Enantioselective amino acid recognition using acyclic thiourea receptors

Article abstract of DOI:10.1039/b102298a

A series of acyclic thiourea derivatives, designed to create a cleft with four hydrogen bond donors suitable for carboxylate recognition, have been prepared, and their ability to bind to N-protected amino acid carboxylate salts has been investigated. The

Full text of DOI:10.1039/b102298a

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Enantioselective amino acid recognition using acyclic thiourea
receptors
Graham M. Kyne,a Mark E. Light,a Mike B. Hursthouse,a Javier de Mendozab and
Jeremy D. Kilburn*a
1
a Department of Chemistry, University of Southampton, Southampton, UK SO17 1BJ
b Departamento de Química Orgánica, Universidad Autónoma de Madrid, Cantoblanco,
E-28049, Madrid, Spain
Received (in Cambridge, UK) 9th March 2001, Accepted 25th April 2001
First published as an Advance Article on the web 11th May 2001
A series of acyclic thiourea derivatives, designed to create a cleft with four hydrogen bond donors suitable for
carboxylate recognition, have been prepared, and their ability to bind to N-protected amino acid carboxylate salts
has been investigated. The crystal structure of one of the thioureas has been determined showing that it forms a
hydrogen bonded centrosymmetric dimer in the solid-state, in a conformation appropriate for the desired binding
of carboxylates. The thioureas show good discrimination between different amino acids and those thioureas
incorporating chiral moieties show moderate enantioselectivity for a range of amino acid derivatives.
deficient aromatic amide to provide aromatic interactions with
guests with aromatic sidechains.
Introduction
Enantioselective recognition remains a major challenge for
host–guest chemists.1 The ability to discriminate between
enantiomers, using synthetic receptors, would, for example,
allow separation of racemates by selective transport across a
membrane.2 We, and others, have previously utilised thioureas
as a binding site for the carboxylate functionality,3 and
incorporation of thioureas into macrocyclic structures has
produced selective receptors for carboxylate derivatives such as
amino acids.4 Such receptors are, however, structurally rather
complex and demanding to synthesise. In an effort to produce
simpler enantioselective receptors for carboxylate derivatives
we have now prepared a series of acyclic thioureas and have
examined their ability to bind a range of amino acids.5 In these
novel receptors the thiourea functionality is linked to aryl
amides to provide additional hydrogen bonds to the carboxylate
moiety and it was anticipated that the formation of a well
defined set of four hydrogen bonds around the carboxylate,
binding to both the syn and anti lone pairs of the carboxylate,6
should also serve to create a cleft which, when chiral building
blocks are incorporated, might discriminate effectively between
enantiomeric guests (Fig. 1). The use of pyridyl amides might
further help to preorganise the chiral cleft since the desired
U-shaped conformation may be stabilised by weak hydrogen
bonding of the amide and thiourea hydrogens to the pyridyl
nitrogen.7 In order to test this hypothesis we prepared the
achiral receptors 5 and 6. We also prepared two chiral pyridyl
receptors 7 and 15, the former incorporating a primary amide,
derived from phenylalanine, to provide further hydrogen
bonding functionality, and the latter incorporating an electron
Results and discussion
The thiourea receptors 5–7 were readily prepared (Scheme 1)
Scheme 1 Reagents and conditions: (i) CSCl2, K2CO3, H2O, CH2Cl2;
(ii) 1, Et3N; (iii) CS2, DCC, DMAP; (iv) nBuNH2; (v) LiOH, dioxane,
H2O; (vi) DPPA, (2S)-2-amino-3-phenylpropanamide.
by coupling of amino esters 1 or 2 to give the corresponding
thiourea esters 3 and 4, which could be converted directly to
the butyl amides 5 and 6, or, in the case of 4, hydrolysed to the
corresponding diacid, and coupled with (2S)-2-amino-3-
phenylpropanamide to give 7. Receptor 15 was prepared from
the protected diamine 8, derived from -phenylalanine.8 Treat-
ment of 8 with hydrazine and coupling of the resulting amine
with p-nitrobenzoyl chloride gave amide 9. Removal of the
Boc-protecting group, and coupling with acid chloride 12
gave 13. Removal of the phthalimide group with hydrazine,
Fig. 1 Proposed binding of carboxylates by a bispyridylthiourea. R*
represents a chiral group.
1258
J. Chem. Soc., Perkin Trans. 1, 2001, 1258–1263
This journal is © The Royal Society of Chemistry 2001
DOI: 10.1039/b102298a

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Table 1 Hydrogen bonds for thiourea 6 [Å and Њ]
D–H ؒ ؒ ؒ Aa
d(D–H) d(H ؒ ؒ ؒ A) d(D ؒ ؒ ؒ A) Є(DHA)
N(1)–H(1N) ؒ ؒ ؒ O(2)#1 0.86(2) 2.24(2)
2.978(2)
2.897(2)
3.089(2)
144.3(18)
157.5(19)
145.9(19)
N(4)–HN4 ؒ ؒ ؒ O(2)#1
N(3)–HN3 ؒ ؒ ؒ O(2)#1
0.85(2) 2.09(2)
0.86(2) 2.34(2)
a Symmetry transformations used to generate equivalent atoms: #1
Ϫx ϩ 2, Ϫy ϩ 1, Ϫz ϩ 1.
conversion of the resulting amine 14 to the isothiocyanate and
coupling with a further equivalent of the amine, gave the
desired thiourea 15 (Scheme 2).
