A sleeping host awoken by its guest: Recognition and sensing of imidazole-containing molecules based on double Cu2+ translocation inside a polyaza macrocycle

Article abstract of DOI:10.1002/anie.200460568

pH-driven double Cu²⁺ ion translocation occurs inside an heteroditopic macrocycle. The movement opens and closes the system allowing or preventing it to function as receptor for bidentate anions. When imidazole is added to a solution buffered at an appropriate pH value, opening of the system and substrate binding can be induced by the substrate itself, which results in a sharp color change (see picture).

Full text of DOI:10.1002/anie.200460568

Angewandte
Chemie
different use of this kind of molecule can be envisaged, which
relies on one of their typical properties: when a molecular
machine moves as a consequence of an external stimulus, an
overall rearrangement of the molecular system takes place
and its shape changes dramatically. According to the well-
established lock-and-key concept, the shape of a molecule,
and in particular the shape of a cavity inside a large molecule,
is connected to its ability to act as a selective receptor, that is,
its ability to recognize a complementary substrate. Provided
that only one of the possible shapes assumed by a given
molecular machine presents a cavity in which a chosen
substrate can selectively fit, molecular machines could
advantageously enter the field of molecular recognition by
acting as receptors with a useful implemented function: to
recognize and bind a given substrate only when the proper
external stimulus is applied. Few examples have been
published that consist of relatively simple photoisomerizable
molecules, in which the stimulus required to make the system
change towards the correct shape and recognize a substrate is
provided by light.[3]
Herein we report the first example of a new kind of
molecular machine capable of molecular recognition. The
stimulus required by the system for changing towards the
correct shape for selective binding can be provided by the
substrate itself. Accordingly, the molecular machine descri-
bed in this paper behaves as a receptor, which merges the
advantages of the lock-and-key principle (selectivity) with
those of its adaptive behavior (activation in the presence of
substrates). Moreover, the drastic color change associated
with movement and recognition events turns this system in a
very efficient colorimetric sensor.
Molecular Devices
[4]
A Sleeping Host Awoken by Its Guest:
Recognition and Sensing of Imidazole-Containing
Molecules Based on Double Cu2+ Translocation
inside a Polyaza Macrocycle**
Macrocycle LH4 contains two couples of polydentate
compartments capable of binding Cu2+ ions, and comprises
two diamide–diamine (DADA) tetradentate and two pyri-
dine–diamine (PDA) tridentate binding sets, which share the
four secondary amino groups.
Potentiometric titrations carried out with standard base,
first on a solution containing LH4 and excess acid, then on a
solution containing LH4, two equivalnents of Cu2+ ions, and
excess acid[5] followed by nonlinear least-squares treatment of
the obtained electrode potential versus added base data,[6]
allowed us to calculate the protonation constants of LH4 and
the formation constants of the copper-containing species
present in solution in the pH range of 2–12 (see caption of
Figure 1).[7] From these data, a distribution diagram for the
LH4/2Cu2+ system was drawn (Figure 1a). Subsequently,
coupled pH-metric and spectrophotometric titrations were
carried out. Two distinct colors and types of band are
observed as a function of pH. A band centered at around
660 nm, responsible for the blue color of the solutions, starts
to form at pH 3 and predominates without dramatic changes
up to pH 9.5. At this pH, the intensity of the band at 660 nm
decreases relative to the band centered at 515 nm then
disappears leaving a purple–pink solution. The interpretation
is straightforward: if the molar absorbance at 660 and 515 nm
versus pH profiles are superimposed onto the distribution
diagram (Figure 1a, filled and empty triangles, respectively):
the relatively short wavelength band at 515 nm correlates with
the [Cu2(L)] neutral complex (form a in Scheme 1)[8] and
Luigi Fabbrizzi,* Francesco Foti, Stefano Patroni,
Piersandro Pallavicini,* and Angelo Taglietti*
Recently, a lot of attention has been focused on artificial
molecular machines,[1] in part for their potential to help
understand and mimic natural biological systems but mainly
from the perspective of developing molecular devices capable
of information processing and data storage.[2] However, a
[*] Prof. L. Fabbrizzi, Dr. F. Foti, Dr. S. Patroni, Dr. P. Pallavicini,
Dr. A. Taglietti
Dipartimento di Chimica Generale
Università di Pavia
viale Taramelli 12, 27100 Pavia (Italy)
Fax: (+39)0382-528-544
E-mail: luigi.fabbrizzi@unipv.it
psp@unipv.it
angelo.taglietti@unipv.it
[**] This work was financially supported by MIUR (Progetto “Dispositivi
Supramolecolari”) and by EU (RT Network Molecular Level Devices
and Machines, contract HPRN-CT-2000-00029).
