Versatile synthesis of end-functionalized thermosensitive poly(2-isopropyl-2-oxazolines)

Article abstract of DOI:10.1021/ma049677n

The synthesis of several end-functionalized poly(2-isopropyl-2-oxazolines) (PiPrOx) has been achieved via cationic ring-opening polymerization of 2-isopropyl-2-oxazoline. Poly(2-isopropyl-2-oxazolines) bearing primary amino groups at one chain end (Me-PiP

Full text of DOI:10.1021/ma049677n

6786
Macromolecules 2004, 37, 6786-6792
Versatile Synthesis of End-Functionalized Thermosensitive
Poly(2-isopropyl-2-oxazolines)
J oon -Sik P a r k ,† Yosh itsu gu Ak iya m a ,† F r a n c¸oise M. Win n ik ,‡ a n d
Ka zu n or i Ka ta ok a *,†
Department of Materials Science, Graduate School of Engineering, The University of Tokyo,
7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, J apan, and Faculty of Pharmacy and
Department of Chemistry, Universite´ de Montre´al, CP 6128 Succursale Centre Ville,
Montreal, Quebec, Canada H3C 3J 7
Received February 16, 2004; Revised Manuscript Received May 30, 2004
ABSTRACT: The synthesis of several end-functionalized poly(2-isopropyl-2-oxazolines) (PiPrOx) has been
achieved via cationic ring-opening polymerization of 2-isopropyl-2-oxazoline. Poly(2-isopropyl-2-oxazolines)
bearing primary amino groups at one chain end (Me-PiPrOx-NH2, with Mn ranging from 3600 to 9700)
were obtained by conversion of hydroxyl-terminated poly(2-isopropyl-2-oxazolines) (Me-PiPrOx-OH) via
phthalimide activation of the hydroxyl groups and subsequent hydrazine treatment. Heterotelechelic
PiPrOx carrying an R-acetal and an ω-hydroxyl group (acetal-PiPrOx-OH) were prepared via cationic
ring-opening polymerization of 2-isopropyl-2-oxazoline initiated with 3,3-diethoxy-1-propyl tosylate. The
polymerizations carried out under mild conditions (40-45 °C) for extended periods of time yielded polymers
of well-controlled molecular weight (MW) and narrow molecular weight distribution (MWD). Analysis of
1
the polymers by H and 13C NMR spectroscopy, ion-exchange HPLC, and MALDI-TOF mass measure-
ments indicated that nearly quantitative end-functionalization was achieved in all cases. Aqueous PiPrOx
solutions (10 mM PBS (pH 7.4) containing 150 mM NaCl) possess a cloud point temperature near 37 °C,
as determined by turbidity. Thermosensitive telechelic PiPrOx offer promising applications as smart
materials including bioconjugates, hydrogels, and drug carriers.
In tr od u ction
enhanced biocompatibility and prolonged blood circula-
tion times, comparable to those of conventional poly-
(ethylene glycol) (PEG) lipopolymers.8,9
Poly(oxazolines) (POx) have emerged recently as
materials of importance in surface chemistry and bio-
materials science, where they may act as nonionic
surfactants, protein modifiers, hydrogels, and drug
carriers.1,2 Under appropriate conditions, the cationic
ring-opening polymerization of oxazolines (Ox) is known
to proceed by a living polymerization process to afford
poly(N-acylethylenimines), often viewed as “pseudopep-
tides”.3 A variety of POx can be prepared by changing
either the alkyl substituent of the starting oxazoline or
the end group. POx possessing a short alkyl substituent,
e.g., methyl or ethyl group, in the side 2-position are
water-soluble. However, the hydrophilicity of POx de-
creases as the length of the alkyl substituent increases,
resulting in insolubility in water at all temperatures or
in certain temperature ranges. Of particular interest are
poly(2-isopropyl-2-oxazolines) (PiPrOx) which possess
an isopropyl group in the side 2-position. These poly-
mers are soluble in cold water, but their aqueous
solutions have a cloud point near physiological condi-
tions,4 like poly(N-isopropylacrylamide) (PNIPAM), the
typical representative of thermosensitive polymers with
numerous applications.5-7 The main advantage of
PiPrOx, which is a POx homologue, is that it is highly
expected to be a biocompatible thermosensitive polymer
and, as such, to be extremely useful in biomedical
applications. It should be noted that POx, as a rule, are
nontoxic and that several POx carry US Food and Drug
