Molecular recognition studies on supramolecular systems. 32. Molecular recognition of dyes by organoselenium-bridged bis(β-cyclodextrin)s

Article abstract of DOI:10.1021/jo001372t

A series of novel bis(β-cyclodextrin)s tethered with organoselenium linkers, i.e., 6,6′-(o-phenylenediseleno)-bridged bis(β-cyclodextrin) (2), 6,6′-[2,2′-diselenobis(benzoyloxy)]-bridged bis(β-cyclodextrin) (3), and 6,6′-[2,2′-diselenobis[2-(benzoylamino)ethylamino]]-bridged bis(β-cyclodextrin) (4), were synthesized from β-cyclodextrin (1). The inclusion complexation behavior of 1-4 with some dyes, such as 8-anilinonaphthalenesulfonate (ANS), Brilliant Green, Crystal Violet, Tropaeolin OO, Auramine O, and Methyl Orange, was investigated in aqueous phosphate buffer solution (pH 7.20) at 25 °C by UV-vis, fluorescence, and circular dichroism spectrometry, as well as fluorescence lifetime measurements. The complex stability constants (K_S) and Gibbs free energy changes (δG°) for the stoichiometric 1:1 inclusion complexation of 1-4 with the dyes were obtained by the spectrophotometric or spectropolarimetric titrations. The bis(β-cyclodextrin)s 2-4 showed much higher affinities toward these guest dyes than native β-cyclodextrin I with fairly good molecular selectivities. The cooperative binding abilities of these bis(β-cyclodextrin)s are discussed from the viewpoints of size/shape-fit interaction, induced-fit concept, and multiple recognition mechanism.

Full text of DOI:10.1021/jo001372t

J . Org. Chem. 2001, 66, 225-232
225
Molecu la r Recogn ition Stu d ies on Su p r a m olecu la r System s. 32.1
Molecu la r Recogn ition of Dyes by Or ga n oselen iu m -Br id ged
Bis(â-cyclod extr in )s
Yu Liu,*,†,‡ Bin Li,†,§ Chang-Cheng You,† Takehiko Wada,§ and Yoshihisa Inoue*,‡,§
Department of Chemistry, Nankai University, Tianjin 300071, China, Inoue Photochirogenesis Project,
ERATO, J ST, 4-6-3 Kamishinden, Toyonaka, Osaka 565-0085, J apan, and Department of Molecular
Chemistry, Faculty of Engineering, Osaka University, 2-1 Yamadaoka, Suita 565-0871, J apan
yuliu@public.tpt.tj.cn
Received September 15, 2000
A series of novel bis(â-cyclodextrin)s tethered with organoselenium linkers, i.e., 6,6′-(o-phenylene-
diseleno)-bridged bis(â-cyclodextrin) (2), 6,6′-[2,2′-diselenobis(benzoyloxy)]-bridged bis(â-cyclodextrin)
(3), and 6,6′-[2,2′-diselenobis[2-(benzoylamino)ethylamino]]-bridged bis(â-cyclodextrin) (4), were
synthesized from â-cyclodextrin (1). The inclusion complexation behavior of 1-4 with some dyes,
such as 8-anilinonaphthalenesulfonate (ANS), Brilliant Green, Crystal Violet, Tropaeolin OO,
Auramine O, and Methyl Orange, was investigated in aqueous phosphate buffer solution (pH 7.20)
at 25 °C by UV-vis, fluorescence, and circular dichroism spectrometry, as well as fluorescence
lifetime measurements. The complex stability constants (KS) and Gibbs free energy changes (∆G°)
for the stoichiometric 1:1 inclusion complexation of 1-4 with the dyes were obtained by the
spectrophotometric or spectropolarimetric titrations. The bis(â-cyclodextrin)s 2-4 showed much
higher affinities toward these guest dyes than native â-cyclodextrin 1 with fairly good molecular
selectivities. The cooperative binding abilities of these bis(â-cyclodextrin)s are discussed from the
viewpoints of size/shape-fit interaction, induced-fit concept, and multiple recognition mechanism.
In tr od u ction
cyclodextrin dimers have been synthesized to investigate
the inclusion complexation behavior which is significantly
different from that of the monocyclodextrin counterparts.
Such studies have helped us understanding the multiple
recognition mechanism and the induced-fit interactions
between the host bis(cyclodextrin)s and guest mole-
cules.10-17 Recently, the molecular recognition behavior
of cyclodextrin dimers has been reviewed comprehen-
sively by Breslow and co-workers.18 However, little at-
tention has been paid to the molecular recognition by
organoselenium-bridged bis(cyclodextrin)s as synthetic
receptors. Recently, we have reported a study on the
molecular recognition of some dyes by several organo-
selenium bridged bis(â-cyclodextrin)s and their platinum-
(IV) complexes, which has provided some insights into
the cooperation of several weak interactions working
between host and guest.19,20
Extensive studies on molecular recognition by native
and modified cyclodextrins as molecular receptors have
documented that the truncated corn-shaped hydrophobic
cavity of cyclodextrins can selectively bind diverse sub-
strates to form host-guest complexes or supramolecular
species,2-4 which provide an excellent model system
mimicking the substrate-specific interaction of enzymes.5
Possessing a good structural variety, dyes have been
widely investigated as typical guest molecules, which can
form stable inclusion complexes with cyclodextrins.6-9 As
a new family of cyclodextrin derivatives, bridged cyclo-
dextrin dimers are known to greatly enhance the original
molecular binding ability of native cyclodextrins through
the cooperative binding of dual hydrophobic cavities
located in a close vicinity. Consequently, a number of
We now wish to report our study on the syntheses and
molecular recognition behavior of organoselenium-bridged
â-cyclodextrin dimers (2-4), shown in Chart 1. A simple
* To whom correspondence should be addressed. Tel: +86-22-
23503625. Fax: +86-22-23504853.
† Nankai University.
