Synthesis, structure, and binding properties of lipophilic cavitands based on a calix[4]pyrrole-resorcinarene hybrid scaffold

Article abstract of DOI:10.1021/jo5007224

We report the synthesis, structural characterization, and binding properties of a series of unprecedented cavitands based on a meso-dodecyl-calix[4]pyrrole-resorcin[4]arene hybrid scaffold. The reported structural and conformational features of the prepared cavitands are derived from results obtained in solution, solid state, and molecular modeling studies. In the solid state, these cavitands are exclusively observed in the kite C ₄ structure and as a racemic mixture of two cyclochiral conformers, which are interconverting fast on the ¹H NMR time scale, according to solution studies. In agreement, molecular modeling studies assign an energy preference for the kite conformer of the cavitands. The polar interior of the synthesized containers allows for the inclusion of a series of pyridine N-oxide derivatives. This results in the formation of 1:1 complexes that are kinetically and thermodynamically highly stable. The putative switching process between the vase and kite forms of these cavitands is investigated in solution by means of variable temperature ¹H NMR experiments. N-Oxide guests that are size and shape complementary to the volume of the cavity of the vase form are also employed to facilitate its emergence. All of the results obtained indicate the existence of a remarkable preference toward the kite conformation both in free and bound calix[4]pyrrole-based cavitands.

Full text of DOI:10.1021/jo5007224

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Article
Synthesis, Structure and Binding Properties of Lipophilic Cavitands
based on a Calix[4]pyrrole-Resorcinarene Hybrid Scaffold
Albano Galan, Eduardo C. Escudero-Adán, Antonio Frontera, and Pablo Ballester
J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/jo5007224 • Publication Date (Web): 20 May 2014
Downloaded from http://pubs.acs.org on May 26, 2014
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Synthesis, Structure and Binding Properties of
Lipophilic Cavitands based on a Calix[4]pyrrole-
Resorcinarene Hybrid Scaffold
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Albano Galán,‡ Eduardo C. Escudero-Adán,† Antonio Frontera,# and Pablo Ballester*,‡,§
‡ Institute of Chemical Research of Catalonia (ICIQ), Avgda. Països Catalans 16, 43007 Tarragona,
Spain
† X-ray Diffraction Unit, Institute of Chemical Research of Catalonia (ICIQ), Avgda. Països Catalans
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16, 43007 Tarragona, Spain
§Catalan Institution for Research and Advanced Studies (ICREA), Passeig Lluís Companys, 23, 08010
Barcelona, Spain
# Departament de Química, Universitat de les Illes Balears, Crta. Valldemossa Km. 7.5, 07122- Palma,
Spain
pballester@iciq.es
RECEIVED DATE (to be automatically inserted after your manuscript is accepted if required
according to the journal that you are submitting your paper to)
SYNOPSIS TOC
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ABSTRACT
We report the synthesis, structural characterization and binding properties of a series of unprecedented
cavitands based on a meso-dodecyl-calix[4]pyrrole-resorcin[4]arene hybrid scaffold. The reported
structural and conformational features of the prepared cavitands are derived from results obtained in
solution, solid state and molecular modeling studies. In the solid state, these cavitands are exclusively
observed in the kite C4 structure and as racemic mixture of two cyclochiral conformers, which are
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interconverting fast on the H NMR timescale, according to solution studies. In agreement, molecular
modeling studies assign an energy preference for the kite conformer of the cavitands. The polar interior
of the synthesized containers allows for the inclusion of a series of pyridine N-oxide derivatives. This
results in the formation of 1:1 complexes that are kinetically and thermodynamically highly stable. The
putative switching process between the vase and kite forms of these cavitands is investigated in solution
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by means of variable temperature H NMR experiments. N-oxide guests that are size and shape
complementary to the volume of the cavity of the vase form are also employed to facilitate its
emergence. All the results obtained indicate the existence of a remarkable preference toward the kite
conformation both in free and bound calix[4]pyrrole-based cavitands.
INTRODUCTION
Cavitands are synthetic molecules that contain an intrinsic cavity capable of including sizeable guests
(molecules or ions).1 In this sense, the elaboration of the shallow cavity provided by the cone conformer
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of resorcin[4]arene 1 through the incorporation of aromatic panels at its upper rim constitutes a valuable
synthetic strategy for the construction of deep cavitands.1,2,3,4 The upper rim functionalization of 1 is
easily accessible synthetically due to the presence of the eight hydroxyl groups. The construction of
deeper cavities has the dual effect of increasing the shielding of bound guests from the solvent and
restricting the conformational flexibility of the initial resorcinarene scaffold. In addition, extending the
cavity of a receptor molecule may confer new binding properties or modulate its kinetic and
thermodynamic binding features.5 Examples of functional deep cavitands based on multiply bridged
resorcin[4]arene structures are abundant in literature.6 These compounds have been extremely useful
both in the self-assembly of molecular capsules7,8 and in the construction of covalent cages.9
Supramolecular sensors,10 catalysts11 or even molecular grippers12,13 have also been derived from
resorcinarene based cavitands. Unfortunately, the extensive use of aromatic panels to shape and
elaborate the cavity in resorcinarene based deep cavitands renders their interiors deprived from polar
functional groups. Nevertheless, attempts to functionalize the interiors of resorcinarene based deep
cavitands with one carboxylic group14,15,16 or several pyrrole units17 have been reported.
