Degradation properties and pH-responsive guest-release of cyclophanes having four ester side chains with terminal choline residues

Article abstract of DOI:10.1246/cl.130964

Water-soluble cationic cyclophane 1 having four ester side chains with terminal choline residues was synthesized as a pH-responsive degradable host. Alkaline hydrolysis of the ester bonds of 1 gave the corresponding tetraanionic cyclophane 2, which exhibited guest-binding affinities that led to release of the entrapped anionic guest molecules into the bulk aqueous phase.

Full text of DOI:10.1246/cl.130964

CL-130964
Received: October 16, 2013 | Accepted: November 7, 2013 | Web Released: November 13, 2013
Degradation Properties and pH-responsive Guest-release of Cyclophanes
Having Four Ester Side Chains with Terminal Choline Residues
Osamu Hayashida* and Daisuke Sato
Department of Chemistry, Faculty of Science, Fukuoka University, 8-19-1 Nanakuma, Fukuoka 814-0180
(E-mail: hayashida@fukuoka-u.ac.jp)
Water-soluble cationic cyclophane 1 having four ester side
Cyclophane 1 was prepared by the condensation of a
tetraamine derivative of cyclophane 37 with the carboxylic acid
of choline derivative 48 in the presence of water-soluble
carbodiimide (WSC) and 1-hydroxybenzotriazole (HOBt). Tetra-
carboxylic acids of cyclophane 2 were prepared from 3 by a
reaction with succinic anhydride. New compounds were
characterized by means of spectroscopy (13C NMR and ESI-
MS) and elemental analysis (see the Supporting Information,
SI).9 Cyclophanes 1 and 2 had good H2O-solubility of >0.33
and >0.25 g mL¹1, respectively. These results indicate that
cyclophanes 1 and 2, which contained a hydrophobic cavity,
were expected to act as water-soluble cationic and anionic hosts,
respectively, and exhibit molecular discrimination10 between
guest molecules through hydrophobic and electrostatic inter-
actions.
chains with terminal choline residues was synthesized as a pH-
responsive degradable host. Alkaline hydrolysis of the ester
bonds of 1 gave the corresponding tetraanionic cyclophane 2,
which exhibited guest-binding affinities that led to release of the
entrapped anionic guest molecules into the bulk aqueous phase.
Currently, much effort is being devoted to develop
functionalized host vehicles having stimuli-responsive binding
capabilities1 for targeted drug delivery2 and controlled release.3
A number of cyclophane derivatives have been developed as
hosts, and their guest-binding behaviors have been widely
investigated for such purposes.4 In the preceding paper, we have
developed cleavable cyclophanes having disulfide moieties.5
The guest molecules entrapped by the hosts were released to
the bulk phase on treatment with dithiothreitol as a reducing
agent. Reduction of disulfide bonds of the hosts afforded its
reduced form, which exhibits reduced guest-binding affinity.5
In the course of our ongoing research on stimuli-responsive
degradable cyclophanes, we became interested in utilizing the
hydrolysis of ester linkages6 of cyclophanes having four cationic
side chains, with an aim to achieve pH-responsive guest release.
By increasing the pH of the aqueous solution containing the
host, the cationic cyclophane is expected to convert to anionic
cyclophane having four polar side chains with terminal carbox-
ylate residues upon hydrolysis. In this regard, we have adopted
a simple strategy by introducing terminal choline moieties into
the tetraaza[6.1.6.1]paracyclophane skeleton via degradable
ester linkages. We describe herein the synthesis of the water-
soluble cyclophane 1 having four ester side chains with terminal
choline residues, its guest-binding behavior, and the pH-
responsive degradation properties with an emphasis on the
release of guest molecules.
Alkaline hydrolysis of the ester bonds of 1 was investigated
by 1H NMR spectroscopy. Methylene proton signals at 4.52 and
3.69 ppm, originating from the side chains of 1, fully disap-
peared after 3 h of incubation with NaOD in D2O at 298 K, as
shown in Figure 1. Newly appeared proton signals at 3.96 and
3.35 ppm were attributable to the corresponding protons of
choline, which was identified (Figure 1b). In addition, ESI-TOF
MS showed that 2 was predominantly obtained by a treatment
¹
identical to NaOH in H2O (see the SI):9 m/z 1255 [M ¹ Na] .
(a)
O
O
O
N+
N
H
d
N
N
O
O
b
d
f
c
c
h
H
N
+
a N
O
O
O
g
e
2
CH3OH
f
a
e
g
b
h
Water-soluble cyclophanes 1 and 2 were synthesized by
following the reaction sequence shown in Scheme 1.
d
(b)
O
O
O
H
N
R
-
N
CO2
R
N+
Cl-
1: R =
2: R =
N
H
d
O
N
c
N
N
+
aN
O
O
O
O
HO
b
H
N
CO2 Na+
-
a
f
H
N
h
N
-
CO2
b
O
e
O
g
2
CH3OH
f,g
N
NH2
O
3: R =
R
a, b
c
R
e
h
O
aWSC, HOBt, 4, DMF
N+
Cl-
4.5
4.0
3.5
3.0
2.5
2.0
HO2C
O
b1) Succinic anhydride, DCM,
2) Ion exchange
ppm
4
1
Figure 1. Partial H NMR spectra of 1 in D2O (a) and in 4%
NaOD/D2O after 3 h incubation at 298 K (b) (400 MHz, 298 K,
CH3OH as an external standard).
Scheme 1. Preparation of cationic and anionic cyclophanes 1
and 2.
322 | Chem. Lett. 2014, 43, 322–324 | doi:10.1246/cl.130964
© 2014 The Chemical Society of Japan

