95, 108 This behavior arises from interactions among the hydrogen bonding

networks that arise between the urethane groups.

Shape memory polymers (SMPs) are defined as materials that can be deformed to

a temporary shape that is ‘locked in’ until an external stimulus is applied (i.e.

electromagnetic field,109 light,110 and heat111) which allows the material to return to the

original shape. Several clever synthetic and architectural strategies have utilized shape

memory triggered by pH changes for drug delivery112 while others have used temperature

for stent deployment following placement in vivo.113 For materials seeking use thermally

induced shape memory for biomedical applications, the source of thermally stimulated

shape memory recovery should ideally come from the physiological temperature of the

44 human body (~ 37 °C). For this reason, the thermally-induced SMPs have been widely studied.26, 77

Thermal and mechanical properties of most well-known polyurethanes (PUs) can be modified by changing the hard segments (from diisocyanate derivatives), soft segments (from diols), and chain extenders (from oligomeric ester diols).114 The wide- ranging types of hydrolytically degradable segments including esters, anhydrides have been widely incorporated as well.21 The conventional polymerization method to obtain polyurethanes usually includes the use of a diisocyanate and a diol with a tin catalyst in an anhydrous system. Unfortunately, unstable carbamic acid could be formed from unremoved water with isocyanate, then rapidly decomposes into carbon dioxide and an amine, which can react with additional isocyanate to afford a urea linkage that could cause the undesired properties from uncontrolled and unrepeatable molecular weight distribution.47 Accordingly, the search for a simple and tolerant method to produce polyurethanes (PUs) with biodegradable, and/or resorbable units for long-term implant has been of interest for the materials science community.

Altering stereochemistry in nature has evolved into a way to control bulk mechanical properties (i.e., natural rubber and gutta-percha).115, 116 Property differences among these materials are due to the stereochemistry78 (cis vs. trans) of the alkene moiety within the polymer backbone and the elastic properties from natural rubber in particular arise from its high cis content which enhances the interchain packing. Stereo- chemically controlled addition within synthetic polymers has recently been accomplished

45 by nucleophilic thiol-yne Michael addition yielding , - unsaturated carbonyl moieties along with the polymer backbone.48-50, 77, 79 These materials have been found to have tunable thermal and mechanical properties which are highly desirable. Regarded as an efficient synthetic tool, the cis/trans ratio can be altered by choices of or organic base catalyst and solvent polarity without changing the stoichiometric composition of the final polymer materials. Additionally, the polymerization is easily scalable without an anhydrous reaction environment.48-50

Herein, we strategically designed and synthesized a urethane-based dipropiolate,

5,14-dioxo-4,15-dioxa-6,13-diazaoctadecane-1,18-diyl dipropiolate, as a novel monomer made from bromo-substituted carbamate and further nucleophilic substitution. The subsequent polymerization can be achieved by utilizing the efficient thiol-yne Michael reaction between the urethane-based dipropiolate and distilled 1,6-hexanedithiol to yield an unsaturated polyurethane. The control over stereochemistry of ,- unsaturated carbonyl moiety along the backbone provides tunable thermal and mechanical properties of which we showcased by exploiting different solvents (DMSO and/or CHCl3) and amine catalysts (Et3N or DBU) to achieve various configuration of cis double bond content. The urethane unit of the polymer chain displayed high potential shape memory behavior based on hydrogen bonding networks. Moreover, the hydrogen bonding interaction of thiol-yne polyurethanes was found to enhance mechanical properties with relatively low cis content. Significantly, polymers with various cis content were all found to be

amorphous and in vitro investigation including accelerated degradation and cell viability

indicated excellent biostability and biocompatibility.

4.3. Experimental

Materials. All reagents and solvents were used as received without further

purification except 1,6-hexanedithiol. The chloroform-d (CDCl3) and was purchased from

Cambridge Isotopes Laboratories, Inc (Tewksbury, MA). Chloroform was purchased from

VWR (Raleigh, NC; ACS Grade, stabilized with amylene). Diethyl ether (Et2O) and isopropyl

alcohol (iPrOH) were purchased from EMD Millipore (Burlington, MA). Anhydrous

methylene chloride (CH2Cl2), ethyl acetate (EtOAc), N,N-dimethylformamide (DMF),

dimethyl sulfoxide (DMSO), methanol (MeOH), sodium hydroxide (NaOH), sodium sulfate

(Na2SO4), sodium bicarbonate (NaHCO3), ammonium chloride (NH4Cl), propiolic acid, 3-

bromo-1-propanol, hexamethylene diisocyanate (HDI), dibutyltin dilaurate, 1,8-

diazabicyclo[5.4.0]undec-7-ene (DBU), triethylamine (Et3N), hexanes, butylated

hydroxytoluene (BHT), were purchased from Sigma-Aldrich (Milwaukee, WI).

Characterization. 1H (300 MHz) and 13C (75MHz) NMR spectra were obtained using

a Varian Mercury NMR spectrometer operated at 303 K. All chemical shifts are reported

1 in ppm (δ) and referenced to the chemical shifts of residual solvent resonances (CDCl3 H:

δ = 7.26 ppm, 13C: δ = 77.16 ppm).

Differential scanning calorimetry (DSC) was performed using a TA Instruments

Q200 DSC (TA Instruments – Waters L.L.C., New Castle, DE) on sample sizes between 5 –

10 mg using temperature heating ramps of 10 °C∙min-1 and a cooling rate of 10 °C∙min-1 from -30 °C to 150 °C. The glass transition temperature (Tg) was determined from the midpoint in the second heating cycle of DSC. The crystalline temperature (Tc) and the

-1 melting temperature (Tm) were obtained at 1 °C∙min in the second heating cycle.

Thermogravimetric analysis (TGA) was performed using a TA Instruments TGA Q50

(TA Instruments – Waters L.L.C, New Castle, DE) on sample sizes of ca. 10 mg using a

-1 heating ramp of 20 °C∙min from r.t. to 800 C. The decomposition temperature (Td) was determined at 5% weight loss.

Size exclusion chromatography (SEC) was performed on all samples using an

EcoSEC HLC-8320 GPC (Tosoh Bioscience LLC, King of Prussia, PA) equipped with a TSKgel

GMHHR-M mixed bed column and refractive index (RI) detector. Molecular masses were calculated using a calibration curve determined from polystyrene standards (PStQuick

MP-M standards, Tosoh Bioscience, LLC) with DMF with 0.1 M LiBr as eluent flowing at

1.0 mL∙min-1 at 323K, and a sample concentration of 3 mg∙mL-1 from DMF/ DMSO (v/ v

1:1).

Infrared (IR) spectra of 0.5 mm 82% cis polymer thin films (before and after stretched) were collected on a Nicolet i550 FT-IR (Thermo Scientific) (32 scans, 8 cm−1 resolution).

Mechanical Property Measurements. Tensile Tests at Different Strain Rates: Thin films of each polymer were fabricated using a vacuum compression machine (TMP

Technical Machine Products Corp.). The machine was preheated to 150 °C. Polymer was

48 then added into the 50 × 50 × 0.5 mm mold and put into the compression machine with vacuum on. After 15 minutes of melting, 5 lbs*1000, 10 lbs*1000, 20 lbs*1000 of pressure were applied for 5 minutes, respectively. Following compression, the mold was cooled while maintaining 20 lbs*1000 of pressure to prevent wrinkle formation on the film’s surface. The films were visually inspected to ensure that no bubbles were present.

Dumbbell-shaped samples were cut using a custom ASTM Die D-638 Type V. Strain rates of 0.1, 1, 5, 10, 20 mm/min were used and a rate of 10 mm/min was determined to be appropriate. Tensile tests at different stretching velocities were carried out using an

Instron 5567 Universal Testing Machine at 25 °C. The gauge length was set as 7 mm and the neck dimensions of the specimens were 7.11 mm in length, 1.70 mm in width and

0.50 mm in thickness.

Tensile Tests at 10 mm/min: Dumbbell-shaped samples were prepared using the same method as noted above. Tensile tests were carried out using Instron 5567 Universal

Testing Machine at 25 °C. The gauge length was set as 7 mm and the crosshead speed was set as 10 mm/min. The dimensions of the neck of the specimens were 7.11 mm in length,

1.70 mm in width and 0.50 mm in thickness. Modulus was obtained from the slope of the initial linear region. The reported results are average values from three individual measurements.

Dynamic Mechanical Analysis (DMA). Temperature Sweep Data: Rectangular DMA specimens (25 x 5 x 0.5 mm) were prepared by compression molding. Single frequency (1

Hz), strain-based (15 µm amplitude), temperature sweep (-50 to 120 °C at a rate of 3

°C∙min-1) experiments were conducted on three independent samples.

