Unprecedented access to functional biodegradable polymers and coatings

Article abstract of DOI:10.1021/ma200824k

The ever-growing biomedical technology such as tissue engineering, regenerative medicine, and controlled drug release intimately relies on the development of advanced functional biomaterials. Here, we report on versatile and robust synthesis of novel vinyl sulfone (VS)-functionalized biodegradable polymers that offer unprecedented access to advanced functional biodegradable polymers and coatings through selective Michael-type conjugate reaction with thiol-containing molecules. VS-functionalized biodegradable polymers including poly(ε-caprolactone) (PCL), poly(l-lactide) (PLA), and poly(trimethylene carbonate) (PTMC) were conveniently prepared with controlled molecular weights and functionalities through ring-opening copolymerization of ε-caprolactone (ε-CL), l-lactide (LA), or trimethylene carbonate (TMC) with a new cyclic carbonate monomer, vinyl sulfone carbonate (VSC), in toluene at 110 °C using isopropanol as an initiator and stannous octoate as a catalyst. Interestingly, these VS-functionalized biodegradable polymers allowed quantitative modification, without aid of a catalyst, with various thiol-containing molecules including 2-mercaptoethanol, cystamine, cysteine, GRGDC peptide, and thiolated poly(ethylene glycol) (PEG-SH) at a ligand-SH/VS molar ratio of 2/1 in DMF at room temperature, confirming that the Michael-type conjugate addition to VS is highly selective and tolerant to most other functional groups including hydroxyl, carboxyl, and amine. Remarkably, results of contact angle measurements, X-ray photoelectron spectroscopy (XPS), and fluorescence studies showed that biodegradable coatings based on these VS-functionalized polymers allowed direct, efficient, and clean (without catalyst and byproduct) surface functionalization with thiol-containing molecules in aqueous conditions, which is unprecedented and opens a new avenue to surface functionalization of medical implants as well as cell and tissue scaffolds. The preliminary cell culture studies using MG6 cells showed that unmodified VS-functionalized PCL films, similar to tissue culture plate, could well support cell attachment and growth, indicating that VS-functionalized PCL film is nontoxic and biocompatible. The surface of VS-functionalized PCL films could be elegantly engineered with thiolated nonfouling polymers (e.g., PEG and glycol chitosan) or cell adhesive motif (GRGDC peptide) to control cell attachment and growth. We are convinced that these vinyl sulfone-functionalized biodegradable polymers have a tremendous potential in biomedical engineering.

Full text of DOI:10.1021/ma200824k

ARTICLE
pubs.acs.org/Macromolecules
Unprecedented Access to Functional Biodegradable Polymers and
Coatings
Rong Wang,† Wei Chen,† Fenghua Meng, Ru Cheng, Chao Deng, Jan Feijen, and Zhiyuan Zhong*
Biomedical Polymers Laboratory, and Jiangsu Key Laboratory of Advanced Functional Polymer Design and Application, Department of
Polymer Science and Engineering, College of Chemistry, Chemical Engineering and Materials Science, Soochow University, Suzhou,
215123, P. R. China
S Supporting Information
b
ABSTRACT:
The ever-growing biomedical technology such as tissue engineering, regenerative medicine, and controlled drug release intimately
relies on the development of advanced functional biomaterials. Here, we report on versatile and robust synthesis of novel vinyl
sulfone (VS)-functionalized biodegradable polymers that offer unprecedented access to advanced functional biodegradable
polymers and coatings through selective Michael-type conjugate reaction with thiol-containing molecules. VS-functionalized
biodegradable polymers including poly(ε-caprolactone) (PCL), poly(L-lactide) (PLA), and poly(trimethylene carbonate) (PTMC)
were conveniently prepared with controlled molecular weights and functionalities through ring-opening copolymerization of ε-
caprolactone (ε-CL), L-lactide (LA), or trimethylene carbonate (TMC) with a new cyclic carbonate monomer, vinyl sulfone
carbonate (VSC), in toluene at 110 °C using isopropanol as an initiator and stannous octoate as a catalyst. Interestingly, these VS-
functionalized biodegradable polymers allowed quantitative modification, without aid of a catalyst, with various thiol-containing
molecules including 2-mercaptoethanol, cystamine, cysteine, GRGDC peptide, and thiolated poly(ethylene glycol) (PEG-SH) at a
ligand-SH/VS molar ratio of 2/1 in DMF at room temperature, confirming that the Michael-type conjugate addition to VS is highly
selective and tolerant to most other functional groups including hydroxyl, carboxyl, and amine. Remarkably, results of contact angle
measurements, X-ray photoelectron spectroscopy (XPS), and fluorescence studies showed that biodegradable coatings based on
these VS-functionalized polymers allowed direct, efficient, and clean (without catalyst and byproduct) surface functionalization with
thiol-containing molecules in aqueous conditions, which is unprecedented and opens a new avenue to surface functionalization of
medical implants as well as cell and tissue scaffolds. The preliminary cell culture studies using MG6 cells showed that unmodified VS-
functionalized PCL films, similar to tissue culture plate, could well support cell attachment and growth, indicating that VS-
functionalized PCL film is nontoxic and biocompatible. The surface of VS-functionalized PCL films could be elegantly engineered
with thiolated nonfouling polymers (e.g., PEG and glycol chitosan) or cell adhesive motif (GRGDC peptide) to control cell
attachment and growth. We are convinced that these vinyl sulfone-functionalized biodegradable polymers have a tremendous
potential in biomedical engineering.
