The effect of alkyl side groups on the secondary structure and crystallization of poly(ethylene glycol)-block-polypeptide copolymers

Article abstract of DOI:10.1016/j.polymer.2013.02.038

The solid state and melt nanoscale structures of a series of poly(ethylene glycol)-block-poly(γ-alkyl-l-glutamate)s (PEG-b-PALG) copolymers bearing different alkyl side groups, i.e., n-butyl, n-hexyl, n-octyl, and n-dodecyl, have been investigated in the present work. An interesting effect of alkyl side groups on the secondary conformation of polypeptide block, crystallization of PEG segments and morphology of the copolymers was observed. With increasing the length of alkyl side groups or raising the temperature, the predominant secondary structure gradually changed from α-helix to β-sheet conformation. Differential scanning calorimetry, wide-angle X-ray diffraction analysis and polarized optical microscopy indicated that the crystallization and morphology of PEG segments were markedly restricted by the polypeptide block. Therefore, the alkyl side groups not only exhibited significant influence on the conformation of polypeptide block, but also affected the crystallization of PEG segments. The secondary structure, crystallization and spherulite morphology of these synthetic biomaterials, which were significantly influenced by alkyl side groups, may play an important role in their physicochemical properties, such as self-assembly, as well as in some biomedical applications.

Full text of DOI:10.1016/j.polymer.2013.02.038

Polymer 54 (2013) 2466e2472
Contents lists available at SciVerse ScienceDirect
Polymer
journal homepage: www.elsevier.com/locate/polymer
The effect of alkyl side groups on the secondary structure and crystallization
of poly(ethylene glycol)-block-polypeptide copolymers
,
,
,
a
a
,
a
Kaixuan Ren a b, Yilong Cheng a b, Chaoliang He a , Chunsheng Xiao , Gao Li
*
**
, Xuesi Chen
a Key Laboratory of Polymer Ecomaterials, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, PR China
b Graduate University of Chinese Academy of Sciences, Beijing 100039, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
The solid state and melt nanoscale structures of a series of poly(ethylene glycol)-block-poly(g-alkyl-L-
Received 28 October 2012
Received in revised form
13 February 2013
Accepted 20 February 2013
Available online 27 February 2013
glutamate)s (PEG-b-PALG) copolymers bearing different alkyl side groups, i.e., n-butyl, n-hexyl, n-octyl,
and n-dodecyl, have been investigated in the present work. An interesting effect of alkyl side groups on
the secondary conformation of polypeptide block, crystallization of PEG segments and morphology of the
copolymers was observed. With increasing the length of alkyl side groups or raising the temperature, the
predominant secondary structure gradually changed from a-helix to b-sheet conformation. Differential
scanning calorimetry, wide-angle X-ray diffraction analysis and polarized optical microscopy indicated
that the crystallization and morphology of PEG segments were markedly restricted by the polypeptide
block. Therefore, the alkyl side groups not only exhibited significant influence on the conformation of
polypeptide block, but also affected the crystallization of PEG segments. The secondary structure, crys-
tallization and spherulite morphology of these synthetic biomaterials, which were significantly influ-
enced by alkyl side groups, may play an important role in their physicochemical properties, such as self-
assembly, as well as in some biomedical applications.
Keywords:
Secondary structure
Crystallization
Polypeptide
Ó 2013 Elsevier Ltd. All rights reserved.
1. Introduction
were greatly determined by the length of the side chains [16e21].
The conformation changes, side chain crystallization and thermal
Natural proteins and synthetic polypeptides have drawn
increasing attention as building blocks for construction of nano-
materials and devices for biomedical applications [1e6]. The
interest in peptides and proteins is driven by their excellent
biocompatibility, biodegradability, and especially hierarchical or-
dered structures (e.g., secondary structure, tertiary and quaternary
superstructures) [7]. The conjugation of peptides to synthetic
polymers could be a versatile strategy to fabricate novel materials
with hybrid physicochemical properties [8e10]. Short peptides
attached to synthetic polymers have been shown to be capable of
guiding the copolymers in forming superstructures, and the self-
assembly of the peptides can be influenced by synthetic polymers.
transitions of PALGs have revealed that they could form either
crystalline or amorphous domains depending on the length of the
side chains and their organization. Shao and co-workers have
investigated the surface characteristics of PALG LangmuireBlodgett
films [22,23]. Protein adsorption and platelets adhesion on these
LangmuireBlodgett films could be attributed to the specific effect
of different alkyl groups on hydrophilicity/hydrophobicity of the
polyglutamate surfaces. Alkyl side chains also have significant in-
fluence on thermodynamic stability and biocompatibility of these
polypeptides [16]. Another interesting property of PALGs is the
coexistence of two structural units: rod-like backbones and flexible
side chains, which create a hydrophobic semi-rigid rod. The former
ensures rigidity of polymer structure, and the latter gives them
solubility and processability [11]. Hence, they may form liquid
crystals at temperatures above the melting point since the molten
alkyl side chains could be regarded as a solvent.
