Self-assembled helical ribbon and tubes of alanine-based amphiphiles induced by two different formation mechanisms

Article abstract of DOI:10.1016/j.tet.2007.11.062

Three long chain alanine-based amphiphiles (1-3) possessing either a saturated long alkyl chain group or unsaturated groups as the self-assembling unit of a highly organized molecular architecture were synthesized. Their self-assembling properties were investigated using EF-TEM, SEM, CD, XRD, and FTIR. The d-form (1) and the l-form (2) enantiomers showed the opposite CD signals. Furthermore, electron micrographs of the self-assembled 1 and 2 exhibited right- and left-handed helical structures, respectively. The helical structures of amphiphiles 1 and 2 were developed into tubular structures by slow cooling. On the other hand, in the helical ribbon growth process, the helical pitch and ribbon width were changed in fast cooling process. The findings strongly imply that the helical ribbon growth of alanine-based amphiphiles can be induced by two different pathways. This mechanism, quite different from that observed for sugar- and cholesterol-based tubes, is a rare example for a tube formation process.

Full text of DOI:10.1016/j.tet.2007.11.062

Available online at www.sciencedirect.com
Tetrahedron 64 (2008) 1301e1308
www.elsevier.com/locate/tet
Self-assembled helical ribbon and tubes of alanine-based amphiphiles
induced by two different formation mechanisms
Soo Jin Lee a, Eunjeong Kim a, Moo Lyong Seo a, Youngkyu Do b, Young-A Lee c,
d,e
d,e,
Shim Sung Lee a, Jong Hwa Jung a, , Masaki Kogiso , Toshimi Shimizu
*
*
a Department of Chemistry, Research Institute of Natural Science, Gyeongsang National University,
900 Gazwa-dong, Jinju 660-701, Republic of Korea
b Department of Chemistry, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 305-701, Republic of Korea
c Department of Chemistry, Chonbuk National University, Jeonju 561-756, Republic of Korea
d SORST, Japan Science and Technology Agency, Tsukuba Central 5, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8565, Japan
e Nanoarchitectonics Research Center, National Institute of Advance Industrial Science and Technology,
Tsukuba Central 5, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8565, Japan
Received 21 September 2007; received in revised form 19 November 2007; accepted 20 November 2007
Available online 22 November 2007
Abstract
Three long chain alanine-based amphiphiles (1e3) possessing either a saturated long alkyl chain group or unsaturated groups as the
self-assembling unit of a highly organized molecular architecture were synthesized. Their self-assembling properties were investigated using
EF-TEM, SEM, CD, XRD, and FTIR. The D-form (1) and the L-form (2) enantiomers showed the opposite CD signals. Furthermore, electron
micrographs of the self-assembled 1 and 2 exhibited right- and left-handed helical structures, respectively. The helical structures of amphiphiles
1 and 2 were developed into tubular structures by slow cooling. On the other hand, in the helical ribbon growth process, the helical pitch and
ribbon width were changed in fast cooling process. The findings strongly imply that the helical ribbon growth of alanine-based amphiphiles can
be induced by two different pathways. This mechanism, quite different from that observed for sugar- and cholesterol-based tubes, is a rare
example for a tube formation process.
Ó 2007 Elsevier Ltd. All rights reserved.
Keywords: Alanines; Amphiphiles; Helical ribbons; Self-assembly
1. Introduction
Helical ribbon growth occurs by two different routes. It arise
from exclusive consequence happenings, which are the short-
ening of the helical pitch of the ribbon and the increase in
the tape width. Thus, one occurs with the former while the
latter does not change and vice versa for the other. For example,
sugar-based amphiphiles3,4 produced tubular structures by the
former process whereas cholesterol-based amphiphiles induced
tubule formation by the latter one.3,5 However, cases of helical
ribbon growth by widening of the tape with no concomitant
change in helical pitch (second route) are more common in
the literature than the former route. In almost all cases, the
importance of pitch or width changes in self-assembled amphi-
philes is not clear in the helical ribbon growth mechanism.
Helical ribbon structures in various systems of chiral organic
molecules, including diacetylenic lipids, surfactant, bile, and
The importance of chirality, or molecular handedness, has
long been recognized in many areas of chemistry and physics.
The interactions between chiral molecules change dramati-
cally when one molecule is replaced by its corresponding mir-
ror image structure. The advantage of chiral specificity has
been recognized by the pharmaceutical industry and is the
basis of a major effort to synthesize chiral drugs.1 Likewise,
in the area of liquid crystals, molecular chirality leads to the
formation of phases with long-range helical modulations,
such as the cholesteric phase.2
* Corresponding authors. Tel.: þ82 55 751 6027; fax: þ82 55 758 6027.
E-mail address: jonghwa@gnu.ac.kr (J.H. Jung).
0040-4020/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2007.11.062

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S.J. Lee et al. / Tetrahedron 64 (2008) 1301e1308
glutamates, were grown into tubules.6 In particular, the fabrica-
tion of tubular structures in phospho- and sugar-based lipids has
been investigated by the introduction of diacetylenic groups.7
However, the importance of diacetylenic units in the formation
of other tubular structures has not been studied. Only a few
examples of the mechanism of growth of tubules have appeared
in the literature.8 For examples, Kunitake and co-workers9
found that certain amphiphiles can form tubular structures by
way of a helical ribbon structure in aqueous solution, with
growth of the tubular structure occurring by an increase of
the width of the helical ribbon.
