MECHANICAL AND SWELLING - DiVA portal571374/FULLTEXT01.pdf · MECHANICAL AND SWELLING PROPERTIES OF...

MECHANICAL AND SWELLING

PROPERTIES OF HYDROGELS

Ting Yang

AKADEMISK AVHANDLING

Som med tillstånd av Kungliga Tekniska högskolan i Stockholm, framlägges till offentlig

granskning för avläggande av teknisk doktorsexamen torsdag 6 december 2012, kl. 10:00 i

sal F3, Lindstedtsvägen 26, KTH, Stockholm. Avhandlingen försvaras på engelska.

Fakultetsopponent: Professor Jöns Hilborn från Uppsala University, Sweden

Copyright © 2012 Ting Yang

All rights reserved

Paper I © 2011 Wiley Periodicals, Inc.

Paper II © 2012 Wiley Periodicals, Inc.

TRITA-CHE-Report 2012: 63

ISSN 1654-1081

ISBN 978-91-7501-471-5

ABSTRACT

Hydrogels have been used as one of the novel soft materials in many biomedical

applications such as drug delivery and tissue engineering for recent decades.

In the main part of this work, bi-functional poly(ethylene glycol) (PEG) precursors with

either thiols (PEG-SH) or allyls (PEG-Al) , covering molecular weights from 3 kDa to 8

kDa were synthesized and thoroughly characterized by 1H NMR,

13C NMR, FT-Raman

and MALDI-TOF techniques. By combining PEG precursors with complementary

trifunctional crosslinkers, a library of well-defined single-network hydrogels was

efficiently constructed via the robust UV-initiated thiol-ene coupling (TEC) chemistry.

Novel sequential interpenetrating network (seqIPN) hydrogels based on PEG were

fabricated by diffusing and afterwards crosslinking secondary-network precursors within

dense (2 kDa) to loose (8 kDa) primary networks. The impacts of polymer chain length

and diffusion time on the swelling and mechanical properties were assessed for the seqIPN

hydrogels. Additionally, disperse red 13 decorated PEG 2 kDa and 8 kDa were synthesized

and used as probes to monitor the secondary-network precursor diffusion rate by UV/Vis

spectroscopy.

FT-Raman and leaching tests were conducted to evaluate the efficiency of the TEC

reaction for the development of PEG networks and their gel fractions. All gels were fully

crosslinked within 5 minutes and with the gel fraction above 84%. The chain length of

PEG, location of functional groups of PEGs, solvents, solid content were found to have

directly influence on the mechanical and swelling properties of PEG single-network

hydrogels. The utilization of the diffusion time dependent seqIPN strategy enabled further

freedom to control the swelling and mechanical properties of PEG hydrogels, with the

degree of water swelling ranged from 280 – 870% and the tensile modulus ranging from

1135 kPa to 175 kPa.

Furthermore, the seqIPN strategy was utilized for fiber reinforced free radical polymerized

hydrogels. N, N-dimethylacrylamide (DMA) with crosslinker poly(ethylene glycol)

diacrylate were diffused in bacterial cellulose (BC) aerogel thereafter UV crosslinked to

form BC-DMA hydrogels. FT-Raman and leaching tests were conducted to evaluate the

efficiency of the free radical polymerization and the BC-DMA gel fractions. After UV cure

for 10 minutes, robust DMA networks were formed within BC aerogels with over 94% gel

fraction. The high porosity and robust interpenetrating DMA network within BC fibers

were further analysed with FE-SEM. Compression tests showed that fiber reinforced DMA

hydrogels have higher compression modulus than DMA hydrogels, ranging from 4.4 to 8.3

MPa with water content from 78 to 70%.

SAMMANFATTNING

Under de senaste decennierna har hydrogeler studerats som nytt mjukt material och även

tillämpats i biomedicinska applikationer såsom produktion av läkemedel och inom

vävnadsteknik.

I detta arbete har bifunktionell poly(etylenglykol) (PEG) med antingen tioler (PEG-SH 3 kDa till 8 kDa) eller allyler (PEG-Al 3 kDa) syntetiserats och karakteriserats med hjälp av

tekniker såsom 1H-NMR,

13C-NMR, FT-Raman samt MALDI-TOF. Kompletterande

trifunktionella tvärbindare användes för att tvärbinda dessa bifunktionella PEG-system till

väldefinierade nätverk (hydrogeler) med hjälp av robust UV-initierad tiol-en kemi (TEC).

Av PEG-Al-systemet (2 kDa till 8 kDa) tillverkades även sekventiella interpenetrerande

nätverk (seqIPN), där monomanerna till det sekundära nätverket fick diffundera in i ett

tidigare tvärbundet nätverk, varpå det sekundära nätverket också tvärbands. Påverkan av

diffusionstiden hos det sekundära nätverket på seqIPN hydrogelernas egenskaper

utvärderades. Vidare utnyttjades disperse red 13-funktionaliserad PEG 2 kDa och 8 kDa

(PEG-röd) för att utvärdera diffusionshastigheten hos det sekundära nätverket med hjälp av

UV/Vis spektroskopi. Mätningar visade att diffusionshastigheten påverkades av tätheten

hos det primära nätverket, med en lägre diffusionshastighet för tätare tvärbundna nätverk.

Effektiviteten hos TEC-reaktionen utvärderades genom att studera omsättningen och

gelfraktionen av de tillverkade hydrogelerna med FT-Raman och urlakning av oreagerad

monomer. Studien visade att samtliga geler var helt tvärbundna efter 5 minuter med en

gelfraktion på över 84 %. Det visade sig även att längden på PEG-kedjan, placeringen av

funktionella grupper längs PEG-kedjan, vilket lösningsmedel som användes samt andelen

fast material i gelerna hade en direkt inverkan på både svällningsegenskaperna och de

mekaniska egenskaperna hos singelnätverk av PEG. Genom att utnyttja den

diffusionsstyrda seqIPN-strategin erhölls större frihet att kontrollera svällningen och de

mekaniska egenskaperna hos hydrogelerna. Detta resulterade i geler med en svällning i

vatten från 280 till 870 % samt en E-modul mellan 175 och 1135 kPa.

SeqIPN-metoden användes även för att tillverka fiberarmerade friradikalpolymeriserade

hydrogeler. N, N-dimetylakrylamid (DMA) och poly(etylenglykol)diakrylat fick

diffundera in i aerogeler av bakteriell cellulosa (BC), varpå systemet tvärbands genom

exponering för UV-ljus och fiberförstärkta BC-DMA-hydrogeler bildades. Rena

polyDMA-hydrogeler (DMA) tillverkade med samma tvärbindningsdensitet användes som

referens. Omsättningen och gelfraktionen hos nätverk av polyDMA utvärderades med FT-

Raman och urlakning. Efter UV-härdning i 10 minuter erhölls robusta nätverk av DMA i

BC-aerogeler med gelfraktioner på över 94 %. Den höga porositeten hos IPN av DMA i

BC-aerogeler analyserades sedan med FE-SEM. Kompressionstest visade att de

fiberförstärkta nätverken hade bättre kompressionegenskaper än hydrogeler av endast DMA. Mätningar visade att BC-DMA hydrogelerna hade kompressionsmoduler mellan 4,4

och 8,3 MPa samt en vattenhalt mellan 78 och 70 %.

献给爷爷

Dedicated to My Grandfather

LIST OF PAPERS

This thesis is a summary of the following papers:

I. ‘Characterization of Well-Defined Poly(Ethylene Glycol) Hydrogels

Prepared By Thiol-Ene Chemistry’ Yang, T.; Long, H.; Malkoch, M.;

Gamstedt, E. K.; Berglund, L.; Hult, A. Journal of Polymer Science Part A:

Polymer Chemistry 2011, 49, 4044–4054.

II. ‘Sequential Interpenetrating PEG Hydrogels Prepared by UV initiated Thiol-

Ene Coupling Chemistry’ Yang, T.; Malkoch, M.; Hult, A. Journal of

Polymer Science Part A: Polymer Chemistry 2012, published. DOI:

10.1002/pola.26393.

III. ‘The Influence of Diffusion Time on the Properties of Sequential

Interpenetrating PEG Hydrogels’ Yang, T.; Malkoch, M.; Hult, A. Journal of

Polymer Science Part A: Polymer Chemistry 2012, submitted.

IV. ‘Mechanical properties of N, N-dimethylacrylamide hydrogels reinforced

with bacterial cellulose aerogel’ Yang, T.; Hult, A. manuscript.

This thesis contains unpublished results.

ABBREVIATIONS

AEOBA 4-(2-(allyloxy)ethoxy)-4-oxobutanoic acid

Al Allyl

BC Bacterial cellulose

BC-DMA Bacterial cellulose reinforced n, n-dimethyl acrylamide hydrogel

C2,r Mass concentration of polymer in solution before crosslinking Cn Characteristic ratio of the polymer

CDCl3 Deuterated chloroform

Da Daltons

DCC Dicyclohexylcarbodiimide

DCM Dichloromethane

DMA N, N-dimethyl acrylamide

DMAP 4-dimethylaminopyridine

DPTS 4-(dimethylamino)pyridinium 4-toluenesulfonate

E Young’s modulus from tensile test or tensile modulus

Ec Young’s modulus from compression test or compression modulus

EtOH Ethanol

FE-SEM Field emission scanning electron microscopy FRP Free radical polymerization

FT Fourier transform

G Shear modulus

G′ Shear storage modulus

Gf Gel fraction

Gf I Primary network gel fraction

Gf II Secondary network gel fraction

HABA 2-(4-hydroxyphenylazo) benzoic acid

IPN Interpenetrating polymer network

II% Secondary network solid content

Irgacure 651 UV-initiator l Average bond length

Ma Methacrylate

MALDI-TOF MS Matrix-assisted laser desorption/ionization mass spectrometry

Mc Molecular weight between crosslinks

Mc,S Molecular weight between crosslinks estimated from swelling

Mc,T Molecular weight between crosslinks estimated from tensile

modulus

MeOH Methanol

Mn Number molecular weight

Mr Molecular weight of the repeat unit

n Number of repeating units md Weight of dry hydrogel

ms Weight of equilibrium swelling hydrogel

mc Weight of cured hydrogel before dry

NaHSO4 Succinic anhydride, sodium hydrogen sulfate

NMR Nuclear magnetic resonance

PEG Polyethylene glycol

PEG-Al Polyethylene glycol functionalized with allyl group

PEG-Ma Polyethylene glycol functionalized with methacrylate group

PEG-SH Polyethylene glycol functionalized with thiol group

PTSA P-toluene sulfonic acid

Q Volume-swelling ratio

qF After cure weight swelling ratio

qW Equilibrium weight swelling ratio

R Gas constant semiIPN Semi interpenetrating polymer network

seqIPN Sequential interpenetrating polymer network

SH Thiol

T’ Temperature of tensile test

Tdiffusion Secondary network precursors diffusion time

TEC Thiol-ene coupling

THF Tetrahydrofuran

TMP Trimethylolpropane

TTT 1, 3, 5-triallyl-1, 3, 5-triazine-2, 4, 6(1H, 3H, 5H)-triazine

UV Ultra violet

V1 Molar volume of solvent Vis Visible

v2,r Volume fraction of polymer in relaxed gel

v2,s Volume fraction of swollen gel

wt% Weight percent

εB Stress-at-break

ν Poisson’s ratio

ξ Mesh size

ρc Crosslinking density

ρp Polyethylene glycol density

ρsol Solvent density

ρwater Water density

σB Strain-at-break χ Polymer solvent interaction parameter

Specific volume of polymer

TABLE OF CONTENTS

1 PURPOSE OF THE STUDY ................................................................................ 1

2 INTRODUCTION ................................................................................................ 2

2.1 HYDROGELS .................................................................................................... 2 2.2 HYDROGEL CROSSLINKING CHEMISTRY............................................................. 3

