Cholesterol-armed cyclens for helical metal complexes offering chiral self-aggregation and sensing of amino acid anions in aqueous solutions

Article abstract of DOI:10.1021/jo0478297

(Chemical Equation Presented) Cholesterol-armed cyclens worked as octadentate receptors for Na⁺, Ca²⁺, and Y³⁺ complexes in which four chiral cholesterol-functionalized sidearms were bundled and asymmetrically twisted abov

Full text of DOI:10.1021/jo0478297

Cholesterol-Armed Cyclens for Helical Metal Complexes Offering
Chiral Self-Aggregation and Sensing of Amino Acid Anions in
Aqueous Solutions
Satoshi Shinoda,† Tsuyoshi Okazaki,† Tomoko N. Player,† Hitomi Misaki,† Kenzi Hori,‡ and
Hiroshi Tsukube*,†
Department of Chemistry, Graduate School of Science, Osaka City University, Sugimoto,
Sumiyoshi-ku, Osaka 558-8585, Japan, and Department of Applied Chemistry and Chemical
Engineering, Faculty of Engineering, Yamaguchi University, Tokiwa-dai, Ube 755-8611, Japan
tsukube@sci.osaka-cu.ac.jp
Received December 10, 2004
Cholesterol-armed cyclens worked as octadentate receptors for Na+, Ca2+, and Y3+ complexes in
which four chiral cholesterol-functionalized sidearms were bundled and asymmetrically twisted
above cyclen-metal complex platforms. Since the resulting helical metal complexes included chiral,
hydrophobic cholesterol residues and charged, hydrophilic metal sites as well as asymmetric
coordination geometries, they exhibited unique amphiphilic properties and provided chiral self-
aggregates in aqueous solutions. Light scattering, fluorescence, and TEM characterizations
demonstrated that Na+ complex with cholesterol-armed cyclen gave a particularly stable self-
aggregate in aqueous solution and offered supramolecular environments effective for sensing and
detection of amino acid anions. Various dansylamino acid derivatives (dansyl ) 5-(dimethylamino)-
1-naphthalenesulfonyl) were nicely accommodated in the helicate aggregates to give highly enhanced
fluorescence signals, which could be detected by the naked eye at 10-7 mol/L level. Their inclusion
behaviors were analyzed by a Langmuir-type equation, indicating that enantiomer-selective
inclusion occurred. MM/MD calculations and circular dichroism (CD) studies further suggested
that cholesterol-armed cyclen helicates have chiral and hydrophobic cavities upon self-aggregation,
in which the dansylamino acid anions were specifically accommodated. Since these helicates
exhibited nonselective binding abilities in solvent extraction experiments of dansylamino acid anions,
uncommon chiral recognition and sensing functions were generated by supramolecular alignments
of the chiral metal helicates in the aqueous solutions.
Introduction
transition metal complexes and octa-/nonacoordinate
lanthanide complexes have received considerable atten-
tion as functional helicates.3 Armed cyclens having four
Various types of receptor molecules were recently
designed and their metal complexes had highly organized
structures and sophisticated functions.1 Representatives
of these are the helical metal complexes, so-called “he-
licates”, with one or more coordinating receptors, which
have well-defined coordination topology and high stability
even in solution states. In addition to various oligopyri-
dine ligand-metal complexes,2 a series of hexacoordinate
(1) (a) Swiegers, G. F.; Malfetese, T. F. Chem. Rev. 2000, 100, 3483.
(b) Fujita, M.; Umemoto, K.; Yoshizawa, M.; Fujita, N.; Kusukawa,
T.; Biradha, K. Chem. Commun. 2001, 509. (c) Holloday, B. J.; Mirkin,
C. A. Angew. Chem., Int. Ed. 2001, 113, 2076. (d) Lehn, J. P. Proc.
Natl. Acad. Sci. 2002, 99, 4763. (e) Seideel, S. R.; Stang, P. J. Acc.
Chem. Res. 2002, 35, 972. (f) Schalley, C. A.; Lutzen, A.; Albrecht, M.
Chem. Eur. J. 2004, 10, 1072. (g) Wurthner, F.; You, C.-C.; Saha-
Moller, C. R. Chem. Soc. Rev. 2004, 33, 133. (h) Pagliari, S.; Corradini,
R.; Galaverna, G.; Sforza, S.; Dossena, A.; Montalti, M.; Prodi, L.;
Zaccheroni, N.; Marchelli, R. Chem. Eur. J. 2004, 10, 2749.
† Osaka City University.
‡ Yamaguchi University.
10.1021/jo0478297 CCC: $30.25 © 2005 American Chemical Society
Published on Web 02/08/2005
J. Org. Chem. 2005, 70, 1835-1843
1835

Shinoda et al.
cation-ligating sidearms were typically reported to form
helical complexes with alkali, alkaline earth and lan-
thanide metal cations, which often had twisted square
anti-prismatic structures.4 When chiral substituents are
attached to the cyclen system, the four sidearms stand
up in the same direction above the macrocyclic ring and
are asymmetrically arranged in a quadruply helical
fashion. Wainwright et al. reported that Cd2+ complex
with chiral alcohol-armed cyclen further formed ternary
complexes with D- and L-histidinate anions in DMSO
solution.5a,b Parker et al. recently employed Yb3+ complex
with chiral, heptadentate amide-armed cyclen in recogni-
tion of lactate and alaninate anions in aqueous solu-
tions.5c,d Since these chiral helicates have potential as a
new class of metallo-receptors, synthetic approaches are
required at not only the molecular level but also the
supramolecular level, so that they can have defined
cavities effective for precise recognition.
As frequently found in protein-based supramolecular
systems,6 the highly structured helicates are expected to
exhibit their sophisticated functionalities via supramo-
lecular aggregation through noncovalent interactions.
Since there are many structural variations, synthetic
metal helicates can be versatile building blocks for new
supramolecular receptors, catalysts, devices, and related
systems along this line. In earlier communications,7 we
demonstrated that cholesterol-armed cyclen 1-Na+ com-
plex exhibited chiral and amphiphilic natures and spon-
taneously formed a stable self-aggregate in aqueous
solution. Although several kinds of metal complex-based
amphiphiles have been developed,8 this helicate provided
specific environments for guest accommodation based on
the following chiroptical features: (1) assembling of
chiral, hydrophobic cholesterol residues; (2) helicity of
octadentate cyclen-metal complex; and (3) supramolecu-
lar alignment of chiral helicates. The aggregate of cyclen
1-Na+ complex nicely accommodated dansylglycine an-
ion and asymmetrically fixed its conformation to induce
the circular dichroism (CD) signals.7 Here, we combine
armed cyclen chemistry with self-aggregation technology
for supramolecular architectures, in which a series of
cholesterol-armed cyclen complexes with Na+, Ca2+ and
Y3+ cations are integrated at the supramolecular level.
Some of them are applicable in sensing and detection of
dansylamino acid anions of 1 × 10-7 mol/L by the naked
eye (see picture in Abstract). Enantiomer determination
of the employed dansyl amino acids was reported using
capillary electrophoresis and fluorescence methods, in
which glycopeptide antibiotics and an R-acid glycoprotein
were typically used as chiral selectors.9 Although the
cholesterol-armed cyclen metal complexes rarely exhib-
ited such anion recognition and sensing behaviors in
homogeneous solution systems, they offered the sophis-
ticated functions upon self-aggregation in aqueous media.