’ INTRODUCTION
scaffolds,8,9 drug-eluting stents,10 and nanoparticles for targeted
drug release.11,12 However, common biodegradable polymers are
often challenged by their high hydrophobicity, improper degra-
dation profile, and in particular absence of reactive centers for the
covalent immobilization of bioactive molecules such as drugs,
peptides, and proteins. This “inert” nature has largely hampered
The ever-growing biomedical technology such as tissue
engineering, regenerative medicine, and controlled drug release
intimately relies on the development of advanced functional
biomaterials.1À3 Aliphatic polyesters and polycarbonates due to
their unique biocompatibility, biodegradability, and approved
use in biomedical devices by the US food and drug administra-
tion (FDA) are the prime synthetic biomaterials.4 For example,
they have been applied for absorbable orthopedic devices,5
microparticles for controlled protein release,6,7 cell and tissue
Received: April 8, 2011
Revised:
July 5, 2011
Published: July 14, 2011
r
2011 American Chemical Society
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Scheme 1. Synthetic Pathway for VSC Monomera
a Conditions: (i) THF, dropwise addition of 40% HBr at 0 °C, then 25 °C for 5 h; (ii) NaSH, 75 °C, 17 h, DMF; (iii) divinyl sulfone, 30 °C, methanol;
(iv) ethyl chloroformate, Et3N, 0 °C, 4 h, THF.
their design of biologically active biomaterials. In the past decade,
significant efforts have been directed to development of func-
tional aliphatic polyesters and polycarbonates,13 containing e.g.
hydroxyl,14À16 carboxyl,16À18 amine,19 allyl,20À23 alkyne/
azide,24À28 and acryloyl pendant groups.29À33 The postpolymer-
ization modification based on these functional polymers may
provide entry to a variety of sophisticated materials.34 In our
laboratory, we are interested in the development of functional
biodegradable polymers in which polymer synthesis does not
involve protection and deprotection procedures and postpoly-
merization modification proceeds quantitatively under mild
conditions without aid of any toxic catalyst and without genera-
tion of any byproduct. In this way, polymer degradation as well as
possible contamination with toxic catalysts and byproducts may
be prevented. We and Jerome’s group found that acryloyl
functional groups amenable to the Michael-type conjugate addi-
tion with thiolated molecules appears to be one of the most ideal
choices.30,35 However, acryloyl functional groups tend to under-
go radical polymerization during storage, (co)polymerization,
work-up, and modification processes. Jerome and co-workers
have reported controlled atom transfer radical polymerization of
γ-acryloyl-ε-caprolactone.29 Recently, we reported facile photo-
cross-linking of biodegradable micelles containing poly(acryloyl
carbonate) block.36 We hypothesized that vinyl sulfone (VS)
functional group with a significantly higher reactivity and selec-
tivity toward the Michael-type conjugate addition with thiolated
molecules,37 much lower reactivity toward radical polymeriza-
tion, and highly hydrophilic nature (divinyl sulfone has a water
solubility g100 mg/mL at 17 °C) might overcome problems
associated with acryloyl functional groups and more importantly
may enable direct modification of the corresponding polymer
films in water. It has been a long challenge to develop a
straightforward and efficient modification strategy for biomater-
ials and coatings that preferably takes place in aqueous conditions
without the help of a catalyst.
and compositions can be readily prepared without protection
and deprotection steps; (ii) postpolymerization modification
with thiol-containing molecules proceeds in a quantitative manner
under extremely mild conditions (in the absence of catalyst),
minimizing possible degradation and contamination; (iii) the
Michael-type conjugate addition of VS groups with thiol-contain-
ing molecules is highly selective and tolerant to a variety of
functional groups including hydroxyl, carboxyl, and amine,
enabling facile conjugation of different types of biologically active
molecules;37 and (iv) most spectacularly, biodegradable coatings
based on these VS-functionalized polymers allow for the fist time
direct, efficient, and clean (without catalyst and byproduct)
surface functionalization in aqueous conditions, which opens a
new avenue to surface functionalization of medical implants as
well as cell and tissue scaffolds.