Shape Memory Characterization: Cyclic thermomechanical testing was conducted using a DMA Q800 instrument. Testing was completed in controlled force mode with heating and cooling rates of 10 °C∙min-1. The fixity and recovery parameters for shape memory were calculated using the standard shape memory equations shown in previous literature.66

In Vitro Accelerated Degradation. A film in 0.5 mm thickness of each elastomer was prepared from vacuum compression mold using the same method as stated above.

Discs with 4 mm in diameter were cut from the film and placed in 5 M NaOH solution in the incubator (37 ⁰C + 5% CO2 humidified atmosphere) for up to 12 weeks. The films absorbed, degraded, swelled and the 5 M NaOH solution was changed every week to ensure the degradation process. At specified intervals (1st, 2nd, 4th, 8th, and 12th week respectively), the samples were removed, dried and weighed. The results of mass changes are the average values of four individual samples for each material at each time point.

The surface changes during degradation were characterized using field emission scanning electron microscopy (SEM, JSM-7401F, JEOL, Peabody, MA).

Cell Viability Studies. Cell seeding onto polymer thin films: Mouse fibroblast cells

(L929, passage 8) were cultured using Eagle’s Minimum Essential Medium (ATCC) supplied with 10 vol. % horse serum, 100 units/mL penicillin, and 100 μg/mL streptomycin at 37 C

+ 5% CO2. Cells were subcultured every 3 days with 0.25% (w/v) trypsin and 0.5% (w/v)

EDTA solution. Films were sterilized by ethylene oxide (EtO) using an Anprolene AN74i sterilizer (Andersen Products) for 12 h followed by a purge cycle for 48 h. Samples were washed 3× with PBS prior to cell seeding. Cells were seeded on the polymer thin films at

30,000 cells/cm2. For positive and negative controls, cells were seeded on glass slides.

LIVE/DEAD imaging of cells seeded on polymer thin films: 48 h after cell seeding, polymer thin film and positive control glass slides were washed 3× with PBS and soaked in a LIVE/DEAD solution (Molecular Probes, Invitrogen) containing 5 μL of 4 mM calcein

AM and 5 μL of 2 mM ethidium homodimer-1 in 5 mL PBS for 30 minutes. Samples were then mounted with mounting media and imaged at 20× magnification using a Keyence

BZ-X700 microscope with filters for Texas Red and GFP. For the negative control, cells on glass slides were soaked in 70% EtOH for 1 h prior to staining, then processed identically to the experimental and positive control groups.

Cell viability as measured by XTT: The XTT (2,3-bis-(2-methoxy-4-nitro-5- sulfophenyl)-2H-tetrazolium-5-carboxanilide) assay (Biotium) was used to measure cell viability. For the experimental and positive controls, 48 h after seeding, growth media was replaced with 300 μL fresh media. For negative controls, growth media was replaced with 70% EtOH for 1 h, followed by replacing the 70% EtOH with 300 μL fresh media. Next,

75 μL of XTT solution was added and the sample was then incubated for 4 h at 37 C + 5%

CO2. Following incubation, the absorbance at 500 nm (A500) of the growth media solution was measured. Percent viability of the cells seeded on the polymer sample was calculated by the equation below,

퐴500(퐸푥푝) − 퐴500(푁푒푔) 푉푖푎푏푖푙푖푡푦 = × 100 퐴500(푃표푠) − 퐴500(푁푒푔)

where “Exp”, “PC”, and “NC” refer to the experimental, positive control, and negative control groups respectively.

Syntheses of Monomer and Polymers. Bis(3-bromopropyl) hexane-1,6- diyldicarbamate (2): Under anhydrous condition, hexamethylene diisocyanate (24.00 mL,

0.150 mol), 3-bromo-1-propanol (30.00 mL, 0.330 mol) were dissolved in anhydrous DCM

(200 mL) and the mixture was cooled to 0 °C for 10 min before a drop of dibutyltin dilaurate as catalyst was added in one aliquot. The reaction was kept at 0 °C for additional

10 min then warmed to room temperature and stirred overnight. The reaction solution was precipitated in diethyl ether (500 mL), filtered, washed again by additional diethyl ether (500 mL), and collected. The collected precipitate was dried by high vacuum system at room temperature for 24 h to obtain the product as a white powder (52.00 g, 78%). 1H

NMR (300 MHz, CDCl3) δ 4.69 (br, 2H), 4.19 (t, J = 6 Hz, 4H), 3.46 (t, J = 6 Hz, 4H), 3.16 (q,

J = 6 Hz, 4H), 2.16 (quint, J = 6 Hz, 4H), 1.54-1.45 (m, 4H), 1.36-1.30 (m, 4H). 13C NMR (75

MHz, CDCl3) δ 156.47, 62.53, 40.88, 32.29, 29.95, 29.74, 26.30. ESI-MS for

+ + C14H26Br2N2O4Na, m/z theoretical: [M+Na] = 469.01 Da, observed: [M+Na] = 469.0 Da.

5,14-Dioxo-4,15-dioxa-6,13-diazaoctadecane-1,18-diyl dipropiolate (U6): In the dark hood, bis(3-bromopropyl) hexane-1,6-diyldicarbamate (2) (10.00 g, 0.022 mol) and sodium propiolate80 (5.150 g, 0.056 mol) were dissolved in DMF (150 mL), then the mixture was heated up to 50 °C, and stirred for 24 h. After the reaction was cooled to room temperature, a saturated solution of NH4Cl (200 mL) was added to the mixture and 52 stirred for 10 min. The mixture was extracted with ethyl acetate (150 mL × 3) and the organic extracts were combined. The extracted organic solution was extracted further with a saturated solution of NaHCO3 (150 mL× 3) to remove excess acid. The organic layer was combined and dried over anhydrous Na2SO4, filtered, and concentrated. The residue was purified by flash column chromatography on silica gel (EtOAc/ hexanes 2:1; Rf = 0.35) twice. After removal of the solvent, the final product was dried under high vacuum to

1 yield the product as a white powder (6.66 g, 70%). H NMR (300 MHz, CDCl3) δ 4.74 (br,

2H), 4.27 (t, J = 6 Hz, 4H), 4.14 (t, J = 6 Hz, 4H), 3.15 (q, J = 6 Hz, 4H), 3.12 (s, 2H), 1,99

13 (quint, J = 6 Hz, 4H), 1.51-1.47 (m, 4H), 1.35-1.32 (m, 4H). C NMR (75 MHz, CDCl3) δ

156.46, 152.77, 75.05, 74.67, 63.15, 61.05, 40.89, 29.96, 28.17, 26.63. ESI-MS for

+ + C20H28N2O8Na, m/z theoretical: [M+Na] = 447.17 Da, observed: [M+Na] = 447.1 Da.

General procedure of thiol-yne step-growth polymerization for polymer U6T6:

Synthesis of 82% cis content was taken as example. 5,14-Dioxo-4,15-dioxa-6,13- diazaoctadecane-1,18-diyl dipropiolate (U6, 2.820 g, 6.600 mmol) and 1,6-hexanedithiol

(T6, 1.000 g, 6.600 mmol) were added to a 100 mL round bottom flask with 20 mL CHCl3.

The solution was then cooled to -15 °C with stirring for 15 min before 1,8- diazabicyclo[5.4.0]undec-7-ene (10.00 μL, 0.066 mmol) was added in one aliquot. Notably, the addition of DBU caused the solution to boil due to an exothermic reaction. After 10 min, the reaction was allowed to warm to room temperature. After 1 h, excess of 5,14- dioxo-4,15-dioxa-6,13-diazaoctadecane-1,18-diyl dipropiolate was dissolved in 5 mL

CHCl3 and added into reaction solution to prevent residual thiol as an end group. After

53 another 0.5 h, the solution was diluted with CHCl3 (20 mL) and butylated hydroxytoluene

(BHT) (0.110 g, 0.500 mmol) was added to limit interchain crosslinking. The polymer solution was then precipitated into diethyl ether (300 mL) and collected by decanting the supernatant. The polymer was then dissolved in CHCl3 (50 mL) and reprecipitated into diethyl ether (300 mL), collected by decanting the supernatant, and dried in vaccuo at room temperature for 24 h to obtain the pale-yellow polymer (3.43 g, 90 %). 1H NMR

(CDCl3, 300 MHz): % cis: % trans = 82 %: 18 %, and cis content was determined by vinyl proton resonances in Figure 7.30. SEC (DMF + 0.1M LiBr, based on PS standards): Mn =

52.9 kDa, Mw = 94.4 kDa, Ð M = 1.8. DSC: Tg = 12 °C, Tc = 78 °C, Tm = 108 °C. TGA: Td = 347

°C . Tensile tests: E0 = 3.6 ± 0.1 MPa, break = 1393 ± 11 %, UTS = 39.1 ± 0.7 MPa.