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Calix[4]pyrroles are tetrapyrrolic macrocyclic compounds known to act as effective receptors for
anions and electron rich neutral compounds.18 The substitution at each of the four meso positions of the
calix[4]pyrrole core by a phenyl group affords aryl-extended calix[4]pyrroles containing deep cavities
defined by fixed aromatic walls.19 In 2007, our group reported the synthesis of the ,,,-isomer of
aryl-extended calix[4]pyrrole 2a decorated with eight hydroxyl groups at its upper rim (Figure 1).20 The
structure of 2a reminded us of resorcinarene 1 and was described as a resorcinarene-calix[4]pyrrole
hybrid. We envisioned that calix[4]pyrrole-resorcinarene hybrid 2a could be a suitable scaffold for the
construction of deep cavitands functionalized with four pyrrole units at their closed ends. Our
expectations were that the synthetic strategies used in the preparation of deep cavitands derived from
resorcin[4]arene 1 could be directly applied to further elaborate the aromatic cavity of 2. Most of these
synthetic strategies involved the bridging of vicinal hydroxyl groups at the upper rim with suitable
aromatic spacers. To the best of our knowledge, there is only one example of cavitands derived from
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aryl-extended calix[4]pyrrole scaffolds in literature.21 However, the described compounds do not qualify
as deep cavitands. Their structures correspond to doubly bridged phosphonated cavitands derived from a
tetraol aryl extended calix[4]pyrrole. These compounds are reminiscent of the phosphonate-
calix[4]arene cavitands of Dalcanale22,23,24 and Dutasta.25,26 Here, we describe the synthesis of a series
of tetrabridged deep cavitands derived from calix[4]pyrrole hybrid 2b. We also report the
conformational properties of the synthesized cavitands, both in solution and in solid state. Finally, we
discuss the experimental results of the binding studies of these novel cavitands with pyridine N-oxide
derivatives.
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Figure 1. Molecular structures of resorcin[4]arene 1 and calix[4]pyrrole-resorcin[4]arene hybrid 2.
RESULTS AND DISCUSSION
Synthesis of lipophilic calix[4]pyrrole-resorcin[4]arene hybrid 2b. All synthetic attempts to elongate
the cavity of built-in calix[4]pyrrole 2a led to reaction mixtures with low solubility in most common
organic solvents. The limited solubility of the crude reaction product made the analysis of the mixture
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and its purification troublesome. To overcome this limitation, we decided to use the more lipophilic
calix[4]pyrrole derivative 2b as the initial building block. Calix[4]pyrrole 2b is a structural analog to 2c
but contains an extra carbon atom in its meso-alkyl chains. A synthetic procedure for calix[4]pyrrole 2c
was recently reported by Cohen.27 Inspired by the reported synthetic methodology, we prepared
dodecyl-3,5-dihydroxy-phenyl ketone 6 in 3 steps. The nucleophilic addition of dodecyl magnesium
bromide to 3,5-dimethoxybenzaldehyde 3 afforded the benzylic alcohol 4 in 95% yield. Oxidation of
alcohol 4 with PCC provided ketone 5, which was demethylated with molten pyridinium chloride
affording the dihydroxyphenyl-dodecyl ketone 6 in 14% overall yield. Finally, the acid catalyzed
condensation of phenyl ketone 6 with pyrrole afforded, after column chromatography purification and
crystallization from acetonitrile, the calixpyrrole-resorcinarene hybrid 2b as a white solid in 19% yield.
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Scheme 1. Synthetic scheme for the preparation of the calix[4]pyrrole-resorcin[4]arene hybrid 2b.
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As shown in Figure 2, the H NMR spectrum of 2b in CDCl3 solution shows broad proton signals. We
assume that similarly to the reported findings for 2c this is mainly due to aggregation phenomena.
Conversely, in polar organic solvents that disrupt intermolecular hydrogen-bonding interactions, like
MeOH-d4 or THF-d8, the 1H-NMR spectrum of 2b shows sharp and well-defined proton signals.
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Figure 2. Selected downfield region of H NMR spectra of 10 mM solutions of calixpyrrole 2b a) in
MeOH-d4 and b) in CDCl3. * Residual peak of non-deuterated solvents. See Scheme 1 for proton
assignment.
Single crystals of 2b suitable for X-ray diffraction were grown from an acetonitrile solution. The
analysis of the diffraction data revealed that in the solid state calixpyrrole 2b adopts the cone
conformation and features one molecule of acetonitrile included in its aromatic cavity. The cone
conformation of 2b is stabilized by the formation of intramolecular hydrogen bonds between the
hydroxyl groups at its upper rim. In addition, the included acetonitrile molecule is hydrogen bonded
simultaneously to the four convergent pyrrolic NHs of 2b. The binding geometry of the solvate
CH3CN2b is a key factor in stabilizing the calix[4]pyrrole core in the cone conformation. The
averaged N···N distance of the four hydrogen-bonds is 3.22 Å. The acetonitrile solvate CH3CN2b
packs into rows through the intermediacy of water molecules that are hydrogen bonded to two adjacent
solvates in the same row.
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Figure 3. a) X-ray structure of the solvate CH3CN2b; b) adjacent solvates of CH3CN2b form rows;
c) three rows of adjacent CH3CN2b solvates pack to form a layer with the polar heads directed in
opposite directions; d) side and e) front views of the bilayers resulting from the packing of layers of
solvates. For clarity, non-polar hydrogen atoms are removed in a) and all hydrogen atoms in b), c) and
d). Selected acetonitrile molecules are shown as CPK models and 2b is depicted in stick representation.
The meso-alkyl substituents (tails) belonging to adjacent CH3CN2b complexes are involved in van-
der-Waals interactions. Each row of CH3CN2b solvates is flanked by two analogous parallel rows
with the polar upper rims (head) of 2b oriented in opposite directions. This arrangement of rows forms a
layer with alternating lipophilic and hydrophilic strips. Layers pack on top of each other by confronting
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strips of the same polarity. Thus, head-to-head interactions between CH3CN2b solvates in different
layers are promoted through water-mediated hydrogen bonding interactions while tail-to-tail contacts
are mainly of van-der-Waals nature. Each molecule of 2b is involved in head-to-head interactions with
two other molecules of 2b pertaining to two contiguous rows of an adjacent layer. Acetonitrile
molecules fill the voids produced in the described packing. The crystal packing demonstrates the
existence of bilayers of calixpyrrole molecules 2b, in which the polar and non-polar sections segregate.