Administration (FDA) approval. Zalipsky et al. reported
that poly(2-ethyl-2-oxazoline)-modified liposomes exhibit
Our group has established synthetic strategies, via
an anionic ring-opening polymerization catalyzed by
functionalized initiators, toward quantitative and selec-
tive syntheses of heterotelechelic PEG, leading to func-
tionalized telechelic polymers with high potential utility
in the biomedical field.10-13 Applying a similar approach
toward the cationic polymerization of Ox derivatives, it
should be possible to prepare a variety of heterotelech-
elic PiPrOx derivatives. Moreover, end-functionalized
block copolymers can be prepared by using heterotelech-
elics as macroinitiators, as in the case of PEG.14 Thus,
the objective of this work was to design end-function-
alized PiPrOx homologues, which can be widely used
as intelligent biomaterials bearing thermosensitivity
near the body temperature of human beings.15
On the basis of this synthetic strategy, we devised
two synthesis routes toward end-functionalized PiPrOx,
as depicted in Scheme 1 and Scheme 2. One process
leads to semitelechelic PiPrOx possessing primary
amino groups at one chain end (Scheme 1). This amine-
terminated PiPrOx may be used for bioconjugation or
as a macroinitiator to incorporate a second monomer
and create a diblock copolymer, opening up the facile
generation of new biofunctional block copolymers. Fur-
thermore, heterotelechelic PiPrOx of greater versatility,
having R-acetal and ω-hydroxyl groups, can be obtained
by the route depicted in Scheme 2. The acetal group is
easily converted to an aldehyde group under mild acidic
conditions. This group is readily amenable to conjuga-
tion with proteins, as it is stable in water and reacts
rapidly with primary amines. The synthesis of these two
new families of polymers is reported here, together with
the polymer characterization by 1H and 13C NMR
† The University of Tokyo.
‡ Universite´ de Montre´al.
* To whom correspondence should be addressed: Tel +81-5841-
7138, Fax +81-5841-7139; e-mail kataoka@bmw.t.u-tokyo.ac.jp.
10.1021/ma049677n CCC: $27.50 © 2004 American Chemical Society
Published on Web 08/05/2004

Macromolecules, Vol. 37, No. 18, 2004
Poly(2-isopropyl-2-oxazolines) 6787
Sch em e 1. Syn th esis of Me-P iP r Ox-NH2 by
F u n ction a liza tion of ω-Hyd r oxyl Ter m in a ls
(TOSOH HLC-8220) system equipped with two TSK gel
columns (G4000HHR and G3000HHR) and an internal refractive
index (RI) detector. Columns were eluted with DMF containing
lithium bromide (10 mM) and triethylamine (30 mM) with a
flow rate of 0.8 mL/min and at a temperature of 40 °C.
Molecular weights were calibrated with poly(ethylene glycol)
standards (Polymer Laboratories, Ltd., UK). Mass measure-
ments were performed on a MALDI-TOF mass spectrometer
(Bruker REFLEX III), operating at an acceleration voltage of
23 kV in the reflection mode. UV-vis spectra were obtained
with a V-550 UV/vis J ASCO spectrophotometer. The function-
ality of amino group was determined by using an ion-exchange
HPLC system equipped with TSK gel column (TOSOH SP-
5PW) and an internal refractive index (RI) detector. The
column was eluted with phosphate-buffered solution (2 mM
PBS (pH 6.5)) at a flow rate of 0.5 mL/min and at a temper-
ature of 30 °C.
Syn th esis of 3,3-Dieth oxy-1-p r op yl Tosyla te. A solution
of p-toluenesulfonyl chloride (9.61 g, 50.4 mmol, 1.5-fold molar
excess vs OH) in chloroform (50 mL) was added to a stirred
solution of 3,3-diethoxy-1-propanol (4.98 g, 33.60 mmol) and
triethylamine (23.5 mL, 168 mmol, 5-fold molar excess vs OH)
in anhydrous chloroform (150 mL) cooled to 0 °C and kept
under an argon atmosphere. At the end of the addition, the
reaction mixture was brought to room temperature and stirred
for 24 h. The mixture was filtered, and the filtrate was
concentrated under reduced pressure. The residue was purified
by chromatography on a silica gel column eluted with chloro-
form. Fractions containing the desired product were dried over
anhydrous sodium sulfate, filtered, and concentrated to dry-
ness. The product was stored at -20 °C under an argon
atmosphere (Figure S4).
