Guest releasing from solution to solid-state triggered by cyclomaltohexaose (α-cyclodextrin) aggregation.

Abstract

Supramolecuar aggregations 1 and 2 were prepared by complexing cyclomaltohexaose with two azodipyridine isomers: 4,4'-azodipyridine and 2,2'-azodipyridine, and their binding abilities and assembly behaviors were investigated comprehensively by X-ray crystallography, 2D NMR spectroscopy, and isothermal titration calorimetry. In solution, 1:1 host-guest complexation is generally assumed, whereas in the solid state, a 2:1 stoichiometry is observed. Crystal structures reveal that channel-type aggregation exists in complex 1, while a layer-type manner is the dominant packing mode in complex 2. The disparity of nitrogen atom position leads to the different binding modes and further affects the aggregation types in complexes 1 and 2.

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Guest releasing from solution to solid-state triggered by cyclomaltohexaose
(a-cyclodextrin) aggregation
Zhuo-Yi Gu, Dong-Sheng Guo, Yu Liu⇑
Department of Chemistry, State Key Laboratory of Elemento-Organic Chemistry, Nankai University, Tianjin 300071, PR China
a r t i c l ei n f o
Article history:
Received 18 March 2010
Received in revised form 14 September
2010
Accepted 23 September 2010
Available online 29 September 2010
Keywords:
Cyclomaltohexaose
Azodipyridines
Single crystal
Aggregation
a b s t r a c t
Supramolecuar aggregations 1 and 2 were prepared by complexing cyclomaltohexaose with two azodi-
pyridine isomers: 4,40-azodipyridine and 2,20-azodipyridine, and their binding abilities and assembly
behaviors were investigated comprehensively by X-ray crystallography, 2D NMR spectroscopy, and iso-
thermal titration calorimetry. In solution, 1:1 host–guest complexation is generally assumed, whereas in
the solid state, a 2:1 stoichiometry is observed. Crystal structures reveal that channel-type aggregation
exists in complex 1, while a layer-type manner is the dominant packing mode in complex 2. The disparity
of nitrogen atom position leads to the different binding modes and further affects the aggregation types
in complexes 1 and 2.
? 2010 Elsevier Ltd. All rights reserved.
1. Introduction
Cyclic oligosaccharides with 6, 7, or 8 (a-, b-, c-) a-glucose units
linked by a-(1?4)-glucose bonds,1called cyclomaltooligosaccha-
rides (cyclodextrins, CDs), have been extensively investigated in
not only molecular recognition but also self-assembly with various
well-defined nanoarchitectures.2,3Nowadays, research on CDs and
derivatives becomes more and more significant benefiting from
their benign water solubility, their biological compatibility, and
from an economics perspective, their relatively low cost. In addi-
tion, their capability of including various molecules in both the
solution and the solid state enhances their importance.4The cavi-
ties of CDs accommodate guest molecules via synergistic contribu-
tion of several non-covalent interactions, such as van der Waals,
hydrophobic, and hydrogen-bonding interactions. In order to ob-
tain direct evidence for the information of the inclusion com-
plexes,5a number of crystallographic complexes of CDs with
guests have been reported during the past few decades.6,7Many
of these studies elucidated the relationship between the structures
of the CD complexes and the structures of the guest molecules, and
further revealed that the geometrical complementarity and/or size/
shape matching between host and guest is certainly one of the
most important factors in determining the complex conformation
and packing mode.8In this context, it seems to be more fascinating
to compare the influence of subtle differences between guest mol-
ecules over the binding and overall structures of CD complexes,
especially in the case of isomeric compounds. Kamitori et al. re-
ported two crystal structures of cyclomaltohexaose (a-cyclodex-
trin) upon complexation with isomeric phenol derivative, which
give different types of aggregations resulting from the disparity
in position of bromine substituted to benzene ring.7In our
previous work,9,10we also studied a series of cyclomaltoheptaose
(b-cyclodextrin) complex crystals with similar structural guests
to explore how and to what extent the slight differences affect
the inclusion modes and aggregation structures.11,12
In the present study, we investigated the inclusion complexa-
tion of cyclomaltohexaose with 4,40-azodipyridine (4-ADP) and
2,20-azodipyridine (2-ADP) isomers by NMR spectroscopy, isother-
mal titration calorimetry (ITC), and X-ray crystallography. Similar
binding behavior was detected in solution, whereas appreciable
differences were observed in the solid state between the com-
plexes of 4-ADP (1) and 2-ADP (2). More interestingly, a process
of releasing guest molecules was found to be occurring with the
aggregation of cyclomaltohexaose units from the solution state to
the solid state.