‡ ERATO.
§ Osaka University.
(1) Part 30: J in, L.; S.-X., Sun; Liu, Y. Chem. J . Chin. 2000, 21,
412.
(10) Zhang, B.; Breslow, R. J . Am. Chem. Soc. 1993, 115, 9353.
(11) Venema, F.; Rowan, A. E.; Nolte, R. J . M. J . Am. Chem. Soc.
1996, 118, 257.
(12) French, R. R.; Wirz, J .; Woggon, W.-D. Helv. Chim. Acta 1998,
81, 1521.
(13) Breslow, R.; Zhang B. J . Am. Chem. Soc. 1996, 118, 8495.
(14) Breslow, R.; Zhang B. J . Am. Chem. Soc. 1992, 114, 5882.
(15) Zhang B.; Breslow, R. J . Am. Chem. Soc. 1997, 119, 1676.
(16) Haskard, C. A.; Easton, C. J .; May, B. L.; Lincoln, S. F. J . Phys.
Chem. 1996, 100, 14457.
(2) Wenz, G. Angew. Chem., Int. Ed. Engl. 1994, 33, 803.
(3) (a) Szejtli, J . Cyclodextrins and Their Inclusion Complexes;
Akademia Kiado: Budapest, 1982. (b) Szejtli, J .; Osa, T. In Compre-
hensive Supramolecular Chemistry; Atwood, J . L., Davies, J . E. D.,
MacNicol, D. D., Vo¨gtle, F., Eds; Elsevier: Oxford, U.K., 1996; Vol. 3.
(4) Nepogodiev, S. A.; Stoddart, J . F. Chem. Rev. 1998, 98, 1959.
(5) Breslow, R.; Dong, S. D. Chem. Rev. 1998, 98, 1997. Breslow, R.
Acc. Chem. Rev. 1995, 28, 146.
(6) Hirai, H.; Toshima, N.; Uenoyama, S Bull. Chem. Soc. J pn. 1985,
58, 1156.
(7) Matsui, Y.; Ogawa, K.; Mikami, S.; Yoshimoto, M.; Mochida, K.
Bull. Chem. Soc. J pn. 1987, 60, 1219.
(8) (a) Buss, V. Angew. Chem., Int. Ed. Engl. 1991, 30, 869. (b) Buss,
V.; Reichardt, C. J . Chem. Soc., Chem. Commun. 1992, 1636.
(9) Choppinet, P.; J ullien, L.; Valeur, B. Chem. Eur. J . 1999, 5, 3666
(17) Chiu, S.-H.; Myles, D. C.; Garrell, R. L.; Stoddart, J . F. J . Org.
Chem. 2000, 65, 2792.
(18) Breslow, R.; Halfon, S.; Zhang, B. Tetrahedron 1995, 51, 377.
(19) Liu, Y.; Li, B.; Wada, T.; Inoue, Y. Supramol. Chem. 1999, 10,
279.
(20) Liu, Y.; You, C.-C.; Chen, Y.; Wada, T.; Inoue, Y. J . Org. Chem.
1999, 64, 7781.
10.1021/jo001372t CCC: $20.00 © 2001 American Chemical Society
Published on Web 12/16/2000

226 J . Org. Chem., Vol. 66, No. 1, 2001
Liu et al.
Ch a r t 1
Ch a r t 2
reason for choosing the (di)selenium tether in bis(â-
cyclodextrin)s is that selenium, possessing a larger radius
and lower electronegativity than carbon, can provide an
Se-Se bond that is longer and more flexible than a C-C
bond. A more serious reason is that selenium compounds
are known to function as mimics of glutathione peroxi-
dase. Therefore, the molecular recognition studies on
selenium-bridged bis(cyclodextrin)s will be essential for
understanding the interaction between the cyclodextrin-
based selenium-containing enzyme model and substrates.
The inclusion complexation behavior has been investi-
gated at 25 °C in aqueous phosphate buffer solution (pH
7.20) by means of spectrophotometry, spectrofluorimetry,
and spectropolarimetry as well as fluorescence lifetime
measurement. The complex stability constants (log KS)
and Gibbs free energy changes (∆G°) obtained for some
structurally related dye molecules, shown in Chart 2, are
discussed from the viewpoints of the cooperative binding
and the induced-fit interaction between the guest and
dimeric host.
Exp er im en ta l Section
In str u m en ts. Combustion analyses were performed on a
Perkin-Elmer-240 instrument. 1H NMR spectra were recorded
in dimethyl sulfoxide-d6 on a Bruker AM200 instrument
operated at 200 MHz. FT-IR spectra were obtained on a Nicolet
560 E.S.P. FT-IR spectrometer. Circular dichroism (CD)
spectra were measured in a conventional quartz cell (10 × 10

Molecular Recognition Studies on Supramolecular Systems
J . Org. Chem., Vol. 66, No. 1, 2001 227
× 45 mm) on a J ASCO J -720W spectropolarimeter equipped
p lexes. It has been amply demonstrated that inclusion
of chromophoric achiral guest in a chiral host such as
cyclodextrin produces induced circular dichroism (ICD)
signals at the wavelengths absorbed by the guest
chromophore.23-25 The sign and magnitude of the ICD
signals are critical functions of the relative orientation
of the chromophore against the host cavity. An empirical
rule that interprets the ICD observed for a guest posi-
tioned inside or outside of the cyclodextrin cavity has
been proposed by Kajta´r et al.,26 Harata and Uedaira,27
and Kodaka.28 This widely accepted sector rule predicts
that, when a guest molecule is accommodated in the
cavity, the transition dipole moment of the guest chro-
mophore, which is parallel to the host axis, produces a
positive ICD band, whereas the transition moment
perpendicular to the axis gives a negative ICD.27 In
contrast, a guest located outside the cavity is known to
show the completely opposite ICD behavior. Thus, the
transition dipole moment parallel to the host axis affords
a negative ICD signal, while the perpendicular transition
gives the opposite signal.28 Our previous study29,30 has
shown that this rule can be applied successfully to the
analysis the conformation of the chromophore attached
to cyclodextrin, although little effort has been devoted
to the conformational analysis of cyclodextrin dimers.