This result highlights the amphiphilic character of 2b.
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Synthesis of calix[4]pyrrole based deep cavitands. The construction of deep cavitands based on
resorcin[4]arene 1 relies in the elaboration of its aromatic cavity by incorporation of four aromatic flaps
at its upper rim. The additional aromatic flaps are easily introduced through nucleophilic aromatic
substitution reactions between the hydroxyl groups at the upper rim of 1 and ortho-disubstituted fluoro
and chloro electron-deficient six-membered aromatic and hetero-aromatic rings.28
Using analogous synthetic procedures to the ones applied for the elaboration of the cavity of 1, we set
out to synthesize deep cavitands derived from the calix[4]pyrrole-resorcin[4]arene hybrid 2b. The
reaction of hybrid 2b with 4 equivalents of ortho-1,2-difluoro-4,5-dinitro benzene 7 carried out by
heating a dimethylformamide (DMF) solution at 85 ºC in the presence of an excess of triethylamine
afforded, after column chromatography purification, the four-folded bridged cavitand 8 as a yellow solid
(mp > 251 ºC, decomp) in 47 % yield. We did not detect the presence of partially (reacted) bridged
products in the reaction mixture.
Similarly, the reaction of 2b with 1,2-difluoro-4,5-dicyano benzene 9 produced octanitrile
calix[4]pyrrole cavitand 10 as a white solid (mp > 292 ºC, decomp) in 69% yield (Scheme 3).
The synthesis of cavitands 12 and 14 required the modification of the reaction conditions. Thus, 2b
was reacted with 4 equivalents of 2,3-dichloroquinoxaline 11 using potassium carbonate as a base in
DMF. After purification of the crude reaction, cavitand 12 was isolated as a white solid in a 51% yield
(mp > 328 ºC, decomp). Analogously, the reaction of 2,3-dichloro-1,4-pyrazine 13 with 2b afforded
cavitand 14 as a white solid (mp > 275 ºC, decomp) in 12% yield.
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Scheme 2. Synthetic schemes for the preparation of deep cavitands 8, 10, 12 and 14.
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Structures of the vase and kite conformers of calix[4]pyrrole based deep cavitands,
thermodynamic stabilities and interconversion processes.
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Molecular modeling studies indicate that in cavitands 8, 10, 12, and 14 the base of four phenyl rings,
which are connected at the meso positions of the calix[4]pyrrole unit, and the four fifteen-membered
rings formed an enforced concave cavity closed at one end by the four pyrrole rings. Regardless of the
starting geometry assigned to the calix[4]pyrrole core of the four cavitands, the energy minimized
structure always converged to a cone conformation. In short, it was not possible to energy minimize any
of these cavitands with the calix[4]pyrrole core exhibiting 1,2- or 1,3-alternate conformations. The
energy minimized models showed that the four phenyl bridging groups attached at the upper rims of 8
and 10, the diazanaphthalenes of 12 and the four 1,3-pyrazine units in 14 resemble flaps that can adopt
axial (a) or equatorial (e) orientations. Consequently and in close analogy to the cavitands derived from
resorcin[4]arene 1,29 we expected that cavitands 8, 10, 12 and 14 could also adopt kite and vase
conformations. The structures of the new cavitands in both conformations were energy minimized using
MM3 (see Figure 4 for cavitand 10 including one molecule of DMF).
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Figure 4. Side and top views of the MM3 energy minimized structures of: a) vase (a,a,a,a) conformer of
cavitand 10 modeled with one molecule of DMF (shown as CPK model) and one molecule of
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chloroform (stick representation) included in its cavity, and b) kite (e,e,e,e) conformer with C4v-like
symmetry. Non-polar hydrogen atoms are omitted for clarity. The kite conformer is modeled with just
one molecule of DMF included. The average orientation of the lone-pairs on the oxygen atoms (Lp) in
the two structures is indicated by green vectors included in the plane defined by the Lp1-oxygen atom-
Lp2 and bisecting the angle they define. Lp = lone-pair electrons of the oxygen atom.
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The MM3 energy minimized vase (a,a,a,a) conformer of the calix[4]pyrrole cavitands possess C4v-like
symmetry. In this conformation, the fixed aromatic cavity defined by the meso-phenyl substituents is
significantly increased by the four bridging aromatic panels. For example, in the energy minimized
structure of the vase conformer of 10 one molecule of DMF and one molecule of chloroform could be
included in its deep aromatic cavity (Figure 4a). The MM3 energy minimized kite structures of the
cavitands also displayed C4v-like symmetry. As shown for the kite conformer of 10 (Figure 4b) the lone
pairs of the eight oxygen atoms are outwardly directed with respect to its fixed aromatic cavity. Using
the MM3 force-field as implemented in the Scigress Program,30 the energy difference for the two
conformers of 10 containing one DMF molecule included in their aromatic cavity is 12.7 kcal/mol more
favorable as the kite.
Figure 5. Energy minimized structure of the kite conformer of 10 displaying C4 symmetry at the BP86-
D3/def2-SVP level of theory.