It is very well known that the various constituent molecules
play very important role in the formation of the helical ribbon
and tubular structures.3,6,8e10 Currently, most of the research
on the self-assembly of peptide amphiphiles were focused on
the preparation and physical properties of self-assembled nano-
fibers, helical ribbon, and tubular structures.11e13 However, the
mechanism of growth of these helical ribbon structures derived
from peptide-based amphiphiles was not conducted in any
them.9a,14 In this study, the growing mechanism and morpho-
logical control of the self-assembled peptide amphiphiles
were investigated.
absence of diacetylene units, was synthesized as a reference
compound to confirm the critical roles for the hydrophobic
interactions between alkyl chain groups in the self-assembly
formation. We herein report on the morphological control
and the self-assembly behaviors of the self-assembled 1 and
2, respectively. We also provide clear evidence regarding the
growth mechanism of the helical ribbon structure of peptide-
based amphiphiles by two different mechanisms. This is the
first study that examines the effect of the introduction of diac-
etylenic units on tubule growth that ultimately leads to the
formation of the diacetylenic peptide-based amphiphiles.
The self-assembled morphologies were strongly dependent
on the stereochemical enantiomer and unsaturated bonds in
the lipophilic region. Furthermore, on the basis of circular
dichroism and powder X-ray diffraction experiments, we pro-
vide evidence for the chiral packing structures of self-assem-
bling superstructures of alanine-based amphiphiles.
2. Results and discussion
2.1. Synthesis of alanine-based amphiphiles
With these objectives in mind, we have newly synthesized
a stereochemical series of alanine-based amphiphiles 1 and 2
as shown in Figure 1 and Scheme 1. Compound 3, in the
Two enantiomer alanine-based amphiphiles 1 and 2 were
synthesized as shown in Scheme 1. Coupling of 7 with
10,12-heptadecadiynoic acid 6 provides 4 in 70% yield. Simi-
lar reaction gives 5 with 75% yield. Then, treatment of 4 or 5
with NaOH gave desired 1 and 2 in 95 and 92% yield, respec-
tively. The products were purified using silica gel column
chromatography. Also, compound 3 possessing saturated
long alkyl chain group instead of the diacetylene unit was pre-
pared as a reference by similar methods to confirm the role of
the diacetylene unit (Supplementary data: Scheme S1).
O
HO
N
H
O
1: D-form
2: L-form
O
HO
N
2.2. Self-assembly and morphological observations
H
O
3: D-form
The self-assembled nanostructures of amphiphiles 1e3
were fabricated by two different methods as follows. Each al-
anine-based amphiphiles (1.0 mg) was dispersed into water
(10.0 mL) at temperatures above the corresponding melting
point (Tm) of the hydrated sample. In general, heating the mix-
ture at 95 ꢀC for 30 min was sufficient to obtain a homoge-
neous transparent solution. During the heating process, the
sample flasks were wrapped with Al foil to prevent the
polymerization.
Figure 1. Chemical structures of 1e3.
O
O
a)
NH2
NH2
HO
O
HCl
7 or 8
9 (D- form) or
10 (L-form)
O
HO
+
7 or 8
b)
The aqueous solutions were allowed to cool to room tem-
perature at two different rates, 5.0 and 0.5 ꢀC/min. Then, we
observed and compared the morphologies of the self-assem-
blies prepared with fast and slow cooling rates at same aging
step.
To precisely characterize the resultant morphologies and
their size dimensions, we observed the individual self-assem-
bled structures using energy-filtering transmission electron
microscopy (EF-TEM) and scanning electron microscopy
(SEM). In SEM and EF-TEM images of the self-assembled
1 prepared by cooling at 5.0 ꢀC/min in aqueous solution
exhibited a right-handed helical ribbon structure with a width
of 4e10 mm and 24 mm of helical pitch (Fig. 2a), with all the
6
O
O
N
H
O
4 or 5
c)
1: D-Form
2: L-Form
Scheme 1. Synthetic routes for amphiphiles 1 and 2. (a) SOCl2, EtOH, 86%;
(b) DCC, HOBt, EA, 95%; (c) NaOH, MeOH, H2O, 95%. DCC¼1,3-dicyclo-
hexylcarbodiimide, HOBt¼1-hydroxybenzotriazole, EA¼ethyl acetate.

S.J. Lee et al. / Tetrahedron 64 (2008) 1301e1308
1303
Figure 2. SEM images of self-assembled (a) 1 and (b) 2 prepared by fast cooling with 5.0 ꢀC/min. SEM images of self-assembled (c) 1 prepared by slow cooling
with 0.5 ꢀC/min [inset, TEM image]. (d) SEM image of the self-assembled 3 prepared by fast cooling.
helicity possessing the right-handed helical motif. The self-
assembled 2, in contrast, exhibited all left-handed helical ribbon
structure, 5e10 mm wide and with an 18 mm helical pitch
(Fig. 2b). These results indicate that the helicity of self-assem-
bled nanostructures is strongly influenced by the stereoiso-
meric alanine unit.
c)
b)
On the other hand, when self-assembled 1 was prepared by
slow cooling at a rate of 0.5 ꢀC/min, a helical ribbon morpho-
logy occurred, with an outer diameter of the intermediate mor-
phological construction equal to 260 nm and the morphology
of the final nanotubular structure exhibiting an inner diameter
of ca. 120 nm and a wall thickness of 70 nm (Fig. 2c). There-
fore, the cooling rate is an important factor in the production
of nanotubes through self-assembly. The helical pitch for the
tubular structure of self-assembled 1 prepared by slow cooling
has almost same value, indicating that the growth mechanism
for the self-assembled tubes 1 in slow cooling process is
different from that in fast cooling. Also, the similar result
was observed for the self-assembled 2 (Supplementary data:
Fig. S1).
