Preparation of a High-Strength Hydrogel with Slidable and Tunable Potential Functionalization Sites

Article abstract of DOI:10.1021/acs.macromol.5b02359

A hydrogel with tunable potential functionalization sites has been successfully prepared. As potential functionalization sites, (2-hydroxypropyl)-α-CDs (Hy-α-CDs) were introduced into the network of tetrahedron-like poly(ethylene glycol) (tetra-PEG) gel t

Full text of DOI:10.1021/acs.macromol.5b02359

Article
pubs.acs.org/Macromolecules
Preparation of a High-Strength Hydrogel with Slidable and Tunable
Potential Functionalization Sites
Zhao Li, Zhen Zheng, Shan Su, Lin Yu, and Xinling Wang*
School of Chemistry and Chemical Engineering, State Key Laboratory of Metal Matrix Composites, Shanghai Jiao Tong University,
800 Dongchuan Road, Shanghai 200240, China
S
* Supporting Information
ABSTRACT: A hydrogel with tunable potential functionaliza-
tion sites has been successfully prepared. As potential
functionalization sites, (2-hydroxypropyl)-α-CDs (Hy-α-
CDs) were introduced into the network of tetrahedron-like
poly(ethylene glycol) (tetra-PEG) gel through supramolecular
chemistry. In the stage of complexation, poly-pseudo-rotaxane
consisting of tetra-PEG macromonomer and Hy-α-CD formed
in pregel solution. The dynamic complexation process and the
structure of the poly-pseudo-rotaxane were investigated by
NMR experiment. In the stage of gelation, some cross-linking reactions were completed through click chemistry. The structures
and mechanical properties of the resultant hydrogels were characterized by ATR-FTIR, XPS, SEM, and compression test in
detail. The number of Hy-α-CD introduced into the hydrogel is related closely to the structure of the poly-pseudo-rotaxane and
can be controlled easily by tuning the feed ratio. Anthracene as an example of function was introduced into the hydrogel through
Hy-α-CD to preliminarily demonstrate the validity of the potential functionalization site in the last, and the hydrogel also has the
capacity for further diverse functionalization.
seven, or eight glucose units are α-, β-, or γ-CDs. One of the
significant features of these CDs is that they are able to be
chemically modified by a wide variety of methods,23,24 which
figures CD to be one of the essential roles in chemistry and
material science. On the other hand, CDs can form inclusion
complexes with many kinds of polymers.25 In 1990, it was
found that PEG and α-CD could form crystalline poly-pseudo-
rotaxane and separated out from water.26 The size matching of
the cross-sectional area of PEG chain and the interior cavity of
α-CD make formation of the poly-pseudo-rotaxane possible, and
the major driving forces of the complexation are hydrophobic
and van der Waals interactions between the inner surface of the
α-CD ring and the hydrophobic sites on PEG.25
α-CD seems to be the appropriate candidate for the potential
functionalization site. This is similar to slide-ring gel, the
network chains in which are interlocked by figure-of-eight
cross-links.27 The figure-of-eight structure is made up of two
linked α-CDs.28 However, the solubility of α-CD/PEG poly-
pseudo-rotaxane is limited by the formation of hydrogen
bonding through hydroxyl groups between adjacent α-CDs.
The homogeneous network structure of tetra-PEG gel would
be undoubtedly affected if crystalline poly-pseudo-rotaxane
exists. (2-Hydroxypropyl)-α-CD is a common α-CD derivative
broadly used in drug formulation due to its high water solubility
and excellent biocompatibility,29 and we found that it could
INTRODUCTION
■
Hydrogels are three-dimensional polymer networks containing
a large amount of water inside.1 They play important roles in
many areas, such as medical treatment,2,3 actuators,4
cosmetics,5 sewage disposal,6 and so on. To be applied in a
more complicated and sophisticated way, hydrogel ought to be
stronger and more functional. On the other hand, the network
structure of hydrogel should be regular and clear so that the
properties of it can be controlled easily.
Over the past decade, hydrogels with high mechanical
strength came through fast development.7 Nanocomposite gel,8
macromolecular microsphere composite gel,9 double network
gel,10 slide-ring gel,11−13 and tetrahedron-like poly(ethylene
glycol) gel (tetra-PEG gel)14 all own excellent mechanical
properties. Among them, tetra-PEG gel has high strength due
to its ideally homogeneous network structure.15−20 To expand
the application fields of the tetra-PEG gel, it is necessary to
functionalize it. However, chemical inertness of PEG backbone
limits its modification, which could only be realized through
linking functional groups to termini of tetra-PEG macro-
monomer.21 With this method, not only is the number of
functional group limited, but the mechanical properties of the
hydrogel are impacted.21 Hence, introducing enough function-
alization sites through an effective way is critical to functionalize
tetra-PEG gel.
Cyclodextrins (CDs) are toroid-shaped cyclic oligomers of
anhydroglucopyranose22 full of hydroxyl groups, with the
primary hydroxyl groups at the narrow side and the secondary
hydroxyl groups at the wide side. Those CDs consisting of six,
Received: October 30, 2015
Revised: December 19, 2015
© XXXX American Chemical Society
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and dried in a vacuum overnight. 1H NMR (DMSO-d6 with 0.03% v/v
TMS, 400 MHz): δ 7.795 (d, 8H, 2-H of phenyl), 7.494 (d, 8H, 3-H of
phenyl), 4.109 (t, 8H, Ts−O−CH2−), 3.508 (m, 1794H,
[CH2CH2O]n), 2.424 (s, 12H, CH3−phenyl−).
Preparation of Tetraazido-Terminated PEG (TAPEG). TTPEG (4.0
g, 0.19 mmol) and sodium azide (0.50 g, 7.76 mmol) were mixed in
DMF (100 mL), and the solution was stirred under argon at 80 °C for
24 h. The solid salts were removed via filtration through Celite. The
polymer was recovered via precipitation with diethyl ether and
filtration thrice. The product was dried in vacuum overnight. 1H NMR
(DMSO-d6 with 0.03% v/v TMS, 400 MHz): δ 3.506 (m, 1794H,
[CH2CH2O]n), 3.372 (t, 8H, N3−CH2−).
form poly-pseudo-rotaxane with PEG with any molar ratio
without crystalline during research. Therefore, we chose it as
the potential functionalization site.
Herein, a novel hydrogel with high strength, ideally
homogeneous network structure, and slidable potential
functionalization site, (2-hydroxypropyl)-α-CD (Hy-α-CD),
was developed. We abbreviated it to αSSS hydrogel based on
the bold words above. The hydrogel’s skeleton is based on
tetra-PEG gel. Hy-α-CD was introduced into the hydrogel
through supramolecular chemistry. Two stages including
complexation and gelation were involved in the preparation
of the hydrogel. In the stage of complexation, Hy-α-CD and
tetra-PEG macromonomers formed poly-pseudo-rotaxane in a
pregel solution. In the stage of gelation, the equilibrated pregel
solution was cross-linked in situ by copper(I)-catalyzed azide−
alkyne cycloaddition (CuAAC) click chemistry.30,31 The
number of Hy-α-CD introduced into the αSSS hydrogel is
related to the structure of the poly-pseudo-rotaxane and can be
controlled easily by tuning the feed ratio. In addition, the
existence of Hy-α-CD changes the morphology and increases
the mechanical properties of the hydrogel. Since the constitu-
tional tetra-PEG macromonomers and Hy-α-CD are biocom-
patible, the αSSS hydrogel has prospect to be used as a
biomaterial. In the last, anthracene as an example of function
unit was introduced into the αSSS hydrogel to briefly
demonstrate the validity of the potential functionalization
sites of Hy-α-CD in the hydrogel. The obtained functionalized
αSSS hydrogel (An-αSSS hydrogel) possesses fluorescence.
Preparation of Tetrapropargyl-Terminated PEG (TPPEG). BTC
(2.04 g, 6.87 mmol) was dissolved in chloroform (40 mL), and the
solution was cooled to −10 °C under argon via an additional funnel.