Results and Discussion
1. Cholesterol-Armed Cyclens for Chiral Heli-
cates. Several armed cyclens were reported to bind Na+,
Ca2+, and trivalent lanthanide cations quite strongly via
octa-coordination from parent cyclen nitrogen atoms and
donor sidearms.4 NMR, CD, X-ray, and CPL character-
izations have shown that only single C4 orientation of
the armed cyclen around the metal center predominated
when chiral substituents were introduced on the four
sidearms. Although cholesterol-armed cyclens 1 and 2
(Figure 1) similarly formed chiral helicates, their metal
complexes were further furnished with hydrophobic
cholesterol groups and the trapped metal cation as chiral
amphiphiles for self-aggregation. NaCl complex with
cyclen 1 is large (10 Å × 10 Å × 25 Å) with an inner void
and exhibited high stability in solution (log K ) 11.0 in
ethanol).4f Figure 2 indicates 13C NMR spectra of cyclen
1-NaCl complex measured in CDCl3 at various temper-
atures. The signals for two cyclen ring carbons separately
resonated at 53.0 and 48.4 ppm at <308 K, while the
singlet signal was observed at 75.5 ppm for N-CH2-CO-
carbons of four sidearms. Since cyclen 1 itself gave no
diastereomeric signal for cyclen ring carbons even at 233
K, its Na+ complex was confirmed to have quadruply
helical structure with C4 symmetry. Increasing the
(2) (a) Baxter, P. N.; Lehn, J.-M.; Kneisel, B. O.; Fenske, D. Angew.
Chem., Int. Ed. Engl. 1997, 19788. (b) Mamula, O.; von Zelewsky, A.
Coord. Chem. Rev. 2003, 242, 87. (c) Hofmeier, H.; Schubert, U. S.
Chem. Soc. Rev. 2004, 33, 373.
(3) (a) Bluhm, M. E.; Hay, B. P.; Kim, S. S.; Dertz, E. A.; Raymond,
K. N. Inorg. Chem. 2002, 41, 5475. (b) Bowyer, P. K.; Cook, V. C.;
Gharib-Naseri, N.; Gugger, P. A.; Rae, A. D.; Swiegers, G. F.; Willis,
A. C.; Zank, J.; Wild, S. B. Proc. Natl. Acad. Sci. 2002, 99, 4877. (c)
Prabaharan, R.; Fletcher, N. C. Dalton Trans. 2003, 2558. (d) Koeller,
S.; Bernardinelli, G.; Bocquet, B.; Piquet, C. Chem. Eur. J. 2003, 9,
1062. (e) Pope, S. J. A.; Kenwright, A. M.; Heath, S. L.; Faulkner, S.
Chem Commun. 2003, 1550. (f) Mizukami, S.; Houjou, H.; Kanesato,
M.; Hiratani, K. Chem. Eur. J. 2003, 9, 1521. (g) Miyake, H.; Yoshida,
K.; Sugimoto, H.; Tsukube, H. J. Am. Chem. Soc. 2004, 126, 6524.
(4) (a) Wainwright, K. P. Coord. Chem. Rev. 1997, 166, 35. (b)
Parker, D.; Dickins, R. S.; Puschmann, H.; Crossland, C.; Howard, J.
A. K. Chem. Rev. 2002, 102, 1977. (c) Tsukube, H.; Mizutani, Y.;
Shinoda, S.; Okazaki, T.; Tadokoro, M.; Hori, K. Inorg. Chem. 1999,
38, 3506. (d) Woods, M.; Aime, S.; Botta, M.; Howard, J. A. K.; Moloney,
J. M.; Navet, M.; Parker, D.; Port, M.; Rousseaux, O. J. Am. Chem.
Soc. 2000, 122, 9781. (e) Aoki, S.; Kawatani, H.; Goto, T.; Kimura, E.;
Shiro, M. J. Am. Chem. Soc. 2001, 123, 1123. (f) Shinoda, S.;
Nishimura, T.; Tadokoro, M.; Tsukube, H. J. Org. Chem. 2001, 66,
6104. (g) Gunnlaugsson, T.; Leonard, J. P.; Senechal, K.; Harte, A. J.
J. Am. Chem. Soc. 2003, 125, 12062. (h) Liu, S.; He, Z.; Hsieh, W.-Y.;
Fanwick, P. E. Inorg. Chem. 2003, 42, 8831. (i) Woods, M.; Kiefer, G.
E.; Castillo-Muzquiz, A.; Eshelbrenner, C.; Michaudet, L.; McMillan,
K.; Mudigunda, S. D. K.; Ogrin, D.; Tircso, G.; Zhang, S.; Zhao, P.;
Sherry, A. D. J. Am. Chem. Soc. 2004, 126, 9248.
(5) (a) Smith, C. B.; Wallwork, K. S.; Weeks, J. M.; Buntine, M. A.;
Lincoln, S. F.; Taylor, M. R.; Wainwright K. P. Inorg. Chem. 1999, 38,
4986. (b) Smith, C. B.; Stephenes, A. K. W.; Wallwork, K. S.; Lincoln,
S. F.; Taylor, M. R. Wainwright K. P. Inorg. Chem. 2002, 41, 1093. (c)
Bruce, J. L.; Dickins, R. S.; Govenlock, L. J.; Gunnlaugsson, T.;
Lopinski, S.; Lowe, M. P.; Parker, D.; Peacock, R. D.; Perry, J. J. B.;
Aime, S.; Botta, M. J. Am. Chem. Soc. 2000, 122, 9674. (d) Dickins, R.
S.; Batsano, A. S.; Howard, J. A. K.; Parker, D.; Puschmann, H.;
Salamano, S. Dalton Trans. 2004, 70.
(6) (a) McDermott, G.; Prince, S. M.; Freer, A. A.; Hawthornthwaite-
Lawless, A. M.; Papiz, M. Z.; Cogdell, R. J.; Isaacs, N. W. Nature 1995,
374, 517. (b) Yu, S. M.; Conticello, V. P.; Zhang, G.; Kayser, C.;
Fournier, M. J.; Mason, T. L.; Tirrell, D. A. Nature 1997, 389, 167. (c)
Tsukube, H.; Shinoda, S. Bull. Chem. Soc. Jpn. 2004, 77, 453.
(7) (a) Shinoda, S.; Okazaki, T.; Nishimura, T.; Hori, K.; Tsukube,
H. Chem. Commun. 2001, 976. (b) Nishimura, T.; Shinoda, S.; Tsukube,
H. Chirality 2002, 14, 555.
(8) (a) Fyles, T. M.; Straten-Nijenhuis, W. F. Comprehensive Su-
pramolecular Chemistry; Pergamon Press: Oxford 1996, Vol. 10, pp
53. (b) Tanaka, Y.; Miyachi. M.; Kobuke, Y. Angew. Chem., Int. Ed.
1999, 38, 504. (c) Andre, J. P.; Toth, E.; Fischer, H.; Seelig, A.; Macke,
H. R.; Merbach, A. E. Chem. Eur. J. 1999, 5, 2977. (d) Pidwell, A. D.;
Collinson, S. R.; Coles, S. J.; Hursthouse, M. B.; Schroder, M.; Bruce,
D. M. Chem. Commun. 2000, 955. (e) Gebbink, R. J. M.; Sandee, A. J.;
Peters, F. G. A.; van der Gaast, S. J.; Feiters, M. C.; Nolte, R. J. M. J.
