An approach to biodegradable star polymeric architectures using disulfide coupling

Article abstract of DOI:10.1039/b817037a

The straightforward synthesis of biodegradable star polymers via both in situ polymerization from a trifunctional RAFT agent and post-polymerization conjugation of pyridyldisulfide-ended linear polymers to a trithiol precursor is described. The Royal Society of Chemistry 2008.

Full text of DOI:10.1039/b817037a

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An approach to biodegradable star polymeric architectures using
disulfide couplingw
Jingquan Liu,* Huiyun Liu, Zhongfan Jia, Volga Bulmus and Thomas P. Davis*
Received (in Cambridge, UK) 29th September 2008, Accepted 30th October 2008
First published as an Advance Article on the web 12th November 2008
DOI: 10.1039/b817037a
The straightforward synthesis of biodegradable star polymers
via both in situ polymerization from a trifunctional RAFT agent
and post-polymerization conjugation of pyridyldisulfide-ended
linear polymers to a trithiol precursor is described.
studied and utilized for the synthesis of functional polymers
and biomolecule–polymer conjugates.23,24,26–30 However, to
the best of our knowledge, the well-defined multi-armed star
architecture containing disulfide intra-linkages has not been
reported yet. In this communication we report the direct
synthesis of three-arm star polymeric architectures using both
‘‘core-first’’ and ‘‘arm-first’’ methodologies to generate a
three-armed architecture containing biodegradable disulfide
linkages, as outlined in Scheme 1. In the ‘core-first’ approach a
trifunctional initiator was first synthesized by covalently at-
taching a trithiocarbonate RAFT agent, 3-[pyridyldisulfide]-
ethyl, 3-benzyltrithiocarbonate propionate (PDEBP) to a
trithiol precursor, trimethylolpropane tris[3-mercaptopropio-
nate] (TMPMP) through disulfide coupling via their z-group,
followed by the direct chain propagation from the RAFT core
to afford three-armed star polymers. The same structure could
also be achieved by ‘arm-first’ methodology, where a linear
polymeric chain with PDS functionality was synthesized first
using PDEBP, followed by the covalent coupling to TMPMP
via disulfide linkage.
Multi-armed star polymeric architectures have attracted
increasing interest due to their potential applications in a
number of areas, e.g. encapsulation, sensing, catalysis, elec-
tronics, optics, biological engineering, coatings, additives,
drug- and gene delivery.1,2 Generally speaking, these compli-
cated architectures could be achieved by ‘arm-first’3–6 or ‘core-
first’ methodologies. Recently, the more straightforward
‘‘core-first’’ strategy has drawn an increasing interest for
generating multi-armed polymeric architectures in a more
controllable mode.7–11 Star polymers consisting of miktoarms
have also been tailored to achieve different properties.6,12,13
In addition to ionic and coordination ring-opening poly-
merization,14,15 living radical polymerizations (LRPs) e.g.
atom transfer radical polymerization (ATRP),16,17 nitroxide-
mediated radical polymerization (NMRP)18 and reversible
addition fragmentation chain transfer (RAFT) polymeriza-
tion19–21 have been exploited extensively to generate multi-
armed structures with predetermined molecular weights and
narrow molecular weight distributions. The combination of
different polymerization methods were also used for genera-
tion of more complicated polymeric architectures.3,13
The three-armed RAFT agent was first characterized by 1H
NMR. As shown in Fig. 1 the disappearance of the peaks at
Most multi-armed polymeric structures have their arms
joined to a core by stable covalent bonding. Therefore it is
usually difficult to cleave the arms under mild conditions. The
creation of complex polymeric structures with degradable
linkages, (e.g. disulfide is cleavable in the presence of
GSH22–24 and acetal is acid labile25) is of great importance
in biological applications. The disulfide bond is a reversible
linkage that exists ubiquitously in proteins and enzymes. It is
biodegradable in vivo in the presence of glutathione (GSH), the
most abundant intracellular thiol (0.2–10 mM) in most mam-
malian and many prokaryotic cells.22 Therefore a polymeric
structure intra-linked by disulfide bonding could be easily
cleaved in vivo and excreted subsequently over a much shorter
period compared with its precursor.
Linear polymers and hydrogels with pyridyldisulfide (PDS)
ended groups or disulfide intra-linkages have been extensively
Centre for Advanced Macromolecular Design (CAMD), The
University of New South Wales, Sydney, NSW 2052, Australia.
E-mail: jingquan.liu@unsw.edu.au; t.davis@unsw.edu.au;
Fax: +61-2-9385 6250; Tel: +61-2-9385 4371
w Electronic supplementary information (ESI) available: Full experi-
mental details, kinetic plots, 1H NMR, ESI-mass and UV-Vis spectra.
See DOI: 10.1039/b817037a