To this solution, propargylamine (1.10 g, 20 mmol) and TEA (4.07 g,
40.3 mmol) that were all dissolved in chloroform (40 mL) were added
dropwise in 45 min. The mixture was stirred at −10 °C for another 45
min under argon. Then the temperature was increased to 30 °C, and
the mixture was stirred for 1 h. The solution was concentrated and
washed with 0.5 M HCl aqueous solution. The organic layer was dried
with magnesium sulfate and filtered. The filtrate was pour into
THPEG’s chloroform solution [THPEG (5.0 g, 0.25 mmol) and
chloroform (100 mL)]. After stannous octanoate (0.005 g) was added,
the solution was stirred at 60 °C under argon for 5 h. The solution was
concentrated, and an excess amount of diethyl ether was added. The
precipitate was precipitated twice in diethyl ether, filtered, and dried in
1
a vacuum overnight. H NMR (DMSO-d6 with 0.03% v/v TMS, 400
MHz): δ 7.650 (t, 4H, −O−CO−NH−), 4.071 (t, 8H, −NHCOO−
CH2−), 3.682 (t, 8H, −NHCOO−CH2−CH2−), 3.508 (m, 1794H,
[CH2CH2O]n), 3.077 (s, 4H, CHC−).
Preparation of 2-Anthracenecarbamyl-Hy-α-CD (An-Hy-α-
CD). First, 2-anthraceneisocyanate (2-An-NCO) was synthesized
through isocyanation of 2-amineanthracene by the BTC method.
BTC (1.25 g, 2.12 mmol) was dissolved in DCM, and the solution was
cooled to −10 °C under argon. 2-Amineanthracene (1.22 g, 6.34
mmol) and TEA (1.25 g, 12.37 mmol) dissolved in DCM were added
dropwise to this solution. The reaction was stirred at −10 and 30 °C
for 1 h. The solution was washed with HCl aqueous solution, and the
organic layer was dried with magnesium sulfate. Evaporation of all the
DCM gave dark green solid of raw product of 2-An-NCO which was
used directly in reaction with Hy-α-CD.
EXPERIMENTAL SECTION
■
Materials. Tetrahydroxyl-terminated PEG (THPEG) (Mw
=
20 000) was obtained from Sinopeg Biotech Co., Ltd., and dried in
a vacuum with magnetic stirring at 85 °C for 2 h before use. (2-
Hydroxypropyl)-α-CD (Hy-α-CD) (Mw ∼ 1180) with substitution
degree about 3.6 was obtained from Sigma-Aldrich Corp. and dried in
a vacuum at 85 °C before use. Propargylamine, 4-(dimethylamino)-
pyridine (DMAP), p-toluenesulfonyl chloride (TsCl), sodium azide
(NaN3), triphosgene (BTC), deuterium oxide (D2O), tetramethy-
lammonium (TMA) chloride, copper sulfate pentahydrate (CuSO4·
5H2O), and sodium ascorbic acid of analytical grade were obtained
from Sigma-Aldrich Corp. and used as-received. 2-Amineanthracene
was synthesized through the reduction of 2-aminoanthraquinone
(Aladdin Industrial Co., Ltd.) according to the method in ref 32.
Dichloromethane (DCM) and chloroform were obtained from Sigma-
Aldrich Corp. and dried with 4A molecular sieve. Dimethylformamide
(DMF) and trimethylamine (TEA) were obtained from Sigma-Aldrich
Corp. and distilled with CaH2 before use.
Then 2-anthracenecarbamyl-Hy-α-CD (Scheme 3) was synthesized
through the reaction between 2-An-NCO and Hy-α-CD. The 2-An-
NCO obtained above and Hy-α-CD (2.49 g, 2.11 mmol) were mixed
in DMF. After stannous octanoate (0.01 g) was added, the solution
was stirred at 60 °C under argon for 24 h. The solution was
concentrated and precipitated in an excess amount of diethyl ether
thrice. After the filtration, the precipitate washed with diethyl ether
until the supernatant was colorless. Drying in vacuum overnight gives
1
the product of 2-anthracenecarbamyl-Hy-α-CD. H NMR (DMSO-d6
with 0.03% v/v TMS, 400 MHz): δ 9.78 (−NH−COO−), 7.10−8.60
(anthracene), 5.30−6.20 O(2,2′,2″,3,3′,3″)H of Hy-α-CD, 5.00 O(8)
H of Hy-α-CD, 4.60−5.20 C(1,1′,1″)H of Hy-α-CD, 4.53 O(6)H of
Hy-α-CD, 3.10−4.20 C(2,2′,2″,3,3′,3″,4,4′,4″,5,5′,5″,6,6′,6″,7,8)H of
Hy-α-CD, 1.03 C(9)H of Hy-α-CD.
Preparation of Terminal Modified Tetra-PEG Macromono-
mers. Tetra-toluenesulfonate-terminated PEG and tetra-azido-termi-
nated PEG were prepared according to the method in refs 33 and 34.
Tetra-propargyl-terminated PEG was prepared through the efficient
reaction between −NCO on propargyl isocyanate and −OH on
THPEG (Scheme S1). The structures of the products were
determined by 1H NMR (400 MHz) spectroscopy recorded on a
Varian MERCURY plus-400 spectrometer at 25.0 0.5 °C.
Preparation of Tetratoluenesulfonate-Terminated PEG (TTPEG).
THPEG (5.0 g, 0.25 mmol), DMAP (3.66 g, 30 mmol), and TEA (7.5
mL) were dissolved in DCM (50 mL), and the solution was cooled to
0 °C under argon via an additional funnel. To this solution, TsCl (5.72
g, 30 mmol) dissolved in DCM (50 mL) was added dropwise in 15
min. The reaction was allowed to warm to room temperature
overnight with magnetic stirring under argon. The solution was
concentrated and washed with 0.5 M HCl aqueous solution. The
organic layer was dried with magnesium sulfate and filtered. An excess
amount of diethyl ether was added in filtrate to precipitate. After
filtration, the product was precipitated twice in diethyl ether, filtered,
Complexation between Hy-α-CD and Tetra-PEG Macro-
monomers in Pregel Solution. Nuclear magnetic resonance
spectroscopy was used to characterize the complexation between
1
Hy-α-CD and tetra-PEG macromonomers in a pregel solution. H
NMR (400 MHz) and 13C NMR (100 MHz) spectra were recorded
on a Varian MERCURY plus-400 spectrometer at 25.0 0.5 °C with
64 and 1024 scans, respectively, for every sample. 2D NOESY NMR
(400 MHz) spectra were obtained using a Bruker AVANCEIII 400
spectrometer with a mixing time of 300 ms at 25.0 0.5 °C. The 2048
experiments were performed with four scans per experiment. THPEG
was used as model macromonomer. Deuterium oxide was used as
solvent with chloroform sealed in glass capillary of 1.0 mm diameter as
an external reference. Samples for NMR experiment were listed in
Table 1. 1a to 6a were used to simulate pregel solutions of the αSSS
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Table 1. Compositions of Samples for NMR Experiment
a
sample
0a
solute
THPEG
solvent
nHy‑α‑CD:nTHPEG
mD O:mTHPEG
cHy‑α‑CD (mmol/g)
2
D2O
D2O
D2O
D2O
D2O
D2O
D2O
D2O
D2O
D2O
D2O
D2O
D2O
9
9
9
9
9
9
9
1a
THPEG, Hy-α-CD
THPEG, Hy-α-CD
THPEG, Hy-α-CD
THPEG, Hy-α-CD
THPEG, Hy-α-CD
THPEG, Hy-α-CD
Hy-α-CD
5
10
0.02689
0.05213
0.1193
0.2097
0.2799
0.3362
0.02689
0.05213
0.1193
0.2097
0.2799
0.3362
2a
3a
25
4a
50
5a
75
6a
100
1a(Blank)
2a(Blank)
3a(Blank)
4a(Blank)
5a(Blank)
6a(Blank)
Hy-α-CD
Hy-α-CD
Hy-α-CD
Hy-α-CD
Hy-α-CD
a
cHy‑α‑CD = nHy‑α‑CD/(mHy‑α‑CD + mD O).
2
Table 2. Some Parameters of Hydrogel Samples 0ag to 6ag
a
sample
nTAPEG:nTPPEG
M
Ninitial
NHy‑α‑CD
SR (%)
EWC (%)
0ag
1ag
2ag
3ag
4ag
5ag
6ag
1
1
1
1
1
1
1
9
9
9
9
9
9
9
0
5
0
0
95.4 0.6
94.6 0.5
95.3 1.1
95.3 1.0
94.7 1.0
94.8 1.2
94.8 0.7
5.0 0.6
9.9 0.5
100 12.0
99 5.0
10
25
50
75
100
22.0 1.0
30.0 2.1
36.9 3.2
48.3 5.0
88.4 4.0
60.4 4.2
49.6 4.3
48.7 5.0
a
M represents the mass ratio of DI water to tetra-PEG macromonomers (TAPEG and TPPEG) (mDI water:mmacromonomer).
hydrogels and had same composition and equal feed ratios
corresponding to the hydrogel samples 1ag to 6ag in Table 2. The
used to determine the structure of the poly-pseudo-rotaxane.