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We computed the energy difference between the empty vase and kite conformers of 10 at the
BP8631,32-D333/def2-SVP level of theory using TURBOMOLE version 6.4.34 C4 symmetry restrictions
were imposed to both structures in order to speed up the calculation time. Interestingly, the energy
minimized kite conformer of 10 with C4 symmetry featured the lone pairs of four of the eight oxygen
atoms inwardly directed with respect to the aromatic cavity representing an example of an inherently
chiral concave molecule (Figure 5).35 The cyclochiral conformer shown in Figure 5 is defined to have M
axial chirality assuming that the oxygen atom with the inwardly directed lone pair takes priority over the
one having outwardly directed lone pairs.36,37 This structure is reminiscent to the kite conformation of
resorcinarene cavitands, however in this latter case they exhibited C2v symmetry.1,2 At this level of
theory, the energy difference between the inclusion complexes of 4-phenyl-pyridine-N-oxide 17 with
the vase and kite conformers of 10 is 8 kcal/mol in favor of the kite (see SI). Taken in concert, the
results of the theoretical calculations indicated that cavitand 10 should adopt exclusively the kite
conformation, either displaying C4v or C4 symmetry. A reverse trend of energies was computed for the
two forms of the inclusion complex of the same N-oxide and the pyrazine-based cavitand 1714 (E =
8.8 kcal/mol in favor of the vase) (SI). Moreover, diazanaphthalene cavitand 12 shows a significant
energy preference for the vase form of the inclusion complex with phenyl pyridine N-oxide, 1712 (E
= 22 kcal/mol in favor of the vase) (SI).
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The 1H NMR spectra of the deep cavitands 10, 12 and 14 taken in different solvents (CD2Cl2, toluene-
d8) and at different temperatures (-60 ºC to 100 ºC) show sharp proton signals, the number of which is
consistent with a C4v symmetry. The signals do not experience substantial changes in their chemical
shifts and shapes as the temperature is changed. At room temperature and in CDCl3 solutions the -
pyrrolic protons (H2) of the aromatic bridged cavitands resonate at  ~ 5.7 ppm (see Scheme 2 and
Figure 4 for proton assignment). Thus, they appear upfield ( = - 0.32 ppm) from the -pyrrolic
protons (H2) of the conformationally rigid methylene bridged cavitand 15 (Figure 6) that resonate at  =
6.02 ppm. In addition, the doublet of the H3 protons in the resorcinol units of the aryl bridged cavitands
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( = 5.84, 5.87, 5.76, and 5.54 ppm for 8, 10, 12 and 14 respectively) are also shifted upfield ( ~ -
0.4-0.7 ppm) with respect to the same signal in 15 ( = 6.3 ppm). In short, the -pyrrolic protons H2 and
the aromatic protons H3 in cavitands 8, 10, 12, and 14 must be close to the shielding cone exerted by the
bridging aromatic ring. In sum, these NMR observations suggest that in solution and in the range of
studied temperatures the aromatic bridged cavitands are locked in the kite (e,e,e,e) conformation. A
result that is in not in agreement with the theoretical calculations discussed above.
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Figure 6. Synthetic scheme of the methylene bridged cavitand 15.
Moreover, a kite conformer displaying C4v symmetry would explain the reduced number of proton
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signals observed in the H NMR spectra. However, the putative existence of a chemical exchange
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process (cycloenantiomerization) between two cyclochiral C4 kite conformers that is fast on the H
NMR chemical shift time scale at any of the investigated temperatures (213-378 K) could also explain
the absence of signal splitting for diastereotopic protons, i.e. H4 and H4’, which can be expected for a
kite structure with C4 symmetry (Figure 7).
Luckily, single crystals of cavitands 8, 10 and 12 suitable for X-ray diffraction analysis grew from
separate solutions of acetonitrile or acetonitrile/dichloromethane mixtures. In the three cases, the
cavitands, in the solid state, adopted a kite conformation with C4 symmetry having one molecule of
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solvent included in their fixed aromatic cavites: DMF for 10 and CH3CN for 8 and 12. The solvent
molecule is hydrogen bonded simultaneously to the four pyrrole N-H hydrogen atoms and the
calix[4]pyrrole core adopts a cone conformation (Figure 7).
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Figure 7. Side and top views of the X-ray structures of the C4 kite conformer of cavitands: a) DMF10
and b) CH3CN12. Only the M cyclochiral conformer is shown and the length of the meso-alkyl groups
is truncated to three methylenes for clarity. The cavitands are displayed in stick representation and the
included solvent molecules are shown as CPK models.
For the three different cavitands, as could be expected, the two cyclochiral (M and P) enantiomers of
the C4 kite conformer are present in the packing of the lattice. Most likely, in solution the kite structures
of the cavitands are also C4 symmetric but the two cyclochiral conformers are rapidly interconverting on
the 1H NMR timescale (Scheme 3).
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The Journal of Organic Chemistry
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Scheme 3. Cycloenantiomerizaton equilibrium between the two cyclochiral M and P kite C4 conformers
of 8. The structures of the two cyclochiral enantiomers were extracted from the packing of the X-ray
structure of the crystal. The meso-alkyl groups are truncated to three carbons for clarity. In solution the
interconversion process is fast on the 1H NMR chemical shift timescale and the diasterotopic protons H4
and H4’ resonate as one signal even at temperatures below 220 K
The structure of the transition state for the cycloenantiomerization process was investigated by means
of theoretical calculations at the BP86-D3/def2-SVP level of theory for the interconversion between the
cyclochiral conformers of the octacyano cavitand 10 (see SI). The energy of the transition state lies just
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0.9 kcal/mol higher than the C4 kite ground state of the cyclochiral conformers. The structure of the
transition state is similar to that of a kite conformer with C4v symmetry obtained in the energy
minimization runs using MM3 (Figure 4b and SI). The low energy barrier computed for the transition
state suggests that it is not possible to induce a slow exchange interconversion process on the NMR
timescale by lowering the temperature. Consequently, in agreement with experiment, the diastereotopic
protons H4 and H4’ (see Scheme 2 and Scheme 3 for proton assignment) of the C4 kite conformers are
observed as one signal.
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Molecular recognition studies of pyridine N-oxides derivatives by calix[4]pyrrole based deep
cavitands. Attempts to change the relative thermodynamic stabilities of the vase and kite forms by
guest inclusion.