Synthesis of 71% cis content: The polymer was prepared by the general procedure described above by using DMSO/CHCl3 (v/v: 1/3) co-solvent system with Et3N as organic

1 base. H NMR (CDCl3, 300 MHz): % cis: % trans = 71 %: 29 %, and cis content was determined by vinyl proton resonances in Figure 7.32. SEC (DMF + 0.1M LiBr, based on PS standards): Mn = 51.3 kDa, Mw = 98.2 kDa, Ð M = 1.9. Tg = 9 °C. TGA: Td = 341 °C . Tensile tests: E0 = 2.1 ± 0.1 MPa, break = 1656 ± 27 %, UTS = 38.1 ± 2.5 MPa.

Synthesis of 62% cis content: The polymer was prepared by the general procedure described above by using DMSO/CHCl3 (v/v: 1/4) co-solvent system with Et3N as organic

1 base. H NMR (CDCl3, 300 MHz): % cis: % trans = 62 %: 38 %, and cis content was determined by vinyl proton resonances in Figure 7.34. SEC (DMF + 0.1M LiBr, based on PS

standards): Mn = 56.0 kDa, Mw = 93.5 kDa, Ð M = 1.7. Tg = 3 °C. TGA: Td = 346 °C . Tensile

tests: E0 = 1.7 ± 0.1 MPa, break = 1669 ± 5 %, UTS = 26.0 ± 0.9 MPa.

Synthesis of 46% cis content: The polymer was prepared by the general procedure

described above by using DMSO/CHCl3 (v/v: 1/5) co-solvent system with Et3N as organic

1 base. H NMR (CDCl3, 300 MHz): % cis: % trans = 46 %: 54 %, and cis content was

determined by vinyl proton resonances in Figure 7.36. SEC (DMF + 0.1M LiBr, based on PS

standards): Mn = 45.8 kDa, Mw = 100.6 kDa, Ð M = 2.2. Tg = 1 °C. TGA: Td = 315 °C . Tensile

tests: E0 = 5.0 ± 0.5 MPa, break = 1056 ± 22 %, UTS = 5.4 ± 0.5 MPa.

Synthesis of 32% cis content: The polymer was prepared by the general procedure

described above by using DMSO/CHCl3 (v/v: 1/6) co-solvent system with Et3N as organic

1 base. H NMR (CDCl3, 300 MHz): % cis: % trans = 32 %: 68 %, and cis content was

determined by vinyl proton resonances in Figure 7.38. SEC (DMF + 0.1M LiBr, based on PS

standards): Mn = 34.5 kDa, Mw = 74.0 kDa, Ð M = 2.1. Tg = -1 °C. TGA: Td = 320 °C . Tensile

tests: E0 = 4.0 ± 0.1 MPa, break = 785 ± 66 %, UTS = 2.6 ± 0.2 MPa.

4.4. Results and Discussion

Monomer and Polymer Syntheses

As depicted in Figure 4.1.A, the synthesis of the urethane-based dipropiolate

monomer, 5,14-dioxo-4,15-dioxa-6,13-diazaoctadecane-1,18-diyl dipropiolate (U6),

began from 3-bromo-1-propanol (1) reacted with hexamethylene diisocyanate (HDI) with

catalytic dibutyltin dilaurate to afford the dibromo-substituted urethane compound,

55 bis(3-bromopropyl) hexane-1,6-diyl dicarbamate (2). Further nucleophilic substitution

(SN2) of dibromo-substituted urethane (2) was successfully accomplished by reacting with sodium propiolate (C3HO2Na) to obtain the urethane-based dipropiolate monomer (U6).

Figure 4.1. Characterization of stereocontrolled polyurethanes. A) A base-directed thiol- yne step-growth polymerization for different %cis content polymers was carried out with 5,14-dioxo-4,15-dioxa-6,13-diazaoctadecane-1,18-diyl dipropiolate (U6) and the distilled 1 1,6-hexanedithiol (T6) in different solvents and catalytic bases. B) Stacked H NMR spectra in CDCl3 demonstrated two clear vinyl proton doublets at δ = 7.1 ppm (cis, 9 Hz) and δ = 7.7 ppm (trans, 15 Hz), respectively. Significantly, the ratio of the cis to trans is determined by the resonance H1 to reveal the stereochemistry can be controlled under thiol-yne step-growth polymerization with specific solvent and catalytic base. C) SEC chromatograms determined by polystyrene (PS) standards in CHCl3 for polyurethanes with tunable stereochemistry.

The combination of 5,14-dioxo-4,15-dioxa-6,13-diazaoctadecane-1,18-diyl dipropiolate (U6), the distilled 1,6-hexanedithiol (T6), and 1.0 mol% 1,8- diazabicyclo[5.4.0]undec-7-ene (DBU) in chloroform yielded U6T6 polymer with 82% cis content (Mn = 52.9 kDa, Mw = 94.4 kDa, Ð M = 1.8). (see Figure 4.1.A and Table 4.1).

Correspondingly, the %cis content can be identified by J coupling (Jcis = 9 Hz and Jtrans = 15

Hz), and be quantitatively calculated by the integration of cis and trans vinyl protons H1

1 using the H NMR spectra shown in Figure 4.1.B. Prototypical conditions (Et3N as basic catalyst and DMF as co-solvent in chloroform) were utilized to achieve relatively low %cis content polymers.48 Unfortunately, the polymer gradually precipitated out due to poor solubility. In order to achieve the targeted %cis content with high molar mass, we replaced DMF with DMSO as a co-solvent in CHCl3 while maintaining Et3N as the organic base catalyst. Four distinct volume ratios of DMSO (25%, 20%, 16.7%, and 14.3%) were used to provide various %cis content polymers with high molar mass. All polymerization conditions and molar masses are listed in Table 4.1.

Thermal and Mechanical Properties

The thermal properties of the materials were investigated by using differential scanning calorimetry (DSC) to observe the glass transition (Tg), crystalline (Tc), and melting

(Tm) temperatures and by using thermogravimetric analysis (TGA) to characterize the degradation temperature (Td). All five different %cis U6T6 stereoelastomers exhibited glass transitions (Tg) while no melting nor crystalline temperatures were observed during

nd st the 2 heating and 1 cooling cycle at 10 °C/min ramp speed. Moreover, the Tg trend shown in Figure 4.2.A displayed a positive correlation with %cis content. 82% cis content stereoelastomer possessed the highest glass transition temperature (Tg = 12 °C) while 32% cis content analog had the lowest glass transition (Tg = -1 °C). The glass transition of 82% cis was then screened using three different ramp speeds (10 °C/min, 5 °C/min, and 1

°C/min) which showed the Tg gradually decreased from 12 °C to 7 °C (see Figure 4.2.B).

Interestingly, crystalline (Tc = 78 °C) and melting (Tm = 108 °C) temperatures were only detected for the 82% cis stereoelastomer during the heating cycle. The thermal-induced crystallinity of 82% cis at slow ramp speeds (1 °C/ min) revealed the highest %cis content which enables better hydrogen bonding interactions between urethane units on polymer backbones. These results suggested that all materials were predominantly amorphous.

%Cis Solvent Base Mn (kDa) Mw (kDa) Ð M Tg (°C) Td (°C)

82 CHCl3 DBU 52.9 94.4 1.8 12 347

71 DMSO/CHCl3 (1:3) Et3N 51.3 98.2 1.9 9 341

62 DMSO/CHCl3 (1:4) Et3N 56.0 93.5 1.7 3 346

46 DMSO/CHCl3 (1:5) Et3N 45.8 100.6 2.2 1 315

32 DMSO/CHCl3 (1:6) Et3N 34.5 74.0 2.1 -1 320

Mechanical properties were investigated for U6T6 with various stereochemistry are listed in Table 4.2. The Young’s modulus, extension at break, and strain at break were all determined from the stress vs. strain curves obtained using tensile testing. The differences of Young’s modulus (E) of all polymers are not distinctive. This is attributed to each of the

59 species having a glass transition temperature (Tg) below room temperature while also being predominantly amorphous. The 82% cis content displayed significant strain-induced crystallization and strain-hardening after it was elongated over 600% of strain. Additionally,

Fourier-transform infrared (FT-IR) spectroscopy was used to study hydrogen bonding interactions. The stretches at 3334 cm-1 and 1686 cm-1 from the 82% cis stereoelastomer are associated with hydrogen bonding between amide N-H and C=O groups, respectively.117 Thus, the results from FT-IR support the hypothesis of hydrogen bonding between urethane linkages shown in Figure 7.41. The 32% cis content exhibits the lowest strain at break (break%) 785 ± 66 % (see Figure 4.3.C) due to reduced hydrogen bonding effect resulting from twisting configuration of polymer backbones based on increase of

%trans content. Notably, mechanical properties of 71% cis not only revealed exactly same

UTS (39.1 ± 0.7) as 82% cis but also performed elongation at strain break (break%) 1656 ±

27 % which is similar as 71% cis. Mechanical performance of 71% cis based on representative stress vs strain curves suggested that 71% cis stereochemistry would be enough to concomitantly obtain maximum UTS and elongation for the species under study once hydrogen bonding system incorporated.

Table 4.2. Thermal and mechanical properties for five different %cis U6T6 polymers.