Complexation between Hy-α-CD and THPEG needed time to reach
the equilibrium after which the structure of the poly-pseudo-rotaxane
was characterized.
Preparation of αSSS Hydrogel. A series of hydrogels 1ag to 6ag
were prepared (Table 2). The molar ratio of TAPEG to TPPEG
(nTAPEG:nTPPEG) of them was kept constant of 1:1, and the mass ratio
of DI water to macromonomers (TAPEG and TPPEG)
(mDI water:mmacromonomers) was kept at 9:1. The molar ratios (feed
ratios) of Hy-α-CD to macromonomers (NInitial) changed from 5 to
100. Tetra-PEG hydrogel 0ag without any Hy-α-CD was as blank
sample to compare with αSSS hydrogel samples.
To a series of small vials were added TAPEG (0.1000 g, 4.975
μmol), TPPEG (0.1011 g, 4.975 μmol), and DI water (1.7599 g).
After the macromonomers were dissolved, a different amount of Hy-α-
CD was added to form pregel solution. The solutions were incubated
at 37 °C. After the pregel solution reached the equilibrium, copper
sulfate (1.592 mg, 9.950 μmol) in DI water (48.41 mg) and sodium
ascorbate (9.8 mg, 49.5 μmol) were added. The vials were placed in an
oven present to 37 °C. The hydrogels formed within minutes and
stood for another 48 h, which were then treated with 0.5 M aqueous
ethylenediaminetetraacetic acid (EDTA) solution followed by plenty
of water to remove the copper catalysis and unthreaded Hy-α-CD. The
hydrogels were incubated in DI water to fully swell before
characterizations.
Characterizations of αSSS Hydrogel. ATR-FTIR Measurement.
Attenuated total reflectance−Fourier transform infrared (ATR-FTIR)
spectra were recorded on a Paragon 1000 instrument equipped with
ATR accessory. The scan range was 4000 to 660 cm−1, and seven scans
were collected for each sample with a resolution of 2 cm−1. The
hydrogel samples were frozen in liquid nitrogen and broken. After
freeze-drying, the sectioned surfaces were caught out ATR
examination. Peak intensities of the αSSS hydrogels 1ag to 6ag were
normalized to the CO in carbamate group of pristine tetra-PEG
hydrogel 0ag.
mass ratio of D2O to THPEG (mD O:mTHPEG) was kept at 9, whereas
2
the molar ratio of Hy-α-CD to THPEG (nHy‑α‑CD:nTHPEG) changed
from 5 to 100. Sample 0a was THPEG’s D2O solution without Hy-α-
CD, and it was the reference to calculate the chemical shift variation
(Δδ) of THPEG. Hy-α-CD could form intermolecular inclusion
complexes in aqueous solution and its complexation behavior had
concentration dependency. Thus, blank samples 1a(Blank) to
6a(Blank) of Hy-α-CD’s D2O solution without THPEG were
prepared, the concentrations of which were equal to samples 1a to
6a, respectively. They were used as references to elucidate the changes
of Hy-α-CD’s structure before and after the addition of THPEG under
different Hy-α-CD concentrations.
The chemical shifts of all protons of CDs decrease linearly with the
increase of CD concentration.35 In addition, with the existence of CDs,
internal reference compounds were reported to show significant
downfield shifts during NMR titration experiments.36 During this
research, it was found not only small molecules but polymers like
THPEG of which all protons decreased linearly with the increase of
Hy-α-CD’s concentration. Furthermore, 13C of Hy-α-CD, THPEG,
and internal references also showed downfield shifts during NMR
experiments. These phenomena may be the consequence of changes in
the magnetic susceptibilities of water due to hydrogen bonding with
the glucose units of the cyclodextrin.37 Accordingly, chloroform was
1
used as external reference to correct the data from H and 13C NMR
spectra.
Monitoring the Complexation Process between Hy-α-CD and
THPEG. 1H NMR spectrometry was used to monitor the process of the
complexation between Hy-α-CD and THPEG. During the experiment,
certain amounts of Hy-α-CD and THPEG were dissolved in D2O
according to the compositions of 1a to 6a, and these six samples were
incubated at 37 °C. 1H NMR spectra were taken every a certain period
of time until 254.0 h.
XPS Measurement. Chemical composition of the sectioned surfaces
of the hydrogel samples were analyzed using a Shimadzu-Kratos (AXIS
Characterization of the Poly-Pseudo-Rotaxane at Dynamic
Equilibrium. 1H, 13C, and 2D NOESY NMR spectrometry were
C
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Scheme 1. (a) Fabrication Process of the αSSS Hydrogel and (b) the Photograph of Hydrogel Sample 5ag
Ultra) X-ray photoelectron spectrometer (XPS) equipped with
monochromatic Al Kα X-rays. A takeoff angle of 90° was employed
for each sample. The scan area is 300 × 700 μm. At least three random
positions on the sectioned surface of every hydrogel sample were
chosen to be executed with XPS measurement. Initial representative
survey scans were acquired from 0 to 1200 eV for three sweeps (sw).
For further elemental analysis, high-resolution scans were acquired for
the C 1s (∼276 to 298 eV, 3 sw), O 1s (∼524 to 543 eV, 2 sw), and N
1s (∼392 to 412 eV, 12 sw) regions. For all spectra, the backgrounds
of all regions were subtracted using the Shirley background.38 Peak
positions were normalized to the C 1s peak at 285.0 eV. The data
analysis was carried out by using XPSPeak 4.1 software.
diameter and 5 mm in initial thickness was placed on a metal plate
coated with silicon oil to decrease the friction.39 The cross-head speeds
was 1.0 mm/min. At least five parallel samples were recorded for each
specimen the test.
Preparation of An-αSSS Hydrogel. TAPEG and TPPEG with
equal moles were dissolved in DI water to get a 10 wt % solution.
Then An-Hy-α-CD was added to the solution. After incubation at 37
°C for 120 h, copper sulfate and sodium ascorbate were added, and the
solution was incubated at 37 °C for another 48 h to obtain a hydrogel.
The hydrogel was treated with EDTA aqueous solution, DI water, and
DMF to remove the copper catalysis and unthreaded An-Hy-α-CD.
The gel was treated with DI water again to displace DMF and fully
swollen in DI water before characterization.
Characterizations of An-αSSS Hydrogel. UV−Vis DRS Meas-
urement. A PerkinElmer Lambda 750S UV/vis/NIR spectropho-
tometer equipped with a 60 mm integrating sphere was used to record
the UV−vis diffuse reflectance spectrum of An-αSSS hydrogel. The
scan range is 200−700 nm with step of 1 nm. The slit width is 2.0 nm.
Polytetrafluoroethylene (PTFE) was used as a reflection standard. A
cylinder-shaped hydrogel sample with diameter of 15 mm and height
of 3 mm was sandwiched in two quartz plates, and the “sandwich” was
placed in the sample compartment of the spectrometer.
Fluorescence Spectroscopy. Fluorescent excitation and emission
spectra were recorded at room temperature using a PerkinElmer LS 55
fluorospectrometer with a solid sample holder with emission and
excitation wavelength of 440 and 360 nm, respectively. The working
voltage was 725 V, and the slit width was 7.5 nm. A cylinder-shaped
hydrogel sample with diameter of 15 mm and height of 3 mm was
sandwiched in two quartz plates, and the “sandwich” was placed in the
sample holder.
Swelling Measurement. The hydrogel samples were freeze-dried
and then immersed in DI water for 3 days at room temperature to fully
swell. The equilibrium water contents (EWCs) of the hydrogels were
calculated by the following equation:
mwet − mdry
EWC =
× 100%
mwet
(1)
At least three parallel samples were recorded for each hydrogel
specimen.
Internal Morphology. The internal morphologies of the sectioned
surfaces of the freeze-dried hydrogel samples were observed using a
Philips Sirion 200 instrument scanning electron microscope (SEM).
Photographs were taken with a Canon IXUS 800IS digital camera. The
dried hydrogel samples were mounted on metal holder and vacuum
coated with a gold layer prior to SEM examination.