Among various synthetic polypeptides, poly(g-alkyl-L-gluta-
mate)s (PALGs) containing various pendant alkyl groups have
attracted considerable attention due to their unusual physico-
chemical properties [11e15]. The pioneering work made by Wata-
nabe et al. revealed that the physicochemical properties of PALGs
However, although lots of work has been focused on the PALG
homopolymers, only limited attention has been devoted to the
bulk properties of conjugates of PALG and synthetic polymers.
Therefore, increasing efforts have been made on the block co-
polymers comprising polypeptides and other biocompatible syn-
thetic polymers, such as biodegradable aliphatic polyesters and
* Corresponding author. Tel./fax: þ86 431 85262116.
** Corresponding author. Tel./fax: þ86 431 85262667.
E-mail addresses: clhe@ciac.jl.cn (C. He), ligao@ciac.jl.cn (G. Li).
0032-3861/$ e see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.polymer.2013.02.038

K. Ren et al. / Polymer 54 (2013) 2466e2472
2467
poly(ethylene glycol) (PEG). PEG is a nonionic hydrophilic polymer
that exhibits excellent anti-fouling properties. PEG also crystallizes
over different length scales: from the monoclinic unit cell to
lamellar stacks separated by amorphous chains and to the
macroscopic superstructures known as spherulites [10,17e24]. In
the present work, various poly(ethylene glycol)-block-poly-
(
g
-alkyl-
alkyl side groups were synthesized by the ring-opening poly-
merization (ROP) of -alkyl- -glutamate N-carboxyanhydrides
L-glutamate)s (PEG-b-PALG) copolymers with different
g
L
(ALG NCA) using amino-terminated PEG as a macroinitiator. The
influence of length of alkyl side groups and temperature on the
secondary structure of the polypeptide block was investigated by
fourier transform infrared spectroscopy. Differential scanning
calorimetry and wide-angle X-ray diffraction analysis were
applied to reveal the crystallization behavior and thermo-induced
changes of the secondary structure of the block copolymers.
Crystallization morphology of PEG-b-PALG was observed by
polarized optical microscopy. The study of mutual influence of
each block within the PEG-polypeptide block copolymers may be
valuable for exploring the physicochemical and biological prop-
erties of these synthetic polymers.
Scheme 1. Synthesis route of PEG-b-PALG copolymers with different alkyl side groups.
2.3. Synthesis of poly(ethylene glycol)-block-poly(
g-alkyl-L-
glutamate)s
In order to compare the influence of different alkyl side groups
on the secondary structure and crystallization of the copolymers,
PEG-b-PALG copolymers with similar backbone structures and
different flexible side groups, i.e., n-butyl, n-hexyl, n-octyl, and
n-dodecyl, were synthesized by the ring-opening polymerization
2. Experimental section
2.1. Materials
(ROP) of
ing mPEG45-NH2 as a macroinitiator. A typical procedure for the
preparation of PEG-b-PALG copolymers was as follows: mPEG45
NH2 was dried by an azeotropic distillation with dry toluene.
Anhydrous DMF and ALG NCA were then added. The reaction
mixture was allowed to stir at 25 ꢁC for 3 days under a dry nitrogen
atmosphere. Then the polymer was purified by precipitation in
glacial diethyl ether, followed by filtration. The obtained product
was further washed twice with diethyl ether and dried under
g-alkyl-L-glutamate N-carboxyanhydrides (ALG NCA), us-
The
g-alkyl-L-glutamate N-carboxyanhydride (ALG NCA)
monomers were synthesized according to our previous work [25].
Polyethylene glycol monomethyl ether (mPEG, Mn ¼ 2000) was
purchased from Aldrich and used without further purification. The
amino-terminated poly(ethylene glycol) monomethyl ether
(denoted as mPEG-NH2) was synthesized according to the literature
procedure [26]. Tetrahydrofuran (THF) and toluene were refluxed
with sodium and distilled under nitrogen prior to use. N,N-dime-
thylformamide (DMF) was stored over calcium hydride (CaH2) and
purified by vacuum distillation. All the other reagents and solvents
were of analytical grade and used as obtained.