’ EXPERIMENTAL SECTION
Materials. 3-Methyl-3-oxetanemethanol (97%, Alfa), hydrobromic
acid (40%, SCRC), sodium hydrosulfide hydrate (68%, Acros), divinyl
sulfone (95%, Dalian Guanghui, China), triethylamine (Et3N, 99%, Alfa
Aesar), stannous octoate (Sn(Oct)2, 95%, Sigma), 2-mercaptoethanol
(>99%, Amresco), 2-mercaptoethylamine hydrochloride (99%, Alfa
Aesar), L-cysteine (>99%, Alfa Aesar), and fluorescein isothiocyanate
(FITC, 98%, Sigma) were used as received. GRGDC was purchased
from Suzhou China Tech Peptide Co., Ltd. Thiolated glycol chitosan
(GC-SH, Mn = 80 000, DS = 6.85) and thiolated PEG (PEG-SH, Mn =
5000) were synthesized according to previous reports.38,39 Ethyl chlor-
oformate (>96%, SCRC) was distilled prior to use. Isopropanol and
ε-caprolactone (ε-CL, 99%, Alfa Aesar) were dried over CaH2 and
distilled before use. Toluene was dried by refluxing over sodium wire
under an argon atmosphere prior to distillation. L-Lactide (L-LA, >99%,
Purac) and trimethylene carbonate (TMC, Jinan Daigang Co. Ltd.,
China) were recrystallized from dry toluene.
Synthesis of Vinyl Sulfone Carbonate (VSC) Monomer.
VSC was synthesized in four steps (Scheme S1). First, HBr (40%,
40 mL) was dropwise added to a solution of 3-methyl-3-oxetanemetha-
nol (10.2 g, 0.10 mol) in THF (100 mL) under stirring at 0 °C. The
reaction mixture was warmed to 25 °C and stirred for 5 h. The reaction
mixture was then diluted with H2O (150 mL) and extracted with diethyl
ether (4 Â 150 mL). The organic phase was dried over anhydrous
MgSO4 and concentrated to give the desired product (bromo-diol) as a
white solid (16.38 g, 90%). 1H NMR (400 MHz, CDCl3): δ 3.68 (s, 4H,
In this paper, we report on versatile and robust synthesis of
VS-functionalized biodegradable polymers that offer unprece-
dented access to advanced functional biomaterials and coatings
through selective Michael-type conjugate reaction with thiol-
containing molecules (e.g., 2-mercaptoethanol, cystamine,
cysteine, GRGDC, PEG-SH, etc.). This novel functionalization
approach presents several remarkable features: (i) VS-function-
alized biodegradable polymers with vastly different structures
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ÀC(CH2OH)2), 3.55 (s, 2H, ÀCH2Br), 2.13 (s, 2H, À(OH)2), 0.93 (s,
3H, ÀCH3).
(s, C(CH2)2O2À, PVSC), 3.22 (m, SO2CH2À, PVSC), 2.86 (m, ÀCH2SÀ,
PVSC), 2.75 (s, ÀSCH2C, PVSC), 2.04(s, ÀCH2CH2CH2À, PTMC), 1.22
(d, (CH3)2CÀ), 1.22 (d, (CH3)2CÀ), 1.03 (s, ÀCH3, PVSC).
To a solution of bromo-diol (16.38 g, 0.09 mol) in DMF (150 mL)
under stirring was added NaSH (22.23 g, 0.27 mol). The reaction
mixture was stirred at 75 °C for 17 h, cooled to 25 °C, diluted with DI
water (1.0 L), and extracted with EtOAc (3 Â 250 mL). The organic
phase was dried over anhydrous MgSO4 and concentrated. Residual
DMF was removed by distillation under reduced pressure to yield
Postpolymerization Modification of VS-Functionalized
Biodegradable Polymers. The postpolymerization modification
of VS-functionalized polymers was carried out using Michael-type
conjugate addition reaction in DMF at room temperature under a
nitrogen atmosphere. Various thiol-containing molecules (R-SH) in-
cluding 2-mercaptoethanol, 2-mercaptoethylamine hydrochloride,
L-cysteine, PEG-SH, GC-SH, or GRGDC were employed. The SH/VS
molar ratio was set at 2/1, and the reaction proceeded for 1 day. The
modified polymers were isolated by precipitation from cold diethyl
ether/ethanol (1/4, v/v) and dried in vacuo at room temperature.
The 1H NMR spectra of thus modified P(CL-co-VSC)8.7% are
given in Figure 3. It could be concluded that the modification was
quantitative.
1
mercapto-diol as a yellowish oil (5.26 g, 43%). H NMR (400 MHz,
CDCl3): δ 3.64 (s, 4H, ÀC(CH2OH)2), 2.67 (d, 2H, ÀCH2SH), 2.27
(s, 2H, À(OH)2), 1.31 (t, 1H, ÀSH), 0.85 (s, 3H, ÀCH3).