Sch em e 2. Syn th esis of Heter otelech elic
Aceta l-P iP r Ox-OH
1H NMR (DMSO-d6, 400 MHz): δ (ppm) ) 1.10 (t, J ) 9.6
Hz, 3H, CH3CH2O), 2.80 (q, J ) 9.6 Hz, 2H, CH3CH2O), 4.50
(t, J ) 9.6 Hz, 1H, CHCH2CH2), 1.80 (q, J ) 9.6 Hz, 2H,
CHCH2CH2), 4.0 (t, J ) 9.6 Hz, 2H, CHCH2CH2), 7.50 (d, J )
8.5 Hz, 2H, C6H4CH3), 7.80 (d, J ) 8.5 Hz, 2H, C6H4CH3), 2.45
(s, J ) 8.5 Hz, 3H, C6H4CH3).
spectroscopy, MALDI-TOF mass spectrometry, and gel
permeation chromatography. Moreover, the tempera-
ture sensitivity of PiPrOx aqueous solutions was exam-
ined via turbidity measurements.
Syn th esis of P oly(2-isop r op yl-2-oxa zolin e) Ha vin g a n
Hyd r oxyl Gr ou p a t th e ω-Ter m in a l En d (Me-P iP r Ox-
OH). 2-Isopropyl-2-oxazoline (10 g, 88.4 mmol) was added via
a syringe to a solution of methyl p-tosylate (0.186 g, 1.0 mmol)
in acetonitrile (30 mL). The polymerization mixture was stirred
at 42 °C for ca. 506 h under an argon atmosphere. The mixture
was cooled to room temperature and treated with methanolic
NaOH (1 M) to quench the poly(2-isopropyl-2-oxazoline) ox-
azolinium living end groups. The solution was dialyzed for 2
days against distilled water to introduce an hydroxyl group
at one of the chain ends. Freeze-drying of the dialyzed solution
yielded a white colorless powder. In addition, eight samples
were collected in the course of the polymerization. They were
subjected to the same treatment and analyzed by GPC in order
to determine the conversion yield (total yields: 5.1 g, 51%).
Con ver sion of Ter m in a l Hyd r oxyl Gr ou p s in to P r i-
m a r y Am in o Gr ou p s via th e Mitsu n obu Rea ction (Me-
P iP r Ox-NH2) (Sch em e 1).18 Me-PiPrOx-OH (100 mg, 0.022
mmol) was added to a stirred solution of triphenylphosphine
(TPP) (57.7 mg, 0.22 mmol) and phthalimide (PI) (32.37 mg,
0.22 mmol) in THF (1 mL). Then, diethyl azodicarboxylate
(DEAD) (38.32 mg, 0.22 mmol) was added dropwise to the
mixture kept under an argon atmosphere. After being kept at
room temperature for 24 h, the mixture was dialyzed first
against ethanol and then against distilled water using a
Spectrapor dialysis membrane with a 1000 Mr molecular
weight cutoff value. The solution lyophilized for 2 days yielded
the phthalimide-activated polymer “Me-PiPrOx-PI”. The yield
was 90 mg and the conversion efficiency was ca. 70%,
Exp er im en ta l Section
Ma ter ia ls. 2-Isopropyl-2-oxaxoline was synthesized from
isobutyric acid (Wako Pure Chemical Industries, Ltd., Osaka,
J apan) and 2-aminoethanol (Wako Pure Chemical Industries,
Ltd., Osaka, J apan) as described previously (see Supporting
Information, Figures S1 and S2).16 Methyl p-tosylate (Tokyo
Kasei Kogyo Co., Ltd., Tokyo, J apan) was distilled from
calcium hydride under reduced pressure. 3,3-Diethoxy-1-
propanol (Aldrich Chemical Co. Ltd., Milwaukee, WI) and
p-toluenesulfonyl chloride (Wako Pure Chemical Industries,
Ltd., Osaka, J apan) were used as received. Acetonitrile (Wako
Pure Chemical Industries, Ltd., Osaka, J apan) was distilled
from calcium hydride. Tetrahydrofuran (THF) (Wako Pure
Chemical Industries, Ltd., Osaka, J apan) and chloroform
(Wako Pure Chemical Industries, Ltd., Osaka, J apan) were
purified by distillation following conventional procedures.17
Other chemicals, such as 1 N NaOH aqueous solution,
methanol, and ethanol, were purchased from Wako Pure
Chemical Industries, Ltd., Osaka, J apan, and used without
further purification. Triphenylphosphine (TPP) (Tokyo Kasei
Kogyo Co., Ltd., Tokyo, J apan), phthalimide (PI) (Wako Pure
Chemical Industries, Ltd., Osaka, J apan), diethyl azodicar-
boxylate (DEAD) (40 wt % in toluene; Tokyo Kasei Kogyo Co.,
Ltd., Tokyo, J apan), and hydrazine monohydrate (Wako Pure
Chemical Industries, Ltd., Osaka, J apan) were used as re-
ceived.