Fig. 2 Thermal ellipsoid plot (30% probability level) of 6 showing the
hydrogen bonded centrosymmetric dimer.
Scheme 2 Reagents and conditions: (i) H2NNH2, EtOH; (ii) p-
NO2C6H4COCl, Et3N, DMAP; (iii) TMSCl, NaI, CH3CN; (iv) SOCl2;
(v) TFA, CH2Cl2; (vi) 12, DMAP; (vii) CS2, DCC, DMAP, then further
addition of 14.
We were able to obtain a crystal structure9 of the achiral
pyridyl receptor 6 which revealed that it forms a centro-
symmetric dimer in the solid-state, with one amide carbonyl
forming a triple hydrogen bond motif (Table 1) with both of the
thiourea NH’s and one amide NH of the other monomer unit
(Fig. 2). The molecule adopts a folded conformation such that
the three NH groups point inwards to an essentially planar
cavity, into which the carbonyl of the second molecule can
dock. Two of the pyridine rings experience intermolecular π–π
stacking interactions with their centroids separated by 3.968 Å
and the angle between the least squares planes being only 0.03Њ.
Thus the crystal structure reveals, in part, the desired hydrogen
bond arrangement required for carboxylate recognition
depicted in Fig. 1. Dilution studies with NMR samples of all
the thioureas, however, showed that the dimerisation observed
in the solid state is not occurring to any significant extent
in solution. Furthermore, in neat CDCl3, the signals of the
NH’s for the pyridyl receptors 6 and 7 are considerably more
downfield in comparison to those for receptor 5, consistent with
the notion that there are weak hydrogen bonds between the
pyridyl nitrogen and the NH’s in the former case (Fig. 1).
Addition of 10% (by volume) CD3SOCD3 to these samples
leads to significant downfield shifts for the signals for both the
amide and thiourea NH’s of receptor 5, but only a small down-
field shift for the amide NH of the pyridyl receptor 6, and an
upfield shift for the thiourea NH of this compound (Fig. 3).
Fig. 3 Comparison of the NMR shifts for the NH signals of thioureas
5–7 in CDCl3 and 10% CD3SOCD3–CDCl3.
Similarly for the chiral pyridyl receptor 7, addition of 10% (by
volume) CD3SOCD3 to a sample in CDCl3 gives just a small
downfield shift for the thiourea NH signal and an upfield shift
for the secondary amide NH signal, all of which is consistent
with the notion that in the case of the pyridyl receptors 6 and
7 there is substantial intramolecular hydrogen bonding.
The benzo compound 5 proved to be rather insoluble in
deuterochloroform and binding studies with this compound
were carried out in 10% CD3SOCD3–CDCl3 as solvent,
whereas the pyridyl compounds were sufficiently soluble to
allow studies also in neat CDCl3. Initially we compared the
J. Chem. Soc., Perkin Trans. 1, 2001, 1258–1263
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Table 2 Binding constants (Kass) and free energies of complexation
(Ϫ∆Gass) for the 1 : 1 complexes formed between thioureas 5 and 6
and tetrabutylammonium phenylacetate, in 10% CD3SOCD3–CDCl3 at
20 ЊC
Table 3 Binding constants (Kass) and free energies of complexation
(Ϫ∆Gass) for the 1 : 1 complexes formed between thiourea 7 and
tetrabutylammonium salts of various amino acid derivatives, in CDCl3
at 20 ЊC
Receptor
Substrate
Kassa/MϪ1
Ϫ∆Gassb/kJ molϪ1
Entry
Substrate
Kassa/MϪ1
Ϫ∆Gassb/kJ molϪ1
ϩ
ϩ
ϩ
ϩ
5
6
PhCH2CO2Ϫ NBu4
PhCH2CO2Ϫ NBu4
740
420
16.1
14.7
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
N-Ac--Ala-CO2Ϫ NBu4
3450
2520
4770
2990
1690
800
9000
4520
1190
810
380
480
12400
14800
3140
2225
19.8
19.1
20.6
19.5
18.1
16.3
22.2
20.5
17.3
16.3
14.5
15.0
23.0
23.4
19.6
18.8
N-Ac--Ala-CO2Ϫ NBu4
ϩ
N-Ac--Phe-CO2Ϫ NBu4
N-Ac--Phe-CO2Ϫ NBu4
N-Ac--Asn-CO2Ϫ NBu4
a Errors were estimated as <10%. b Errors were estimated as <0.2 kJ
ϩ
ϩ
ϩ
molϪ1
.