Chem. Soc., Dalton Trans. 2001, 3056. (f) Yuan, J.; Liu, M. J. Am.
Chem. Soc. 2003, 125, 5051. (g) Nakai, T.; Kanamori, T.; Sando, S.;
Aoyama, Y. J. Am. Chem. Soc. 2003, 125, 8465. (h) Larpent, C.;
Laplace, A.; Zemb, T. Angew. Chem., Int. Ed. 2004, 43, 3163.
(9) (a) Kang, J.; Bischoff, D.; Jiang, Z.; Bister, B.; Su¨ssmuth, R. D.;
Schurig, V. Anal. Chem. 2004, 76, 2387. (b) Yan, Y.; Myrick, M. L.
Anal. Chem. 1999, 71, 1958.
1836 J. Org. Chem., Vol. 70, No. 5, 2005

Cholesterol-Armed Cyclens
complex exhibited similar NMR spectral profiles to those
with Na+ complex, though Y3+ complex gave broad and
1
unidentified H NMR signals.
When four planar and hydrophobic cholesterol moieties
stand up above the Na+ - cyclen complex platform, a
substantial box-like cavity can arise from juxtaposition
of the four cholesterol planes. Although the suitable
crystal for X-ray structural determination was not ob-
tained, modeling experiments combined with molecular
mechanics and molecular dynamics calculations were
carried out. These support that cyclen 1-Na+ complex
can have a chiral, hydrophobic cavity suitable for guest
accommodation and self-aggregation. As schematically
illustrated in Figure 3, two types of three-dimensional
structures were postulated for cholesterol-armed cyclen-
Na+ complex: a saucer-shaped open-form and a stewpan-
shaped closed-form. We have reported that some ester
and amide-armed cyclen-Na+ complexes had twisted
anti-prismatic structures in solid states and preferred
saucer-shaped structures due to the steric factors involv-
ing four sidearm-substituents.4c,f Therefore, cyclen 1-Na+
complex was first optimized as a saucer-shaped structure
using the CAChe MM2 force field. Two diastereomeric
structures with clockwise and anticlockwise helicities
(Figure 2) were optimized to have similar stabilities.
When these saucer-shaped structures were used for MD
calculations, the stewpan-shaped structures were newly
generated from both clockwise and anticlockwise precur-
sors. Effective interactions between the cholesterol arms
are believed to be the driving force for structural changes
from the saucer-shaped to the stewpan-shaped forms.
Five initial geometries of the stewpan-shaped close-form
were extracted from trajectories of MD simulations. An
optimized structure of this close-form is shown in Figure
3. In this complex, the four cholesterol moieties stand
up in the same direction to provide a chiral, hydrophobic
cavity. Since both interior and exterior surfaces of the
complex are composed of hydrophobic planes, this can be
effective not only for guest accommodation but for self-
aggregation. Similar stewpan-shaped structures had been
established in other types of armed cyclen complexes.5
Although various cholesterol derivatives are known to
provide chiral, hydrophobic environments for molecular
recognition and self-aggregation,10,11 the present type of
armed cyclen helicates is suggested to offer effective
arrangements of the chiral, four cholesterol sidearms for
supramolecular recognition and aggregation.
FIGURE 1. Cholesterol-armed cyclen 1 and its derivatives 2
and 3.
2. Self-Aggregation of Cholesterol-Armed Cy-
clen-Metal Complexes. When cyclen 1-NaCl complex
was dispersed in the aqueous ethanolic solution
(H2O/EtOH ) 80/20, v/v), this spontaneously formed very
stable self-aggregate and gave no precipitate from the
aqueous solution after 10 days. Its aggregation behaviors
were characterized by fluorescence titrations, light-
(10) Cholesterol assemblies: (a) Davis, A. P.; Bonar-Law, R. P.;
Sanders, J. K. M. Comprehensive Supramolecular Chemistry; Perga-
mon Press: Oxford 1996, Vol. 4, p 257. Recent examples: (b) Janout,
V.; Giorgio, C. D.; Regen, S. L. J. Am. Chem. Soc. 2000, 122, 2671. (c)
Janout, V.; Jing, B.; Staina, I. V.; Regen, S. L. J. Am. Chem. Soc. 2003,
125, 4436. (d) Klein, C.; Nazeeruddin, M. K.; Censo, D. D.; Liska, P.;
Gratzel, M. Inorg. Chem. 2004, 43, 4216. (e) Sugiyasu, K.; Fujita, N.;
Shinkai, S. Angew. Chem., Int. Ed. 2004, 43, 1229.
(11) (a) Gokel, G. W.; Murillo, O. Acc. Chem. Res. 1996, 29, 425. (b)
Shinkai, S.; Murata, K. J. Mater. Chem. 1998, 8, 485. (c) Ariga, K.;
Terasaka, Y.; Sakai, D.; Tsuji, H.; Kikuchi, J. J. Am. Chem. Soc. 2000,
122, 7835.
FIGURE 2. Two possible diastereomers and 13C NMR spectra
of cyclen 1-NaCl complex. Conditions: cyclen 1-NaCl, 4 ×
10-2 mol/L in CDCl3.
sample temperature to 318 K caused disappearance of
the cyclen ring signals, suggesting that intramolecular
ligand exchange occurred at higher temperature. Ca2+
and Y3+ complexes were prepared by mixing cyclen 1 and
corresponding metal triflates in CH3CN/CHCl3. Its Ca2+
J. Org. Chem, Vol. 70, No. 5, 2005 1837

Shinoda et al.
FIGURE 3. Schematic illustration of two preferred conformations of cyclen 1-Na+ complex and their optimized structures (top
views).
dansyl-L-phenylalanine anion. When cyclen 1-Na+ com-
plex was gradually added to the aqueous solution of
dansyl-L-phenylalanine anion (1 × 10-5 mol/L), its fluo-
rescence maximum continuously shifted from 538 to 498
nm and the intensity recorded at 498 nm was gradually
increased. Pronounced changes in the fluorescence in-
tensity occurred around 4.0 × 10-6 mol/L of cyclen 1-Na+
complex (Figure 4A). Dynamic light scattering experi-
ments showed that the resulting self-aggregate had a
mean hydrodynamic radius of 60 nm, when 1 × 10-4
mol/L of the Na+ complex was dispersed (Figure 4B). A
TEM picture taken after treatment of uranyl acetate
indicates that dispersed self-aggregates of 60 nm or
larger were observed (Figure 4C). When the size distri-
butions were measured at 20-60 °C, the average distri-
bution was confirmed as almost constant, revealing that
the formed self-aggregate was stable even at a wide range
of temperature. Ca2+ and Y3+ complexes with armed
cyclen 1 similarly formed self-aggregates in the aqueous
ethanolic solutions, but their particle sizes were some-
what different from that with Na+ complex: 112 nm for
Ca2+ complex and 29 nm for Y3+ complex. Since these
three cations have almost the same ion radii,12 the charge
state of the metal center played an important role in the
self-aggregation processes.
FIGURE 4. Self-aggregate of armed cyclen 1-NaCl complex
in H2O/EtOH (80/20 v/v, pH ) 7.2). (A) Fluorescence titration
with dansyl-L-phenylalanine. (B) Size distribution by dynamic
light scattering. (C) TEM picture.
