Synthesis and inclusion ability of anthracene appended β- cyclodextrins: Unexpected effect of triazole linker

Article abstract of DOI:10.1016/j.carres.2010.09.031

A new fluorescent β-cyclodextrin has been synthesized by coupling an anthracene moiety to the cyclic oligosaccharide via click chemistry. The influence of the triazole spacer was compared to the simple amino and amido linkers. While a sensing ability toward adamantan-1-ol was observed with the latter two spacers, the absence of inclusion capacity prevents the triazole modified cyclodextrin from showing any fluorescence variations. The difference in the binding behaviors studied by Isothermal Titration Calorimetry, UV-vis and fluorescence spectroscopies, was highlighted by the NOESY NMR spectra of the modified cyclodextrins: whereas a free cavity was observed for the amino and amido linkers, an important obstruction was obtained in the case of the triazole.

Full text of DOI:10.1016/j.carres.2010.09.031

Carbohydrate Research 346 (2011) 35–42
Contents lists available at ScienceDirect
Carbohydrate Research
journal homepage: www.elsevier.com/locate/carres
Synthesis and inclusion ability of anthracene appended b-cyclodextrins:
unexpected effect of triazole linker
Isabelle Mallard a,b, , David Landy a,b, Nadia Bouchemal c, Sophie Fourmentin a,b
⇑
a Univ Lille Nord de France, F-59000 Lille, France
b ULCO, UCEIV, F-59140 Dunkerque, France
c Univ Paris XIII, ANBiophy FRE 3207, F-93017 Bobigny, France
a r t i c l e i n f o
a b s t r a c t
Article history:
A new fluorescent b-cyclodextrin has been synthesized by coupling an anthracene moiety to the cyclic
oligosaccharide via click chemistry. The influence of the triazole spacer was compared to the simple
amino and amido linkers. While a sensing ability toward adamantan-1-ol was observed with the latter
two spacers, the absence of inclusion capacity prevents the triazole modified cyclodextrin from showing
any fluorescence variations. The difference in the binding behaviors studied by Isothermal Titration Cal-
orimetry, UV–vis and fluorescence spectroscopies, was highlighted by the NOESY NMR spectra of the
modified cyclodextrins: whereas a free cavity was observed for the amino and amido linkers, an impor-
tant obstruction was obtained in the case of the triazole.
Received 24 June 2010
Received in revised form 20 September
2010
Accepted 28 September 2010
Available online 23 October 2010
Keywords:
Anthracene
b-Cyclodextrins
Formation constants
Fluorescent sensors
Ó 2010 Elsevier Ltd. All rights reserved.
1. Introduction
compound, CD1, consists of an anthracene moiety connected to
b-CD via an imidazole ring spacer. The triazole ring is introduced
To evaluate environmental pollution, new efficient systems of
detection should be developed. For several years, we have focused
on the design and synthesis of novel cyclodextrin sensors for the
detection of volatile organic compounds (VOCs).1–3 Within this
scope, cyclodextrins should be modified by a chromophore to be
used as a tool for detection.4–6
via a Hugsen [1,3] dipolar cycloaddition, with an attempt to carry
out the click chemistry on the anthracene derivative rather than
on cyclodextrin. To validate the eventual efficiency of the triazole
ring arm, the linker between anthracene moiety and cyclodextrin
was varied. Sensor CD2 was constructed according to the same
design as CD1, except that the spacer was an amide linkage
without the triazole ring. The last sensor, CD3, was based on
an amine linkage and was prepared by means of a Schiff base
reaction.
The inclusion ability of CD1, CD2, and CD3 toward adamantan-
1-ol as a model for organic compounds was studied by UV–vis and
fluorescence spectroscopy and also by Isothermal Titration Calo-
rimetry (ITC). On the basis of these results, we were able to eluci-
date the effect of the spacer on the binding ability of these
fluorescent cyclodextrins.
In the past few years, click chemistry has received much atten-
tion in the cyclodextrin field, as it allows for the development of
new synthetic methods, leading to modified cyclodextrins used in
catalysis,7,8 chiral stationary phases9, and cation detection.10
According to the literature, triazole cyclodextrin derivatives have
not been exploited for organic compound detection. A question thus
arises: could we take advantage of the triazole ring for the detection
process? As a consequence, we decided to investigate the effect of
the triazole ring associated with a fluorescent anthracene moiety
as a chromophoric model. Some anthracene derivatives have high
quantum yield11 and are used for metal detection12–14 or medical
diagnosis.15 The synthesis of anthracene cyclodextrin derivatives
has been reported for controlling stereochemistry of bimolecular
interactions and reactions,16 photophysical studies17, and the
molecular recognition.18
2. Results and discussion
2.1. Synthesis of b-CD precursors
Mono 6I-amino-6I-deoxy-b-cyclodextrin was synthesized accor-
ding to well known described procedures:19–21 tosylation of the
native b-CD followed by sodium azide substitution gave mono
6I-azido-6I-deoxy-b-cyclodextrin which was reduced to mono
6I-amino-6I-deoxy-b-cyclodextrin.