Pyridine N-oxides are important targets for molecular recognition studies due to their pronounced
biological activity.38,39 We reported that aryl extended calix[4]pyrroles40 and their supramolecular
capsular derivatives41,42,43 bind pyridine N-oxides and other aromatic and aliphatic N-oxides with high
affinity. The oxygen atom of the N-oxide forms four simultaneous hydrogen bonds with the pyrrole NHs
of the calix[4]pyrrole core. The N-oxide guests are bound deep in the aromatic cavity of the host and
experience additional CH- and - intermolecular interactions with the meso-aryl substituents. Based
on our previous findings, we decided to investigate the interaction of the calix[4]pyrrole deep cavitands
synthesized in this work with a series of pyridine N-oxides (Figure 8).
Figure 8. Molecular structures of the pyridine N-oxide derivatives used in this work
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The Journal of Organic Chemistry
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We first probed the binding of the N-oxides by these cavitands in CDCl3 solution using H NMR
spectroscopy. The addition of 0.5 equivalents of 4-dimethylaminopyridine N-oxide 16 to a 1 mM CDCl3
solution of octanitro cavitand 8 resulted in two sets of signals of equal intensity for the protons of the
host (Figure 9). The two sets of signals belong to the protons of free and bound host 8, indicating that
the chemical exchange process is slow on the 1H NMR timescale. The large downfield shift (Δ = 1.29
ppm) observed for the pyrrole NHs of bound 8 is indicative of the formation of hydrogen bonds with the
oxygen atom of the bound N-oxide 16. Protons H2, H3 and H4 of host 8 experience reduced changes of
their chemical shifts with respect to the free state, while H1 shifts slightly upfield (Δ = -0.23 ppm).
This effect is probably due to the shielding effect of the bound N-oxide. The signals assigned to the
aromatic protons of bound 16 moved upfield (Ha Δ = -3.13 ppm and Hb Δ = -0.42 ppm) supporting
the inclusion of the N-oxide in the fixed aromatic cavity of 8. In the presence of 1 equivalent of 16 only
the proton signals assigned to the bound host and the bound guest were detected. This observation
indicates the quantitative formation of a 1:1 complex, 168, for which a stability constant value larger
than 104 M-1 can be estimated. The addition of an excess of 16 did not induce any change in the
chemical shifts of the protons assigned to the 1:1 complex. However, signals for the protons of free 16
became visible in the 1H NMR spectrum of the mixture. A 2D-EXSY experiment performed on a 1 mM
solution of cavitand 8 containing 1.5 equivalent of N-oxide 16 revealed the absence of cross peaks
between the signals of the free and bound N-oxide. This result shows that the exchange rate between the
free and bound N-oxide is too slow to be measured with this technique and suggest that the 168
complex is kinetically very stable (> 5 s).
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It is worth noting that the incremental addition of N-oxide 16 to the CDCl3 solution of 8 induced the
emergence and subsequent growth of the proton signals characteristic for DMF. We rationalized this
observation assuming that cavitand 8 is isolated as a DMF solvate. The DMF molecule is included in the
cavity of 8 and it is bound to the four pyrrole NHs through the oxygen atom (see X-ray structure of 10
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in Figure 10). The included DMF could not be removed even after heating a sample of 8 for 24 h under
high vacuum conditions. Most likely, the octanitro cavitand 8 binds N-oxide 16 considerably stronger
than DMF inducing the exchange of guests and the release of free DMF to the bulk solution. The
exchange of a bound DMF molecule by the included N-oxide is observed in all titration experiments
performed with the cavitand series. The elemental analysis of the yellow solid isolated as cavitand 8 is
in agreement with the presence of one molecule of DMF for each cavitand.
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Figure 9. Selected region of 1H NMR spectra acquired during the titration of a 1 mM CDCl3 solution of
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cavitand 8 with incremental amounts of DMAP N-oxide 16: a) 0 eq. b) 0.5 eq. c) 1.5 eq. Panel d) H
NMR spectra of free N-oxide 16 in CDCl3 solution. Primed letters and numbers indicate protons in the
168 complex. See Scheme 2 and Figure 8 for proton assignments.
The minimal chemical shift changes observed for the H2 and H3 protons of 8 upon binding suggest
that in the 168 complex the cavitand maintains the kite conformation. Single crystals of the 168
complex suitable for X-ray diffraction grew from the CDCl3 solution. The solid state structure of the
168 complex is in complete agreement with that assigned in solution based on the observed chemical
shift changes. In the crystal, the bound octanitro cavitand 8 is in the kite form with the calix[4]pyrrole
core in the cone conformation displaying an overall C4-like symmetry. The oxygen atom of the bound
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The Journal of Organic Chemistry
N-oxide 16 is hydrogen bonded to the four pyrrole NHs. The aromatic protons, alpha to the nitrogen
atom, are included in the fixed aromatic cavity of 8. The pyridyl group of the N-oxide experiences
simultaneous - and CH- interactions with the meso-aryl walls of the cavitand.
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Figure 10. Side and top views of the X-ray structure of the 168 complex. Non polar hydrogen atoms
of 8 are removed and alkyl chains are pruned for clarity. N-oxide 16 is shown as CPK model and 8 in
stick representation.
Completely analogous results were obtained in the titration of octacyano cavitand 10 with 4-
dimethylaminopyridine N-oxide 16.