Armed cyclen 2-Na+ complex also formed self-ag-
gregate in the aqueous ethanol solutions, though up to
30% ethanol content is required to give a spectroscopi-
scattering experiments, and TEM observations (Figure
4). Its critical aggregation concentration (cac) was esti-
mated as 4.0 × 10-6 mol/L by titration with fluorescent
(12) Shannon, R. D. Acta Crystallogr. 1976, A32, 751.
1838 J. Org. Chem., Vol. 70, No. 5, 2005

Cholesterol-Armed Cyclens
FIGURE 5. Fluorescence sensing of dansyl-L- and -D-leucine anions with cyclen 1-metal complex aggregates: (a) dansyl-L-
leucine (bound), (b) dansyl-D-leucine (bound), (c) dansyl-L- and -D-leucine (free). Conditions: cyclen metal complex, 3.3 × 10-5
mol/L; dansylleucine, 1.0 × 10-6 mol/L; in H2O/EtOH (80/20 v/v); pH ) 7.2.
TABLE 1. Fluorescence and Binding Behaviors of Dansylamino Acid Anions in Armed Cyclen 1-NaCl Self-Aggregate
fluorescencea
intensity (λmax/nm)
bindingc
K/M-1
dansylamino acid
no aggregate
aggregate
enhancementb
n/m
log D
L-valine
27 (536)
26 (537)
37 (534)
36 (537)
32 (533)
32 (532)
32 (539)
32 (538)
25 (538)
25 (539)
213 (504)
185 (505)
258 (504)
226 (506)
207 (507)
158 (505)
200 (514)
250 (504)
229 (498)
203 (505)
7.9
7.1
7.0
6.3
6.5
4.9
6.3
7.8
9.2
8.1
(2.3 ( 0.8) × 105
(1.7 ( 0.3) × 105
(2.8 ( 0.4) × 105
(1.9 ( 0.3) × 105
(3.1 ( 0.4) × 105
(1.4 ( 0.2) × 105
(4.5 ( 0.7) × 105
(8.8 ( 1.7) × 105
(4.2 ( 0.6) × 105
(6.0 ( 1.4) × 105
0.59
0.38
0.55
0.30
0.35
0.30
0.67
0.65
0.53
0.55
-1.35
D-valine
L-t-leucine
-1.05
-0.88
-1.03
-0.60
D-t-leucine
L-leucine
D-leucine
L-phenylglycine
D-phenylglycine
L-phenylalnine
D-phenylalnine
a Conditions: cyclen 1-NaCl, 1.0 × 10-4 mol/L; dansylamino acid, 1.0 × 10-5 mol/L; in H2O/EtOH (80/20, pH ) 7.2). b Fluorescence
intensity at λmax in the presence of aggregate/fluorescence intensity at λmax in the absence of aggregate. c Conditions: see the Experimental
Section.
cally clear solution. Its cac value (1.6 × 10-5 mol/L) and
averaged particle-size (30 nm) were different from those
of cyclen 1-Na+ complex in H2O/EtOH (70/30, w/w),
indicating that the sidearm substitution altered the
detailed profile of the self-aggregate. When CD spectrum
of cyclen 2-Na+ complex in ethanol was compared with
that in aqueous ethanol solution, the sign of CD signal
around 230 nm changed drastically (see the Supporting
Information, Figure S1): a negative sense in EtOH f a
positive sense in H2O/EtOH. These CD spectral changes
clearly indicate that the self-aggregation greatly altered
chiral environments around the phenyl chromophore of
the cyclen complex. Probably, the three cholesterol-
functionalized sidearms of cyclen 2 were compressed
upon self-aggregation to stabilize the stewpan-shaped
conformation (see Figure 3). Similar cavity shape conver-
sion of the receptor was reported in the monolayer system
composed of steroid cyclophane,11c in which four steroid
moieties were rearranged upon monolayer formation at
the air/water interface to offer the hydrophobic cavity
effective for guest accommodation.
nature of the metal center as well as mole ratio of guest
to complex amphiphile. Figure 5 compares the enhanced
fluorescence profiles of dansyl-L- and D-leucine anions
upon inclusion into the self-aggregates of cyclen 1-Na+,
-Ca2+, and -Y3+ complexes. Among them, cyclen 1-Na+
complex exhibited the largest fluorescence enhance-
ment: 6.1 times for Na+ complex; 2.5 times for Ca2+
complex; and 3.6 times for Y3+ complex. The intensity
difference between the guest enantiomers was observed
in each helicate aggregate, and Na+ complex also offered
a greater difference than Ca2+ and Y3+ complexes. Several
dansylamino acids of 10-7 mol/L were detected by the
naked eye13 in the presence of cyclen 1-Na+ complex
aggregate (see picture in Abstract).
Cyclen 1-Na+ complex gave the most stable self-
aggregate in the aqueous media, and its anion recognition
and sensing behaviors were characterized by fluorescence
(13) Naked eye detection: (a) Miyaji, H.; Sessler, J. L. Angew. Chem.,
Int. Ed. 2001, 40, 154. (b) Yamaguchi, S.; Akiyama, S.; Tamao, K. J.
Am. Chem. Soc. 2001, 123, 11372. (c) Ros-Lis, J. V.; Martinez-Manez,
R.; Sto, J. Chem. Commun. 2002, 2248. (d) Piatek, P.; Jurczak, J. Chem.
Commun. 2002, 2450. (e) Mahajan, R. K.; Kaur, I.; Kaur, R.; Uchida,
S.; Onimaru, A.; Shinoda, S.; Tsukube, H. Chem. Commun. 2003, 2238.
(f) Yamada, T.; Shinoda, S.; Sugimoto, H.; Uenishi, J.; Tsukube, H.
Inorg. Chem. 2003, 42, 7932. (g) Rusin, C.; St. Luce, N. N.; Agbaria,
R. A.; Escobedo, J. O.; Jiang, S.; Warner, I. M.; Dawan, F. B.; Lian,
K.; Strongin, R. J. Am. Chem. Soc. 2004, 126, 438. (h) Vazquez, M.;
Fabbrizzi, L.; Taglietti, A.; Pedrido, R. M.; Gonzalez-Noya, A. M.;
Bermejo, M. R. Angew. Chem., Int. Ed. 2004, 43, 1962.