This paper describes the synthesis and the inclusion ability of
three anthracene cyclodextrin derivatives (Fig. 1). Our target
⇑
Corresponding author. Tel.: +33 (0) 328658257; fax: +33 (0) 328237605.
E-mail address: isabelle.mallard@univ-littoral.fr (I. Mallard).
0008-6215/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2010.09.031

36
I. Mallard et al. / Carbohydrate Research 346 (2011) 35–42
N
N
N
O
O
HN
HN
HN
OH
6
OH
6
OH
6
O
O
O
O
2 OHO
O
2 OH O
O
2 OH O
O
O
OH
O
4
4
HO
HO
4
5
3
5
3
HO
5
3
OH
1
1
OH
1
HO
HO
HO
n
n
n
n=6
n=6
n=6
CD2
CD1
CD3
Figure 1. Structure of modified CDs.
2.2. Synthesis of CD1
Moreover, because of the size of anthracene, the reaction was ap-
plied to anthracene and not on cyclodextrin.30–33
The formation of CD1 was conducted in six steps as shown in
Scheme 1. Commercial anthracene 1 was converted to anthracene-
9-carbaldehyde 2 via a slightly modified Vilsmeyer reaction.22
Reduction11 of 2 with NaBH4 leads to compound 3. Nucleophilic sub-
stitution23 with thionyl chloride gave 4. Azidation of 4 was slightly
different from the usual treatment.11 Following HSAB theory, the
reaction was performed in the presence of quaternary tetrabutyl
ammonium bromide salt,24 used in catalytic amount to reduce reac-
tion time from 5 h to 1 h. Propynoic acid-4-nitrophenylester was
synthesized in 60% yield according to literature precedent,25 by
reacting para-nitrophenol and propiolic acid with dicyclohexyl-car-
bodiimide as activating agents and dimethylaminopyridine.
Huigsen 1,3-dipolar cycloaddition using copper iodide as a cata-
lyst was performed between azide 5 and propynoic acid-4-nitroph-
enylester in a THF–water (1:1) solvent mixture at room temperature
for one week to give 6. Without heating, the cycloaddition pro-
ceeded regioselectively, however, a much longer reaction was re-
quired.26 Although the click azide–alkyne cycloaddition reaction
was reported with various different solvents27,28 and copper salt
catalysts,28,29 we decided to use copper iodide and heterogeneous
conditions to make the purification process less complicated instead
of reduction in situ with copper sulfate and sodium ascorbate.
The formation of the fluorescent triazole product is easily visi-
ble upon irradiation at 365 nm with a hand-held UV lamp. Copper
catalyst was removed by washing with EDTA, then compound 6
was purified on silica gel to afford the pure product with a 72%
yield. The triazole ring protons produced a singlet at 7.75 ppm.
No other singlet was observed, confirming the formation of only
one regioisomer. CDNH2 was treated with 7 at 60 °C for two days;
CD1 was formed with a yield of 80%.
2.3. Synthesis of CD2
Our attempts to synthesize CD2 with anthracene-9-carboxylic
acid as a precursor and 6I-amino-6I-deoxy-b-cyclodextrin using
DCC–DMAP provided moderate yield, so we reverted to fluoride
acid synthesis. Commercial carboxylic anthracene acid was con-
verted to compound 8 in 80% yield using cyanuric fluoride,34 pyri-
dine, and dichloromethane. Condensation of 8 with CDNH2 leads to
compound CD2 in a 73% of yield (Scheme 2).
It should be noted that CD2 was prepared under mild conditions
and that the purification methods were less fastidious than those
required for carboxylic or chloride acid methods.
CHO
CH2OH
CH2Cl
ii
i
iii
1
2
4
3
iv
CH2N3
v
vi
N
N
N
N
N
O
5
N
O
6
O
HN
OH
6
O
NO2
O
2 OH O
O
4
HO
5
OH
1
3
HO
n
n=6
CD1
Scheme 1. (i) POCl3, N-methylformanilide, o-dichlorobenzene, 363–368 K, 2 h; (ii) NaBH4, ethanol, rt, 4 h; (iii) SOCl2, CH2Cl2, RT, 2 h; (iv) NaN3, tetrabutylammonium
bromide, DMF, 353 K, 1 h; (v) propynoic acid-4-nitrophenylester, H2O–THF, CuI, rt, one week; (vi) CDNH2, 333 K, two days.

I. Mallard et al. / Carbohydrate Research 346 (2011) 35–42
37
COOH
COF
O
HN
i
ii
OH
6
O
O
2 OH O
O
4
HO
5
3
OH
1
7
8
HO
n
n=6
CD2
Scheme 2. (i) Cyanuric fluoride, pyridine, CH2Cl2, rt; (ii) CDNH2, rt, overnight.