Compression Test. A MTS Criterion 43 universal texting machine
was used to measure mechanical performance of the αSSS hydrogel.
Compression mode was used. A cylindrical sample about 7 mm in
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at around 1100 cm−1. A peak at 1721 cm−1 is for the CO in
carbamate group, which was used as the internal reference.
Spectra in Figure 1 are normalized spectra. With the
introduction of Hy-α-CD, the broad peak of hydroxyl group
at around 3370 cm−1 appears. C−O−C stretching of Hy-α-CD
arises at 1030 cm−1. These results indicate that Hy-α-CDs are
confined in hydrogel’s network. It is also found that with
increasing the amount of fed Hy-α-CD, peaks of hydroxyl and
C−O−C group of Hy-α-CD strengthen gradually from 1ag to
6ag. This suggests that with the increase of fed Hy-α-CD in
pregel solution, the amount of it confined in network of the
hydrogel increases.
XPS was used to quantitatively determine the amount of Hy-
α-CD introduced into the αSSS hydrogel. Typical high-
resolution narrow scans of the C 1s regions of 0ag to 6ag
are shown in Figure 2. After spectral fitting, in 0ag spectrum,
the peak at 285.0 eV is attributed to contaminate carbon and
the peak at 286.6 eV is C(C−O/C−N) region of poly(ethylene
oxide) network. In spectra of 1ag to 6ag, new peaks at 288.1 eV
appear which are attributed to anomeric C(O−C−O) of Hy-α-
CD, and the peaks at 286.5 eV correspond to C−O/C−N of
poly(ethylene oxide) and Hy-α-CD. The peaks at 285.0 eV are
attributed to C−C/C−H of Hy-α-CD and contaminate carbon.
The number of Hy-α-CD corresponding to every tetra-PEG
macromonomer (TAPEG or TPPEG) (NHy‑α‑CD) is used to
express the amount of Hy-α-CD confined in the αSSS
hydrogel’s network. NHy‑α‑CD is defined as
RESULTS AND DISCUSSION
■
Matrix of αSSS Hydrogel. The matrix of the αSSS
hydrogel is tetra-PEG gel with regular and homogeneous
network structure which is the origin of high strength and
enables Hy-α-CDs to distribute evenly. To avert the influence
of hydroxyl group on Hy-α-CD, classical “CuAAC” click
chemistry was used as cross-linking reaction considering its
unique characteristics of quantitative yields and high tolerance
of functional groups. As for the synthesis of TAPEG, −OH
groups at termini of THPEG were first activated by the tosyl
group and then reacted with NaN3. As for the synthesis of
TPPEG, propargylamine was converted to propargyl isocyanate
and then linked to termini of THPEG (Scheme S1). It takes the
advantage of high efficiency of the reaction between −NCO
and −OH and makes the preparation of TPPEG easy. In the
meantime, CO in the carbamate group was taken as effective
internal reference in ATR-FTIR examination to normalize the
spectra.
Preparation of αSSS Hydrogel. Two symmetrical
tetrahedron-like PEG macromonomers, TAPEG and TPPEG,
derived from THPEG were dissolved in DI water together with
different amount of Hy-α-CD to form a series of pregel
solutions. After these pregel solutions reached dynamic
equilibrium (determination of the time to reach the equilibrium
is discussed in the latter part of NMR analysis), gelation was
carried out. The resultant αSSS hydrogel is a colorless hydrogel
with excellent transparency (Scheme 1b). That partly indicates
the homogeneous network of tetra-PEG gel is inherited by the
αSSS hydrogel. That also indicates introduction of Hy-α-CD
does not cause any crystalline in the hydrogel.
nHy‐α‐CD
NHy‐α‐CD
=
nmacromonomer
(2)
Quantification of the Amount of Hy-α-CD Introduced
in αSSS Hydrogel. As the potential functionalization site, Hy-
α-CD in the αSSS hydrogel will certainly act as a key role on
the functionalization of the hydrogel. Thus, at this stage
determining how many Hy-α-CDs are introduced in the αSSS
hydrogel appears particularly important. Here ATR-FTIR and
XPS spectroscopy were used to determine this parameter.
ATR-FTIR spectroscopy was used to semiquantitatively
determine the amount of Hy-α-CD introduced into the αSSS
hydrogel. Figure 1 shows the ATR-FTIR spectra of the
sectioned surfaces of freeze-dried hydrogel samples 0ag to 6ag.
In the figure, tetra-PEG hydrogel 0ag provides a baseline. C−
O−C stretching of poly(ethylene oxide) network appears peak
Here nHy‑α‑CD and nmacromonomer stand for moles of Hy-α-CD and
tetra-PEG macromonomer in the hydrogel’s network.
The αSSS hydrogel contains equal amount of TAPEG and
TPPEG macromonomers. One TAPEG has 907 C−O/C−N
type carbons. One TPPEG has 919 C−O/C−N type carbons.
The average number of C−O/C−N type carbon contained in a
tetra-PEG macromonomer is 913 (NM).
One α-CD consists of NCD = 6 glucopyranose units. In a Hy-
α-CD molecule with 2-hydroxypropyl-substituted degree of 3.6,
one glucopyranose unit contains 1 O−C−O type carbon, 6.2
C−O type carbons, and 0.6 C−C/C−H type carbons on
average. In the XPS spectrum, peak area is in proportion to
atom mole. Thus, in a αSSS hydrogel’s XPS C 1s spectrum, the
area corresponding to macromonomer’s C−O/C−N type
carbon is (A286.6 eV − A288.1 eV × 6.2), where A286.6 eV and
A
288.1 eV represent areas of peaks at 286.6 and 288.1 eV. NHy‑α‑CD
can be written as
A288.1 eV
NCD
A286.6 eV − A288.1 eV × 6.2
NM
NHy‐α‐CD
=
(3)
(4)
Putting NM = 913 and NCD = 6 into eq 3:
A288.1 eV × 152.2
NHy‐α‐CD
=
A286.6 eV − A288.1 eV × 6.2
According to eq 4, NHy‑α‑CD values of samples 1ag to 6ag were
calculated and are listed in Table 2. The threading ratio (SR) of
the αSSS hydrogel is defined as
NHy‐α‐CD
SR =
× 100%
Figure 1. ATR-FTIR spectra of the sectioned surfaces of freeze-dried
hydrogel samples 0ag to 6ag.
Ninitial
(5)
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Figure 2. Typical XPS (C 1s) spectra for the sectioned surfaces of freeze-dried hydrogel samples 0ag to 6ag.
Here Ninitial is the number of Hy-α-CD corresponding to every
macromonomer added initially in pregel solution.
From Table 2, it can be found that with the increase of the
amount of fed Hy-α-CD in pregel solution, the number of Hy-
α-CD confined in the αSSS hydrogel’s network increases, and
the threading efficiency of Hy-α-CD reduces.
Mechanism of Complexation between Hy-α-CD and
Tetra-PEG Macromonomers in Pregel Solution. Complex-
ation between Hy-α-CD and tetra-PEG macromonomers in
pregel solution is the critical step for the preparation of the
αSSS hydrogel. Clarifying the mechanism of the complexation
is significant for preparation of the hydrogel with tunable
amount of Hy-α-CD. On the basis of the obtained parameter of
NHy‑α‑CD above, we are able to discuss the complexation
mechanism combining it with the results of NMR experiment.
TAGEG and TPPEG are derived from THPEG, and they have
the same molecular weights as THPEG. So in the NMR
experiment, THPEG was used as model macromonomer to
complex with Hy-α-CD. NMR samples 1a to 6a have the same
feed ratios of Hy-α-CD to tetra-PEG macromonomer and
solute concentrations as the corresponding hydrogel samples
1ag to 6ag and were used to simulate the pregel solutions of
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1ag to 6ag. The parameter of NHy‑α‑CD thus can be regarded as
the amount of Hy-α-CD threaded on THPEG at dynamic
equilibrium.
Although many reports on the preparation, characterization,
and application of polymer/CD based poly-pseudo-rotaxanes
and polyrotaxanes have been published, the threading
mechanism and dynamics of a linear polymer in CDs have
yet to be revealed.40 However, this issue is essential and
significant for dynamics of polymers in solution.41,42 On the
other hand, physical and chemical characters as well as
behaviors in aqueous solution of Hy-α-CD are different from
that of α-CD due to the substitution. And there are few
researches about the complexation between polymers and
substituted CD. Thus, clarification of complexation mechanism
between Hy-α-CD and tetra-PEG is valuable to improve the
theory of supramolecules.