2.2.1 Thiol-ene coupling chemistry ................................................................... 3 2.2.1 Functionalized poly(ethylene glycol) ........................................................ 3 2.2.1 Free radical polymerization ..................................................................... 4

2.3 HYDROGEL NETWORKS .................................................................................... 5 2.3.1 Single network ......................................................................................... 5 2.3.2 Interpenetrating polymer networks (IPN) ................................................. 6 2.3.1 Bacterial cellulose (BC)........................................................................... 7 2.3.2 Cellulose-reinforced hydrogels ................................................................ 9

2.4 IMPORTANT PROPERTIES OF HYDROGEL ............................................................ 9 2.4.1 Swelling properties .................................................................................. 9 2.4.2 Mechanical properties ........................................................................... 10

2.5 APPLICATIONS ............................................................................................... 11

3 EXPERIMENTAL .............................................................................................. 13

3.1 MATERIALS ................................................................................................... 13 3.2 INSTRUMENTATION ........................................................................................ 14 3.3 FABRICATION OF PEG HYDROGELS WITH TRIAZINE-BASED CROSSLINKER......... 15 3.4 FABRICATION OF SEQUENTIAL-IPN HYDROGELS WITH TMP-BASED CROSSLINKER18

3.4.1 Preparation of single-network TEC hydrogels ........................................ 18 3.4.2 Preparation of chain length influenced seqIPN TEC hydrogels............... 18 3.4.3 Preparation of diffusion time influenced seqIPN and assessment of the

diffuse rate ........................................................................................................... 20 3.5 PROPERTY ASSESSMENT OF PEG HYDROGELS ................................................. 21

3.5.1 Gel fraction determination by leaching test ............................................ 21 3.5.2 Swelling test .......................................................................................... 22 3.5.3 Structure evaluation .............................................................................. 23

3.5.3.1 Determination of the average molecular weight between crosslinks (Mc,S) and mesh size (ξ) from swelling profile ................................................................................. 24 3.5.3.2 Determination of the average molecular weight between crosslinks (Mc,T) and mesh size (ξ) from the tensile modulus ........................................................................... 24

3.6 FABRICATION OF BACTERIAL-CELLULOSE-REINFORCED HYDROGELS ................ 26 3.6.1 Preparation of BC aerogels ................................................................... 26

3.6.2 Fabrication of DMA and BC-DMA hydrogel .......................................... 26 3.1 PROPERTY ASSESSMENT OF DMA AND BC-DMA HYDROGEL .......................... 28

4 RESULTS AND DISCUSSION .......................................................................... 30

4.1 POLYMER PRECURSORS .................................................................................. 30 4.2 PEG HYDROGELS AND PROPERTY ASSESSMENT ............................................... 30

4.2.1 Influence of solvent and functional group location on hydrogels mechanical

properties ............................................................................................................. 32 4.2.2 Influence of UV-irradiation cure time and PEG chain length on hydrogel

properties ............................................................................................................. 33 4.2.3 PEG-Al and PEG-SH hydrogel structure evaluation............................... 34

4.3 SEQUENTIAL-IPN HYDROGELS PERPARATION AND PROPERTIES ........................ 35 4.3.1 Single-network hydrogels and assessment of their properties .................. 35 4.3.2 PEG chain length influence on seqIPN................................................... 37 4.3.3 Secondary-network diffusion time influence on seqIPN........................... 41

4.4 PROPERTIES OF BACTERIAL-CELLULOSE-REINFORCED DMA HYDROGEL .......... 49

5 CONCLUSIONS ................................................................................................. 56

6 FUTURE WORK ................................................................................................ 58

7 ACKNOWLEDGEMENTS ................................................................................ 59

8 REFERENCES ................................................................................................... 61

Purpose of study

1

1 PURPOSE OF THE STUDY

Hydrogels as novel soft material have been studied for bio-application for over fifty years.

The good biocompatibility of hydrogels emanates from their higher water content (over

70%), but it is also a limitation to the mechanical properties so as the applications. Hereby,

establishing a database relating initial materials, fabrication factors and methods to

swelling and mechanical properties of hydrogels has been the main purpose of this study.

Preparing and assessing well-defined novel hydrogels have been the main part of the study

in which thiol and allyl bi-functionalized poly(ethylene glycol) (PEG) (Mn = 3 kDa to 8k

Da) were used to form hydrogels with the complementary trifunctional crosslinkers via

UV-initiated thiol-ene coupling chemistry. To improve while control the mechanical

properties of PEG hydrogels, sequential interpenetrating networks (seqIPN) were

introduced by diffusing secondary PEG network into dense (2k Da) to loose (8k Da)

crosslinked single networks. To further assess the properties of seqIPN, the influence of

secondary-network diffusion time in dense (2k Da) and loose (8k Da) network hydrogels

was studied. Swelling and tensile tests were conducted to evaluate the swelling and

mechanical properties of PEG hydrogels. Furthermore, PEGs decorated with disperse red

13 were used to investigate the diffusion rate by UV/Vis spectroscopy.

An alternative seqIPN approach was further developed based on fiber reinforced

hydrogels. N, N-dimethylacrylamide (DMA) was used as monomer and combined with

poly(ethylene glycol)diacrylate crosslinker to fabricate hydrogels within the bacterial

cellulose (BC) aerogels via UV-initiated free radical polymerization (BC-DMA).

Compression and rheology tests were conducted to assess the mechanical properties of

BC-DMA hydrogels.

Introduction

2

2 INTRODUCTION

2.1 HYDROGELS

Hydrogels are hydrophilic three-dimensional (3D) networks that are chemically

crosslinked or physically entangled with excellent water swelling capacity.1 On a

molecular level, water in a hydrogel is either bonding to polar hydrophilic groups as ‘bond

water’ or is filling the space between the network chains, pores or voids as ‘free water’.2

As a kind of rapidly developing new material, many scientific reports have been published

since 1870s with focus on the preparation, characterization and applications of hydrogels.2

Hydrogels are characterized as soft material with high water content, which is similar to

soft tissue, so they have good biocompatible properties and have been exploited in many

fields such as food additives, pharmaceuticals, cell culture3 and biomedical implants.4 For

example, polyethylene glycol based hydrogels with water content over 90% are used for

wound healing and N, N-dimethylacrylamide based hydrogels are used for commercial soft

contact lens (Figure 1).

Figure 1. A commercial soft contact lens contains N, N-dimethylacrylamide based

hydrogel.

Introduction

3

2.2 HYDROGEL CROSSLINKING CHEMISTRY

2.2.1 Thiol-ene coupling chemistry

Thiol-ene coupling (TEC) chemistry is an organic reaction between a thiol and an alkene

(Figure 2), which was discovered in 1905.5 The advantages of TEC are high efficiency,

high yielding in the presence of oxygen, tolerance of various solvents and functional

groups.5 In recent years, researchers have considered the TEC reaction to be a "click"

reaction.6 This reaction has been extensively applied in coating,7 nano-printing,8 adhesive

technology9 and in dendrimer chemistry or other well-defined structure syntheses.10 The

TEC addition reaction is typically facilitated by UV irradiation and proceeds through a

thiyl radical species. When TEC utilized in polymer chemistry, the polymerization follows

a free radical, step-growth mechanism.11

Figure 2. Scheme of thiol-ene coupling reaction. R and R’ contains thiol and allyl groups

respectively.

2.2.1 Functionalized poly(ethylene glycol)

Poly(ethylene glycol) (PEG) is a common hydrophilic polymer and characterized as a soft

biocompatible material.3,12,13 The U. S. Food and Drug Administration (FDA) approved

PEG usage in the pharmaceutical, food, and cosmetics industries.14 PEG is a commercial

product made from ethylene oxide with water and ethylene glycol or ethylene glycol

oligomers. PEGs can be functionalized as telechelic polymers for hydrogels preparation.15-

18 Figure 3 shows the chemical structure of PEG and the PEG bi-functionalized by thiol,

Introduction

4

allyl and methacrylate groups, which were used for TEC hydrogels preparation in this

work.

Figure 3. Poly(ethylene glycol) (PEG) and thiol, allyl, methacrylate functionalized PEG.

2.2.1 Free radical polymerization

Free radical polymerization (FRP) is a conventional route to prepare hydrogels.19-21 N, N-

dimethylacrylamide (DMA), which has been studied for biomedical applications,22-24 is a

liquid chemical used as monomer to co-polymerize with crosslinkers into a hydrogel.

Figure 4 shows the chemical structure of a DMA monomer, a homo-polymer of DMA and

a network of DMA after copolymerization with acrylate or methacrylate multifunctional

crosslinkers to form a polymer network. The advantages of FRP are easy preparation, fast

curing and can be performed in an aqueous solution.

Introduction

5

Figure 4. N, N-dimethyl acrylamide (DMA), PolyDMA and a DMA network.

2.3 HYDROGEL NETWORKS

Network is the basic structure of a hydrogel, which is covalently crosslinked or physically

entangled. The network components of hydrogels can be either synthetic or natural, as long

as they absorb and maintain large quantities of water.25

2.3.1 Single network

In the main parts of this work, telechelic polymers based on PEG were used to fabricate

TEC networks. The molecular weight between crosslinks (Mc) is the number average

molecular weight between the crosslinking junctions and is a key component correlated to

diffusion properties of a network.26 The length in between the crosslinking junctions is

mesh size (ξ).27 Figure 5 (a) illustrates an ideal network formed by bi-functional PEG and

complementary tri-functional crosslinkers via TEC, mesh size (ξ) or Mc of this network is

controlled by the polymer chain length.

Introduction

6

Figure 5. (a) TEC network formed by tri-functional crosslinker and bi-functional polymer.

(b) FRP network formed by monomers and bi-functional crosslinker.

Another hydrogel network used in this thesis is FRP network, which is formed by

monomers and bi-functional crosslinkers. Figure 5 (b) illustrates the FRP network that has

a random mesh size or Mc because of the different reactivity between the monomer homo-

polymerization and the monomer and crosslinker copolymerization; besides, the

polymerization can be terminated by different termination mechanisms.28

2.3.2 Interpenetrating polymer networks (IPN)

An interpenetrating polymer networks (IPN) is a system containing multiple physically

interlaced inseparable networks without covalent bonding between them.29 IPN

methodology is an approach to improve hydrogel mechanical properties.30-32 IPNs can be

classified according to two schemes: their synthesis, simultaneous IPN or sequential IPN

(seqIPN), or their structure, full IPN33 and semiIPN.34 Simultaneous IPN is prepared by

forming polymer networks independently in one system via non-interfering reaction

mechanisms,35 which is not included in this work. The seqIPN is prepared by forming the

secondary network after the primary network formation by diffusion.36-38

A seqIPN

hydrogel can be prepared by diffusing either polymer precursors or monomers with

crosslinker into a primary network, thereby crosslinking a secondary network within the

primary network. The semiIPN is prepared by diffusing a linear polymer into the primary

Introduction

7

network without crosslinking.39,40 Figure 6 shows full seqIPN formation, by TEC and FRP,

use UV curing and semiIPN formation use linear polymer. In all cases the components of

the secondary network are diffused into the primary network.