2.4. Synthesis of CD3
Data analysis revealed that formation constants were equal to
5962 and 4353 Mꢀ1 for CD2/adamantan-1-ol and CD3/adaman-
tan-1-ol complexes, respectively (Table 1). These values were low-
er than those observed for the native b-cyclodextrin inclusion
compound (34354 Mꢀ1),37 indicating that the appending anthra-
cene is not beneficial for inclusion ability. One can think that the
anthracene moiety may obstruct the primary rim, if a capped con-
formation is sometimes occurring for CD2 and CD3. Such a struc-
ture, which may only constitute one conformation among others
and which differs from an auto-inclusion, has been suggested by
Balan and Gopidas,38 for a very similar anthracene appended
b-cyclodextrin. It implies that the primary rim is not as available
as in the case of native b-cyclodextrin, and that the inclusion
ability of CD2 and CD3 is reduced.
Synthesis of CD3 was readily achieved in two steps via forma-
tion of a Schiff’s base (Scheme 3).35,36 The Schiff’s base was synthe-
sized by the reaction between an aromatic aldehyde and an amino
group in which a nucleophilic addition and elimination of water
occurred, giving an imine. The imine intermediate was generated
in situ by reacting
(CDNH2) in methanol at room temperature for four days and
reduced with NaBH4 to afford CD3 with a yield of 66%.
2
with 6I-amino-6I-deoxy-b-cyclodextrin
2.5. Inclusion ability of anthracene cyclodextrin derivatives
To study the inclusion ability of these new anthracene cyclo-
dextrin derivatives, we investigated the complexes formed with
adamantan-1-ol, as a model guest compound. Adamantan-1-ol
was found to be well recognized by the b-CD cavity.37
Table 1
Formation constants (Mꢀ1) obtained for the b-cyclodextrins/adamantan-1-ol inclu-
sion complexes, by means of UV–vis, ITC and fluorescence measurements
2.5.1. UV studies
The complexation phenomena were studied by means of UV–
vis absorption spectroscopy. In the case of CD1, no proof of inclu-
sion was obtained as spectral variation was not observed. On the
contrary, increasing concentrations of adamantan-1-ol in the pres-
ence of a fixed concentration of cyclodextrin lead to measurable
variations in the spectra of CD2 and CD3 (Fig. 2).
UV–vis
ITC
Fluorescence
CD1
CD2
CD3
No inclusion detected with any technique
5962 631
4353 843
6162 796
5303 672
4575 827
4823 952
CHO
HN
OH
6
i
ii
O
O
2 OHO
O
4
HO
5
OH
1
2
1
3
HO
n
n=6
CD3
Scheme 3. (i) POCl3, N-methylformanilide, o-dichlorobenzene, 363–368 K, 2 h; (ii) methanol, calcium perchlorate hydrate, CDNH2, four days, rt then NaBH4, four days, rt.
Figure 2. Representative UV–vis titration plot for inclusion of adamantan-1-ol in CD2 (right part) and CD3 (left part). Solid line represents the best theoretical fit to
experimental points.

38
I. Mallard et al. / Carbohydrate Research 346 (2011) 35–42
Figure 3. Representative observed heats (upper part) and corresponding binding isotherm (lower part) for ITC release experiments with adamantan-1-ol complexes of CD2
(left part) and CD3 (right part). Solid line represents the best theoretical fit to experimental points.
2.5.2. ITC studies
enthalpy were equal to ꢀ4853 cal molꢀ1 for CD2, and to ꢀ4634
To gain better insight into the adamantan-1-ol complexation,
release experiments were also conducted with Isothermal Titration
Calorimetry (ITC). Figure 3 describes the ITC enthalpogram for dilu-
tion of adamantan-1-ol complexes with CD2 and CD3 in water
(phosphate buffer). Positive peaks were observed for each addition
as a consequence of the partial dissociation of inclusion com-
pounds. The intensities gradually decreased as such dissociation
weakens as the host and guest concentrations inside the cell in-
creased. Because the experimental conditions were identical for
each complex and complex stabilities are close, the ITC data were
very similar for CD2 and CD3. The fit of these data to the theoret-
ical model allowed the determination of the inclusion thermody-
namic parameters. Formation constants were equal to 6162 Mꢀ1
and 5303 Mꢀ1 for CD2 and CD3, respectively, which were in agree-
ment with the UV–vis results (see Table 1). Values of inclusion
cal molꢀ1 for CD3. The entropic part (expressed as ꢀT
DS at
298 K) was equal to ꢀ314 cal molꢀ1 for CD2, and to ꢀ444 cal molꢀ1
for CD3. Concerning CD1, no signal was again observed, confirming
the UV–vis study. Therefore, it can be concluded that the CD1 cav-
ity is not suited for inclusion phenomena.
2.5.3. NMR studies
To obtain further evidence about the differences of inclusion
behavior, 2D NMR spectroscopy was performed on CD1, CD2, and
CD3 in D2O. Because NOESY cross-peaks are indicative of specific
proximity relationships between adjacent protons, it was possible
to determine if interactions were occurring between the cyclodex-
trin cavity and the appended chromophores. While no interaction
was observed between the anthracene sidearm and the b-CD cavity
for CD2 and CD3, CD1 spectra displayed cross-peaks between the
Figure 4. NOESY spectrum of CD1 in D2O at 298 K with a mixing time of 300 ms.