%Cis E0 (MPa) beak (%) UTS (MPa) 82 3.6 ± 0.1 1393 ± 11 39.1 ± 0.7 71 2.1 ± 0.1 1656 ± 27 38.1 ± 2.5 62 1.7 ± 0.1 1669 ± 5 26.0 ± 0.9 46 5.0 ± 0.8 1056 ± 22 5.4 ± 0.5 32 4.0 ± 0.1 785 ± 66 2.6 ± 0.2 60

DMA Tests and Shape Memory Properties

The shape memory properties were tested by using a DMA Q800 instrument with cyclic thermomechanical testing over three cycles. Testing was completed in controlled force mode with heating and cooling rates of 10 °C/ min. Shape fixity (Rf) and recovery ratios (Rr) were calculated based on the equations shown below and notably the shape recovery percentage of each cycle is based on previous cycle.118 The strain after fixity was recorded as f, the strain before fixity was recorded as s, and the strain after shape recovery was recorded as r).

휀푓 − 휀0 푅푓 = × 100% 휀푠 − 휀0

휀푟 − 휀0 푅푟 = × 100% 휀푠 − 휀0

As a proof of concept for shape memory based on hydrogen bonding, the 82% cis content stereoelastomer was chosen to perform thermomechanical behavior. After investigated by temperature sweep, a storage modulus drop was observed at the glass transition temperature (Tg), followed by a rubbery-like plateau (see red line in Figure

4.2.D). This feature shows that the material has thermally stimulated shape memory properties. The large peak in loss modulus (see blue transition in Figure 4.2.D) revealed activation of molecular mobility and the rubbery-plateau indicated the presence of a network based on hydrogen bonding that is responsible for shape transformations.119-121

The material (82% cis) was examined with three shape memory cycles (see Figure 4.3.A).

The depicted results show the 1st cycle shape recovery was ~24%. Nevertheless, the 2nd and 3rd cycle displayed significant shape recovery improvement (~90%), and all of three cycles presented ideal fixity (~98.5%).

Unlike most shape memory materials based on covalently crosslinked bonding system,122, 123 the properties of the polyurethane stereoelastomers are based on hydrogen bonding within the urethane units which function as physical crosslinking units.117, 124, 125

When the material is heated, the hydrogen bonding orientation is disordered. However, the DSC thermogram of the 82% cis content showed a thermally induced crystalline transition (Tc) indicating that when the material was heated close to the onset of Tc, additional highly ordered hydrogen bonds would be formed. The result of thermomechanical analysis from the 1st cycle matched the observation from DSC as well. 62

Significantly, part of the polymer chains with weaker or disordered hydrogen bonding interaction existed in an amorphous domain which plays a key role in the shape recovery from the 2nd cycle that improve shape recovery performance from 24% to 90%. These features make urethane-based elastomer distinct from other SMP systems that depend on crosslinked chemical bonds, copolymers, polymers with blending, and classic physical crosslinking bond (H-bond) to form the network. In Figure 4.3.B, a visual demonstration with a three-dimensional plot of the 3rd cycle for 82% cis content is shown as it performs an ideal shape recovery process.

In Vitro Accelerated Degradation and Biocompatibility Studies

The hydrolytic stability was assessed for all three different %cis U6T6 polymers and molar masses were listed in Table 4.1. All testing samples for degradation studies were prepared by compression molding of a film with 0.5 mm thickness, cut into discs with 4 mm in diameter, and tested using accelerated degradation conditions in 5 M NaOH solution (n=3) at 37 °C + 5% CO2 for up to 12 weeks. The results show that degradation rates are highly dependent on the %cis conformation of , - unsaturated alkenes on the backbone (see Figure 4.4.A and 4.4.B). The lower %cis content provides more flexibility to increase the chain mobility matched the DSC thermograms (see Figure 4.2.A) and helps to accelerate the degradation rate through increased water penetration into the discs. SEM analysis of tested coupons from the accelerated degradation conditions on the 62% cis

63 polymer indicates uniform degradation and confirms surface erosion as the predominant degradation process (see Figure 4.4.C).

Figure 4.4. A) Mass loss over time with different %cis elastomers (black: 82%; red: 71%; blue: 62%; pink: 46%; olive: 32%, respectively) shows stereochemistry dependent surface erosion behavior. B) Enlarged scale at degradation region. C) SEM analysis of test coupons from 62% cis content exposed to accelerated degradation conditions indicates uniform degradation from surface erosion processes. Cell viability of 82% cis content. D) LIVE/DEAD imaging at 20× magnification of L929 cells on a 82% cis stereoelastomer disc and after exposure to 70% EtOH for 1 h. E) Cell viability as measured by XTT assay.

To assess further potential cytotoxicity from these materials, stereoelastomer samples with 82% cis content were seeded with L929 fibroblasts and cell viability was evaluated for 48 h after cell seeding. LIVE/DEAD imaging showed that cells viability was nearly quantitative as assessed by XTT assay and nearly identical to L929’s seeded on glass slides (Figures 4.4.D and 4.4.E). These data suggest that the urethane-based elastomers are compatible with cells and have potential use as an implantable biomaterial.

4.5. Conclusion

The urethane-based dipropiolate, 5,14-dioxo-4,15-dioxa-6,13-diazaoctadecane-

1,18-diyl dipropiolate, was successfully synthesized and utilized as a new class of dipropiolic monomer that can further be polymerized using stereo-chemically controlled thiol-yne Michael additions to yield polyurethanes. The thermal and mechanical properties are tunable by varying the stereochemistry of the alkene unit from 82% to 32% cis content in the polymer backbone. Amorphous polymers possessing hydrogen bonding along the backbone retain their mechanical properties. The thermomechanical analysis of 82% cis content polymer displayed shape memory behavior. The investigation of accelerated degradation behavior in 5 M NaOH solution up to a three-month period has displayed the materials with reduced %cis content yielded faster degradation rates thus demonstrating that stability of material is highly stereo-chemically dependent. In vitro cell viability of 82% cis content has indicated urethane-based elastomers possess excellent biocompatibility. The combination of biocompatible, biostable, and tunable thermal and

mechanical properties supports our material as potentially new candidates for a long-

term implantable biomaterial and biomedical device. Nonetheless, biodegradable

polymeric materials might be more valuable for soft tissue engineering due to degraded

segments would be expected to be non-toxic and resorbable in biological systems.

Importantly, a well-known degradable unit such as an ester moiety can be easily

incorporated into our polyurethane backbone and formulated as copolymer systems

could provide a series of resorbable polymers in the future.

4.6. Acknowledgement

The authors gratefully acknowledge financial support from the W. Gerald Austen

Endowed Chair in Polymer Science and Polymer Engineering. Mass spectrometry was

performed by Benqian Wei in the lab of Dr. Chrys Wesdemiotis at The University of Akron

Mass Spectrometry Center.

CHAPTER V

BIORESORBABLE ELASTOMERS WITH TUNABLE CRYSTALLINITY AS ENCAPSULATION

LAYER IN WIRELESS BIO-ELECTRONICS TO ENHANCE WATER BARRIER

This work has not yet been submitted for publication. Yen-Hao Hsu, Matthew L. Becker 5.1. Abstract

Bioresorbable materials have attracted tremendous attention in applications of

implantable medical devices. However, materials that possess the mechanical, resorbable

and chemical characteristics have been difficult to match the engineering desires such as

properties of elasticity and hydrophobicity to yield sufficient water barrier as

encapsulation layer and to have tunable degradation rates in flexible wireless medical

electronics. Herein, we report a series of bioresorbable elastomers with the

stereochemically controlled double bond and incorporation of longer aliphatic chain in

the backbone that allows to tune crystalline, physical, and degradation properties.

Significantly, we show semicrystalline elastomers could enhance water barrier

performances that protects the core structures of medical electronics extending desired

lifetime and manipulating degradation rates.