To confirm the diacetylene unit effect in the formation of
helical structures, we introduced the saturated alkyl chain
group into the lipophilic region of 3 instead of diacetylene
unit of 1. The self-assembled 3 exhibits the linear and the
loosely helical structures with 50e200 nm of width
(Fig. 2d), further supporting the view that the morphology for-
mation of the alanine-based amphiphile depends strongly on
the nature of the diacetylene unit in the long alkyl chain group.
Photopolymerization of the self-assembled 1 and 2 by irra-
diation at 254 nm wavelength with 6 W UV lamp at a distance
of 10 cm for 30 min results in a red solution with an absorp-
tion maximum at 575 nm (Fig. 3). During irradiation process,
the self-assembled samples were maintained at 10 ꢀC to pre-
vent the undesired side reaction. The color change from
a)
400
600
Wavelength (nm)
800
Figure 3. Solid UVevis absorption spectra for self-assembled 1 (a) before and
(b) after irradiation for 15 min and (d) 30 min.
colorless to red means that the non-covalented self-assembled
samples were polymerized via the diacetinyl group. So, the
polymerized red product has a reinforced form.
By comparing the physical properties (solubility and color
change) of two compounds before and after polymerization,
we also confirmed that the polymerization occurred. For
example, the product after irradiation became insoluble in or-
ganic solvents such as DMSO, DMF, THF, CHCl3, CH3OH,
and toluene. The insoluble product was isolated and its mor-
phology was observed by SEM. The well-ordered helical
structures of self-assembled 1 and 2 are still maintained after
polymerization (Fig. S2). This finding is a rare example for the
polymerized helical ribbon structure. However, the molecular
weight of the polymerized helical ribbons could not be deter-
mined because of its insolubility in organic solvents such as
DMSO, DMF, THF, CHCl3, CH3OH, and toluene.

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S.J. Lee et al. / Tetrahedron 64 (2008) 1301e1308
2.3. A possible mechanism for the helical ribbon growth
SEM images indicate that the self-assembled 1 prepared by
fast cooling possesses the helical ribbon structure with
various pitches (10e24 mm) and with various ribbon widths
(4e10 mm) in the initial stage (Fig. 4). Once again, both pitch
and ribbon width values were changed during the process of
helical ribbon growth. This finding strongly implies that the
helical ribbon growth of alanine-based amphiphiles can be
induced by two different pathways (Fig. 5). This is a rare
example that examines the effect of the introduction of diace-
tylenic units on tubule growth that ultimately leads to the for-
mation of the diacetylenic peptide-based amphiphiles.
We demonstrated the growth mechanism of the self-assem-
blies based on sugar and cholesterol derivatives in water or
organic solvents. For example, the sugar-based lipid was in-
duced by the mechanism illustrated in Figure 5a,4a whereas
the cholesterol-based amphiphile was induced by that shown
in Figure 5b.5f However, these self-assemblies 1 and 2 differ
markedly from those observed with sugar- and cholesterol-
based helical ribbons, which are formed by the mechanisms
shown in Figure 5a or b. The helical ribbon growth process
in these alanine-based amphiphiles is very rare, perhaps being
influenced by formation conditions, cooling times, and driving
forces related to hydrophobicity and hydrogen bond forma-
tion. However, the main factors that determine the mechanism
of helical ribbon growth are not clear.
Figure 5. Representation of possible helical ribbon growth mechanisms of 1
and 2: (a) the helical ribbon is formed by a change in the pitch length; (b)
the helical ribbon is formed by growth in the width.
which indicates that the tubular structure is occurred by one
process. However, it is not clear whether the tubular structure
is induced by the mechanism shown in Figure 5a or b because
the growth rate of the tubular structure is too fast for the inter-
mediate to observe. Once again, the growth mechanism of the
helical ribbon structure of self-assembled 1 prepared by fast
cooling is quite different from that of the self-assembled 1
prepared by slow cooling.