I. Mallard et al. / Carbohydrate Research 346 (2011) 35–42
39
studies), no environmental change seems to occur for this fluores-
cent probe.
On the contrary, the fluorescence properties of CD2 and CD3
were altered in the presence of various concentrations of adaman-
tan-1-ol (Fig. 6). The inclusion of such a bulky guest constitutes a
constraint on the cyclodextrin dynamic behavior, in such a way
that conformational changes probably occur for the appended
chromophore, thus inducing
fluorescence.
a
moderate modification of
The inclusion ability of CD2 and CD3 was again confirmed, as
formation constant values (4575 Mꢀ1 for CD2, 4823 Mꢀ1 for CD3,
see Table 1) were similar to what was obtained by UV–vis spec-
troscopy and ITC experiments. The sensitivity factors
DF/F0 for ada-
mantan-1-ol concentration of 0,675 mM were equal to +30% for
CD2 and to ꢀ20% for CD3 (data integrated over the 400–440 nm
range). Even if greater values have been observed in the litera-
ture,39,40 one can consider that these two anthracene appended
b-cyclodextrins may be used as sensors for organic compounds,
in contrary to the triazole-anthracene appended CD1.
3. Conclusion
Figure 5. Minimized structure obtained for the intermolecular inclusion occurring
between two CD1 molecules (CPK representation for the included anthracene
moiety).
We have determined that the popular click chemistry is not
necessarily an optimal way to obtain efficient modified cyclodex-
trins. In our attempt to design new cyclodextrin sensors, the tria-
zole spacer associated with an anthracene moiety lead to
cyclodextrin without inclusion ability, as demonstrated by UV–
vis, ITC, and NOESY NMR studies. As a result, sensing ability was
not observed for this modified cyclodextrin, whereas the same
anthracene moiety associated with simple amino or amido spacer
lead to fluorescence variations with inclusion of adamantan-1-ol.
As the click chemistry increasingly focuses attention on cyclodex-
trin, care should be taken to the inclusion ability of new triazole
appended oligosaccharide.
internal protons (3.62 and 3.9 ppm, H3, H5, H6) of the CD cavity
and the protons of the anthracene moiety and triazole ring (7.5–
8 ppm) (Fig. 4).
As the triazole linker is quite rigid, one can think that the
observed interactions probably result from an intermolecular
inclusion between two CD1 molecules, as illustrated by the mini-
mized structure depicted in Figure 5. As a result, the appended
anthracene causes a significant steric hindrance which strongly
reduces the binding ability of CD1. Because CD2 and CD3 do not
show a similar behavior, it seems that the triazole moiety plays
an important role for the stabilization of the intermolecular recog-
nition between CD1 molecules.
4. Experimental section
4.1. General
All chemicals were purchased from Acros and Aldrich Chemicals
in their highest purity. All solvents were used distilled. Distilled
water was used in all experiments. Analytical thin layer chroma-
tography (TLC) was performed on aluminum backed silica gel from
E. Merck (silica gel F254). CM-25 Sephadex (dry bead size: 40–
125 m) was obtained from Sigma Aldrich. Mass spectra were re-
corded on a Maldi-TOF/TOF Bruker Daltonics Ultraflex II in positive
reflectron mode with 2.5-DHB as matrix.
2.5.4. Fluorescence studies
The influence of the addition of adamantan-1-ol on the fluores-
cence spectra of each modified cyclodextrin was studied in order to
establish their sensing potential. As could be expected from the
preceding results, no variation of fluorescence was observed upon
addition of adamantan-1-ol to CD1. The unavailability of the cavity
is indeed prohibitive: as adamantan-1-ol does not induce any dis-
placement of the anthracene moiety (according to UV–vis and ITC
Figure 6. Representative fluorescence variations and corresponding titration plot (inset), for CD2 (left part) and CD3 (right part) upon inclusion with adamantan-1-ol
(concentrations equal to 0, 0.056, 0.113, 0.225, 0.450 and 0.675 mM). Solid lines represent the best theoretical fit to experimental points.