Supporting information for this article is available on the WWW
under http://www.angewandte.org or from the author.
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DOI: 10.1002/anie.200460568
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nating, and the two Cu2+ ions
move to the two available sepa-
rated tridentate PDA units. This
form
is
the
form
complex
in
[Cu2(LH4)]4+,
b
Scheme 1, in which each Cu2+ is
tricoordinated by the ligand
(lmax = 660, deep blue), with the
other coordination positions of
the Cu2+ ion occupied by water
molecules.[10] On raising the pH
(6–9.5), two water molecules are deprotonated to give the
blue species [Cu2(LH4)(OH)]3+ and [Cu2(LH4)(OH)2]2+.
According to these data, when a 1:2 molar ratio of LH4/
Cu2+ is chosen, this system is capable of a pH-driven double
cation translocation; in a solution with 4 < pH < 5 the two
Cu2+ ions are coordinated by the two PDA moieties to give
[Cu2(LH4)]4+ (form b in Scheme 1, 95% of the relative
abundance, RA, in the distribution diagram), but if the pH is
raised above 10.5 the four amide protons are released and
each Cu2+ moves inside one of the two deprotonated DADA
moieties to give the neutral complex [Cu2(L)] (form a in
Scheme 1, 99% in the distribution diagram). The movement
is reversible and several translocation cycles have been
carried out in various experiments, without significant varia-
tions (except those due to dilution) in the spectra of the a and
b forms. To check the correctness of the interpretation of
spectral and potentiometric data, we carried out the same
measurement set on the reference compound L’H4, which
lacks the pyridine nitrogen atoms and should not give rise to
the same behavior. Infact, no blue was seen in the whole pH
range, and only a very weak shoulder in the spectra between
600–700 nm was observed at low pH, which completely
disappeared before pH 7. On the other hand, pink, pertinent
to the band centered at 510 nm, started to appear at a pH of
around 5 and the intensity of this band reached a plateaux at
pH 7 (see the Supporting Information), which did not change
with an increase in pH, thus showing that: 1) no blue dicopper
complex is formed; 2) a pink complex [Cu2(L’)] is completely
formed (> 95%) at pH 7, which is more than two pH units
below the pH at which the analogous complex with LH4 exists.
These results clearly show that the nitrogen atom of the
pyridine moiety is essential to the formation of the blue
complex, giving rise to two binding cavities in which the two
copper ions can be positioned in a wide pH range. These
cavities have such a good affinity for Cu2+ ions that they
prevent them from entering the DADA ligand sets before
pH 9.