N-Ac--Asn-CO2Ϫ NBu4
ϩ
N-Ac--Gln-CO2Ϫ NBu4
ϩ
N-Ac--Gln-CO2Ϫ NBu4
N-Boc--Gln-CO2Ϫ NBu4
ϩ
ϩ
N-Boc--Gln-CO2Ϫ NBu4
ϩ
N-Ac--Ser-CO2Ϫ NBu4
ϩ
N-Ac--Ser-CO2Ϫ NBu4
ϩ
N-Ac--Trp-CO2Ϫ NBu4
ϩ
N-Ac--Trp-CO2Ϫ NBu4
Fig. 4 Crabtree’s anion receptors.
N-Boc--Trp-CO2Ϫ NBu4
ϩ
ϩ
N-Boc--Trp-CO2Ϫ NBu4
a Errors were estimated as <10%. b Errors were estimated as <0.2 kJ
molϪ1
.
binding properties of the achiral receptors 5 and 6. Using
NMR titration experiments, monitoring the shift of both the
NH and ArH signals of the receptors, we obtained binding
constants for both 5 and 6 with the tetrabutylammonium salt
of phenylacetic acid in 10% CD3SOCD3–CDCl3 (Table 2).10 For
both receptors, binding with phenylacetic acid carboxylate
leads to significant downfield shifts of the signals for both the
urea NH (∼2 ppm for 5, >3 ppm for 6) and amide NH (∼1 ppm
for 5, ∼0.5 ppm for 6), indicating hydrogen bond formation,
and smaller shifts (∼0.2 ppm) of signals for the aromatic
protons. The two binding constants are similar although
binding with the pyridyl receptor 6 is slightly weaker. Thus the
pyridyl receptor 6 may be more preorganised for binding
as desired, but absolute binding of a carboxylate anion may be
disfavoured by electrostatic repulsion between the anion and
the pyridine nitrogen lone-pair—as suggested by Crabtree’s
studies on binding of various anions with related receptors 16
and 17 (Fig. 4).11
Table 4 Binding constants (Kass) and free energies of complexation
(Ϫ∆Gass) for the 1 : 1 complexes formed between thiourea 15 and
tetrabutylammonium salts of various amino acid derivatives, in CDCl3
at 20 ЊC
Entry
Substrate
Kassa/MϪ1
Ϫ∆Gassb/kJ molϪ1
ϩ
ϩ
1
2
3
4
N-Boc--Trp-CO2Ϫ NBu4
1925
3785
1570
1870
18.4
20.1
17.9
18.4
N-Boc--Trp-CO2Ϫ NBu4
(R)-Naproxen-CO2Ϫ NBu4
ϩ
ϩ
(S)-Naproxen-CO2Ϫ NBu4
a Errors were estimated as <10%. b Errors were estimated as <0.2 kJ
molϪ1
.
Binding studies with the chiral pyridyl receptor 7 and a range
of amino acid derivatives, as the tetrabutylammonium salts,
in neat CDCl3, were carried out and the results are presented in
Table 3. The receptor is strongly selective for different amino
acids (e.g. >30 : 1 selectivity for N-Ac--Trp-CO2Ϫ over N-Ac--
reagents. Thus, for example, we prepared a CDCl3 solution
of receptor 6 with the tetrabutylammonium salt of racemic
N-Ac-Phe, which leads to two cleanly resolved signals for the
NCOCH3 group of the two resulting diastereomeric complexes
at 1.86 and 1.88 ppm.
Ϫ
Ser-CO2 ), but in general the receptor shows only moderate
enantioselectivity with a general preference for -amino acids
and a highest enantioselectivity for N-Ac-Gln-CO2 ( :  ≈
2 : 1). Binding constants were also highest for amino acids with
Thiourea 15 showed similar downfield shifts for the thiourea
NH (∼2 ppm), pyridyl amide NH (∼0.5 ppm) and nitrobenz-
amide NH (∼1.1 ppm) on titration with the enantiomers of
N-Boc-Trp, and the receptor now showed a modest selectivity
for the -enantiomer (Table 4). A probable contribution to
the binding from aromatic interactions is evidenced by
downfield shifts (∼0.2 ppm) of the signals for both aromatic
protons from the p-nitrobenzamide on complexation. The
receptor also bound the enantiomers of naproxen (as the
tetrabutylammonium salts), with associated shifts (∼0.2 ppm)
for the signals for the protons from the p-nitrobenzamide,
but was unable to discriminate effectively between the
enantiomers.
In summary we have prepared a series of simple acyclic
thiourea receptors which are both sidechain selective and
moderately enantioselective for a range of amino acid deriva-
tives. Despite the modest enantioselectivity observed the
receptors are simple to prepare, and evidence from the NMR
binding studies does suggest that the tweezer receptors do
indeed wrap around the substrates as desired using a com-
bination of several hydrogen bonding interactions. These
results indicate that more selective receptors may result from
the introduction of suitable functionality for more specific
interactions with the sidechain functionality of the amino acid
guests.
