Selective binding of l-glutamate derivative in aqueous solvents

Article abstract of DOI:10.1039/b803089h

A chiral bisguanidinium macrocycle binds N-Boc-l-glutamate in a 1: 1 stoichiometry with significant selectivity in competitive solvent (DMSO-H ₂O).

Full text of DOI:10.1039/b803089h

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PAPER
www.rsc.org/obc | Organic & Biomolecular Chemistry
Selective binding of L-glutamate derivative in aqueous solvents†
Sandra Bartoli, Tariq Mahmood, Abdul Malik, Sally Dixon and Jeremy D. Kilburn*
Received 22nd February 2008, Accepted 25th March 2008
First published as an Advance Article on the web 25th April 2008
DOI: 10.1039/b803089h
A chiral bisguanidinium macrocycle binds N-Boc-L-glutamate in a 1 : 1 stoichiometry with significant
selectivity in competitive solvent (DMSO–H2O).
crystal structure of acyclic pyridyl thiourea analogues of 2.10 In
Introduction
chloroform, macrocycle 1 does not bind carboxylates at all because
the energy required to reorganise the macrocycle into a suitable
binding conformation (i.e. to break the intramolecular hydrogen
bonds) is not compensated by binding interactions that would
thereby be established. In DMSO, solvation of the intramolecular
hydrogen bonds weakens the preference for a tightly wrapped
conformation unsuitable for binding a carboxylate guest, but
binding of the first carboxylate group may still require a significant
reorganisation of the molecule and consequent energetic penalty,
which does not then have to be paid when a second carboxylate is
bound.
In order to develop similarly selective receptors able to bind
glutamate in more competitive aqueous solvent systems, we have
now prepared bisguanidinium macrocycle 3 and investigated its
binding properties with glutamate and aspartate derivatives. In
50 : 50 DMSO–H2O solution, macrocycle 3 forms a strong
1 : 1 complex with N-Boc-L-glutamate, but 1:2 complexation is
preferred with N-Boc-D-glutamate or either enantiomer of N-Boc-
aspartate.
Recognition of polar molecules by synthetic receptors in aqueous
solvents is a formidable challenge because of the high degree of
solvation that has to be overcome. The development of synthetic
receptors for enantioselective binding of L-glutamate and L-
aspartate derivatives has been of particular interest because of
the critical role these amino acids play in the central nervous
system as excitatory transmitters. Several groups have successfully
developed receptors able to bind a range of dicarboxylates1–8
(including glutamate and aspartate) in protic solvents, typically
using polyamine macrocyclic receptors (which generally exist,
at physiological pH, as a mixture of several protonated forms,
all of which may contribute to binding),1,2 the metal complexes
derived from such polyamines,3 or appropriately functionalised
guanidinium or amidinium derivatives.4–6 However, examples
of enantioselective receptors for such dicarboxylates in protic
solvents remain rare.7
We recently described bisthiourea macrocycles 1 and 2 which
it was anticipated could serve as enantioselective receptors for di-
carboxylates, such as glutamate or aspartate, using two thiourea–
carboxylate interactions with additional hydrogen bonds between
the macrocyclic amides and the carboxylate groups (Fig. 1).
Macrocycles 1 and 2 proved to be highly selective for the 1 :
1 binding of N-Boc-L-glutamate in both acetonitrile and the
more competitive solvent dimethylsulfoxide (DMSO),9 with the
glutamate guest accommodated within the macrocyclic cavity
[Fig. 1, i)]. Binding of N-Boc-D-glutamate on the other hand was
predominantly 1 : 2 (host–guest) stoichiometry in DMSO [Fig. 1,
ii)], with a weak 1 : 1 (host H + guest G) binding constant and a
significantly stronger stepwise 1 : 2 (HG + G) binding constant.
The apparent cooperativity with stronger binding of the second
anionic guest than the first, a positive allosteric effect, is at first
sight surprising but the two anionic guests may be a significant
distance apart and therefore experience little electrostatic repul-
sion when both are bound. Furthermore, macrocycle 1 adopts
a wrapped conformation in chloroform that is stabilised by a
number of intramolecular hydrogen bonds, particularly from an
amide carbonyl to the thiourea NHs and adjacent amide NH, a
hydrogen bonding motif previously described by the dimeric pair
Results
Synthesis
Direct conversion of bisthiourea macrocycle 1 to bisguanidinium
macrocycle 3, via treatment of an intermediate S-alkyl bisthiouro-
nium macrocycle with ammonia, was not successful. The desired
macrocycle was instead assembled via carbodiimide mediated
condensation of a biscarbamoyl thiourea with a bisamine;11 via
an efficient synthetic strategy involving use of the bisamine as
both precursor and condensation partner to the bis-carbamoyl
thiourea (Scheme 1).