Figure 1. a) Distribution diagram (relative abundance% (RA), of spe-
cies vs pH) for a system that contains Cu(CF3SO3)2 at 10À3 m and LH4
at 5”10À4 m. The calculated formation constants (expressed as logb)
for the species indicated (with uncertainties in parenthesis) are:
LH5+ =7.66(0.01), LH62+ =14.80(0.01), LH73+ =21.55(0.01),
LH84+ =27.73(0.01), [Cu2(LH4)]4+ =25.31(0.02),
[Cu2(LH4)(OH)]3+ =18.95(0.02), [Cu2(LH4)(OH)2]2+ =10.97(0.02),
[Cu2(L)]=À8.17(0.02). The full and empty triangles report the molar
absorptivity of the complex at 660 and 515 nm vs pH, respectively, rel-
ative to [Cu2(LH4)]4+ and to [Cu2(L)]; b) Distribution diagram (RA of
species vs pH) for a system that contains Cu(CF3SO3)2 at 10À3 m, and
LH4 and IMH at 5”10À4 m. The full and empty triangles report the
molar absorptivity of the complex at 650 and 515 vs pH profiles,
respectively, relative to [Cu2(LH4)(Im)]3+ and to [Cu2(L)]. The calculated
formation constants are the same as in (a), to which these equilibria
and constants have been added: LH4 + 2Cu2+ + ImH=[Cu2-
(LH4)(Im)]3+ + H+, logK=38.05(0.02); ImÀ + H+ =ImH,
[Cu2(L)] is a neutral, quite-rigid molecule in which each
Cu2+ centre is four-coordinated and square-planar. Moreover,
it is well-established that inside a similar donor set the Cu2+
ion is coordinatively saturated, that is, it does not interact with
any ligand added to the solution:[11] [Cu2(L)] can be consid-
ered the “closed” form of the system. On the other hand, in
the [Cu2(LH4)]4+ form, each Cu2+ is only three-coordinated by
the ligand and the water molecules that complete the
coordination sphere can be easily substituted by stronger
ligands. Thus, the system is in its “open” form and resembles a
whole category of dicopper complexes inside ditopic macro-
logK=10.82(0.02); ImÀ + 2H+ =ImH2+, logK=19.2(0.02).
arises from the typical d–d transition for a tetra coordinated
Cu2+ ion with square-planar geometry inside a bis-deproto-
nated DADA donor set.[9]
On changing pH to lower values (3–6), the deprotonated
amide groups are reprotonated, thus becoming noncoordi-
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pled pH–spectrophoto-
metric titrations evidence
the formation of the blue
complex
[Cu2(LH4)]4+
(lmax = 660 nm,
A =
the
204mÀ1 cmÀ1)
in
expected range of 3 <
pH < 4.5, while a rise in
the pH up to 6 induces the
shift of the maximum to
646 nm and an increase in
the intensity of this band
(A = 302mÀ1 cmÀ1), which
displays l and A values
consistent with a {Cu2+-
Scheme 1. pH dependent intramolecular dislocation of the Cu2+ ions.
(ImÀ)-Cu2+}
moiety.[16]
cyclic ligands,[12] which are able to bind in cascade several
bidentate ligands in a bridging fashion. Consequently, we
investigated the behavior of the system in the presence of
typical copper-coordinating bridging substrates.
A solution of the complex, buffered at pH 7 with HEPES
(4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) (0.05m)
was titrated with several anions such as dicarboxylates
(oxalate and malonate), phosphates (phosphate and pyro-
phosphate), azide and imidazolate. At the chosen pH the
[Cu2(LH4)(OH)]3+ species is predominant (> 70%). In all
cases, (except azide) the formation of 1:1 adduct was
observed. Following our interests on imidazole (ImH) and
histidine detection,[13] we decided to further investigate the
behavior of the complex in the presence of imidazole.
Potentiometric titrations were carried out with standard
base on solutions containing LH4, Cu2+, and ImH in a 1:2:1
molar ratio with excess acid. The formation constants of the
species in solution for this system have been calculated and
the relative distribution diagram drawn (Figure 1b, see
caption for additional equilibrium and the relative con-
stants).[14] In the range of 3 < pH < 4.5 [Cu2(LH4)]4+ is
predominant, as it is in the absence of ImH, whereas an
increase in pH above 4.5 result in the gradual formation of
[Cu2(LH4)(Im)]3+ (form c in Scheme 2), which predominates
in the pH range of 6.0 to 10.0. This latter species contains a
bridging imidazolate anion (ImÀ), which forms thanks to the
particularly stable[15,13c] {Cu2+-(ImÀ)-Cu2+} disposition. Cou-
This band persists in the pH range in which
[Cu2(LH4)(Im)]3+ is prevalent (Figure 1b, full triangles). On
further increasing the pH over 10.0, the blue disappears,
leaving the pink of the [Cu2(L)] species, which displays a band
with the same l and e values found when no ImH is present. It
has to be stressed that in the presence of imidazole no OH-
containing species are found in the 2–12 pH range and that
[Cu2(LH4)(Im)]3+ is so stable that it exists in a very large pH
range (< 90% in the 5.8–10.2 pH range), thus pushing the
formation of [Cu2(L)] to a pH value one unit higher than in
the absence of imidazole. Double cationic translocation also
occurs in the presence of imidazole, as the correct and
reversible spectral changes are observed on changing pH
from, for example, 11.0 to 7.0 and back again. Moreover, it has
to be pointed out that at pH 11.0 ImH is free in solution and
does not interact with [Cu2(L)] (form a, closed), while at
pH 7.0 the two Cu2+ ions translocate, the system is in it open
form and it is able to bind bridging anions such as ImÀ, thus
representing an example of controllable binding action
obtained inside a molecular machine after a chemical
(DpH) stimulus.
