Oligocholate foldamer with 'prefolded' macrocycles for enhanced folding in solution and surfactant micelles

Article abstract of DOI:10.1016/j.tet.2013.05.088

A cholate oligomer was synthesized with six linear cholate groups sandwiched by two tricholate macrocycles at the two termini. Two pyrenyl labels on the compound allowed its conformation to be studied by fluorescence spectroscopy. The linear/macrocyclic h

Full text of DOI:10.1016/j.tet.2013.05.088

Tetrahedron 69 (2013) 6051e6059
Contents lists available at SciVerse ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Oligocholate foldamer with ‘prefolded’ macrocycles for enhanced
folding in solution and surfactant micelles
Xueshu Li, Yan Zhao *
Department of Chemistry, Iowa State University, Ames, IA 50011-3111, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A cholate oligomer was synthesized with six linear cholate groups sandwiched by two tricholate mac-
rocycles at the two termini. Two pyrenyl labels on the compound allowed its conformation to be studied
by fluorescence spectroscopy. The linear/macrocyclic hybrid oligomer was found to fold consistently
better than a linear analogue in mixed organic solvents and surfactant micelles. The enhanced folding
was hypothesized to occur as the tricholate macrocycles, by their prefolded configuration, facilitate the
preferential solvation of the facially amphiphilic cholate groups. In general, the folding-enhancing effect
was stronger as the environment became more challenging for the oligocholate to fold. Most unusually,
the hybrid oligocholate folded better in a more challenging binary methanol/ethyl acetate mixture than
in the methanol/hexane/ethyl acetate ternary solvents that have been considered the most ‘folding-
friendly’ to all other cholate foldamers.
Received 18 March 2013
Received in revised form 16 May 2013
Accepted 20 May 2013
Available online 24 May 2013
Keywords:
Foldamer
Conformational control
Macrocycle
Amphiphile
Ó 2013 Elsevier Ltd. All rights reserved.
1. Introduction
of a foldamer host can be exploited to "magnify" its binding affinity
for a guest that triggers additional intrahost interactions during the
1
4
Foldamers as synthetic mimics of conformationally controlled
binding. Such cooperative binding is seen frequently in bi-
molecular recognition including the extraordinary binding be-
biopolymers have attracted many chemists’ attention in recent
1
e7
15,16
years.
In comparison to preorganized macrocycles that are the
tween biotin and streptavidin.
favorites of supramolecular chemists, foldamers are highly tunable
due to their conformational mobility. There are several immediate
benefits resulting from this tunability. First, as a linear oligomer
undergoes a conformational change, its size, shape, and distribu-
tion of functional group could change accordingly, often in re-
sponse to temperature, solvents, and/or the presence of other
molecules and ions in the solution. For this reason, conformational
control can be used as a rational way to create environmentally
Given the highly efficient binding, catalysis, and molecular
transport displayed by biofoldamers, chemists have every reason to
believe that, with increasing knowledge in the construction and
utilization of conformationally controlled molecules, superior ma-
1
e7
terials with protein-like functions may emerge.
Foldamer-based
17e21
synthetic antimicrobial agents,
protein surface-binders and
2
2e25
26
inhibitors,
vesicles and organogellators, and biomimetic
2
7
enantioselective catalysts have all been reported in recent years.
In some cases, synthetic foldamers having quaternary structural
8
9
responsive materials including sensors, ‘smart’ catalysts, and
10e12
28,29
even mechanically responsive materials.
Second, the confor-
order were even synthesized.
mational energy stored in the foldamer backbone may be used to
modulate the molecular recognition of foldamer-based receptors.
Whereas a macrocyclic multidentate ligand may bind a metal ion
with extraordinary affinity as a result of the chelate effect, the co-
operatively folded counterpart may have either very tight or very
loose binding depending on the environment. This tunability is
immensely useful in the controlled binding and release of guest
molecules or ions.13 Third, the cooperative conformational change
Our group has developed a class of foldamers from a naturally
3
0e32
occurring facial amphiphile, cholic acid.
The oligocholates are
characterized by several unusual features: (a) they fold into helical
structures with nanometer-sized internal hydrophilic cavities.
Cavities of this size are typically found in the tertiary and quater-
nary structures of proteins but are formed in our foldamers pre-
pared in a few steps from the cholate monomer (Fig. 1); (b) their
conformation is extremely sensitive to environmental stimuli.
Changes in the solvent composition by a few percent have been
shown to trigger complete unfolding of the folded helix, making the
oligocholates particularly useful as sensors and environmentally
responsive materials; (c) their synthesis is highly modular and their
*
Corresponding author. Tel.: þ1 515 294 5845; fax: þ1 515 294 0105; e-mail
address: zhaoy@iastate.edu (Y. Zhao).
0
040-4020/$ e see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tet.2013.05.088

6052
X. Li, Y. Zhao / Tetrahedron 69 (2013) 6051e6059
Fig. 1. Synthesis of an oligocholate and its solvophobic folding of the oligocholate.
functionalization can be achieved easily by the incorporation of
natural -amino acids; (d) their solvophobic folding has out-
that the macrocycles would concentrate the polar solvent into their
interior and induce the cholate groups of the linear segment of 1 to
fold.
a
3
3
standing tolerance for structural perturbation ; (e) their confor-
mational change is highly cooperative, similar to the two-state
Second, unlike the headetail arrangement of the repeat units in
typical oligocholates, symmetry is introduced in the structure of 1 to
maximize synthetic efficiency. As shown in Scheme 1, the linear
hexacholate in the mid-section of the molecule was obtained by
couplingtwo identical trimeracids (i.e., thecarboxylic acidderivative
of 6) to para-xylylenediamine, doubling the chain length of the
product (7) in one step. Not only so, trimer 6 already had an azido
group at one end of the oligomer, as a result of the azido-
functionalized cholate monomer (Fig. 1). Hence, the azideealkyne
3
4
folding/unfolding of many proteins.
Learning to control the conformation of a molecule is the most
fundamental aspect of foldamer chemistry. In the past, hydrogen-
bonded salt-bridges,34 metaleligand complexation,
13,35
aromatic
3
6
33
donoreacceptor interactions, flexible tethers, and the prefer-
3
4
ential solvation of charged functionalities by polar solvent have
been demonstrated to enhance the solvophobic folding of the oli-
gocholates in organic solvents. However, due to the strong envi-
ronmental dependency of the folding, some of these ‘folding-
enhancing’ methods became ineffective or even detrimental in
4
0,41
click reaction
could be employed conveniently to ‘click’ two
alkyne-functionalized macrocycles (12) directly onto 7, doubling the
chain length again in another step. Note that the alkyne in 12 was
introduced via the phenol side chain of a tyrosine inserted in the
cholate macrocycle. In the end, the highly convergent synthesis and
3
4,37
other environments such as surfactant micelles.