1
confirmed by comparing integral ratio obtained from H NMR
analysis. This material (50 mg, 0.011 mmol) was then dissolved
in ethanol (5 mL) and treated with hydrazine monohydrate
(5 mL). The mixture was kept at room temperature for 24 h.
After ethanol was evaporated, an aqueous sodium hydroxide
was added up to pH 9-10. The polymer was extracted with
methylene chloride and then purified by dialysis against
ethanol and distilled water using a Spectrapor dialysis mem-
Tech n iqu es. 1H NMR and 13C NMR spectra were recorded
on a J EOL EX 300 spectrometer (J EOL, Tokyo, J apan) at 400
MHz. Chemical shifts are reported in parts per million (ppm)
downfield from tetramethylsilane. Molecular weights and
molecular weight distributions were determined using a GPC

6788 Park et al.
Macromolecules, Vol. 37, No. 18, 2004
brane with a 1000 Mr cutoff value. Lyophilization of the
dialyzed solution gave the amine-terminated polymer “Me-
PiPrOx-NH2”. The yield was 30 mg and the conversion
efficiency, viz., functionality of amino group, was ca. 89%,
determined by the analysis of ion-exchange HPLC.
Syn th esis of Heter otelech elic P oly(2-isop r op yl-2-ox-
a zolin e) Con ta in in g Aceta l a n d Hyd r oxyl Ter m in a ls
(Aceta l-P iP r Ox-OH) (Sch em e 2). 2-Isopropyl-2-oxazoline
(9.74 g, 86 mmol) was added to a solution of 3,3-diethoxy-1-
propyl tosylate (0.30 g, 1.0 mmol) in acetonitrile (30 mL). The
resulting mixture was stirred at 45 °C for 240 h under an
argon atmosphere. An aliquot taken from the reaction mixture
(1 mL) was cooled to room temperature. It was added to
methanolic NaOH (3 mL, 1 M) and subsequently dialyzed for
2 days against distilled water to yield heterotelechelic PiPrOx
with hydroxyl groups at the ω-end (acetal-PiPrOx-OH). The
solution was then evaporated, and the residue was dissolved
in methanol (20 mL) and dialyzed for 2 days against distilled
water. The polymer was recovered as a white colorless powder
by freeze-drying of the dialyzed solution.
F igu r e 1. Gel permeation chromatograms of three Me-
PiPrOx-OH samples having different molecular weights (PEG
standard; eluent: DMF (containing 10 mM LiCl and 30 mM
TEA); temperature: 40 °C; RI detection).
MALDI-TOF Ma ss Sp ectr om etr y. An external calibra-
tion was performed by using poly(ethylene glycol) standards
(MeO-PEG-OH; MW ) 5000, NOF Corp.). Ions were generated
by laser desorption at 337 nm (N2 laser, 3 ns pulse width, 106-
107 W/cm2). For each spectrum, approximately 400 transients
were accumulated, and all spectra were recorded in the
reflection mode. Data evaluation was performed with the
Bruker XMASS program, using the reflection spectra only to
achieve better signal-to-noise ratio. 1,8,9-Trihydroxyanthracene
(Aldrich, Milwaukee, WI) was selected as a suitable matrix.19
To prepare the matrix, a solution of 1,8,9-trihydroxyan-
thracene (20 µL, 25 mg mL-1) was mixed with a solution of
the polymer in THF (20 µL, 6.5 mg/mL). Finally, a solution of
lithium trifluoroacetate in THF (2 µL, 2 mg/mL) was added.
The resulting mixture was shaken for few seconds. An aliquot
of the mixture (1 µL) was placed on the target plate and
applied to the probe. The lithium salt was added to the matrix
in order to promote selective and quantitative ionization of
the sample.
Tu r bid ity Mea su r em en ts by Usin g UV-Vis Sp ectr o-
p h otom eter . Cloud points were determined by spectropho-
tometric detection of the changes in transmittance (λ ) 500
nm) of aqueous polymer solutions (0.07-1.0 wt %) heated at
a constant rate (0.2 °C min-1). The samples were thermostated
with a temperature-controlled circulating water bath. Values
for the cloud point of polymer solutions were determined as
the temperature corresponding to 1.0% decrease in optical
transmittance.
F igu r e 2. MALDI-TOF mass spectrum of (a) Me-PiPrOx-
OH (after 119 h) and (b) its expanded spectrum in the region
of 5300-5800.