After the addition of 1 equivalent of DMAP N-oxide 16 to a 1 mM CDCl3 solution of cavitand 12, the
formation of a 1:1 inclusion complex was observed in the 1H NMR spectrum. The large downfield shift
of the pyrrolic NHs (Δ = 1.59 ppm) and the upfield shift of the protons for the included guest clearly
support the formation of the inclusion complex 1612. In contrast with the titration results obtained for
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the previous cavitands, 8 and 10, the formation of the 1:1 inclusion complex between the DMAP N-
oxide 16 and the tetradiazanaphthalene cavitand 12 generated a reduced upfield shift for the H1 proton
(Δ = -0.08 ppm for 1612 vs -0.23 ppm 168 and 1610). Concomitantly, the H3 proton of bound
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cavitand 12 moved downfield (Δ = 0.15 ppm). The chemical shift of the H3 proton in cavitands 8 and
10 was almost unaffected upon N-oxide binding. These unparalleled chemical shift changes observed
for the H1 and H3 protons in the titration of tetradiazanaphthalene cavitand 12 with 16 suggested to us
the possible existence of a conformational switch of the diazanaphthalene flaps, from the axial to
equatorial position, induced by the binding of the N-oxide. That is, the 1612 complex could exist in
solution as an undetermined mixture of vase and kite forms of the cavitand involved in a fast chemical
exchange on the 1H NMR timescale.
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To further explore the putative complexation induced conformational switch of 12, we decided to use
4-phenylpyridine-N-oxide 17 as a guest. Our expectations were that the para-phenyl substituent of N-
oxide 17 should favor the formation of the 1712 complex in vase form. On the one hand, the phenyl
substituent is shape and size complementary to the additional aromatic cavity present in the vase
conformer of 12 compared to the kite. Additionally, the vase conformer of complex 1712 should be
energetically favored by additional π-π and CH- interactions established between the para-phenyl
substituent of 17 and the diazanaphthalene flaps in the axial conformation (Figure 11). As expected, the
interaction of phenyl N-oxide 17 with cavitand 12 featured a slow chemical exchange process on the 1H
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NMR timescale. The H NMR analysis of an equimolar mixture of 17 and 12 in CDCl3 solution
revealed the quantitative formation of the 1712 complex. Unexpectedly, the magnitude of the
downfield shift experienced by the H3 proton in the 1712 complex (Δ = 0.15 ppm) was coincident
with the one observed for inclusion of the shorter N-oxide 16. This result constituted an indication of the
failure to provoke a notable guest-induced switch of the kite to the vase form in the inclusion complex
1712.
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Figure 11. MM3 energy minimized structure of the putative 1712 complex in the vase form.
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In addition, the signals of the protons pertaining to the para-phenyl substituent of included 17 did not
shift noticeably with respect to those of the free N-oxide. To evaluate if the modification of the
electronic properties of the p-phenyl ring of the pyridine N-oxide had any effect on the possible
conformational switch of the bound cavitand 12, we prepared 4-(4-methoxyphenyl)pyridine N-oxide 18
and 4-(4-nitrophenyl)pyridine N-oxide 19 (Figure 8). Both pyridine N-oxide derivatives produced 1:1
complexes with 12. The stability constant values for these complexes can be estimated to be larger than
104 M-1 in CDCl3 solution. In the case of the methoxy-N-oxide 18, the signal of the H3 proton of 12 in
the inclusion complex 1812 shifts upfield 0.14 ppm, a value that coincides with the one observed for
the 1712 complex. Interestingly, the upfield shift experienced by the H3 proton in the 1912 complex
with the nitro N-oxide is larger, 0.21 ppm. Additionally, the signal of the ortho protons of the p-
nitrophenyl ring of 19 in the latter complex move somewhat upfield (Δδ = -0.23 ppm). Taken together,
these results suggest, that in solution, the nitro N-oxide 19 is capable of inducing a kite to vase
conformational switch of the bound cavitand 12 in a larger, although undetermined, extent than the
other pyridine N-oxide derivatives 17 and 18. In contrast with these findings, the analysis of the X-ray
diffraction data of single crystals of complexes 1712, 1812 and 1912 revealed that, in the solid
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state, cavitand 12 exclusively adopts the kite form in the three inclusion complexes (Supporting
Information).
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Pyridine N-oxides 18 and 19 were also used to investigate the putative guest induced conformational
switch using the shorter aryl bridged cavitands 10 and 14. The obtained results point to exclusive
formation of inclusion complexes in which the shorter cavitands are in the kite form (Supporting
Information).
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Tetramethylene bridged cavitand 15 is deprived of aromatic flaps and constitutes an ideal reference to
determine the expected chemical shift changes for the protons of the host induced by the inclusion of N-
oxides. The equimolar mixture of methylene bridged cavitand 15 and 4-phenylpyridine N-oxide 17 in
CDCl3 solution at 1 mM concentration produced the quantitative formation of the 1715 complex. The
1H NMR spectrum of the mixture indicated that, in response to guest inclusion, the H1 proton in 15
moves upfield (Δ = -0.53 ppm). Most likely, the observed chemical shift change is due to the shielding
effect produced by the para-phenyl group of the included N-oxide. In addition, the chemical shift values
of protons H2 and H3 in 15 are insensitive to the inclusion process. These results provide strong support
to the hypothesis that the detected changes in chemical shifts for equivalent protons in cavitand 12 are
indeed caused by a conformational change in the aromatic flaps.
We wanted to evaluate the effect of temperature in the kite to vase conversion of cavitand 12 when
involved in the formation of inclusion complexes with N-oxides. It is known that in related cavitands
derived from a resocin[4]arene scaffold, the vase form is favored at high temperatures owed to an
increase in the negative entropic term over the positive enthalpy component of the kite to vase
conversion.44 The variable temperature experiments were performed in solutions of tetrachloroethane-
d2, a solvent with a boiling point of 146 ºC. In this solvent and at 298 K, the chemical shift changes
experienced by the protons of the complexes between cavitand 12 and the series of pyridine N-oxides
17, 18 and 19 were similar to those observed in CDCl3 solution.
Increasing the temperature to 398 K amplifies the chemical shift changes. For the three inclusion
complexes, the downfield shift experienced by H3 in bound 12 increases two-fold. Similarly, the upfield
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shift of the ortho-proton, Hc, in the phenyl substituent of the included N-oxides doubles its magnitude.