In this paper,
we report a new strategy to enhance the folding of the oligocho-
lates, through incorporation of ‘prefolded’ cholate macrocycles in
the structure. The macrocycles were shown to enhance the folding
of the remaining, linear cholate groups. This effect has been shown
to persist over organic solution and surfactant micelles and be-
comes stronger as the environment is more challenging for the
folding. Most amazingly, the macrocycles enabled the hybrid fol-
damer to do what seemed to be impossible, i.e., to fold better in
a less ‘folding-friendly’ solvent.
strategic usage of naturally functionalized a-amino acids made it
possible to synthesize a complex, conformationally controlled mac-
romolecule over 6000 Da in M.W. in just a few steps from trimer 3.
Third, two pyrenyl labels were introduced at the two ends of
hexacholate 7 via two L-ornithines. Due to its exceptionally long
fluorescence lifetime, pyrene forms excimer readily at a sufficiently
high concentration or if two pyrenyl groups are held together in
4
2
proximity. Because the trimeric periodicity of the oligocholate
foldamer, pyrenyl groups at the ends of a folded hexacholate can be
2
2
. Results and discussion
3
6,38
used to monitor the folding.
It should be mentioned that the above synthetic designs are
only possible because the solvophobic folding of the oligocholates
.1. Molecular design and syntheses
3
8
has extraordinary tolerance for structural heterogeneity. Since
the folding is essentially driven by a solvent effect, there is no re-
quirement for precise alignment of functional groups as in
hydrogen-bonding interactions. In fact, flexible spacers have been
shown in a previous work to facilitate the folding of oligocholates in
mixed organic solvents, presumably because the folded helices
have lower strain than those formed by the more rigid, parent
Synthesis of the linear/macrocyclic hybrid dodecamer (1) is
shown in Scheme 1. An all-linear analogue (2) was prepared pre-
3
8
viously and used as the control compound for the conformational
study. Several considerations went into the design of the hybrid
foldamer.
First, the compound consists of six linearly arranged cholates in
the middle and two macrocyclic tricholate units at the ends. The
driving force for the folding of the oligocholates derives from the
preferential solvation of the cholate polar groups by the polar sol-
vent molecules that phase-separate from the bulk mixture into the
nanocavity (Fig. 1).31 The size, shape, and polarity of the solvent
molecules are all known to impact the folding strongly. Recently,
we established that the same solvophobic driving force to help
a linear oligocholate to fold could make cholate macrocycles such as
3
3
foldamers.
2.2. Folding in mixed organic solvents
The essence of the preferential solvation in Fig. 1 is the sol-
vophobic interactions of the polar faces of the oligocholate in
a largely nonpolar solvent mixture. In a nonpolar-dominant sol-
vent, the extended conformer is disfavored because of its exposed
polar faces to the nonpolar environment. By folding into a helix
with introverted hydrophilic groups, the oligocholate creates a hy-
drophilic internal cavity filled disproportionally with the polar
solvent, allowing the polar faces to largely avoid exposure to the
nonpolar solvent.
3
9
1
3 stack on top of one another in lipid membranes. Because the
folded helix of a linear oligocholate has three cholate groups per
3
0
turn, 13 essentially represents the cross-section of the folded
helix. Since a cholate macrocycle is already fixed into the folded
state by the covalent framework, having two of such units in 1 is
analogous to ‘prefolding’ half of the dodecamer. The anticipation is

X. Li, Y. Zhao / Tetrahedron 69 (2013) 6051e6059
6053
Scheme 1. Syntheses of oligocholate 1.
The most ‘folding-friendly’ solvent systems for the oligocholates
are ternary mixtures such as 2:1 hexane/ethyl acetate with a small
amount of methanol.31 Hexane is immiscible with methanol but
miscible with ethyl acetate. A large amount of hexane in the mix-
ture, thus, makes it easy to phase-separate methanol from the bulk
and reduces the energetic cost associated with the folding. As the
amount of methanol increases, however, the folded oligocholate
intermolecular aggregation in similar solvents at tens of micro-
molar in concentration. Because of the strong fluorescence in-
tensity of pyrene, we performed the solvent titration at 0.0014 and
3
0
0.014
mM, a concentration at which intermolecular aggregation
should be absent. The similar unfolding/unfolding transitions ob-
tained supported the absence of aggregation.
Fig. 2b shows the excimer/monomer ratio of 1 as a function of
the percentage of methanol in the solvent mixture (,). The sig-
moidal curve is characteristic of a cooperative foldingeunfolding
tends to unfold, due to the better solvation of the polar groups by
the bulk solvent.3
0e32
Essentially, when both the hydrophilic and
4
3,44
the hydrophobic faces of the oligocholate are well-solvated, the
unfolded conformation becomes more favorable because of its
higher conformational entropy.
Fig. 2a shows the normalized emission spectra of 1 in 2:1 hex-
ane/ethyl acetate with different percentages of methanol. In low-
methanol solvents, the emission from the pyrene excimer was
clearly visible at 450e500 nm. As the amount of methanol in-
creased, the excimer decreased in intensity, suggesting that the
pyrenyl labels were farther away as the solvent mixture became
more polar. Typical oligocholates were shown to be free of
transition that is frequently seen in proteins.
Such transitions
can be analyzed by a two-state model (Eq. 1), in which the folded
and unfolded states exist simultaneously and interconvert as the
environmental conditions are varied.
K
eq
FoldedðhelixÞ%UnfoldedðcoilÞ
(1)
A two-sate transition normally has a linear dependence of free
energies on the denaturant concentration, i.e., G¼ ꢀm[MeOH],
D
DG
0
in which G0 is the unfolding free energy of the foldamer in the
D

6054
X. Li, Y. Zhao / Tetrahedron 69 (2013) 6051e6059
(a) 1.20
(b)
1
0
.0
.8
1−20%MeOH
0
0
0
.6
.4
.2
0
0
0
.80
.40
.00
0.6
0
0
.4
.2
0.0
0.0
3
50
400
450
500
550
600
0
5
10
15
20
λ
(nm)
% MeOH
Fig. 2. (a) Normalized emission spectra of 1 in 2:1 hexane/ethyl acetate with different percentage of methanol. (b) Excimer/monomer ratio (Fex/Fmon) and fraction of the unfolded
conformer (f ) as a function of volume percentage of methanol in 2:1 hexane/ethyl acetate. The theoretical curves are nonlinear least-squares fitting to a two-state transition model.
Details for the curve-fitting can be found in Ref. 34. Fex and Fmon were the emission intensities at 474 and 385 nm, respectively. [1]¼0.014 M.