Figure 6. Full IPN and Semi IPN formation via sequential diffusion process.

2.3.1 Bacterial cellulose (BC)

Bacterial cellulose (BC) hydrogel is one of the most common natural cellulose which is

produced by acetic bacterium "Acetobacter xylinum".41,42 BC was first discovered in

1886,43-45 it has been intensively studied since 1947 and has ever since attracted scientists’

attention.46 Although cellulose is insoluble in water and most organic solvents,47 the long

and refined nanoscale fibers of BC can form a physically entangled network and contains

around 98% water.48 Different from polymeric hydrogels, the hydrogen bonding between

water molecules and BC are mostly on the fiber surface, large amounts of free water are

filling the voids in between BC fibers, so the water in a cellulose hydrogel can be squeezed

Introduction

8

out easily. The advantages of BC hydrogel are not only green and sustainable49 with high

average molecular weight and crystallinity,50

but also are high dimensional stability after

converting the BC hydrogel into the respective aerogels.51,52 Figure 7 (a) and (b) is a

photograph of the raw BC hydrogels and the freeze-dried BC aerogels respectively. Figure

8 (c) demonstrates the FE-SEM picture of a BC network. Figure 7 (d) illustrates a scheme

of the hierarchical structure of a BC cellulose fiber, from the basic glucose chain to the

crystalline micro fibril and cellulose fiber.53,54

Figure 7. (a) Raw BC hydrogels, (b) freeze-dried BC aerogel, (c) FE-SEM picture of cross

section from BC aerogel, (d) scheme of BC fibrils structure: glucose chains, micro fibrils,

and cellulose fibers.

Introduction

9

2.3.2 Cellulose-reinforced hydrogels

Cellulose whiskers have been used as reinforcement to improve the mechanical properties

of hydrogels.55,56 Different from the cellulose whiskers, BC aerogel with the long fiber and

large voids in between the fibers is a suitable scaffold to improve the mechanical

properties of the brittle FRP hydrogels.

2.4 IMPORTANT PROPERTIES OF HYDROGEL

2.4.1 Swelling properties

A crosslinked polymer hydrogels swell but not dissolve when water or a solvent enters it.

The swelling properties, which usually use degree of swelling to define hydrogels, depend

on many factors such as network density, solvent nature, polymer solvent interaction

parameter.1 The properties of water swelling of PEG hydrogels and PEG diffusion in PEG

networks were studied in this work.

Because water acts as a plasticizer in a hydrophilic polymer network system, the swelling

process of the hydrogel can be considered under rubbery state and can be described by the

free energy of mixing ΔGmix from the polymer and solvent interaction and the elastic free

energy ΔGelastic from the crosslinked network:57

ΔGsystem = ΔGmix + ΔGelastic

At the beginning of swelling, the ΔGmix << 0, ΔGelastic > 0, ΔGmix + ΔGelastic < 0, so the

swelling is favoured and the solvent diffuses into the network. During the processing of

swelling, the ΔGmix and ΔGelastic both increased until │ΔGmix │ =│ΔGelastic │ and ΔGsystem =

ΔGmix + ΔGelastic = 0, so that the driving force for swelling is gone: equilibrium swelling is

reached and swelling stops.

Introduction

10

2.4.2 Mechanical properties

Mechanical tests were conducted to assess the hydrogel properties. To establish a library of

mechanical properties of hydrogels is to gather the information of the hydrogel network

and to determine the range of application. 58

Tensile properties were the mainly studied mechanical properties of PEG based hydrogels

in this work. Figure 8 displays a photo of a hydrogel sample in a tensile testing machine.

The hydrogel samples are cut into a dumbbell shape prior to assessment.

Figure 8. Tensile test of a hydrogel sample in an Instron tensile testing machine.

The compression and rheology properties were used to assess the mechanical properties of

fiber-reinforced hydrogels in this work. Rheology is the study of the flow of matter,

measurements being prepared by shearing the sample, as shown in Figure 9.

Introduction

11

Figure 9. Rheology measurements on a hydrogel sample.

2.5 APPLICATIONS

The hydrogels prepared in this work covered a large range of mechanical properties, from

a soft hydrogel such as PEG to a hard hydrogel such as BC-DMA. Because of the high

water content and biocompatibility of hydrogels, many applications are related to

biomedical usage.59,60

Drug delivery is a method of administering a pharmaceutical compound to achieve a

therapeutic effect in or at a certain location in the human body.61,62 Materials used as drug

delivery systems need to have controlled properties, such as absorption and release profile

and no toxicity.63 PEG hydrogels in this work can be used as biodegradable drug delivery

systems, because of the biodegradable ester bond in the structure, the well-controlled

structure and properties;64 most importantly, PEG is a FDA approved polymer for use

inside the human body.65

Because of the low toxicity of its precursors66

and crosslinkers,

PEG can be used to form a hydrogel in situ via TEC. The seqIPN PEG hydrogels can also

change the mechanical properties to suit the applications. Figure 10 pictures a diagram of a

degradable PEG hydrogel as a drug-delivery system.

Introduction

12

Figure 10. Diagram of a drug-delivery system uses a biodegradable PEG hydrogel carrying

drugs and releasing at certain location.

Tissue engineering involves improving or replacing biological functions of the human

body with artificial materials.67,68 Tissue engineering requires certain mechanical and

structural properties for proper functioning for the repair or replacement of portions of the

human body.69 For example, a hydrogel for artificial cartilage needs to have a compression

stress of 0.5 to 10 MPa.70 It needs to be biocompatible and to have interconnected macro-

pores for cell culture.71 Bacterial-cellulose-reinforced DMA hydrogels could be good

candidates for this application.

Experimental

13

3 EXPERIMENTAL

3.1 MATERIALS

PEG (Mn: 2 kDa, 3 kDa, 6 kDa, 8 kDa), trimethylolpropane tris(3-

mercaptopropionate) (TMP-tris-thiol), 2,2-dimethoxy-1,2-diphenylethan-1-one

(IRGACURE 651) and tris[2-(3-mercaptopropionyloxy) ethyl] isocyanurate (TMI),

disperse red 13, 3-mercaptopropionic acid, 99% (MPA), p-toluene sulfonic acid

(PTSA), dicyclohexylcarbodiimide (DCC), 1,3,5-triallyl-1,3,5-triazine-

2,4,6(1H,3H,5H)-triazine (TTT), tris[2-(acryloyloxy)ethyl] isocyanurate13,4-

dimethylaminopyridine, D-glucose, sodium hydroxide, N,N-dimethylacrylamide

(DMA), alpha-ketoglutaric acid, polyethylene glycol diacrylate (Mn=700 Da) were

purchased from Sigma-Aldrich. Chloroform, toluene-4-sulfonic acid monohydrate,

dichloromethane (DCM), diethyl ether, 2-(4-hydroxyphenylazo) benzoic acid

(HABA) were purchased from Fisher Scientific Merck. Ethanol (EtOH, 96% and

100%) was purchased from VWR. Succinic anhydride, sodium hydrogen sulfate

(NaHSO4), 4-dimethylaminopyridine (DMAP), toluene-4-sulfonic acid

monohydrate, diethylether, methanol (MeOH), tetrahydrofuran (THF), toluene and

deuterated chloroform (CDCl3) were purchased from Fisher Scientific Merck. All

the chemicals were used as received. 4-(dimethylamino)pyridinium 4-

toluenesulfonate (DPTS) was prepared as reported elsewhere.72 The synthesis of 4-

(2-(allyloxy)ethoxy)-4-oxobutanoic acid is described in a previous publication.73

Experimental

14

3.2 INSTRUMENTATION

Nuclear Magnetic Resonance (NMR) spectra were recorded with a Bruker Avance

400 MHz instrument using CDCl3 as solvent. The solvent signal was used as

internal standard.

Field Emission Scanning Electron Microscopy (FE-SEM) pictures were collected

from a Hitachi S-4800 scanning electron microscope operated at 1 to 1.5 kV.

MALDI-TOF MS spectra (MALDI-TOF) were conducted on a Bruker UltraFlex

MALDI-TOF MS with SCOUT-MTP Ion Source (Bruker Daltonics, Bremen)

equipped with a N2 laser (337 nm), a gridless ion source and reflector design

FT-Raman spectra were acquired for all samples using a Perkin-Elmer Spectrum

2000 NIR FT-Raman instrument. Each spectrum was based on 128 scans using 800

mW laser powers. The conversion of thiols (2573–2568 cm-1) or vinyl acrylate

(1635 cm-1), allyl (1640-1643 cm-1) was determined using the carbonyl bond (1760-

1763 cm-1) as an internal reference.

UV curing was performed using a Blak-Ray xx-15BlB UV bench lamp for PEG

hydrogels, where the wavelength of the lamp was 365 nm and the intensity of the

UV radiation was 28.5 mW/cm2. Fushion UV cure system model F300 equipped

with Fusion electrodeless bulbs standard type BF9 (Lamp power 300 W/inch, 1800

W total) was used for cuing DMA and BC-DMA hydrogels.

Tensile and compression tests were conducted with a universal testing machine

Instron 5944 with an advanced non-contacting video extensometer (Instron Korea

LLC.) at 23°C and 50% relative humidity using a cross-head speed of 100 mm/min.

For tensile tests, the swollen hydrogels were cut into a dumbbell shape to prevent

unwanted fracture close to the clamping region and to avoid grip slippage.

Compression tests on water swollen hydrogels were performed at 50% relative

humidity and 23°C, using the Miniature materials tester Minimat 2000 with load

cells of 20 N and 200 N.

Rheology tests were performed in TA AR2000 rheometer. An 8 mm sample holder

disk is used for all the samples.

Experimental

15

UV/Vis absorption spectrum was determined by SHIMADZU UV-2550 UV/Vis

spectroscopy. The PEG and PEG-red mixture solution concentration was set to 50

wt% in EtOH. PEG-red-2k (0.05 wt%) and PEG-red-8k (0.01 wt%) as marked

concentration was determined by software UVprobe 2.34.

3.3 FABRICATION OF PEG HYDROGELS WITH TRIAZINE-

BASED CROSSLINKER

To study the formation of the hydrogels using TEC chemistry, PEGs were

functionalized with thiol (PEG-SH) and allyl (PEG-Al) to prepared TEC hydrogels

with the complementary tri-functional triazine based crosslinkers. All the hydrogels

were prepared in 3 to 6 replicas with a thiol to allyl molar ratio set to 1:1 with 3

wt% initiator (Irgacure 651). Solvents and solid content were varied to study the

efficient of TEC in different reaction conditions. Figure 11 presents the scheme of

PEG hydrogels fabrication; Table 1 details the sample codes for the single network

hydrogels.