1
Result of 2D NOESY H NMR Measurement. 2D NOESY
1H NMR spectroscopy is a straightforward method to
determine the existence of host−guest interaction of organic
compounds. It establishes correlation between nuclei of proton
through nuclear Overhauser enhancement (NOE) effect. Hy-α-
CD and THPEG were dissolved together in D2O with different
molar ratios (Table 1), and the solutions were incubated at 37
°C. After these NMR samples reached the dynamic equilibrium
(a detailed description of the time to reach the dynamic
equilibrium and the complexation process between Hy-α-CD
and THPEG is being involved in the following part), 2D
NOESY NMR spectra of them were taken.
1
Figure 3 shows a typical 2D NOESY H NMR spectrum of
the Hy-α-CD/THPEG system. In the 1D 1H NMR spectrum in
the figure, the signal around 3.73 ppm is assigned to the
methylene proton of THPEG’s monomeric CH2CH2O unit,
and the signals around 4.00, 3.82, and 1.19 ppm are assigned to
the protons of C(3)H, C(5)H, and C(9)H of Hy-α-CD,
respectively (Scheme 2). As can be seen in the figure, signals of
C(3)H and C(5)H of Hy-α-CD correlate with the resonance of
the methylene proton of THPEG, which indicates that the
host−guest interaction exists between Hy-α-CD and THPEG
and the chains of THPEG are included in Hy-α-CDs’ cavities.
This complexation is driven by hydrophobic and van der Waals
interactions between the inner surface of the Hy-α-CD ring and
the hydrophobic sites on the chains of THPEG. Moreover, it is
found that signal of C(9)H of Hy-α-CD correlates with the
signal of methylene proton of THPEG. That indicates when
Hy-α-CD threads on THPEG’s chain, 2-hydroxypropyl groups
of Hy-α-CD change their conformations to approach THPEG’s
chain and parallel with it at the equilibrium. In Figure 3b, it can
be seen that signal of C(9)H also correlates with signals of
C(3)H and C(5)H. When the feed ratio of Hy-α-CD to
THPEG is greater than 10, the threading ratio (SR) of Hy-α-
CD is less than 100% (Table 2). That means not all the fed Hy-
α-CDs can thread on THPEG at the equilibrium, and there are
unthreaded Hy-α-CDs in solution. These “free” Hy-α-CDs
interact with each other to form intermolecular inclusion
complexes in which 2-hydroxypropyl groups of one Hy-α-CD
can insert into cavities of other Hy-α-CDs. Methyl parts on the
2-hydroxypropyl groups have hydrophobic interaction with C3
and C5 methines in the interior cavity of CD, which leads to
the cross-peaks between C(9)H and C(3,5)H. This behavior of
Hy-α-CD is also being demonstrated with the results of 13C
NMR experiment and discussed in the following part.
Figure 3. Partial 2D NOESY 1H NMR spectra of sample 6a at
dynamic equilibrium.
of THPEG’s monomeric CH2CH2O unit exhibits signal around
3.7 ppm (see Supporting Information). Figure 4 shows the
chemical shift variations of the methylene protons (ΔδTHPEG
)
as functions of time and molar ratio of Hy-α-CD to THPEG
(nHy‑α‑CD:nTHPEG). The positive variation indicates an increase
in chemical shift δ, viz., a shift toward downfield, and vice versa.
When THPEG was added to Hy-α-CD’s aqueous solution,
1
ΔδTHPEG values exhibited positive; that is, the H resonance of
the methylene proton shifted downfield. This indicates that Hy-
α-CD threads on THPEG’s chain and the exchange rate of the
1
complex are faster than H NMR time scale. Besides, the main
driving force of the complexation is the hydrophobic
interaction between the inner surface of Hy-α-CD ring and
the hydrophobic site on CH2CH2O unit. This hydrophobic
interaction causes deshielding effect on the methylene proton
of the CH2CH2O unit and leads to signal of the methylene
proton shifting downfield.
From curves in Figure 4a, it can be inferred that the
methylene proton of THPEG for all samples shifts first upfield
and then downfield and finally reaches equilibrium. This
suggests that there is an assembly process between Hy-α-CD
and THPEG. And it is time-consuming, which may be caused
by the rearrangement process of threaded Hy-α-CDs on
THPEG’s chains. Hy-α-CDs thread onto THPEG’s chains in
despite of their positions and conformations as soon as THPEG
is added. This process is fast and leads to large downfield shift
at the beginning. Some Hy-α-CDs have improper state which is
not energy minimum. Therefore, they dethread from THPEG’s
chains, adjust their conformations and positions, and then
rethread. This process is relatively slow and time-consuming. It
Complexation Process between Hy-α-CD and THPEG. In
the 1H NMR spectra of samples 1a to 6a, the methylene proton
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Scheme 2. Process of Complexation between THPEG and Hy-α-CD in Aqueous Solution and the Equilibrium State of the Poly-
Pseudo-Rotaxane
Figure 4. Chemical shift variations of THPEG’s monomeric unit methylene protons (ΔδTHPEG) as functions of (a) incubation time of samples 1a−
6a and (b) molar ratio of Hy-α-CD to THPEG (nHy‑α‑CD:nTHPEG) with different incubation times.
is also found that different samples spend different time to
reach the equilibrium. With increasing the value of
nHy‑α‑CD:nTHPEG, the time spent turns longer. This phenomenon
can be easily understood that more Hy-α-CDs need more time
to adjust their positions and conformations.
off the chains of THPEG freely and quickly with very few
hampers. The exchange rate of THPEG between free and
associated forms is high. Thus, the signal of methylene proton
of THPEG locates relatively downfield. The methylene proton
of THPEG of sample 2a locates more downfield than that of
1a. This is caused by the more Hy-α-CDs held by the poly-
pseudo-rotaxane of 2a than that of 1a at the equilibrium. With
With the increase of the feed ratio of Hy-α-CD to THPEG
(nHy‑α‑CD:nTHPEG) from samples 1a to 6a, the value of NHy‑α‑CD
increases monotonically in Table 2. However, the trend of the
value of ΔδTHPEG from 1a to 6a is not consistent with that of
NHy‑α‑CD. Curves in Figure 4b show a strange pattern. In spite of
differences on time, all the curves show an M-shaped trend.
When THPEG is added to Hy-α-CD’s aqueous solution,
they form poly-pseudo-rotaxane. Hy-α-CD can thread on and
off of THPEG’s chain dynamically, and this exchange is fast on
n
Hy‑α‑CD:nTHPEG increasing to 25, the average number of Hy-α-
CD threaded on every arm of THPEG increases to 5.5
thereupon. More Hy-α-CDs bring the threading/dethreading
process some difficulties because of crowdedness, which slows
the exchange of the poly-pseudo-rotaxane, and this is the main
factor comparing with the increase of threaded Hy-α-CDs to
determine the trend of the curve of ΔδTHPEG − nHy‑α‑CD:nTHPEG
.
1
a H NMR time scale. When the value of nHy‑α‑CD:nTHPEG is
So the methylene proton of THPEG shifts upfield. When
nHy‑α‑CD:nTHPEG increases further, free THPEG disappears.
THPEG’s chains are covered by some Hy-α-CDs all along,
which shortens the length of effective chains. Hy-α-CDs outside
the chains can thread on and off, and this exchange turns fast
again. Therefore, the methylene proton shifts downfield again.
When nHy‑α‑CD:nTHPEG continuously increases from 5a to 6a, the
main factor determine of trend of ΔδTHPEG turns to the reduced
small, all the fed Hy-α-CDs can thread on THPEG’s chains.
With the increase of nHy‑α‑CD:nTHPEG, the threading ratio (SR)
of Hy-α-CD decreases. When the value of nHy‑α‑CD:nTHPEG is
small, the average number of Hy-α-CD corresponding to every
arm of THPEG is small. For nHy‑α‑CD:nTHPEG is 5, the average
number is 1.25. For nHy‑α‑CD:nTHPEG is 10, and the average
number is 2.5. In these circumstances, Hy-α-CDs thread on and
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Figure 5. (a) Partial 13C NMR spectra of samples 0a to 6a at dynamic equilibrium. (b) Chemical shift variations of 13C of THPEG monomeric unit
methylene [Δδ(13C)THPEG] as a functions of nHy‑α‑CD:nTHPEG at the equilibrium.