Here we try to explain the ICD signals of dimeric hosts
2-4 using the above rule.
As can be seen from Figure 1, hosts 2-4 display
substantially different CD spectra in the absence of guest,
indicating that there exist significant, but different
degrees of, interactions between the aromatic tether and
the two chiral cavities of cyclodextrin dimer. As we
reported previously,19 the CD spectrum of dimeric host
2 in aqueous solution shows a strong negative Cotton
effect peak at 259 nm, which is ascribed to the substitu-
ent effect of the diseleno moiety and also to the coopera-
tive binding by dual cyclodextrin cavities. It is interesting
to compare the CD spectral behavior of hosts 3 and 4.
Host 3 displays two negative Cotton effect peaks at 233
nm (∆ꢀ ) -4.03) and 266 nm (∆ꢀ ) -0.32) and a weak
positive peak at 325 nm (∆ꢀ ) 0.35). As a higher
homologue of 3, â-cyclodextrin dimer 4 gives the opposite
Cotton effects; i.e., a positive Cotton effect peak at 232
nm (∆ꢀ ) 27.6) with a shoulder at 260 nm (∆ꢀ ) 8.64)
and a weak negative peak at 318 nm (∆ꢀ ) -1.43).
According to the sector rule,26-28 we can deduce that the
benzene ring in the tether of host 3 is shallowly self-
included in the cavity, where both of the transition
moments of the 1La and 1Lb bands at 233 and 266 nm
are nearly perpendicular to the axis of cyclodextrin,
resulting in the two negative Cotton effect peaks. On the
other hand, the Se-Se moiety is located outside the
cavity, but situated between the two primary rims of dual
cyclodextrin cavities. Hence, the transition dipole of the
Se-Se bond is perpendicular to the host axis, giving the
with a PTC-348WI temperature controller to keep the tem-
perature at 25 °C. UV-vis spectra were recorded in
a
conventional quartz cell (10 × 10 × 45 mm) at 25 °C on a
J ASCO UV-550 spectrometer.
Fluorescence lifetimes were determined by the time-cor-
related single-photon-counting method using a Horiba NAES-
550 instrument with a time resolution of 0.5 ns. A self-
oscillating discharge lamp filled with hydrogen gas was
employed as the pulsed light source, and the excitation light
was made monochromatic by a 10 cm monochromator. The
emission from the sample was passed through an appropriate
filter (Toshiba UV-33) placed before the detector unit in order
to eliminate scattered excitation light. Maximum counts of up
to 10 000 were collected in each measurement. The ac-
cumulated signals were then processed and the lifetime
determined by deconvolution with nonlinear least-squares fit.
Ma ter ia ls. 8-Anilinonaphthalenesulfonate (ANS), Brilliant
Green, Crystal Violet, Tropaeolin OO, Auramine O, and
Methyl Orange were purchased from Wako. All chemicals were
reagent grade and used without further purification unless
noted otherwise. â-Cyclodextrin of reagent grade (Shanghai
Reagent Works) was recrystallized twice from water and dried
in vacuo at 95 °C for 24 h prior to use. N,N-Dimethylforma-
mide (DMF) was dried over calcium hydride for 2 days and
then distilled under reduced pressure prior to use. 6,6′-o-
Phenylenediseleno-bridged bis(â-cyclodextrin) (2) was prepared
according to the reported procedures.19 Disodium hydrogen
phosphate and sodium dihydrogen phosphate were dissolved
in distilled, deionized water to make a 0.10 M phosphate buffer
solution of pH 7.20, which was used in spectral measurements.
Syn th esis of 6,6′-[2,2′-Diselen obis(ben zoyloxyl)]-Br idged
Bis(â-cyclod extr in ) (3). To a solution of DMF (100 mL)
containing 2,2′-diselenobis(benzoic acid)21 (0.4 g) and dicyclo-
hexylcarbodiimide (DCC) (0.33 g) was added 2.5 g of â-cyclo-
dextrin and 25 mL of dry pyridine in the presence of molecular
sieves 4 Å. The resultant mixture was stirred for 12 h in an
ice bath and another 18 h at room temperature and then
allowed to stand for 3 days until no more precipitation
deposited. The precipitate was removed by filtration, and the
filtrate was evaporated to dryness under reduced pressure. The
residue was dissolved in a minimum amount of hot water and
then poured into 150 mL of acetone. The precipitate formed
was collected by filtration to obtain a white powder, which was
purified on a column of Sephadex G-25 to give 0.5 g (19% yield)
1
of 3 as a light yellow solid. FAB-MS: m/z 2636 (M + H+). H
NMR (DMSO-d6, TMS): δ 2.9-4.1 (m, 80H), 4.2-4.6 (m, 4H),
4.8-5.1 (m, 14H), 7.3-8.0 (m, Ar 8H). IR (KBr): ν 3405.7,
2925.8, 2150.3, 1638.8, 1415.1, 1369.3, 1336.2, 1302.1, 1257.1,
1200.9, 1156.9, 1079.8, 1028.9, 972.4, 938.0, 860.3, 755.9,
707.5, 609.0, 579.0, 530.5 cm-1. Anal. Calcd for C98H146O72Se2‚
7H2O: C, 40.78; H, 6.03. Found: C, 40.83; H, 6.32.