Taken in concert, these data suggest that the increase in temperature favors the vase form of the cavitand
12 in the inclusion complexes. Unfortunately, the available data do not allow the determination of an
accurate percentage of kite and vase forms of the cavitand in these complexes at different temperatures.
Similar variable temperature experiments performed with an equimolar mixture of the shorter
tetrapyrazine cavitand 14 and methoxyphenyl N-oxide 18, revealed that in the 1814 complex the
chemical shifts of protons H3 and Hc are not modified by temperature changes. This result underlines a
clear conformational preference for the kite form in bound cavitand 14. This is in contrast with the
observations made with related pyrazine cavitands based on resocin[4]arene scaffolds, which are locked
in the vase form.2
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From the results obtained in the H NMR titration experiments, the stability constants of the 1:1
inclusion complexes formed between cavitands, 8, 10, 12 and 14, and pyridine N-oxide derivatives, 16,
17, 18, 19 and 20, were estimated to be larger than 104 M-1 (quantitative formation of the 1:1 complex in
the presence of equimolar amounts of host and guest at mM concentration). For this reason, we
undertook the calculation of accurate values for the stability constants of these 1:1 complexes using
isothermal titration calorimetry (ITC) experiments. The ITC data obtained for the titrations of
designated cavitands (12 and 10) with phenyl pyridine N-oxide 17 pictured double sigmoid curves.
Figure 12 displays the plot of the normalized integrated heat (black diamonds) vs molar ratio [17]/[12]
for the titration of 12 with phenyl-N-oxide 17.
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Figure 12. Plot of the normalized integration heat (black diamonds) vs [17]/[12] molar ratio obtained
for the ITC experiment of 12 (0.99 mM) with 17 (9.8 mM) in CHCl3. Theoretical binding isotherm (red
line) for a binding model that considers three species, 12, DMF, 17, and two complexes DMF12 and
1712, fit to the experimental data is also shown.
The two inflexion points of the double sigmoid binding isotherm are centered at values close to 0.25
and 1 for the molar ratio [17]/[12]. ITC dilution experiments performed on both the cavitand and the
pyridine N-oxide showed insignificant release of heat. In addition, a DOSY NMR experiment carried
out with a CDCl3 solution containing a 2:1 mixture of cavitand 12 and phenyl N-oxide 17 provided
similar diffusion coefficient values for the free and bound cavitand. These findings ruled out the
existence of dimerization/oligomerization processes for 12 or 17 in solution in the range of used
concentrations. As we already commented above, the solid isolated as cavitand 12 was in fact, to a great
extent, a DMF solvate of 12. Based on this precedent and using the HypDH software, we were able to fit
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the double sigmoid binding curve to a theoretical binding model that considers the existence of three
species and five stoichiometric states: free 12, free DMF, free 17, DMF12 and 1712 (Figure 12). The
concentration of DMF in the calorimetric cell was determined to be 0.85  [12].45 In short, only 15% of
the molecules in the solid isolated as 12 do not include a molecule of DMF. The fit of the ITC data
returned the values of H(DMF12) = - 4.1 kcal/mol, H(1712) = - 6.3 kcal/mol, K(DMF12) = 5.5
 105 M-1 and K(1712) = 6.3  107 M-1. The two orders of magnitude difference measured for the
stability constants of the complexes of 12 indicates the almost exclusive formation of the 1712
complex when DMF and N-oxide 17 are in solution in an equimolar ratio with respect to 12. The
speciation profiles derived from the ITC experiments are in complete agreement with the observations
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made in the H NMR titrations of 12 with 17 (release of approximately one equivalent of DMF to the
bulk solution on addition of one equivalent of 17). We attempted to remove the DMF from the solid
isolated as cavitand 12 by continuous heating under high vacuum. However, the ITC experiments
performed with samples of 12 treated as indicated above also produced double-sigmoid binding curves
with reduced changes in the molar ratio of the first inflexion point. The deep inclusion of DMF in the
conformationally rigid aromatic cavity of 12 means that its release is energetically highly disfavored
because it produces an empty cavity (vacuum). The deep inclusion nature of complexes 1712 and
DMF12 is also responsible of the favorable entropic terms (3.3 kcal/mol and 3.7 kcal/mol,
respectively) determined for their complexation processes. Both host and guest must experience strong
desolvation processes prior to the formation of the inclusion complex. The release of solvating
chloroform molecules to the bulk solution assists binding entropically, even in such a non-polar solvent,
overcoming the loss of translational and rotational entropy of one of the partners and of the
conformational flexibility of host and guest bound in the complex.
We decided to grade the stability constant values of some inclusion complexes of cavitands 12 and 10
with designated phenyl-N-oxides by performing pairwise competitive experiments and analyzing them
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using H NMR spectroscopy. An equimolar mixture of cavitand 12 with phenyl-N-oxide 17 and
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methoxyphenyl-N-oxide 18 in CDCl3 produced inclusion complexes 1712 and 1812 in an exact 1:1
ratio, demonstrating that the stability constant values of the two complexes are independent of the
electronic nature of the para-phenyl substituent of the N-oxide. We also performed a pairwise
competitive experiment by mixing equimolar amounts of cavitands 10 and 12 with methoxyphenyl- N-
oxide 18. The obtained CDCl3 solution was analyzed by 1H NMR spectroscopy indicating the formation
of complexes 1810 and 1812 in equal amounts. This latter result proves that the stability constants of
the caviplexes resulting from calix[4]pyrrole cavitands 10 and 12 and pyridine N-oxides are not
significantly influenced by the characteristics of the bridging aromatic ring.