A comparison of the distribution diagrams in the absence
and presence of imidazole (Figure 1a and b) shows that in a
sharp pH range (10.0 < pH < 10.4) in the absence of imidazole
the [Cu2(L)] species is at 95%, whereas in the presence of
imidazole, it is at 10% and the [Cu2(LH4)(Im)]3+ species is
around 90%. Thus, we can expect that when a pink solution
buffered at pH 10.2 is
treated with one equiva-
lent of imidazole, trans-
location takes place with
imidazolate
according to the equili-
brium described in
binding,
Scheme 2. Although it is
somewhat slow, this
process
is
indeed
observed, as can be seen
by the series of spectra
obtained from a solution
of [Cu2(L)] buffered at
pH 10.2 after addition of
1 equivalent of ImH.
Scheme 2. The imidazole-induced translocation equilibrium at pH 10.2.
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Figure 2 shows the formation of the band typical of
[Cu2(LH4)(Im)]3+ with time. The absorbance versus time
profile corresponds to first-order kinetic, with k =
transport across apolar membranes (by using lipophilic
tails), histamine removal from biological fluids or its control-
lable release from nanodevices.
a
The Supporting Information for this article (distribution
diagram for ligand LH4 and pH–spectrophotometric titration
of the reference compound L’H4 in presence of two equiv-
alents of CuII ions) is available on the WWW under http:/
www.angewandte.org or from the author.
Received: May 6, 2004
Keywords: copper · molecular devices · molecular recognition ·
.
receptors · sensors
[1] V. Amendola, L. Fabbrizzi, C. Mangano, P. Pallavicini, Acc.
Chem. Res. 2001, 34, 488 – 493.
[2] V. Balzani, A. Credi, M. Venturi, Molecular Devices and
Machines—A Journey into the Nanoworld, Wiley-VCH, Wen-
heim, 2003.
[3] S. Shinkai in Molecular Switches (Ed.: B. L. Feringa), Wiley-
VCH, Wenheim, 2001, pp. 281 – 307.
Figure 2. The effects on absorption spectra of addition of one equiva-
lent of imidazole to a solution (7.5”10À4 m) of the complex buffered at
pH 10.2 with CAPS (3-cyclohexylamino-1-propanesulfonic acid)
(0.05m). Sample spectra showed are taken at t=0 (dotted line), 20,
50, 90, 150 min (solid line).
[4] Macrocyclic ligand LH4 and L’H4 were synthesized through a
2+2 condensation via Schiff base formation of 6-benzyl
À1,4,8,11-tetraazaundecane-5,7-dione and, respectively, 2,6-pyr-
idine dicarboxyaldehyde (for LH4) and isophtaladehyde (for
L’H4). A solution of 6-benzyl-1,4,8,11-tetraazaundecane-5,7-
dione (0.8 mmol) in acetonitrile (30 mL) was added dropwise
to a solution of the corrsesponding dialdheyde (0.8 mmol) in the
same solvent (60 mL) at room temperature under a nitrogen
atmosphere over a period of 30 min. The reaction mixture was
stirred overnight, then was reduced in situ with an excess of
NaBH4 (0.6 g). The solvent was removed from the reaction
mixture on a rotary evaporator and the solid residue was treated
with water (50 mL). Extraction with CH2Cl2, drying with MgSO4,
and removal of the solvent under vacuum gave the desired
product as a solid in 72% yield (LH4) and 73% (L’H4).