5.2. Introduction

In the modern clinical medicine, the implantable pressure sensors have been

widely used to monitor the conditions that range from acute damages to traumatic

injuries; especially in intraocular, intracranial, and intravascular pressure measurements

based on the highly precise and stable signals.51-53 However, the traditional technologies

of which require a second surgical procedure to remove electronic hardware after

patients fully recovered that could potentially cause the immune-inflammatory response

and the fatal infections.54-57, 126 To solve this issue, the new technology using

bioresorbable constituent materials as building elements that diminish the inflammatory

response and eliminate the need for an additional surgical extraction has been

developed.58-62 The performance with consistency through a clinically monitoring

timescale with devices that resorb completely over relatively longer period via biological

metabolism represents a substantial target. Bioresorbable strategy means the devices

immediately begin to hydrolyze and dissolve the components after implantation.127

Recently, materials making use of the encapsulation layers based on polymers

128 60 60, 65 (PLGA, polyanhydride ), and inorganic materials (silicon dioxide (SiO2) and silicon

65 nitride (Si3N4) ) that formed by either physical or chemical vapor deposition (PVD or CVD)

did not perform excellent water barrier property except SiO2 with thermal process by

61, 129, 130 calcination (t-SiO2) that grows ultrathin and defect-free film on the surface to

perform promising result. Yet, the perfect encapsulation layer by t-SiO2 provides the

unpredictable degradation rate (over years) that sustained longer than clinical need and

68 the less elasticity limits the applications in flexible bio-electronics.61, 129 Elastomeric materials provide the need in elasticity while there has been no systematic methodology to yield biodegradable and bioresorbable properties under a controlled period. We recently reported a series of bioresorbable elastomers with tunable thermal and mechanical properties and degradation rates that have been produced by organocatalytic thiol-yne step-growth polymerization with different amount of succinate-based moiety incorporation50; however, these materials as encapsulation layer are insufficient to protect active elements to achieve desired lifetime. Recently, Woodard and Grunlan proposed the mechanism to show the polymers with crystalline domain could efficiently delay the water penetration.16 To our best knowledges, designing the biodegradable polymers with tunable crystallinity has not been systematically established yet.

Herein, we strategically replace the 1,6-hexane dithiol with 1,10-decanedithiol while maintain 1,3-propane diyl dipropiolate and bis(3-mercaptopropyl) succinate as monomers to undergo thiol-yne step-growth (co)polymerization yielding a new series of elastomers with semicrystalline property resulting from better polymer chain packing.

Meaningly, the molar fraction control between 1,10-decanedithiol and bis(3- mercaptopropyl) succinate enables the unbiased tuning of mechanical, and degradation properties and the intrinsic semicrystalline property that could potentially protect the active elements to achieved or extend the desired device lifetime.

5.3. Experimental

Materials. All commercial reagents and solvents were used as received without

further purification except 1,10-decanedithiol. The chloroform-d (CDCl3) was purchased

from Cambridge Isotopes Laboratories, Inc (Tewksbury, MA). Diethyl ether (Et2O),

i isopropyl alcohol ( PrOH), and dichloromethane (CH2Cl2) were purchased from EMD

Millipore (Burlington, MA). Ethyl acetate (EtOAc), sodium sulfate (Na2SO4), sodium

bicarbonate (NaHCO3), ammonium chloride (NH4Cl), propiolic acid, sulfuric acid, 1,8-

diazabicyclo[5.4.0]undec-7-ene (DBU), hexanes, butylated hydroxytoluene (BHT), succinic

acid, 1,3-propanediol (>98%) were purchased from Sigma-Aldrich (Milwaukee, WI).

Chloroform (CHCl3) was purchased from VWR Chemicals (99% with amylene as inhibitor)

and Fischer Scientific (HPLC grade, 99.5+% with amylene as inhibitor), 3-mercapto-1-

propanol (>97%), 1,10-decanedithiol (>98%), were purchased from Tokyo Chemical

Industry Ltd (Philadelphia, PA).

Characterization. 1H NMR (500 Hz) and 13C NMR (125 Hz) spectra were obtained

using a Bruker NMR spectrometer operated at 303 K. All chemical shifts are reported in

1 ppm (δ) and referenced to the chemical shifts of residual solvent resonances (CDCl3 H: δ

= 7.26 ppm, 13C: δ = 77.16 ppm).

Differential scanning calorimetry (DSC) was performed using a TA Instruments

Q200 DSC (TA Instruments – Waters L.L.C., New Castle, DE) on sample sizes between 5 –

10 mg using temperature ramps for heating of 10 °C∙min-1 and a cooling rate of 10 °C∙min-

1 . from -30 °C to 130 °C. The glass transition temperature (Tg) was determined from the

70 midpoint in the second heating cycle and the crystalline temperature was determined from the second cooling cycle.

Thermogravimetric (TGA) analysis was performed using a TA Instruments TGA Q50

(TA Instruments – Waters L.L.C, New Castle, DE) on sample sizes of ca. 10 mg using a

-1 heating ramp of 20 °C∙min from r.t. to 800 °C . The decomposition temperature (Td) was determined at 5% mass loss.

Size exclusion chromatography (SEC) was performed on all samples using an

EcoSEC HLC-8320GPC (Tosoh Bioscience LLC, King of Prussia, PA) equipped with a TSKgel

GMHHR-M mixed bed column and refractive index (RI) detector. Molecular masses were calculated using a calibration curve determined from polystyrene standards (PStQuick C and D standards, Tosoh Bioscience, LLC, King of Prussia, PA) as eluent flowing at 0.5

-1 -1 mL∙min at 313K, and a sample concentration of 3 mg∙mL by HPLC grade CHCl3.

Mechanical Property Measurements. Tensile Tests at Different Strain Rates: Thin films of each polymer were fabricated using a heat press machine with manual force

(ROSINER product). The machine was preheated to 125 °C. Polymer was then added into the 50 × 50 × 0.5 mm mold and put into the heat press machine. After 15 minutes of melting, the force was applied by hand for 5 minutes then released. The press process mentioned above was repeated twice to enable removal of air bubbles. Following the press, the mold was cooled before removed. The films were visually inspected to ensure that no bubbles were present. Dumbbell-shaped samples were cut using a custom ASTM

Die D-638 Type V. Rates were tested by 1, 2, 3, 5, 10, 20 mm/min. A rate of 10 mm/min

71 was determined to be appropriate. Tensile tests at different stretching velocities were carried out using an Instron 5965 Universal Testing Machine at 75 °F (25 °C). The gauge length was set as 7 mm and the neck dimensions of the specimens were 7.11 mm in length,

1.70 mm in width and 0.50 mm in thickness.

Tensile Tests at 10 mm/min: Dumbbell-shaped samples were prepared using the same method as stated previously. Tensile tests were carried out using Instron 5965

Universal Testing Machine at 75 °F (25 °C). The gauge length was set as 7 mm and the crosshead speed was set as 10 mm/min. The dimensions of the neck of the specimens were 7.11 mm in length, 1.70 mm in width and 0.50 mm in thickness. Modulus was obtained from the slope of the initial 1-5% of linear region. The reported results are average values from three individual measurements.

In Vitro Accelerated Degradation. Films (50 m thickness) of each elastomer were prepared from the compression molding using the conditions stated above. Round discs

(4 mm in diameter) were punched from the respective films and placed in 5 M NaOH solution in the incubator (37 ⁰C + 5% CO2 humidified atmosphere) for up to 16 weeks. The films absorbed water, degraded, swelled under these conditions and the 5 M NaOH solution was changed weekly to maintain sink conditions. At specified intervals, the samples were removed, dried and weighed. The results of mass changes are the average values of four individual samples for each material at each time point.

Syntheses for Monomers and Polymers.

1,3-Propane diyl dipropiolate (C3A): The synthetic procedure was followed by previous work.48 1,3-propanediol (20.00 g, 0.263 mol), propiolic acid (50.00 g, 0.714 mol), and two drops of H2SO4 were added to a 500 mL one-neck round bottom flask with 200 mL toluene. The mixture was heated to reflux (110 °C) with Dean-Stark trap for overnight.

After that, the reaction solution was cooled to room temperature, and extracted with saturated solution of NaHCO3 (100 mL × 3) to remove the excess of propiolic acid. Then the organic phase was collected, dried over Na2SO4, filtered and reduced in volume to dryness collected. The collected crude product was purified by flash column

st chromatography on silica gel (EtOAc/ hexanes 1:3; Rf = 0.45) to collect 1 fraction. After removal of the solvent, the final product was further purified by distillation under high vacuum (0.02 Torr, 90 oC) to yield the colorless oil product (39.80 g, 84.0 %). 1H NMR (500

MHz, CDCl3) δ 4.29 (t, J = 10 Hz, 4H), 2.91 (s, 2 H), 2.07 (quint, J = 10 Hz, 2 H).