-
vacuum. Poly(ethylene glycol)-block-poly(
(PEG-b-PBLG), poly(ethylene glycol)-block-poly(
mate) (PEG-b-PHLG), poly(ethylene glycol)-block-poly(
glutamate) (PEG-b-POLG), and poly(ethylene glycol)-block-poly
-dodecyl- -glutamate) (PEG-b-PDLG) were synthesized with the
g
-butyl-
-hexyl-
-octyl-
L
-glutamate)
g
L-gluta-
g
L-
(g
L
2.2. Characterization
yields of 72, 69, 64, and 68%, respectively.
1H NMR spectra of PEG-b-PALG copolymers were recorded on a
Bruker AV 400 NMR spectrometer in deuterated trifluoroacetic acid
(CF3COOD) with tetramethylsilane as internal standard. The mo-
lecular weights and polydispersity indexs (PDI) were determined
by gel permeation chromatography (GPC) using a series of liner
Tskgel Super columns (AW3000 and AW5000) and Waters 515
HPLC pump with OPTILAB DSP Interferometric Refractometer
(Wyatt Technology) as the detector. The eluent was DMF containing
0.01 M LiBr at a flow rate of 1.0 mL minꢀ1 at 50 ꢁC. Monodispersed
polystyrene standards purchased from Waters Co. with a molecular
weight ranging from 1790 to 2.0 ꢂ 105 were used to generate the
calibration curve. Fourier-transform infrared (FTIR) spectra were
recorded on a Bio-Rad Win-IR instrument using a potassium bro-
mide (KBr) wafer method. To analyze their secondary structure
quantitatively, FTIR spectra in the amide I band region of 1600e
1700 cmꢀ1 were recorded for a solid film by casting a 5.0 wt% co-
polymers solution in CHCl3. The deconvoluted spectra were fitted
with GaussianeLorentzian sum function (20% Gaussian and 80%
Lorentzian) using XPSPEAK software 4.1. Temperature-dependent
FTIR spectra were investigated as a function of temperature in a
range of 30e140 ꢁC by an increment of 10 ꢁC per step. The samples
were prepared by casting a 5.0 wt% polymer solution in CHCl3 on a
KBr plate in a temperature-controlled chamber and were equili-
brated for 20 min at each temperature.
Fig. 1. 1H NMR spectra of (A) PEG45-b-PBLG8, (B) PEG45-b-PHLG8, (C) PEG45-b-POLG8,
and (D) PEG45-b-PDLG8 in CF3COOD.

2468
K. Ren et al. / Polymer 54 (2013) 2466e2472
Table 1
Characteristics of PEG-b-PALG copolymers.
DPa
Mn
PDIb
a
Polymers
Feeding molar ratio of
NCA/mPEG45-NH2
PEG45-b-PBLG8
PEG45-b-PHLG8
PEG45-b-POLG8
PEG45-b-PDLG8
10
10
11
12
8
8
8
8
3500
3700
3900
4400
1.1
1.2
1.1
1.3
a
Determined by 1H NMR in CF3COOD.
Determined by GPC. The eluent was DMF.
b
2.4. Differential scanning calorimetry (DSC)
The thermal properties of PEG-b-PALG copolymers were
examined by differential scanning calorimeter (Q100, TA In-
struments, USA) under nitrogen atmosphere. To ensure reliability
of the data obtained, heat flow and temperature were calibrated
with standard indium (Tm ¼ 156.6 ꢁC and
D
H ¼ 28.5 J/g). Mea-
Fig. 3. Deconvolution of the amide I band in FTIR spectra of PEG45-b-PBLG8.
surements during the first heating from ꢀ50 to 140 ꢁC and then the
first cooling from 140 to ꢀ50 ꢁC as well as the second heating
from ꢀ50 to 140 ꢁC at 10 ꢁC/min were performed. The crystallinity
was calculated based on the melting enthalpies according to the
following equation [7,27e29]:
Zeiss, Germany) equipped with a video CCD camera. The samples
were prepared by casting a 1 wt% polymer solution in dichloro-
methane on a clean cover glass and then airing at 25 ꢁC for
30 min.
À Á
Xm
%
¼
D
Hm=
D
Hmꢁ ꢂ 100%
3. Results and discussion
where
DHm (melting enthalpy) is the enthalpy value of the PEG
crystallites in PEG-b-PALG copolymers obtained from the second
3.1. Synthesis and characterization of the block copolymers
heating curves, and
the PEG crystals which have infinite thickness (
[30,31].
