Branched cationic polyurethane prepared by polyaddition of chloromethylated five-membered cyclic carbonate and diethylenetriamine in molten salts

Article abstract of DOI:10.1002/pola.24969

A branched cationic polymer was synthesized by polyaddition of 4-chloromethyl-1,3-dioxolan-2-one and diethylenetriamine in molten salts. The polyaddition proceeded through nucleophilic addition of the primary amino group to the cyclic carbonate structure and quarternization of the secondary amino group with the chloromethyl group. The resulting cationic polymer was a good and recyclable catalyst for the reaction of carbon dioxide and epoxides owing to the ammonium structure. The complexation with DNA was confirmed by dynamic light scattering and zeta potential measurements, and it suggests the potential application as a carrier for drug delivery system.

Full text of DOI:10.1002/pola.24969

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Branched Cationic Polyurethane Prepared by Polyaddition of
Chloromethylated Five-Membered Cyclic Carbonate and
Diethylenetriamine in Molten Salts
Bungo Ochiai,1 Kazuhiro Koda,1 Takeshi Endo2
1Department of Chemistry and Chemical Engineering, Faculty of Engineering, Yamagata University, 4-3-16 Jonan,
Yonezawa, Yamagata 992-8510, Japan
2Molecular Engineering Institute, Kinki University, 11-6 Kayanomori, Iizuka, Fukuoka 820-8555, Japan
Correspondence to: T. Endo (E-mail: tendo@mol-eng.fuk.kindai.ac.jp)
Received 23 June 2011; accepted 12 August 2011; published online 30 September 2011
DOI: 10.1002/pola.24969
ABSTRACT: A branched cationic polymer was synthesized by
polyaddition of 4-chloromethyl-1,3-dioxolan-2-one and di-
ethylenetriamine in molten salts. The polyaddition pro-
ceeded through nucleophilic addition of the primary amino
group to the cyclic carbonate structure and quarternization
of the secondary amino group with the chloromethyl group.
The resulting cationic polymer was a good and recyclable
catalyst for the reaction of carbon dioxide and epoxides
owing to the ammonium structure. The complexation with
DNA was confirmed by dynamic light scattering and zeta
potential measurements, and it suggests the potential appli-
C
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cation as a carrier for drug delivery system.
2011 Wiley
Periodicals, Inc. J Polym Sci Part A: Polym Chem 50: 47–51,
2012
KEYWORDS: addition polymerization; biomaterials; branched;
carbonate; catalysis; catalyst; cationic polymer; drug delivery
systems; gene delivery; polyurethane
INTRODUCTION Ionic polymers have wide applications for
carriers for drug delivery system,1–12 biomaterials,13 cata-
lysts,14–17 electrolytes,18,19 and so forth.20 For instance, pol-
y(ethyleneimine) is a branched and cationic polymer, and it
has been used as an efficient gene delivery carrier by ionic
complexation.1–9 However, its severe disadvantage is the tox-
icity, probably because of the permeation ability into cell
menbranes.21,22 A polymer with branched and cationic struc-
tures is a potential candidate of useful biomaterials, espe-
cially if it is less toxic. Cationic moieties play an important
role in complexing with drug candidates, and branched
structures are effective to higher loading of ligands inside.
We thought that such branched and cationic polymers bear-
ing biocompatible structures would have better properties.
We focused on the reaction of five-membered cyclic carbo-
nates with amines to produce hydroxyurethanes, which is
very chemoselective.23–31 The focused chemoselective reac-
tions were (a) selective nucleophilic addition of the primary
amino groups in diethylenetriamine (DETA) to the cyclic car-
bonate group remaining the secondary amine structure
unreacted29 and (b) the selective nucleophilic addition of
primary amines to the cyclic carbonate structure in a five-
membered cyclic carbonate bearing chloromethyl group
(Cl5CC) without quarternization.26 The low toxicity of DETA
may also be mentioned (e.g., the patch testing of 0.05%
aqueous solution did not cause stimulation, and the oral
LD50 values for rats are 819–2600 mg/kg).32 Our strategy
for synthesizing branched and cationic polymers bearing bio-
compatible structures is the polyaddition of DETA and the
cyclic carbonate bearing a halomethyl group, in which the
nucleophilic addition of the primary amino group to the five-
membered cyclic group produces a biocompatible urethane
main chain, and the successive quarternization of the sec-
ondary amine structure with the halomethyl group produces
cationic structure.33–36 We examined this polyaddition under
various conditions and the application of the resulting
branched cationic polymer as a catalyst and a DNA capsule.