Efficient coupling of 3-cyanobenzoic acid with (1S,2S)-
diphenylethylene diamine, followed by nitrile reduction, also
in good yield, gave bisamine 7. Conversion to a bis-carbomyl
thiourea 8, upon treatment of
7 with benzyloxycarbonyl
isothiocyanate,12 was straightforward. Coupling of bisthiourea
8 with bisamine 7 gave the protected bisguanidine macrocycle
9, in a modest optimised yield of 29% critically dependent
upon the use of 2 equivalents of DMAP in this step; the first
demonstration of carbamoyl thiourea–amine condensation utility
in macrocyclisation. Removal of the Cbz protecting groups was
readily achieved by hydrogenolysis to give the desired macrocycle
3, as its bis(hexafluorophosphate) salt.
School of Chemistry, University of Southampton, Southampton, UK SO17
1BJ. E-mail: jdk1@soton.ac.uk; Fax: +44 (0)2380 596805; Tel: +44
(0)2380 593596
† Electronic supplementary information (ESI) available: NMR and ITC
titration data for all binding studies. See DOI: 10.1039/b803089h
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Fig. 1 i) 1 : 1 complex of bisthiourea macrocycle 1 with N-Boc-L-glutamate (Ka = 2.7 × 104 M−1, Ka = 3.9 × 102 M−1 in DMSO); ii) 1 : 2 complex
1:1
1:2
1:1
1:2
of bisthiourea macrocycle 1 with N-Boc-D-glutamate (Ka = 2.6 × 102 M−1, Ka = 1.9 × 104 M−1 in DMSO).
Scheme 1 Reagents and conditions: i) (PhO)2P(O)Cl, NEt3, CH2Cl2, then (1S,2S)-diphenylethylene diamine, K2CO3, H2O, 90%; ii) H2, Pd/C,
TFA–DMF–MeOH, then NEt3, 83%; iii) PhCH2O2CNCS, DMF–CH2Cl2, 73%; iv) 7, EDC, CH2Cl2–DMF, DMAP, 29%; v) H2, Pd/C, MeOH–DMF,
then HPF6, CH2Cl2–MeOH, 77%.
1:2
Binding characterisation
G) (Ka ∼ 2 × 103 M−1). A similar downfield shift of the amide
NH resonance was observed on addition of 5.6 equiv. N-Boc-D-
glutamate, (also as the bis(tetrabutylammonium) dicarboxylate
salt, Ddmax ∼ 1.2 ppm), to macrocycle 3 and, once again, a
sigmoidal data plot was obtained which could not be fitted
assuming 1 : 1 binding and indicates two binding stoichiometries.15
Fitting using a 1 : 2 (host–guest) binding isotherm indicated that
binding of N-Boc-D-glutamate is dominated by 1 : 2 binding
Macrocycle 3 gave a well-resolved 1H NMR spectrum in d6-DMSO
and in 5% CD3OD–CDCl3, consistent with the expected D2
symmetry. Binding studies with macrocycle 3 were first conducted
1
by H NMR titration in d6-DMSO (Fig. 2).†13,14 Addition of N-
Boc-L-glutamate (as the bis(tetrabutylammonium) dicarboxylate
salt) led to downfield shifts of the guanidinium NH resonance
(Ddmax > 1.3 ppm) and amide NH resonance (Ddmax > 1.0 ppm),
consistent with the formation of hydrogen bonds. The titration
curve did not reach saturation (6.4 equiv. added guest) and data
could not be fitted to a simple 1 : 1 (host–guest) binding isotherm,
but to a 1 : 2 (host–guest) binding isotherm in which binding is
1:1
1:2
(Ka ∼ 103 M−1, Ka > 104 M−1). Hence macrocycle 3 shows
similar binding selectivity, and preference for 1 : 1 binding with
N-Boc-L-glutamate, in DMSO, as the thiourea analogue 1.