To a solution of divinyl sulfone (10 mL, 99.6 mmol) in MeOH
(350 mL) under stirring was dropwise added mercapto-diol (5.26 g, 38.7
mmol) at room temperature. The reaction mixture was warmed to 30 °C
and stirred overnight in the dark. The solution was concentrated under
reduced pressure, and the residue was purified by column chromatog-
raphy (eluent: ethyl acetate/petroleum ether = 4/1, v/v) to yield vinyl
sulfone-diol (5.89 g, 60%). 1H NMR (400 MHz, CDCl3): δ 6.23À6.64
(m, 3H, ÀCHdCH2), 3.64 (s, 4H, ÀC(CH2OH)2), 3.27 (m, 2H,
ÀSCH2CH2À), 2.95 (m, 2H, ÀSCH2CH2À), 2.75 (s, 2H, ÀCCH2SÀ),
2.49 (s, 2H, À(OH)2), 0.85 (s, 3H, ÀCH3).
Preparation of VS-Functionalized Biodegradable Films
and Direct Modification with Thiol-Containing Molecules.
Biodegradable films were prepared on the microscope slides using 0.2 wt
% solution of VS-functionalized copolymers in chloroform by dip-
coating. The films, after thoroughly dried, were immersed in the
phosphate buffered aqueous solution of a thiol-containing molecule
(such as 2-mercaptoethanol, 2-mercaptoethylamine hydrochloride, L-
cysteine, PEG-SH, GC-SH, and GRGDC) at a concentration of 1 mg/mL
for 24 h. The resulting modified films were thoroughly rinsed with
deionized water and dried over phosphorus pentoxide under reduced
pressure. The contact angles of both modified and unmodified films
were determined on an SL-200C optical contact angle meter (Solon
Information Technology Co.) using the sessile drop method. For XPS
analysis, films were prepared on silicon wafers (0.076 Ω/0).
To a solution of vinyl sulfone-diol (5.89 g, 23.2 mmol) and ethyl
chloroformate (4.6 mL, 48.7 mmol) in dry THF (200 mL) at 0 °C under
stirring was dropwise added a solution of Et3N (7 mL, 51.1 mmol) in
THF (5 mL). The reaction was allowed to proceed for 5 h at 0 °C. The
reaction mixture was filtered, and the filtrate was concentrated under
reduced pressure. The crude product was recrystallized from THF to
1
yield vinyl sulfone carbonate monomer (4.61 g, 71%). H NMR (400
MHz, CDCl3): δ 6.23À6.64 (m, 3H, ÀCHdCH2), 4.17À4.26 (s, 4H,
ÀC(CH2)2CÀ), 3.27 (m, 2H, ÀSCH2CH2À), 2.95 (m, 2H,
ÀSCH2CH2À), 2.75 (s, 2H, ÀCCH2S-), 1.10 (s, 3H, ÀCH3). 13C
NMR (400 MHz, CDCl3): δ 145.21, 133.12, 129.27, 72.40, 51.67, 34.65,
30.17, 23.58, 15.75. Anal. Calcd for C10H16O5S2: C, 42.84; H, 5.75; S,
22.87. Found: C, 43.24; H, 5.70; S, 22.37. TOF-MS (m/z): calcd for
C10H16O5S2 280.0439; found 280.0231.
1
Characterization. H NMR spectra were recorded on the Unity
Inova 400 operating at 400 MHz. CDCl3 and DMSO-d6 were used as
solvents, and the chemical shifts were calibrated against residual solvent
signals. The molecular weight and polydispersity of the copolymers were
determined by a Waters 1515 gel permeation chromatograph (GPC)
instrument equipped with two linear PLgel columns (500 Å and Mixed-C)
following a guard column and a differential refractive-index detector.
The measurements were performed using THF as the eluent at a flow
rate of 1.0 mL/min at 30 °C and a series of narrow polystyrene standards
for the calibration of the columns. The static water contact angle
measurements were performed on an SL-200C optical contact angle
meter (Solon Information Technology Co.) using the sessile drop
method.
X-ray photoelectron spectroscopy (XPS) measurements were carried
out on a Kratos AXIS UltraDLD instrument equipped with an evapora-
tion chamber (base pressure <5 Â 10À10 Torr) and an analysis chamber
(3 Â 10À10 Torr). XPS analysis was undertaken under high vacuum on
films prepared on silicon wafers (0.076 Ω/0). The samples were
irradiated with monochromatic Al KR (hν = 1486.6 eV, spot size 400
μm  700 μm) and a takeoff angle of 45° with respect to the sample
surface. All spectra were measured at room temperature and calibrated
by setting the C 1s (CÀC) peak at 284.5 eV.