Syn th esis of 6,6′-[2,2′-Diselen obis[2-(ben zoyla m in o)-
eth ylen ea m in o]]-Br id ged Bis(â-cyclod extr in ) (4). â-Cy-
clodextrin dimer 4 was prepared in 25% yield from 2,2′-
diselenobis(benzoic acid)21 and mono[6-(2-aminoethyleneamino)-
6-deoxy]-â-cyclodextrin,22 according to the similar procedures
described above. FAB-MS m/z: 2722 (M + H+). 1H NMR
(DMSO-d6, TMS): δ 2.9-4.1 (m, 88H), 4.2-4.6 (m, 4H), 4.8-
5.1 (m, 14H), 7.3-8.0 (m, Ar 8H). IR (KBr): ν 3384.7, 2928.5,
1635.2, 1541.0, 1403.3, 1335.0, 1302.8, 1258.0, 1200.1, 1155.7,
1079.2, 1028.3, 946.2, 856.6, 753.5, 706.8, 578.9, 530.2 cm-1
.
Anal. Calcd for C102H158O70N4Se2‚8H2O: C, 42.79; H, 6.08; N,
1.96. Found: C, 42.98; H, 6.20; N, 2.04.
(23) Connors, K. A. Chem. Rev. 1997, 97, 1325.
(24) Rekharsky, M. V.; Inoue, Y. Chem. Rev. 1998, 98, 1875.
(25) Zhdanov, Y. A.; Alekseev, Y. E.; Kompantseva, E. V.; Vergeichik,
E. N. Russ. Chem. Rev. 1992, 61, 563.
Resu lts a n d Discu ssion
(26) Kajta´r, M.; Horvath-Toro, C.; Kuthi, E.; Szejtli, J .Acta Chim.
Acad. Sci. Hung. 1982, 110, 327.
In d u ced Cir cu la r Dich r oism Sp ectr a of Cyclo-
(27) Harata, K.; Uedaira, H. Bull. Chem. Soc. J pn. 1975, 48, 375.
(28) Kodaka, M. J . Am. Chem. Soc. 1993, 115, 3702.
(29) Inoue, Y.; Yamamoto, K.; Wada, T.; Everitt, S.; Gao, X.-M.; Hou,
Z.-J .; Tong, L.-H.; J iang, S.-K.; Wu, H.-M. J . Chem. Soc., Perkin Trans.
2 1998, 1807.
(30) Tong, L.-H.; Hou, Z.-J .; Inoue, Y.; Tai, A. J . Chem. Soc., Perkin
Trans. 2 1992, 1253.
d extr in Dim er s (2-4) a n d Th eir In clu sion Com -
(21) Kamigata, N.; Iizuka, H.; Izuoka, A.; Kobayashi, M. Bull. Chem.
Soc. J pn. 1986, 59, 2179.
(22) May, B. L.; Kean, S. D.; Easton, C. J .; Lincoln, S. F. J . Chem.
Soc., Perkin Trans. 1 1997, 3157.

228 J . Org. Chem., Vol. 66, No. 1, 2001
Liu et al.
F igu r e 1. (a) Circular dichroism and (b) UV-vis absorption
spectra of 6,6′-(o-phenylenediseleno)-bridged bis(â-cyclodex-
trin) 2, 6,6′-[2,2′-diselenobis(benzoyloxy)]-bridged bis(â-cyclo-
dextrin) 3, and 6,6′-[2,2′-diselenobis[2-(benzoylamino)ethyl-
amino]]-bridged bis(â-cyclodextrin) 4 (0.1 mM) in phosphate
buffer solution (pH 7.2) at 25 °C.
F igu r e 2. Induced circular dichroism spectra of Methyl
Orange (10.7 µM) (a) in the absence and in the presence of (b)
â-cyclodextrin 1 (1.5 mM) and (c) 6,6′-(o-phenylenediseleno)-
bridged bis(â-cyclodextrin) 2, 6,6′-[2,2′-diselenobis(benzoyl-
oxy)]-bridged bis(â-cyclodextrin) 3, and 6,6′-[2,2′-diselenobis-
[2-(benzoylamino)ethylamino]]-bridged bis(â-cyclodextrin) 4
(0.25 mM) in aqueous buffer solution at pH 7.20.
positive Cotton effect at 325 nm. However, it is difficult
to be self-included into the cavity for the benzene ring of
host 4, which is tethered to each cyclodextrin with a
longer linker. Consequently, the transition moments of
1
1
the La and Lb bands become perpendicular to the axis
of cyclodextrin, affording two strong positive Cotton
effects. On the contrary, the transition dipole moment
of the Se-Se bond in host 4 is inferred to be parallel to
the host axis, as the corresponding transition at 318 nm
gives a negative Cotton effect.
F igu r e 3. Illustration of the transition dipole moment for the
πfπ* (solid arrow) and nfπ* (open arrow) electronic transi-
tions of the azo chromophore of Methyl Orange.
Using the CD spectrometric titration technique, the
inclusion complexation of â-cyclodextrin derivatives with
some azo dye guests has been investigated.28,31 We have
also reported that native cyclodextrin and some cyclo-
dextrin dimers give distinctly different ICD signals upon
inclusion complexation of Methyl Orange.20 In this study,
we further investigate the inclusion behavior of Methyl
Orange with the other cyclodextrin dimers linked by
different organoselenium tethers. As reported previ-
ously,20 native â-cyclodextrin 1 can induce an appreciable
positive CD signal at 420 nm, which corresponds to the
πfπ* transition band of the azo group of Methyl Orange
(Figure 2, trace b), while no CD signals were observed
in the absence of cyclodextrin (Figure 2, trace a). The
positive CD signal observed indicates that Methyl Orange
molecule penetrates into the cavities in the longitudinal
direction,26,28 where the πfπ* transition moment of the
azo chromophore in Methyl Orange is parallel to the host
axis, exhibiting a positive ICD signal at 420 nm.