Experimental

16

Figure 11. Schematic of PEG hydrogel networks formation. Thiol-ene hydrogel

prepared by P-Al: allyl-functionalized PEG; P-SH: thiol functionalized PEG; P-Ma:

methacrylate hydrogel prepared by methacrylate functionalized PEG.

Experimental

17

Table 1. Single-network PEG hydrogels based on triazine crosslinker.

Sample code Mn of

PEG

(Da)

Functional

group

Solvent used

in hydrogels

formation

UV-

irradiation

Time

(minutes)

Solid

content

(wt%)

M-P3-SH-15-1 3000 Thiol (SH) Methanol (M) 15 56

M-P3-SH-30-1 3000 Thiol Methanol 15 56

E-P3-SH-15-1 3000 Thiol Ethanol (E) 15 56

E-P3-SH-30-1 3000 Thiol Ethanol 15 56

M/W-P3-SH-

15-1

3000 Thiol Methanol/water

(W)

(50:50 wt%)

15 56

T-P3-Al-1 3000 Allyl (Al) THF 1 56

T-P3-MA-1 3000 Methacrylate THF 1 56

T-P3-SH-1 3000 Thiol THF 1 56

E-P3-Al-1 3000 Allyl Ethanol 1 56

E/B-P3-Al-1 3000 Allyl Ethanol/butyl

acetate (B)

(50:50 wt%)

1 56

E/B-P3-Al-15 3000 Allyl Ethanol/butyl

acetate (B)

(50:50 wt%)

15 56

E-P3-SH-1 3000 Thiol Ethanol 1 56








Experimental

18

3.4 FABRICATION OF SEQUENTIAL-IPN HYDROGELS

WITH TMP-BASED CROSSLINKER

3.4.1 Preparation of single-network TEC hydrogels

Four molecular weights PEGs (2 kDa, 3 kDa, 6 kDa and 8 kDa), equimolar ratio

(1:1) with crosslinker TMP-tris-thiol and Irgacure 651 (3 wt%) were dissolve in

ethanol (EtOH) (50 wt% solid content) respectively to prepare single-network

hydrogels. Using TMP based crosslinker TMP-tris-thiol instead of triazine based

crosslinker was because the better solubility in EtOH for the hydrogel preparation

process. Each mixture was vortexed to homogeneous and poured into a Teflon mold

(thickness: 0.10 cm, width: 1.0 cm) and was covered with a glass slide to prevent

EtOH evaporation. The mixture solution was allowed to gel for 5 minutes under UV

exposure (365 nm, 28.5 mW/cm intensity, Blak-Ray xx-15BlB UV bench lamp) at

room temperature. A small part of the cured hydrogel was cut and dried overnight

for gel fraction tests. The rest of the hydrogels were immersed in deionized water

for 5 hours and immersed in ethanol (100%) for 3 hours to remove residues. The

cleaned hydrogels were dried in air overnight and then placed in a vacuum oven at

40 °C for 1 hour. The single-network hydrogel sample code is S2 prepared from

PEG-Al, Mn = 2 kDa (PEG-Al-2k); hydrogel S3 prepared from PEG-Al, Mn = 3

kDa; hydrogel S6 prepared from PEG-Al, Mn = 6 kDa; and hydrogel S8 prepared

from PEG-Al, Mn = 8 kDa, all hydrogels were prepare in 6 to 10 replicas, the S8

hydrogels were prepared in at least 12 replicas.

3.4.2 Preparation of chain length influenced seqIPN TEC

hydrogels

The dried single-network hydrogels, S2, S3, S6 and S8, were used as primary

network to prepare seqIPN hydrogels. All the primary networks were immersed in

ethanol solutions containing (50 wt% solid content) PEG-Al precursors, crosslinker

TMP-tris-thiol and initiator (3 wt%), the molar ratio of thiol to allyl was set to 1:1.

Experimental

19

The precursors in solutions were allowed to diffuse into the primary networks for 2

hours at 40 °C. The excess of solution on the surface of primary network hydrogels

was removed with tissue paper and was placed in a Teflon mold covered by a glass

slide to prevent solvent evaporation. The secondary networks were allowed to

crosslink for 5 minutes under UV exposure (365nm, 28.5 mW/cm). The crosslinked

seqIPN hydrogels were dried in air overnight and then placed in a vacuum oven at

40 °C for 1 hour. Figure 12 shows the scheme of seqIPN hydrogel preparation.

Figure 12. Preparation of seqIPN, using allyl functionalized PEG (Mn= 2k, 3k, 6k,

8k Da).

Experimental

20

3.4.3 Preparation of diffusion time influenced seqIPN and

assessment of the diffuse rate

Dense and loose single network, 2 kDa (S2) and 8 kDa (S8); hydrogels were used to

assess the diffusion time influence on mechanical properties of seqIPN hydrogels.

S2 and S8 hydrogels were immerged in the ethanol solution of PEG-Al precursors

(2 kDa and 8 kDa) with crosslinker TMP-tris-thiol and initiator (50 wt% solid

content) in an oven at 40°C for 2h, 4h, 20h and 44 hours diffusion time. The

secondary networks were allowed to cure for 5 minutes under UV exposure (365

nm, 28.5 mW/cm2). All seqIPNs were prepared in 12 replicas with a thiol to allyl

molar ratio set to 1:1 with 3 wt% initiator to assess the properties at totally four

diffusion times. The cured seqIPN hydrogels with four different diffusion times

were dried in air overnight and then placed in a vacuum oven at 40°C for 1 hour

respectively.

PEG 2kDa and 8 kDa were dissolved in EtOH to prepare 54.5 mg/mL and 138 mg/l

PEG solution with PEG-red-2k (0.05 wt%) and PEG-red-8k (0.01 wt%)

respectively. S2 and S8 hydrogels were immersed in a 1 mL PEG-red-2k and PEG-

red-8k solution respectively. A total of 24 pieces of samples were placed in an oven

at 40 ° C, the remaining solutions were used to analyse the PEG diffusion after 2, 4,

20, 25, 28 and 44 hours by UV/Vis absorption spectroscopy.

The scheme of diffusion time dependence seqIPN and semiIPN preparation is

demonstrated in Figure 13.

Experimental

21

Figure 13. Preparation of diffusion time dependent IPNs and semi IPN hydrogel

from 2 kDa and 8 kDa PEG.

3.5 PROPERTY ASSESSMENT OF PEG HYDROGELS

3.5.1 Gel fraction determination by leaching test

Leaching tests were conducted to identify any unreacted starting materials and to

determine the gel fraction. The hydrogels were dried in the vacuum oven to remove

the solvent directly after curing, totally three replicas of each hydrogel sample

(thickness: 0.10 cm, length: 1.0 cm, width: 1.0 cm) were weighed (dry weight M)

and immersed in chloroform, one in deuterated chloroform for NMR analysis. All

Experimental

22

samples were swelled for 5 hours with three cycles of solvent exchange. The

leached hydrogels were dried overnight in air and placed in a 40°C vacuum oven

for half hour to remove water and the dried weight was measured as M’. The

leachate in deuterated chloroform was analyzed by NMR to determine remaining

functional groups. The gel fraction of the hydrogel was calculated by the following:

Gel fraction = M’/M ×100% (1)

where M is the mass of the dry hydrogels after cure and M' is the mass of the dry

hydrogels after leaching.

3.5.2 Swelling test

Three or four replicas of each dried hydrogel were swollen in deionized water at

room temperature for 3 days to achieve equilibrium swelling. The degree of

swelling of hydrogels were measured after 5 min, 10 min, 20 min, 0,5 h, 1,5 h, 1

day, 2 days and 3 days. Totally three replicas were measured, the standard

deviations were marked with error bars in the swelling profile charts. The degree of

swelling was calculated as the following:

Degree of swelling = [(Wet weight – Dry weight) / Dry weight] ×100% (2)

The water content of hydrogels were calculated after the equilibrium swelling by

Water content = (Wet weight / Dry weight) ×100% (3)

The molecular weight between cross-links (Mc), effective crosslinking density (ρ),

and mesh size (ξ) were estimated according to the equilibrium swelling result.74

The water-induced volume-swelling ratio75 of hydrogel was calculated as:

Experimental

23

(4)

where v2,r is the volume fraction of polymer in the relaxed gel (hydrogels after cure,

before dry) and v2,s is the volume fraction polymer of swollen gel. These variables

are related to weight fractions by:

,

(5)

[

]

(6)

and [

]

(7)

where is the weight swelling ratio of hydrogels after equilibrium swelling, is

the weight of equilibrium swollen gel, is the weight of dry gel, is the weight

of gel after curing, is the weight ratio after curing. The PEG density is denoted

(1.12 g/cm3), is the density of water (1.00 g/cm3), and is the density

of solvent used in hydrogel cure process (EtOH: 0.79 g/cm3, EtOH/butyl acetate:

0.93 g/cm3).76

3.5.3 Structure evaluation

The structure evaluation is based on swelling and tensile tests result, the average

molecular weight between crosslinks (Mc) and mesh size (ξ) of PEG hydrogel were

calculated. Chain lengths between cross-links of PEG networks were assumed to

follow Gaussian distribution and the equilibrium swelling theory of Flory for

crosslinked polymers.

Experimental

24

3.5.3.1 Determination of the average molecular weight between crosslinks (Mc,S)

and mesh size (ξ) from swelling profile

The Mc,S was calculated with the following equation where ‘S’ indicates swelling77

–

( ) ( ) ( )

( ) (

)

–

(8)

where Mn is the number-average molecular weight of the polymer, is the specific

volume of polymer (0.84 cm3/g for PEG),78 V1 is molar volume of solvent (18

cm3/mol for water) χ is the polymer solvent interaction parameter (0.43 for PEG -

water)76 and is assumed constant in this work.

3.5.3.2 Determination of the average molecular weight between crosslinks (Mc,T)

and mesh size (ξ) from the tensile modulus

Due to the isotropic homogeneous crosslinking of PEG hydrogels, the shear

modulus G can be calculated from the tensile Young’s modulus E according to79:

(9)

where v is Poisson's ratio. When the hydrogel material is fully swollen, its

mechanical behavior is similar to that of an incompressible rubber-like material.

Hence, the Poisson ratio of the hydrogels was assumed to be 0.5 in this work.80

Mc,T, where ‘T’ indicates tensile stiffness,77 was calculated from the equation:

(10)

where C2,r is the mass concentration of polymer in solution before crosslinking (0.9

kg/l in ethanol, 0.86 kg/l in ethanol/butyl acetate mixture), is the gas constant

Experimental

25

(8.31 kPa l/mol K), and T’ is the temperature 298 K at which the tensile testing was

carried.

The crosslinking density ρc was determined 81 by:

(11)

The mesh size ξ was then determined as the root-mean-square of the end-to-end

distance of the polymer chain in the unperturbed state is

(12)

where l is the average bond length (0.146 nm), is the characteristic ratio of the

polymer (typically 4.0 for PEG)82 and n is the number of repeating units in the

crosslink:

(13)

where is the molecular weight of the repeating unit (44 for PEG). The mesh size

ξ can then be expressed as82

(14)

With the above expressions, structural evaluation of the molecular network can be

estimated by tensile measurements and swelling.