D
Hmꢁ is the theoretical enthalpy value for
D
Hmꢁ ¼ 208 J/g)
Ring-opening polymerization (ROP) of NCA monomers is a
common approach to prepare polypeptide-based copolymers. The
most used initiators for ROP of NCA monomers are primary amines
owing to their ‘‘living’’ polymerization characteristics and better
control of the degree of polymerization by varying the NCA/amine
feed ratio [25,32]. In this study, ALG NCA monomers with different
alkyl groups, including n-butyl, n-hexyl, n-octyl and n-dodecyl,
were synthesized by Fuchs-Farthing method, and PEG-b-PALG co-
polymers were then prepared by ROP of ALG NCA monomers with
mPEG45-NH2 as a macroinitiator (Scheme 1). The typical 1H NMR
spectra of PEG-b-PALG copolymers are shown in Fig. 1. The degree
of polymerization (DP) of the polypeptide block was calculated by
comparing the integration of methylene peak of PEG (eCH2CH2Oe,
b) at 3.6e3.8 ppm with that of the methylene peak of glutamate
units (eCH2CH2C(O)e, d) at about 2.1 ppm, and the results are
summarized in Table 1. GPC results showed that the obtained co-
polymers had relatively narrow molecular weight distributions in
the range of 1.1e1.3, indicating that PEG-b-PALG block copolymers
were well prepared.
2.5. Temperature-dependent wide-angle X-ray diffraction (WAXD)
Temperature-dependent WAXD measurements were carried
out on a Bruker D8 Advance X-ray diffractometer with a Ni-filtered
Cu K
a
radiation (
l
¼ 0.154 nm). The selective voltage and current
were 40 kV and 200 mA, respectively. The scattering angle (2
q
)
ranged from 2ꢁ to 45ꢁ at a rate of 2ꢁ
2q/min. A typical heating
process was performed from 30 ꢁC to 140 ꢁC at a rate of 10 ꢁC/min.
WAXD measurements were performed at 30, 35, 40, 45, 50, 55, 60,
70, 100 and 140 ꢁC, respectively.
2.6. Polarized optical microscopy (POM)
Crystal morphology of PEG-b-PALG copolymers was observed
by polarized optical microscope (Zeiss Axio Imager A2m, Carl
3.2. Secondary structure of the block copolymers
The secondary structure of PEG-b-PALG copolymers in solid
state was examined using FTIR spectroscopy. It is well-known that
the secondary structure of polypeptides could be identified by the
peak locations of the amide I band (C]O stretching) and the amide
II band (CeN stretching coupled with NeH bending) [33e36]. For a
typical a-helical structure, the absorbance peak of the amide I band
Table 2
Percentage content of secondary structure of the PEG-b-PALG copolymers.
Polymers
a
-Helix (%)
b-Sheet (%)
Random coil (%)
Others (%)
PEG45-b-PBLG8
PEG45-b-PHLG8
PEG45-b-POLG8
PEG45-b-PDLG8
57
32
24
26
27
47
53
58
6
12
12
11
10
9
11
5
Fig. 2. FTIR spectra of (A) PEG45-b-PBLG8, (B) PEG45-b-PHLG8, (C) PEG45-b-POLG8, (D)
PEG45-b-PDLG8.

K. Ren et al. / Polymer 54 (2013) 2466e2472
2469
Table 3
quantify the contents of the secondary structures (Fig. 3) and the
results are listed in Table 2. As alkyl side groups changed from n-
Melting and crystallization properties of PEG45 and PEG-b-PALG copolymers.
Polymers
Tc/ꢁC
DHc/J/g
Tm/ꢁC
DHm/J/g
Xm/%
butyl to n-dodecyl, the fraction of
b-sheet conformation increased
from 27% to 58% while the content of
from 57% to 26%. Although the polypeptide segments of all the
block copolymers adopted a mixture of secondary structures, the
a
-helix structure decreased
PEG45
20.9
10.2
ꢀ0.7
ꢀ0.8
ꢀ5.2
142.2
57.9
52.7
44.1
39.5
53.7
46.9
42.2
38.9
37.7
151.4
61.0
60.7
52.8
52.2
74.1
29.3
29.2
25.4
25.1
PEG45-b-PBLG8
PEG45-b-PHLG8
PEG45-b-POLG8
PEG45-b-PDLG8
predominant secondary structure gradually changed from
a-helix
to -sheet conformation with the increase in the length of alkyl side
b
Tc denotes the crystallization temperature of PEG segments in the cooling run. Tm
denotes the maximal melting temperature of PEG segments in the second heating
chain. The reason may be attributed to the fact that the polypeptide
block with longer side chains preferred to induce intermolecular
run.