Figure 6. (a) Chemical shifts of carbons C3 [δC3(Blank)] and C5 [δC5(Blank)] of Hy-α-CD without THPEG as a function of mass concentration of
Hy-α-CD (cHy‑α‑CD). (b) Chemical shifts of carbons C3 (δC3) and C5 (δC5) of Hy-α-CD with THPEG as a function of cHy‑α‑CD at dynamic
equilibrium.
exchange rate of the poly-pseudo-rotaxane again, so the value of
ΔδTHPEG decreases again.
and the peaks also broaden with the increase of nHy‑α‑CD:nTHPEG.
And the dependence of Δδ(13C)THPEG on nHy‑α‑CD:nTHPEG in
Figure 5b is similar to curves in Figure 4b.
In short, though the trend of the value of ΔδTHPEG is
inconsistent with that of NHy‑α‑CD with the increase of the feed
ratio of Hy-α-CD to THPEG, this does not mean there is a
mistake of calculated value of NHy‑α‑CD. There are two factors
determining the trend of ΔδTHPEG, the amount of threaded Hy-
α-CD, and the exchange rate of the poly-pseudo-rotaxane.
These two factors have opposite effects on the trend of the
curve of ΔδTHPEG − nHy‑α‑CD:nTHPEG. The M-shaped curve
pattern of ΔδTHPEG − nHy‑α‑CD:nTHPEG is the result of synergistic
effect of the two factors.
The Hy-α-CD used in this work has a substituted degree of
3.6, that is, 3.6 2-hydroxypropyl groups substitute randomly on
C6, C3, or C2 positions on α-CD (Scheme 2). Unlike α-CD,
hydrogen bonding within and between Hy-α-CDs is weakened
due to the substitution. Therefore, in aqueous solution, Hy-α-
CD has unique molecular conformations and behaviors with or
without THPEG.
In 13C NMR spectra, CD backbone carbons C3 and C5 of
Hy-α-CD exhibit signals at around 73.2 and 71.8 ppm (Figure
S4a,c), and methyl carbon C9 of the 2-hydroxypropyl group of
Hy-α-CD exhibits a complex pattern at around 18.1 ppm
(Figure S4b,d). Before the study of the poly-pseudo-rotaxane,
behaviors of Hy-α-CDs in aqueous solution should be revealed
because they can form intermolecular inclusion complexes with
each other.
Structure of Hy-α-CD/THPEG-Based Poly-Pseudo-Rotax-
1
ane at Dynamic Equilibrium. In H NMR spectra of samples
0a to 6a at the dynamic equilibrium (Figure S1g,h), peaks of
the methylene proton of THPEG broaden with the increase of
nHy‑α‑CD:nTHPEG, which indicates the chains of THPEG are
covered by Hy-α-CD and the amount of Hy-α-CD threaded
increases with the rise of nHy‑α‑CD:nTHPEG. The broadness is
related to the threading dynamics of PEO chain into the cavity
of Hy-α-CD. When THPEG is threaded by Hy-α-CDs, the
movements of all the molecules are restricted, and the flexibility
of the PEO chains of THPEG reduces. In 13C NMR spectra of
0a to 6a at the equilibrium (Figure 5a), the monomeric
methylene carbon of THPEG exhibits signal around 69.4 ppm,
Figure 6a shows chemical shifts of carbons C3 [δC3(Blank)]
and C5 [δC5(Blank)] of Hy-α-CD as a function of Hy-α-CD
mass concentration (cHy‑α‑CD) of blank references 1a(Blank) to
6a(Blank). With the increase of cHy‑α‑CD, δC3(Blank) and
δC5(Blank) shift downfield gradually, and the curve of
δC3(Blank) − cHy‑α‑CD is steeper than the curve of δC5(Blank)
− cHy‑α‑CD. These results indicate that Hy-α-CDs form inclusion
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Figure 7. Chemical shifts of carbons C3 (a), C5 (b), and C9 (c) of Hy-α-CD with and without THPEG as functions of mass concentration of Hy-α-
CD (cHy‑α‑CD).
complexes. In addition, δC3(Blank) and δC5(Blank) show
concentration dependency, so the chemical shift variations
cannot be attributed to the formation of intramolecular
complex.41 The methyl part on the 2-hydroxypropyl group of
one Hy-α-CD can insert into the cavity of another Hy-α-CD
shallowly from both wide and narrow rims driven by
hydrophobic interaction. Owing to sizes’ difference, the wide
rim of Hy-α-CD is able to accommodate more than one methyl
group while the narrow rim does only one. Carbon C3 which is
closer to the wide rim interacts with more methyl groups than
C5 does, causing larger gradient of the curve. δC9(Blank) shows
similar concentration dependency which also demonstrates the
existence of the intermolecular inclusion complexes (Figure 7c,
black line).
narrow rim of Hy-α-CD and methyl. So on addition of
THPEG, change of guest group from methyl of 2-
hydroxypropyl to ethylene oxide unit of THPEG leads to
signal of carbon C5 shifting upfield. For the methine of C3, the
methyl of 2-hydroxypropyl group complex with it loosely in the
inclusion complex formed by Hy-α-CDs because the size of
wide rim of Hy-α-CD is slightly larger for the methyl group. On
addition of THPEG, Hy-α-CD threads on the PEO chain and
complexes with it tightly due to the formation of the
pseudorotaxane mechanical bond. So the interaction between
the methine of C3 and ethylene oxide of THPEG is stronger
compared with that of methine of C3 and the methyl group in
the inclusion complex. This results in signal of C3 shifting
downfield after the addition of THPEG. Curves of δC3
−
When THPEG is added, for samples 1a to 6a, in Figure 7,
δC3 and δC9 shift downfield, whereas δC5 shifts upfield for all
cHy‑α‑CD. In aqueous solution without THPEG, Hy-α-CDs form
inclusion complexes. When THPEG is added, the inclusion
complexes dissociate, and the free Hy-α-CDs complex with
THPEG to form poly-pseudo-rotaxane. In this process, both
methines of C3 and C5 experience the transition that the group
they interact with changes from methyl (−CH3) of 2-
hydroxypropyl to ethylene oxide unit (CH2CH2O) of
THPEG. In the cavity of Hy-α-CD, methine of C3 is closer
to the wide rim, whereas methine of C5 is closer to the narrow
rim. For the methine of C5, the interaction between it and the
methyl group is stronger than that between it and the ethylene
oxide group due to the better size matchment between the
cHy‑α‑CD and δC5 − cHy‑α‑CD show M-shaped trend, which is
similar to curves of ΔδTHPEG − nHy‑α‑CD:nTHPEG. Moreover, the
slope of the δC5−cHy‑α‑CD curve surpasses that of the δC3−
c
Hy‑α‑CD curve (Figure 6b). These results indicate that Hy-α-CD
threads on THPEG’s chains.
From the results of ATR-FTIR and XPS above, it reaches a
conclusion that Hy-α-CD is confined in the network of the
αSSS hydrogel and the confined Hy-α-CD cannot be washed
away by water. This is most likely caused by the formation of
poly-pseudo-rotaxane between Hy-α-CD and tetra-PEG macro-
monomers in pregel solution, and the threaded Hy-α-CD is
confined in the network of the hydrogel after gelation of the
1
poly-pseudo-rotaxane. And the results of 2D NOESY, H and
13C NMR confirmed this speculation. Based on the ATR, XPS,
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and NMR results and the discussion above, the clarified
mechanism of the complexation between THPEG and Hy-α-
CD in aqueous solution is depicted in Scheme 2. Before
THPEG is added, Hy-α-CDs form intermolecular inclusion
complexes. The methyl part on 2-hydroxypropyl group of one
Hy-α-CD inserts shallowly in the cavity of another Hy-α-CD
from wide or narrow rims. The wide rim can accommodate
more than one methyl group, whereas the narrow rim can hold
only one. Hy-α-CDs thread onto THPEG’s chains as soon as
THPEG is added irrespective of their positions and
conformations. After that, some Hy-α-CDs dethread from
THPEG’s chains, regulate their conformations and positions,
and rethread. This process is time-consuming. The time needed
to reach dynamic equilibrium increases with the increase of the
feed ratio (nHy‑α‑CD:nTHPEG). At the equilibrium, THPEG is
covered by Hy-α-CDs 2-hydroxypropyl groups of which are
parallel with THPEG’s chain and have interaction with it. The
amount of Hy-α-CD threaded on THPEG at the equilibrium
increases with the increase of nHy‑α‑CD:nTHPEG, though the
threading ratio of Hy-α-CD decreases with the increase of the
feed ratio. When the amount of fed Hy-α-CD is small, all of
them can thread onto THPEG’s chains. This is the result of
strong interaction between Hy-α-CD and poly(ethylene oxide)
chain. When more Hy-α-CDs are added, not all of them can
thread onto THPEG’s chains, which is the consequences of
crowdedness effect and dynamic equilibrium between threading
and dethreading Hy-α-CD. Some Hy-α-CDs unthread all along.