Experimental

26

3.6 FABRICATION OF BACTERIAL-CELLULOSE-

REINFORCED HYDROGELS

3.6.1 Preparation of BC aerogels

The bacterial cellulose (BC) was obtained by cultivating the bacterium Acetobacter

in a pre-culture HS medium. This cultivating medium was prepared by pre-

cultivating Acetobacter aceti strain in the Hestrin–Schramm (HS) medium 27 for 7

days at 27 °C. This pre-culture medium (5 mL) was used to inoculate 30 mL of

fresh HS medium for bacterium cultivation. The BC hydrogels were harvested after

7 days of culture at 27 °C under static conditions. They were treated with 0.1 M

NaOH at 80°C for 3 h and washed with de-ionized water. This process was repeated

3 times and the BC hydrogels were finally washed with de-ionized water for several

days until neutrality was reached. After the purified BC has been obtained, scissors

was used to cut the bacterial cellulose into 1x1x1 cm cubic shape and they were

then freeze-dried for 2 days to prepare the BC aerogels.

3.6.2 Fabrication of DMA and BC-DMA hydrogel

N, N-dimethyl acrylamide (DMA) hydrogel was prepared by FRP in aqueous

solution with polyethylene glycol diacrylate (Mn= 700 Da, PEGDA) as crosslinker

and 2-oxo-ketoglutaric acid (5 wt%) as a UV initiator. The DMA monomer (10, 40

and 70 wt%) and crosslinker PEGDA (2 or 4 mol%) were cured in aqueous solution

under 10 minutes UV irradiation (Lamp powder 300 W/inch). Figure 14 presents a

diagram of DMA and its network chemical structure.

Experimental

27

Figure 14. PolyDMA network formation via UV-initiated FRP.

The BC aerogels were used as templates for the preparation of fiber reinforced

DMA hydrogels. All BC aerogels were immersed in deionized aqueous solutions

containing 10, 40, 70 wt% DMA, respectively and initiator (5 wt%). The molar

ratio of DMA to PEGDA was set to 4 mol% for 10, 70 wt% DMA solution and 2

mol% to 40 wt% DMA solution. The solutions were allowed to diffuse into the BC

aerogels for one week at room temperature. Any excess of solutions on the surface

of the hydrogels were thereafter dried with a piece of tissue paper and the gels were

carefully placed into a Teflon mold with a glass slide on top to prevent solvent

evaporation. The DMA networks were allowed to crosslink with PEGDA for 10

minutes under Fusion UV irradiation belt with a total dose of 500 mJ/cm2, three

replicas of each samples were prepared for further analysis. One of three cured BC-

DMA replicas for each sample was subsequently freeze-dried for FT-Raman

analysis and gel fraction assess. The other 2 replicas were immersed in water for

one week to ensure the removal of any unreacted precursors and reaction residues,

water was changed several time each day. The purified hydrogels were dried in air

for 2 days and then placed in a vacuum oven at 40°C overnight. Figure 15

illustrates the diagram of BC-DMA hydrogel formation.

Experimental

28

Figure 15. Scheme of BC-DMA hydrogel networks formation.

3.1 PROPERTY ASSESSMENT OF DMA AND BC-DMA

HYDROGEL

To obtain the gel fraction of DMA network a leaching study was conducted. One

replica of 3 samples freeze-dried BC-MDA hydrogel with mass (M) weighed (170

to 230 mg) including any unreacted starting materials was immersed in 200 mL of

deionized water. The deionized water was exchanged every day for one week. All

swollen hydrogels were air dried for 2 days and placed at 50°C in a vacuum oven

overnight. The fully leached and dried sample mass M’ was recorded. The gel

fraction (crosslinking efficiency) of all samples was acquired from mass before and

after leaching using the formula below:

Gel fraction = (M’/M) ×100% (15)

Content of the DMA network within the BC-DMA hydrogel was calculated from

the mass increase after leaching the samples from any unreacted precursors. In this

Experimental

29

case, the dried BC aerogel mass is marked as MBC, the dried BC-DMA mass is

marked as MBC-DMA, enabling the calculation of the secondary-network content:

DMA network content = [(MBC-DMA – MBC) / MBC-DMA] × 100% (16)

Water content of the swollen DMA and BC-DMA hydrogels were collected on

three replicas. The hydrogel samples were dried in air for 2 days and further dried at

50°C in vacuum oven overnight. The drying procedure was completed upon

reaching constant mass loss values. Then after the samples were immersed in

deionized water for one week and the wet weight of each sample was recorded, the

water content was calculated from the formula:

Water content = (Wet weight / Dry weight) ×100% (17)

Results and Discussion

30

4 RESULTS AND DISCUSSION

4.1 POLYMER PRECURSORS

PEGs were functionalized with thiol (3 kDa to 8 kDa) (PEG-SH), allyl (3 kDa) (PEG-Al),

and methacrylate PEG (3 kDa). PEG based precursors were used to study the effect of

chain length, location of functional groups of PEGs, solvents, solid content of TEC

hydrogels. PEG-Al (2 kDa to 8 kDa) was prepared to study the effect of chain length on

single and seqIPN hydrogels. PEG-Al (2 kDa and 8 kDa) was prepared to study the effect

of diffusion time of seqIPN hydrogels. Additionally, PEGs (2 kDa and 8 kDa) were

decorated with disperse red 13 to probe the diffusion rate of PEGs in single-network

hydrogels. 1H NMR and 13C NMR and MALDI-TOF were used to confirm the fully

functionalization of PEG precursors. FT-Raman revealed the S-H stretching peak at 2571

cm-1 and C=C stretching at 1645cm-1. The UV/Vis absorption spectrum of disperse red 13

evidenced a characteristic absorption peak at 503 nm.

4.2 PEG HYDROGELS AND PROPERTY ASSESSMENT

All PEG hydrogels were prepared with a ratio of PEG functional group to crosslinker

functional group of 1:1 with 3wt% UV-initiator. The UV crosslinking of the three systems

was initially conducted in THF (56 wt%). The FT-Raman spectroscopy was used to assess

the conversion for P3-Al, P3-SH, and P3-MA after 1 minute UV-irradiation time. As

shown in Figure 16(I) there is no trace of the thiol stretching peak at 2570 cm-1 for the

PEG-SH hydrogel and no trace of allyl C=C stretching peak at 1645 cm-1 from PEG-Al

hydrogel, Figure 16 (II) and not for PEG-Ma hydrogel at 1645 cm-1, Figure 16 (III). The


31

gel fraction tests of hydrogels were performed by leaching in chloroform. Hydrogels

prepared in ethanol reveals over 92% gel fraction, which is much higher than hydrogels

prepared in THF, 68% and 72% (T-P3-Al-1, T-P3-SH-1), presented in Table 2. The P3-

MA hydrogel (T-P3-Ma-1) was brittle and it broke into small pieces during leaching

process, therefore it was not possible to assess the gel fraction.

Figure 16, FT-Raman spectra of I A, P3-Al, B, PEG-Al with trithiol-triazine, C, Cured

Hydrogel T-P3-Al; II A, P3-SH, B, PEG-Thiol with triallyl-triazine, C, Cured Hydrogel T-

P3-SH; III A, P3-Ma, B, PEG-Methacrylate with triacrylate-triazine, C, Cured hydrogel T-

P3-Ma.


32

Table 2. Gel fraction of hydrogels prepared by TEC and FRP. Abbreviation: T:

tetrahydrofuran, E: ethanol, E/B: ethanol/ butyl acetate (50:50 wt%), 1: UV irradiation

time: 1 minute; 15: UV irradiation time: 15 minutes; 30: UV irradiation time: 30 minutes.

Sample

code

T-P3-

Al-1

T-P3-

SH-1

T-P3-

Ma-1

E-P3-

Al-1

E/B-P3-

Al-1

E/B-P3-

Al-15

E-P3-

SH-1

E-P3-

SH-15

E-P6-

SH-1

E-P6-

SH-15

E-P8-

SH-1

E-P8-

SH-15

E-P8-

SH-30

Gel

fraction

(%)

68 72 - 95 92 97 99 99 99 99 99 99 99

4.2.1 Influence of solvent and functional group location on

hydrogels mechanical properties

The influence of solvent and solvent content using during polymerization on the hydrogel

network formation was studied. Table 3 presents the properties of hydrogels prepared with

56 wt% solid content. The hydrogel (E/B-P3-Al-1) prepared in EtOH/ butyl acetate

exhibits a tensile modulus of 444, which is much higher than the hydrogels prepared in

THF (T-P3-Al-1, 0.15 kPa). For hydrogels, PEG-Al system hydrogels E-P3-Al-1 revealed

higher mechanical properties compare to the same chain length PEG-SH hydrogels E-P3-

SH-1, which were assessed in this work. Because thiol-ene reaction has one side reaction

that is thiyl-thiyl radical coupling into disulfide, which was revealed to have more

influence of PEG-SH based hydrogels than PEG-Al based hydrogels. The disulfide

formation between PEG-SH based hydrogel E-P3-SH-1 (tensile moduli 139 kPa)

decreased the network density and demonstrated lower mechanical properties compare to

PEG-Al hydrogel E-P3-Al-1 (tensile moduli 311 kPa).


33

Table 3. Tensile modulus (E), stress-at-break (σB), strain-at-break (εB) of hydrogels

prepared from PEG-Al, and PEG-SH and PEG-Ma in different solvents with 56 wt% solids

content.

Sample code E

(kPa)

σB

(kPa)

εB

(%) Solvent

T-P3-Al-1 0.15 - - THF

T-P3-SH-1 23 30 81 THF

T-P3-MA-1 388 111 35 THF

E/B-P3-Al-1 444 411 171 EtOH / butyl acetate

(50:50 wt%)

E/B-P3-Al-15 214 199 135 EtOH / butyl acetate

(50:50 wt%)

E-P3-Al-1 311 281 152 EtOH

E-P3-SH-1 139 127 127 EtOH

E-P3-SH-15 104 89 126 EtOH

Abbreviation: 1: UV irradiation time: 1 minute, 15: UV irradiation time: 15 minutes.

4.2.2 Influence of UV-irradiation cure time and PEG chain length

on hydrogel properties

After 1 or 15 minutes of UV irradiation, properties of PEG-SH hydrogels (3 kDa, 6 kDa, 8

kDa) are detailed in Table 4. Gel fraction is 99% for all hydrogels, which is evidence of a

high conversion of the TEC network for both 1 and 15 minutes UV-irradiation cure time.

For longer chain length PEG-SH, longer cure time needed for better mechanical properties,

for hydrogel E-P3-SH-1 (PEG-SH, 3 kDa), after 1 minute cure time, the tensile moduli is

139 kPa, after 15 minutes cure time (E-P3-SH-5), tensile moduli decreased to 104 kPa.

For 6 kDa hydrogel E-P6-SH-15, after 1 minutes cure time, the tensile moduli is 2 kPa,

after 15 minutes, the tensile moduli is 20 kPa. The tensile modulus decreased from 139

kPa to 3 kPa, when the chain length of PEG-SH increased from 3 kDa to 8 kDa (E-P3-SH-

1 to E-P8-SH-30).