40
I. Mallard et al. / Carbohydrate Research 346 (2011) 35–42
where CD and Ad represent the cyclodextrin and adamantan-1-ol
concentration, respectively, H corresponds to the inclusion enthal-
4.2. NMR spectroscopy
D
py, the subscripts t, b, and f stand for total, bound, and free concen-
trations, respectively. Superscript ‘syr’ refers to the concentrations
of the corresponding species in the syringe. Xsyr represents the mole
fraction of the guest concentration in the syringe. qdil is the heat of
dilution which is accounted for by performing blank titrations (for
both host and guest). The partial differentials of the bound guest
concentration with respect to total guest concentration and to total
host concentration are calculated on the basis of the following
equations (for 1:1 complexes):
All NMR experiments were carried out on a Varian Unity Inova
spectrometer (11.7 T) operating at a proton frequency of 500 MHz
and at a carbon frequency of 125 MHz with a 5 mm gradient indi-
rect probe. The experiments were performed at 298 K and spectral
widths of 5000 Hz for proton and 32 kHz for carbon were used. 1
H
spectra (32 transients) were obtained with a 90° pulse length
(7.3 s), 1 s relaxation delay, 5000 Hz spectral width and 16 K data
l
size. The water signal was suppressed by a presaturation pulse
during the relaxation delay. 0.3 Hz line-broadening was applied
to the free induction decay (FID) prior to Fourier transform. 2D
NOESY proton NMR experiments were recorded with the phase
sensitive mode using the hypercomplex scheme. Mixing times of
300 ms were used for NOESY experiment. Typical two-dimensional
spectra were acquired with 1024 complex points in t2 and 512 in
t1, employing the TPPI scheme (Marion and Wuthrich 1983). The
8
9
>
<
>
=
@Adb
@Adt
1
2
ðCDt þ Adt þ 1=Kf Þ ꢀ 2CDt
ðCDt þ Adt þ 1=Kf Þ ꢀ 4CDtAdt
qffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi
¼
¼
1 ꢀ
1 ꢀ
>
:
>
;
2
8
9
>
<
>
=
@Adb
@CDt
1
2
ðCDt þ Adt þ 1=Kf Þ ꢀ 2Adt
qffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi
*
>
:
>
;
2
recycling delay was 3 s. Data were zero filled to 4 K 2 K points
ðCDt þ Adt þ 1=Kf Þ ꢀ 4CDtAdt
in both dimensions and apodized with skewed phase-shifted sine
bell functions. For the Heteronuclear Single Quantum Coherence
(HSQC) experiment, a delay t/2 was set to 15 ms for optimal polar-
ization transfer.
The theoretical heats are then fitted to the experimental ones by
varying the Kf and
D
H values (v2 minimisation process, with New-
ton–Raphson algorithm). Each release experiment was performed
three times to ensure reproducibility of the results.
4.3. UV spectroscopy
4.5. Fluorescence spectroscopy
Spectra were recorded using a Perkin Elmer Lambda 2S double
beam spectrometer and a quartz cell with optical path length of
1.00 cm at 298 K. All compounds were dissolved in phosphate buffer
at pH 6.0. The temperature was controlled by the use of a thermo-
stated bath linked to the cell holder (accuracy: 0.1 K). The stability
of the complexes formed between the adamantan-1-ol and the hosts
(CD1, CD2, and CD3) was measured by the use of a direct titration
method. Spectra were recorded between 300–450 nm for a cyclo-
dextrin concentration fixed at 0.05 mM, with varying adamantan-
1-ol concentrations (from 0 to 0.75 mM). The 1:1 formation
constant Kf was obtained by applying a dedicated algorithmic treat-
ment to the first derivatives of the UV spectra in order to avoid any
spectral influence of diffraction phenomena.41 Each titration exper-
iment was performed three times to ensure reproducibility of the
results. Reported values for Kf are the mean standard deviation
of the three experiments.
The measurements were carried out with a Perkin Elmer LS-50B
fluorimeter and a quartz cell with optical path length of 1.00 cm at
298 K. The temperature was controlled by the use of a thermo-
stated bath linked to the cell holder (accuracy: 0.1 K). All spectra
were obtained with degassed aqueous buffer solutions (phosphate
buffer, pH 6). The excitation wavelength of the fluorescence spec-
tra was 346 nm for CD2 and 348 nm for CD3 (10 nm excitation and
emission slits). The spectra were collected from 360 nm to 520 nm
with a scan rate fixed to 60 nm/mn, for cyclodextrin concentrations
equal to 2 ꢁ 10ꢀ6 M and 5 ꢁ 10ꢀ7 M for CD2 and CD3, respectively.
Sensitivity factors and 1:1 Formation constants were calculated by
integrating data between 400 and 440 nm. The sensitivity factors
were defined as
D
F/F0, where
D
F corresponds to F ꢀ F0, F, and F0
being respectively the integrated fluorescence intensity in the
presence and absence of adamantan-1-ol. The employed adaman-
tan-1-ol concentration was equal to 0.675 mM. 1:1 formation con-
stants were obtained by nonlinear fitting (Newton–Raphson
algorithm integrated to Excel) of the well known equation:44
4.4. Isothermal Titration Calorimetry
An isothermal calorimeter (ITC200, MicroCal Inc., USA) was used
for simultaneously determining the formation constant and the
inclusion enthalpy and entropy. The release protocol was used.42,43
F
F0
Kf ½titrating speciesꢂT
¼ 1 þ ðF =F0 ꢀ 1Þ
1
1 þ Kf ½titrating speciesꢂT
where F (calculated at each algorithmic step for a given Kf value) is
1
204.5
pH 6) was titrated at 298 K with cyclodextrin/adamantan-1-ol
solution using a 40 L syringe. Concentrations of cyclodextrins
and adamantan-1-ol were equal to 2 mM and 0.4 mM, respectively.