Ϫ
sidechains incorporating hydrogen bonding functionality (N-
Ϫ
Ac-Gln-CO2 ) or an electron rich aromatic sidechain (N-Ac-
Ϫ
Trp-CO2 ). The corresponding N-Boc derivatives gave reduced
binding constants, but gave better enantioselectivity in the case
Ϫ
of N-Boc-Trp-CO2 . For all substrates tested, binding resulted
in significant downfield shifts of the NH signals from the
thiourea (1.6–2.1 ppm), the secondary amide (0.3–0.7 ppm) and
for one of the primary amide protons (1.2–1.5 ppm), and an
upfield shift (0.3–0.7 ppm) of the other primary amide proton.
This suggests that binding of the amino acids involves hydrogen
bonds to the thiourea, secondary amide and one of the primary
amide NH’s, but also involves breaking of an intramolecular
hydrogen bond to the other primary amide NH. Conversely,
binding of the tryptophan derivatives led to a significant
upfield shift for the indole NH signal (>1 ppm) presumably
reflecting the breaking of an intramolecular hydrogen bond
between the indole NH and the carboxylate on binding, and
similarly binding of the glutamine and asparagine derivatives
led to upfield shifts for one of the primary amide NH’s in
each case.
As well as showing modest enantioselectivity in binding
amino acid derivatives, the receptors can be used as chiral shift
1260
J. Chem. Soc., Perkin Trans. 1, 2001, 1258–1263

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and 1535; m/z (ES)ϩ 455.4 (M ϩ H)ϩ (Found: C, 63.98; H, 7.59;
N, 11.74. C25H34N4O2S.MeOH requires C, 64.17; H, 7.87; N,
11.51%).
Experimental
General procedures
All reactions requiring anhydrous conditions were conducted
in flame dried glassware under a static, inert atmosphere unless
otherwise stated. CH2Cl2 was distilled from calcium hydride,
and petrol was distilled and the fraction boiling between 40
and 60 ЊC was used. All other solvents were of commercial
grade and were used without further purification. Thin layer
chromatography was performed on plastic backed sheets
(Camlab) coated with silica gel (SiO2: 0.25 mm). Flash column
chromatography was performed on Sorbil C60, 40–60 mesh
silica.
Ethyl 6-({[({[6-(ethoxycarbonyl)-2-pyridyl]methyl}amino)thio-
carbonyl]amino}methyl)pyridine-2-carboxylate 4
Based on a procedure originally reported by Anslyn,12 DMAP
(0.57 g, 4.70 mmol) was added to a suspension of amine hydro-
chloride salt 2 (0.41 g, 1.88 mmol), DCC (0.19 g, 0.94 mmol)
and carbon disulfide (0.40 mL, 6.58 mmol) in dry CHCl3
(10 mL) at Ϫ10 ЊC. The solution was stirred at rt for 1 hour
then heated at reflux for 12 hours. The solvent was removed in
vacuo and the resultant orange oil was purified by flash column
chromatography, eluting with ethyl acetate–petroleum ether
(70 : 30 to 100 : 0 v/v) to give thiourea 4 as a yellow solid
(0.25 g, 64%): mp 103–104 ЊC (ethyl acetate–petroleum ether);
Rf 0.44 (ethyl acetate); δH (300 MHz, CDCl3) 1.45 (6H, t, J 7,
CH3), 4.49 (4H, q, J 7, CH2CH3), 5.02 (4H, br s, CH2N), 7.50
(2H, d, J 8, pyrH), 7.83 (2H, t, J 8, pyrH), 8.02 (2H, d, J 8,
pyrH), 8.56 (2H, br s, NH); δC (75.5 MHz, CDCl3) 14.4, 49.5,
62.2, 124.0, 125.7, 138.0, 147.2, 157.4, 165.1, 183.3; νmax/cmϪ1
(neat) 3545, 3300, 1705, and 1530; m/z (ESϩ) 403.0 (M ϩ H)ϩ,
424.9 (M ϩ Na)ϩ (Found: C, 56.69; H, 5.68; N, 13.63.
C19H22N4O4S requires C, 56.70; H, 5.51; N, 13.92%).
Infra-red spectra were recorded on a Perkin-Elmer 1600
series FT-IR spectrometer. Proton NMR spectra were obtained
at 270 MHz on a JEOL GX 270, at 300 MHz on a Bruker AC
300, and at 360 MHz on a Bruker Aspect 3000 spectrometer.
Spectra were referenced with respect to the residual solvent
peak for the deuterated solvent concerned. 13C NMR spectra
were obtained at 75 MHz on a Bruker AC 300 and at 100 MHz
1
on a Bruker DPX 400. COSY spectra and H–13C correlation
spectra were performed on a Bruker AC 300. Mass spectra were
obtained on a VG analytical 70-250-SE normal geometry
double focusing mass spectrometer or on a Micromass Platform
quadrupole mass analyser with an electrospray ion source.