3. Fluorescence Sensing of Amino Acid Anions in
Aqueous Media. Dansylamino acid anions were nicely
accommodated in armed cyclen 1-metal complex ag-
gregates and exhibited characteristic fluorescence en-
hancements. The fluorescence profiles of the bound
dansylamino acid anions were largely dependent on the
J. Org. Chem, Vol. 70, No. 5, 2005 1839

Shinoda et al.
method (Table 1). When an excess of the self-aggregate
(1.0 × 10-4 mol/L) was added to an aqueous ethanol
solution of dansyl-L- or D-phenylalanine anion (1.0 × 10-5
mol/L), the fluorescence maxima of the bound anions
shifted from 538 to 498 nm for L-isomer and to 505 nm
for D-isomer. Under the employed conditions (see Table
1), the fluorescence intensities were enhanced 9.2-fold for
dansyl-L-phenylalanine anion and 8.1-fold for its D-isomer
upon inclusion, indicating that the two enantiomers were
located under somewhat different circumstances. In
contrast, the enantiomers of aliphatic amino acid anions
exhibited the enhanced fluorescence signals upon inclu-
sion, but their maxima appeared at almost the same
wavelengths: 507 and 505 nm for dansylleucine; 504 and
506 nm for dansyl-tert-leucine; and 504 and 505 nm for
dansylvaline. These observations suggest that the phenyl
residues of aromatic amino acids occupy unique positions
in the self-aggregate. The fluorescence maxima of the
dansyl derivatives generally relate to the polar natures
of the employed solvents. The fluorescence maxima of
dansyl-L-leucine were recorded in the absence of self-
aggregate at 533 nm in H2O (pH ) 7.2), 503 nm in
ethanol, and 490 nm in CHCl3. Therefore, cyclen 1-Na+
complex aggregate provided an ethanol-like environment
around the dansyl chromophores. With an increase of
ethanol content in the aqueous aggregate solution, the
intensity of the observed fluorescence signal decreased,
and there was no enhancement in 60% of ethanol aqueous
solution. Since the particle sizes could not be determined
under such ethanol-rich conditions by light scattering
method, cyclen 1-Na+ complex in a nonaggregated form
did not have an effective cavity to accommodate dan-
sylamino acid anions. Although several chiral disk-
shaped molecules were reported to exhibit chirality-
amplification by stacking in highly ordered chiral col-
umns in the solutions,14 the present type of helicates
generated unique anion-sensing functions upon self-
aggregation in aqueous media. Cationic dansylethylene-
diamine hydrochloride was also employed as a fluores-
cence probe. Since its fluorescence intensity was rarely
enhanced in the presence of cyclen 1-Na+ complex
aggregate, the electrostatic attraction between the metal
fitting method (see the Supporting Information). As
summarized in Table 1, the self-aggregate of cyclen
1-Na+
complex preferred L-isomers of aliphatic amino
acids to their D-isomers but favored D-isomers of aromatic
amino acids. The K values (L/mol) were estimated as 2.3
× 105 and 1.7 × 105 for dansyl-L- and -D-valine anions,
2.8 × 105 and 1.9 × 105 for dansyl-L- and -D-t-leucine
anions, 3.1 × 105 and 1.4 × 105 for dansyl-L- and
-D-leucine anions, 4.5 × 105 and 8.8 × 105 for dansyl-L-
and -D-phenylglycine anions; and 4.2 × 105 and 6.0 × 105
for dansyl-L- and -D-phenylalanine anions. The mole ratio
of n/m ranged from 0.30 to 0.67, and two possible guest
accommodation modes of intra- and inter-complex bind-
ings were postulated. The log D values were calculated
at pH ) 7.2 using the PALLAS program (version 3.0,
CompuDrug Chemistry Ltd.)15 as measures of hydropho-
bicity for dansylamino acid anions (Table 1). No clear
relationship between K values and log D values of the
guests was found, but aromatic amino acid anions with
large log D values were more strongly bound than
aliphatic amino acids with small log D values. The
highest L/D selectivity was recorded as 2.2 in the case of
leucine derivative, while the highest D/L one was esti-
mated as 2.0 with phenylglycine anion. As described
above, dansylamino acid anions were bound by cyclen
1-Na+ complex via electrostatic attraction and hydro-
phobic interaction. Since aromatic and aliphatic amino
acid derivatives exhibited different trends in K values
and enantiomer selectivity, aromatic residues of amino
acid anions provided further interactions with cyclen
metal complex.
Circular dichroism (CD) analysis provided structural
information of the dansylamino acid anions incorporated
in the chiral helicate aggregates. The employed dansyl-
amino acid anions changed CD signs upon incorporation.
Typically, dansyl-L-leucine anion exhibited an intense CD
signal around 260 nm with a negative sign in the aqueous
solution but a positive sign in the self-aggregate of cyclen
1-Na+ complex (Figure 6). In contrast, dansyl-D-leucine
anion gave a positive sign in the aqueous solution and a
negative sign in the self-aggregate. Polonski et al. had
reported that the Cotton effect observed with chiral
dansylamino acid was originated from asymmetrical
twisting of the sulfonamide group in relation to the
naphthalene plane under the influence of hydrogen atom
in the peri-position.16 Based on this model, the observed
inversion of the CD signs suggests the conformational
change between A and B in Figure 6. Since the most
bulky substituent R of the leucine anion predominantly
occupies the sterically least hindered position, dansyl-L-
leucine anion favors an “anti-clockwise” form to offer the
negative CD signal in the bulk aqueous solution (A in
Figure 6). When this anion is included in the self-
aggregate, the CO2- group becomes more bulky than the
substituent R, because of electrostatic interaction with
large cyclen 1-Na+ cation. This means that dansyl-L-
leucine anion has a “clockwise” form and gives the
positive CD signal in the self-aggregate (B in Figure 6).
Since the D-isomer has apparently reversed profiles, the
-
center of the cyclen complex and -CO2 group of the
dansylamino acid anion was essentially involved as well
as hydrophobic interaction between cholesterol moieties
and dansyl group in the aggregate inclusion processes.
4. Quantitative Studies of Anion Inclusion. The
inclusion behaviors of dansylamino acid anions with
cyclen 1-Na+ complex aggregate were quantitatively
analyzed using the following Langmuir-type equation
m[A]b/[1 - Na+]o ) n K [A]u/(1 + K [A]u)
where [A]b and [A]u are concentrations of bound and
unbound guest anions, and m and n are numbers of
cyclen 1-Na+ complex and saturated guests in the single
self-aggregate. According to this equation, K values were
estimated for enantiomers of dansyl-valine, tert-leucine,
leucine, phenylglycine, and phenylalanine by curve-
(14) (a) Palmans, A. R. A.; Vekemans, J. A. J. M.; Havinga, E. E.;
Meijer, E. W. Angew. Chem., Int. Ed. Engl. 1997, 36, 2648. (b) Prins,
L. J.; Timmerman, P.; Reinhoudt, D. N. J. Am. Chem. Soc. 2001, 123,
10153. (c) Brunsveld, L.; Lohmeijer, B. G. G.; Vekemans, J. A. J. M.;
Meijer, E. W. Chem. Commun. 2000, 2305.
(15) Tryar, N. E.; Tsai, R. S.; Carrupt, P. A.; Testa, B. J. Chem. Soc.,
Perkin Trans. 2 1992, 79.
(16) Polonski, T.; Chimiak, A.; Kochman, M. Tetrahedron 1974, 30,
641.
1840 J. Org. Chem., Vol. 70, No. 5, 2005

Cholesterol-Armed Cyclens
hanced fluorescence signals of the dansylamino acid
anions in the neutral aqueous solutions. This exhibited
larger fluorescence signal for L-leucine than D-leucine,
but smaller signal for L-t-leucine than D-t-leucine. Since
this protein has several different binding sites for sub-
strate accommodation, dansylamino acids were confirmed
to locate in the different cavities. Therefore, the present
type of cyclen metal complexes was demonstrated to be
unique building blocks for supramolecular aggregates,
which offered enantiomer recognition and visual sensing
of amino acid anions in aqueous solutions. The proper
organization of cholesterol-functionalized sidearms, oc-
tadentate chiral armed cyclen platform, and charged
metal center offered water-soluble aggregates with no-
table anion recognition and sensing functions. Since
similar phenomena were not observed at the molecular
level, self-aggregation of the helicates can be considered
an effective method of generating sophisticated function-
alities.