Investigation of the binding properties of macrocycle 3 in a
more competitive solvent system, 50% H2O–DMSO in which
the macrocycle gives a poorly resolved 1H NMR spectrum,16
was made using isothermal calorimetry (ITC).17–19 Titration of 3
dominated by a large 1 : 1 association (H + G) (Ka > 104 M−1)
1:1
with a small contribution from sequential 1 : 2 binding (HG +
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binding constant, Ka 1.4 × 104 M−1 (DG1:1 = −24.0 kJ mol−1),
and a smaller 1 : 1 binding component, Ka1:1 2.9 × 103 M−1 (Table 1,
entry 2). ITC titration of macrocycle 3 with the two enantiomers
of N-Boc-aspartate was also carried out, and in both cases gave
1:2
predominantly 1 : 2 binding (Ka > 104 M−1) (Table 1, entries
1:2
3 and 4), and a titration curve very similar to that observed for
N-Boc-D-glutamate. In each of these three cases the dominant 1 : 2
binding was largely driven by the enthalpic contribution, whereas
the smaller 1 : 1 binding was enthalpically very unfavourable, and
was driven by a large entropic contribution.
Overall the data suggest that there is a precise and tight fit
within the macrocyclic cavity for the preferred guest, N-Boc-L-
glutamate, with strong hydrogen bonding interactions to both
carboxylate groups of the glutamate (Fig. 4) and enthalpically
favourable 1 : 1 binding. Sequential binding, to form the 1 : 2
complex, presumably involves displacement of one carboxylate
functionality of the internally bound glutamate and binding to
the carboxylate of a second glutamate, with no net increase in
hydrogen bonding, but leading to a much more flexible complex.
Formation of the 1 : 2 complex from the 1 : 1 complex might
therefore be expected to be enthalpically neutral but entropically
favourable, as observed experimentally. Tight binding of N-
Boc-D-glutamate (and similarly N-Boc-L- and D-aspartate) is
presumably not possible within the cavity of macrocycle 3 and
instead initial 1 : 1 binding may only involve a single carboxylate–
thiourea interaction on the outside of the macrocycle. The data
indicate that this process is both highly endothermic and en-
tropically favourable, presumably because this first binding event
involves substantial reorganisation of macrocycle 3, breaking of
a number of intramolecular hydrogen bonds and desolvation of
tightly bound solvent molecules (in contrast binding of N-Boc-
L-glutamate described above leads to a much more rigid 1 :
1 complex with significantly greater hydrogen bonding leading
to an enthalpically much more favourable process). Sequential
binding of a second molecule of N-Boc-D-glutamate (and sim-
ilarly N-Boc-L- and D-aspartate) then involves an enthalpically
favourable formation of a second carboxylate–thiourea interaction
on the outside of the macrocycle and without any substantial
further reorganisation of the macrocycle. While it is important
not to over-interpret the thermodynamic data obtained from
the ITC experiments or rule out other binding stoichiometries,
this explanation satisfactorily rationalises the experimental data
and is entirely consistent with the binding properties observed
with the original thiourea macrocycle when binding the two
enantiomers of N-Boc-glutamate in DMSO. Unfortunately we
were unable to obtain any detailed structural information about
the complexation from NMR studies due to the poor solubility and
Fig. 2 NMR binding titration of macrocycle 3 with N-Boc-L-glutamate
(ꢀ), [3]/mM = 1.75, and N-Boc-D-glutamate (᭺), [3]/mM = 1.30, in
d6-DMSO.
with the bis(tetrabutylammonium) dicarboxylate salt of N-Boc-L-
glutamate gave a titration curve (Fig. 3)† which could be fitted
using a two-site binding model, giving a large 1 : 1 binding
constant, Ka 3.8 × 104 M−1 (DG1:1 = −26.1 kJmol−1), and a
1:1
1:2
smaller sequential 1 : 2 (host–guest) binding constant, Ka = 5.3 ×
103 M−1. The data indicate that the predominant 1 : 1 binding is
both enthalpically and, to a lesser extent, entropically favourable
(Table 1, entry 1), whereas the subsequent binding of the second
equivalent is largely driven by favourable entropy.