N-1-Butyl-6-[({[({6-[(butylamino)carbonyl]-2-pyridyl}methyl)-
amino]thiocarbonyl}amino)methyl]pyridine-2-carboxamide 6
Methyl 3-{[({[3-(methoxycarbonyl)benzyl]amino}thiocarbonyl)-
amino]methyl}benzoate 3
Diester 4 (26 mg, 65 µmol) was dissolved in butylamine (4 mL)
and the solution was heated at reflux for 12 hours. The excess
butylamine was removed in vacuo and the resultant yellow solid
was recrystallised from ethyl acetate–petroleum ether to give
thiourea 6 as colourless crystals (32 mg, 100%): mp 95–96 ЊC
(ethyl acetate–petroleum ether); Rf 0.31 (ethyl acetate); δH (300
MHz, CDCl3) 0.80 (6H, t, J 7, CH3), 1.21 (4H, sextet, J 7,
CH2CH3), 1.44 (4H, quintet, J 7, CH2CH2CH3), 3.26 (4H,
Based on a procedure originally reported by Anslyn,12 a suspen-
sion of amine hydrochloride 1 (0.84 g, 4.17 mmol) in CH2Cl2
(15 mL) was added dropwise to a vigorously stirred mixture
of potassium carbonate (1.14 g, 8.34 mmol) in water (45 mL)
and thiophosgene (0.73 mL, 9.52 mmol) in CH2Cl2 (45 mL).
The mixture was stirred at rt for 1 hour then heated at reflux for
12 hours. After cooling to rt the organic layer was separated,
dried (MgSO4), filtered and concentrated in vacuo to give crude
isothiocyanate as an orange oil which was used in the next
step without further purification. Amine 1 (0.65 g, 3.24 mmol)
was added to a stirred solution of the isothiocyanate (0.67 g,
3.31 mmol) and triethylamine (1.32 mL, 9.72 mmol) in CH2Cl2
(30 mL). The mixture was heated at reflux for 12 hours and
after allowing to cool to rt was washed with 2.0 M HCl (3 ×
20 mL), dried (MgSO4) and the solvent removed in vacuo to
give thiourea 3 as an orange viscous oil (1.15 g, 74%): Rf 0.47
(ethyl acetate); δH (300 MHz, CDCl3) 3.83 (6H, s, CH3), 4.70
(4H, d, J 5, ArCH2), 6.74 (2H, br s, NH), 7.32 (2H, t, J 8, ArH),
7.45 (2H, d, J 8, ArH), 7.83 (2H, s, ArH), 7.84 (2H, d, J 8,
ArH); δC (75.5 MHz, CDCl3) 48.2, 52.4, 128.7, 129.0, 129.1,
130.6, 132.4, 137.9, 167.1, 182.9; m/z (ESϩ) 373.5 (M ϩ H)ϩ,
395.5 (M Ϫ Na)ϩ HRMS: found 372.1223. C19H21N2O4S
(M ϩ H)ϩ requires 373.1222.
q, J 7, CH CH N), 4.98 (4H, s, CH NHC᎐S), 7.48 (2H, d,
᎐
2
2
2
J 8, pyrH), 7.77 (2H, t, J 8, pyrH), 7.96 (2H, d, J 8, pyrH), 8.02
(2H, br s, NH), 8.29 (2H, br s, NH); δC (75.5 MHz, CDCl3)
13.8, 20.3, 31.8, 39.6, 49.5, 121.3, 125.2, 138.8, 149.0, 155.9,
164.1; νmax/cmϪ1 (neat) 3360, 3300, 2950, 2860, 2360, 1670,
1640, 1570 and 1530; m/z (ESϩ) 457.4 (M ϩ H)ϩ, 479.5
(M ϩ Na)ϩ; HRMS (FAB): found 457.2384. C23H33N6O2S
requires 457.2386. An X-ray crystal structure was obtained
(see text).
N-[(1S)-2-Amino-1-benzyl-2-oxoethyl]-6-({[({[6-({[(1S)-2-
amino-1-benzyl-2-oxoethyl]amino}carbonyl)-2-pyridyl ]methyl}-
amino)thiocarbonyl]amino}methyl)pyridine-2-carboxamide 7
LiOH (20.1 mL of a 1 M soln, 20.1 mmol) was added to diester
4 (0.81 g, 2.01 mmol) in 1,4-dioxane (23 mL) and the mixture
was stirred for 1 hour. KHSO4 (20.1 mL of a 1 M soln, 20.1
mmol) was added and the water was removed in vacuo and by
azeotroping with acetonitrile (20 mL). Methanol (5 mL) was
added to the resultant solid, the insoluble material was removed
by filtration and the excess methanol was removed in vacuo
to yield the diacid as a pale yellow solid (0.70 g, 100%) which
was used in the next reaction without further purification.