On the other hand, the self-assembled tube 1 prepared by
slow cooling possesses a constant ribbon width (ca. 200 nm),
2.4. CD measurement
CD spectroscopy is very useful for ascertaining the chiral
molecular arrangement of self-assemblies.3e,6h For example,
recently, Schnur and our group have studied on the chiral
molecular architectures for self-assemblies of sugar or diace-
tylenic lipids by CD at different temperatures or solvents.3e,6h
In order to further test the effects of chirality, we have ob-
served the CD and UVevis spectra of the self-assembled 1e3
in water (see Fig. 6 and Fig. S3). In the CD spectra, lq¼0 for
the self-assembled 1 and 2 appeared at around 232 nm, which
are originated from the amide group of alanine moiety by
nep* transition.4a,15 On the other hand, the lmax value in the UV
absorption spectrum of 1 cannot be clearly observed due to the
strong water peak at around 200 nm, however, the shoulder
peak was observed at around 255 nm (Fig. S3). Thus it is
not clear that CD spectra of the self-assembled 1e3 were
induced by exciton coupling effect. However, the D-form (1)
and the L-form (2) enantiomers showed the opposite CD
signals as shown in Figure 6a and b, respectively. This result
indicates that the helical ribbon structures of the two enantio-
mers have chiral molecular architectures with the opposite
handedness, as would be expected. In addition, only weak
CD signal of the self-assembled 1 at temperatures above
Figure 4. SEM or TEM images of the self-assembled helical ribbons with vari-
ous pitches and various widths obtained from 1.

S.J. Lee et al. / Tetrahedron 64 (2008) 1301e1308
1305
30000
20000
10000
0
a)
b)
b)
c)
c)
e)
-10000
-20000
-30000
-40000
-50000
a)
d)
200
300
400
Wavelength (nm)
500
600
2
4
2- theta degree
6
Figure 6. CD spectra of the self-assembled (a) 1 and (b) 2 prepared by fast
cooling with 5.0 ꢀC/min at 25 ꢀC, (c) 1 at 70 ꢀC, (d) 1 prepared by slow cool-
ing with 0.5 ꢀC/min, and (e) 3 prepared by fast cooling with 5.0 ꢀC/min. The
Figure 7. Powder XRD patterns of (a) self-assembled 1 prepared by fast cool-
ing with 5.0 ꢀC/min, (b) self-assembled 1 prepared by slow cooling with
0.5 ꢀC/min, and (c) self-assembled 3 prepared by fast cooling with 5.0 ꢀC/min.
molar ellipticity [q] is given in deg cm2 dmolꢁ1
.
a phase transition temperature (70 ꢀC) was observed (Fig. 6c),
which is probably composed of the monomers or small aggre-
gates, such as micelles or vesicles. A strong CD signal reap-
peared when the self-assembled helical ribbon structure
formed again after several hours.
We also measured CD spectrum of the self-assembled 1
prepared by slow and fast cooling to demonstrate different chi-
ral molecular arrangements. The CD signal intensity of self-
assembled 1 prepared by slow cooling (Fig. 6d) was much
stronger than that by fast cooling. These findings indicate
that the tubular form possesses the relatively well-ordered
helical packing structure. On the other hand, the CD spectrum
of the self-assembled 3 in aqueous solution showed a much
weaker negative band (Fig. 6e) in comparison to 1 and 2, sug-
gesting that the self-assembled 3 forms disordered chiral pack-
ing structures.
mechanism. The d values correspond to the distance for regu-
larly arranged molecular packing. Particularly, if the self-
assemblies form bilayered structures with a same orientation,
the d value gives the relative distance for the interdigitated
hydrophobic interaction between alkyl chain groups, which
is related to the hydrophobic interaction. First, the molecular
lengths of three amphiphiles were calculated using CPK space
filling models and were determined to be highly similar. Sec-
ondly, X-ray diffraction patterns were measured. The d-spac-
ing value of crystalline 1 from the self-assembled structures
is shown in Figure 7. The small-angle diffraction patterns of
helical ribbons 1 and 2 prepared by fast cooling appeared at
4.02 nm (Fig. 7a and Fig. S4), whereas the self-assembled
tubes 1 and 2 prepared by slow cooling appeared at
4.29 nm. These values are smaller than twice the extended
molecular length of 1 or 2 (2.80 nm using the CPK space
filling model) but larger than the length of a single molecule.
These results strongly suggest that the self-assembled 1 and 2
form a bilayered structure with a relative small region interdig-
itated by hydrophobic interaction as shown in Figure 7, due to
the higher d value. Also, the hydrophobic interaction of the
self-assembled tube was relatively weaker than that of the
self-assembled helical ribbon.
2.5. Powder XRD measurement
We demonstrated the growth mechanism of the self-assem-
blies based on sugar and cholesterol derivatives in water or
organic solvents. For example, the sugar-based lipid was in-
duced by the mechanism illustrated in Figure 5a,4a whereas
the cholesterol-based amphiphile was induced by that shown
in Figure 5b.5f However, these self-assemblies 1 and 2 differ
markedly from that observed with sugar- and cholesterol-
based helical ribbons, which formed by the mechanisms in
Figure 5a or b. The helical ribbon growth process in these
alanine-based amphiphiles is very rare, perhaps being influ-
enced by formation conditions, cooling times, and driving
forces related to hydrophobicity and hydrogen bond forma-
tion. However, the main factors that determine the mechanism
of helical ribbon growth are not clear.
On the other hand, the microcrystalline solid 3 (one molec-
ular length: 2.84 nm) formed a bilayer structure with 3.69 nm,
supporting the idea that 3 maintains a much larger interdigi-
tated bilayer (Fig. 7c and Fig. S4) structure between the lipo-
philic regions than do those of 1 and 2. The finding indicates
that the relative strength of the hydrophobic interaction was
dependent on the formation of the helical structure.