2. Results and discussion
2.1. Binding modes in solution
2D NMR spectroscopy, as an important method for identifying
compound structures,9,13is frequently used to investigate the
inclusion geometry of CDs with guests, where the cross-peak be-
tween the protons that are closer than 0.4 nm in space will be ob-
served in NOESY spectrum, and the relative intensities of these
0008-6215/$ - see front matter ? 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2010.09.030
⇑Corresponding author.
E-mail address: yuliu@nankai.edu.cn (Y. Liu).
Carbohydrate Research 345 (2010) 2670–2675
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cross-peaks depend on the spaces between the corresponding pro-
tons.10In order to get structural information about complexes 1
and 2 in aqueous solution, 2D NMR spectra were, therefore, em-
ployed as shown in Figure 1a and b. Clear cross-peaks between
H3of cyclomaltohexaose and Ha/Hbof the 4-ADP (Peaks A and B,
Fig. 1a) as well as H5and Hb(Peak B, Fig. 1a) were observed, indi-
cating that 4-ADP is included into the hydrophobic cavity of
cyclomaltohexaose. The correlations between H6and Ha/Hb(Peak
B, Fig. 1a) illustrate that one pyridine ring of 4-ADP locates at the
narrow-rim of cyclomaltohexaose. A similar phenomenon was ob-
served in the case of 2-ADP, presenting correlations not only be-
tween H3of cyclomaltohexaose and Hc/Hdof 2-ADP (Peaks C and
D, Fig. 1b) but also between H5and Hd(Peak D, Fig. 1b), as well
as the correlated signals between H6and Ha/Hd(Peaks A and D,
Fig. 1b), with the absence of correlation between Hband cyclomal-
tohexaose protons. The reasonable binding modes were, therefore,
deduced: 4-ADP and 2-ADP penetrate axially into the cavity of
cyclomaltohexaose in almost the same manner, in which the azo
group is thoroughly immersed, while two pyridine rings point
out of both narrow and of wide-rims to some extent (Fig. 1c and
d). This is easily acceptable by taking the molecular length of guest
and height of host into account, which will be discussed more in
detail in the crystal section.
2.2. Complexation stabilities and thermodynamics
To obtain a quantitative insight on the binding stabilities (KS)
and the thermodynamic parameters (DH? and DS?) of complexes
1 and 2, ITC has been performed at 25 ?C in phosphate buffer solu-
tion (pH 7.2), and a typical titration curve of cyclomaltohexaose
with 4-ADP is shown in Figure 2. For complex 1, the titration data
could be well fitted using the ‘one set of binding sites’ model and
repeated as a 1:1 complex formation; thereby, higher order com-
plexes did not need to be postulated.14,15However, we could not
obtain reliable thermodynamic parameters for complex 2, owing
to no significant heat effect16upon cyclomaltohexaose complexing
with 2-ADP (see Section 4). Two independent measurements were
performed for complex 1, and the data obtained are as follows:
n = 0.9,
KS= 272.9 M?1
with DH? = ?3.9 kJ mol?1
?0.6 kJ mol?1for the first time, and n = 1.0, KS= 279.8 M?1with
DH? = ?3.7 kJ mol?1and TDS? = ?0.4 kJ mol?1for the second time.