Synthesis of VS-Functionalized Polyesters and Polycarbo-
nates. The copolymerizations of VSC with ε-CL, L-LA, and TMC were
carried out in toluene at 110 °C for 1 day using isopropanol as an
initiator and Sn(Oct)2 as a catalyst. The following is a typical example on
synthesis of P(CL-co-VSC)4.2% copolymer. In a glovebox under a
nitrogen atmosphere, to a solution of ε-CL (1.160 g, 10.18 mmol)
and VSC (0.15 g, 0.54 mmol) in toluene (11 mL) under stirring was
quickly added the stock solutions of isopropanol (4 mg, 0.06 mmol) and
Sn(Oct)2 (22 mg, 0.05 mmol) in toluene. The reaction vessel was sealed
and placed in an oil-bath thermostated at 110 °C. After 24 h polymer-
ization, the reaction was terminated by two drops of acetic acid. A sample
was taken for the determination of monomer conversion using 1H NMR.
The resulting P(CL-co-VSC) copolymer was isolated by precipitation in
1
ethanol, filtration, and drying in vacuo. H NMR (400 MHz, CDCl3,
Figure 2A): δ 6.23À6.64 (m, ÀCHdCH2, PVSC), 5.01 (m,
(CH3)2CHÀ), 4.05À4.13 (s, ÀCH2OÀ, PCL; C(CH2)2O2À, PVSC),
3.22 (m, SO2CH2À, PVSC), 2.86 (m, ÀCH2SÀ, PVSC), 2.75 (s,
ÀSCH2C, PVSC), 2.30 (t, ÀCOCH2À, PCL), 1.64 (m,
ÀCH2CH2CH2À, PCL), 1.37 (m, ÀCH2CH2CH2À, PCL), 1.22 (d,
(CH3)2CÀ), 1.03 (s, ÀCH3, PVSC).
Fluorescence Observation on Cystamine-Functionalized
Films Treated with FITC. To confirm the immobilization of cysta-
mine to VS-functionalized degradable polymer films and to test the
chemical reactivity of the amine groups at the surface, cystamine-
functionalized PCL film was further treated with FITC in phosphate
buffered saline (PBS, 20 mM, pH 9.0) and then visualized with
fluorescence microscopy. Briefly, VS-functionalized degradable polymer
films following treatment with cystamine as above-described were
immersed in 0.5 mg/mL FITC solution in phosphate buffered saline
(PBS, 20 mM, pH 9.0, 4 mL) at 37 °C for 24 h in the dark. The films were
thoroughly rinsed with deionized water and then visualized using
fluorescence microscope (Leica DM4000M).
P(LA-co-VSC) and P(TMC-co-VSC) copolymers were synthesized
in a similar manner. 1H NMR spectra as well as signal assignments of the
copolymers are shown in Figure 2B,C. 1H NMR (400 MHz, CDCl3) of
P(LA-co-VSC): δ 6.23À6.64 (m, ÀCHdCH2, PVSC), 5.16 (m,
CH3CHÀ, PLA), 4.05 (s, C(CH2)2O2À, PVSC), 3.22 (m, SO2CH2À,
PVSC), 2.86 (m, ÀCH2SÀ, PVSC), 2.75 (s, ÀSCH2C, PVSC), 1.58 (m,
1
ÀCHCH3, PLA), 1.22 (d, (CH3)2CÀ), 1.03 (s, ÀCH3, PVSC). H
NMR (400 MHz, CDCl3) of P(TMC-co-VSC): δ 6.23À6.64
(m, ÀCHdCH2, PVSC), 4.25 (s, ÀCH2CH2CH2À, PTMC), 4.05
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Cell Culture on Surface Engineered Biodegradable Poly-