2.6. IR measurement
Powder X-ray diffraction crystallography, a recently
reported technique for ascertaining the molecular packing of
self-assemblies,4a,10,16 was used to clarify the self-assembling
To confirm the morphological influences of the self-assem-
bled 1 and 2 prepared at different pH conditions, we observed

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S.J. Lee et al. / Tetrahedron 64 (2008) 1301e1308
can be easily controlled by the alanine units. Interestingly,
a)
b)
they form the tubular structure by slow cooling. However,
the saturated long chain alanine-based amphiphile revealed
the linear ribbon structure.
O
H
N
R
O
O
To the best of our knowledge, these results are a rare exam-
ple of a systematic study on the influence of the alanine unit as
a hydrophilic moiety and diacetylene unit in a hydrophobic
portion on self-assembled morphologies. Based on the CD
results, the self-assembled helical ribbon and nanotubular
structure showed a relatively stronger intensity than that of
the linear ribbon structure, indicating that the helical ribbon
and nanotube formed relatively well-ordered chiral packing
structures compared to the linear ribbon structure. Further-
more, in the XRD experiments, the relatively weak hydropho-
bic interactions of 1 and 2 formed with bilayered structures
between the lipophilic groups led to the formation of the heli-
cal ribbon and nanotubular structures.
c)
O
H
N
R
OH
O
3400
3200
3000
2800 1800 1600
Wavenumber (cm-1)
1400
1200
1000
Figure 8. IR spectra of solution 1 at (a) pH¼9, and of self-assembled 1
prepared at (b) pH¼7 and (c) pH¼4.
4. Experimental
IR spectra of the self-assembled 1 formed at pH¼4, 7, and 9,
respectively (Fig. 8). Clearly, the helical ribbon formed at
pH¼7 with strong hydrogen bonding between the amide group
and the highly organized, closely packed hydrocarbon chains
produced sharper IR bands relative to the monomer species
in aqueous solution at pH¼9. Prominent differences were
observed in a few regions. First, the helical ribbon 1 obtained
at pH¼7 displayed strong bands at 1701 and 1410 cmꢁ1 for
COOH and COOꢁ species, respectively. In contrast, the linear
fiber 1 obtained at pH¼4 showed only a single strong band at
1701 cmꢁ1 for COOH. An alkaline solution of 1 (pH¼9)
shows a band for only the ionized COOꢁ species at
1410 cmꢁ1, which showed no solidified morphology. The
results indicate that the intermolecular hydrogen bonding in-
teractions between the COOH and COOꢁ species display
a critical role to induce self-assembled superstructure of 1.
Secondly, amide function is characterized by three infrared
absorption bands corresponding to the NeH stretch
(nNHz3322 cmꢁ1), the C]O stretch called the Amide I
band (nCOz1638 cmꢁ1) and a combination of the NeH defor-
mation and the CeN stretch called the Amide II
(dNHþnCNz1566 cmꢁ1). The intensities and wavenumbers
of these bands are directly related to the hydrogen bond
strength of the system. In the case of a transition from an
associated amide to a free amide, the Amide I band is shifted
to higher wavenumbers and the Amide II band to higher wave-
numbers. Furthermore, the self-assembled 3 was confirmed by
the intermolecular hydrogen bonding interaction between
amide groups (Fig. S5).
4.1. General
1H and 13C NMR spectra were recorded with a DRX-500
(Bruker, 500 MHz) or an Avance-300 (Bruker, 300 MHz)
NMR spectrometer. Preparative column chromatography was
performed on silica gel. The chromatographic purity of the
intermediates was monitored by thin-layer chromatography
(Kiesel gel 60 F254, Merck). The compounds were visualized
by spraying the plates with 5% sulfuric acid in methanol and
then charring them on a hot plate. The molecular lengths were
estimated by molecular mechanics calculations performed
with the CONFLEX-MM2 force field as implemented in
CAChe, version 4.1.1 (Fujitsu Co. Ltd., Japan). Circular
dichroism (CD) spectra were measured on a JASCO J-715
spectrophotometer (cell diameter 1 mm).
4.1.1. TEM observations
The aqueous dispersions of the nanostructures
(0.1 mg mLꢁ1) were dripped onto an amorphous carbon grid
and excess water was blotted with filter paper. TEM was per-
formed with a Carl-Zeiss LEO912 instrument operated at
50 keV. Images were recorded on an imaging plate (Fuji Photo
Film Co. Ltd. FDL5000 system) with 20-eV energy windows
at 3000e250,000ꢂ and were digitally enlarged.
4.1.2. FTIR measurements
The FTIR spectra of the self-assembled nanostructures
were measured with a Fourier transform IR spectrometer
(JASCO FT-620) operated at 4 cmꢁ1 resolution with an unpo-
larized beam, attenuated total reflection (ATR) accessory
system (Diamond MIRacle, horizontal ATR accessory with
a diamond crystal prism, PIKE Technologies, USA) and a mer-
cury cadmium telluride (MCT) detector. Several drops of the
aqueous dispersions of the self-assembly were dripped onto
the prism and dried under nitrogen stream prior to
measurement.