DHc denotes the crystallization enthalpy of PEG segments in the cooling run.
hydrogen bonding in
intramolecular hydrogen bonding in
b
-sheet conformation rather than form
-helix structure due to steric
D
Hm denotes the melting enthalpy of PEG segments in the second heating run. Xm
denotes the degree of crystallinity of PEG segments according to the melting
enthalpy in the second heating run.
a
hindrance [37]. The results of FTIR spectroscopy demonstrated that
the side groups have notable influence on the conformation of the
PEG-polypeptide block copolymers. It’s worth mentioning that the
polypeptide blocks within the PEG-b-PALG copolymers are not long
enough to form a stable helical structure, which has stronger
is at w1650 cmꢀ1 and the amide II band is at w1550 cmꢀ1. The
absorbance peak of the amide I band at w1630 cmꢀ1 and the amide
II band at w1530 cmꢀ1 is identified for a
b-sheet conformation
dependence on chain length than
Therefore, in this work, the short chain length of the polypeptide
blocks has more influence on helical structure than -sheet.
b-sheet conformation [3,34].
[25,35]. As shown in Fig. 2, the strong amide I absorption band at
1628 cmꢀ1 and the amide II band at 1530 cmꢀ1 were observed,
b
indicating the presence of a parallel b-sheet conformation in the
polypeptide blocks of all copolymers. On the other hand, the amide
I absorption band at 1659 cmꢀ1 and the amide II band at 1548 cmꢀ1
3.3. Thermal and crystallization behaviors of the block copolymers
suggested the co-existence of an
weaker amide I band at 1694 cmꢀ1 was ascribed to an anti-parallel
-sheet conformation. Meanwhile, FTIR spectra were used to
a-helix structure. In addition, the
DSC measurements were performed to characterize the thermal
behavior and supramolecular organization of the PEG-polypeptide
block copolymers in solid state [38e40]. As listed in Table 3 and
Fig. 4, the crystallization temperature (Tc), melting temperature
b
(Tm), crystallization enthalpy (DHc) and melting enthalpy (DHm) of
the PEG segments within the copolymers were markedly lower
than that of the PEG homopolymer (Mn ¼ 2000, denoted as PEG45).
Notably, the Tc of PEG45-b-PHLG8, PEG45-b-POLG8, and PEG45-b-
PDLG8 were even below 0 ꢁC. The crystallization of the PEG seg-
ments within the block copolymers at large supercoolings implied a
restricted crystallization process [21,41]. As the length of the alkyl
side chain increased, obvious decreases in Tc and Tm of the co-
polymers were observed. The crystallinity (Xm) of all the block
copolymers were found to be less than 30%, which was significantly
lower than that of PEG45 (74.1%). This further confirmed that the
crystallization of the PEG segments within the copolymers was
markedly suppressed due to the confinement of PEG chains by the
polypeptide blocks. In addition, the PEG crystallinity of the block
copolymers decreased from 29.3% to 25.1% with increasing the
length of alkyl side groups (Table 3), which revealed that the longer
alkyl side chain exhibited more seriously inhibition on the crys-
tallinity of the PEG segments.
Fig. 4. DSC curves of the second heating process: (A) PEG45, (B) PEG45-b-PBLG8, (C)
PEG45-b-PHLG8, (D) PEG45-b-POLG8, (E) PEG45-b-PDLG8.
Fig. 5. WAXD patterns obtained at T ¼ 30 ꢁC. The results of PEG-b-PALG copolymers show Bragg peaks of PEG crystals with different crystallinities (A) and secondary structures of
polypeptide blocks (B).