On the basis of extensive investigation of the mechanism of
complexation between Hy-α-CD and tetra-PEG macromono-
mer, we are able to understand the relationship between the
structure of Hy-α-CD/tetra-PEG macromonomer based poly-
pseudo-rotaxane and the composition of αSSS hydrogel. And
the amount of Hy-α-CD introduced into the αSSS hydrogel can
be controlled easily by tuning the feed ratio.
Figure 9. Molar ratio of Hy-α-CD to THPEG (NInitial) dependence of
breaking stress (red square) and breaking strain (blue triangle).
represent αSSS hydrogels 1ag, 2ag, 3ag, 4ag, 5ag, and 6ag. The
breaking stresses of the αSSS hydrogels are in the same order of
magnitude as that of the tetra-PEG gel, which also partly
indicates the homogeneous network of tetra-PEG gel is
inherited by the αSSS hydrogel, for the ideal network structure
is the origin of the high mechanical strength of the tetra-PEG
gel. We can find that the breaking stresses of the αSSS
hydrogels 1ag to 4ag are slightly lower than that of 0ag. But
when more Hy-α-CDs are added, the strength of the αSSS
hydrogel surpasses that of tetra-PEG gel and even up to 3.67
MPa. The existence of large amount of Hy-α-CD increases the
stiffness of the network chains in the hydrogel, which leads to
the high strength. Both the pristine tetra-PEG gel and the αSSS
hydrogel have high breaking strain over 90%, which indicates
these hydrogels of excellent flexibilities.
Sakai and co-workers reported in 2010 a tetra-PEG gel with
compression breaking strength of 27 MPa.43 Compression
properties of tetra-PEG hydrogel 0ag and αSSS hydrogels 1ag
to 6ag in present study are not as good as the tetra-PEG gel of
Sakai’s. This may be the results of different cross-linking
reactions and preparing conditions of the hydrogels and
different samples’ states in compression test. The tetra-PEG
gel reported by Sakai et al. was prepared through termini cross-
coupling of amine and NHS-glutarate groups, whereas the
tetra-PEG and αSSS hydrogel here were prepared through
“CuAAC” click gelation reaction. The cross-linking media of
Sakai’s tetra-PEG gel is mixture of phosphate and phosphate−
citric acid buffers, whereas the cross-linking media of tetra-PEG
and αSSS hydrogels here is DI water. The last but perhaps most
important factor is that the hydrogel specimen being executed
with compression test was used in as-prepared state in the work
reported by Sakai and co-workers, whereas here the tetra-PEG
and αSSS hydrogels were fully swollen before the compression
test. So the compression strengths of hydrogels reported here
are different from that of tetra-PEG gel reported by Sakai et al.
Introduction of a Function into αSSS Hydrogel. In the
last part, a function was introduced into the αSSS hydrogel to
preliminarily study the validity of the potential functionalization
sites of Hy-α-CD. Anthracene was chosen as an example of the
function. It was chemically linked to Hy-α-CD, and the
resultant An-Hy-α-CD (Scheme 3) was complexed with tetra-
PEG macromonomers TAPEG and TPPEG. The resulted poly-
pseudo-rotaxane was then cross-linked through “CuAAC” click
chemistry to obtain the An-αSSS hydrogel. The hydrogel was
treated with EDTA-2Na aqueous solution to remove the
catalyst and then repeated washed with water and DMF until
Internal Morphology of the αSSS Hydrogel. Figure 8
shows typical SEM images of sectioned surfaces of pristine
Figure 8. SEM images of the sectioned surfaces of freeze-dried
hydrogels (a) 0ag and (b) 5ag.
tetra-PEG gel 0ag and αSSS hydrogel 5ag. 0ag has
homogeneous network with thick and loose texture, whereas,
5ag’s network appears slim and fine texture. Poly(ethylene
oxide) has strong tendency to get together and crystallize. In
freeze-dried tetra-PEG hydrogel’s network, chains of poly-
(ethylene oxide) gather to form thick texture. When Hy-α-CDs
are introduced, they cover poly(ethylene oxide) chain of the
network, hinder the approach among the chains, and weaken
the crystallization. Moreover, the rigidity of the chain rises due
to the coverage of Hy-α-CDs.
Mechanical Properties of the αSSS Hydrogel. Com-
pression test was performed to investigate mechanical proper-
ties of the αSSS hydrogel.
In Figure 9, the points at NInitial = 0 represent pristine tetra-
PEG gel 0ag; the points at Ninitial = 5, 10, 25, 50, 75, and 100
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Scheme 3. Molecular Structure of An-Hy-α-CD
Figure 10. Photographs of αSSS hydrogel and An-αSSS hydrogel under (a) daylight and (b) 365 nm UV illumination. (c) UV−vis DRS spectrum of
An-αSSS hydrogel. (d) Fluorescent excitation spectrum of An-αSSS hydrogel. (e) Fluorescent emission spectrum of An-αSSS hydrogel.
no unthreaded An-Hy-α-CD residue was remained. The
hydrogel was fully swollen in DI water before characterizations.
Figure 10a,b shows photographs of αSSS hydrogel and An-
αSSS hydrogel under daylight and 365 nm UV illumination.
Under daylight An-αSSS hydrogel presents brown color
compared with the colorless αSSS hydrogel. Under the UV
illumination An-αSSS hydrogel emits blue fluorescent light. In
the UV−vis DRS spectrum of An-αSSS hydrogel (Figure 10c),
indicate that the functional unit of anthracene was successfully
introduced into the αSSS hydrogel through functionalization
site of Hy-α-CD and the obtained An-αSSS hydrogel possesses
fluorescent character. Extensive investigation of preparation,
characterization, and functional feature of the An-αSSS
hydrogel system is in progress in our laboratory.
Functionalizing the αSSS hydrogel with anthracene unit is an
attempt to utilize the potential functionalization site of Hy-α-
CD in the hydrogel. Further study of functionalization of the
αSSS hydrogel with other diverse functions is under way.
1
the low-energy band of the Bb transition of anthracene unit
appears around 240 nm, and the 1La band appears between 260
and 360 nm.44 Figure 10d shows excitation spectrum of An-
αSSS hydrogel. From the figure, a excitation wavelength of 360
nm was chosen to further record the emission spectrum of An-
αSSS hydrogel in Figure 10e which is in agreement with the
emission curve of anthracene unit.45 The results above all
CONCLUSION
■
A novel tetra-PEG/Hy-α-CD hydrogel with a homogeneous
network and tunable potential functionalization sites has been
successfully prepared. Tetra-PEG gel as the skeleton endows
L
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(7) Shibayama, M. Soft Matter 2012, 8, 8030−8038.
the hydrogel with an ideal network structure and good
mechanical properties. Evenly distributed Hy-α-CDs as the
functionalization sites thread on poly(ethylene oxide) chain and
can shuttle between two cross-linking points. Formation of
poly-pseudo-rotaxane with Hy-α-CD and tetra-PEG macro-
monomers is the critical stage for the preparation of the
hydrogel, for the amount of Hy-α-CD introduced is in relation
with the structure of the poly-pseudo-rotaxane. The amount of
Hy-α-CD introduced into the hydrogel can be controlled easily
by tuning the feed ratio. The got αSSS hydrogels composed by
biocompatible tetra-PEG macromonomers and Hy-α-CD have
higher mechanical strengths. So it is believed that the hydrogel
has great potential in biomedical applications.
Besides, anthracene as an example of function was
successfully introduced into the αSSS hydrogel through Hy-
α-CD, which preliminarily verifies the feasibility of using Hy-α-
CD as the functionalization site. The constructed An-αSSS
hydrogel has character of fluorescence.