Diphenylphosphoryl azide (DPPA) (0.42 mL, 1.92 mmol)
followed by triethylamine (0.53 mL, 3.83 mmol) were added to
a stirred mixture of the diacid (0.30 g, 0.87 mmol) and (2S)-
2-amino-3-phenylpropanamide (0.31 g, 1.92 mmol) in DMF
(0.5 mL) at 0 ЊC. The mixture was stirred at 0 ЊC for 2 hours
and at rt for 2 days. The solvent was removed in vacuo and the
crude oil was suspended in CH2Cl2 (15 mL), washed with water
(15 mL), dried (MgSO4) and concentrated in vacuo to give an oil
which was purified by flash column chromatography eluting
with methanol–CH2Cl2 (1 : 99 to 10 : 90 v/v) to give thiourea 7
as a white solid (84 mg, 15%): mp 126–128 ЊC (ethanol–water);
N-1-Butyl-3-({[({3-[(butylamino)carbonyl]benzyl}amino)-
thiocarbonyl]amino}methyl)benzamide 5
Diester 3 (50 mg, 0.13 mmol) was dissolved in butylamine
(5 mL) and heated at reflux for 12 hours. After cooling to rt the
excess butylamine was removed in vacuo to yield a white solid
which was purified by crystallisation (ethyl acetate–methanol)
to give thiourea 5 as colourless crystals (25 mg, 41%): mp 88–
90 ЊC (ethyl acetate–methanol); Rf 0.47 (ethyl acetate); δH (300
MHz, CDCl3) 0.94 (6H, t, J 7, CH3), 1.37 (4H, sextet, J 7,
CH2CH2CH3), 1.55 (4H, quintet, J 7, CH2), 3.32 (4H, q, J 7,
CH2NHCO), 4.7 (4H, d, J 5, CH2NHCS), 6.38 (2H, br s,
NHCS), 6.98 (2H, br s, NHCO), 7.32 (2H, d, J 8, ArH), 7.40
(2H, d, J 8, ArH), 7.53 (2H, d, J 8, ArH), 7.54 (2H, s, ArH);
δC {100 MHz, [(CD3)2SO–CDCl3 10 : 90 v/v]} 11.5, 18.1, 29.4,
37.8, 45.8, 124.0, 124.1, 126.7, 128.5, 133.0, 136.7, 166.8, 182.4;
νmax/cmϪ1 (neat) 3285, 3225, 3060, 2960, 2930, 2865, 1635, 1585
J. Chem. Soc., Perkin Trans. 1, 2001, 1258–1263
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Rf 0.43 [methanol–CH2Cl2 (10 : 90 v/v)]; [α]2D0 Ϫ36.0Њ (c 1 in
CH2Cl2); δH (300 MHz, CDCl3) 3.12 (2H, dd, J 7, 14, CHA-
HBPh), 3.17 (2H, dd, J 7, 14, CHAHBPh), 4.87 (2H, apparent q,
J 7, CONHCH), 4.99 (4H, m, CSNHCH2), 6.34 (2H, br s,
NHAHB), 6.47 (2H, br s, NHAHB), 7.15–7.25 (10H, m, ArH),
7.43 (2H, d, J 8, ArH), 7.74 (2H, t, J 8, pyrH), 7.91 (2H, d, J 8,
pyrH), 8.00 (2H, br s, CSNH), 8.80 (2H, d, J 8, CONHCH);
δC (75.5 MHz, CDCl3) 39.4, 50.0, 55.6, 121.5, 125.9, 127.8,
129.4, 130.2, 130.4, 138.0, 139.3, 149.6, 158.6, 165.8 and 175.69;
νmax/cmϪ1 2395, 2360, 1660, 1650 and 1515; m/z (ESϩ) 639.6
(M ϩ H)ϩ, 661.6 (M ϩ Na)ϩ, 677.5 (M ϩ K)ϩ; HRMS (FAB):
found 661.2311. C33H34N8O4NaS requires 661.2321.
0 ЊC and the mixture was stirred for 12 hours at rt. The solvent
was removed in vacuo and the resulting oil was triturated with
ether (2 × 10 mL) to give the amine TFA salt as a pale yellow
oil. Acid 11 (0.19 g, 0.68 mmol) was heated at reflux in thionyl
chloride (2 mL) for 8 hours then cooled and stirred at rt for
12 hours. The excess thionyl chloride was removed in vacuo to
give acid chloride 12 as a white solid, which was dissolved in dry
DMF (5 mL) and added dropwise to a solution of amine TFA
salt and DMAP (70 mg, 0.57 mmol) in DMF and the mixture
was stirred at rt for 2 days. Solvent was removed in vacuo and
the brown residue was purified by filtration through a pad of
silica gel, eluting with ethyl acetate, to give amide 13 as a white
solid (0.23 g, 79%): Rf 0.59 (ethyl acetate); mp 174–176 ЊC
(CHCl3–hexane); [α]2D0 Ϫ11.3Њ (c 1 in CH2Cl2); δH (400 MHz,
CDCl3) 2.78 (1H, dd, J 8, 15, CHAHBPh), 2.82 (1H, dd, J 7, 15,
CHAHBPh), 3.38 (1H, ddd, J 3, 4, 14, NHCHACHB), 3.63 (1H,
ddd, J 5, 10, 14, NHCHACHB), 4.37 (1H, m, NHCHCH2), 4.97
(2H, s, CH2Phth), 7.11–7.25 (5H, m, ArH), 7.40 (2H, d, J 8,
pyrH), 7.72 (2H, dd, J 3, 5, ArH), 7.76 (1H, t, J 8, pyrH), 7.84
(2H, dd, J 3, 5, ArH), 7.86–7.90 (3H, m, ArH and NHCH2CH),
7.96 (1H, d, J 8, pyrH), 8.12 (1H, d, J 8, NHCHCH2), 8.17 (2H,
d, J 9, ArH); δC (100 MHz, CDCl3) 39.3, 43.0, 47.3, 51.5, 121.6,
124.0, 124.1, 125.3, 127.5, 128.7, 129.5, 129.3, 132.4, 134.9,
136.9, 139.0, 140.1, 149.0, 149.9, 154.7, 165.7, 166.1, 168.4;
νmax/cmϪ1 (neat) 3350, 2360, 1685, 1644, 1600 and 1550; m/z
(ESϩ) 564.3(M ϩ H)ϩ. HRMS: found 564.1879. C31H26N5O6
(M ϩ H)ϩ requires 564.1883.