2470
K. Ren et al. / Polymer 54 (2013) 2466e2472
Table 4
the alkyl side chain increased, the peak intensities gradually
Spacings of peaks observed in WAXD patterns.
declined. Therefore, WAXD results indicated that the crystallization
of PEG segments was seriously restricted by the polypeptide blocks,
which is consistent with the DSC analysis. Meanwhile, the Bragg
peak in Fig. 5B, reflecting the (020) plane of the orthogonal unit cell
Polymers
d1(Å)
d2(Å)
d3(Å)
d4(Å)
d5(Å)
PEG45
4.6
4.6
4.6
4.6
4.5
3.8
3.8
3.8
3.8
3.8
3.4
2.5
PEG45-b-PBuLG8
PEG45-b-PHLG8
PEG45-b-POLG8
PEG45-b-PDLG8
17.0
20.3
22.4
28.5
with the b
-sheet conformation, gradually shifted from 0.37 Åꢀ1 to
0.22 Åꢀ1. It was pointed out that this peak corresponded to the
predominant secondary structure in the oligopeptides with low
degrees of polymerization (<10) [17,44]. The intensity of this
diffraction peak indicated that the content of b-sheet conformation
increased with increasing the length of the alkyl side chains.
Meanwhile, the corresponding spacing d5 (Table 4), which repre-
sents the width of a lamellae organization with a b-sheet confor-
mation, changed from w1.7 nm to w2.85 nm [45].
Temperature-dependent WAXD pattern were then performed to
explore the thermo-induced changes of the PEG crystals and the
secondary structure of polypeptide blocks [46,47]. As shown in
Fig. 6A, both strong peaks ascribed to the PEG homopolymer dis-
appeared at above 50 ꢁC, indicating the melting of PEG crystals. In
contrast, the peaks for the PEG crystals within the copolymers
WAXD analysis is a useful method to analyze the crystalline
structure of PEG segments and secondary structure of polypeptide
blocks within the copolymers [42]. As shown in Fig. 5A, two strong
characteristic peaks of PEG crystals in a monoclinic unit cell at about
1.35 and 1.64 Åꢀ1 could be easily recognized at room temperature.
These peaks are listed in Table 4 as d1, which was indexed as the
(120) reflection and d2, which was indexed as an overlap of (112),
(032), (132), and (212) reflections, respectively [5,43]. Compared
with PEG45, obviously weaker crystalline diffraction peaks were
observed for the PEG-polypeptide block copolymers. As the length of
Fig. 6. Temperature-dependent WAXD patterns of samples obtained from 30 to 140 ꢁC (A) PEG45, (B) PEG45-b-PBLG8, (C) PEG45-b-PHLG8, (D) PEG45-b-POLG8, (E) PEG45-b-PDLG8.

K. Ren et al. / Polymer 54 (2013) 2466e2472
2471
Fig. 7. Temperature-dependent FTIR spectra of (A) PEG45-b-PBLG8, (B) PEG45-b-PHLG8.
disappeared at lower temperatures (Fig. 6(BeE)). At temperatures
above the Tm of PEG crystals, a broad amorphous halo centered at
amide I absorption band at 1659 cmꢀ1 and amide II band at
1548 cmꢀ1 reduced gradually. The results suggested that the con-
around 1.46 Åꢀ1(2
q
¼ 20.6ꢁ) originated from the amorphous PEG
tent of
reduced as the temperature rose from 30 ꢁC to 140 ꢁC. The increase
in temperature led to a secondary structure transition from -helix
to -sheet conformation.
b-sheet conformation increased while the a-helix content
and polypeptide chains was observed [17]. The peak intensity cor-
responding to
rose, implying the enhancement of the content of
mation. In addition, the retainment of the peaks for
conformation at all experimental temperatures also suggested that
the -sheet conformation of polypeptide blocks was not destroyed
b
-sheet conformation increased as the temperature
-sheet confor-
-sheet
a
b
b
b
3.4. Morphology of the block copolymers
b
during the crystallization and melting process of PEG segments.
Meanwhile, temperature-dependent FTIR spectra were recorded to
further investigate the thermostability of secondary structure of
polypeptide blocks within the copolymers. As shown in Fig. 7, the
intensities of amide I absorption band at 1628 cmꢀ1 and amide II
band at 1530 cmꢀ1 increased markedly when temperature was
raised from 30 ꢁC to 140 ꢁC. On the other hand, the intensities of
The crystal morphology of the PEG-b-PALG copolymer films was
revealed by polarized optical microscopy and the images are shown
in Fig. 8. The samples of PEG45-b-PBLG8, PEG45-b-PHLG8 and PEG45
-
b-POLG8 showed characteristic spherulite morphology from PEG
crystallization [21,22]. In contrast, no typical spherulitic crystalliza-
tion was observed for the sample of PEG45-b-PDLG8 bearing dodecyl
side groups, attributed to the severely restricted crystallization of
Fig. 8. POM micrographs (A) PEG45-b-PBLG8, (B) PEG45-b-PHLG8, (C) PEG45-b-POLG8, (D) PEG45-b-PDLG8. The images were taken from air-dried films at 25 ꢁC for 30 min.