Experimental Section
Materials. Dansyl-L-valine, -L-leucine, and -L-phenylalanine
were commercially available and used after recrystallization
from CH3OH/H2O. Other dansylamino acids were synthesized
by the reported methods.19 Their optical purities were deter-
mined as >99% by chiral HPLC analysis (SUMICHIRAL
OA-3200). The [R]D values of the synthesized derivatives were
as follows: dansyl-D-valine, -22.1 (c ) 0.5, CH3OH); dansyl-
D-leucine, +29.2 (c ) 0.5, CH3OH); dansyl-L-tert-leucine, +56.2
(c ) 0.5, CH3OH); dansyl-D-tert-leucine, -55.5 (c ) 0.5,
CH3OH); dansyl-D-phenylalanine, +55.5 (c ) 0.5, CH3OH);
dansyl-L-phenylglycine, +50.6 (c ) 0.5, CH3OH); dansyl-D-
phenylglycine, -50.7 (c ) 0.5, CH3OH).
FIGURE 6. CD spectral changes of dansyl-L-leucine and its
preferred conformations (A) in aqueous solution and (B) in self-
aggregate of cyclen 1-NaCl aggregate.17
bound amino acid guest was thought to have a “frozen”
conformation in the self-aggregate.
1,4,7,10-Tetrakis(cholesteryloxycarbonylmethyl)-1,4,7,-
10-tetraazacyclododecane (1). After a solution of cyclen
tetrahydrochloride (644 mg, 2.02 mmol) and Cs2CO3 (9.80 g,
30.1 mmol) in CH3CN (15 mL) was refluxed for 1 h, a solu-
tion of cholesterol choloroacetate (5.58 g, 12.0 mmol) in CHCl3
(20 mL) was added dropwise. The resulting mixture was
refluxed for 5 h and then filtered. After solvent was evapo-
rated, the residue was washed with ether and recrystallized
from CHCl3/ether, CHCl3/acetone and then CHCl3/CH3CN
(14%): mp 196-202 °C decomp; [R]D ) -29.3 (c ) 0.5, CHCl3);
The cholesterol-armed cyclen metal complexes also
interacted with dansylamino acid anions in organic media
but exhibited no enantiomer selectivity. When an aque-
ous solution of racemic dansyl-L- and D-leucine anion (2.0
× 10-4 mol/L, each, 2.0 mL) was shaken with a CHCl3
solution of cyclen 1-NaCl complex (2.0 × 10-4 mol/L, 2.0
mL), the guest anion was quantitatively but not enan-
tiomer-selectively extracted. Although cyclen 1-Na+
complex has an asymmetric helical structure even in the
organic media, its nonaggregated form simply behaved
like a cationic surfactant and rarely responded to the
guest chirality.5 Thus, supramolecular alignments of the
helical complexes generated characteristic anion recogni-
tion and sensing functions. When cholesteroxycarbonyl-
4-methylmorpholine 3 (2.0 × 10-5 mol/L) was dispersed
with NaCl (5.0 × 10-6 mol/L) in aqueous ethanol solution
(20/80, v/v, pH ) 7.2), TEM experiments confirmed its
self-aggregation. Since the addition of this aggregate
rarely influenced the fluorescence spectra of the dansyl-
amino acid anions, it could not provide an effective
environment for anion inclusion. Albumin and γ-cyclo-
dextrin were also employed as receptors for dansylamino
acids.18 Although they are well-known to accommodate
aromatic guest anions, only albumin exhibited the en-
1
IR (KBr) ν 1738 cm-1 (br); H NMR (CDCl3) δ 0.68 (s, 12H),
0.80-2.20 (m, 104H), 0.85 (d, J ) 1.2 Hz, 12H), 0.87 (d, J )
1.5 Hz, 12H), 0.91 (d, J ) 6.1 Hz, 12H), 1.01 (s, 12H), 2.31 (d,
J ) 8.1 Hz, 8H), 2.82 (s, 16H), 3.36 (s, 8H), 4.66 (br m, 4H),
and 5.36 (br, 4H); 13C NMR (CDCl3) δ 11.85, 18.72, 19.32,
21.04, 22.55, 22,81, 23.85, 24.28, 27.93, 28.00, 28.33, 31.86,
31.91, 35.80, 36.19, 36.58, 37.02, 38.26, 39.51, 39.74, 42.31,
50.03, 52.07, 56.11, 56.16, 56.70, 73.96, 122.66, 139.59, and
171.14. Anal. Calcd for C124H2O4N4O8: C, 79.26; H, 10.94; N,
2.98. Found: C, 79.09; H, 10.97; N, 2.97.
1,4,7,10-Tetrakis(cholesteryloxycarbonylmethyl)-1,4,7,-
10-tetraazacyclododecane (1)-NaCl Complex. After a
solution of cyclen tetrahydrochloride (0.65 g, 2.05 mmol) and
Na2CO3 (3.22 g, 30.4 mmol) in CH3CN (15 mL) was refluxed
for 1.5 h, a solution of cholesterol chloroacetate (5.56 g, 12.0
mmol) in CHCl3 (40 mL) was added dropwise. The resulting
mixture was refluxed for 7 h and then filtered. After the
solvent was evaporated, the residue was washed with ether
and recrystallization from CHCl3/CH3CN gave white needle
crystals of 1-NaCl‚5H2O complex (71%): mp 151-152 °C dec;
[R]D ) -83.4 (c ) 0.5, CHCl3); IR (neat) ν 1725 cm-1; 1H NMR
(CDCl3) δ 0.70 (s, 12H), 0.80-2.40 (m, 112H), 0.86 (d, 12H, J
(17) In the presence of aggregate, the concentrations of bound and
unbound dansylleucine were estimated to be 0.8 × 10-4 and 1.0 × 10-4
M, respectively. The [θ] value used in the graph was obtained by
dividing the θ value by total concentration of dansylleucine, 1.8 ×
10-4 M.
(18) (a) Lightner, D. D.; Wijekoon, W. M. D.; Zhang, M. H. J. Biol.
Chem. 1988, 263, 16669. (b) Kano, K.; Nishiyabu, R.; Asada, T.;
Kuroda, Y. J. Am. Chem. Soc. 2002, 124, 9937.
(19) (a) Chimiak, A.; Polonski, T. Org. Prep. Proced. Int. 1973, 5,
117. (b) Ikeda, H.; Nakamura, M.; Ise, N.; Toda, F.; Ueno, A. J. Org.
Chem. 1997, 62, 1411.
J. Org. Chem, Vol. 70, No. 5, 2005 1841

Shinoda et al.
1.74 mmol) with concd HCl (1.5 mL) in MeOH (10 mL). After
stirring for 2 h, the solvent was removed. White hygroscopic
precipitate was obtained (571 mg, 65%) by adding Et2O to the
MeOH solution: IR (KBr) ν 1653, 1551, and 1071 cm-1 1H
;
NMR (D2O) δ 2.8 (br s, 8H), 2.9-3.1 (8H), 3.36 (s, 2H), 4.22
(s, 2H), 7.08 (d, 2H, J ) 8.1 Hz) and 7.41 (d, 2H, J ) 8.2 Hz);
13C NMR (D2O) δ 42.9, 43.3, 45.1, 50.4, 56.0, 129.7, 132.3,
137.4, and 173.8; MS (FAB, pos) m/z 398 (M + H+). Anal. Calcd
for C17H28N5OBr‚3HCl‚1.5H2O: C, 38.18; H, 6.41; N, 13.10.