34

Table 4. Gel fraction (Gf), tensile modulus (E), stress-at-break (σB), strain-at-break (εB) of

PEG-SH hydrogels with 3k, 6k, 8k molecular weight and different UV irradiation times (1,

15, 30 minutes).

Hydrogel sample code UV-irradiation time

(minute)

Gf

(%)

E

(kPa)

σB

(kPa)

εB

(%)

E-P3-SH-1 1 99 139 127 127

E-P3-SH-15 15 99 104 89 126

E-P6-SH-1 1 99 2 9 171

E-P6-SH-15 15 99 20 15 95

E-P8-SH-1 1 99 3 - Over 200

E-P6-SH-30 30 99 17 20 153

E-P8-SH-15 15 99 2 4 169

E-P8-SH-30 30 99 3 6 177

4.2.3 PEG-Al and PEG-SH hydrogel structure evaluation

The structural properties of a selection of hydrogel systems were evaluated using swelling

and tensile tests on the fully cured PEG-Al, PEG-SH (3 kDa and 6 kDa) hydrogels. As can

be seen in Table 5, the average molecular weight between crosslinks (Mc) for 3 kDa

system hydrogels prepared by PEG-Al is 1179 g/mol, which is a bit lower than PEG-SH,

ca. 1438 g/mol. For PEG-SH system, 6 kDa hydrogel E-P6-SH-15 Mc (2512 g/mol) is ca. 2

times higher than 3 kDa system. The volume-swelling ratio (from 2.7 to 8.6) and the mesh

size (4.8 to 11.7 nm) increased with increasing PEG chain length, from 3 kDa to 6 kDa, the

crosslinking density decreased from 10 to 4.1.


35

Table 5. The average molecular weight between crosslinks (Mc), volume-swelling ratio

(Q), crosslinking density (ρc), and mesh size (ξ) of PEG allyl and PEG thiol hydrogels.

Indices ‘S’ and ‘T’ denote whether the property was determined from swelling or from

tensile tests, respectively.

Hydrogel

sample code

Mc,S

(g/mol)

Mc,T

(g/mol)

Q ρc,S

(×10-4

)

ρc,T

(×10-4

)

ξS

(nm)

ξT

(nm)

E-P3-Al-1 1179 1367 2.7 10 8.7 4.8 5.1

E-P3-SH-1 1438 1420 5.7 5.6 5.6 6.7 6.7

E-P6-SH-15 2512 2934 8.6 4.1 4.7 11.7 11.7

4.3 SEQUENTIAL-IPN HYDROGELS PERPARATION AND

PROPERTIES

The seqIPNs were introduced to the PEG hydrogel systems by diffusing secondary

networks into the crosslinked primary network. To prepare robust TEC PEG hydrogels,

PEG-Al system was used to crosslink with tri-thiol functionalized crosslinker (TMP-tir-

SH) for seqIPN hydrogel fabrication. Although the other TEC side reaction that is head to

head coupling of the carbon centred radicals from allyl could happen in PEG-Al system,

because of the auto-inhibition of the allylic compounds, this side reaction is much less

likely to happen during network formation.83

4.3.1 Single-network hydrogels and assessment of their properties

Single-network hydrogels as primary hydrogel were prepared by PEG-Al (Mn= 2, 3, 6, 8

kDa) with TMP-tir-Allyl crosslinker in EtOH. The solids content of the precursor solution

was 50 wt% with 3 wt% UV-initiator and the molar ratio of functional groups were set to

1:1. Figure 17 shows the FT-Raman spectra for PEG-Al (8 kDa) and the single-network

hydrogels S2, S3, S6 and S8. There is no trace of the C=C vibrations of allyl group at 1640


36

cm-1 in the cured hydrogel. Also in Table 6 evinces that the gel fraction reached 97% in all

single-network hydrogels, which correspond to the full TEC conversion.

Figure 17. FT-Raman spectra of 8 kDa PEG-Al and single-network hydrogel 2, 3, 6 and 8

kDa (S2, S3, S6 and S8).

Figure 18. (I), the tensile modulus and (II), degree of swelling of the single-network

hydrogel.


37

As shown in Figure 18 (I), the tensile modulus decreased from 555 to 175 KPa with

increasing PEG chain length from 2 kDa to 8 kDa, the water content of hydrogels

increased from 79 to 90% as the corresponding degree of swelling increased from 377 to

871%, Figure 18 (II). Table 6 details the relevant structure properties of the hydrogels

including the gel fraction based on the PEG-Al system and the average molecular weight

between crosslinks. As can be seen, the mesh size increased while the crosslinking density

decreased with increasing chain length of PEG precursors.

Table 6. Gel fraction (Gf), tensile modulus (E), stress-at-break (σB), strain-at-break (εB),

average molecular weight between crosslinks (Mc), mesh size (ξ), volume-swelling ratio

(Q), crosslinking density (ρc) and mesh size (ξ) of single-network hydrogel. Indices ‘S’ and

‘T’ indicate values determined from swelling or tensile tests, respectively.

Sample

code

Gf

%

E

(kPa)

σB

(kPa)

εB

(%)

Q

Mc,S

(g/mol)

Mc,T

(g/mol)

ρc,S×10-4

(mol/mL)

ρc,T×10-4

(mol/mL)

ξS

(nm)

ξT

(nm)

S2 95 555±18 226 65 2.3 766 902 15.5 13.2 3.7 4.0

S3 97 403±9 159 67 2.7 1141 1332 10.4 8.9 4.8 5.2

S6 97 279±22 153 75 3.7 2230 2515 5.3 4.7 7.4 7.9

S8 97 175±1 121 102 4.6 3104 3407 3.8 3.5 9.5 9.9

4.3.2 PEG chain length influence on seqIPN

PEG-Al system (Mn = 2, 3, 6, 8 kDa), PEG-Al-2k, PEG-Al-3k, PEG-Al-6k and PEG-Al-

8k were used to fabricate seqIPN hydrogels. All IPNs were prepared by diffusion of

mixtures containing PEG precursors, crosslinker (1:1 molar ratio) and initiator (3 wt%) in

EtOH (50 wt% solid content) for 2 hours at 40 °C, followed by 5 minutes of UV irradiation

at 365 nm to reach a fully cured system.


38

Figure19. The FT-Raman spectra of PEG, crosslinker and dry hydrogels. a PEG-Al-8k, b

PEG-Al-8k with thiol corsslinker, c single-network hydrogel S8 and d seqIPN hydrogel 8-

8.

The FT-Raman spectra in Figure 19 show that the C=C vibrations of allyl group at 1640

cm-1

were not detected for the dried primary network S8 so as the seqIPN 8-8 hydrogels

based on 8 kDa PEG-Al. Table 7 demonstrates that the secondary-network gel fraction (Gf

II) was higher than 85%, which indicates that seqIPN hydrogels were successfully

prepared by TEC chemistry. The secondary-network solid content decreased with the

increasing chain length of the secondary-network precursor and decreasing primary

network mesh size. The volume fraction of relaxed hydrogel (vr) and volume fraction of

swollen hydrogel (vs) were used to calculate the volume-swelling ratio (Q) and apparent

crosslinking density (ρc,a). The Q value is increased and the crosslinking density decreased

with increasing secondary precursor chain length. These values indicate that the secondary

PEG network structure is also well controlled by the TEC chemistry.

The seqIPN hydrogel swelling kinetic were studied by measuring the degree of swelling

with time. Figure 20 depicts the degree of swelling as a function of time for the IPN

hydrogels based on different PEG chain length. The general trend for all IPNs is a reduced

swelling degree with the introduction of secondary networks based on shorter PEG chains.


39

Figure 20. Degree of swelling of seqIPN based on S8 hydrogel (a) based on S6 hydrogel

(b) based on S3 hydrogel (c) based on S2 hydrogel (d).


40

Seq

IPN

sam

ple

cod

e

Prim

ary

netw

ork

(Da)

Secon

dary

netw

ork

(Da

)

Gf I

(%)

Gf

II

(%)

II

netw

ork

soli

d

con

ten

t

(%)

Sw

ell

ing

(%)

Wa

ter

con

ten

t

(%)

E

(kP

a)

σB

(kP

a)

ε B

(%)

vr

vs

Q ρ

c,a×

10

-4

(mol/

ml)

8-2

8

00

0

2000

97

95

3

4

58

4±

31

86

3

65

±6

0

12

7±

20

54

±2

0.6

0

.1

4.5

4.3

8-3

8

000

3000

97

94

3

2

74

4±

34

87

2

17

±4

6

72

±1

8

41

±9

0.6

0

.1

5.7

2.6

8-6

8

000

6000

97

92

2

5

79

2 ±

4

89

1

92

±2

1

98

±4

65

±9

0.6

0

.1

5.9

2.4

8-8

8

000

8000

97

85

2

1

81

0±

13

89

2

06

±1

6

13

2±

17

85±

15

0.6

0

.1

5.9

2.4

6-2

6

000

2000

94

95

3

9

52

8±

37

84

3

81

±8

5

11

6±

33

36

±5

0.6

0

.1

4.2

4.1

6-3

6

000

3000

94

92

3

2

66

1±

12

87

3

13

±3

7

10

6±

19

44±

10

0.6

0

.1

4.8

3.5

6-6

6

000

6000

94

91

2

2

64

6±

23

89

3

52

±2

3

17

4±

31

69±

19

0.6

0

.1

5.0

4.0

3-2

3

000

2000

97

93

2

6

39

7±

9

80

5

95

±3

0

28

3±

20

62±

37

0.7

0

.2

3.7

5.5

3-3

3

000

3000

97

97

2

8

39

0±

12

80

5

88

±4

0

26

4±

77

58±

11

0.6

0

.2

3.2

5.8

2-2

2

000

2000

97

94

2

7

31

5 ±

8

76

8

89

±5

0

29

2±

26

38±

5

0.7

0

.2

3.0

7.8

Tab

le 7

. S

eqIP

Ns

hydro

gel

s pri

mar

y n

etw

ork

gel

fra

ctio

n (

Gf

I),

seco

ndar

y g

el f

ract

ion (

Gf

II),

the

seco

ndar

y (

II)

net

work

mas

s

con

ten

t an

d t

he

deg

ree

of

swel

ling,

tensi

le m

od

ulu

s (E

), s

tres

s at

bre

ak (

σB),

str

ain a

t b

reak

(ε B

), v

olu

me

frac

tion o

f re

laxed

hy

dro

gel

(v r

), v

olu

me

frac

tion o

f sw

oll

en h

yd

rog

el (

v s),

vo

lum

e sw

elli

ng

rat

io (

Q),

ap

par

ent

cro

ssli

nk

ing d

ensi

ty (ρ c

,a).

*T

he

app

aren

t cr

oss

linkin

g d

ensi

ty i

s dif

fere

nt

from

th

e cr

oss

lin

kin

g d

ensi

ty c

alcu

late

d f

rom

Mc.


41

Figure 21 renders the overall plot of the tensile modulus and water content data for the

PEG chain influence on the seqIPN hydrogels.

Figure 21. The overall plot displaying the range of tensile modulus (top) and the water

content (bottom) of primary and seqIPNs hydrogels.