Fig. 3 ITC data for macrocycle 3 with N-Boc-L-glutamate (ꢀ) and
N-Boc-D-glutamate (᭺) in 50 : 50 DMSO–H2O.
Titration of 3 with the bis(tetrabutylammonium) dicarboxylate
salt of N-Boc-D-glutamate indicated a large 1 : 2 (host–guest)
Table 1 ITC binding data for macrocycle 3 with biscarboxylate salts in 50 : 50 DMSO–H2Oa
DG1:1/kJ
DH1:1/kJ
TDS1:1/kJ
DG1:2/kJ
DH1:2/kJ
TDS1:2/kJ
Entry
Guest
Ka1:1/M−1
mol−1
mol−1
mol−1
Ka1:2/M−1
mol−1
mol−1
mol−1
1
2
3
4
N-Boc-L-Glu
N-Boc-D-Glu
N-Boc-L-Asp
N-Boc-D-Asp
3.8 × 104
2.9 × 103
1.1 × 103
2.5 × 103
−26.1
−19.8
−17.4
−19.4
−18.1
39.3
61.7
8.0
59.1
79.1
66.3
5.3 × 103
1.4 × 104
1.1 × 104
1.4 × 104
−21.6
−24.0
−23.3
−23.9
−2.2
−22.5
−19.2
−19.5
19.4
1.5
4.1
4.4
46.9
a Association constants are reported to two significant figures with estimated errors of 20%. Ka1:2 refers to the binding constant for stepwise association
1:1
1:2
of the 1 : 1 complex with a second equivalent of guest, the overall binding constant is therefore the product of reported Ka and Ka
.
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Fig. 4 Proposed mode of sequential binding of i) N-Boc-L-glutamate and ii) N-Boc-D-glutamate by macrocycle 3.
poorly resolved spectra of the complexes obtained in 50% DMSO–
H2O.
as guest in 50% H2O–DMSO (using a tris buffer) but in this case
indicated very weak binding (Ka < 102 M−1). Studies are under
1:1
way to synthesise macrocycles which are both more water soluble
and incorporate additional functionality to stabilise interactions
with the ammonium group of unprotected glutamate.
Conclusions
Bisguanidinium macrocycle 3 is able to bind strongly with
biscarboxylates in a highly competitive aqueous solvent system,
demonstrating an important advance over the class of bisthiourea
receptors we have reported previously. Macrocycle 3 is also
able to discriminate for the 1 : 1 binding of N-Boc-L-glutamate
against both its enantiomer N-Boc-D-glutamate and the smaller
aspartate derivatives. ITC binding studies were also carried out
with macrocycle 3 and using unprotected, zwitterionic L-glutamate
Experimental
General techniques
Reactions were carried out in solvents of commercial grade and,
where necessary, distilled prior to use. Reactions requiring a
dry atmosphere were conducted in oven dried glassware under
nitrogen. CH2Cl2 was distilled from calcium hydride, as was
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petroleum ether where the fraction boiling between 40 and
60 ◦C was used. Where degassed solvents were used, a stream of
nitrogen was passed through them immediately prior to use, unless
otherwise stated. TLC was conducted on foil backed sheets coated
with silica gel (0.25 mm) and containing fluorescent indicator
UV254. Column chromatography was performed on Sorbsil C60,
40–60 mesh silica.
1H and 13C NMR spectra were recorded on Bruker AV300,
AM300 or DPX400 spectrometers. 1H chemical shifts are re-
ported as values in ppm referenced to residual solvent. The
following abbreviations are used to denote multiplicity and may
be compounded: s = singlet, d = doublet, t = triplet, q =
quartet, fs = fine splitting. Coupling constants, J, are measured
in Hertz (Hz). 13C spectra were proton decoupled and referenced
to solvent. The number of adjacent protons was determined by
DEPT experiments. Low resolution mass spectra were recorded
on a Waters ZMD mass spectrometer, single quadrupole, 2700
autosampler in methanol or acetonitrile. Accurate mass spectra
were recorded on a VG analytical 70–250-SE double focussing
mass spectrometer. Melting points were determined in open
capillary tubes using a Gallenkamp Electrothermal melting point
apparatus and are uncorrected. Microanalyses were performed by
MEDAC Ltd., Surrey.