mer Films. The influence of surface chemistry on cell adhesion and
growth was studied using P(CL-co-VSC)8.7% films. The films were
prepared and modified with varying thiol-containing molecules includ-
ing PEG-SH, GC-SH, and GRGDC as above-described. The parent
unmodified and modified films were placed into a 6-well tissue culture
plate and sterilized by radiation prior to use. MG63 osteoblasts were
cultured on the films using Dulbecco’s Modified Eagle Medium
(DMEM), containing 10% FBS at a density of 1 Â 104 cells/well in a
humidified 5% CO2 atmosphere at 37 °C. The culture media was set as
4 mL per well and replaced each day. After 1, 4, and 6 days culture, the
cells were observed on an inverted fluorescence microscope (Axiovert
40 CFL microscope equipped with an AxioCam MR3 camera cable).
mercapto-diol; (iii) the Michael-type conjugate addition be-
tween mercapto-diol and excess vinyl sulfone gave rise to vinyl
sulfone-diol; and (iv) similar to preparation of acid-labile cyclic
carbonate monomer,40 cyclization of vinyl sulfone-diol was
performed in the presence of ethyl chloroformate in dilute
anhydrous THF solution at 0 °C via dropwise addition of
triethylamine (Scheme 1). The resulting VSC monomer was
purified by recrystallization from dry THF. 1H NMR spectrum
showed clearly signals at δ 6.23À6.64 attributable to VS protons,
δ 4.17À4.26 to methylene protons next to the carbonate, δ 3.27
to methylene protons neighboring to the sulfone group, δ 2.95
and 2.75 to methylene protons next to the thiol ether, and δ 1.10
to methyl protons (Figure 1). The integral ratio of resonances at
δ 6.23À6.64 (VS protons) and δ 4.17À4.26 (methylene protons
next to the carbonate) was close to the theoretical value (3:4),
which combined with results of mass and elemental analyses fully
confirmed successful synthesis of VSC monomer.
’ RESULTS AND DISCUSSION
Synthesis of Vinyl Sulfone Cyclic Carbonate Monomer. To
obtain VS-functionalized biodegradable polymers, here we de-
signed and prepared a new cyclic carbonate monomer, vinyl
sulfone carbonate (VSC), in four straight steps: (i) 3-methyl-3-
oxetanemethanol was treated with HBr to afford bromo-diol; (ii)
bromo-diol was reacted with sodium hydrosulfide to yield
Synthesis of Vinyl Sulfone-Functionalized Biodegradable
Polymers. VSC was readily copolymerized with ε-caprolactone
(ε-CL) in toluene at 110 °C using isopropanol as an initiator and
stannous octoate as a catalyst to afford VS-functionalized poly(ε-
caprolactone) (PCL) in good yields (78.6À95.0%) (Scheme 2a).
1H NMR showed resonances of VS protons at δ 6.23À6.64
(Figure 2A), indicating that VS group was intact during copo-
lymerization and subsequent work-up procedures. Notably,
copolymer compositions were nicely controlled by the VSC/
ε-CL molar feed ratio, in which P(CL-co-VSC) copolymers with
VSC contents (FVSC) ranging from 4.1 to 34.5 mol % were
obtained at VSC monomer in the feed (fVSC) of 5 to 40 mol %,
respectively (Table 1). The number-average molecular weights
(Mn) estimated from 1H NMR end-group analysis were in good
agreement with the theoretical data. The Mn values determined
by gel permeation chromatography (GPC) measurements using
polystyrene as standards were shown to increase in proportion to
the monomer-to-initiator ratios (Table 1, entries 1À3). These
P(CL-co-VSC) copolymers had moderate polydispersities (PDI)
of 1.34À1.81 (Table 1).
In the same way, we synthesized VS-functionalized poly-
(L-lactide) (PLA) (Scheme 2b) and VS-functionalized
Figure 1. 1H NMR spectrum (400 MHz, CDCl3) of VSC.
Scheme 2. Synthesis of VS-Functionalized Biodegradable Polymers Using Isopropanol as an Initiator and Stannous Octoate as a
Catalyst in Toluene at 110 °C: (a) VS-Functionalized Poly(ε-caprolactone), (b) VS-Functionalized Poly(L-lactide), and (c) VS-
Functionalized Poly(trimethylene carbonate)
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Table 1. Synthesis of VS-Functionalized Biodegradable PCL
through Ring-Opening Copolymerization of ε-CL and VSCa
Mn  10À3
c
d
fVSC
FVSC
1H
PDI
yield
(%)
entry M/Ib
(%)
(%) theory NMRe GPCf GPCf
1
2
3
4
5
6
100/1
160/1
320/1
150/1
150/1
120/1
5
5
4.8 12.2
4.2 20.9
4.1 39.1
8.7 19.6
16.3 22.0
34.5 21.6
11.3
19.8
36.4
18.1
19.4
19.5
17.7
26.4
46.7
21.0
21.3
24.0
1.78
1.34
1.66
1.50
1.62
1.80
84.8
91.5
95.0
80.0
81.4
78.6
5
10
20
40
aThe copolymerization was carried out in toluene at 110 °C using
isopropanol as an initiator and Sn(Oct)2 as the catalyst for 1 day. b Total
monomer-to-initiator molar ratio. c Molar fraction of VSC monomer in
the feed. d Molar fraction of VSC units in the resulting copolymer
e
determined by 1H NMR. Estimated by 1H NMR end-group analysis.
f Determined by GPC (eluent: THF, flow rate: 1.0 mL/min, standards:
polystyrene).
calibration in our GPC measurements. Therefore, through
copolymerization with VSC monomer, a range of VS-function-
alized biodegradable polymers can be conveniently prepared
with controlled functionalities.