Bis(3-mercaptopropyl) succinate (CSS): The synthetic procedure was similar to methods described previously.50 Succinic acid (15.00 g, 0.127 mol), 3-mercapto-1- propanol (25.00 g, 0.271 mol), and two drops of H2SO4 were added into 250 mL one-neck round-bottom flask with 180 mL toluene, and the mixture was heated to reflux (110 oC) with Dean-Stark trap for overnight to remove the water. Then, the reaction solution was cooled to room temperature, and extracted with saturated solution of NaHCO3 (100 mL ×

3) to remove the residual acid. The organic phase was collected, dried over Na2SO4, filtered and reduced in volume to dryness collected. The collected crude product was purified by flash column chromatography on silica gel (EtOAc/ hexanes 2:3; Rf = 0.45) to

73 collect 1st fraction. After removal of the solvent, the final product was further purified by distillation under high vacuum (0.02 Torr, 120 °C) to yield the colorless oil product (26.70

1 g, 79.1 %). H NMR (500 MHz, CDCl3) δ 4.20 (t, J = 10 Hz, 4 H), 2.62-2.57 (m, 8 H), 1.93

(quint, J = 10 Hz, 4 H), 1.39 (t, J = 10 Hz, 2 H). 13C NMR

80-82% Cis content of polymer with 0% bis(3-mercaptopropyl) succinate (CSS) taken as example of general procedure of thiol-yne step growth polymerization: 1,3-Propane diyl dipropiolate (C3A, 1.56 g, 8.60 mmol) and bis(3-mercaptopropyl) succinate (C10S, 1.78 g, 8.60 mol) were added to a 100 mL round bottom flask with 16 mL CHCl3. The solution was then cooled to -15 °C with stirring for 15 min before DBU (13 μL, 0.086 mmol) was added in one portion. Notably, the addition of DBU caused the solvent to reflux due to the vigorous exothermic reaction and the reaction was allowed to warm to room temperature slowly. After 1 h, excess of C3A was dissolved in 5 mL CHCl3 and added into reaction solution. After another 0.5 h, the solution was diluted with CHCl3 (20 mL) and

BHT (0.14 g, 0.65 mmol) was added to prevent crosslinking. The polymer solution was then precipitated into diethyl ether (400 mL) and collected by decanting the supernatant and dried under high vacuum conditions at room temperature for 24 h to yield the pale-

1 yellow polymer (3.08 g, 92 %). SEC (CHCl3) Mn = 32.6 kDa, Mw = 68.6 kDa, Ð M = 2.24. H

NMR (CDCl3, 500 MHz) % cis: % trans = 81 %: 19 % (see Figure 7.43). DSC: Tg = -6.8 C, Tc

= 61.4 C, Tm = 112.4 C. TGA: Td = 359 C. Tensile tests: E0 = 65.8 ± 0.6 MPa, break = 1457

± 112 %, UTS = 32.2 ± 3.9 MPa.

20% bis(3-mercaptopropyl) succinate (CSS) incorporation as general procedure for thiol-yne step-growth random copolymerization: CSS (0.715 g, 2.680 mmol), C3A (2.420 g,

13.43 mmol) and C10S (2.210 g, 10.70 mmol) were added to a 100 mL round bottom flask with 25 mL CHCl3. The solution was then cooled to -15 °C with stirring for 15 min before

DBU (20 μL, 0.013 mmol) was added in one portion. Notably, the addition of DBU caused the solvent to bubble due to an exothermic reaction. After 10 min, the reaction was allowed to warm to room temperature. After 1 h, excess of C3A was dissolved in 5 mL

CHCl3 and added into reaction solution. After another 0.5 h, the solution was diluted with

CHCl3 (25 mL) and BHT (0.225 g, 10.22 mmol) was added. The polymer solution was then precipitated into diethyl ether (400 mL) and collected by decanting the supernatant and dried by high vacuum system at room temperature for 24 h to yield the pale-yellow

1 polymer with 20% CSS (4.81 g, 90 %). H NMR (CDCl3, 500 MHz) % cis: % trans = 80 %: 20

% (see Figure 7.45). SEC (CHCl3, based on PS standards) Mn = 32.5 kDa, Mw = 72.8 kDa, Ð M

= 2.10. DSC: Tg = -7.3 C, Tc = 30.3 C, Tm = 99.3 C. TGA: Td = 348C. Tensile tests: E0 = 57.2

± 4.4 MPa, break = 1700 ± 31 %, UTS = 40.5 ± 2.2 MPa.

Random copolymer with 30% CSS incorporation: CSS (1.073 g, 4.030 mmol), C3A

(2.420 g, 13.43 mmol) and C10S (1.941 g, 9.402 mmol) were added to a 100 mL round bottom flask with 25 mL CHCl3. The solution was then cooled to -15 °C with stirring for 15 min before DBU (20 μL, 0.013 mmol) was added in one portion. Notably, the addition of

DBU caused the solvent to bubble due to an exothermic reaction. After 10 min, the reaction was allowed to warm to room temperature. After 1 h, excess of C3A was dissolved

75 in 5 mL CHCl3 and added into reaction solution. After another 0.5 h, the solution was diluted with CHCl3 (25 mL) and BHT (0.225 g, 10.22 mmol) was added. The polymer solution was then precipitated into diethyl ether (400 mL) and collected by decanting the supernatant and dried by high vacuum system at room temperature for 24 h to yield the

1 pale-yellow polymer with 30% CSS (4.84 g, 89 %). H NMR (CDCl3, 500 MHz) % cis ~ 80 with

30% CSS incorporation (see Figure 7.47). SEC (CHCl3) Mn = 29.1 kDa, Mw = 60.4 kDa, Ð M =

2.08. DSC: Tg = -7.3 C, Tc = 18.5 C, Tm = 88.9 C. TGA: Td = 349 C. Tensile tests: E0 = 40.9

± 3.5 MPa, break = 1638 ± 33 %, UTS = 28.3 ± 1.2 MPa.

Random copolymer with 40% CSS incorporation: CSS (1.431 g, 5.373 mmol), C3A

(2.420 g, 13.43 mmol) and C10S (1.664 g, 8.060 mmol) were added to a 100 mL round bottom flask with 25 mL CHCl3. The solution was then cooled to -15 °C with stirring for 15 min before DBU (20 μL, 0.013 mmol) was added in one portion. Notably, the addition of

DBU caused the solvent to bubble due to an exothermic reaction. After 10 min, the reaction was allowed to warm to room temperature. After 1 h, excess of C3A was dissolved in 5 mL CHCl3 and added into reaction solution. After another 0.5 h, the solution was diluted with CHCl3 (25 mL) and BHT (0.225 g, 10.22 mmol) was added. The polymer solution was then precipitated into diethyl ether (400 mL) and collected by decanting the supernatant and dried by high vacuum system at room temperature for 24 h to yield the

1 pale-yellow polymer with 40% CSS (4.96 g, 90 %). H NMR (CDCl3, 500 MHz) % cis ~ 80 with

40% CSS incorporation (see Figure 7.49). SEC (CHCl3) Mn = 26.7 kDa, Mw = 57.0 kDa, Ð M =

2.13. DSC: Tg = -7.4 C, Tc = 12.7 C, Tm = 82.5 C. TGA: Td = 349 C. Tensile tests: E0 = 36.2

± 2.0 MPa, break = 2110 ± 50 %, UTS = 36.2 ± 4.1 MPa.

Random copolymer with 50% CSS incorporation: CSS (1.789 g, 6.716 mmol), C3A

(2.420 g, 13.43 mmol) and C10S (1.386 g, 6.716 mmol) were added to a 100 mL round bottom flask with 25 mL CHCl3. The solution was then cooled to -15 °C with stirring for 15 min before DBU (20 μL, 0.013 mmol) was added in one portion. Notably, the addition of

1 pale-yellow polymer with 50% CSS (4.92 g, 88 %). H NMR (CDCl3, 500 MHz) % cis ~ 80 with

50% CSS incorporation (see Figure 7.51). SEC (CHCl3) Mn = 45.4 kDa, Mw = 94.9 kDa, Ð M =

2.09. DSC: Tg = -9.7 C, Tc = 21.5 C, Tm = 74.2 C. TGA: Td = 348 C. Tensile tests: E0 = 36.2

± 2.0 MPa, break = 1943 ± 155 %, UTS = 37.4 ± 4.5 MPa.

Random copolymer with 60% CSS incorporation: CSS (2.147 g, 8.060 mmol), C3A

(2.420 g, 13.43 mmol) and C10S (1.109 g, 5.373 mmol) were added to a 100 mL round bottom flask with 25 mL CHCl3. The solution was then cooled to -15 °C with stirring for 15 min before DBU (20 μL, 0.013 mmol) was added in one portion. Notably, the addition of

DBU caused the solvent to bubble due to an exothermic reaction. After 10 min, the

reaction was allowed to warm to room temperature. After 1 h, excess of C3A was dissolved

in 5 mL CHCl3 and added into reaction solution. After another 0.5 h, the solution was

diluted with CHCl3 (25 mL) and BHT (0.225 g, 10.22 mmol) was added. The polymer

solution was then precipitated into diethyl ether (400 mL) and collected by decanting the

supernatant and dried by high vacuum system at room temperature for 24 h to yield the

1 pale-yellow polymer with 60% CSS (5.17 g, 91 %). H NMR (CDCl3, 500 MHz) % cis ~ 80 with

60% CSS incorporation (see Figure 7.53). SEC (CHCl3) Mn = 29.0 kDa, Mw = 71.7 kDa, Ð M =

2.47. DSC: Tg = -12.2 C. TGA: Td = 342 C. Tensile tests: E0 = 12.0 ± 1.4 MPa, break = 2077

± 78 %, UTS = 20.5 ± 0.6 MPa.