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K. Ren et al. / Polymer 54 (2013) 2466e2472
[3] Chen C, Wang Z, Li Z. Biomacromolecules 2011;12(8):2859e63.
[4] Lin YC, Kuo SW. Polymer Chemistry 2012;3(1):162e71.
[5] Rosler A, Klok HA, Hamley IW, Castelletto V, Mykhaylyk OO. Biomacromolecules
2003;4(4):859e63.
PEG segments. Therefore, the results implied that the variation of
alkyl side groups displayed notable influence on the crystallization
morphology of the PEG-polypeptide block copolymers.
[6] Tang HY, Zhang DH. Biomacromolecules 2010;11(6):1585e92.
[7] Hua C, Dong CM, Wei Y. Biomacromolecules 2009;10(5):1140e8.
[8] Krysmann MJ, Funari SS, Canetta E, Hamley IW. Macromolecular Chemistry
and Physics 2008;209(9):883e9.
4. Conclusions
[9] Castelletto V, Newby GE, Merino DH, Hamley IW, Liu D, Noirez L. Polymer
Chemistry 2010;1(4):453e9.
[10] Nguyen MK, Lee DS. Macromolecular Bioscience 2010;10(6):563e79.
[11] Jeon S, Choo J, Sohn D, Lee SN. Polymer 2001;42(25):9915e20.
[12] Daly WH, Negulescu II, Russo PS, Poche DS. Acs Symposium Series 1992;493:
292e9.
[13] Gitsas A, Floudas G, Dietz M, Mondeshki M, Spiess HW, Wegner G. Macro-
molecules 2007;40(23):8311e22.
[14] Schmidt A, Lehmann S, Georgelin M, Katana G, Mathauer K, Kremer F, et al.
Macromolecules 1995;28(16):5487e97.
[15] Lee JL, Pearce EM, Kwei TK. Macromolecules 1997;30(22):6877e83.
[16] Luo CH, Zhao B, Li ZB. Polymer 2012;53(8):1725e32.
[17] Floudas G, Papadopoulos P, Klok HA, Vandermeulen GWM, Rodriguez-
Hernandez J. Macromolecules 2003;36(10):3673e83.
[18] Ge L, Guo R, Zhang X. The Journal of Physical Chemistry B 2009;113(7):1993e
2000.
[19] Wang T, Wang H, Li H, Gan Z, Yan S. Physical Chemistry Chemical Physics
2009;11(10):1619e27.
[20] Kuwabara K, Gan Z, Nakamura T, Abe H, Doi Y. Biomacromolecules 2002;3(2):
390e6.
[21] He C, Sun J, Ma J, Chen X, Jing X. Biomacromolecules 2006;7(12):3482e9.
[22] He C, Sun J, Zhao T, Hong Z, Zhuang X, Chen X, et al. Biomacromolecules
2005;7(1):252e8.
[23] Gan Z, Jim TF, Li M, Yuer Z, Wang S, Wu C. Macromolecules 1999;32(3):590e4.
[24] He LH, Xue R, Yang DB, Liu Y, Song R. Chinese Journal of Polymer Science
2009;27(04):501e10.
[25] Xiao C, Zhao C, He P, Tang Z, Chen X, Jing X. Macromolecular Rapid Com-
munications 2010;31(11):991e7.
[26] Ding J, Shi F, Xiao C, Lin L, Chen L, He C, et al. Polymer Chemistry 2011;2(12):
2857e64.
This paper discusses the secondary conformation, crystallization
and spherulite morphology of a series of poly(ethylene glycol)-block-
poly(g-alkyl-L-glutamate)s (PEG-b-PALG) copolymers with different
alkyl side groups including n-butyl, n-hexyl, n-octyl, and n-dodecyl.
Fourier transform infrared spectroscopy revealed that the poly-
peptide segments of all the block copolymers adopted a mixture of
secondary structures. As the alkyl side groups changed from n-butyl
to n-dodecyl, the fraction of
27% to 58% while the content of
to 26%, which indicated that the predominant secondary structure
gradually changed from -helix to -sheet conformation. The result
b-sheet conformation increased from
a-helix structure decreased from 57%
a
b
may be attributed to the fact that the polypeptide block with longer
side chains preferred to induce intermolecular hydrogen bonding in
b
-sheet conformation rather than form intramolecular hydrogen
bonding in -helix structure due to steric hindrance. Temperature-
dependent FTIR spectra indicated a secondary structure transition
from -helix to -sheet conformation with the increase in temper-
a
a
b
ature. As determined by differential scanning calorimetry, the
melting temperature and crystallinity of the PEG segments within
the block copolymers were much lower than that of PEG homopol-
ymer. Temperature-dependent wide-angle X-ray diffraction analysis
revealed that the content of b-sheet conformation of polypeptide
blocks increased and PEG crystals disappeared when the tempera-
ture rose. These results indicated that the crystallization of PEG
segments was significantly restricted by rigid polypeptide blocks.