Scheme 1 ‘Core-first’ and ‘arm-first’ methodologies to generate
identical three-armed polymers and the subsequent cleavage into
single-armed linear chains.
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Fig. 1 1H NMR of three-armed RAFT agent.
8.49, 7.67 and 7.11 ppm from the PDS groups on PDEBP
precursor (Fig. S1, ESIw) evidenced the successful coupling of
the RAFT to trithiol precursor through the thiol–disulfide
exchange reaction. The ratio of integration of protons from
the phenyl groups at 7.33 ppm to those from TMPMP
confirmed that each TMPMP molecule attached three RAFT
molecules. The successful synthesis of the three-armed RAFT
core was also supported by the presence of a parent sodium
ion at m/z 1411.07 (calc. 1411.04) in its electrospray ionization
(ESI) mass spectrum. (Fig. S2, ESIw)
The direct polymerizations of NIPAAm and styrene using
the three-armed RAFT core proved to be well controlled
by RAFT mechanism. As shown in Fig. S3(a,b)w and
Fig. S4(a,b)w the monomer conversion increased with increas-
ing polymerization time and the radical concentration was
constant during the whole polymerization as indicated by
pseudo-first order plot. The polydispersity indices (PDI) for
purified PNIPAAm and PSt were 1.28 and 1.24, respectively.
The low PDI values and the well-defined chromatograms from
gel permeation chromatography (GPC) analysis strongly
suggested that the chain propagation on the three arms was
synchronous. The NMR signals from the RAFT residue could
be observed clearly in the 1H NMR spectra of purified
PNIPAAm and PSt in Fig. S3(c)w and Fig. S4(c)w, evidencing
the integrity of RAFT moieties after polymerization.
Fig. 2 (a) GPC traces of three-armed PNIPAAm (Mw 10 000 from 1H
NMR, PDI 1.28) (filled circles) and cleaved mixture in 0.1 M DTT
(empty circles) and (b) GPC traces of linear PSt (empty circles),
conjugation mixture with TMPMP (filled circles) and conjugation
mixture after cleavage in 0.1 M DTT for 2 h (filled diamonds).
star was calculated to be 2.3 times of its one-armed precursors.
This observation is consistent with the result obtained from
theoretical simulation.32 The star polymer was cleaved in the
presence of DTT. After cleavage, the molecular weight dis-
tribution overlapped with that of the original one-armed
precursor, consistent with complete cleavage. The covalent
coupling of the linear polymeric chain to the trithiol core was
monitored by UV-Vis spectroscopy. Upon reaction with
TMPMP, the release of 2-pyridinethione was observed clearly
at 370 nm, its characteristic absorption wavelength in dimethyl
acetamide.33 (Fig. S6, ESIw).
The cleavability of three-armed PNIPAAm was first checked
by DL-dithiothereitol (DTT). 10 mM PNIPAAM was cleaved
into one-armed chains in 2 hours in 0.1 M DTT aq. solution
(pH 6.5). (Fig. 2a). After cleavage, the PDI of the single-armed
chain decreased to 1.20, furnishing strong evidence for successful
living polymerization. The Mw of the single-armed chain after
cleavage was calculated to be 3800 from GPC, which is approxi-
mately one third of its precursor. The bio-degradability of three-
armed PNIPAAm was tested by incubating 5 mg polymer in
GSH solution (40 mM in 1 ml pH 5.0 phosphate buffer). After
48 hours, 30% of three-armed polymer was found to be cleaved
(Fig. S5, ESIw). Apparently, the disulfide-cleavability of GSH
was much weaker than that of DTT. The adoption of lower pH
was based on the principle that lower pH will increase the
reduction potential of GSH according to the Nernst equation.31
The ‘arm-first’ methodology was also employed to synthe-
size the identical three-armed structure. One-armed linear PSt
with terminal PDS groups was sythesized first, followed by
covalent coupling to TMPMP. GPC traces indicated an
increase of Mw after conjugation. (Fig. 2b) The Mw of the
The coupling reaction of linear PNIPAAm with TMPMP
was also characterized using ESI mass spectroscopy. The mass
1
spectrum of linear PNIPAAm (Mn 820 from H NMR, PDI
1.12) is shown in Fig. 3a. After coupling the Mw of the star
polymer increased significantly. (Fig. 3b) It should be noted
that any three of the linear chains could couple to one
TMPMP to form a three-armed structure. However the
three-armed structure formed by the same linear chains was
still identified from the spectrum of the reaction mixture, as
shown in Fig. 3.
In summary, we have successfully demonstrated the synth-
esis of bio-degradable three-armed polymers using both
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of conjugates using a variety of biomolecules such as proteins,
peptides, oligoneucleotides, si-RNA, and genes for the appli-
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TPD thanks Australian Research Council for Federation
Fellowship Award. JL acknowledges the UNSW Vice
Chancellor’s Post-doctoral Research Fellowship.
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