5.4. Results and Discussion

Monomer and Polymer Syntheses

In this work, the commercially available 1,10-decandedithiol (C10S), a 10 carbon

aliphatic chain instead of 1,6-hexandithiol, was redistilled to afford the monomer with

99%+ purity to serve as a key functional monomer for a stereocontrolled thiol-yne step-

growth polymerization with 1,3-propane diyl dipropiolate (C3A) to yield the elastomeric

polymer (Mn = 32.6 kDa, Mw = 68.6 kDa, Ð M = 2.24) with high cis content (80-81 %,

identified by J coupling of resonances of vinyl protons in 1H NMR spectrum shown in

Figure 5.1.C and Figure 7.43) by using DBU as a catalyst in chloroform based on our

conditions developed previously.48, 50

In order to investigate the potential crystallinity while maintaining degradation of these elastomeric systems, a copolymerization strategy that incorporates bis(3- mercaptopropyl) succinate (CSS) varied from 20% to 60% feed ratios with the 1,3-propane diyl dipropiolate (C3A) and the distilled 1,10-hexanedithiol (C10S) were employed to afford a series of high cis content copolymers with controlled high molar mass (see Figure 5.1.B and 5.1.C). The % incorporation of bis(mercaptopropyl) succinate (CSS) was calculated quantitatively by the integration ratio of the resonances at 2.64 ppm (from monomer CSS)

1 to the resonances at 1.39 ppm (from monomer C10S) based on the H NMR spectra of the

(co)polymers in solution (CDCl3) (see Figure 5.1.C). The resulting stoichiometry-controlled copolymers each maintain high cis content (80-82 % cis).

Thermal and Mechanical Properties

(Tm). Notably, no melting temperature (Tm) or crystalline temperature (Tc) was observed at 60% CSS incorporated polymer in heating/cooling cycles at 10 C/ min ramp speed. The polymer with 0% incorporation of CSS possessed the highest glass transition temperature

(Tg = -68 °C), melting temperature (Tm = 112.4 °C), and crystalline temperature (Tc = 61.4 o C) while 60% CSS incorporated polymer had the lowest glass transition (Tg = -12.2 °C) and no melting and crystalline temperature which means the material is amorphous instead of semicrystalline (see Figure 5.2.A and 5.2.B). Particularly, the trend of thermal transition from DSC displayed a positive correlation with %C10S content that presents longer aliphatic chain monomer could enhance crystalline property. Thermal stability tests from

TGA provided that the decomposition temperatures at 5 % weight loss of all polymers are above 340 oC presented all materials are thermally stable. All thermal transition data were listed in Table 5.1.

Table 5.1. Molecular masses, thermal and properties of (co)polymers with different molar fraction of bis(3-mercaptopropyl) succinate (CSS) incorporation. [a] % of CSS Mw (kDa) Ð M Tg (C) Tm (C) Tc (C) Td (C)

0 68.6 2.24 -6.8 112.4 61.4 359

20 72.8 2.10 -7.3 99.3 30.3 348

30 60.4 2.08 -7.3 88.9 18.5 349

40 57.0 2.13 -7.4 82.5 12.7 349

50 94.9 2.09 -9.7 74.2 21.5 348

60 71.7 2.47 -12.2 - - 342

1 [a] % of bis(3-mercaptopropyl) succinate (CSS) was quantitatively determined by H NMR spectrometry.

Mechanical properties were investigated for all high cis content elastomers with various % CSS incorporation are listed in Table 5.2. The Young’s modulus, extension at break, and strain at break were all determined from the stress vs. strain curves obtained using uniaxial tensile testing. The Young’s modulus (E) of all polymers are distinctive and highly reverse correlative to % amount of CSS incorporation. The modulus of elastomers ranged from 65.8 ± 0.6 (0% CSS) to 12.0 ± 1.4 (60% CSS) and the strain at break varied from

1457± 112% (0% CSS) to 2077 ± 78 (0% CSS). Mechanical property is highly tunable and dependent on the amount of CSS content resulted from increasing the amount of CSS which is a longer, bulkier comonomer providing the flexibility of the chain packing that relatively elongated the strain at break (break) but decreased modulus based on the uniaxial tensile tests.

Figure 5.2. Thermal and mechanical properties of stereocontrolled polymers with different % of bis(3-mercaptopropyl) succinate (CSS) incorporation. Stacked thermogram of differential scanning calorimetry (DSC) at A) second heating cycle shows that the glass transition temperatures (Tg) and melting temperature (Tm) have a positive correlation with %CSS incorporation and B) second cooling cycle displays the fraction of crystallinity have the same trend. C) Overlapped data of thermogravimetric analysis (TGA) were performed to determine the degradation profile for each species. D) Increasing the amount of (Css) which is a longer, bulkier comonomer relatively reduced the UTS and the modulus of the resulting elastomers based on the uniaxial tensile tests. Mechanical property is highly tunable depending on the amount of Css content.

Table 5.2. Mechanical properties of (co)polymers with different molar fraction of bis(3- mercaptopropyl) succinate (CSS) incorporation.

% of CSS  cis break UTS Elasticity (%) (MPa) (%) 0 81 1457 ± 112 32.2 ± 3.9 15 20 80 1700 ± 31 40.5 ± 2.2 20 30 80 1638 ± 33 28.3 ± 1.2 20 40 80 2110 ± 50 36.2 ± 4.1 25 50 80 1943 ± 155 37.4 ± 4.5 20 60 80 2077 ± 78 20.5 ± 0.6 28

In Vitro Accelerated Degradation Studies

The hydrolytic stability was assessed for each of the %CSS elastomers and the respective molar masses were listed in Table 5.1. Degradation studies were conducted using thin films prepared by compression molding (50 m thickness) and tested using accelerated degradation conditions in 5 M NaOH solution (n=3) at 37 °C + 5% CO2. The time for the polymers to be fully degraded ranged from 4 weeks to 15 weeks. The results show that degradation rates are highly dependent on the amount of CSS incorporation

(see Figure 5.3.A). The highest %CSS content polymer (60% CSS) provides more flexibility to increase the chain mobility matched the DSC thermograms (see Figure 5.2.A and 5.2.B) and helps to accelerate the degradation rate through increased water penetration while the polymer with 80% C10S incorporation with relative higher molar fraction of crystallinity increases the hydrophobicity that allows to decrease the degradation rate. The proposed mechanism of water penetration and degradation process are shown in Figure 5.3.B.

5.5. Conclusion

A longer chain dithiol monomer, 1,10-decanedithiol, was used in combination with

bis(3-mercaptopropyl) succinate and 1,3-propane diyl dipropiolate to undergo thiol-yne

step-growth random copolymerization yielding a series of biodegradable elastomers. The

polymerization condition by DBU with chloroform was used to direct the stereochemistry

to a consistent 80-82%. Significantly, the respective polymers containing different molar

extents of % succinate incorporation were not only capable of varying semicrystalline

property but also tuning in vitro degradation over a period of ranging from 4 weeks to 15

weeks. These materials will provide a new choice for encapsulation layer in wireless bio-

electronic devices to target desired lifetime with varied mechanical properties including

elasticity and flexibility as well as controlled degradation rate. Future studies on these

materials will include investigation of material water barrier properties from surface

energy, water transport rates from polymer thin film, in vivo degradation, and both in

vitro and in vivo medical device fabrications to test real work functions.

5.6. Acknowledgement

The authors gratefully acknowledge financial support from Duke University.

CHAPTER VI

CONCLUSION

Crosslinked Internal Alkyne-based StereoElastomers: Polymers with Tunable Mechanical

Properties

Methodologies to enhance mechanical properties of polymers and elastomers for industrial applications include “vulcanization” by which polymer chains are crosslinked through chemical bonds. In this work, we introduce a new method to crosslink well- defined, synthetic elastomers using “click” reactions. Specifically, 2-butyne-1,4-diyl dipropiolate which possesses an internal alkyne, was synthesized as a functional monomer and further copolymerized to yield a series of elastomeric materials. Notably, the glass transition temperature and mechanical properties of the resulting copolymers can be tuned by changing the molar fraction of 2-butyne-1,4-diyl dipropiolate. The alkyne functionalities at the polymer chain ends and within the backbone allow for post- polymerization end-group functionalization and interchain crosslinks which form polymer networks using a ruthenium-catalyzed azide-alkyne cycloaddition (RuAAC). Hysteresis tests have shown that tensile modulus and recovery can be controlled by the density of the crosslinking within the network. These findings herein have tremendous potential to

87 be applied to a wide range of biomaterial applications where methods to toughen degradable materials has been extremely limited. The features described in this work are of critical importance for implementing these materials into biomedical devices and efforts toward this end are currently underway.

Shape Memory Behavior of Biocompatible Polyurethane StereoElastomers Synthesized via

Thiol-yne Michael Addition

The current formulation of polyurethanes to yield pure “urethane” linkage without any side product such as urea formation has not been investigated. In this work, the urethane-based dipropiolate, 5,14-dioxo-4,15-dioxa-6,13-diazaoctadecane-1,18-diyl dipropiolate, was successfully synthesized and utilized as a new class of activated dialkyne monomer that can further be polymerized using stereochemically controlled thiol-yne

Michael additions to yield polyurethane elastomers without urea generation. Significantly, the thermal and mechanical properties are tunable by varying the stereochemistry of the alkene unit from 82% to 32% cis content in the polymer backbone. Amorphous polymers possessing hydrogen bonding along the backbone retain their mechanical properties. The thermomechanical analysis of 82% cis content polymer displayed shape memory behavior.