U
m
absence of the denaturant and m represents the sensitivity of the
free energy to the concentration of the denaturant. The two-state
model works particularly well for solvophobic foldamers with
0
energy (DG ) than its linear analogue 2 in the ternary mixture
(entries 1 and 2). For the oligocholates, methanol plays a dual
role: when located inside the folded helix, it is needed to solvate
the introverted polar groups and helpful to the folding; when
located outside, it diminishes the solvophobic driving force to the
4
5
30e32
relatively rigid repeat units including oligocholates.
As re-
ported previously,34 the model allows one to correlate an experi-
mentally measured parameter R (¼Fex/Fmon in this case) with the
concentration of the denaturant (methanol in our study) by Eq. 2:
folding by increasing the environmental polarity. Because
0
DG is
the unfolding free energy of the foldamer extrapolated to 0%
methanol, we can view this parameter as a measure of the in-
herent foldability of the oligocholate. For hybrid oligocholate 1,
since only the linear segment can fold and unfold in response to
R ¼ R ꢀ f ðR ꢀ R Þ
F
U
F
U
¼
R ꢀ ðR ꢀ R Þ=f1 þ 1=exp½ ꢀ ð G0 ꢀ m½MeOHꢁÞ=RTꢁg (2)
D
F
F
U
methanol,
0
DG should be the inherent foldability of the linear
in which R
F
is the Fex/Fmon ratio for the fully folded conformer, R
U
segment.
the Fex/Fmon ratio for the fully unfolded conformer, and f
U
the
Once the above point is made clear, it is not surprising to see
that G was higher for 2 than for 1, as the parameter represents
fraction of the unfolded conformer in the mixture.
D
0
As shown in Fig. 2b, the experimental data fitted extremely well
to the two-state model. Fig. 3 shows the curve fittings for 1 and 2 in
MeOH/ethyl acetate. This binary mixture represents a more chal-
lenging condition for the oligocholates to fold because, without
hexane in the mixture, phase separation of methanol from ethyl
the foldability of 12 cholate groups in the linear dodecamer but
only 6 in the linear/macrocyclic hybrid. On the other hand, 2 was
clearly less resistant to methanol denaturation, evident from its
higher m than that of 1. Interestingly, because of the opposite di-
rections of
0
DG and m, the percentage of methanol to trigger 50%
3
3
acetate is energetically more costly.
unfolding happened to be identical for the two compounds. In this
(a)
(b)
1
0
.0
.8
1.0
0.8
0.6
0.4
0.2
0.0
0
0
0
0
.6
.4
.2
.0
0.14
0
0
.13
.12
0.6
0
0
.4
.2
0
.11
0.0
0.10
0
5
10
15
20
0
5
10
15
20
%
MeOH
% MeOH
U
Fig. 3. Excimer/monomer ratio (Fex/Fmon) and fraction of the unfolded conformer (f ) as a function of volume percentage of methanol in ethyl acetate for dodecamer 1 (a) and 2 (b).
The theoretical curves are nonlinear least-squares fitting to a two-state transition model. Details for the curve-fitting can be found in Ref. 34. Fex and Fmon were the emission
intensities at 474 and 385 nm, respectively.
Table 1 summarizes the parameters for the two-state confor-
mational transition obtained from the nonlinear least-squares
fitting. Dodecamer 1 was shown to have a lower unfolding free
sense, the macrocycles did not help the folding too much in the
highly folding-friendly ternary solvent mixture.
In the binary methanol/ethyl acetate mixture, the situation was
dramatically different. As expected, the unfolding free energy at 0%
0
methanol (DG ) decreased for both oligocholates, from 4.6 to 2.4 in
Table 1
Values of
D
G
0
and m determined from solvent denaturation curves
(kcal/mol) m (kcal/mol) % MeOH at
1 and from 6.0 to 2.5 in 2. Thus, the inherent foldability of both
dodecamers decreased significantly. Meantime, both foldamers
became less sensitive to methanol, as demonstrated by their lower
m values in comparison to those in the ternary solvents. The data
seem to suggest that, when the inherent foldability decreased, the
folding was less dependent on methanol. Once again, m is smaller
for 1 than for 2, indicating that the linear/macrocyclic hybrid
dodecamer is more resistant to methanol denaturation than the
Entry Foldamer Solventa
DG
0
50% unfolding
1
2
3
4
1
2
1
2
A
A
B
B
4.6ꢂ0.3
6.0ꢂ0.5
2.4ꢂ0.1
2.5ꢂ0.4
0.74ꢂ0.05
0.96ꢂ0.09
0.30ꢂ0.01
0.73ꢂ0.20
6.2
6.2
7.8
2.9
a
A¼MeOH in 2:1 hexane/ethyl acetate; B¼MeOH in ethyl acetate.

X. Li, Y. Zhao / Tetrahedron 69 (2013) 6051e6059
6055
linear control. The percentage of methanol for 50% unfolding was
.8% for 1 and only 2.9% for 2. Hence, in the more challenging binary
mixture, the folding-enhancing effect of the macrocycles was far
more pronounced than that in the ternary mixture.
Flexible tethers that reduce the strain in the folded helix in solu-
tion were found to allow the oligocholate to satisfy its solvation
needs in the unfolded form on the surface of the micelles. It was
particularly important to us, therefore, to see whether the folding-
enhancing effect of the macrocycles would remain in surfactant
micelles.
7
37
The most surprising result was seen when entries 1 and 3 are
compared. In the solvent denaturation, it took more methanol to
unfold 1 in the less folding-friendly binary solvents than in the
ternary solvents. In other words, with the macrocycles, this com-
pound folded better in a less favorable solvent mixture, following
a completely opposite trend from linear dodecamer 2 (compare
The parent cholate hexamer (without any spacers in between
4
6
the cholates) folds well in sodium dodecyl sulfate (SDS) micelles.
The folding, however, follows a mechanism completely different
from that in solution. The SDS micelle accommodates the folded
conformer much better than the unfolded due to its small size (ca.
3 nm in diameter). Each cholate unit is about 1.4 nm in length.
According to the CPK model, a fully folded cholate hexamer is less
than 2 nm in dimension but an unfolded conformer can stretch to
several nanometers. Because the SDS micelles prefer to maintain
the spherical shape in water, solubilizing an oligocholate within
the micelle is analogous to pushing a snake into a small cage: the
result is that the snake (oligocholate) has no choice but to coil up
(fold).
3
4
entries 2 and 4) and all previously studied (linear) oligocholates.