3. Conclusions
The present study has demonstrated that D- and L-alanine-
based amphiphiles possessing a diacetylene unit in the lipho-
philic region form right- and left-handed helical ribbon
structures, respectively, depending on the stereochemical en-
antiomer. Thus, the self-assembled morphological helicity

S.J. Lee et al. / Tetrahedron 64 (2008) 1301e1308
1307
4.1.3. XRD measurements
saturated NaHSO4, and saturated NaHCO3. After drying
with MgSO4 the solvents were removed under reduced pres-
sure. This crude product was purified by column chromatogra-
phy (ethyl acetate/hexane, 1/2) to give the product (70%). Mp
The XRD of a freeze-dried sample was measured with
a Rigakudiffractometer (Type 4037) using graded d-space
elliptical side-by-side multilayer optics, monochromated Cu
Ka radiation (40 kV, 30 mA) and an imaging plate (R-Axis
IV). The typical exposure time was 10 min with an 150-mm
camera length. Freeze-dried samples from 1 to 3 were vacuum
dried to constant weight and then put into capillary tubes,
without being powdered.
1
81.3e82.0 ꢀC. H NMR (300 MHz, CDCl3, 25 ꢀC): d 5.82 (s,
1H, NH), 4.58 (q, 1H, J¼6 Hz), 4.17 (q, 2H, J¼7.2 Hz), 2.18
13
(t, 2H, J¼6.9 Hz), 2.05 (m, 4H), 1.54e1.30 (m, 22H), 0.91 (t,
3H, J¼7.2 Hz); C NMR (75 MHz, CDCl3, 25 ꢀC): d 176,
174, 71, 65, 60, 45, 32, 31e29, 21, 17, 15; IR (KBr):
~n¼3322, 2957, 2917, 2849, 1742, 1648, 1537, 1472, 1421,
1030, 951; MS (FAB) m/z 362 (MþH)þ. Anal. Calcd for
C23H45NO3: C, 73.09; H, 9.76; N, 3.87. Found C, 73.15; H,
10.01; N, 3.64.
4.1.4. CD measurements
Circular dichroism (CD) studies were performed on
a JASCO J-715 spectropolarimeter operating between 190
and 500 nm. The samples (5.0ꢂ10ꢁ3 M) were placed in wa-
ter-jacketed quartz cells with path lengths of 0.1 mm. Temper-
ature control was provided by a water circulator, which
provided a thermal stability of about 0.2 ꢀC. The spectrometer
4.1.9. Compound 5
Compound 5 was prepared by a method similar to that used
1
for 4 in 75% yield. Mp 80.8e82.0 ꢀC. H NMR (300 MHz,
was calibrated with ammonium-D-camphorsulfonate ([q]291
7910 deg cm2 dmolꢁ1) and D-pantonyllactone ([q]219¼ꢁ16,140
¼
CDCl3, 25 ꢀC): d 5.82 (s, 1H, NH), 4.58 (q, 1H, J¼6 Hz),
4.17 (q, 2H, J¼7.2 Hz), 2.18 (t, 2H, J¼6.9 Hz), 2.05 (m,
4H), 1.54e1.30 (m, 22H), 0.91 (t, 3H, J¼7.2 Hz); 13C NMR
(75 MHz, CDCl3, 25 ꢀC): d 176, 174, 71, 65, 60, 45, 32,
31e29, 21, 17, 15; IR (KBr): ~n¼3322, 2957, 2917, 2849,
1742, 1648, 1537, 1472, 1421, 1030, 951; MS (FAB) m/z
362 (MþH)þ; Anal. Calcd for C23H45NO3: C, 73.09; H,
9.76; N, 3.87. Found C, 73.01; H, 9.57; N, 3.63.
in water, [q]223¼ꢁ12,420 in methanol).
4.1.5. pH adjustment of the self-assembled 1e3
Aqueous dispersion (10 mL) was prepared by adding de-
gassed distilled water to a weighted powder of 1e3 and
succeeding sonication at 25 ꢀC. The pH values of aqueous dis-
persions were then adjusted with 0.1 M NaOH or 0.1 M HCl
solution. During the heating and irradiation processes, the
sample flask was wrapped with Al foil to prevent the
polymerization.
4.1.10. Compound 1
Compound 4 (120 mg, 0.33 mmol) was dissolved in meth-
anol (5 mL) and 1 M NaOH (1 mL, 1 mmol) was added. The
mixture was stirred at room temperature for 4 h and 1 M
HCl (2.5 mL, 2.5 mmol) was then added. The methanol was
removed under reduced pressure and the solution thus ob-
tained was cooled in an ice bath and acidified with 1 M HCl
(3 mL) with vigorous stirring and extracted with ethyl acetate
(50 mL). The organic layer was dried over MgSO4. Removal
of the organic solvent in vacuo afforded a white solid. This
crude product was purified by column chromatography (etha-
4.1.6. Compound 7
Thionyl chloride (12.5 mL) was added dropwise to stirred
and cooled (ꢁ10 ꢀC) ethanol (50 mL), followed by the addi-
tion of D-alanine (4.46 g, 50 mmol). The temperature of the
solution was increased and maintained at 40 ꢀC for 4 h. The
solvent was removed and crystallization from methanol/ethyl
ether yielded 4.65 g (86%). Mp 76e78 ꢀC.17 1H NMR
(300 MHz, DMSO-d6, 25 ꢀC): d 8.66 (s, 3H, NH3), 4.17 (q,
1H, J¼7.2 Hz), 4.00 (q, 2H, J¼7.2 Hz), 1.40 (d, 3H,
J¼7 Hz), 1.22 (t, 3H, J¼7.2 Hz).