Modification of VS-Functionalized Biodegradable Poly-
mers or Coatings. The VS groups in the biodegradable poly-
mers allowed facile modification, without aid of a catalyst, with
various thiol-containing molecules including 2-mercaptoethanol,
cystamine, cysteine, GRGDC, and PEG-SH at a ligand-SH/VS
molar ratio of 2/1 under mild conditions (Scheme 3). 1H NMR
analysis displayed complete vanishing of signals assignable to the
VS protons and appearance of new peaks due to the conjugated
ligands with corresponding intensities (Figure 3), indicat-
ing quantitative conjugation. These results confirm that the
Michael-type conjugate addition to VS is highly selective and
tolerant to most other functional groups including hydroxyl,
carboxyl, and amine.
In the next, we tried to prepare functional biodegradable
coatings by directly treating VS-functionalized PCL films with
thiol-containing water-soluble molecules under aqueous condi-
tions. Remarkably, results of the contact angle measurements
demonstrated that VS-functionalized PCL films following treat-
ment with 2-mercaptoethanol, cystamine, cysteine, thiolated
glycol chitosan, GRGDC, or thiolated PEG all became signifi-
cantly more hydrophilic, in which significant reduction of contact
angles by 13°À24° was observed depending on the nature of
thiol-containing molecules (Figure 4). In contrast, no change of
contact angles was detected for the control films of acryloyl-
functionalized PCL under otherwise the same conditions (data
not shown). These results strongly suggest that VS-functional-
ized biodegradable coatings are amenable to facile derivatization
with thiol-containing ligands.
To confirm the covalent immobilization of thiol-containing
molecules, VS-functionalized PCL films fabricated on a support-
ing silicon substrate before and after treatment with cystamine
were analyzed with X-ray photoelectron spectroscopy (XPS).
The XPS spectrum of VS-functionalized PCL films after treating
with cystamine displayed besides prominent carbon (around
284.5 eV) and oxygen signals (around 532.4 eV) also two sets of
peaks at around 399.5, 401.5 eV and around 163.7, 164.4, 167.5
eV due to NÀ(C,H) nitrogen unique to the cystamine moieties
Figure 2. 1H NMR spectra (400 MHz, CDCl3) of VS-functionalized
biodegradable polymers: (A) P(CL-co-VSC), (B) P(LA-co-VSC), and
(C) P(TMC-co-VSC).
poly(trimethylene carbonate) (PTMC) (Scheme 2c). 1H NMR
spectra corroborated successful incorporation of VSC moieties
into PLA and PTMC (Figure 2B,C). The VSC contents in P(LA-
co-VSC) copolymers were determined to be about 3.5 and 7.4
mol % at fVSC of 5 and 10 mol %, respectively (Table 2, entries
1À3). Very similarly, for P(TMC-co-VSC) copolymer, a VSC
content of 3.5 mol % was obtained at fVSC of 5 mol % (Table 2,
entry 4). The Mn data determined by GPC were in general higher
1
than those determined by H NMR (Table 2), which is most
likely due to use of polystyrene standards for molecular weight
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Table 2. Synthesis of VS-Functionalized PLA and PTMC through Ring-Opening Copolymerization of LA or TMC with VSCa
Mn  10À3
entry
copolymer
M/Ib
fVSCc (%)
FVSCd (%)
theory
1H NMRe
GPCf
PDI GPCf
yield (%)
1
2
3
4
P(LA-co-VSC)
P(LA-co-VSC)
P(LA-co-VSC)
P(TMC-co-VSC)
140/1
300/1
130/1
180/1
5
5
3.5
3.4
7.4
3.5
21.1
45.2
20.5
19.9
20.2
39.8
18.5
15.1
58.3
86.2
40.1
39.7
1.67
1.56
1.71
1.87
91.3
70.6
88.1
79.0
10
5
aThe copolymerization was carried out in toluene at 110 °C using isopropanol as an initiator and Sn(Oct)2 as the catalyst for 1 day. bTotal monomer-to-
initiator molar ratio. cMolar fraction of VSC monomer in the feed. dMolar fraction of VSC units in the resulting copolymer determined by 1H NMR.
eEstimated by 1H NMR end-group analysis. fDetermined by GPC (eluent: THF, flow rate: 1.0 mL/min, standards: polystyrene).
Scheme 3. Postpolymerization Modifications of P(CL-co-
VSC)8.7% Copolymer by Michael-Type Addition with
Varying Thiol-Containing Molecules (Ligand-SH/VS = 2/1
mol/mol, DMF, rt, 1 day)a
a Functionality was defined as mole percentage of functional groups
relative to CL and VSC units in total.
and SÀ(C,O) sulfur atom, respectively (Supporting Informa-
tion, Figure S1A). In comparison, nitrogen atom was not
detected for the untreated VS-functionalized PCL control
(Supporting Information, Figure S1B). It is evident, therefore,
Figure 3. 1H NMR spectra (400 MHz) of P(CL-co-AC)8.7% copoly-
mer after modification with thiol-containing molecules: (A) PEG-SH
(CDCl3), (B) 2-mercaptoethylamine (CDCl3), (C) 2-mercaptoethanol
(CDCl3), (D) L-cysteine (DMSO-d6), and (E) RGDC (DMSO-d6).
that cystamine has successfully immobilized to the surface.