Synthetic procedures
(1S,2S)-(−)-1,2-Diphenyl-1,2-ethylenediamine-L-tartrate
was
prepared according to the method of Corey and co-workers.20
Benzyloxycarbonyl isothiocyanate was prepared according to
the method of Groziak and Townsend.12 Preparation of N,Nꢀ-
(1S,2S)-diphenyl-ethyl-1,2-diylbis-3-aminomethylbenzamide (7)
has been reported previously.9b
N,Nꢀ-(1S,2S)-Diphenyl-ethyl-1,2-diylbis-[3-(Nꢀ-benzyl-Nꢀꢀ-
carbobenzyloxy-thioureido)-benzamide 8
N,Nꢀ -(1S,2S)-Diphenyl-ethyl-1,2-diylbis-3-aminomethylbenza-
mide (7) (250 mg, 0.52 mmol) was taken into a mixture of
CH2Cl2 (10 mL) and dimethylformamide (DMF) (1 mL). Ben-
zyloxycarbonyl isothiocyanate (252 mg, 1.3 mmol) was added
and the mixture stirred for 8 h, during which time formation of
a precipitate was observed, before concentration in vacuo. The
residue was suspended in CH2Cl2 (5 mL) and the solid collected
by filtration and washed with CH2Cl2 (2 × 5 mL) and Et2O (2 ×
5 mL). Drying in vacuo gave the title compound as a white solid
(330 mg, 73%): Rf = 0.25 (3% MeOH in DCM); mp = 155–156 ◦C;
1H NMR (400 MHz, d6-DMSO): d = 4.30 (4H, brs, CH2NH),
5.17 (4H, s, CH2O), 5.65 (2H, d, J = 6.5 Hz, CH), 7.10–7.50
(26H, m, ArH), 7.63 (2H, s, ArH), 9.04 (2H, d, J = 7.0 Hz,
NHCH), 10.24 (2H, br s, NHCH2), 11.23 (2H, s, NHCbz) ppm;
13C NMR (100 MHz, d6-DMSO): d = 47.8 (CH2), 57.2 (CH), 67.0
(CH2), 125.9 (CH), 126.7 (CH), 127.0 (CH), 127.4 (CH), 128.0
(CH), 128.4 (CH), 128.5 (CH), 128.6 (CH), 130.4 (CH), 135.2
(C), 135.7 (C), 138.2 (C), 140.8 (C), 153.4 (C), 166.6 (C), 180.1
(C) ppm; MS (ES+): m/z: 865 ([M + H]+, 90%) 887 ([M + Na]+,
85%).
1H NMR titration experiments13
1H NMR titration experiments were conducted on a Bru¨ker AM
300 spectrometer at 298 K. A sample of host was dissolved in the
deuterated solvent. A portion of this solution was used as the host
NMR sample and the remainder used to dissolve a sample of the
guest, so that the concentration of the host remained constant
throughout the titration. Guest stock solutions were typically
prepared such that 10 lL of that solution contained 0.1 equivalents
of guest with respect to host, unless otherwise stated. Successive
aliquots of the guest solution were added to the host NMR sample
and the 1H NMR spectrum recorded after each addition. Changes
in chemical shift of guanidinium and amide NH host signals, as
a function of guest concentration, were analysed using NMRTit
HG software, assuming a 1 : 1 or 1 : 2 binding stoichiometry.
Macrocycle 9
N,Nꢀ-(1S,2S)-Diphenyl-ethyl-1,2-diylbis-[3-(Nꢀ-benzyl-Nꢀꢀ-carbo-
benzyloxy-thioureido)-benzamide 8 (60 mg, 0.069 mmol) was
taken into CH2Cl2 (10 mL) and DMF (10 mL). 1-Ethyl-3-(3ꢀ-
dimethylaminopropyl)carbodiimide (EDC) (53 mg, 0.276 mmol)
and 4-dimethylaminopyridine (DMAP) (15 mg, 0.14 mmol) were
then added. To this vigorously stirred solution, a solution of N,Nꢀ-
(1S,2S)-diphenyl-ethyl-1,2-diylbis-3-aminomethylbenzamide (7)
(34 mg, 0.071 mmol) in CH2Cl2–DMF (15 mL of a 2 : 1 mixture)
was added and the resulting mixture stirred for 36 h before
concentration in vacuo. Purification by column chromatography.