1
nol/hexane) to give the product (95%). Mp 92.0e93.1 ꢀC. H
NMR (300 MHz, CDCl3, 25 ꢀC): d 8.01 (s, 1H), 4.45 (q, 1H,
J¼6 Hz), 2.24 (t, 2H, J¼6.9 Hz), 2.10 (m, 4 H), 1.58e1.35 (m,
19H), 0.93 (t, 3H, J¼7.2 Hz); 13C NMR (75 MHz, CDCl3,
25 ꢀC): d 175, 174, 74, 68, 31, 31e29, 20, 16, 15; IR (KBr):
~n¼3315, 2915, 2839, 1696, 1633, 1565, 1518, 1449, 1405,
1300, 1231, 1078, 910, 705, 631, 565, 533; MS (FAB) m/z
334 (MþH)þ. Anal. Calcd for C20H31NO3: C, 72.04; H,
9.37; N, 4.20. Found C, 72.51; H, 9.51; N, 4.41.
4.1.7. Compound 8
Compound 8 was prepared by a method similar to that used
for 7 in 78% yield. Mp 76e78 ꢀC. 1H NMR (300 MHz,
DMSO-d6, 25 ꢀC): d 8.66 (s, 3H, NH3), 4.17 (q, 1H,
J¼7.2 Hz), 4.00 (q, 2H, J¼7.2 Hz), 1.40 (d, 3H, J¼7 Hz),
1.22 (t, 3H, J¼7.2 Hz).
4.1.8. Compound 4
4.1.11. Compound 2
Compound 2 was prepared by a method similar to that used
Compound 6 (100 mg, 0.4 mmol) was dissolved in ethyl
acetate (4 mL), and DCC (105.6 mg, 0.51 mmol) and HOBt
(69.2 mg, 0.51 mmol) were added after cooling to 0 ꢀC. The
mixture was stirred for 30 min whereupon compound 7
(55 mg, 0.36 mmol) and triethylamine (40 mg, 0.4 mmol)
were added. The reaction mixture was stirred at room temper-
ature overnight. The insoluble material was filtered off and the
solution was washed successively with saturated NaHCO3,
1
for 1 in 92% yield. Mp 91.5e93.5 ꢀC. H NMR (300 MHz,
CDCl3, 25 ꢀC): d 8.01 (s, 1H), 4.45 (q, 1H, J¼6 Hz), 2.24 (t,
2H, J¼6.9 Hz), 2.10 (m, 4H), 1.58e1.35 (m, 19H), 0.93
(t, 3H, J¼7.2 Hz); 13C NMR (75 MHz, CDCl3, 25 ꢀC): d 175,
174, 74, 68, 31, 31e29, 20, 16, 15; IR (KBr): ~n¼3315, 2915,
2839, 1696, 1633, 1565, 1518, 1449, 1405, 1300, 1231, 1078,
910, 705, 631, 565, 533; MS (FAB) m/z 334 (MþH)þ. Anal.

1308
S.J. Lee et al. / Tetrahedron 64 (2008) 1301e1308
Masuda, M.; Minamikawa, H. Chem. Rev. 2005, 105, 1401; (d) John,
G.; Mason, M.; Ajayan, P. M.; Dordick, J. S. J. Am. Chem. Soc. 2004,
126, 15012; (e) Jung, J. H.; John, G.; Yoshida, K.; Shimizu, T. J. Am.
Chem. Soc. 2002, 124, 10674.
Calcd for C20H31NO3: C, 72.04; H, 9.37; N, 4.20. Found C,
72.55; H, 9.53; N, 4.31.
4.1.12. Compound 11
4. (a) Jung, J. H.; John, G.; Yoshida, K.; Shimizu, T. J. Am. Chem. Soc. 2002,
124, 10674; (b) John, G.; Jung, J. H.; Minamikawa, H.; Yoshida, K.;
Shimizu, T. Chem.dEur. J. 2002, 8, 5494; (c) John, G.; Jung, J. H.;
Masuda, M.; Shimizu, T. Langmuir 2004, 20, 2060.
5. (a) Ono, Y.; Kanekiyo, K.; Inoue, K.; Hojo, J.; Shinkai, S. Chem. Lett.
1999, 23; (b) Jung, J. H.; Ono, Y.; Shinkai, S. J. Chem. Soc., Perkin
Trans. 2 1999, 1289; (c) Jung, J. H.; Ono, Y.; Shinkai, S. Langmuir
2000, 16, 1643; (d) Jung, J. H.; Ono, Y.; Shinkai, S. Angew. Chem.,
Int. Ed. 2000, 39, 1862; (e) Jung, J. H.; Ono, Y.; Sakurai, K.; Sano,
M.; Shinkai, S. J. Am. Chem. Soc. 2000, 122, 8648; (f) Jung, J. H.;
Kobayashi, H.; Masuda, M.; Shimizu, T.; Shinkai, S. J. Am. Chem. Soc.
2001, 123, 8785.