And then the calculated average data with reasonable errors are as
follows: KS= 276.4 ± 3.4 M?1with DH? = ?3.8 ± 0.1 kJ mol?1and
TDS? = ?0.5 ± 0.1 kJ mol?1. Especially, both titrations show excel-
lent n values, indicating a 1:1 complexation between cyclomalto-
hexaose and 4-ADP. As can be seen from the thermodynamic
parameters, the complexation of cyclomaltohexaose with 4-ADP
is driven by a favorable enthalpy change, over-ruling the unfavor-
able entropy change. Such a favorable enthalpy change may origi-
nate from the collective contributions of the hydrophobic
interactions, the van der Waals interactions, as well as the release
of high-energy water molecules, arising from the inclusion of the
guest molecules.17,18Further, the observed negative entropy
change may be attributed to the conformational freedom limita-
tions of the 4-ADP molecule where the cis- and trans-forms of 4-
ADP may co-exist before inclusion into the CD cavity, while only
the trans form is preferred in the inclusion complex.
and
TDS? =
2.3. Crystal structures
In both of the solid-state complexes 1 and 2, the inclusion struc-
tures are in accordance with the binding modes in solution inferred
Figure 1. (a) ROESY spectra of complexes 1 and (b) 2 in D2O (5.0 mM) at 25 ?C with mixing time of 250 ms, and (c) the corresponding binding modes of cyclomaltohexaose
with 4-ADP and (d) 2-ADP.
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from the NMR studies (Fig. 3). 4-ADP and 2-ADP penetrate into the
cavity of cyclomaltohexaose with almost parallel orientation to the
cavity axis, which is somewhat different from those cyclomalto-
heptaose cases that have been reported (where guest molecules
are 2,20-/4,40-dipyridines, 4-hydroxyazobenzene and 4-aminoazo-
benzene, respectively).10,11Cyclomaltohexaose is 33.6% smaller in
cavity volume than cyclomaltoheptaose, and therefore, the depar-
ture of inclusion orientation from the cavity axis is relatively re-
strained. We measured the guest lengths and host height as
follows: 4-ADP,9.42 Å;2-ADP,
?8 Å.19These data reinforced the deduced binding modes deter-
mined by NMR spectroscopy. In addition, the distances between
the pyridine nitrogen atoms are 9.42 Å in 4-ADP and 5.67 Å in
2-ADP, which illustrates the large disparities of the aforemen-
tioned thermodynamic parameters to some extent. It can also be
clearly seen that the pyridine nitrogen atoms of 4-ADP are located
outside the cavity of cyclomaltohexaose, while the nitrogen atoms
of 2-ADP are positioned at the narrow and wide rims, respectively.
The host–guest molar ratio in the solid state is dramatically dif-
ferent from the binding stoichiometry in solution. In complex 1,
there are 1 cyclomaltohexaose unit and 0.5 4-ADP molecules in
the crystal cell and in complex 2, there are 2 cyclomaltohexaose
units and 1 2-ADP molecule. Both solid-state complexes present
a 2:1 host–guest ratio. This is an interesting and infrequent phe-
nomenon in CD complexes, where the stoichiometries are gener-
ally the same in solution and in the solid state.5–12In the present
case, the guest-release process from solution to solid state can be
well interpreted in view of aggregation of CD, guest lengths, and
binding abilities. It is well-known that CD molecules aggregate into
channels, layers, or cages through the intermolecular hydrogen
bonds donated by narrow-rim and wide-rim hydroxyl groups.8
However, it seems to be complicated for complexes 1 and 2, and
three theoretical possibilities can be assumed as illustrated in
Scheme 1 when taking into account the excessive lengths of the
ADP guests. Pattern b can be reasonably eliminated because the
cavity of the cyclomaltohexaose unit cannot accommodate two
ADP guests simultaneously according to its size (diameters of only
5.3 and 4.7 Å for the wide and narrow rims, respectively). Conse-
quently, a competition between guest inclusion and aggregation
of CD themselves emerges in fact (Patterns a and c). As proved
by ITC measurements, cyclomaltohexaose offers weak (medium
at most) binding affinities to ADP guests, and therefore, the host–
guest interactions are overcome by the hydrogen bonds leading
10.85 Å; cyclomaltohexaose,
Figure 2. ITC experiment of 4-ADP with cyclomaltohexaose in aqueous solution at 25 ?C: (left) heat effects of dilution and of complexation of cyclomaltohexaose with 4-ADP
for each injection during titration of micro-calorimetric experiment; (right) ‘net’ heat effect obtained by subtracting the heat of dilution from the heat of reaction, which was
analyzed by computer simulation with the use of the ‘one set of binding sites’ model.