4.3.3 Secondary-network diffusion time influence on seqIPN

Single-network hydrogels S8 and S2 were used as primary networks to prepare seqIPNs

for diffusion-time studies. The secondary-network precursors PEG-Al-2k and PEG-Al-8k

were allowed to diffuse into primary network hydrogels for 2, 4, 20 and 44 hours.

Neither the FT-Raman spectra in Figure 22 evidence that the allyl group at 1640 cm-1 was

detected for the dried primary network S8 nor the 2 kDa PEG-Al based IPN 8-2.


42

Figure 22. The FT-Raman spectra of PEG-Al-2k and diffusing time dependent IPN

hydrogels use S8 as primary network and PEG-Al-2k as secondary-network precursor (8-

2). The diffusion time of secondary-network precursor is shown in the brackets.


43

Figure 23. The secondary-network content and tensile modulus of IPN hydrogels (a) 8-2,

(b) 8-8 and degree of swelling of 8-2 (c) and 8-8 at different diffusion time.

In Table 8, the TEC reaction enabled efficient secondary network build up with excellent

gel fraction values of the secondary network (Gf II) of 84% and higher.

For the loosely crosslinked S8 system, in the case of the PEG 2 kDa secondary network,

Figure 23 (a), the percentage of solid content increased from 34% after 2 hours diffusion

(8-2:2) when compared to the maximum obtained level of 64% after 44 hours (8-2:44).

Remarkably, in Table 8, the tensile modulus was found to increase ca. 4 folds when

comparing the fully saturated IPN hydrogel (8-2:44) of 758 kPa with the primary network

S8 (175 kPa) and 2 folds in comparison to the unsaturated seqIPN (8-2:2; 365 kPa). These

results were further corroborated as can be seen in Table 8 by 3 folds enhanced stress-at-


44

break (σB = 320 kPa) and 2 folds of strain-at-break values (εB = 106%) for seqIPN 8-2:44

and in relationship to 8-2:2.

Figure 23 (b), the diffusion of 8 kDa secondary-network precursors in S8 scaffolds reached

a maximum solid content of 38% after 44 hours of diffusion (8-8:44). These results

demonstrated a 60% lower solid content of 8 kDa secondary network when compared to

the 2 kDa secondary network. Nonetheless, with the increasing solid content within the

seqIPN 8-2, from 18% to 38% (2 to 44 hours of diffusion), the tensile modules increased

from 206 kPa to 324 kPa (8-2:2 to 8-2:44).

Figure 23 (c) displays the degree of swelling of 8-2 systems supressed by increasing

content of denser secondary network (2 kDa) in 8-2 system, which was the result of

increasing diffusion time. Figure 24 (c) shows that with the same crosslinking density of

primary and secondary network, the increasing secondary solid content still increased the

crosslinking density and decreased the degree of swelling.


45

Figure 24. The secondary-network content and tensile modulus of seqIPN hydrogels (a) 2-

2, (b) 2-8 and degree of swelling of 2-2 (c) and 2-8 at different diffusion time.

Similar diffusion behaviour was displayed for the 2 kDa secondary-network precursors

within densely crosslinked S2 primary network, Figure 24 (a) and (b). In Table 8, seqIPN

2-2:2 (with 2 hours diffusion time) exhibited 27% secondary-network solid content and

tensile moduli of 889 kPa. The maximum diffusion was reached after 44 hours and the

fabricated seqIPN 2-2:44 revealed a secondary-network solid content of 49% and a water

swelling capacity of 280%. The seqIPN 2-2:44 displayed the highest tensile modulus of

all fabricated networks with a value of 1135 kPa, which can be compared with the primary

network S2 of 555 kPa. Interestingly, the strain-at-break first decreased from 65% for S2


46

to 38% for the seqIPN 2-2:2 and then increased to 78% after 44 hours of diffusion (seqIPN

2-2:44).

For the 2-8 system, Figure 24 (b), is different from the other, since the secondary network

is looser than the primary network. From 2 to 44 hours diffusion time, secondary-network

content of seqIPN 2-8 increased from 8% and reached equilibrium to 22%. However, the

tensile modulus increased from S2, 555 kPa to 829 kPa with 8% secondary network and

dropped to 640 kPa with 22% secondary-network content.

The same suppression of degree of swelling is noticeable in Figure 24 (c) for seqIPN 2-2

system with increasing diffusion time and secondary-network content. For seqIPN 2-8,

Figure 24 (d) the degree of swelling depicts decreased and increased with increased

secondary-network content.


47

Sa

mp

le

cod

e

Prim

ary

netw

ork

(Da)

Gf I

(%)

Secon

dary

netw

ork

(Da)

Td

iffu

sio

n

(Hou

r)

II n

etw

ork

soli

d c

on

ten

t

(%)

Gf

II

(%)

Sw

ell

ing

(%)

Wa

ter

con

ten

t

(%)

E

(kP

a)

σB

(kP

a)

ε B

(%)

S8

8000

95

-

- -

- 8

70

±3

0

90

1

75

±1

97±

15

102±

5

8-2

:2

8000

95

2000

2

34

95

58

0±

30

86

36

5±

60

95

±3

3

54

±2

8-2

:4

8000

95

2000

4

53

9

4

52

0±

20

84

4

00

±2

3

120±

20

99±

47

8-2

:20

8000

95

2000

20

62

9

4

41

0±

30

82

7

43

±3

1

276±

22

106±

9

8-2

:44

8000

95

2000

44

64

9

4

38

0±

2

79

7

58

±5

3

320±

62

106±

12

8-8

:2

8000

95

8000

2

18

85

79

0±

30

89

20

6±

16

130±

17

85

±1

5

8-8

:4

8000

95

8000

4

30

8

5

77

0±

80

88

2

89

±1

130±

2

87±

11

8-8

:20

8000

95

8000

20

34

8

4

67

0±

5

86

2

90

±1

130±

3

102±

11

8-8

:44

8000

95

8000

44

38

8

4

66

0±

20

86

3

24

±3

0

260±

5

110±

4

S2

2000

97

-

- -

- 3

80

±4

79

5

55

±1

8

226±

1

65±

5

2-2

:2

2000

97

2000

2

27

9

4

32

0±

8

76

8

89

±5

0

292

±26

38

±5

2-2

:4

2000

97

2000

4

39

9

4

32

0±

2

75

8

90

±3

4

410±

12

73±

5

2-2

:20

2000

97

2000

20

45

94

30

0±

3

75

90

0±

30

44

5±

50

76

±1

2-2

:44

2000

97

2000

44

49

9

4

28

0±

8

74

1

13

5±

23

410±

19

78±

1

2-8

:2

2000

97

8000

2

9

90

30

0±

2

77

8

29

±5

374±

14

83±

6

2-8

:4

2000

97

8000

4

12

9

0

32

0±

4

78

6

55

±1

8

274±

10

83±

1

2-8

:20

2000

97

8000

20

22

8

6

32

0±

8

82

6

59

±1

1

226±

16

81±

15

2-8

:44

2000

97

8000

44

22

8

6

33

0±

4

82

6

40

±1

0

226±

17

81±

4

Tab

le 8.

Sum

mar

y of

pro

per

ties

of

single

an

d S

eqIP

N hy

dro

gel

s in

clu

din

g gel

fr

acti

on (G

f I)

, se

con

dar

y net

work

pre

curs

or

dif

fusi

on

tim

e (T

dif

fusi

on),

sec

ondar

y g

el f

ract

ion

(G

f II

), s

eco

ndar

y (

II)

net

wo

rk s

oli

d c

on

ten

t (I

I%),

th

e eq

uil

ibri

um

deg

ree

of

swel

lin

g, te

nsi

le m

odulu

s (E

), s

tres

s at

bre

ak (σ

B)

and

str

ain

at

bre

ak (ε B

).


48

An overall plot of tensile modulus and water content is displayed in Figure 25.

Figure 25. The tensile modulus (top) and water content (bottom) of the seqIPNs with

different secondary-network diffusion time.

The diffusion rate of secondary-network precursors PEG-red 2 kDa and 8 kDa in PEG

hydrogels were traced by PEG-red (2 kDa and 8 kDa) and assessed by UV/Vis

spectrometry. Figure 26 displays the concentration of the remaining PEG-red in EtOH for

all semiIPNs, because of the single-network hydrogel absorption of the PEG-red, after 44

hours of diffusion the remaining PEG-red is less than the initial solution concentration,

after 2 hours of diffusion, an increase in the concentration are shown in most solutions. For

the PEG-red-8k, the equilibrium was reached after 20 hours of diffusion for the semi-2-8,

after 28 hours the semi-8-8 also reached equilibrium.


49

Figure 26. PEG-red-2k and PEG-red-8k remaining concentration in EtOH versus diffusion

time of semi-8-2, semi-8-8 and semi-2-2 and semi-2-8. UV/Vis absorption peak of PEG-

red at 503 nm.

4.4 PROPERTIES OF BACTERIAL-CELLULOSE-REINFORCED

DMA HYDROGEL

Bacterial-cellulose (BC) aerogels were used as fiber reinforcement with DMA as monomer

and PEGDA as crosslinker in the preparation of BC-DMA hydrogels. As can be seen in

Figure 27 (a), SEM pictures reveal the fiber network of bacterial cellulose nanofibers.

Figure 27 (b) demonstrates a cross-section of BC aerogel and evidences the ca. 2 nm of

diameter and hundreds of micrometers in length of an individual nanofiber.


50

Figure 27. FE-SEM pictures of the freeze-dried BC microstructures (a) surface and (b)

cross section.

DMA hydrogel were also prepared as reference via FRP in aqueous solution under Fushion

UV cure system for 10 minutes. The hydrogels reached a water swelling equilibrium after

one week of swelling and seven solvent exchanges to remove any unreacted residues. The

FT-Raman spectra of cured DMA hydrogels are shown in Figure 28 (a). The C=C

vibrations at 1615-1661 cm-1 from unreacted DMA monomer or PEGDA crosslinker were

not detected in the dried hydrogels, supporting a high reaction conversion i.e. below the

detection limit using FT-Raman. For the BC-DMA hydrogels, there is a small trace of the

cellulose peaks from BC at 1157 - 1095 cm-1 in the FT-Raman spectra due to the small

fraction of BC present in the sample.


51

Figure 28, (a) The FT-Raman spectra of DMA and PEGDA, dry DMA hydrogel P-1, P-2,

P-3 and BC, dry BC-DMA hydrogel F-P-1, F-P-2 and F-P-3. (b) Photo of (I) swollen and

(II) dried BC-DMA hydrogel F-P-1, F-P-2 and F-P-3.

The gel fraction, compression modulus and other properties are presented in Table 9. With

the same crosslinker molar ratio (4 mol%) and the DMA monomer concentration

increasing from 10% (P-1) to 70% (P-3), the water content decreased from 92% to 77%,


52

which indicates that P-3 has a higher crosslinking density than P-1. However, with the

DMA monomer concentration increasing from 10% to 40% but crosslinker molar ratio

decreasing from 4 mol% (P-1) to 2 mol% (P-2), the water content still decreased from 92%

to 85%, which also gave evidence of the higher crosslinking density. These could due to

the network formation of FRP is random, the initial monomer concentration exhibits more

dominant effect of crosslinking density compared to crosslinker concentration.84 The

higher initial DMA monomer concentrations showed the higher crosslinking density of the

cured DMA hydrogels.85,86

Table 9. Sample code and properties of DMA hydrogels and BC-reinforced DMA

hydrogels: gel fraction (Gf), concentration (wt%) of DMA monomer, crosslinker

(PEGDA) molar ratio (mol%) to DMA, BC fiber content in DMA monomer and

crosslinker aqueous solution (BCas), DMA network content, BC fiber content in swollen

BC-DMA hydrogel (BCBC-DMA), equilibrium water content, and compression modulus (Ec).