Aliquots of 8.5 L of the inclusion compound solution were deliv-
lL of degassed aqueous buffer solution (phosphate buffer,
the integrated fluorescence intensity when all the cyclodextrin
molecules have been complexed by adamantan-1-ol molecules.
[titrating species]T represents the total adamantan-1-ol concentra-
tions (which were equal to 0, 0.056, 0.113, 0.225, 0.450, and
0.675 mM). Each titration experiment was performed three times
to ensure reproducibility of the results.
l
l
ered over 17 s and the corresponding heat flow was recorded as
a function of time. The time interval between two consecutive
injections was 150 s with an agitation speed of 1000 rpm for all
experiments. The area under the peak following each injection of
the inclusion compound solution (obtained by integration of the
raw signal) was then expressed as the heat effect per mole of added
adamantan-1-ol. Data analysis was carried out according to Ref. 42,
by implementing the corresponding equation in Excel software.
The observed heat (qobs) can be given by:
4.6. Molecular modeling
Molecular modeling has been employed in order to illustrate
the intermolecular inclusion between CD1 molecules. Simulations
were realized by means of Macromodel with MMFF force field and
GB/SA simulation of water.45,46 A symmetric b-CD structure was
used to construct the CD1 molecule. The appended anthracene
with the triazole linker, which was added to the b-CD structure,
has been submitted to a Polak–Ribiere Conjugate Gradient minimi-
zation (convergence fixed to 0.05 kJ/Å-mol). The docking of this
"
#
@Adb
@Adt
@Adb
@CDt
Adsbyr
qobs
¼
D
H Xsyr
þ ð1 ꢀ Xsyr
Þ
ꢀ
þ qdil
Adstyr þ CDstyr

I. Mallard et al. / Carbohydrate Research 346 (2011) 35–42
41
appended anthracene inside a second CD1 molecule (which was
kept rigid at this stage) was then realized by means of Monte Carlo
searches, with generation of 10,000 conformations. The most sta-
ble conformation was then completely relaxed. All minimizations
have been realized with Polak–Ribiere Conjugate Gradient
algorithm (convergence fixed to 0.05 kJ/Å-mol).
353 K for 1 h after which the solution was cooled and the DMF
was removed. The crude product was purified on silica gel (SiO2,
CH2Cl2, Rf = 0.83) to obtain a yellow powder (1.02 g, 99%). 1H
NMR d (500 MHz, CDCl3): 8.55 (s, 1H), 8.33 (d, J = 8.75 Hz, 2H),
8.09 (d, J = 8.75 Hz), 7.60 (t, J = 6.6 Hz, 2H), 7.54 (t, J = 6.6 Hz, 2H),
5.38 (s, 2H) ppm. 13C NMR d (125 MHz, CDCl3): 131.4, 130.7,
129.3, 129.0, 126.9, 125.8, 125.2, 123.5, 46.4 ppm. MS: calcd for
4.7. Synthesis
C15H11N3 233.10; found 204.00.
4.7.1. Anthracene-9-carbaldehyde (2)
4.7.5. (1-Anthracen-9-ylmethyl-1H-[1,2,3]triazol-4-yl)-(4-
nitrophenyl)-methanone (6)
A 100 mL round bottomed flask fitted with a mechanical stirrer
and reflux condenser was filled with N-methylformanilide (7.58 g,
56 mmol), phosphorus oxychloride (7.61 g, 49.6 mmol), and
anthracene 1 (5 g, 28 mmol) in o-dichlorobenzene (4.5 mL). The
flask was heated in a steam bath with stirring to 363–368 K over
a period of 20 min. During this time, the anthracene dissolved to
give a deep red solution and the hydrogen chloride evolved. The
heating was continued for 1–2 h, after which the reaction mixture
was cooled to room temperature. A solution of 31 g of crystalline
sodium acetate in 56 mL of water was added and followed by
dichloromethane. The aqueous phase was washed several times
with dichloromethane. The organic phase was dried on magnesium
sulfate, filtered, and the solvent evaporated. The crude product was
recrystallized in ethanol to give 2 as yellow crystals (5.11 g, 89%);
1H NMR d (500 MHz, CDCl3): 11.60 (s, 1H), 9.02 (d, J = 9 Hz, 2H),
8.74 (s, 1H), 7.74 (d, J = 8 Hz, 2H), 7.69 (t, J = 7.5 Hz, 2H), 7.59 (t,
J = 7.5 Hz, 2H) ppm. 13C NMR d (125 MHz, CDCl3): 193.0, 135.3,
132.2, 131.1, 129.3, 129.1, 125.7, 124.8, 123.6 ppm. MS: calcd for
Propynoic acid-4-nitrophenylester (0.331 g, 1.73 mmol) and 5
(0.606 g, 2.6 mmol) were dissolved in a mixture of water–THF
(8 mL:16 mL). Copper iodide (3.3 mg) was added and the reaction
mixture was stirred at room temperature for one week. The crude
product was dissolved in dichloromethane then washed with water.