Orange (11 µM) caused by host 2 (50.6 µM) shows a
strong negative Cotton effect at 467 nm (∆ꢀ ) -3.82) and
a moderate positive peak at 408 nm (∆ꢀ ) 2.19). However,
host 3 (0.3 mM) induced a weak positive Cotton effect at
440 nm (∆ꢀ ) 0.56) and a weak negative Cotton effect at
369 nm (∆ꢀ ) -0.94) (Figure 2, trace d), while host 4
(0.3 mM) also induced a weak positive Cotton effect at
423 nm (∆ꢀ ) 0.40) and a weak negative peak at 377 nm
(∆ꢀ ) -0.26) (Figure 2, trace e). Although the complete
and comprehensive rationalization of these spectral
features would be difficult, one possible explanation is
that the broad absorption band of Methyl Orange at 350-
500 nm is composed of two allowed transitions and their
transition dipole moments are perpendicular to each
other. Indeed, the transition moment of the πfπ* (paral-
lel to the NdN bond)28 and nfπ* (perpendicular to the
NdN bond)31 of azo chromophore are perpendicular to
each other, as illustrated in Figure 3. The inverted sign
of the split ICD spectra observed for host 2 and hosts 3
and 4 and the substantially weak ICD intensities for the
latter two hosts would also be accounted for in terms of
the depth of guest penetration, or the host-guest dis-
tance. Possessing the shortest tether, host 2 can include
most part of the azo chromophore in the cavities, giving
rise to the strong ICDs for both πfπ* (parallel to the
Compared with native â-cyclodextrin 1, the bridged bis-
(â-cyclodextrin)s 2-4 possessing dual hydrophobic cavi-
ties, displayed completely different ICD spectral behavior
upon complexation of dyes, giving two ICD extreme with
the opposite sign for each host-guest combination. As
shown in Figure 2 (trace c), the ICD spectrum of Methyl
(31) Zhang, X.-Y.; Nau, W. M. Angew. Chem., Int. Ed. 2000, 39, 544.

Molecular Recognition Studies on Supramolecular Systems
J . Org. Chem., Vol. 66, No. 1, 2001 229
Ta ble 1. F lu or escen ce Lifetim e (τ) a n d Rela tive
Qu a n tu m Yield (Φ) of Am m on iu m
8-An ilin o-1-n a p h th a len esu lfon a te (ANS) in th e P r esen ce
a n d Absen ce of Na tive â-Cyclod extr in a n d
Bis(â-cyclod extr in )s 1-4
ANS/µM host equiv τS/ns ΦS/% τL/ns ΦL/%
ø2
ref
10
500
10
250
10
none
none
0.4
0.4
0.5
1.5
2.9
0.6
1.2
100
100
96.5
67.6
45.6
88.1
1.42
1.46
3.5 1.00
a
b
a
b
b
a
a
1
1
2
3
4
40
10
40
20
20
3.1
3.2 32.4 1.24
9.9 54.4 1.11
7.4 11.9 1.04
10
10
75.2 10.1 24.8 1.46
a
This work. Measured in the presence of host in aqueous buffer
solution of pH 7.20 at 25.0 °C; the fluorescence decay profile could
be successfully deconvoluted by double exponential function, giving
short and long lifetimes, denoted by subscripts S and L, respec-
b
tively, along with the fitting parameter ø2. Reference 20.
host axis) and nfπ* (perpendicular to the host axis) with
the signs predicted for the chromophore inside the cavity
from the sector rule. On the contrary, hosts 3 and 4 are
linked by longer tethers and therefore most part of the
azo chromophore cannot be fully accommodated in the
cavities, resulting in the weak ICDs with the opposite
signs, which are anticipated for the chromophore outside
the cavity.
F lu or escen ce Lifetim e. Bright32 and Reinsborough33
have demonstrated that the fluorescent dyes accom-
modated in the hydrophobic cavity of cyclodextrin leads
to fluorescence enhancement and peak shifts19 as well
as significantly elongated fluorescence lifetimes. Re-
cently, we have reported the fluorescence spectra and
lifetimes of ANS in the presence of native cyclodextrin
and some cyclodextrin dimers, which enabled us to
elucidate the complex structure.19,20 In the present study,
we performed the nanosecond time-resolved fluorescence
measurements with ANS in aqueous buffer solution (pH
7.20) in the presence or absence of â-cyclodextrin 1 and
bis(â-cyclodextrin)s 2-4 in order to assess the micro-
environmental polarity around the included ANS and to
understand the inclusion complexation behavior of host
compounds in further detail.
F igu r e 4. (A) ICD spectral changes of phosphate buffer
solution of Methyl Orange (10.7 µM) in the presence of 6,6′-
(o-phenylenediseleno)-bridged bis(â-cyclodextrin) 2. The con-
centration of host 2 (from a to n): 0, 10, 20, 30, 40, 50.6, 101.1,
151.7, 202.2, 252.8, 303.4, 353.9, 404.4, and 455 µM, respec-
tively. (B) Least-squares curve-fitting analysis for the com-
plexations of Methyl Orange with 2.