6-[(1,3-Dioxo-2,3-dihydro-1H-isoindol-2-yl)methyl]pyridine-
2-carboxylic acid 11
Based on a procedure originally reported by Olah,13 chloro-
trimethylsilane (16.6 mL, 0.13 mol) was added to a solution
of ester 1014 (10.1 g, 32.7 mmol) and sodium iodide (19.6 g,
0.13 mol) in acetonitrile (50 mL) at 0 ЊC. The reaction mixture
was allowed to warm to rt and heated at reflux for 4 days. After
allowing to cool to rt, water (200 mL) was added, the reaction
mixture was diluted with CH2Cl2 (100 mL) and was washed
successively with water (100 mL) followed by aqueous sodium
thiosulfate (150 mL). The organic phase was extracted with
saturated sodium hydrogen carbonate (50 mL) and the aqueous
phase acidified to pH 3 using 2.0 M HCl. The acidic solution
was extracted with methanol–CH2Cl2 [3 × 150 mL (5 : 95 v/v)],
dried (MgSO4) and the solvent removed in vacuo to give acid 11
as a white solid (3.12 g, 34%): mp 219–221 ЊC (from methanol–
CHCl3); Rf 0.21 [methanol–CH2Cl2 (10 : 90 v/v)]; δH {400 MHz,
[(CD3)2SO–CDCl3 10 : 90 v/v]} 5.03 (2H, s, PhthNCH2), 7.33
(1H, dd, J 1, 8, pyrH), 7.73 (2H, dd, J 3, 5, ArH), 7.77 (1H,
t, J 8, pyrH), 7.82 (2H, dd, J 3, 5, ArH), 7.96 (1H, dd, J 1, 8,
pyrH); δC (100 MHz, CD3OD) 42.5, 123.1, 123.2, 123.7, 131.5,
134.0, 137.8, 147.6, 155.5, 165.7, 167.5; νmax/cmϪ1 (neat) 3360,
1750, 1705, 1600 and 1420 (Found: C, 64.07; H, 3.55; N, 9.95.
C15H10N2O4 requires C, 63.83; H, 3.57; N, 9.92%).
N-2-{(1S)-1-Benzyl-2-[(4-nitrobenzoyl)amino]ethyl}-6-
[({6-[({(1S)-1-benzyl-2-[(4-nitrobenzoyl)amino]ethyl}amino)-
carbonyl]-2-pyridyl}methyl)amino]thiocarbonyl}amino)-
methyl]-2-pyridinecarboxamide 15
Hydrazine hydrate (17 µL, 0.35 mmol) was added to a solution
of phthalimide 13 (0.20 g, 0.35 mmol) in ethanol (5 mL) and the
mixture heated at reflux for 12 hours. The insoluble material
was removed by filtration and the filtrate concentrated in vacuo
to give amine 14 as a yellow residue which was used in the
next step without further purification. Carbon disulfide (74 µL,
1.21 mmol) was added to a solution of the amine (75 mg,
1.73 µmol), DCC (36 mg, 174 µmol) and DMAP (43 mg, 0.35
mmol) in dry CH2Cl2 (2 mL) at Ϫ10 ЊC and the mixture was
stirred for 1 hour. After warming to rt the excess solvent and
carbon disulfide were removed in vacuo. The resultant orange
oil was dissolved in dry CH2Cl2 (2 mL) and further amine
14 (75 mg, 1.73 µmol) was added. After stirring at rt for 5 days
the solvent was removed in vacuo and the brown residue
was purified by flash column chromatography eluting with
methanol–CH2Cl2 (1 : 99 to 10 : 90 v/v) to give thiourea 15 as
a pale yellow solid (60 mg, 25%): Rf 0.59 (ethyl acetate);
[α]2D0 ϩ 36.0Њ (c 2 in CH2Cl2); δH (400 MHz, CDCl3) 2.90 (1H,
dd, J 8, 14, CHAHBPh), 3.22 (1H, dd, J 6, 14, CHAHBPh), 3.58
(1H, dd, J 3, 14, CHAHBNH), 3.80 (1H, dd, J 10, 14, CHAHB-
NH), 4.51 (2H, m, CHCH2Ph), 5.02 (2H, d, J 17, CHAHBN-
HCS), 5.10 (2H, d, J 17, CHAHBNHCS), 7.22–7.35 (10H, m,
ArH), 7.56 (2H, d, J 8, pyrH), 7.73 (2H, br s, CH2NHCO),
7.83 (4H, d, J 9, ArH), 7.89 (2H, t, J 8, pyrH), 8.07 (2H, d,
J 8, pyrH), 8.11 (4H, d, J 8.5, ArH), 8.44 (2H, br m, NHCS),
8.89 (2H, d, J 5, NHCH); δC (100 MHz, CDCl3) 37.3, 42.7, 47.2,
51.0, 120.0, 122.3, 123.9, 125.5, 126.7, 127.4, 127.7, 135.5,
137.7, 146.4, 148.1, 154.1, 162.6, 165.9, 182.3; νmax/cmϪ1 (neat)
3285, 1650, 1595 and 1518; m/z (ESϩ) 910.4 (M ϩ H)ϩ (Found:
C, 62.38; H, 5.10; N, 15.46. C47H44N10O8SؒMeOH requires C,
62.10; H, 4.88; N, 15.41%).