1
The H NMR spectrum of Me-PiPrOx-OH in CDCl3
(Figure S3 in Supporting Information) presents a broad
singlet at 3.4 ppm attributed to the methylene protons
of polymer backbone, two broad singlets at 2.6-2.8 ppm
ascribed to the side-chain isopropyl methine proton, and
a strong broad singlet due to the resonance of the side-
chain methyl protons (1.1 ppm). The polymer structure
is supported also by the 13C NMR spectrum of the
polymer in CDCl3, shown in Figure S5a in the Support-
ing Information. End-group analysis of the polymer was
performed from the MALDI-TOF mass spectra re-
corded for a Me-PiPrOx-OH (polymerization time: 119
h, Figure 2). Assuming that only parent ions are
detected by the MALDI-TOF mass analysis,21 and
given the molecular mass of the iPrOx monomer, the
molar mass of the end groups, and the metal ions
present, the MW of the Me-PiPrOx-OH samples can be
determined. These values were then compared to Mn
and Mw derived from GPC analysis of the same poly-
mers. The data are summarized in Table 1, where Mp
is the mass at the peak maximum of the strongest signal
in the MALDI-TOF mass of each sample, except in the
case of the polymer obtained after 119 h (sample 3), for
Resu lts a n d Discu ssion
Syn th esis of Me-P iP r Ox-OH a n d Me-P iP r Ox-
NH2. Cationic ring-opening polymerization of iPrOx,
initiated with methyl p-tosylate, led to poly(2-isopropyl-
2-oxazoline) carrying a hydroxyl group at one end (Me-
PiPrOx-OH). For the polymerization to proceed in good
yield and without undesired side reactions, mild condi-
tions needed to be applied. We found that the polym-
erization of Me-PiPrOx-OH proceeded best, if done in
acetonitrile, at a temperature (42 °C) lower than the
temperature conditions (usually >70 °C) reported for
the polymerization of conventional poly(2-alkyl-2-ox-
azolines) (POx).3,20 Under these conditions, the polym-
erization had to be left to proceed for lengths of time
up to 506 h, but no noticeable side reactions occurred.
From the GPC trace shown in Figure 1, it was ascer-
tained that the number-average molecular weight (Mn)
and the molecular weight distribution (MWD) were
consistent with a living polymerization process: the
polydispersity index was low (1.02), and the experimen-
tal Mn value was close to the value predicted from the
initial monomer/initiator ratio (Mn,calc ) 10 000).

Macromolecules, Vol. 37, No. 18, 2004
Poly(2-isopropyl-2-oxazolines) 6789
Ta ble 1. An a lytica l Da ta As Der ived fr om th e Resp ective MALDI-TOF -MS a lon g w ith th e Resu lts fr om th e GP C
Mea su r em en ts for Com p a r ison (MWD ) Mw /Mn ; Molecu la r Weigh t Distr ibu tion )
MALDI-TOF-MS
GPC
b
b
c
c
polymer
∆Mtheor
∆Mexp
RMtheor
RMexp
Mtheor
[g/mol]
Mn
[g/mol]
Mw
[g/mol]
Mp
[g/mol]
Mn
[g/mol]
Mw
[g/mol]
samplesa
[g/mol]
[g/mol]
[g/mol]
[g/mol]
MWD
MWD
1 (78 h)
2 (95 h)
113.158
113.158
113.19
113.19
113.07
113.16
113.14
113.15
55.032
38.983
24.957
38.983
55.032
38.983
53.912d
39.07e
4015.562
4678.461
5456.541
5470.567
5486.616
6262.673
3800
4400
4000
4600
4016.95
4679.86
5457.72
5471.68
5487.95
1.04
1.05
3600
4300
3800
4400
1.03
1.03
29.181f
38.887e
57.23d
3 (119 h)
113.158
113.158
5400
6200
5700
6400
1.05
1.03
5100
5200
1.03
4 (135 h)
5 (506 h)
39.423e
5900
9700
6000
9900
1.03
1.02
a
b
Taken at different times during the process of polymerization. ∆M ) mass of monomer unit. c RM ) residual mass, Mtheor ) calculated
mass with a degree of polymerization nearest to the measured value (Mtheor ) ∆Mtheorn + RMtheor for the species given in footnotes d-f.
Me-[iPrOx]n-OH + Na+. e Me-[iPrOx]n-OH + Li+. f H-[iPrOx]n-OH + Li+, n ) DP).
d
which the second and third strongest signals were
included as well. The mass of sample 3 was about Mn,MS
) 5400 (m/z) (Mw,MS/Mn,MS ) 1.05), a value in close
agreement with the GPC results. This coincidence of
GPC and MS derived mass values was observed for the
other polymers as well. The mass difference in the series
of signals registered by MALDI-TOF mass of sample
3 (m/z 113.158, Figure 2b) agrees well with the mass of
the polymer repeating unit, confirming that the ions
form a homologue series originating from end-function-
alized polymers. The theoretical mass of each Me-
PiPrOx-OH is expressed by the following equation:
Mtheor ) ∆Mtheorn + RMtheor
where Mtheor is the calculated mass of a polymer of
degree of polymerization nearest to the measured value,
∆Mtheor is the mass of the monomer unit, and RMtheor
represents the calculated residual mass.