Found: C, 38.33; H, 6.41; N, 13.08.
1-(4-Bromophenyl)methylcarbamoylmethyl-4,7,10-tris-
(cholesteryloxycarbonylmethyl)-1,4,7,10-tetraazacy-
clododecane (2)-NaCl complex was prepared as described
below. 1-(4-Bromophenyl)methylcarbamoylmethyl-1,4,7,10-
tetraazacyclododecane‚3HCl (151 mg, 0.298 mmol) and
Na2CO3 (328 mg, 3.09 mmol) were refluxed in CH3CN (7 mL)
for 1 h under N2. Cholesterol chloroacetate (631 mg, 1.36
mmol) dissolved in CHCl3 (10 mL) was added dropwise and
refluxed for 50 h. After solvent was removed, CH2Cl2 was
added, and the insoluble inorganic materials were filtered
using cerite. After evaporation, the residue was repeatedly
reprecipitated from CH2Cl2/hexane followed by centrifugation
to remove excess cholesterol chloroacetate. Colored impurities
were removed using charcoal (160 mg × 2) in CH2Cl2 solution
which finally gave colorless crystals from CH2Cl2/Et2O (230
mg, 42%): mp 198-200 °C; [R]27D ) -63 (c ) 0.50, CHCl3); IR
(KBr) ν 1732, 1668, and 1213 cm-1; 1H NMR (CDCl3, 50 °C) δ
0.68 (s, 3H), 0.70 (s, 3H × 2), 0.87 (d, 18H, J ) 6.0 Hz), 0.92
(d, 6H, J ) 6.7 Hz), 0.94 (d, 3H, J ) 6.4 Hz), 0.96 (s, 3H), 0.98
(s, 6H), 0.9-1.2 (33H), 1.2-1.7 (33H), 1.7-2.1 (15H), 2.1-3.4
(25H), 3.62 (s, 2H), 4.28 (br s, 1H), 4.48 (br s, 1H), 4.62 (m,
3H), 5.10 (br s, 1H), 5.31 (m, 2H), 7.24 (m, 2H), 7.31 (m, 2H),
and 9.99 (br s, 1H); 13C NMR (CDCl3, 50 °C) δ 11.9, 12.1, 18.8,
18.9, 19.0, 19.2, 19.3, 21.2, 21.3, 22.6, 22.8, 22.8, 23.9, 24.0,
24.1, 24.1, 24.4, 24.5, 27.5, 27.9, 28.0, 28.1, 28.1, 28.2, 28.3,
31.9, 31.9, 32.0, 32.1, 32.3, 32.5, 35.8, 35.9, 35.9, 36.0, 36.3,
36.4, 36.6, 36.6, 36.7, 37.0, 37.1, 37.3, 38.0, 38.2, 38.3, 39.7,
39.9, 40.0, 40.1, 42.4, 42.5, 42.5, 50.1, 50.3, 50.4, 55.2, 55.2,
56.2, 56.4, 56.5, 56.9, 56.9, 57.1, 57.2, 75.5, 75.2, 75.6, 120.2,
123.0, 123.1, 129.6, 131.2, 139.1, 139.4, 139.9, 172.6, and
172.9; MS (FAB, pos) m/z 1699 (2 + Na+). Anal. Calcd for
FIGURE 7. Synthesis of armed cyclen 2.
) 1.5 Hz), 0.88 (d, 12H, J ) 1.7 Hz), 0.92 (d, 12H, J ) 6.3 Hz),
0.99 (s, 12H), 2.15-2.35 (br m, 4H), 2.40-2.70 (br m, 8H), 2.98
(br d, 4H, J ) 17.3 Hz), 3.25-3.45 (br m, 4H), 3.45 (br d, 4H,
J ) 17.1 Hz), 4.53 (br m, 4H), and 5.26 (br 4H); 13C NMR
(CDCl3) δ 11.93, 18.69, 19.08, 21.03, 22.54, 22.77, 23.57, 24.26,
27.64, 27.97, 28.13, 31.63, 32.49, 35.69, 36.20, 36.36, 36.90,
38.15, 39.57, 39.90, 42.32, 48.46, 50.34, 53.01, 54.80, 56.07,
57.23, 75.48, 122.49, 139.71, and 173.49. Anal. Calcd for
C
124H204N4O8‚NaCl‚5H2O: C, 73.46; H, 10.64; N, 2.76. Found:
C, 73.76; H, 10.49; N, 2.76.
1-(4-Bromophenyl)methylcarbamoylmethyl-4,7,10-tris-
(cholesteryloxycarbonylmethyl)-1,4,7,10-tetraazacy-
clododecane (2)-NaCl Complex (See Figure 7). N-(4-
Bromophenyl)methyl-2-chloroethanamide20 was synthe-
sized by mixing a CH2Cl2 (100 mL) solution of 4-(bromophen-
yl)methylamine (1.96 g, 8.81 mmol) and triethylamine (2.24
g, 22.1 mmol) with a CH2Cl2 solution (50 mL) of 2-chloroacetyl
chloride (1.51 g, 13.4 mmol) at 0 °C. After 1.5 h stirring at 0
°C, the organic phase was washed with water (100 mL), 1 M
hydrochloric acid (100 mL × 2) and sat. NaCl solution (100
mL), evaporated, and dried over MgSO4. Recrystallization from
CH2Cl2/hexane gave needle crystals (1.33 g, 5.07 mmol): 58%
yield; mp 124-125 °C; IR (KBr) ν 3270, 1661, 1558, 1245, and
796 cm-1; 1H NMR (CDCl3) δ 4.11 (s, 2H), 4.45 (d, 2H, J ) 6.1
Hz), 6.88 (br s, 1H), 7.18 (m, 2H), and 7.48 (m, 2H); 13C NMR
(CDCl3) δ 42.6, 43.2, 121.7, 129.5, 131.9, 136.3, and 165.9; MS
(FAB, pos) m/z 262 (M + H+).
C104H166BrN5O7‚NaCl‚(H2O)3.5: C; 69.40, H; 9.69, N; 3.89.
Found: C; 69.19, H; 9.38, N; 3.82.
Synthesis of Cholesteryl Morpholinoacetate (3). This
was similarly prepared by reaction of morpholine and choles-
terol chloroacetate (65%): mp 123-124 °C; [R]25 ) -30.4 (c
D
) 2.0, CHCl3); IR (KBr) ν 1729 cm-1; 1H NMR (CDCl3) δ 0.68
(s, 3H), 0.85 (d, 3H, J ) 1.3 Hz), 0.87 (d, 3H, J ) 1.3 Hz), 0.91
(d, 3H, J ) 6.4 Hz), 1.02 (s, 3H), 0.8-2.1 (m, 26H), 2.33 (d,
2H, J ) 7.7 Hz), 2.59, 3.75 (t x 2, 8H, J ) 4.7 Hz), 3.18 (s,
2H), 4.66 (br m, 1H), and 5.38 (d, 1H, J ) 4.4 Hz); 13C NMR
(CDCl3) δ 11.84, 18.70, 19.29, 21.01, 22.54, 22.80, 23.81, 24.26,
27.78, 28.00, 28.20, 31.84, 31.89, 35.78, 36.16, 36.57, 36.93,
38.11, 39.50, 39.71, 42.30, 50.00, 53.28, 56.12, 56.67, 59.90,
66.80, 74.73, 122.82, 139.44, and 169.49. Anal. Calcd for
C33H55NO3: C, 77.14; H, 10.79; N, 2.73. Found: C, 77.01; H,
10.77; N, 2.73.