that the ANS molecule is located in a more hydrophobic
environment rather than in the bulk water, forming a
1:1 inclusion complex with â-cyclodextrin. Interestingly,
the shorter lifetime of ANS (τS ) 2.9 ns) in the presence
of 2 is in good agreement with the longer lifetime (τL )
3.1-3.2 ns) in the presence of 1. The significantly long
τS (2.9 ns) and much longer τL (9.9 ns) of ANS in the
presence of 2 may indicate the coexistence of “intramo-
lecular” 1:1 and 2:1 host-guest complexes, in which the
ANS molecule is accommodated in one and two cavities
of host 2, respectively.20 Similarly, the τS’s of ANS (0.6-
1.2 ns) in the presence of hosts 3 and 4 are appreciably
longer than the original τ of ANS (0.4 ns) of ANS in water
but shorter than that (2.9 ns) obtained with 2, while the
τL’s of ANS (7.4-10.1 ns) in the presence of 3 and 4 are
comparable to that in the presence of 2 (9.9 ns). As higher
hydrophobicity around the fluorophore leads in general
to a longer lifetime (τL), we may deduce that host 4 with
a longer tether gives the most hydrophobic environment
around ANS upon “sandwich” complexation. The smaller
relative fluorescence intensities (ΦL/ΦS) for 3 and 4 than
for 1 may indicate that the longer tethers in 3 and 4 are
unfavorable in general for the sandwich complexation.
Sp ectr op ola r im etr ic Titr a tion s. The quantitative
ICD spectral study enables us not only to elucidate the
conformation of the chromophore attached to the host but
also to determinate the binding constants for various
guests. Thus, the addition of hosts 1-4 to an aqueous
dye solution induced substantial CD spectral changes at
wavelengths that are absorbed by the dye. As can be seen
from Figure 4A, the stepwise addition of host 2 of up to
Since the rates of complexation/decomplexation are
much slower than that of the fluorescence decay, the
decay profile of fluorescence intensity (F(t)) can be
described as a sum of unimolecular decays for all
fluorescing species present in the solution
n
F(t) ) A exp(-t/τ ) (n ) 1, 2, etc.)
∑
i
i
i)1
where Ai and τi represent the initial abundance and
lifetime of the ith species. In the absence of host, the
fluorescence decay curve observed for ANS in aqueous
buffer solution was perfectly fitted to a single-exponential
function, giving a lifetime of 0.4 ns. On the contrary, the
decay profile of ANS in the presence of â-cyclodextrin or
bis(â-cyclodextrin)s could be analyzed only by a linear
combination of two exponential functions. The short and
long fluorescence lifetimes (τS and τL) and relative
quantum yields (Φ) observed for ANS in the presence of
1-4 are summarized in Table 1. The longer lifetime of
ANS (3.1-3.2 ns) in the presence of 1 clearly indicated
(32) Bright, F. V.; Catena, G. C. Anal. Chem. 1989, 61, 905.
(33) J obe, D. J .; Verrall, R. E.; Paleu, R.; Reinsborough, V. C. J .
Phys. Chem. 1988, 92, 3582.

230 J . Org. Chem., Vol. 66, No. 1, 2001
Liu et al.
Ta ble 2. Com p lex Sta bility Con sta n t (Ks) a n d Gibbs F r ee En er gy Ch a n ge (-∆G°) for 1:1 In clu sion Com p lexa tion of
Va r iou s Gu est Dyes w ith â-Cyclod extr in a n d Bis(â-cyclod extr in )s (1-4) in Aqu eou s Bu ffer Solu tion (p H 7.20) a t 25.0 °C
host
guest
KS/M-1
KS(X)/KS(1)a
log Ks
-∆G°/kJ mol-1
R
methodb
ref
1
Brilliant Green
Crystal Violet
Methyl Orange
Auramine O
Tropaeolin OO
ANS
Brilliant Green
Crystal Violet
Methyl Orange
Auramine O
ANS
Brilliant Green
Crystal Violet
Methyl Orange
Auramine O
Brilliant Green
Crystal Violet
Methyl Orange
Tropaeolin OO
2190
1870
3560
819
5530
≡1
3.34
3.27
3.55
2.91
3.74
2.01
4.10
4.52
4.81
4.14
3.11
3.49
4.31
4.44
3.52
4.12
4.18
4.17
4.17
19.1
18.7
20.3
16.6
21.4
11.5
23.4
25.8
27.4
23.6
17.7
19.9
24.6
25.3
20.1
23.5
23.9
23.8
23.8
13520
13570
UV
UV
CD
UV
UV
FL
UV
UV
CD
UV
FL
UV
UV
CD
UV
UV
UV
CD
UV
c
c
d
c
c
e
c
c
c
c
e
c
c
c
c
c
c
c
c
≡1
≡1
≡1
4770
1770
634450
8250
17460
426960
3000
883570
7910
21150
52950
3170
28150
15080
52060
1090
≡1
103
≡1
2
12700
33100
63900
13700
1280
5.8
17.7
17.9
16.7
12.4
1.4
10.9
7.7
4.1
6.1
8.2
4.2
2.7
3
4
3060
20400
27300
3330
13300
15300
14800
14800
a
b
Relative selectivity for each dye, where host X ) 1, 2, 3, or 4. Method employed: CD, circular dichroism spectrometry; UV, UV-vis
spectrophotometry; FL, fluorimetry. c This work. Reference 20. e Reference 19.
d
0.45 mM to a dilute aqueous buffer solution of Methyl
Orange (11 µM) causes a gradual increase of the ICD
intensity over the wavelength range 420-520 nm. These
ICD spectral changes can be used for the quantitative
analysis to determine complex stability constant (KS).
Assuming the 1:1 host/guest stoichiometry, the com-
plexation of guest (G) with cyclodextrin host (H) is
expressed by eq 1
KS
H + G y
\
z H‚G
(1)
The stability constant (KS) can be determined using a
nonlinear least-squares method according to the curve
fitting eq 229
F igu r e 5. (A) UV-vis spectral changes of phosphate buffer
solution of Brilliant Green (11 µM) in the presence of 6,6′-[2,2′-
diselenobis(benzoylaminoethyleneamino)]-bridged bis(â-cyclo-
dextrin) 4. The concentration of host 4 (from a to j): 0, 50,
100, 150, 200, 250, 300, 350, 400, and 450 µM, respectively.