The most intense signal can be assigned to the lithium
F igu r e 3. MALDI-TOF mass spectra of poly(2-isopropyl-2-
oxazoline) having (a) hydroxyl groups (Me-PiPrOx-OH), (b)
adduct of Me-PiPrOx-OH, while the second most intense
signal is due to the sodium adduct of Me-PiPrOx-OH.
The occurrence of weak signals to the left side of the
lithium adduct signal in the spectrum of the Me-PiPrOx-
OH (H-[iPrOx]n-OH + Li+) reflects the presence of a
small fraction of polymer chain initiated by p-toluene-
sulfonic acid (H+), formed by inadvertent hydrolysis of
the methyl p-tosylate initiator.
phthalimide groups (Me-PiPrOx-PI), and (c) primary amino
groups (Me-PiPrOx-NH2) at ω-terminal ends.
imide groups at ω-terminals calculated to ∆m/z )
-130.096 (Table 2). Differences between the theoretical
and experimental mass (Mtheor - Mexp) are also very
small (<2.0 amu, Table 2).
There has been an argument that the coexistence of
ester-amines with hydroxyl terminals is possible unless
the hydrolysis of an oxazolinium salt is conducted under
the relatively high temperature condition even for a long
time.3 However, we have selected the mild conditions
for avoiding the inadvertent side reaction of the dimer-
ization of two oxazolinium living ends as well as the
formation of undesirable proton initiated polymers,
inclined to occur easily under the high-temperature
conditions (>60 °C) just in the case of the polymerization
of iPrOx. Note that neither the 1H NMR nor the 13C
NMR spectrum of Me-PiPrOx-OH (Figures S3 and S5a
in the Supporting Information) showed any undesirable
signals derived from ester-amines in our system which
terminated with methanolic NaOH (1 M) at room
temperature. Moreover, the high conversion efficiency
(ca. 70%) into phthalimide terminals and functionality
of amino groups (ca. 89%) also supported the reliability
about more favorable formation of hydroxyl terminals
than ester-amines.
We set out next to convert the terminal hydroxyl
groups of Me-PiPrOx-OH into primary amine groups
since these groups are more amenable to further con-
jugation with bioactive agents. This transformation was
performed in good yields via the Mitsunobu reaction,
which proceeds in two steps: (1) conversion of the
terminal hydroxyl into a phthalimide and (2) treatment
of the phthalimide-terminated polymer with hydrazine
to generate the free amine (Scheme 1). The success of
1
the reaction was assessed by H NMR analysis compar-
ing the integral ratio of signals and MALDI-TOF mass
analysis, comparing the molar mass of the strongest
signals in the spectra of the hydroxyl-terminated poly-
mer, Me-PiPrOx-OH, to that of the phthalimide termi-
nated polymer, Me-PiPrOx-PI (step 1). A similar com-
parison was also applied to the polymer obtained after
hydrazine treatment, Me-PiPrOx-NH2 (step 2) (Figure
3). Thus, the signals (n ) 43-46) in the two spectra (b
and c in Figure 3) can be assigned to homologues of the
sodium adducts of Me-PiPrOx-PI and Me-PiPrOx-NH2,
respectively (Table 2). We note that the difference of
mass value between Mexp in Me-PiPrOx-PI and Mexp in
Me-PiPrOx-NH2 is, within experimental error, equal to
the mass value corresponding to the loss of the phthal-
Syn th esis of Aceta l-P iP r Ox-OH. The preparation
of heterotelechelic PiPrOx was based on the cationic
ring-opening polymerization of 2-isopropyl-2-oxazoline
as well, but to attach a functional group at each end of
the polymer, it was necessary to initiate the polymeri-