1000
1000
500
250
1000
500
1000
500
(a)
(b)
0
0
200 400
[1] / µM
200 400
[2] / µM
500
500
0
350
0
350
0
450
550
450
550
100
50
150
Wavelength / nm
Wavelength / nm
Time / min
Figure 2. Fluorescence spectral changes for an aqueous
solution of TNS (1.0 ¯M) upon addition of 1 (a) and 2 (b) in
HEPES buffer at 298 K; [1] = [2] = 0, 40, 80, 120, 160, 200,
240, 280, 320, 360, and 400 ¯M (from bottom to top). Ex.
322 nm. Inset: the corresponding titration curves.
Figure 3. Time course for change of fluorescence intensity
originating TNS (1.0 ¯M) in HEPES buffer in the presence of 1
(200 ¯M) upon addition of a NaOH solution: pH was adjusted to
11.0.
Table 1. Half life of host-guest complexes of 1 with TNS (t1/2
)
at various pH conditions
These results indicate that the alkaline hydrolysis of the ester
bonds of 1 gave the corresponding tetraanionic cyclophane 2
quantitatively.
pH
t
1/2/min
10.5
10.6
10.7
10.8
10.9
11.0
120
80
61
33
30
22
The guest-binding behavior of cationic cyclophane 1 toward
anionic fluorescence guests such as 6-p-toluidinonaphthalene-
2-sulfonate (TNS) and 6-anilinonaphthalene-2-sulfonate (2,6-
ANS), whose emissions are extremely sensitive to changes in
microenvironmental polarity experienced by the molecules,11
was examined by fluorescence spectroscopy in aqueous HEPES
(2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid) buffer
(0.01 M, pH 7.4, 0.15 M with NaCl) at 298 K. Upon addition of
a large excess of 1 to an aqueous solution containing TNS, the
fluorescence intensity originating from TNS increased along
with a concomitant blue shift of the fluorescence maximum, as
shown in Figure 2a. The relative fluorescence quantum yield
increases about 71 times from 0.001 in water12 to 0.071 in 1
(400 ¯M). The binding constant for the formation of inclusion
complexes of 1 with TNS in a 1:1 molar ratio13 (K) was
evaluated on the basis of the Benesi-Hildebrand relationship
(K = 2.0 © 103 M¹1). A similar guest-binding behavior of 1 was
observed when 2,6-ANS was used as a guest (see the SI):9
K = 4.7 © 103 M¹1. On the other hand, TNS and 2,6-ANS were
weakly bound to anionic cyclophane 2, reflecting the electro-
static repulsion between 2 and the anionic guests (Figure 2b).
The K values of 2 with TNS and 2,6-ANS were not determined
due to the low affinity.
the SI).9 Hence, the alkaline hydrolysis of the ester linkages
present in 1 is responsible for the release of guest molecules.
In conclusion, cyclophane 1 having four ester side chains
with terminal choline residues was successfully synthesized. The
guest-binding affinities of 1 toward TNS and 2,6-ANS were
much larger than those of cyclophane 2, a product of hydrolysis,
reflecting the electrostatic repulsion. Accordingly, the guest
molecules entrapped by 1 were released into the bulk phase on
treatment with pH-stimuli.
The present work is partially supported by Grant-in-Aid
(No. 24550166) from the Ministry of Education, Culture,
Sports, Science and Technology of Japan.
References and Notes
pH-Stimulated guest-releasing behavior of 1 was inves-
tigated by fluorescence spectroscopy. Upon addition of NaOH to
a HEPES buffer solution containing host-guest complexes of 1
with TNS, the fluorescence intensity originating from the guest
molecules decreased, as shown in Figure 3. These results
indicate that 2 having poor guest-binding affinity14 was
generated by the alkaline hydrolysis of ester bonds of 1.
Accordingly, almost all the guest molecules entrapped by 1 were
released into the bulk aqueous phase after 180 min incubation.
Moreover, similar pH-responsive guest-releasing behavior of 1
was also observed on using 2,6-ANS as the guest (see the SI).9
1
2
3
4
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to 11.0, by fluorescence spectroscopy and are summarized in
Table 1. It is noteworthy that the t1/2 values decreased gradually
on increasing the pH value of the buffers from 10.5 to 11.0 (see
5
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SI).9
14 The K values of 2 with TNS and 2,6-ANS were not
determined due to the low affinity in CAPS buffer (0.01 M,
pH 11.0, 0.15 M with NaCl) by fluorescence spectroscopy.
7
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9
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324 | Chem. Lett. 2014, 43, 322–324 | doi:10.1246/cl.130964
© 2014 The Chemical Society of Japan