(B) Least-squares curve-fitting analyses for complexations of
Brilliant Green with 4.
∆∆ꢀ ) [R([H]0 + [G]0 + 1/KS) (
R2([H] + [G] + 1/K )2 - 4R2[H] [G] ]/2 (2)
x
0
0
S
0
0
where [G]0 and [H]0 refer to the total concentrations of
dyes and â-cyclodextrin derivatives, respectively, R is the
proportionality coefficient for the effective CD intensity
change induced by host complexation, which may be
taken as a sensitivity factor for the CD change, and ∆∆ꢀ
denotes the change in CD intensity upon stepwise addi-
tion of host. For each host examined, the plot of ∆∆ꢀ as
a function of [H]0 gave an excellent fit to the theoretical
curve, verifying the validity of the 1:1 complex stoichi-
ometry assumed above. As shown in Figure 4B, the
observed ∆∆ꢀ value (open circle) are plotted against [H]0
to give an excellent fit without serious deviations from
the calculated values (small dots). In the repeated
measurements, the KS values were reproducible within
an error of (5%. The KS and R values obtained by the
curve fitting are listed in Table 2, along with the free
energy changes of complex formation (-∆G°).
Sp ectr op h otom etr ic Titr a tion s. To further inves-
tigate the molecular binding abilities and selectivities of
dimeric hosts, complexation behavior of some azo and tri/
diphenylmethane dyes possessing related structures, i.e.,
Tropaeolin OO, Auramine O, Brilliant Green, and Crystal
Violet, was examined by differential UV-vis spectral
titrations.
In the spectrophotometric titration experiments, the
absorption of dyes gradually decreased in intensity upon
addition of varying amounts of hosts 1-4. Typical UV-
vis spectral changes upon addition of host 4 to Brilliant
Green solution are shown in Figure 5A. The absorbance
at ca. 625 nm decreases with increasing concentration
of 4, indicating the formation of inclusion complex with
dye. With the assumption of the 1:1 stoichiometry, the
inclusion complexation can also be expressed by eq 1. The
stability constant (KS) may be determined using a
nonlinear least-squares method according to the curve
fitting eq 329
∆A ) [R([H]0 + [G]0 + 1/KS) (
R2([H] + [G] + 1/K )2 - 4R2[H] [G] ]/2 (3)
x
0
0
S
0
0
where all the abbreviations are the same as eq 2, except
for A which denotes the change in absorbance upon
stepwise addition of host. For each host-guest combina-
tion examined, the plot of observed A (open circle) as a
function of the initial host concentration [H]0 gave an
excellent fit to the theoretical value (small dots), as shown

Molecular Recognition Studies on Supramolecular Systems
J . Org. Chem., Vol. 66, No. 1, 2001 231
in Figure 5B. In the repeated measurements, the KS
values were reproducible within an error of (5%. The
KS and R values obtained by the curve fitting are also
listed in Table 2, along with the free energy change of
complex formation (-∆G°).
KS value obtained with 2 decreases in the order: Methyl
Orange > Crystal Violet > Auramine O > Brilliant Green
> ANS. It is noted that particularly Brilliant Green is
not fully benefited from the cooperative binding. This is
evident from the comparison of the relative binding
constants, KS(2)/KS(1) ) 5.8, for Brilliant Green with the
corresponding values of 12.4-17.9 for the other dyes
(Table 2). This dramatic change in selectivity sequence
and the much decreased cooperative effect observed for
Brilliant Green may be ascribed to the different inclusion
complexation behavior of native cyclodextrin and cyclo-
dextrin dimers. Harata’s studies34 on the inclusion com-
plexation behavior of native cyclodextrins clearly dem-
onstrate that guest molecule usually penetrates into
cyclodextrin cavity from the wider secondary side. How-
ever, 6,6′-bridged bis(cyclodextrin)s are considered to bind
guest in a cooperative manner from the primary side of
dual cyclodextrin moieties. Since one of the major struc-
tural differences between Brilliant Green and Crystal
Violet or Auramine O is the alkyl group attached to the
anilino nitrogen, it is inferred that the larger diethyl-
amino substituent in Brilliant Green causes steric hin-
drance upon penetration into the cavities from the
narrower primary side, preventing effective operation of
the cooperative binding by dual cyclodextrins.
Molecu la r Bin d in g Ability a n d Molecu la r Selec-
tivity. It has been demonstrated in the preceding studies
on molecular recognition by cyclodextrin dimers10-18 that,
possessing dual hydrophobic cavities, tethered bis(cyclo-
dextrin)s exhibit greatly enhanced molecular binding
ability through the cooperative binding and multiple
recognition mechanism. In the present study on the
inclusion complexation of several dyes, we obtained much
higher stability constants (KS) for dimeric hosts 2-4 than
for native â-cyclodextrin 1. Thus, the KS values for the
dimeric hosts are enhanced by factors of 5.8-18 for 2, of
1.4-11 for 3, and of 2.7-8.2 for 4, respectively. There is
a fairly general tendency of KS, decreasing with increas-
ing tether length. Thus the binding constants for Crystal
Violet, Methyl Orange, and Auramine O gradually de-
crease in the order 2 > 3 > 4, although Brilliant Green
is exceptional, giving the lowest KS with 3. This tendency
is entropic in origin and is attributable to the gradually
declining cooperative effect upon extending the distance
between two cyclodextrin moieties.