6790 Park et al.
Macromolecules, Vol. 37, No. 18, 2004
Ta ble 2. Assign m en ts of th e Ch em ica l Com p osition for Most In ten sive Sign a ls in MALDI-TOF Ma ss Sp ectr a of
Me-P iP r Ox-OH, Me-P iP r Ox-P I, a n d Me-P iP r Ox-NH2
(a) Me-PiPrOx-OH + Na+
(b) Me-PiPrOx-PI + Na+
(c) Me-PiPrOx-NH2 + Na+
adduct
adduct
adduct
a
a
a
n
Mtheor
Mexp
[g/mol]
Mtheor
Mexp
-
Mtheor
Mexp
[g/mol]
Mtheor
Mexp
-
Mtheor
Mexp
[g/mol]
Mtheor
Mexp
-
∆[Mtheor(b) ∆[Mexp(b) ∆[Mtheor(c) ∆[Mexp(c)
- Mtheor(a)] - Mexp(a)] - Mtheor(b)] - Mexp(b)]
(DP) [g/mol]
[g/mol]
[g/mol]
43 4920.826 4922.38 -1.55 5049.940 5050.33 -0.39 4919.844 4921.77 -1.93
44 5033.984 5035.56 -1.58 5163.098 5164.37 -1.27 5033.002 5034.97 -1.97
45 5147.142 5149.80 -2.66 5276.256 5276.65 -0.39 5146.160 5148.15 -1.99
46 5260.300 5262.88 -2.58 5389.414 5390.67 -1.26 5259.318 5261.36 -2.04
+129.114
+129.114
+129.114
+129.114
+127.95
+128.81
+126.85
+127.79
-130.096
-130.096
-130.096
-130.096
-128.56
-129.4
-128.5
-129.31
a
Mtheor ) calculated mass with a degree of polymerization nearest to the measured value (Mtheor ) ∆Mtheorn + RMtheor for the following
species: Me-[iPrOx]n-OH + Na+, Me-[iPrOx]n-PI + Na+, Me-[iPrOx]n-NH2 + Na+, RM ) residual mass).
1
by analysis of the H NMR spectrum of acetal-PiPrOx-
OH (Figure 4), isolated after a polymerization time of
240 h, using spectral data gathered by analyzing spectra
of PiPrOx and 3,3-diethoxy-1-propyl tosylate used as
references. In this spectrum, the signals at 4.6 and 1.2
ppm can be ascribed to the methine and methyl protons
of the acetal group. Conversion of the ω-end group into
a hydroxyl group was confirmed by the absence, in the
1H NMR spectrum of acetal-PiPrOx-OH, of signals
characteristic of the tosylate phenyl protons (7.5-7.8
ppm). Further support to the assigned structure was
given by analysis of the 13C NMR spectrum of acetal-
PiPrOx-OH (Figure S5b in the Supporting Information).
A MALDI-TOF mass analysis was carried out using
an acetal-PiPrOx-OH sample obtained after a 160 h
polymerization (Mn,GPC ) 5200, Mw,GPC/Mn,GPC ) 1.25).
The difference in mass of the signals in the major mass
agrees well with the expected value series (m/z 113.158,
Figure 5). The experimental value of acetal-P iPrOx-OH
(Li+ adduct) is in agreement with the theoretical values,
defined as Mtheor ) ∆Mtheor(113.158)n + RMtheor(131.197
+ 17.008 + 6.941), indicating near quantitative incor-
poration of the acetal and hydroxyl groups on the
PiPrOx chain ends. The occurrence of small signals with
a mass difference of m/z ) 130.01 may reflect the
presence of a small fraction of polymer chains, initiated
by toluenesulfonic acid (H+) formed inadvertently by
hydrolysis of the acetal initiator.
Deter m in ation of th e Clou d P oin ts of Me-P iP r Ox-
OH a n d Aceta l-P iP r Ox-OH in Aqu eou s Solu tion .
We determined by turbidimetry the cloud points of
several Me-PiPrOx-OH samples of various molecular
weights (Mn ) 4300, Mw/Mn ) 1.03; Mn ) 7800, Mw/Mn
) 1.02; Mn ) 9700, Mw/Mn ) 1.02), dissolved in
phosphate-buffered saline (10 mM PBS containing 150
mM NaCl (pH 7.4)). They exhibit a remarkable depen-
F igu r e 4. 1H NMR spectrum of acetal-PiPrOx-OH in DMSO-
d6 at 80 °C.
zation with a different initiator, 3,3-diethoxy-1-propyl
tosylate (Scheme 2). This initiator was synthesized by
reaction of 3,3-diethoxy-1-propanol and p-toluenesulfo-
nyl chloride. This material proved to be an effective
cationic ring-opening polymerization initiator, as de-
termined from the GPC trace of a polymer obtained after
a polymerization time of 240 h (Figure S6 in the
Supporting Information). The GPC trace for this poly-
mer presents no shoulder, and the experimental Mn
value (9600) is close to the expected value based on the
initial monomer/initiator ratio (Mn,calc ) 10 000). More-
over, the molecular weight distribution (MWD) (1.1) is
consistent with the proposed cationic polymerization
mechanism. The structure of the polymer was confirmed
F igu r e 5. Enlarged detail of the MALDI-TOF mass spectrum of acetal-PiPrOx-OH sample obtained after 160 h.