(SiO2 eluted with CH2Cl2 → EtOAc–CH2Cl2 (5 : 1) then SiO2
eluted with 2% MeOH in CH2Cl2) gave the title compound as a
white solid (28 mg, 29%); mp 171–172 C; H NMR (400 MHz,
CDCl3): d = 4.24 (8H, brs, CH2NH), 5.09 (4H, s, CH2O), 5.64
(4H, br s, CHPh), 7.04 (4H, d, J = 7.0 Hz, NHCO), 7.12 (4H, br s,
NHCN), 7.20–7.40 (46H, m, ArH) ppm; 13C NMR (100 MHz,
5% CD3OD–CDCl3): d = 44.5 (CH2), 59.3 (CH), 66.8 (CH2),
126.0 (CH), 127.6 (CH), 127.7 (CH), 127.8 (CH), 127.9 (CH),
128.3 (CH), 128.5 (CH), 128.7 (CH), 130.4 (C), 134.8 (C), 137.1
(C), 138.7 (C), 159.9 (C), 163.9 (C), 168.5 (C) ppm; MS (ES+):
m/z: 639 ([M + 2H]2+, 100%) 1276 ([M + H]+, 30%); Found: C,
71.70; H, 5.54; N, 10.57. Calc. for C78H70N10O8·2H2O: C, 71.43;
H, 5.69; N, 10.68%.
Isothermal calorimetry titration experiments19
All experiments were performed using an isothermal titration
calorimeter from Microcal Inc. (Northampton, MA). In a typical
experiment, a 0.6 mM receptor solution is added to the calorimet-
ric cell. A 40 mM solution of guest is introduced in 65 injections:
30 injections of 2.5 lL followed by 35 injections of 5.0 lL to
a total of 250 lL added guest. The solution is continuously
stirred to ensure rapid mixing and kept at 30 ◦C, through the
combination of an external cooling bath and an internal heater.
Dilution effects are determined by performing a blank experiment
in which the same guest solution is added to pure solvent and
the heat signature subtracted from the raw titration to produce
the final binding curve. Binding parameters are determined by
applying either one-site or two-sites models, using the Origin
software provided. These methods rely on standard nonlinear
least-squares regression (Levenberg–Marquard method) to fit the
curves, taking into account the change in volume that occurs
during the calorimetric titration.
◦
1
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Macrocycle 3
J. de Mendoza, A. D. Hamilton and E. Giralt, Chem. Commun., 2000,
1399–1400.
6 (a) L. Sebo, B. Schweizer and F. Diederich, Helv. Chim. Acta, 2000, 83,
80–92; (b) L. Sebo, F. Diederich and V. Gramlich, Helv. Chim. Acta,
2000, 83, 93–113.
Macrocycle 9 (17 mg, 0.013 mmol) was taken into MeOH–DMF
(2 mL of a 1 : 1 mixture). 10% Pd/C (3 mg) was added and
the mixture stirred under H2 (1 atm) for 14 h. The mixture was
filtered through CeliteTM and the filtrate concentrated in vacuo. The
residue obtained was then taken into CH2Cl2–MeOH (6 mL of a 5 :
1 mixture) and HPF6 (10 lL of a 60% v/v aqueous solution, 0.04
mmol) was added. The solution was then concentrated in vacuo
and the residue suspended in H2O, filtration and drying in vacuo
gave◦the title compound as a white solid (13 mg, 77%); mp 183–
7 (a) I. Alfonso, F. Rebolledo and V. Gotor, Chem.–Eur. J., 2000, 6, 3331–
3338; (b) I. Alfonso, B. Dietrich, F. Rebolledo, V. Gotor and J.-M.
´
Lehn, Helv. Chim. Acta, 2001, 84, 280–295; (c) A. Gonza´lez-Alvarez,
I. Alfonso, P. D´ıaz, E. Garc´ıa-Espana and V. Gotor, Chem. Commun.,
2006, 1227–1229.
8 (a) Recent examples of related receptors for tricarboxylates in water:
S. L. Tobey and E. V. Anslyn, J. Am. Chem. Soc., 2003, 125, 10963–
10970; (b) S. L. Wiskur, J. L. Lavigne, A. Metzger, S. L. Tobey, V. Lynch
and E. V. Anslyn, Chem.–Eur. J., 2004, 10, 3792–3804; (c) C. Schmuck
and M. Schwegmann, J. Am. Chem. Soc., 2005, 127, 3373–3379; (d) A.