Figure 3. (a) Crystal cells of complexes 1 and (b) 2 showing host and guest
molecules, only.
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to aggregation. Figure 4 shows the multiple hydrogen bonds be-
tween CD hosts. The heights of the asymmetrical units of CD aggre-
gations are 7.92 Å in 1 and 7.89 Å in 2, about 2 Å shorter than the
ADP lengths, which further confirms the rationality of Pattern c.
As shown in Scheme 2, cyclomaltohexaose forms a 1:1 complex
with 4-ADP in solution, and therein, the aggregation between
cyclomaltohexaoses is weak because the intermolecular hydrogen
bonds between the narrow/wide-rim hydroxyl groups can hardly
form because of the effect of the water medium. That is, the
host–guest inclusion affinity is dominant rather than the aggrega-
tion affinity of CD themselves in aqueous solution. However, going
with the phase transfer from liquid solution to solid state, the
aggregation affinity emerges to be the dominant force in conver-
sion. Once CD molecules arrange into channel-type structure, there
is not enough space to accommodate all the guest molecules, and
then half of the ADP guests are released before crystallization.
For a different construction of the crystal cells of 1 and 2, we in-
ferred that the termini of 4-ADP are hydrophilic nitrogen atoms,
and no other neighboring CD prefers to include the pyridine por-
tion, whereas the termini of 2-ADP are hydrophobic carbon atoms,
and the neighboring CD prefers to include the pyridine portion, al-
beit shallowly (It should be mentioned herein that no specific
host–guest interactions, such as non-conventional hydrogen bonds
and C–H???p interactions, were discussed because the ADP guests
are either the half-occupied or unordered in the CD cavity.). As a
result, all CD units in 1 are equal in position, forming head-to-tail
aggregation with half-occupied 4-ADP guest in each CD cavity.
However in 2, the asymmetrical cell is composed of two head-to-
head CD units. The 2-ADP guest mainly resides in one CD cavity,
although it is unordered and equally distributed in two positions.
The CD dimer mediated by guest molecules further forms the
tail-to-tail aggregation through intermolecular hydrogen bonds.
Taking the host molecules into account only for clarifying the
aggregation behavior of cyclomaltohexaose in complexes 1 and 2,
cyclomaltohexaose forms channel aggregation in 1 and layer
aggregation in 2 (Fig. 4). The channel in 1 extends infinitely along
the crystallographic a direction, jointed by the multiple hydrogen
bonds between 2,3- and 6-hydroxyl groups (O13–H???O15,
2.837 Å; O3–H???O5, 2.864 Å; O8–H???O10, 2.855 Å; O38–H???O30,
2.826 Å; O18–H???O20, 2.836 Å; O2–H???O30, 2.832 Å). Water mol-
ecules are filled in the interspaces between channels, forming
hydrogen bonds with the sidewalls of cyclomaltohexaoses. There
are no hydrogen bonds between cyclomaltohexaoses themselves
that link the sidewalls of the channels directly. Only hydrogen
bonds between 2,3- and 2,3-hydroxyl groups (O28–H???O47,
2.736 Å; O27–H???O48, 2.847 Å; O23–H???O52, 2.827 Å; O8–
H???O38, 2.811 Å; O12–H???O37, 2.864 Å; O18–H???O57, 2.749 Å;
O13–H???O33, 2.761 Å; O17–H???O58, 2.737 Å) were observed in
2, which then lead to the layer aggregation in the crystallographic
a ? b plane together with the hydrogen bonds between 2,3-hydro-
xyl and 2,3-hydroxyl groups on the sidewall direction (Fig. 4b,
Scheme 1. Three possible patterns in the solid-state complexes of cyclomalto-
hexaose with ADP guests: (a) inclusion without aggregation, (b) synergic inclusion
and aggregation, (c) aggregation with partial release of guest.