[27] Langhe DS, Hiltner A, Baer E. Polymer 2011;52(25):5879e89.
[28] Liu JH, Mikhaylov IA, Zou JH, Osaka I, Masunov AE, McCullough RD, et al.
Polymer 2011;52(10):2302e9.
[29] Yoon JG, Zin WC, Kim JH. Polymer 2012;53(13):2744e50.
[30] Mai SM, Abbot A, Norton D, McKean R, Ryan AJ. Macromolecular Chemistry
and Physics 2009;210(10):840e51.
[31] Campbell C, Viras K, Richardson MJ, Masters AJ, Booth C. Makromolekulare
Chemie-Macromolecular Chemistry and Physics 1993;194(3):799e816.
[32] Kim MS, Dayananda K, Choi EK, Park HJ, Kim JS, Lee DS. Polymer 2009;50(10):
2252e7.
Meanwhile, the b-sheet conformation of polypeptide blocks was not
destroyed during the crystallization and melting process of PEG
segments. The macroscopic morphology observed from polarized
optical microscope confirmed the crystallization of PEG segments
within the PEG-b-PALG copolymers was suppressed. The investiga-
tion of the mutual influence of each block within the PEG-
polypeptide block copolymers on the secondary conformation and
crystallization may give valuable insights on the physicochemical
properties, e.g., self-assembly, as well as biological properties of
these synthetic biomacromolecules.
[33] Jackson M, Mantsch HH. Critical Reviews in Biochemistry and Molecular
Biology 1995;30(2):95e120.
ꢀ
ꢀ
[34] baginska K, Makowska J, Wiczk W, Kasprzykowski F, ChmurzynSki L. Journal
of Peptide Science 2008;14(3):283e9.
[35] Cheng Y, He C, Xiao C, Ding J, Zhuang X, Huang Y, et al. Biomacromolecules
2012;13(7):2053e9.
[36] Ruiz-Orta C, Fernandez-Blazquez JP, Pereira EJ, Alamo RG. Polymer 2011;52(13):
2856e68.
Acknowledgments
[37] Chang YC, Frank CW. Macromolecular Symposia 1997;118:641e6.
[38] Wang L, Wan D, Qiu J, Tang T. Polymer 2012;53(21):4737e57.
[39] van Drongelen M, van Erp TB, Peters GWM. Polymer 2012;53(21):4758e69.
[40] Zhuravlev E, Schmelzer JWP, Wunderlich B, Schick C. Polymer 2011;52(9):
1983e97.
[41] Müller AJ, Balsamo V, Arnal ML, Jakob T, Schmalz H, Abetz V. Macromolecules
2002;35(8):3048e58.
[42] Polec IA, Fujiwara T, Kanaya T, Deng C. Polymer 2012;53(16):3540e7.
[43] Takahash Y, Tadokoro H. Macromolecules 1973;6(5):672e5.
[44] Komoto T, Kim KY, Kawai T. Makromolekulare Chemie-Macromolecular
Chemistry and Physics 1978;179(2):373e85.
The authors are grateful for the financial support from the Na-
tional Natural Science Foundation of China (projects 51003103,
21174142, 51073154, 51233004 and 51021003), and the Ministry of
Science and Technology of China (International cooperation and
communication program 2011DFR51090).
References
[45] Caillol S, Lecommandoux S, Mingotaud AF, Schappacher M, Soum A, Bryson N,
et al. Macromolecules 2003;36(4):1118e24.
[46] Huang L, Kiyofuji G, Matsumoto J, Fukagawa Y, Gong C, Nojima S. Polymer
2012;53(25):5856e63.
[47] Abe H, Ishii N, Sato S, Tsuge T. Polymer 2012;53(14):3026e34.
[1] Hamley IW. Angewandte Chemie International Edition 2007;46(43):8128e47.
[2] He C, Zhuang X, Tang Z, Tian H, Chen X. Advanced Healthcare Materials
2012;1(1):48e78.