It is not difficult to understand the inverse correlation between
the folding-enhancing effect of the macrocycles and ‘folding-
friendliness’ of the environments. In a folding-friendly solvent, the
linear dodecamer has a large inherent foldability and the macro-
cycles in a sense are not needed for good folding. The parent cholate
hexamer cannot fold in methanol/ethyl acetate due to the strong
miscibility of the two solvents.14 As the phase separation of
methanol becomes more difficult in the binary methanol/ethyl
acetate mixture, having two prefolded macrocycles becomes more
important to the folding of 1. This is because the amphiphilic
macrocycles, by their numerous inward-facing hydroxyl and amide
groups, could concentrate methanol from the environment without
paying for the loss of conformational entropy, which a folded helix
has to pay. Once two pockets of methanol are formed inside the
macrocycles, it is easier for additional methanol to phase-separate
from the bulk to the vicinity of the linear hexacholate to cause the
latter to fold.
The Fex/Fmon ratio for 1 ranged from w0.8 to w0.4 when the
concentration of SDS increased from 1 to 100 mM. The critical
micelle concentration (CMC) of SDS is 8 mM in water.4 As shown in
Fig. 4a, micellization had no step-changing effect on the relative
distance of the pyrene labels, as the excimer/monomer ratio of 1
7
gradually decreased at higher SDS concentrations. If the high Fex
/
Fmon in 1 below the CMC of SDS was caused by intermolecular
aggregation of the oligocholates, formation of SDS micelles is ex-
pected to dissociate the aggregates, as discovered in a previous
37
How can we rationalize the better folding of 1 in the less folding-
friendly condition? The result seems to contradict logic. When Figs.
study. In such a situation, the excimer/monomer ratio should
have decreased abruptly at the CMC of SDS.
(
b)
(a) 1.00
0
.15
0
0
0
0
0
.80
.60
.40
.20
.00
0.10
0
.05
0.00
0
.5
5
50
0.5
5
50
[
SDS] (mM)
[SDS] (mM)
Fig. 4. Intensity ratio of the excimer over monomer emission (Fex/Fmon) against the concentration of SDS for dodecamer 1 (a) and 2 (b); [oligocholate]¼0.14
mM.
2
b and 3a are examined more closely, it becomes clear that the
If the Fex/Fmon value of 0.4e0.8 in Fig. 4a, on the other hand, was
folding/unfolding transition of this compound was much sharper in
the ternary solvents than in the binary solvents. According to the
data in Table 1, it was the greatly reduced m that allowed 1 to
maintain a higher population of folded conformation in the less
favorable solvents. Thus, the seemingly impossible feat was simply
not caused by the intermolecular aggregation of 1, the alternative
explanation is that a gradual unfolding of the oligocholate has oc-
curred with increasing SDS. The fully folded 1 had an excimer/
monomer ratio of ca. 0.5 in mixed organic solvents (Fig. 2). It is
possible that the SDS-solubilized oligocholate was in a more
‘compressed’ state than the folded helix in solution. A similar ob-
a result of the different magnitudes of the responses of
toward the macrocycles. As long as the positive ‘macrocyclic effect’
on m is larger than the negative effect on , we can expect the
foldamer to appear more stable in the less favorable solvents.
0
DG and m
3
4
servation was made for the parent hexacholate. As mentioned
earlier, the folding of oligocholates in SDS derives from the strong
preference for the anionic micelle to maintain the spherical shape
to minimize charge repulsion of the headgroups. It is questionable
whether the 1eSDS co-assembly would be really spherical but the
low charge repulsion among the surfactant headgroups would still
prefer the dodecamer to be folded for its lower aspect ratiodthis is
the essence of the ‘cage effect’ of SDS surfactants mentioned earlier
DG
0
2
.3. Folding in surfactant micelles
Many of our previously reported folding-promoting methods
become ineffective when the oligocholates move into surfactant
micelles. Intramolecular guanidiniumecarboxylate salt-bridges
lose their effectiveness because guanidinium could form a salt-
bridge with the surfactant headgroup (e.g., sulfate) and charged
functionalities (i.e., guanidinium and carboxylate) prefer the sur-
3
4
on the entrapped oligocholate. At higher concentrations, it is
easier for the micelles to become rod-like in shape and release the
4
6
compressed oligocholate in the unfolded form.
The linear dodecamer (2) behaved quite differently (Fig. 4b).
3
4
48
face of the micelle where they can be well-solvated by water.
Below the CMC of SDS, its Fex/Fmon ratio reached 0.15, slightly

6056
X. Li, Y. Zhao / Tetrahedron 69 (2013) 6051e6059
higher than the highest value (0.13) observed for the folded con-
former in methanol/ethyl acetate (Fig. 3b) and 0.11 in the methanol/
In general, the folding of the oligocholates in mixed organic
solvents is highly predictable, mainly determined by the sol-
vophobic interactions of the polar faces of the cholates in the
nonpolar mixture. What was unusual and most interesting was
that the macrocycle-containing dodecamer folded better in the
less folding-friendly binary solvents than the ternary
mixture. For a cooperatively folded system, structural modifi-
cation could impact both the inherent foldability of the foldamer
and its resistance to denaturant (unfavorable solvents or
whatever other adverse conditions). As shown by the linear/
macrocyclic hybrid oligocholate, because these effects can go to
opposite directions in a nonlinear fashion, structural modifica-
tion could even make a foldamer fold better in a less friendly
environment.
3
8
hexane/ethyl acetate ternary mixture. There could be two pos-
sible reasons for the abrupt decrease of Fex/Fmon upon micellization
of SDS. (a) If the high Fex/Fmon below the CMC of SDS was caused by
intermolecular aggregation of the linear dodecamer, then its drop
at 8 mM SDS would correspond to the disintegration of the ag-
gregates. This effect simply reflected the stronger ability for a mi-
celle to solubilize hydrophobic compounds than individual
surfactants.46 The same effect has been observed in 4-
aminobutyroyl-connected oligocholates that are prone to aggre-
37
gation. (b) To us, however, it seemed too coincidental that a ran-
dom aggregate of could have similar (or even closer)
2
pyreneepyrene distance as that in the folded helix. We, therefore,
prefer the alternative explanation that the high Fex/Fmon of 2 below
the CMC of SDS results from its folding in the surfac-
tanteoligocholate co-assembly. Our previous studies showed that
the folding of an oligocholate in surfactant assemblies is mainly
When solubilized by surfactants in aqueous solution, the needs
for the polar and nonpolar faces of the cholates to receive appro-
priate solvation stay the same. However, these solvation needs may
be met in different ways including intermolecular aggregation,
unfolding, or folding, with surfactant molecules surrounding the
corresponding oligocholates. The conformation of rigid, spacer-free
37,49
determined by the flexibility of the foldamer.
In general, the
parent oligocholates without flexible tethers in between the cho-
lates prefer folding instead of aggregation because the amphiphilic,
rigid, and awkwardly shaped steroid backbone makes it difficult for
the oligomer to pack tightly to form stable intermolecular aggre-
gates. Dodecamer 2 has a rigid, spacer-free, hexacholate in the
4
6
oligocholates in micelles is straightforward to predict, so is that of
37
highly flexible ones. Dodecamers 1 and 2 contain rigid as well as
flexible fragments in the structure. Their folding in the surfactant
assemblies varied with the structure of the oligocholate, as well as
the nature and the concentration of the surfactants.