6. (a) Fuhrhop, J. H.; Spiroski, D.; Boettcher, C. J. Am. Chem. Soc. 1993,
115, 1600; (b) Shimizu, T.; Hato, M. Biochim. Biophys. Acta 1993,
1147, 50; (c) Shimizu, T.; Kogiso, M.; Masuda, M. Nature 1996, 383,
487; (d) Kogiso, M.; Ohnishi, S.; Yase, K.; Masuda, M.; Shimizu, T.
Langmuir 1998, 14, 4978; (e) Kogiso, M.; Okada, Y.; Yase, T.; Shimizu,
T. Biochim. Biophys. Acta 2000, 1475, 346; (f) Kogiso, M.; Zhou, Y.;
Shimizu, T. Adv. Mater. 2007, 19, 242; (g) Spector, M. S.; Easwaran,
K. R. K.; Jyothi, G.; Selinger, J. V.; Singh, A.; Schnur, J. M. Proc.
Natl. Acad. Sci. U.S.A. 1996, 93, 12943; (h) Schnur, J. M.; Ratna,
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J. M.; Price, R. R.; Schnur, J. M. Langmuir 1998, 14, 3493.
To a solution of 7 (1 g, 6.44 mmol) in dry THF (20 mL) at
56 ꢀC under N2 was added TEA (4 g, 39.61 mmol). The reac-
tion mixture was stirred at 56 ꢀC for 10 min. To the reaction
mixture were added stearoyl chloride and
8 (1.5 g,
4.95 mmol). After 1 h, the precipitate was removed by filtra-
tion. Then the solvent was removed by rotary evaporation to
produce a yellow solid. This crude product was purified by
column chromatography (ethyl acetate/hexane, 1/3) to give
the product 9 (1.41 g, 78%). 1H NMR (300 MHz, CDCl3,
25 ꢀC): d 6.04 (s, 1H, NH), 4.60 (q, 1H, J¼6 Hz), 4.21 (q,
2H, J¼7.2 Hz), 2.22 (t, 2H, J¼7.2 Hz), 1.67 (m, 3H), 1.41e
1.27 (m, 33H), 0.89 (t, 3H, J¼6.9 Hz); 13C NMR (75 MHz,
CDCl3, 25 ꢀC): d 173, 172, 61, 47, 36, 31e29, 25, 22, 18,
14; IR (KBr): ~n¼3322, 2957, 2915, 2849, 1742, 1648, 1537,
1472, 1415, 1381, 1201, 1165, 1030, 717, 668; MS (FAB)
m/z 384 (MþH)þ. Anal. Calcd for C23H45NO3: C, 72.01; H,
11.82; N, 3.65. Found C, 71.84; H, 12.01; N, 3.58.
4.1.13. Compound 3
7. (a) Kim, J.-M.; Ji, E.-K.; Woo, S. M.; Lee, H.; Ahn, D. J. Adv. Mater. 2003,
15, 1118; (b) Ahn, D. J.; Chae, E.-H.; Lee, G. S.; Shim, H.-Y.; Chang, T.-E.;
Ahn, K.-D.; Kim, J.-M. J. Am. Chem. Soc. 2003, 125, 8976; (c) Kim, J.-M.;
Lee, Y. B.; Yang, D. H.; Lee, J.-S.; Lee, G. S.; Ahn, D. J. J. Am. Chem. Soc.
2005, 127, 17580; (d) Kim, J.-M.; Lee, J.-S.; Choi, H.; Sohn, D.; Ahn, D. J.
Macromolecules 2005, 38, 9366.
Compound 3 was prepared bya methodsimilar tothatused for
1 in 43% yield. Mp 96.0e97.2 ꢀC. 1H NMR (300 MHz, CDCl3,
25 ꢀC): d 8.51 (s, 1H), 4.39 (q, 1H, J¼6.9 Hz), 3.3 (s, 1H), 2.24
(t, 2H, J¼7.5 Hz), 1.63 (m, 3H), 1.3 (m), 0.92 (t, 3H,
J¼6.9 Hz); 13C NMR (75 MHz, CDCl3, 25 ꢀC): d 174.7, 174.6,
35, 31, 29e27, 25, 22, 16, 13; IR (KBr): ~n¼3316, 2905, 2848,
1707, 1645, 1541, 1467, 1415, 1247, 924, 716; MS (FAB) m/z
356 (MþH)þ. Anal. Calcd for C21H41NO3: C, 70.94; H, 11.62;
N, 3.94. Found C, 70.01; H, 11.2; N, 3.65.
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J. Am. Chem. Soc. 1985, 107, 509; (d) Shoen, P. E.; Yager, P. Mol. Cryst.
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Acknowledgements
10. Sumiyoshi, T.; Nishimura, K.; Nakano, M.; Handa, T.; Miwa, Y.;
Tomioka, K. J. Am. Chem. Soc. 2003, 125, 12137.
This work was supported by KRF (KRF-2005-070-C00068)
and KOSEF (R01-2005-000-10229-0 and M20702000050-
07B-0200-05000). In addition, this work is partially supported
by Korea Ministry of Environment as ‘The Eco-technopia 21
Project’.
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Supplementary data
It includes synthetic routes, SEM images, molecular pack-
ing structure, and IR spectrum of self-assembled 1 and 2. Sup-
plementary data associated with this article can be found in the
online version, at doi:10.1016/j.tet.2007.11.062.
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