The organic phase was stirred twice with EDTA (0.5 N) to remove
copper, then evaporated under vacuum. The crude product was
purified on silica gel (SiO2, CH2Cl2, Rf = 0.7) to offer a yellow powder
(0.528 mg, 72%). 1H NMR d (500 MHz, CDCl3): 8.68 (s, 1H), 8.29 (d,
J = 9.3 Hz, 2H), 8.27 (d, J = 8.95 Hz, 2H), 8.16 (d, J = 8.22 Hz, 2H),
7.75 (s, 1H), 7.71–7.57 (m, 4H), 7.33 (d, J = 8.95 Hz, 2H), 6.69 (s,
2H) ppm.13C NMR d (125 MHz, CDCl3): 157.9, 154.7, 145.5, 138.6,
131.5, 130.8, 130.6, 129.8, 128.3, 128.2, 126.2, 125.6, 125.3, 122.5,
122.4, 122.2, 115.7, 47.0 ppm. MS: calcd for C24H16N4O4 424.12;
found 424.00. Anal. Calcd for C24H16N4O4: C, 67.92; H, 3.80; N,
13.20. Found: C, 67.74; H, 3.83; N, 13.15.
C15H10O 206.24; found 206.17.
4.7.6. Anthracene-9-carbonyl fluoride (8)
Anthracene-9-carboxylic acid 7 (0.5 g, 2.25 mmol) was dis-
solved in pyridine (0.37 mL) and dichloromethane (30 mL) and
cooled to 273 K. Cyanuric fluoride (0.41 mL, 4.77 mmol) was added
dropwise and the reaction mixture was stirred for 90 min. 30 mL of
water was added and the organic phase was extracted, dried on
anhydrous magnesium sulfate, filtered, and concentrated to yield
a cream powder (0.449 g, 89%). 1H NMR d (500 MHz, CDCl3): 9.02
(s, 1H), 8.27 (d, J = 9 Hz, 2H), 8.22 (d, J = 9 Hz, 2H), 7.78 (dd,
J = 5 Hz, 2H), 7.67 (dd, J = 5 Hz, 2H) ppm. 13C NMR d (125 MHz,
CDCl3): 160.6, 133.8, 130.2, 129.2, 126.1, 123.8 ppm. MS: calcd
for C15H9FO 224.06; found 224.00. Anal. Calcd for C15H9FO: C,
80.35; H, 4.05. Found: C, 80.20; H, 4.04.
4.7.2. Anthracen-9-ylmethanol (3)
Sodium borohydride (201.8 mg) was added slowly to the sus-
pension of 2 (1 g, 4.8 mmol) in ethanol (40 mL) at 273 K. The reac-
tion mixture was stirred at room temperature for 4 h. Concentrated
HCl solution was added drop wise to decompose the unreacted
NaBH4 and then water was added until neutral pH. Dichlorometh-
ane was then added and the organic layer was separated, washed
with brine, and dried over anhydrous magnesium sulfate. The sol-
vent was removed under reduced pressure to get a pale yellow
powder (0.99 g, 98.3%). 1H NMR d (500 MHz, CDCl3): 8.50 (s, 1H),
8.45 (d, J = 7.5 Hz, 2H), 8.06 (d, J = 7.5 Hz, 2H), 7.63–7.52 (m, 4H),
5.72 (s, 2H), 1.59 (s, 1H) ppm. 13C NMR d (125 MHz, CDCl3):
131.6, 131.0, 130.2, 129.2, 128.4, 126.50, 125.1, 123.9, 57.5 ppm.
MS: calcd for C15H12O 208.09; found 208.00.