Thus, we think that a feasible method has been found to
prepare a new hydrogel having both better mechanical
properties and diverse functionalities. The αSSS hydrogel
could be a platform to be modified easily and turned to
functional materials due to the tunable functionalization sites of
Hy-α-CDs. And also the resultant slidable functions may bring
the hydrogel new properties. Further work is in progress.
(8) Haraguchi, K.; Takehisa, T. Adv. Mater. 2002, 14, 1120−1124.
(9) Huang, T.; Xu, H. G.; Jiao, K. X.; Zhu, L. P.; Brown, H. R.; Wang,
H. L. Adv. Mater. 2007, 19, 1622−1626.
(10) Gong, J. P.; Katsuyama, Y.; Kurokawa, T.; Osada, Y. Adv. Mater.
2003, 15, 1155−1158.
(11) Okumura, Y.; Ito, K. Adv. Mater. 2001, 13, 485−487.
(12) Bin Imran, A.; Esaki, K.; Gotoh, H.; Seki, T.; Ito, K.; Sakai, Y.;
Takeoka, Y. Nat. Commun. 2014, 5, 5124.
(13) Mayumi, K.; Tezuka, M.; Bando, A.; Ito, K. Soft Matter 2012, 8,
8179−8183.
(14) Sakai, T.; Matsunaga, T.; Yamamoto, Y.; Ito, C.; Yoshida, R.;
Suzuki, S.; Sasaki, N.; Shibayama, M.; Chung, U. I. Macromolecules
2008, 41, 5379−5384.
(15) Matsunaga, T.; Sakai, T.; Akagi, Y.; Chung, U. I.; Shibayama, M.
Macromolecules 2009, 42, 1344−1351.
(16) Matsunaga, T.; Sakai, T.; Akagi, Y.; Chung, U. I.; Shibayama, M.
Macromolecules 2009, 42, 6245−6252.
(17) Nishi, K.; Fujii, K.; Katsumoto, Y.; Sakai, T.; Shibayama, M.
Macromolecules 2014, 47, 3274−3281.
(18) Akagi, Y.; Gong, J. P.; Chung, U. I.; Sakai, T. Macromolecules
2013, 46, 1035−1040.
(19) Shibayama, M. J. Phys. Soc. Jpn. 2009, 78, 041008−1−7.
(20) Nomoto, Y.; Matsunaga, T.; Sakai, T.; Tosaka, M.; Shibayama,
M. Polymer 2011, 52, 4123−4128.
(21) Jia, H.; Li, Z.; Wang, X.; Zheng, Z. J. Mater. Chem. A 2015, 3,
1158−1163.
(22) Wenz, G.; Han, B. H.; Muller, A. Chem. Rev. 2006, 106, 782−
817.
ASSOCIATED CONTENT
* Supporting Information
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.macro-
mol.5b02359.
■
(23) Wenz, G. Angew. Chem., Int. Ed. Engl. 1994, 33, 803−822.
(24) Easton, C. J.; Lincoln, S. F. Modified Cyclodextrins, Scaffolds and
Templates for Supramolecular Chemistry; Imperial College Press:
London, 1999; p 72.
S
(25) Harada, A.; Hashidzume, A.; Yamaguchi, H.; Takashima, Y.
Chem. Rev. 2009, 109, 5974−6023.
Synthesis scheme for tetra-azido-terminated PEG
(TAPEG) and tetra-propargyl-terminated PEG
(26) Harada, A.; Kamachi, M. Macromolecules 1990, 23, 2821−2823.
(27) Murata, N.; Konda, A.; Urayama, K.; Takigawa, T.; Kidowaki,
M.; Ito, K. Macromolecules 2009, 42, 8485−8491.
(28) Noda, Y.; Hayashi, Y.; Ito, K. J. Appl. Polym. Sci. 2014, 131,
40509.
1
(TPPEG), H NMR spectra of samples 0a to 6a with
incubation time involved in Figure 1, 1H NMR spectra of
reference samples 1a(Blank) to 6a(Blank), 13C NMR
spectra of samples 0a to 6a at equilibrium, and 13C NMR
spectra of reference samples 1a(Blank) to 6a(Blank)
(PDF)
(29) Maestrelli, F.; Garcia-Fuentes, M.; Mura, P.; Alonso, M. J. Eur. J.
Pharm. Biopharm. 2006, 63, 79−86.
(30) Uliniuc, A.; Popa, M.; Hamaide, T.; Dobromir, M. Cell Chem.
Technol. 2012, 46, 1−11.
(31) Kolb, H. C.; Finn, M. G.; Sharpless, K. B. Angew. Chem., Int. Ed.
2001, 40, 2004−2021.
AUTHOR INFORMATION
Corresponding Author
*Ph +86-21-54745817; e-mail xlwang@sjtu.edu.cn (X.W.).
■
(32) de Miranda, M. G.; de Oliveira, M. F. G.; Lopes, R. S. C.; da
Silva, A. J. R.; Albert, A. L. M.; Lopes, C. C. Tetrahedron Lett. 2013, 54,
2587−2590.
Notes
(33) Polizzotti, B. D.; Fairbanks, B. D.; Anseth, K. S.
Biomacromolecules 2008, 9, 1084−1087.
The authors declare no competing financial interest.
(34) Xu, L. Q.; Yao, F.; Fu, G. D.; Kang, E. T. Biomacromolecules
2010, 11, 1810−1817.
ACKNOWLEDGMENTS
■
This work is financially supported by National Natural Science
Foundation of China (21274085) and Shanghai Leading
Academic Discipline Project (no. B202). The author is grateful
for support from Evonik Degussa (China) Co., Ltd.
(35) Alston, D. R.; Lilley, T. H.; Stoddart, J. F. J. Chem. Soc., Chem.
Commun. 1985, 1600−1602.
(36) Funasaki, N.; Nomura, M.; Yamaguchi, H.; Ishikawa, S.; Neya, S.
Bull. Chem. Soc. Jpn. 2000, 73, 2727−2728.
(37) Schneider, H. J.; Hacket, F.; Rudiger, V.; Ikeda, H. Chem. Rev.
1998, 98, 1755−1785.
REFERENCES
■
(38) Ennist, J. H.; Gobrogge, E. A.; Schlick, K. H.; Walker, R. A.;
Cloninger, M. J. ACS Appl. Mater. Interfaces 2014, 6, 18087−18097.
(39) Sun, T. L.; Kurokawa, T.; Kuroda, S.; Bin Ihsan, A.; Akasaki, T.;
Sato, K.; Haque, M. A.; Nakajima, T.; Gong, J. P. Nat. Mater. 2013, 12,
932−937.
(1) Kamata, H.; Akagi, Y.; Kayasuga-Kariya, Y.; Chung, U.; Sakai, T.
Science 2014, 343, 873−875.
(2) Lin, C. C.; Metters, A. T. Adv. Drug Delivery Rev. 2006, 58, 1379−
1408.
(3) Kogan, G.; Soltes, L.; Stern, R.; Gemeiner, P. Biotechnol. Lett.
2007, 29, 17−25.
(40) Harada, A.; Takashima, Y.; Yamaguchi, H. Chem. Soc. Rev. 2009,
38, 875−882.
(41) Inoue, Y.; Miyauchi, M.; Nakajima, H.; Takashima, Y.;
Yamaguchi, H.; Harada, A. Macromolecules 2007, 40, 3256−3262.
(42) Duhamel, J. Acc. Chem. Res. 2006, 39, 953−960.
(4) Dong, L.; Agarwal, A. K.; Beebe, D. J.; Jiang, H. R. Nature 2006,
442, 551−554.
(5) Lee, E.; Kim, B. Korean J. Chem. Eng. 2011, 28, 1347−1350.
(6) Li, X. Y.; Lin, Y. Environ. Chem. 2010, 29, 305−309.
M
DOI: 10.1021/acs.macromol.5b02359
Macromolecules XXXX, XXX, XXX−XXX

Macromolecules
Article
(43) Sakai, T.; Akagi, Y.; Matsunaga, T.; Kurakazu, M.; Chung, U. I.;
Shibayama, M. Macromol. Rapid Commun. 2010, 31, 1954−1959.
(44) Yang, C.; Mori, T.; Origane, Y.; Ko, Y. H.; Selvapalam, N.; Kim,
K.; Inoue, Y. J. Am. Chem. Soc. 2008, 130, 8574−8575.
(45) Nakamura, A.; Inoue, Y. J. Am. Chem. Soc. 2003, 125, 966−972.
N
DOI: 10.1021/acs.macromol.5b02359
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