4
6
middle that is known to fold in the SDS micelle. Thus, it is rea-
sonable that 2 could fold under the same condition.
Switching to a nonionic surfactant (Brij 35) has a large impact on
the conformation of the oligocholates, particularly for the linear
dodecamer (Fig. 5). The Fex/Fmon values were nearly constant for
both compounds, averaging 0.7 for 1 and 0.12 for 2. Since the
concentration of the surfactant increased 10,000-fold in these ex-
periments, it was unlikely that these oligocholates were in the
aggregated state in Brij 35 micelles (CMCz0.1 mM). Instead, the
nearly invariant Fex/Fmon should result from a concentration-
independent, intramolecular process such as folding.
4
4
. Experimental
.1. General
All reagents and solvents were of ACS-certified grade or higher,
and were used as received from commercial suppliers.
Millipore water was used to the micelle solutions. Fluorescence
spectra were recorded on a Varian Cary Eclipse Fluorescence
spectrophotometer.
(a) 1.00
(b)
0
0
0
0
.20
.15
.10
.05
0
0
0
0
0
.80
.60
.40
.20
.00
0.00
0.001 0.01
0
.001 0.01
0.1
Brij 35] (mM)
1
10
100
0.1
1
10
100
[
[
Brij 35] (mM)
Fig. 5. Intensity ratio of the excimer over monomer emission (Fex/Fmon) against the concentration of Brij 35 for dodecamer 1 (a) and 2 (b); [oligocholate]¼0.14
mM.
3
. Conclusions
This study shows that marriage between the conformationally
4.2. Synthesis
The syntheses of compounds 2,38 3,30 and 839 were previously
rigid macrocycle and a conformationally mobile foldamer5 is in-
deed powerful in enhancing the folding. Dodecamer 1 folded con-
sistently better than its linear analogue 2, especially when the
environment became less favorable to the folding. In methanol/
ethyl acetate and SDS micelles, the folding-enhancing effect of the
macrocycles was most obvious, presumably because the preferential
solvation needed for the folded conformer was strongly facilitated
by the prefolded macrocycles. In less challenging environments (i.e.,
methanol/hexane/ethyl acetate and Brij 35 micelles), both dodeca-
mers could fold equally well and the benefit of the macrocycles was
small.
0
reported.
0
4.2.1. Compound 4. N,N -Dicyclohexylcarbodiimide (DCC, 0.854 g,
4.14 mmol) was added to a solution of 1-pyrenecarboxylic acid
(1.71 g, 3.94 mmol) and N-hydroxysuccinimide (HOSu, 0.476 g,
ꢃ
4.14 mmol) in anhydrous THF (10 mL) at 0 C. The reaction mixture
was warmed to room temperature and stirred overnight. The white
precipitate was filtered off and the filtrate was dried in vacuo. The
resulting activated ester was added to a solution of N-Boc ornithine
(0.854 g, 4.14 mmol) and N,N-diisopropylethylamine (DIPEA,
4.1 mL, 23.6 mmol) in anhydrous DMF (10 mL). The reaction

X. Li, Y. Zhao / Tetrahedron 69 (2013) 6051e6059
6057
ꢃ
mixture was stirred under N
diluted with CH Cl . The organic phase was washed with 0.5 N HCl,
brine, dried over MgSO , and concentrated in vacuo. After the sol-
vent was removed in vacuo, the residue was purified by column
chromatography over silica gel using 7:1 CH Cl /MeOH as the el-
uent to give a light yellow powder (1.3 g, 72%). H NMR (400 MHz,
CD OD)
8.60e8.05 (m, 9H), 4.77 (m, 1H), 3.14 (m, J¼6.7 Hz, 2H),
.81 (m, 2H), 1.60 (m, 2H), 1.42 (s, 9H). C NMR (100 MHz, CD
175.6, 173.1, 158.7, 133.9, 132.6, 132.3, 132.2, 129.8, 129.7, 129.6,
2
for 24 h at 70 C. The mixture was
hydrochloride (6.7 mg, 32
m
mol), HOBT (19.5 mg, 144
m
mol), BOP
2
2
(63.7 mg, 144
mmol), DIPEA (0.11 mL, 0.58 mmol, 8 equiv), and
ꢃ
4
anhydrous DMF (1.5 mL). The mixture was stirred at 60 C and the
reaction monitored by TLC. After the completion of the reaction, the
reaction mixture was poured into a dilute HCl solution. The pre-
cipitate was collected by suction filtration, dried in air, and purified
2
2
1
3
d
by column chromatography over silica gel using 10:1 CH
as the eluent to give an off-white powder (91 mg, 72%). H NMR
(600 MHz, CDCl /CD OD) 8.52e8.05 (m, 18H), 7.50 (m, 4H), 4.73
2 2
Cl /MeOH
13
1
1
d
3
OD)
3
3
d
1
5
C
28.3, 127.6, 126.9, 126.8, 126.2, 125.8, 125.64, 125.55, 125.45, 80.0,
(m, 2H), 4.33 (s, 4H), 3.96e3.90 (m, 10H), 3.81e3.80 (m, 6H),
3.76e3.74 (m, 2H), 3.64e3.62 (m, 2H), 3.53e3.51 (m, 4H),
þ
4.5, 40.9, 29.8, 28.8 (3C), 27.9. ESI-HRMS (m/z): [MþH] calcd for
13
27
H
29
N
2
O
5
, 461.2076; found 461.2071.
3.25e0.90 (a series of m), 0.69 (s, 3H), 0.68 (s, 3H), 0.67 (s, 3H).
C
NMR (150 MHz, CDCl /CD OD) 175.7, 174.9, 172.0, 171.9, 171.5,
3
3
d
4
.2.2. Compound 5. DIPEA (247
m
L, 1.4 mmol) was added via a sy-
171.4, 169.2, 169.1, 138.1 (2C), 133.3, 131.7, 131.2, 130.7, 129.3, 129.1,
128.2 (4C), 127.6, 126.4, 126.3, 125.4, 125.2, 124.9, 124.8, 124.7, 73.5,
68.6, 68.4, 52.7, 50.0, 49.9, 47.5, 47.4, 47.3, 46.9, 46.8, 43.4, 42.6, 42.5,
42.4, 42.3, 40.0, 39.9, 39.8, 39.6, 36.7, 36.4, 36.1, 35.3, 35.2, 35.1, 33.9,
33.7, 32.6, 32.5, 30.6, 30.5, 30.1, 28.7, 28.6, 28.1, 27.8, 27.0, 26.0, 23.7,
ringe to a mixture of compound 4 (110 mg, 0.237 mmol), compound
3
0
phonium hexafluorophosphate (BOP, 207 mg, 0.47 mmol) in an-
hydrous DMF (3 mL). The reaction was allowed to continue at room
temperature and monitored by TLC. After the completion of the
reaction, the reaction mixture was poured into an aqueous NaCl
solution. The precipitate was collected by suction filtration, dried in
air, and purified by column chromatography over silica gel using
(285 mg, 0.237 mmol), N-hydroxybenzotriazole (HOBT, 64 mg,
.47 mmol), and (benzotriazol-1-yloxy)tris(dimethylamino)phos-
þ
23.0, 23.0, 17.5, 17.4, 12.8. MALDI-TOF MS (m/z): [MþNa] calcd for
200 284
C H N18NaO24, 3345; found 3347.