Frontera, J. Morey, A. Oliver, M. N. Pina, D. Quinonero, A. Costa,
P. Ballester, P. M. Deya and E. V. Anslyn, J. Org. Chem., 2006, 71,
7185–7195.
9 (a) S. Rossi, G. M. Kyne, D. L. Turner, N. J. Wells and J. D. Kilburn,
Angew. Chem., Int. Ed., 2002, 41, 4233–4236 [NB: macrocycle 2 is
incorrectly represented as the (1R,2R)-(+)-1,2-diphenyl enantiomer of
the 1S,2S-macrocycle for which binding studies are reported, in both
Schemes 1 and 2 of reference 9a]; (b) A. Ragusa, S. Rossi, J. M. Hayes,
M. Stein and J. D. Kilburn, Chem.–Eur. J., 2005, 11, 5674–5688.
10 G. M. Kyne, M. E. Light, M. B. Hursthouse, J. de Mendoza and J. D.
Kilburn, J. Chem. Soc., Perkin Trans. 1, 2001, 1258–1263.
11 (a) B. R. Linton, A. J. Carr, B. P. Orner and A. D. Hamilton, J. Org.
Chem., 2000, 65, 1566–1568; (b) R. J. Fitzmaurice, F. Gaggini, N.
Srinivasan and J. D. Kilburn, Org. Biomol. Chem., 2007, 5, 1706–1714.
12 Preparation of CbzNCS was adapted from: M. P. Groziak and L. B.
Townsend, J. Org. Chem., 1986, 51, 1277–1282.
1
185 C; H NMR (300 MHz, d6-DMSO): d = 4.43 (8H, d, J =
4.0 Hz, CH2NH), 5.66 (4H, br s, CHPh), 7.00–7.80 (40H, m,
+
ArH + NH2 ), 8.03 (4H, br s, NHCH2), 9.16 (4H, br s, NHCO)
ppm; 13C NMR (100 MHz, CDCl3–CD3OD): d = 47.5 (CH2), 61.8
(CH), 126.2 (CH), 128.9 (CH), 129.9 (CH), 130.4 (CH), 130.6
(CH), 131.4 (CH), 131.9 (CH), 133.3 (C), 137.5 (C), 138.9 (C),
141.5 (C), 171.5 (C) ppm; MS (ES+): m/z: 504 ([M + 2H]2+, 93%)
526 ([M + 2Na]2+, 100%) 1008 ([M + H]+, 30%); Found: C, 55.55;
H, 4.90; N, 10.77. Calc. for C62H60F12N10O4P2·2H2O: C, 55.77; H,
4.83; N, 10.89%.
References and notes
1 Review: E. Garc´ıa-Espana, P. D´ıaz, J. M. Llinares and A. Bianchi,
Coord. Chem. Rev., 2006, 250, 2952–2986.
13 NMR titration data was analysed using NMRTit HG software: A. P.
Bisson, C. A. Hunter, J. C. Morales and K. Young, Chem.–Eur. J.,
1998, 4, 845–851. All binding data from NMR titration experiments
are provided in the ESI†.
2 (a) Original work on polyamine macrocyclic receptors for carboxylates:
B. Dietrich, M. W. Hosseini, J.-M. Lehn and R. B. Sessions, J. Am.
Chem. Soc., 1981, 103, 1282–1283; (b) E. Kimura, A. Sakonaka, T.
Yatsunami and M. Kodama, J. Am. Chem. Soc., 1981, 103, 3041–3045;
(c) E. Kimura, M. Kodama and T. Yatsunami, J. Am. Chem. Soc., 1982,
104, 3182–3187; (d) M. W. Hosseini and J.-M. Lehn, J. Am. Chem. Soc.,
1982, 104, 3525–3527; (e) M. W. Hosseini and J.-M. Lehn, Helv. Chim.
Acta, 1986, 69, 587–603.
3 (a) Macrocyclic metal complexes: A. E. Martell and R. J. Motekaitis,
J. Am. Chem. Soc., 1988, 110, 8059–8064; (b) C. Bazzicalupi, A.
Bencini, A. Bianchi, V. Fusi, E. Garc´ıa-Espana, C. Giorgi, J. M.