It is not appropriate or even hazardous in general to
ascribe the observed inclusion complexation behavior to
a single cause, since the binding ability and selectivity
are affected by several factors, including size/shape
matching, hydrophobicity, electrostatic interaction, and
hydrogen-bonding ability. However, the guest dyes em-
ployed in the present study share some structural and
functional similarities, possessing an anilino moiety in
common and a quasi-linear or triangular shape with an
anionic sulfonate or cationic iminonium/ammonium group.
All of these guest features are suitable for investigating
interaction with the ditopic hosts. We therefore discuss
the inclusion behavior of bis(cyclodextrin)s with the
structurally related guests.
As can be seen from Table 2, native â-cyclodextrin
displays a selectivity sequence: Tropaeolin OO > Methyl
Orange > Brilliant Green > Crystal Violet > Auramine
O > ANS. The strongest binding of Tropaeolin OO and
Methyl Orange may be ascribed primarily to the quasi-
linear shape and size fitted to the â-cyclodextrin cavity
and additionally to the presence of a sulfonate group that
could interact with the secondary hydroxyl groups of
cyclodextrin through the electrostatic and/or hydrogen-
bonding interactions. It would be somewhat unexpected
that â-cyclodextrin show the lowest binding ability
toward ANS. As we reported,20 examinations with CPK
molecular models indicate that ANS can only partially
penetrate into the cavity to form a weak inclusion
complex as a result of the steric hindrance arising from
the bent structure. The global selectivity sequence is
affected also by the hydrophobicity of the guest molecules,
or the numbers of hydrophobic benzene ring and of
hydrophilic polar/charged group, which may explain the
selectivity sequences within the two families of dyes:
Tropaeolin OO > Methyl Orange and Brilliant Green >
Crystal Violet > Auramine O.
Host 3 with a tether of moderate length also binds all
the examined dyes stronger than native cyclodextrin 1,
but gives appreciably lower KS values with the same
selectivity sequence as host 2, i.e., Methyl Orange >
Crystal Violet > Auramine O > Brilliant Green. The
efficiency of cooperative binding as measured by KS(3)/
KS(1) is certainly lower than that observed for 2 in each
case, but is much more widely spread from 1.4 for
Brilliant Green to 10.9 for Crystal Violet. Consequently,
the molecular selectivity for Methyl Orange over Brilliant
Green is dramatically enhanced from 1.6 for host 1 to
5.0 for host 2 and then to 8.9 for host 3. This global
improvement of molecular selectivity observed for 3, as
a result of the guest-specific enhancement of KS, probably
originates from the moderate conformational freedom of
3, which enables the tailoring of host conformation to fit
to the shape of specific guest dyes. Both the weak ICD
intensity observed for Methyl Orange (Figure 2) and the
shortest τL (7.4 ns) for ANS (Table 1) in the presence of
3 also indicate that not the core but the peripheral part
of aromatic chromophore of dyes is accommodated in the
host cavities, which is consistent with the complexation
behavior of dyad hosts.
It is intriguing that host 4 gives moderately enhanced
(by a factor of 2.7-8.2) but mutually very close binding
constants (KS ) 13300-15300 M-1) for all of the azo and
triphenylmethane dyes examined. The very flat selectiv-
ity profile observed for 4 is not anticipated or understood
at all from a simple extension of the complexation
behavior of its lower homologues 2 and 3, although the
rather limited enhancement of KS seems reasonable in
view of the entropic disadvantage of the longer tether
which requires more extensive conformational changes,
and greater reduction of entropy, upon guest inclusion.
However, it is not an easy task to invent a mechanism
that can rationalize the comparable affinities of 4 toward
the four dyes, as the degree of affinity enhancement
differs from guest to guest. Then, we may tentatively
Host 2 displayed the highest binding ability among the
three dimeric cyclodextrins for all of the dyes examined,
for which the shortest and least flexible tether is most
probably responsible. The selectivity sequence shows an
interesting difference from that observed for 1. Thus, the
(34) (a) Harata, K.; Uedaira, H.; Tanaka, J . Bull. Chem. Soc. J pn.
1978, 51, 1627. (b) Harata, K. Bull. Chem. Soc. J pn. 1980, 53, 2782.

232 J . Org. Chem., Vol. 66, No. 1, 2001
Liu et al.
conclude that the very close KS’s observed are just an
incidental coincidence as a result of the larger enhance-
ment (6.1-8.2 times) for the triphenylmethane dyes and
the smaller enhancement (2.7-4.2 times) for the azo
dyes.
hydrophobicity and substituent effect of guest determine
the stability of host-guest complex through hydrophobic,
van der Waals, and hydrogen-bonding interactions. Fi-
nally, we wish to emphasize that the size/shape-fit
relationship and induced-fit concept as well as multiple
recognition mechanism working between host and guest
also play crucial roles in the molecular binding ability/
selectivity of bis(cyclodextrin)s, inducing the ultimate
conformation of host adjusted to the guest accommodated
in the dual cyclodextrins.
In conclusion, we have demonstrated that several
factors greatly and extensively affect the inclusion com-
plexation of guest dyes with cyclodextrin dimers, includ-
ing not only the shape/charge/substituent of guest and
the tether of host but also the critical changes in host-
guest interactions such as hydrophobic, van der Waals,
and hydrogen-bonding interactions as well as the size/
shape-fit, induced-fit, multiple recognition, and coopera-
tive binding mechanisms. The inclusion complexation
behavior of dimeric hosts mainly depend on the confor-
mation, length, and flexibility of the organoselenium
tether, which may control how the dual cyclodextrin
cavities adjust their orientations and conformation to
cooperatively bind guest molecule. Simultaneously, the
Ack n ow led gm en t. This work was supported by the
National Outstanding Youth Fund (Grant No. 29625203),
Natural Science Foundation of China (Grant No.
29992590-8 and 29972029), and Natural Science Fund
of Tianjin Municipality (Grant No. 993601311), which
are gratefully acknowledged.
J O001372T