Macromolecules, Vol. 37, No. 18, 2004
Poly(2-isopropyl-2-oxazolines) 6791
Con clu sion s
The synthesis of novel thermosensitive telechelic poly-
(2-isopropyl-2-oxazolines) (PiPrOx) bearing different
terminal ends (Me-PiPrOx-OH, Me-PiPrOx-NH2, acetal-
PiPrOx-OH) was successfully accomplished under op-
timum temperature conditions. The molecular weights
of all Me-PiPrOx-OH samples were close to the theoreti-
cal value derived from the monomer/initiator ratio, with
appreciably narrow molecular weight distributions,
suggesting that the polymerization proceeded in a
rather controlled manner without any noticeable side
reactions. Furthermore, the successful conversion of Me-
PiPrOx-OH into Me-PiPrOx-NH2 opens up paths toward
a variety of applications, such as the preparation of
thermosensitive bioconjugates and the synthesis of
biofunctional block copolymers, as they can serve to
initiate the polymerization of a second monomer. The
heterotelechelic acetal-PiPrOx-OH may also act as
utility as hetero-cross-linkers in the preparation of
thermosensitive conjugates of biomacromolecules, in-
cluding proteins and nucleic acid derivatives.
F igu r e 6. Transmittance changes of aqueous solutions as a
function of temperature under several polymer concentrations
(Me-PiPrOx-OH, Mn ) 9700, Mw/Mn ) 1.02; concentration )
0.1, 0.4, and 1.0 wt %; 10 mM PBS (pH ) 7.4) containing 150
mM NaCl (rate: 0.2 deg/min; wavelength: 500 nm)).
Ack n ow led gm en t. This work was financially sup-
ported by Special Coordination Funds for Science and
Technology from the Ministry of Education, Culture,
Sports, Science and Technology of J apan (MEXT) as well
as by the Core Research for Evolution of Science and
Technology (CREST), J apan Science and Technology
Corp. (J ST). We thank Dr. Woo-Dong J ang for invalu-
able suggestions.
Su p p or tin g In for m a tion Ava ila ble: Experimental pro-
cedure for the synthesis of 2-isopropyl-2-oxazoline and its
characterization via 1H, 13C NMR spectra,1H NMR spectra of
Me-PiPrOx-OH and 3,3-diethoxy-1-propyl tosylate, 13C NMR
spectra of Me-PiPrOx-OH and acetal-PiPrOx-OH, and gel
permeation chromatogram of acetal-PiPrOx-OH. This material
is available free of charge via the Internet at http://pubs.ac-
s.org.
F igu r e 7. Relationship between cloud point (Tcp) and polymer
concentration for three Me-PiPrOx-OH samples having dif-
ferent molecular weights (Mn ) 4300 (2), Mw/Mn ) 1.03; Mn )
7800 ([), Mw/Mn ) 1.02; Mn ) 9700 (9), Mw/Mn ) 1.02; 10 mM
PBS (pH ) 7.4) containing 150 mM NaCl (rate: 0.2 deg/min;
wavelength: 500 nm)).
Refer en ces a n d Notes
dence on the solution concentration. For example, in the
case of the polymer of Mn ) 9700 and Mw/Mn ) 1.02,
the cloud point of its aqueous solution decreases from
39.8 to 37.3 °C, as the concentration increases from 0.1
to 1.0 wt %. The generality of this trend was confirmed
by recording the concentration dependence of several
poly(oxazolines) (Figure 6). For all polymer solutions,
the cloud point tends to decrease with increasing
concentration in the low concentration domain (0.1 to
∼0.5 wt %) and to level off for solutions of higher
concentration (Figure 7). This tendency becomes less
pronounced as the molecular weight of the polymer
increases. It is interesting to note that, in all cases, the
cloud points of the poly(oxazolines) dissolved in phos-
phate-buffered saline remain slightly higher than the
physiological temperature, opening up the possibility of
controlling the temperature response of polyoxazoline
aqueous solutions around physiological conditions via
subtle changes in polymer molecular weight. Note that
the cloud point (37.5 °C) of a 1.0 wt % solution of acetal-
PiPrOx-OH (Mn ) 9600, Mw/Mn ) 1.15) is nearly
identical to the cloud point of a Me-PiPrOx-OH (Mn )
9700, Mw/Mn ) 1.02) solution measured under identical
conditions (Figure 6), confirming that molecular weight
is the determinant factor dictating the thermal response
of poly(oxazolines).
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