Llinares, J. A. Ramirez and B. Valtancoli, Inorg. Chem., 1999, 38, 620–
621; (c) M. Boiocchi, M. Bonizzoni, L. Fabbrizzi, G. Piovani and A.
Taglietti, Angew. Chem., Int. Ed., 2004, 43, 3847–3852; (d) B. Verdejo,
J. Aguilar, A. Dome´nech, C. Miranda, P. Navarro, H. R. Jime´nez, C.
Soriano and E. Garc´ıa-Espana, Chem. Commun., 2005, 3086–3088;
(e) F. Li, R. Delgado and V. Fe´lix, Eur. J. Inorg. Chem., 2005, 4550–
4561.
14 The binding constants obtained for N-Boc-L/D-glutamate using NMR
titrations could not be determined with great accuracy. The dominant
binding constant in each case was at the upper limit of detectability
using this method (L. Fielding, Tetrahedron, 2000, 56, 6151–6170) and
the value obtained for the smaller binding constant in each case was
sensitive to the values used at the start of the iterative calculation and
could therefore not be determined to better than 1 significant figure.
Job plots give ambiguous results with broad maxima at stoichiometries
of ∼1–1.5.
15 C. S. Wilcox, J. C. Adrian Jr., T. H. Webb and F. J. Zawacki, J. Am.
Chem. Soc., 1992, 114, 10189–10197.
1
16 Poor resolution of the H NMR spectrum of 3 in 50% H2O–DMSO
results from sparing solubility of the macrocycle. In fact, neither the
amide nor guanidinium NH resonance were visible in the spectrum—
presumably because of fast exchange with the water.
4 (a) Reviews: P. Blondeau, M. Segura, R. Pe´rez-Ferna´ndez and J. de
Mendoza, Chem. Soc. Rev., 2007, 36, 198–210; (b) K. A. Schug and W.
Lindner, Chem. Rev., 2005, 105, 67–113; (c) M. D. Best, S. L. Tobey
and E. V. Anslyn, Coord. Chem. Rev., 2003, 240, 3–15.
17 A. Cooper, Curr. Opin. Chem. Biol., 1999, 3, 557–563 and references
therein.
18 (a) For the specific application of isothermal calorimetry to carboxylate
recognition, see: ref. 5d and 6a; (b) B. Linton and A. D. Hamilton,
Tetrahedron, 1999, 55, 6027–6038; (c) M. Berger and F. P. Schmidtchen,
J. Am. Chem. Soc., 1999, 121, 9986–9993.
19 The ITC data were obtained using a MicroCal VP-ITC isothermal
calorimeter at apparent pH 6.8 and are corrected for the heat generated
from dilution of the host. Data were analysed by non-linear least square
fitting using the Origin software supplied by MicroCal: T. Wiseman,
S. Williston, J. F. Brandts and L.-N. Lin, Anal. Biochem., 1989, 179,
131–137. All binding data from ITC experiments are provided in the
ESI†.
5 (a) B. Dietrich, T. M. Fyles, J.-M. Lehn, L. G. Pease and D. L. Fyles,
J. Chem. Soc., Chem. Commun., 1978, 934–936; (b) B. Dietrich, D. L.
Fyles, T. M. Fyles and J.-M. Lehn, Helv. Chim. Acta, 1979, 62, 2763–
2787; (c) P. Schiebl and F. P. Schmitdchen, Tetrahedron Lett., 1993,
34, 2449–2452; (d) B. R. Linton, M. S. Goodman, E. Fan, S. A. van
Arman and A. D. Hamilton, J. Org. Chem., 2001, 66, 7313–7319; (e) J.
Raker and T. E. Glass, J. Org. Chem., 2002, 67, 6113–6116; (f) H. Ait-
Haddou, S. L. Wiskur, V. M. Lynch and E. V. Anslyn, J. Am. Chem.
Soc., 2001, 123, 11296–11297; (g) for guanidinium derived receptors
able to bind to aspartate or glutamate rich peptides, see, for example:
X. Salvatella, M. W. Peczuh, M. Gair´ı, R. K. Jain, J. Sanchez-Quesada,
20 (a) S. Pikul and E. J. Corey, Org. Synth., 1993, 71, 22; (b) E. J. Corey,
D-H. Lee and S. Sarshar, Tetrahedron: Asymmetry, 1995, 6, 3–6.
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