RESULTS AND DISCUSSION
Polyaddition of Cl5CC and DETA was conducted using 2:1
molar amounts of the monomers (Scheme 1 and Table 1).
When the polymerizations were conducted in conventional
solvents, the yields were better when polar solvents were
used but the molecular weights were almost identical. The
product obtained in the reaction in THF at room temperature
was the 2:1 adduct resulting from the selective nucleophilic
ring opening of the NH2 group in DETA to the cyclic carbon-
ate group in Cl5CC, indicating the first selective ring opening
produced this intermediate prior to the reactions of the
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SCHEME 1 Synthesis of branched cationic polyhydroxyurethane via polyaddition of Cl5CC with DETA.
secondary amine group or the chloromethyl group in a simi-
lar manner to the reported reactions [Fig. 1(a)].26,29
Although typical nucleophilic addition of amines to cyclic
carbonates gives two products with primary and secondary
OH groups, this intermediate has only secondary hydroxyl
groups owing to the electron-withdrawing character of the
chloromethyl group. Then, we tried polymerizations in mol-
ten salts to conduct the polymerization under more polar
environment.37 The products obtained in molten salts, espe-
cially polar chloride salts, had higher molecular weight than
the products obtained in conventional solvents. This result
suggests the importance of polarity of the polymerization
media. The lower amounts of molten salts were effective to
obtain polymers in higher yields, probably because too high
amounts of molten salts resulted in low concentration for ef-
ficient reactions. The structure of the polymer was eluci-
dated by IR and nuclear magnetic resonance (NMR) [Fig.
1(b,c)] spectroscopic analyses. An strong IR absorption at
1695 cmꢀ1 is assignable to the C¼¼O moieties in the ure-
thane group, and the wavenumber is the same value with
that of typical polyhydroxyurethanes.38 The polymers with
very low molecular weight showed NMR spectra very similar
with the nonionic intermediate produced via the nucleophilic
addition of the primary amine group without quarterniza-
tion, whereas the NMR signal of the methylene protons adja-
cent to the secondary amine moieties was very weak in the
spectra of polymers with Mn higher than 3000. This result
also supports that the nucleophilic addition proceeded prior
to the quarternization. The signals observed at 2.5–3.0 ppm
are attributable to the methylene groups in the terminal
groups not quarternized, and the signals were disappeared
by treatment with HCl aq (data not shown). This polymer
may contain the cyclic urethane structure produced via the
intramolecular cyclization of the chloroalkyl groups and the
secondary amine groups, although its presence could not be
confirmed due to the similarity in the structure. In a similar
manner, we could not determine the degree of branching
due to the overlap of the signals.
ether (GPE) proceeded quantitatively, and Cl5CC and 4-phe-
noxymethyl-1,3-dioxolane-2-one were obtained in quantita-
tive yields from the corresponding epoxides. The polymeric
catalyst could be recycled twice keeping the good activity via
precipitation of the polymer with acetone. Contrary to this,
the reaction using a low-molecular-weight polymer (e.g., run
4) was unsuccessful (30% yield at the reaction of 4-phenox-
ymethyl-1,3-dioxolane-2-one for 24 h) due to the lower con-
tents of the ammonium structure serving as the catalyst for
this reaction.
Complexation with DNA was confirmed by dynamic light
scattering (DLS) experiments (Fig. 2). Tris-HCl solutions of
DNA (pH ¼ 8.0 and 10 mg/mL) were mixed with an aqueous
polymer solution (100 mg/mL, 10 mL). The mixtures were
transparent when 1.0 or 0.5 wt % amounts of DNA were
added. Clear scattering patterns indicated the formation of
polyplexes with diameters of 171 and 148 nm, respectively.