4.2.5. Compound 9. Compound 9 was synthesized according to the
51
reported procedure. A mixture of propargyl bromide (1.1 mL,
7.5 mmol), BocNHeTyreOMe (2 g, 6.77 mmol), and K CO (1.87 g,
10:1 CH
2
Cl
2
/MeOH as the eluent to give an off-white powder
2
3
1
(
0.20 g, 69%). H NMR (400 MHz, CDCl
3
/CD
3
OD)
d
8.44e7.96 (m,
13.54 mmol) in DMF (10 mL) was stirred at room temperature
overnight. The mixture was diluted with ethyl acetate. The organic
9
3
CDCl
1
1
H), 4.59 (m, 1H), 3.89 (m, 5H), 3.75 (m, 3H), 3.60 (s, 3H), 3.45 (m,
13
H), 2.35e9.70 (a series of m), 0.58 (m, 9H). C NMR (100 MHz,
/CD OD) 175.3 (2C), 174.4, 174.3 (2C), 157.0, 132.6, 131.1,
30.6, 130.4, 130.2, 128.5, 128.4, 126.9, 126.2, 125.7, 125.6, 124.8,
24.7, 124.5, 124.2, 124.1, 78.9, 72.7, 57.8, 54.5, 51.1, 49.5, 46.7, 46.2,
4
phase was washed with 0.5 N HCl, water, brine, dried over MgSO ,
1
3
3
d
and concentrated in vacuo to give a viscous oil (2.0 g, 89%). H NMR
(400 MHz, CDCl
3
)
d
7.05 (d, J¼8.8 Hz, 2H), 6.90 (d, J¼8.6 Hz, 2H),
4.96 (d, J¼7.2 Hz,1H), 4.66 (d, J¼2.3 Hz, 2H), 4.57e4.52 (m, 1H), 3.71
(s, 3H), 3.08e2.98 (m, 2H), 2.51 (t, J¼2.3 Hz, 1H), 1.41 (s, 9H). A
4
3
2
C
6.1, 42.5, 42.0, 41.9, 41.6, 37.4, 39.3, 36.0, 35.7, 35.6, 35.3, 34.6, 34.5,
4.4, 33.2, 33.19, 31.99, 30.9, 30.8, 28.1, 27.8, 27.4, 27.39, 27.12, 26.43,
2 2
portion of this product (0.53 g) was dissolved CH Cl (2 mL) and
þ
2.9, 22.3, 22.2, 16.7, 16.6, 12.01. ESI-HRMS (m/z): [MþH] calcd for
trifluoroacetic acid (1 mL). After 30 min at room temperature, the
solvents were removed in vacuo and the residue was used in the
next step directly.
100 148 5
H N O14, 1643.1023; found 1643.1017.
4
.2.3. Compound 6. Trifluoroacetic acid (1.0 mL) was added to
a solution of compound 5 (200 mg, 0.12 mmol) in anhydrous CH Cl
4 mL). The mixture was stirred at room temperature and the re-
action monitored by TLC until the starting material was consumed.
After 30 min, the solvents were removed in vacuo. The residue was
2
2
4.2.6. Compound 10. A mixture of compound 8 (122 mg, 0.1 mmol),
compound 9 (41.7 mg, 0.12 mmol), BOP (88 mg, 0.2 mmol), HOBT
(27 mg, 0.2 mmol), and DIPEA (0.11 mL, 0.6 mmol) in anhydrous
(
ꢃ
DMF (2 mL) was allowed to react in a microwave reactor at 100 C
co-evaporated with CH
solved in anhydrous DMF (2 mL), followed by the addition of DIPEA
2
Cl
2
for three times. The residue was dis-
for 45 min. After the completion of the reaction, the reaction
mixture was poured into a dilute HCl solution. The precipitate was
collected by suction filtration, dried in air, and purified by column
(
0.6 mmol) and the HOSu ester of azidoacetic acid (64 mg,
ꢃ
0
.24 mmol). The reaction mixture was stirred at 50 C overnight.
chromatography over silica gel using 12:1 CH
eluent to give an off-white powder (89 mg, 62%). H NMR
(400 MHz, CDCl /CD OD)
2 2
Cl /MeOH as the
1
After the completion of the reaction, the reaction mixture was
poured into a dilute HCl solution. The precipitate was collected by
suction filtration, dried in air, and purified by column chromatog-
3
3
d
7.07 (d, J¼8.8 Hz, 2H), 6.87 (d, J¼8.8 Hz,
2H), 4.65 (d, J¼2.5 Hz, 2H), 3.98e3.91 (m, 4H), 3.80e3.78 (m, 3H),
3.68 (s, 3H), 3.55e3.49 (m, 2H), 3.18 (m, 1H), 3.06 (dd, J¼5.9 ,
13.9 Hz,1H), 2.90 (dd, J¼8.3 ,13.8 Hz,1H), 2.72 (m, 1H), 2.31e0.87 (a
raphy over silica gel using 10:1 CH
an off-white powder (85 mg, 42%). H NMR (400 MHz, CDCl
CD OD) 8.46e7.97 (m, 9H), 4.65 (m, 1H), 3.88 (m, 5H), 3.74 (m,
H), 3.59 (s, 3H), 3.45 (m, 3H), 2.31e0.83 (series m), 0.61 (m, 9H).