Elemental analysis calcd (%) for LH4 C42H54N10O4: C 66.12,
H 7.13, N 18.36; found C 66.48, H 7.06, N 18.15; for L’H4
C44H56N8O4: C 69.45, H 7.42, N 14.72; found C 69.32, H 7.16,
N 14.11. MS(ESI): m/z: 764 [LH4+H+], 762 [L’H4+H+].
[5] Measurements were carried out in a solution of NaClO4 (0.05m)
in 1:4 v/v water/ethanol. The water/ethanol mixture was chosen
instead of pure water for the low water solubility of the less
protonated or neutral forms of the ligand. All the presented
experiments and data were obtained in the aforementioned
solvent mixture. However, it has to be stressed that both the
[Cu2(L)] and the [Cu2(LH4)]4+ complexes are fairly soluble also
in pure water (up to 5 ” 10À3 m).
[6] P. Gans, A. Sabatini, A. Vacca, Talanta 1996, 43, 1739.
[7] The distribution diagram for protonated species (i.e., in the
absence of Cu2+) is available in the Supporting Information.
[8] The two metal/one ligand nature of the complexes responsible of
the 515 and 660 nm bands, that is, [Cu2(L)] and [Cu2(LH4)]4+, has
been checked also by means of spectrophotometric titrations: A
solution of KOH (0.01m; i.e., at pH ꢀ 12) in water/ethanol (1:4
v/v) that contained 5 ” 10À4 m LH4 were titrated with standard
aqueous Cu(CF3SO3)2; a linear increase of the 515 nm band and
a clear end point at 1:2 ligand/metal molar ratio was observed;
on the other hand, solutions buffered at pH 4.5 (acetic/acetate
buffer) that contained 5 ” 10À4 m LH4 were titrated with standard
aqueous Cu(CF3SO3)2; a linear increase of the 660 nm band with
a sharp end point at 1:2 ligand/metal molar ratio was observed.
Solid samples of [Cu2(L)] and [Cu2(LH4)(OH)](CF3SO3)3·5H2O,
which gave satisfactory CHN analysis, were obtained by slow
0.00037 sÀ1 (t = 45 min). According to this result, in this
system it is the substrate itself that makes the cations
translocate and causes the system to open, thus allowing
binding to take place. A high degree of selectivity is found:
when pH is fixed at 10.2 the system is in its closed form (form
À
a), and the addition of potentially bridging anions (N3
,
PO43À, P2O74À, C2O42À) give no spectral change even after the
addition of up to fivefold excess of anions. The same behavior
is observed when several equivalents of representative
aminoacids and other biologically relevant substrates are
added to the solution: no color change is observed upon
addition of several equivalents of glycine, arginine, proline,
glutamate, ADP, ATP. On the other hand, as expected, the
addition of histidine or histamine gives the same effect
already observed with imidazole. The same response (switch
from pink to blue) is observed when titrations with imidazole,
histidine, and histamine are repeated in the presence of a
fivefold excess of the potential interferents. Only histidine
and histamine are recognized and sensed colorimetrically.
The receptor is a sleeping host that is closed to all guests we
have tried so far, except imidazole-containing molecules,
which have the key (the imidazole fragment itself) to switch
the host cavity to the open, binding form. This recognition is
associated to a neat color change, thus providing a signal of
the selective inclusion. The peculiar, extreme case of selec-
tivity relies on the unique energy gain obtained from the
particularly stable {Cu2+-(ImÀ)-Cu2+} moiety, which more
than compensates the energy barrier necessary for the
movement of the molecular machine, that is, double Cu2+
translocation. The design and synthesis of devices based on
this approach are being performed in our laboratory. This
peculiar kind of selectivity can be exploited by simply
changing the groups appended between the two amide
functions for a broader range of applications such as
fluorimetric sensing (appending fluorogenic fragments),
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Chemie
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spectra (ESI) carried out for [Cu2(L)] gave the expected m/z
peak m/z: 887 [[Cu2(L)] + H+].
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imidazole have been recalculated, as the titration experiments
were carried out in 1:4 water/ethanol mixture and not in water.
The formation constants for the species that do not contain
imidazole have been maintained at a constant, with the same
values determined in the previous experiments.
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