TABLE 1 Polyaddition of Cl5CC with DETAa
Run
Solvent (mL)
None
Temp. (ꢁC) Yield (%)b Mn (Mw/Mn)c
1
100
50
51
16
51
40
30
72
44
51
15
86
90
97
46
46
1,200 (1.59)
1,280 (1.24)
1,110 (1.27)
940 (1.29)
2
MeOH (1.0)
MeOH (1.0)
MeOH (1.0)
THF (1.0)
3
80
4
100
100
100
100
100
100
100
100
5
1,660 (1.39)
1,030 (1.42)
1,050 (1.37)
1,000 (1.43)
2,090 (1.18)
1,530 (1.37)
1,680 (1.63)
1,480 (1.32)
2,400 (1.20)
3,120 (1.81)
6
H2O (1.0)
7
DMF (1.0)
8
DMSO (1.0)
BMIBF4 (1.0)
BMIBF4 (0.5)
BMIPF6 (0.5)
9
10
11
12
13
14
EMICF3SO3 (0.5) 100
BMICl (0.5)
100
100
BMICl (0.05)
BMIBF4, 1-n-butyl-3-methylimidazolium tetrafluoroborate; BMIPF6, 1-n-
butyl-3-methylimidazolium hexafluorophosphate; EMICF3SO3, 1-ethyl-3-
methylimidazolium trifluoromethanesulfonate; BMICl, 1-n-butyl-3-meth-
ylimidazolium chloride.
As applications of the resulting polymer, we evaluated the
catalyst activity for the reaction of an epoxide and carbon
dioxide14,15,17,39 and the DNA complexing ability. First, the
branched cationic polymer (run 14) was used as a catalyst
for the reaction of epoxides with carbon dioxide in N-methyl-
pyrrolidone (2 mol/L) (Scheme 2 and Table 2). The carbona-
tion reactions of epichlorohydrin (ECH) and glycidyl phenyl
a
Conditions: Cl5CC (1.83 mmol), diethylenetriamine (0.915 mmol), 24 h,
degassed sealed tube.
b
Isolated yield after precipitation with acetone.
Estimated by SEC [poly(ethylene glycol) standards, CH3CN/0.5
c
N
NaNO3 aq. (v/v ¼ 1/1)].
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FIGURE 1 1H NMR spectra of (a) the intermediate, (b) oligomer obtained in run 4, and (c) polymer obtained in run 14.
The diameters of the polyplexes were shorter than the diam-
eter of the cationic polymer (478 nm). The longer diameter
of the cationic polymer in this buffered solution indicates
that the cationic polymer has self-aggregation character via
dynamic intermolecular ion exchange. The smaller sizes of
the polyplexes may be ascribed to the decreased dynamic
character of the ion exchange originating from the multiva-
lent character of the anionic moieties in DNA. Emulsification
took place when more than 2 wt % amounts of DNA were
added. The DLS measurements were difficult due to the very
broad size distribution, and the diameters of the resulting
complex exceeded 1000 nm. We presume that the emulsifica-
tion originated from the insufficient amounts of the cationic
polymer to cover DNA completely. The zeta potentials, which
are the indexes of the total charges in the complexes,
decreased as the amount of anionic DNA added. The shift to
negative potential suggests that the neutralization between
the cationic polymer and DNA took place.
EXPERIMENTAL
Materials
4-Chloromethyl-1,3-dioxolane-2-one (Cl5CC) was prepared
by the reaction of ECH (Tokyo Kasei Kogyo) and CO2 in N-
methylpyrrolidinone (Kanto Chemical) catalyzed by LiBr
(Kanto Chemical, Tokyo, Japan) and distilled under reduced
pressure.35 The spectroscopic data agreed with those in a lit-
erature.36 DETA (Tokyo Kasei Kogyo) was dried with CaH2
and distilled under reduced pressure. Low-molecular-weight
DNA from salmon sperm (Fluka) was used for the DNA com-
plexation experiment. The DNA was dissolved in a Tris-HCl
solution (0.05 M, pH ¼ 8.0), which is a solvent recom-
mended by the supplier. Other reagents were used as
received.
Measurements
NMR spectroscopy measurements were performed on a JEOL
ECX-400 instrument using tetramethylsilane as an internal
standard at ambient temperatures (400 MHz for 1H, 100
MHz for 13C, and 40 MHz for 15N). Molecular weights were
measured by size exclusion chromatography performed on a
system consisting of a Tosoh DP-8020 pump, a Viscotek TDA
Model 300 detector (refractive index detector was used),
and tandem columns of Tosoh TSK gel G3000PWXL-CP (pore
size: 7 lm) and TSK gel G5000PWXL-CP (pore size: 10 lm).
A mixed solvent of acetonitrile and 0.1 mol/L NaNO3
SCHEME 2 Reaction of epoxides with carbon dioxide catalyzed
by the branched cationic polymer.