1-(4-Bromophenyl)methylcarbamoylmethyl-4,7,10-tris-
(tert-butyloxycarbonyl)-1,4,7,10-tetraazacyclodode-
cane was prepared by the reaction of 1,4,7-tris(tert-butyl-
oxycarbonyl)-1,4,7,10-tetraazacyclododecane (993 mg, 2.10
mmol)21 and N-(4-bromophenyl)methyl-2-chloroethanamide
(822 mg, 3.14 mmol) in the presence of K2CO3 (647 mg, 4.68
mmol) and KI (156 mg, 0.940 mmol) in dry CH3CN (20 mL).
After refluxing for 15 h under N2, the solvent was evaporated
and the residue was dissolved in CH2Cl2 and filtered. The
filtrate was chromatographed on silica gel with CH2Cl2/AcOEt
(1/5, v/v) to give 1.26 g (85%) of amorphous product: IR (KBr)
Preparation of Self-Aggregates. An ethanol solution of
cholesterol-armed cyclen 1-NaCl complex (0.50 µmol/1 mL)
was diluted with Bis-Tris-HCl buffer solution (6.3 × 10-3 M,
pH ) 7.2). The resulting aqueous solution (H2O/EtOH )
80/20, v/v) was spectroscopically clear and gave no precipitate
for several days. Light scattering experiments indicated that
the particle sizes of the self-aggregates were slightly changed
(50-70 nm) after aging, centrifugation or other procedures.
The self-aggregate of morpholine derivative 3 was similarly
prepared, though it did not complex with Na+ cation. Ca2+ and
1
ν 1689, 1540, 1250, and 1167 cm-1; H NMR (CDCl3) δ 1.38
(br s, 18H), 1.48 (s, 9H), 3.19 (s, 2H), 3.37 + 3.47 (br s, 12H),
4.37 (d, 2H, J ) 6.1 Hz), 7.17 (m, 2H), 7.28 (br s, 1H), and
7.43 (m, 2H); 13C NMR (CDCl3) δ 28.3, 28.5, 42.6, 47.6, 50.0,
60.3, 80.0, 121.0, 129.5, 131.6, 137.6, 155.6, and 171.2; MS
(FAB, pos) m/z 698 (M + H+).
1-(4-Bromophenyl)methylcarbamoylmethyl-1,4,7,10-
tetraazacyclododecane‚3HCl was synthesized by deprotec-
tion of 1-(4-bromophenyl)methylcarbamoylmethyl-4,7,10-tris-
(tert-butyloxycarbonyl)-1,4,7,10-tetraazacyclododecane (1.22 g,
Y
3+ complexes with cyclen 1 were in situ prepared. When 1.5
equiv of Ca2+ triflate was typically mixed with free cyclen 1,
the cation complexation process was monitored by ESI-MS and
NMR measurements. After evaporation, the ethanol solution
of the complex was diluted with aqueous buffer solution. Both
(20) Watanabe, S.; Kataoka, T. Jpn. Tokkyo Koho. 1969, JP 44026847.
(21) Kimura, E.; Aoki, S.; Koike, T.; Shiro, M. J. Am. Chem. Soc.
1997, 119, 3068.
1842 J. Org. Chem., Vol. 70, No. 5, 2005

Cholesterol-Armed Cyclens
complexes formed stable aggregates in Bis-Tris-HCl buffer
solutions (pH ) 7.2, H2O/EtOH ) 80/20, v/v). Since the Na+
complex with cyclen 2 required high ethanol contents to
disperse in the solutions, we usually employed aqueous ethanol
solution (H2O/EtOH ) 70/30, w/w) in this case.
K values listed in Table 1 (see Figure S2 in the Supporting
Information).
Liquid-Liquid Extraction. Determination of distribution
percentage for each dansylamino acid was determined by
adding a CHCl3 solution of the cyclen 1-NaCl complex (4.0 ×
10-4 mmol in 2.0 mL) to an aqueous solution of racemic
dansylamino acid (8.0 × 10-4 mmol in 2.0 mL, pH was adjusted
to be 12.2 with NaOH). After the mixture had been stirred for
2 h, the concentration of the dansylamino acid in the aqueous
phase was determined by UV spectroscopic method (monitored
at 320 nm) and its enantiomeric excess % was determined by
chiral HPLC method (SUMICHIRAL OA-3200, eluted with
0.01 mol/L CH3CO2NH4/CH3OH). We confirmed that negligible
amounts of dansylamino acids were distributed into the CHCl3
phase in the absence of cyclen 1-NaCl complex (<1%).
Molecular Modeling of Cyclen 1-Na+ Complex. CAChe
(ver.5.04, Fujitsu Ltd. Japan) was used for molecular modeling.
To obtain initial structures of the large cyclens, ∆- and Λ-types
of Na+ complexes with cyclen having four CH2COOH arms
were optimized using the CAChe MM2 force field. The four H
atoms on the sidearms were replaced by cholesterol arms to
obtain initial structures which were used for optimizing
complexes with the saucer-shaped open-form geometry as
shown in Figure 3. Five structures with different stewpan-
shaped closed-form geometry were extracted from trajectories
of MD simulations of 40 ps at 298 K. MM2 calculations were
performed to optimize the cyclen 1-Na+ complex.
Acknowledgment. We are grateful to Professor
Atsushi Yoshizawa of Hirosaki University and Dr. You
Shimidzu of NAIST, Japan, for their valuable comments
on the characterization of self-aggregates. This research
was supported in part by Grants-in-Aid for Scientific
Research (Nos. 14044090 and 16080217) from the
Ministry of Education, Culture, Sports, Science and
Technology, Japan.
Fluorescence and CD Measurements. Fluorescence
experiments were carried out in a 20% ethanol aqueous
solution (Bis-Tris-HCl buffer solution: 6.3 × 10-3 M; pH )
7.2). The initial concentrations of self-aggregate and guest
dansylamino acid anions are shown in each Table and Figure.
The fluorescence spectra of the dansylamino acid anions were
recorded upon excitation at 327 nm. CD experiments were
similarly performed.
Langmuir-Type Binding Studies. Dansylamino acid (1.0
× 10-6-4.0 × 10-5 mol/L) was added to an aqueous solution
of cyclen 1 - NaCl complex aggregate (3.0 × 10-5 mol/L). The
fluorescence spectral changes of each dansylamino acid were
observed by excitation at 327 nm in H2O/EtOH (80/20, pH )
7.2) and analyzed based on the equation described above. The
obtained Langmuir-type plots were employed to estimate the
Supporting Information Available: CD spectral changes
of cyclen 2-NaCl complex and Langmuir-type analysis data
for inclusion of dansylamino acids. This material is available
free of charge via the Internet at http://pubs.acs.org.
JO0478297
J. Org. Chem, Vol. 70, No. 5, 2005 1843