The investigation of accelerated degradation behavior in 5 M NaOH solution up to a three- month period has displayed the materials with reduced %cis content yielded faster degradation rates thus demonstrating that stability of material is highly stereo-chemically dependent. In vitro cell viability of 82% cis content has indicated urethane-based elastomers possess excellent biocompatibility. The combination of biocompatible, 88 biostable, and tunable thermal and mechanical properties supports our material as potentially new candidates for a long-term implantable biomaterial and biomedical device. Nonetheless, biodegradable polymeric materials might be more valuable for soft tissue engineering due to degraded segments would be expected to be non-toxic and resorbable in biological systems. Importantly, a well-known degradable unit such as an ester moiety can be easily incorporated into our polyurethane backbone and formulated as copolymer systems could provide a series of bioresorbable polyurethane elastomers in the future.

Bioresorbable Elastomers with Tunable Crystallinity as Encapsulation Layer in Wireless

Bio-electronics to Enhance Water Barrier

In this work, the well-designed semicrystalline polymeric system has been introduced by utilizing a relative longer chain dithiol monomer, 1,10-decanedithiol, with degradable bis(3-mercaptopropyl) succinate moiety and 1,3-propane diyl dipropiolate to undergo thiol-yne step-growth random copolymerization yielding a series of biodegradable elastomers with tunable thermal and mechanical properties. Considerably, the variants containing % succinate incorporation were not only capable of varying semicrystalline property but also tuning in vitro degradation over a period of ranging from

4 weeks to 15 weeks. The findings from fundamental characterization of materials demonstrate that the biodegradable elastomers could provide a new choice for encapsulation layer in wireless bio-electronic devices to target desired lifetime with varied mechanical properties including elasticity, flexibility, and controlled degradation 89 rate. Future studies on these materials will include investigation of material water barrier properties from surface energy, water transport rates from polymer thin film, in vivo degradation, and both in vitro and in vivo medical device fabrications and tests.

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APPENDICES

111

APPENDIX A-SUPPORTING FIGURES

Figure 7.1. The 1H NMR spectrum of compound 2 shows only resonance a, which confirms full conversion and high purity (CD3OD, 300 MHz).

112

Figure 7.2. The 13C NMR spectrum of compound 2 shows quantitative resonances, which confirm full conversion and high purity (CD3OD, 75 MHz).

113

Figure 7.3. The 1H NMR spectrum of compound 4 shows appearance of resonances a, b and c, coming from the methyl (CH3) and phenol ring (aromatic) of tosyl groups (CDCl3, 300 MHz).

114

Figure 7.4. The 13C NMR spectrum of compound 4 shows appearance of resonances corresponding to the formation of disubstituted tosylation (CDCl3, 75 MHz).

115

116

117

118

Figure 7.8. The 1H NMR spectrum of 1,6-diazidohexane displays a triplet splitting pattern a corresponding to methylene group (CDCl3, 300 MHz).

119

Figure 7.9. The 13C NMR spectrum of 1,6-diazidohexane displays only three resonances that confirm the disubstituted formation of the diazido compound (CDCl3, 75 MHz).

120

121

122

Figure 7.12. SEC chromatogram of I100 of thiol-yne step-growth polymer with 100% incorporation of I; Mn = 48.5 kDa, Mw = 79.9 kDa, Ð M = 1.7 (SEC DMF with 0.1 M LiBr, based on PS standards).

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124

Figure 7.14. SEC chromatogram of I70A30 of thiol-yne step-growth polymer with 70% incorporation of I; Mn = 49.4 kDa, Mw = 87.6 kDa, Ð M = 1.8 (SEC DMF with 0.1 M LiBr, based on PS standards).

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126

Figure 7.16. SEC chromatogram of I30A70 of thiol-yne step-growth polymer with 30% incorporation of I; Mn = 53.8 kDa, Mw = 108.2 kDa, Ð M = 2.0 (SEC DMF with 0.1 M LiBr, based on PS standards).

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128

Figure 7.18. SEC chromatogram of I10A90 of thiol-yne step-growth polymer with 10% incorporation of I; Mn = 31.4 kDa, Mw = 105.3 kDa, Ð M = 3.4 (SEC DMF with 0.1 M LiBr, based on PS standards).

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131

Figure 7.21. The overlapped thermogravimetric analysis (TGA) data were performed to determine the degradation profile for each species.

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133

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Figure 7.27. The 13C NMR spectrum of compound 2 displayed resonance 4 at 156.47 ppm corresponding to carbonyl group (C=O) of urethane (CDCl3, 75 MHz).

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Figure 7.31. SEC chromatogram of 82% cis content of thiol-yne step-growth polymer; Mn = 52.9 kDa, Mw = 94.4 kDa, Ð M = 1.8 (SEC DMF with 0.1 M LiBr, based on PS standards).

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Figure 7.33. SEC chromatogram of 71% cis content of thiol-yne step-growth polymer; Mn = 51.3 kDa, Mw = 98.2 kDa, Ð M = 1.9 (SEC DMF with 0.1 M LiBr, based on PS standards).

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Figure 7.35. SEC chromatogram of 62% cis content of thiol-yne step-growth polymer; Mn = 56.0 kDa, Mw = 93.5 kDa, Ð M = 1.7 (SEC DMF with 0.1 M LiBr, based on PS standards).

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Figure 7.37. SEC chromatogram of 46% cis content of thiol-yne step-growth polymer; Mn = 45.8 kDa, Mw = 100.6 kDa, Ð M = 2.2 (SEC DMF with 0.1 M LiBr, based on PS standards).

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Figure 7.39. SEC chromatogram of 32% cis content of thiol-yne step-growth polymer; Mn = 34.5 kDa, Mw = 74.0 kDa, Ð M = 2.1 (SEC DMF with 0.1 M LiBr, based on PS standards).

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Figure 7.44. SEC chromatogram of P1 (81% Cis) of thiol-yne step-growth polymer; Mn = 32.5 kDa, Mw = 72.8 kDa, Ð M = 2.24 (SEC CHCl3, based on PS standards).

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153

Figure 7.46. SEC chromatogram of P2 (80% Cis) of thiol-yne step-growth polymer with 20% incorporation of CSS; Mn = 32.6 kDa, Mw = 68.6 kDa, Ð M = 2.10 (SEC CHCl3, based on PS standards).

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155

Figure 7.48. SEC chromatogram of P3 (80% Cis) of thiol-yne step-growth polymer with 30% incorporation of CSS; Mn = 29.1 kDa, Mw = 60.4 kDa, Ð M = 2.08 (SEC CHCl3, based on PS standards).

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157

Figure 7.50. SEC chromatogram of P4 (80% Cis) of thiol-yne step-growth polymer with 40% incorporation of CSS; Mn = 26.7 kDa, Mw = 57.0 kDa, Ð M = 2.13 (SEC CHCl3, based on PS standards).

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1 Figure 7.51. The H NMR spectrum of P5 (with 50% CSS) shows the 80% cis content for the thiol-yne step-growth copolymer with 50% incorporation of CSS in CHCl3 with 1 mol% DBU. The ratio of resonance n (from CSS) to resonance h (from C3A) displays 49%: 51% that affords polymer P5 with 49% incorporation of CSS (CDCl3, 500 MHz).

159

Figure 7.52. SEC chromatogram of P5 (80% Cis) of thiol-yne step-growth polymer with 50% incorporation of CSS; Mn = 45.4 kDa, Mw = 94.9 kDa, Ð M = 2.09 (SEC CHCl3, based on PS standards).

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Figure 7.54. SEC chromatogram of P6 (80% Cis) of thiol-yne step-growth polymer with 60% incorporation of CSS; Mn = 29.0 kDa, Mw = 71.7 kDa, Ð M = 2.47 (SEC CHCl3, based on PS standards).

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Figure 7.55. Exemplar stress vs. strain curves for (co)polymers (0%, 20%, 30%, 40%, 50%, 60% CSS) tested at 10 mm/min. Data for 3 samples are shown to illustrate the reproducibility.

Figure 7.56. Illustration of in vitro medical device fabrication.

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APPENDIX B-SUPPORTING SCHEMES

Scheme 7.2. The synthetic route for urethane-based dipropiolate monomer (U6).

164

Scheme 7.3. The general thiol-yne step-growth polymerization for U6T6 polymers

Scheme 7.4. The synthesis of 1,3-propane diyl dipropiolate (C3A).

Scheme 7.5. The synthesis of Bis(3-mercaptopropyl) succinate (CSS).

165