2 2
Cl /MeOH as the eluent to give
1
3
/
13
3
d
3 3
series of m), 0.67e0.65 (m, 3H). C NMR (100 MHz, CDCl /CD OD)
3
d 175.1, 174.3, 174.2, 172.4, 157.3, 130.0 (2C), 129.4, 114.7 (2C), 78.5,
1
3
C NMR (100 MHz, CDCl
69.4, 169.3, 133.5, 131.9, 131.8, 131.4, 131.0, 129.4, 129.3, 127.8, 127.1,
26.6, 126.5, 125.5, 125.3, 125.0, 124.9, 124.8, 73.6, 73.5, 68.7, 55.3,
3
/CD
3
OD)
d
176.2, 175.1, 172.3, 172.2, 171.8,
78.4, 75.8, 75.5, 72.8, 72.7, 67.9, 67.8, 67.7, 67.6, 61.4, 55.6, 53.8, 53.6,
51.9, 51.7, 46.8, 46.3, 46.2, 41.9, 41.6, 41.5, 39.4, 39.3, 36.1, 35.8, 35.6,
35.5, 35.3, 34.8, 34.6, 34.5, 34.4, 32.7, 32.6, 31.7, 28.2, 28.1, 28.0, 27.5,
1
1
5
3
2
2.7, 51.9, 50.2, 47.6, 47.1, 47.0, 42.7, 42.4, 40.2, 40.1, 39.6, 39.5, 36.9,
6.5, 36.4, 36.1, 35.5, 35.4, 35.3, 35.2, 34.0, 32.8, 31.7, 31.1, 30.7, 30.6,
9.6, 28.9, 27.9, 27.2, 26.4, 24.4, 23.8, 23.1, 17.6, 17.5, 12.9. ESI-HRMS
27.4, 27.2, 27.1, 26.5, 23.0, 22.4, 22.3, 22.2, 16.9, 16.8, 16.7, 16.6, 12.2,
þ
12.1, 12.0. ESI-HRMS (m/z): [MþH] calcd for C85
131 6 12
H N O ,
1427.9825; found 1427.9820.
þ
(m/z): [MþH] calcd for C97
141 8
H N O13,1626.0618; found 1626.0613.
4
.2.7. Compound 11. Compound 10 (200 mg, 0.14 mmol) and Ph
(74 mg, 0.28 mmol) were dissolved in MeOH (10 mL). The solution
was heated to reflux under N overnight. After the solvent was
removed, the residue was purified by column chromatography
over silica gel using 4:1 CH Cl /MeOH and 2:1:0.1 CH Cl /MeOH/
Et N as the eluents to give an off-white power (180 mg, 91%). H
NMR (400 MHz, CDCl /CD OD)
7.07 (d, J¼8.8 Hz, 2H), 6.87 (d,
J¼8.8 Hz, 2H), 3.96e3.92 (m, 3H), 3.81e3.78 (m, 3H), 3.68 (s, 3H),
3
P
4.2.4. Compound 7. An aqueous solution of 2 M LiOH (0.5 mL,
1.0 mmol) was added to a solution of compound 6 (170 mg,
2
0
.1 mmol) in THF (1 mL). The mixture was stirred at room tem-
perature until TLC showed the completion of the reaction. The
solvents were removed and the residue was dissolved in DMF
2
2
2
2
1
3
(
0.5 mL). The DMF solution was poured into a dilute HCl solution.
3
3
d
0
The precipitate was combined with p-xylene-
a
,a
-diammonium

6058
X. Li, Y. Zhao / Tetrahedron 69 (2013) 6051e6059
3
2
3
.55e3.48 (m, 2H), 3.18 (m, 1H), 3.08e3.04 (m, 1H), 2.93e2.82 (m,
excitation wavelength was 342 nm. For fluorescence titrations in
aqueous surfactant solution, stock solutions (0.10 mM) of the ap-
propriate oligocholates in DMSO were prepared. Aliquots (4.2 mL) of
H), 2.71 (t, J¼2.3 Hz, 1H), 2.37e0.89 (a series of m), 0.68e0.64 (m,
13
H). C NMR (100 MHz, CDCl
3
/CD
3
OD)
d
175.6, 174.85, 174.84,
1
5
3
73.2, 157.4, 130.8 (2C), 130.3, 115.5 (2C), 79.3, 76.6, 73.4, 68.6, 56.4,
4.6, 54.5, 52.6, 49.9, 47.5, 47.1, 47.0, 46.8, 47.4, 42.9, 42.4, 40.3,
7.3, 36.9, 36.7, 36.4, 36.3, 29.0, 28.3, 28.0, 27.3, 23.9, 23.3, 17.8,
the stock solution were added to separate samples (3.00 mL) of the
surfactant solution in water. The samples were gently vortexed for
60 s and allowed to sit at room temperature for 2 h before their
fluorescence spectra were recorded. The excitation wavelength was
342 nm.
þ
1
7.7, 17.6, 13.0. ESI-HRMS (m/z): [MþH] calcd for C85
133 4 12
H N O ,
1401.9920; found 1401.9915.
4
.2.8. Compound 12. Compound 11 (180 mg, 0.128 mmol) was
Acknowledgements
dissolved in a mixture of MeOH (3 mL) and THF (1 mL), followed
by the addition of an aqueous solution of 2 M LiOH (0.65 mL).
After the completion of the reaction, the solvents were removed in
vacuo. A dilute solution of HCl was added to the residue and the
precipitate was collected by suction filtration. The precipitate was
then combined with HOBT (86 mg, 0.64 mmol), BOP (283 mg,
We thank NSF (CHE-1303764) for supporting the research.
References and notes
1. Gellman, S. H. Acc. Chem. Res. 1998, 31, 173e180.
0
.64 mmol), DIPEA (174
m
L, 1 mmol), and anhydrous DMF (2 mL).
2
3
4
. Hill, D. J.; Mio, M. J.; Prince, R. B.; Hughes, T. S.; Moore, J. S. Chem. Rev. 2001, 101,
3893e4011.
. Hecht, S.; Huc, I. In Foldamers: Structure, Properties, and Applications; Wiley-
VCH: Weinheim, Germany, 2007.
. Cubberley, M. S.; Iverson, B. L. Curr. Opin. Chem. Biol. 2001, 5, 650e653.
ꢃ
The mixture was allowed to react in a microwave reactor at 100 C
for 60 min. After the completion of the reaction, the reaction
mixture was poured into a dilute HCl solution. The precipitate was
collected by suction filtration, dried in air, and purified by column
5. Kirshenbaum, K.; Zuckermann, R. N.; Dill, K. A. Curr. Opin. Struct. Biol. 1999, 9,
30e535.
6. Goodman, C. M.; Choi, S.; Shandler, S.; DeGrado, W. F. Nat. Chem. Biol. 2007, 3,
52e262.