Sample

code

DMA

(wt%)

PEGDA

(mol%)

BCas

(wt%)

Gf

(%)

DMA-

network

content

(wt%)

BCBC-DMA

(wt%)

Water

content

(%)

Ec

(MPa)

P-1 10 4 0 94 8 - 92 1.0

P-2 40 2 0 93 15 - 85 1.0

P-3 70 4 0 - 23 - 77 2.9

F-P-1 10 4 1 94 13 8 78 4.4

F-P-2 40 2 1 94 23 5 75 5.3

F-P-3 70 4 1 95 30 3 70 8.3

As presented in Table 9, although the initial BC fiber content is the same for all the

samples (1%), the cured and swollen BC-DMA hydrogel fiber content is decreased from

8% to 3% with the DMA network content increased from 13% to 30%. Due to the limited

expansion of swollen BC network, the higher crosslinking density and wt% of DMA

network in BC-DMA hydrogel, the water content of BC-DMA hydrogels have been


53

suppressed ca. 10% compared with pure DMA hydrogels (92% to 77%) with the same

polymer-crosslinking density.

In Table 9, while the initial DMA wt% increased from 10 to 70 wt%, the compression

modulus (Ec) of DMA hydrogels increased from 1 to 2.9 MPa without BC. The Ec of BC-

DMA increased from 4.4 to 8.3 MPa with the DMA wt% increased from 10 to 70 wt% (4

mol% PEGDA). Ec of BC-DMA hydrogel increased ca. 3 times compared with the DMA

hydrogel with the same crosslinking density. However, increasing DMA from 10 to 40

wt% and decreasing PEGDA from 4 to 2 mol%, the BC-DMA exhibits an increase Ec from

4.4 to 5.3 MPa. Further presents the increasing initial DMA wt% will increase the

crosslinking density even with less crosslinker mol%.

DMA hydrogels also contributed with improved shear modulus and changed the water

absorption of BC hydrogels. Figure 29, (I) depicts the shear storage modulus G′ of BC-

DMA hydrogels and BC hydrogel. Shear modulus of BC without polymer network is close

to 0 kPa but after curing the DMA network in the BC network, the G′ of the resulting IPN

hydrogel P-F-3 increased up to ca. 350 kPa. The combination of DMA and BC fibers

locked water in the voids of the BC aerogel, as shown in Figure 29 (II) a, with a finger

force twisting the re-swollen BC hydrogel, water can be squeezed out. Water in swollen

BC-DMA hydrogels (Figure 30 (II) b) on the other hand, can only be removed by

evaporation and not by physical force.


54

Figure 29. I (a): the photo of BC hydrogel and (b): BC-DMA hydrogel under a finger

shear force. II: the shear storage modulus G′ (kPa) of re-swollen BC hydrogel, BC-DMA

hydrogels F-P-1, F-P-2 and F-P- 3. (ω: 0.1 to 20 Hz, γ: 1%)

The microstructures of BC and DMA interpenetrating networks were observed by FE-

SEM. As can be seen in Figure 30, after DMA network cured in the BC aerogels, the

composites of BC fibers and DMA networks show no trace of individual BC fiber compare

to raw BC aerogel, Figure 27. The FE-SEM pictures reveal that there is no trace of phase

separation between fiber and polymer networks of the BC-DMA hydrogels surface, Figure

30 (a), (b), (c), and cross section, Figure 30 (a’), (b’), (c’); also, the pore size of the

hydrogels decreased from the loose crosslinked BC-DMA hydrogel F-P-1 (10 wt% DMA

and 4 mol% PEGDA) to densely crosslinked BC-DMA hydrogel F-P-3 (70 wt% DMA and

4 mol% PEGDA).


55

Fig

ure

30.

FE

-SE

M pic

ture

s of

BC

-DM

A h

ydro

gel

su

rfac

e o

f B

C-D

MA

hy

dro

gel

F

-P-1

(1

0 w

t% D

MA

and 4 m

ol%

PE

GD

A),

F-P

-2 (

40 w

t% D

MA

and 2

mol%

PE

GD

A),

an

d F

-P-3

(7

0 w

t% D

MA

an

d 4

mol%

PE

GD

A):

a,

b,

c an

d c

ross

sect

ion a

’, b

’ an

d c

’.

Conclusions

56

5 CONCLUSIONS

The purpose of this work was to relate and evaluate hydrogel fabrication parameters with

swelling and mechanical properties.

PEG single and sequential interpenetrating polymer network (seqIPN) hydrogels were

successfully prepared and characterized in this work via UV-initiated thiol-ene coupling

(TEC) chemistry.

A library of PEG hydrogels was prepared using PEG-SH or the reversed PEG-Al

precursors in combination with trifunctional crosslinkers. The UV-initiated TEC reaction

proved to be robust for the formation of PEG hydrogels under benign experimental

conditions. The type of solvent, solid content in solution, functional group and PEG chain

length were found to have direct influence on mechanical properties of PEG hydrogels.

Fabricated hydrogels based on PEG-Al displayed stronger mechanical properties than

PEG-SH hydrogels.

Well-defined PEG seqIPN with controllable mechanical properties have also been

prepared and assessed. Properties of a library of seqIPN hydrogel were compiled based on

the information of secondary-network content, gel fraction, swelling kinetic, mechanical

properties and structure evaluations. The seqIPN hydrogels based on the PEG-Al systems

showed 84% and higher gel fraction. The mechanical properties of PEG hydrogel were

increased to reach a tensile modulus over 1 MPa (2-2:44). The range of tensile modulus is

extended from single network, range from 175 kPa to 555 kPa, from seqIPN 365 kPa to

1135 KPa. Water content of PEG hydrogels was supressed by seqIPN from single network,

range 90% to 79% (degree of swelling 870% to 380%), to seqIPN 88% to 74% (degree of

swelling 580% to 280%).

Conclusions

57

The diffusion rate of the disperse red 13 decorated linear PEG with short chain (2 kDa) and

long chain (8 kDa) was evaluated in both a dense (2 kDa) and loose (8 kDa) PEG

crosslinked network.

Finally, BC aerogels were employed as reinforced templates to yield BC-DMA hydrogels

via traditional UV-initiated polymerization. The compression modulus of DMA hydrogel

ranging from 1 MPa - 2.9 MPa increased to 4.4 MPa - 8.3 MPa upon introducing BC

fibers. The water content was supressed by the BC network from DMA hydrogel from

92% to 77% in the pure DMA hydrogels to 78% to 70% in the BC-DMA hydrogels.

Future work

58

6 FUTURE WORK

IPN is an effective way to manipulate mechanical properties of PEG hydrogels. Triple and

multiple networks of PEG IPN will be the next interesting hydrogel to prepare by TEC.

Combining polyDMA with PEG TEC hydrogels could achieve even higher mechanical

properties.

Copper-catalyzed azide–alkyne cycloaddition (CuAAC) could be another interesting

process for preparing hydrogels. FRP with CuAAC in one system could prepare IPN in

one pot.

In order to further explore the potential of TEC chemistry for well-defined hydrogel

preparation, it will be interested to compare dendronized PEG with allyl groups with linear

PEG-Al. The mechanical properties will be expected to improve a lot because of the much

higher crosslinking density.

Dendronized PEG with thiol groups could crosslink without using crosslinker or

dissolution in a solvent.

Nano-febrile cellulose (NFC) could be used to prepare fiber-reinforced hydrogels. The

NFC could be TEMPO-oxidized and surface-treated with polymer to improve the

compatibility with monomers. NFC solid concentrations can be tuned to fit the needs of

the hydrogel applications.

Acknowledgements

59

7 ACKNOWLEDGEMENTS

I would like to thank my supervisors: Prof. Anders Hult is thanked for giving me the

chance to be a PhD student at KTH. Prof. Lars Berglund is thanked for being my co-

supervisor in the first 2 years. Associate prof. Michael Malkoch is thanked for teaching me

from laboratory safety to writing paper and helping me to become a PhD.

All seniors in the department of fiber and polymer technology are thanked for their help.

Thanks to members of biocomposite group. Prof. E. Kristofer Gamstedt and associate prof.

Qi Zhou are thanked for their helps.

The China Scholarship Council (CSC) and the Department of Fiber and Polymer

Technology at KTH are acknowledged for financial support.

Prof. Ling Wang from Chengdu University of Technology is thanked for supporting my

application of CSC scholarship during my master study.

Thanks to members of the coating group: special thanks to Marie, Yvonne for checking my

writings and giving a lot of help when I just arrived in the group. Hui is thanked for

working together with me for my papers and being a nice friend. Kim is thanked for a lot

Acknowledgements

60

of help in the lab. Jan is really thanked for supporting me with the materials to work for the

paper. Oliver, Jonas and Kristina are thanked for helping me with the Swedish abstract

writing. Mats K.G.J is thanked for checking my last paper and being ‘tomten’. Linda is

thanked for being so nice and reviewing my thesis. Camilla is thanked for her nice talks

and warm care. Alireza, Assya and Susanne are thanked for being good office-roommates.

Stacy is thanked for her happy personality. Andreas Fall is thanked for helping me with the

rheometer. Inger is greatly thanked for her help in the department. Big thanks to Eva,

Anna, Emma L, Emma Ö, Bella, Hanna, Sara, Linn, Christian, Carl, Marcus, Martin,

Mauro, Pontus, Carmen, Eric and Petra.

Special thanks to my uncle Jiebing Li and Aunt Yun Yang in Sweden for helping me in

every aspect of my life. Yitong is thanked for her relax outlook on life. Jesper is thanked

for all the fun art things and taping for my niece. Jingyu is thanked for her nice entertain

in Japan and help me a lot in the wedding. Big thanks for Andong Liu, Aihua Pei, Yafang

Yan, Yingzhi Zhu, Wenbin Yu, Xueyu Du, Hongli Zhu, Bi Ran, Lingquan Deng, Yujia

Zhang, Jiayi Yang, Lin Li, Bo Yin, Liming, Zhiqiang Zheng, Chun Ying Shean, Andrew

M., Raquel, Lage, Dany, Pedro and other friends in Sweden, I thank all of you for the

happy moments.

感谢父亲杨竟达和母亲黄果对我的养育之恩。感谢爷爷杨金南对我的栽培和奶奶姜

映南在生活上给我的照顾。感谢外公黄岳飞，外婆王文飞，小姨黄朵，堂哥杨泰宏、

杨经宇，表姐梁晓莉，表哥黄毅对我的关心。

Till familjen, Anders Nilsson (Fluffy) tackas för att vara en omtänksam make; till Rositha,

Leif, Bosse, Elvy, Birgit, Kerstin, Lasse, Ammi, Lisbeth, Michael, Jenny, Stefan, tackas!

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61

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