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The preference exhibited by cavitand 12 to adopt the kite form in the inclusion complexes with
pyridine N-oxides prompted us to investigate the formation of three-component aggregates involving
one molecule of a bis-N-oxide (ditopic guest) and two units of cavitand 12. Molecular modelling studies
indicate that 4,4’-dipyridyl-N,N’-dioxide 20 is a suitable molecular platform to study such assembly
process. The energy minimized structure of the complex 20122 did not show clear signs of attractive
or repulsion interactions between the two units of 12 (Figure 13).
Figure 13. Energy minimized structure of the 20122 complex. One unit of 12 is depicted in stick
representation, the other as van der Waals surface and the bis-N-oxide 20 is shown as CPK model.
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The Journal of Organic Chemistry
1
The binding of bis-N-oxide 20 with 12 was probed in CDCl3 solution using H NMR spectroscopy.
Addition of 0.25 equivalents of bis-N-oxide 20 resulted in the appearance of a new set of signals (Figure
14b). We assigned the new signals to the protons of the cavitand 12 in the 20122 complex. The pyrrole
NHs of the cavitand in the 1:2 complex resonate downfield at δ = 9.27 ppm indicating the formation of
hydrogen bonds with the oxygen atom of bound 20. The protons of bound 20 appear highly upfield
shifted: i.e.  = 5.82 ppm and 7.68 ppm for Ha” and Hb”, respectively and as unique set. These
observations are in agreement with the inclusion of each binding site of 20 in one unit of 12 producing a
1:2 complex, 20122, featuring D4h-like symmetry.
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Figure 14. Selected regions of the H NMR spectra acquired during the titration of a 1 mM CDCl3
solution of tetradiazanaphthalene cavitand 12 with bis N-oxide 20. Equivalents added: a) 0 b) 0.25 c)
0.75 d) 1.5. Panel f) corresponds to the same regions of the 1H NMR spectrum of 20. Primed letters and
numbers indicate protons in the 2012 complex and doubly primed for the 20122 complex.
In the presence of 0.5 equivalents of bis-N-oxide 20 we observed the emergence of a new set of
proton signals for 12 that do not correspond with those assigned to the free host or the 1:2 complex.
These signals must correspond to a new inclusion complex in which the pyrrole NHs resonate at  =
9.40 ppm. Upon further addition of 20, the intensity of the signals of the latter complex grew at the
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expenses of those of 20122 (Figure 14c,d). Interestingly, the protons of bound 20 in the new complex
resonate as four separate signals. Two of them appear at chemical shift values that are in line with those
observed for the included N-oxide in the 20122 complex. The other two, however, are just slightly
upfield shifted with respect to the protons in free 20. The relative integrals for these four signals are
1:1:1:1. When 0.75 equivalents of 20 are added the signals for the protons of free N-oxide start to arise.
In the presence of 1.5 equivalents of 20 the 2012 complex is the major species in solution. Taken
together, these observations indicate that the incremental addition of 20 induces the destruction of the
20122 complex and induces the formation of a new 1:1 complex, 2012. The observation of four
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proton signals for the included N-oxide in the 2012 complex is consistent with its lower symmetry.
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The determination of accurate values for the stability constants of the 1:1 complex from the H NMR
titration data is hampered by the large value of the microscopic binding constants (Km > 104 M-1)
assigned to the interaction of the pyridine N-oxide group with the cavitand 12. Nevertheless, after all the
free cavitand is consumed, it is possible to determine an average value for the equilibrium constant of
the destruction process Kd (equation 1) by integrating the signals assigned to the 2012 and 20122
complexes in the presence of different amounts of 20.
Scheme 4. Equilibria involved in the destruction of the 20122 complex (1) and in the formation of the
2012 (2) and 20122 (3) complexes. The relationship of the equilibrium constants with Km
(microscopic constant),  (the cooperativity factor), and statistical correction factors are shown.
Using the relationship that exists between Kd and the cooperativity factor () of the N-oxide 20
(Equation 3 in Scheme 4) we determined  to be 0.09. A value of  < 1 for 20 indicates the existence of
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a negative cooperativity in the assembly process of the 20122 complex. In other words, the second
binding event (3) leading to the formation of the 20122 complex from the preformed 2012 is less
favorable than the statistical estimate derived from the microscopic constant value of the first binding
event. Most likely, the binding of the second unit of 12 is disfavored by a combination of electrostatic
and steric effects. On one hand, the upfield shift observed for the protons of the diazanaphthalene flaps
in the 20122 complex suggest their involvement in - stacking interactions between adjacent units of
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12. On the other hand, the reduced downfield shift experienced by the pyrrole NHs in the 20122
complex compared to those in 2012 points to the formation of larger hydrogen bonds in the latter. This
can be either a consequence of repulsive interactions between the two units of 12 in the 20122
complex or to a modification of negative density of the free oxygen atom of the N-oxide in the 2012
complex. The existence of a negative cooperativity in the assembly of the 20122 complex is also
supported by the observation of a solution mixture of free 12, 1:1 and 1:2 complexes when working
under strict stoichiometric control (ratio of [20]/[12] = 0.5).
CONCLUSIONS
We report here the synthesis of a novel family of deep-cavitands based on a calix[4]pyrrole scaffold.
The cavitands have been characterized both in the solid state and in solution. Whereas resorcinarene-
based cavitands easily switch between the vase and the kite conformations with temperature, the
calix[4]pyrrole-derived cavitands are conformationally more rigid than their resorcinarene analogs. In
the range of conditions studied they are locked in a C4 kite form as a racemic mixture of two
cycloenantiomers. Molecular modeling studies are in agreement with the experimental observations.
The energy barrier calculated for the interconversion between cycloenantiomers is very low (0.9
kcal/mol) explaining the fast interconversion observed in solution. Pyridine N-oxide derivatives are
included by the deep cavitands yielding 1:1 caviplexes that are thermodynamically and kinetically
highly stable. The putative conformational switch between the kite and vase forms of the calix[4]pyrrole
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cavitands was studied using variable temperatures H NMR experiments and by including N-oxide
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