5
chromatography over silica gel using 12:1 CH
eluent to give an off-white powder (91 mg, 52%). H NMR
400 MHz, CDCl /CD OD)
7.09 (d, J¼8.4 Hz, 2H), 6.87 (d,
J¼8.8 Hz, 2H), 4.64 (d, J¼2.3 Hz, 2H), 4.50e4.42 (m, 1H), 3.92 (s,
H), 3.78 (s, 3H), 3.50e3.41 (m, 3H), 3.08e3.00 (m, 1H), 2.85e2.77
m, 1H), 2.73 (m, J¼2.3 Hz, 1H), 2.24e0.86 (a series of m),
.68e0.67 (m, 9H). 13C NMR (100 MHz, CDCl
/CD OD) 176.4,
75.5 (2C), 172.1, 157.4, 130.9 (2C), 127.1, 115.6, 115.5, 79.3, 76.5,
3.5, 68.5, 56.4, 54.3, 49.9, 47.0, 46.8, 42.8, 42.7, 42.6, 40.3, 40.1,
7.8, 36.9, 36.8, 36.7, 36.6, 36.0, 35.9, 35.5, 35.3, 33.3, 33.2, 33.1,
2 2
Cl /MeOH as the
1
2
(
3
3
d
7. Bautista, A. D.; Craig, C. J.; Harker, E. A.; Schepartz, A. Curr. Opin. Chem. Biol.
2007, 11, 685e692.
8
9
. Arunkumar, E.; Ajayaghosh, A.; Daub, J. J. Am. Chem. Soc. 2005, 127, 3156e3164.
. Yu, J.; RajanBabu, T. V.; Parquette, J. R. J. Am. Chem. Soc. 2008, 130, 7845e7847.
3
(
0
1
7
3
3
10. Barboiu, M.; Lehn, J. M. Proc. Natl. Acad. Sci. U.S.A. 2002, 99, 5201e5206.
11. Kolomiets, E.; Berl, V.; Lehn, J. M. Chem.dEur. J. 2007, 13, 5466e5479.
3
3
d
1
2. Stadler, A. M.; Ramirez, J.; Lehn, J. M. Chem.dEur. J. 2010, 16, 5369e5378.
3. Zhao, Y.; Zhong, Z. J. Am. Chem. Soc. 2006, 128, 9988e9989.
4. Zhong, Z.; Li, X.; Zhao, Y. J. Am. Chem. Soc. 2011, 133, 8862e8865.
15. Meskers, S.; Ruysschaert, J.-M.; Goormaghtigh, E. J. Am. Chem. Soc. 1999, 121,
115e5122.
6. Williams, D. H.; Stephens, E.; Zhou, M. J. Mol. Biol. 2003, 329, 389e399.
7. Porter, E. A.; Weisblum, B.; Gellman, S. H. J. Am. Chem. Soc. 2002, 124,
7324e7330.
1
1
5
2.7, 32.1, 30.3, 29.1, 28.2, 27.5, 27.3, 27.2, 23.8, 23.2, 17.6, 17.5, 13.0,
1
1
þ
1
2.9, 12.8. ESI-HRMS (m/z): [MþH] calcd for C84
129 4 11
H N O ,
1369.9658; found 1369.9652.
18. Mowery, B. P.; Lee, S. E.; Kissounko, D. A.; Epand, R. F.; Epand, R. M.; Weisblum,
B.; Stahl, S. S.; Gellman, S. H. J. Am. Chem. Soc. 2007, 129, 15474e15476.
9. Chongsiriwatana, N. P.; Patch, J. A.; Czyzewski, A. M.; Dohm, M. T.; Ivankin, A.;
Gidalevitz, D.; Zuckermann, R. N.; Barron, A. E. Proc. Natl. Acad. Sci. U.S.A. 2008,
4
.2.9. Compound 1. Compound 7 (19.4 mg, 5.8
mol) were dissolved in a 2:1:1 mixture of
O (2 mL). Copper power (2 mg) and a 0.1 M CuSO
aqueous solution (20 L, 2 mol) were added. The reaction mixture
was stirred at room temperature under N for 3 d. After removal of
the solvents in vacuo, the residue was purified by preparative TLC
using 5:1 ethyl acetate/MeOH and then 3:1 CH Cl /MeOH as the
developing solvents to give a light yellow powder (24 mg, 69%). H
NMR (600 MHz, CDCl /CD OD) 8.50e7.80 (m, 18H), 7.17 (m, 4H),
.10e6.80 (m, 8H), 5.12e5.08 (m, 8H), 4.74e4.68 (m, 4H), 4.30 (s,
mmol) and com-
1
pound 12 (20 mg, 14.6
THF/MeOH/H
m
105, 2794e2799.
2
4
2
0. Choi, S.; Isaacs, A.; Clements, D.; Liu, D.; Kim, H.; Scott, R. W.; Winkler, J. D.;
DeGrado, W. F. Proc. Natl. Acad. Sci. U.S.A. 2009, 106, 6968e6973.
21. Tew, G. N.; Scott, R. W.; Klein, M. L.; DeGrado, W. F. Acc. Chem. Res. 2009, 43,
m
m
2
30e39.
2
2
2
2. Rodriguez, J. M.; Hamilton, A. D. Angew. Chem., Int. Ed. 2007, 46, 8614e8617.
3. Wyrembak, P. N.; Hamilton, A. D. J. Am. Chem. Soc. 2009, 131, 4566e4567.
4. Horne, W. S.; Johnson, L. M.; Ketas, T. J.; Klasse, P. J.; Lu, M.; Moore, J. P.; Gell-
man, S. H. Proc. Natl. Acad. Sci. U.S.A. 2009, 106, 14751e14756.
5. Imamura, Y.; Watanabe, N.; Umezawa, N.; Iwatsubo, T.; Kato, N.; Tomita, T.;
Higuchi, T. J. Am. Chem. Soc. 2009, 131, 7353e7359.
2
2
1
3
3
d
2
2
7
4
3
H), 3.96e3.86 (m, 12H), 3.80e3.72 (m, 12H), 3.64e3.44 (m, 12H),
.02e3.79 (m, 4H), 2.27e0.80 (a series of m), 0.69e0.55 (m, 36H).
6. Cai, W.; Wang, G. T.; Xu, Y. X.; Jiang, X. K.; Li, Z. T. J. Am. Chem. Soc. 2008, 130,
6936e6937.
1
3
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27.6, 127.0, 126.4, 125.8, 125.4, 124.7, 124.3, 124.2, 124.1, 115.1 (2C),
3 3
/CD OD) d 175.0, 174.5, 174.2, 171.0, 166.0,
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1
2
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4
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6.2, 46.1, 45.4, 42.0, 41.9, 41.8, 39.5, 39.2, 36.1, 36.0, 35.8, 35.7, 35.4,
4.7, 34.6, 34.5, 33.4, 33.1, 32.1, 31.9, 29.5, 28.2, 28.1, 27.4, 27.2, 27.0,
3
6.7, 25.4, 23.1, 22.5, 22.4, 22.3, 17.0, 16.9, 16.8, 12.2. MALDI-TOF MS
þ
(
6
m/z): [MþNaꢀH
2
O] calcd for C368
538
H N
26NaO45, 6065; found
3
067.
3
3
3
4
.3. Fluorescence titration
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/
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3
4
4