Figure 4. Views showing the aggregation of cyclomaltohexaose via intermolecular hydrogen bonds in complexes (a) 1 and (b) 2.
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right). No hydrogen bond between the 6-hydroxyl and 6-hydroxyl
groups forms because the narrow rims of cyclomaltohexaoses are
held apart from each other by the 2-ADP guest as a pillar.
3. Conclusions
Two solid-state complexes 1 and 2 were obtained by cyclomal-
tohexaose complexing with two azodipyridine isomers, 4-ADP and
2-ADP, presenting a host–guest molar ratio of 2:1. However, the
binding stoichiometry in solution is determined as 1:1 with a rel-
atively weak binding affinity. We infer that the complexes undergo
a release process of half of the guest molecules in going from solu-
tion to the solid state, owing to the hydrogen-bonding aggregation
of cyclomaltohexaoses themselves. Moreover, complexation of iso-
meric compounds leads to different aggregation modes: cyclomal-
tohexaose presents a 1D channel-type complex with a head-to-tail
arrangement in 1, while there is a 2D layer-type arrangement in 2
with a head-to-head orientation. The present results help us to fur-
ther understand the inclusion and aggregation behavior between
the cyclomaltohexaose host and isomeric guests and help us
understand to what extent the size/shape matching factor affects
the aggregation mode, which is beneficial in the design and con-
structionof diverse supramolecular
cyclomaltohexaose.
assemblies basedon
4. Experimental
4.1. Materials and instruments
4,40-Azodipyridine
according to the literature method.20Reagent grade cyclomalto-
hexaose was recrystallized from water twice and dried in vacuo
at 95 ?C for 24 h prior to use.
1H NMR and 2D ROESY spectra were obtained in D2O using a
Varian Mercury VX300 instrument with a mixing time of 250 ms.
The isothermal titration calorimetry (ITC) experiments were
performed by an isothermal titration microcalorimeter at atmo-
spheric pressure and at 25 ?C in aqueous phosphate buffer solution
(pH 7.2). In each run, a solution of host in a 0.250 mL syringe was
sequentially injected with stirring at 300 rpm into a solution of
guest in the sample cell (1.4227 mL volume). All thermodynamic
parameters reported in this work were obtained by using the
‘one set of binding sites’ model. Two independent titration exper-
iments were performed to afford self-consistent parameters and
to give the averaged values. For complex 1, the concentrations
were used as 40 mM for cyclomaltohexaose and 1 mM for 4-ADP.