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aqueous solution (v/v ¼ 50/50) was used as an eluent at 40
ꢁC. DLS and zeta potential measurements were performed
using a Malvern Zetasizer nano-ZS instrument equipped with
a 4-mW He–Ne laser (633 nm) in 12-mm-square glass cuv-
ettes at 25 ꢁC. Dip cell was used for zeta potential
measurement.
Polyaddition of Cl5CC and DETA (Typical Procedure)
Cl5CC (0.250 g, 1.83 mmol), DETA (0.0945 g, 0.915 mmol),
and solvents were stirred in a degassed sealed tube. The
resulting mixture was diluted with water or methanol and
poured into an excess amount of acetone. The precipitate
was collected by centrifugation and drying under reduced
pressure.
Synthesis of Intermediate from Cl5CC and DETA
FIGURE 2 Size (solid circle) and zeta potential (open square) of
Cl5CC (0.546 g, 4.00 mmol), DETA (0.206 g, 2.00 mmol), and
THF (2.0 mL) were stirred at 25 ꢁC for 24 h in a degassed
sealed tube. The resulting mixture was poured into an excess
amount of diethyl ether. The viscous product, bis(3-chloro-2-
hydroxypropyl)carbamate of DETA was obtained by drying
under reduced pressure (0.646 g, 1.72 mmol, 86%). The
product contained a trace amount of THF to avoid self-quar-
ternization spontaneously took place in the absence of THF.
polymer–DNA complexes under various DNA–polymer ratios.
tone-soluble parts. The polymer was isolated as the acetone-
insoluble part and used as a recycled catalyst.
Complexation of the Branched Ionic Polymer and DNA
An aqueous solution of the polymer (Mn ¼ 3330, 20 mg/mL,
1 mL) was mixed with various amounts of Tris-HCl (0.05 M,
pH ¼ 8.0) aqueous solution of DNA (10 mg/mL, 0–5 wt %
to polymer) at room temperature. The solution was adjusted
to 2 mL by addition of Tris-HCl aq (0.05 M, pH ¼ 8.0). Then,
the mixture was sonicated for 1 min and filtered with a
membrane filter (Whatman GMF-150, 2 lm). The filtered so-
lution was analyzed by DLS and zeta potential
measurements.
1H NMR (400 MHz, d in ppm, CD3OD): 2.70 (t, J ¼ 12.2 Hz,
4H, ANHCH2CH2NHCOOA), 3.23 (t,
J
¼
12.2 Hz, 4H,
ANHCH2CH2NHCOOA), 3.50–3.73 (m, 4H, ClCH2A), 3.92–
4.00 (m, 2H, ACHA), 4.07–4.13 (m, 4H, AOCH2A). 13C NMR
(100 MHz, d in ppm, CD3OD): 40.04 (ANHCH2CH2NHCOOA),
45.55 (ClCH2A), 48.31 (ANHCH2CH2NHCOOA), 65.89
(AOCH2A), 69.31 (ACHA), 157.47 (C¼¼O). 15N NMR (40
MHz, d in ppm, CD3OD): 25 (ANHCH2CH2NHCOOA), 75
(ANHCH2CH2NHCOOA).
CONCLUSIONS
Branched cationic polyurethane was obtained by polyaddi-
tion of a five-membered cyclic carbonate bearing chloro-
methyl group and DETA. Molten salts were good media for
this polyaddition by the high polarity. Owing to the ammo-
nium structure, the polymer could serve as a catalyst and a
capsule for DNA. The potential application of the branched
cationic polymer is a recyclable catalyst and a gene delivery
carrier. Specifically, we believe that the application to gene
delivery system is promising, considering the biocompatible
polyurethane backbone.
Reaction of Epoxides and CO2 Catalyzed by the Branched
Cationic Polymer
Typical procedure: GPE (0.45 g, 3.0 mmol) or ECH (0.28 g,
3.0 mmol) and the polymer (Mn ¼ 970 or 3330, 0.30 mmol
unit) were dissolved in NMP (1.5 mL). The mixture was
stirred for 12 h at 100 ꢁC with bubbling CO2. The composi-
tion in the mixture was confirmed by 1H NMR analysis. The
product from GPE was isolated by precipitation with water
and drying under reduced pressure. The product from ECH
was isolated by Kugelrohr distillation after collecting the ace-
The authors thank Seigou Kawaguchi for his kind advises.
TABLE 2 Reaction of Epoxides with Carbon Dioxide Catalyzed
by the Branched Cationic Polymer
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