4.7.7. Mono-[6-(9-anthryl-1H-[1,2,3]triazole formamido)-
6-deoxy]-b-cyclodextrin (CD1)
Compound 6 (0.25 g, 0.59 mmol) was mixed with 6I-amino-6I-
deoxy-b-cyclodextrin (1 g, 0.88 mmol) in DMF (10 mL). The reac-
tion mixture was stirred at 333 K for two days. The crude mixture
was then added dropwise to acetone. The resulting solid was col-
lected by filtration and subsequently purified on a CM-25 Sepha-
dex column with water as eluent. Evaporation of the solvent
yielded compound CD1 as a yellow powder (0.669 g, 80%). 1H
NMR d (500 MHz, D2O): 8.41 (s, 1H), 8.30–8.16 (m, 2H), 8.10–
7.98 (m, 2H), 7.90 (s, 1H), 7.78–7.30 (m, 4H), 6.55–6.42 (m, 2H),
4.7.3. 9-Chloromethylanthracene (4)
Compound 3 (1 g, 4.8 mmol) was dissolved in freshly distilled
dichloromethane (15 mL) followed by the addition of thionyl
chloride (2.27 mL, 31.2 mmol). The mixture was stirred at room
temperature for 2 h, after which the solvent was removed under
reduced pressure. Dichloromethane was added to the crude prod-
uct and sodium hydrogenocarbonate was introduced until pH 7
was reached. The mixture was filtered, washed with water and
brine, and dried over anhydrous sodium sulfate. The solvent was
removed and the crude product was purified on silica gel (SiO2,
CH2Cl2, Rf = 0.9) to give a yellow powder (0.177 g, 19%). 1H NMR
d (500 MHz, CDCl3): 8.53 (s, 1H), 8.35 (d, J = 8.8 Hz, 2H), 8.07 (d,
J = 8.3 Hz, 2H), 7.64 (t, J = 7.5 Hz, 2H), 7.53 (t, J = 7.5 Hz, 2H), 5.66
(s, 2H) ppm. 13C NMR d (125 MHz, CDCl3): 131.5, 129.9, 129.3,
129.3, 127.4, 126.9, 125.3, 123.4, 39.0 ppm. MS: calcd for
5.08–4.92 (m, 7H), 4.18–3.80 (m, 42H) ppm. 13C NMR
d
(125 MHz, DMSO): 175.0, 159.5, 130.9, 130.3, 129.1, 127.25,
125.4, 125.2, 123.8, 101.9, 81.3, 72.9, 72.4, 71.9, 64.9, 59.9,
40.4 ppm. MS: calcd for [C60H81N4O35+Na]+ 1441.20; found
1441.39. Anal. Calcd for C60H82N4O35ꢃ3H2O: C, 48.91; H, 6.02;
N, 3.80. Found: C, 48.75; H, 6.03; N, 3.77.
C
15H11Cl 226.05; found 191.00.
4.7.8. Mono-[6-(9-formamido)-6-deoxy]-b-cyclodextrin (CD2)
Compound 8 (0.371 g, 1.55 mmol) was added dropwise to
6I-amino-6I-deoxy-b-cyclodextrin (1.46 g, 1.28 mmol) dissolved
in DMF (15 mL) at 273 K. The reaction mixture was stirred at room
temperature overnight. The mixture was poured into acetone and
the resulting solid was collected by filtration. The crude solid
4.7.4. 9-Azidomethylanthracene (5)
Compound 4 (1 g, 4.41 mmol), tetrabutylammonium bromide
(0.071 g, 0.22 mmol) and sodium azide (0.316 g, 4.85 mmol) were
dissolved in DMF (40 mL). The reaction mixture was stirred at

42
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was then passed through a CM-25 Sephadex column by eluting
with water. The pure compound CD2 was obtained as an orange
powder (1.25 g, 73%). 1H NMR d (500 MHz, D2O): 8.61 (s, 1H),
8.24–8.04 (m, 4H), 7.85–7.74 (m, 4H), 5.20–4.90 (m, 7H),
4.05–3.42 (m, 4H) ppm. 13C NMR d (125 MHz, DMSO): 168.8,
133.4, 130.6, 128.2, 127.3, 126.9, 126.1, 125.5, 101.9, 81.3, 72.9,
72.4, 72.0, 69.9, 64.9, 59.8, 40.4 ppm. MS: calcd for [C57H78NO35+-
Na]+ 1360.13; found 1360.33. Anal. Calcd for C57H79NO35ꢃ2H2O: C,
49.82; H, 6.09; N, 1.02. Found: C, 49.78; H, 6.07; N, 1.04.
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4.7.9. Mono-[6-(9-anthrylmethylamino)-6-deoxy]-b-
cyclodextrin (CD3)
Anthracene-9-carbaldehyde 2 (0.25 g, 1.21 mmol) was mixed
with 6I-amino-6I-deoxy-b-cyclodextrin (1.25 g, 1.1 mmol) in dry
MeOH (30 mL). Calcium perchlorate tetrahydrate was added to
the reaction mixture and kept at room temperature for four days.
NaBH4 (0.23 g, 6 mmol) was added and the reaction mixture was
kept at room temperature for four days. The reaction mixture
was neutralized with 1 N HCl and filtered. The crude product was
dissolved in water and added dropwise to acetone. The resulting
solid was collected by filtration and purified on a CM-25 Sephadex
column with water as eluent. The pure compound CD3 was
obtained as a white powder (1.05 g, 66%). 1H NMR d (500 MHz,
D2O): 8.40 (s, 1H), 8.35–8.20 (m, 4H), 7.80–7.69 (m, 4H), 6.05–
5.99 (m, 2H), 5.00–4.79 (m, 7H), 4.20–3.54 (m, 42H) ppm. 13C
NMR d (125 MHz, DMSO): 173.8, 160.2, 130.8, 130.7, 130.4,
128.7, 127.5, 126.1, 125.6, 123.7, 101.8, 81.0, 73.0, 72.0, 71.8,
60.2, 43.2 ppm. MS: calcd for [C57H80NO34+Na]+ 1346.14; found
1346.37. Anal. Calcd for C57H80NO34ꢃ6H2O: C, 47.82; H, 5.69; N,
0.98. Found: C, 48.02; H, 5.81; N, 1.02.
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