For complex 2, we could not obtain reliable thermodynamic
parameters although various conditions were conducted. The con-
centrations selected were as follows: 10 mM for cyclomaltohexa-
ose and 1 mM for 2-ADP; 40 mM for cyclomaltohexaose and
1 mM for 2-ADP; 80 mM for cyclomaltohexaose and 1 mM for 2-
ADP; 80 mM for cyclomaltohexaose and 2 mM for 2-ADP;
100 mM for cyclomaltohexaose and 1 mM for 2-ADP.
and 2,20-azodipyridine wereprepared
The X-ray intensity data were collected on a Rigaku MM-007
rotating anode diffractometer equipped with a Saturn CCD Area
Detector System using monochromated Mo Ka radiation at
T = 113(2) K. Data collection and reduction were performed with
the use of the CYSTALCLEAR21program. The structures were solved
by using direct methods and refined by employing full-matrix least
squares on F2(CrystalStructure, SHELXTL-97).22
4.2. Preparation of cyclomaltohexaose/4-ADP crystal 1
Anaqueous solutionof 4-ADP (1 mmol,2.5 mL)was added drop-
wise to an aqueous solution of cyclomaltohexaose (1 mmol, 2.5 mL)
andstirredat40 ?Cfor5 h.Thesolutionwascooledtoroomtemper-
ature, and the precipitate was filtered and redissolved in hot water
to make a saturated solution and then cooled to room temperature.
After removing the precipitates by filtration, a small amount of
water was added to the filtrate. The resultant solution was kept at
room temperature for about two weeks. The orange-red crystals
formed were collected along with its mother liquor for the X-ray
crystallographic analysis. Yield: 43%.1H NMR (300 MHz, D2O): d
8.85 (d, 4H, py-H), 7.89 (d, 4H, py-H), 4.95 (s, 12H, 1-H), 3.83–3.45
(m, 72H) ppm. Crystal data for 1: C41H76N2O36 M = 1173.04,
monoclinic, space group P21, a = 7.9194(18), b = 13.581(4), c =
24.843(6) Å, a = 90?, b = 90.653(14)?, c = 90?, V = 2671.8(12) Å3,
F(0 0 0) = 1248, Z = 2, Dc= 1.458 g/cm?3, l = 0.129 mm?1, approxi-
mate crystal dimensions, 0.24 ? 0.22 ? 0.20 mm3, h range = 1.64–
26.00?, 18,779 measured reflections, of which 5482 (Rint= 0.0708)
were unique, final R indices [I > 2r (I)]: R1= 0.0909, wR2= 0.2480,
R indices (all data): R1= 0.0957, wR2= 0.2567, goodness of fit on
F2= 1.027.
4.3. Preparation of cyclomaltohexaose/2-ADP crystal 2
Crystal 2 was prepared by a method similar to that for 1, using2-
ADPinsteadof4-ADP.Yield:36%.1HNMR(300 MHz,D2O):d8.66(d,
2H, py-H), 8.10 (t, 2H, py-H), 7.90 (d, 2H, py-H), 7.63 (t, 2H, py-H),
4.95 (s, 12H, 1-H), 3.85–3.43 (m, 72H) ppm. Crystal data for 2:
C82H149.50N4O70.75 M = 2323.56, triclinic, space group P1, a =
13.6989(10), b = 13.9700(10), c = 15.7722(10) Å, a = 93.183(2)?,
b = 91.938(2)?, c = 118.718(2)?, V = 2636.7(3) Å3, F(0 0 0) = 1236,
Z = 2, Dc= 1.463 g/cm?3, l = 0.129 mm?1, approximate crystal
dimensions,0.12 ? 0.10 ? 0.08 mm3,
24,868 measured reflections, of which 20,083 (Rint= 0.0271) were
unique, final R indices [I > 2r (I)]: R1= 0.0487, wR2= 0.1095, R
indices (all data): R1= 0.0568, wR2= 0.1155, goodness of fit on
F2= 1.026.
h
range = 1.70–27.88?,
Supplementary data
Complete crystallographic data for the structural analyses have
been deposited with the Cambridge Crystallographic Data Centre
(CCDC Nos. 768430 for 1 and 768431 for 2). Copies of this informa-
tion may be obtained free of charge from the Director, Cambridge
Scheme 2. An illustration showing the releasing process of ADP guests from solution to solid state.
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Z.-Y. Gu et al./Carbohydrate Research 345 (2010) 2670–2675