To test the reactivity of the amine groups at the surface,
cystamine-functionalized PCL film was further treated with
fluorescein isothiocyanate (FITC) in phosphate buffered saline
(PBS, 20 mM, pH 9.0) and then visualized with fluorescence
microscopy. Interestingly, strong green fluorescence was
observed throughout the whole film (Supporting Information,
Figure S2). Notably, the fluorescence intensities were shown to
increase with increasing the initial VS contents in the P(CL-co-
VSC) copolymers (Supporting Information, Figure S2AÀC). In
contrast, no fluorescence was detected for the VS-functionalized
PCL film treated with FITC directly (Supporting Information,
Figure S2F). Similarly, the treatment of VS-functionalized PLA
or PTMC films with cystamine and FITC resulted in apparent
fluorescence (Supporting Information, Figure S2D,E). It appears
that PTMC film has stronger fluorescence than PCL and PLA
films at comparable VS contents. This is likely due to that the
amorphous nature of PTMC facilitates exposure of VS groups to
the surface. The above results conclude that biodegradable
coatings based on VS-functionalized polymers enable versatile
and efficient surface modification under aqueous conditions,
likely due to the fact that (i) VS groups are hydrophilic, rendering
them readily available at the surface in the aqueous environment,
and (ii) VS groups are highly reactive toward the Michael-type
conjugate addition reaction under aqueous conditions. Notably,
unlike amine- or carboxyl-functionalized counterparts in which
the presence of amine or carboxyl groups in the bulk material
largely alters their physicochemical properties, VS-functionalized
biodegradable polymers showed similar solubility and filming
characteristics to the parent polymers without VS functional
groups.
Influence of Surface Modification on Cell Culture. It is
known that the surface chemistry of biomaterials is of critical
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showed that unmodified VS-functionalized PCL films, similar to
tissue culture plate, could well support cell attachment and
growth (Figure 5A,B), indicating that VS-functionalized PCL
film is nontoxic and biocompatible. The VS-functionalized PCL
films following treatment with thiolated nonfouling polymers
such as PEG and glycol chitosan, however, revealed diminished
cell adhesion and growth as compared to the unmodified control
(Figure 5C,D). Notably, excellent cell adhesion and growth were
observed for VS-functionalized PCL films treated with GRGDC
peptide (Figure 5E). These initial results show that the surface of
VS-functionalized biodegradable polymer-based biomaterials
can be elegantly engineered to meet the demands of a particular
application.
’ CONCLUSIONS
We have demonstrated that novel vinyl sulfone-functionalized
biodegradable polymers, which can be readily prepared with
vastly different structures and compositions by ring-opening
copolymerization with vinyl sulfone cyclic carbonate monomer,
provide an unprecedented and robust access to advanced func-
tional biomaterials as well as coatings. To the best of our
knowledge, this represents a first report on direct, efficient, and
clean (without catalyst and byproduct) modification of biode-
gradable coatings under aqueous conditions. These vinyl sulfone-
functionalized biodegradable polymers open a brand-new avenue
to engineering the surface chemistry of biomedical devices and
coatings. We are convinced that these vinyl sulfone-functional-
ized biodegradable polymers have a tremendous potential in
biomedical engineering.
Figure 4. Contact angles of P(CL-co-VSC)8.7% copolymer films
following postpolymerization modification with varying thiol-containing
molecules (Student’s t test, *p < 0.05, **p < 0.01).
’ ASSOCIATED CONTENT
S
Supporting Information. XPS data and fluorescence
b
images. This material is available free of charge via the Internet
at http://pubs.acs.org.
’ AUTHOR INFORMATION
Corresponding Author
*Tel: +86-512-65880098. E-mail: zyzhong@suda.edu.cn.
Author Contributions
†R.W. and W.C. made equal contributions to this work.
’ ACKNOWLEDGMENT
This work is financially supported by research grants from the
National Natural Science Foundation of China (NSFC
50703028, 20974073, 50973078, and 20874070), a Project
Funded by the Priority Academic Program Development of
Jiangsu Higher Education Institutions, and the Program of
Innovative Research Team of Soochow University.
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Figure 5. Images (Â50) of MG63 cells cultured on the unmodified and
modified P(CL-co-VSC)8.7% films for 1, 4, and 6 days: (A) unmodified
P(CL-co-VSC)8.7% film, (B) PEG-SH modified film, (C) GC-SH
modified film, (D) RGDC modified film, and (E) tissue culture plastic
(blank control).
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