tert-Butyl-N-{(1S)-1-benzyl-2-[(4-nitrobenzoyl)amino]ethyl}-
carbamate 9
Hydrazine hydrate (90 µL, 1.88 mmol) was added to a solution
of phthalimide 88 (0.48 g, 1.26 mmol) in ethanol (25 mL)
and the mixture heated at reflux for 6 hours. After cooling to
rt the insoluble material was removed by filtration and the
excess solvent removed in vacuo to give the amine as a pale
yellow solid, which was suspended in dry CH2Cl2 (20 mL).
4-Nitrobenzoyl chloride (0.52 g, 2.82 mmol) followed by tri-
ethylamine (0.39 mL, 2.82 mmol) and DMAP (50 mg, 0.41
mmol) were added. After stirring at rt for 12 hours solvent
was removed in vacuo and the crude material was purified by
flash column chromatography eluting with ethyl acetate–
petroleum ether (30 : 70 v/v) to give amide 9 as a white solid
(0.31 g, 63%): mp 184–186 ЊC (ethyl acetate); Rf 0.5 [ethyl
acetate–petroleum ether (30 : 70 v/v)]; δH {300 MHz,
[(CD3)2SO–CDCl3 10 : 90 v/v]} 1.29 (9H, s, C(CH3)3), 2.69 (1H,
dd, J 9, 14, CHAHBPh), 2.80 (1H, dd, J 6, 14, CHAHBPh), 3.38–
3.33 (2H, m, CH2N), 3.93 (1H, dt, J 6, 9, CHCH2), 6.81 (1H, d,
J 9, NHCH), 7.31–7.16 (5H, m, ArH), 8.07 (2H, d, J 9, ArH),
8.32 (2H, d, J 9, ArH), 8.80 (1H, t, J 5, NHCH2); δC (100 MHz,
DMSO-d6) 26.5, 36.1, 42.0, 49.7, 75.8, 121.7, 124.2, 126.3,
127.1, 127.4, 137.2, 138.6, 147.2, 153.6, 163.2; νmax/cmϪ1 (neat)
3350, 1690, 1644, 1550 and 1525; m/z (ESϩ) 400.4 (M ϩ H)ϩ
(Found: C, 63.28; H, 6.62; N, 10.44. C21H25N3O5 requires C,
63.15; H, 6.31; N, 10.52%).
Acknowledgements
N-{(1S)-1-Benzyl-2-[(4-nitrobenzoyl)amino]ethyl}-6-[(1,3-dioxo-
2,3-dihydro-1H-isoindol-2-yl)methyl]pyridine-2-carboxamide 13
We thank the EPSRC for a studentship (GK) and the EC
(TMR Network Grant “Enantioselective Separations” ERB
FMRX-CT-98-0233) for financial support.
TFA–CH2Cl2 [5 mL (50 : 50 v/v)] was added to a solution of
protected amine 9 (0.21 g, 0.53 mmol) in CH2Cl2 (10 mL) at
1262
J. Chem. Soc., Perkin Trans. 1, 2001, 1258–1263

View Article Online
8 K. Burgess, J. Ibarzo and D. S. Linthicum, J. Am. Chem. Soc., 1997,
119, 1556.
References
1 For a recent review on synthetic receptors, see: J. H. Hartley,
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2 W. H. Pirkle and W. E. Bowen, Tetrahedron: Asymmetry, 1994,
5, 773; J. T. F. Keurentjes, L. J. W. M. Nabuurs and E. A. Vegter,
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3 For original use of thioureas for carboxylate recognition, see:
E. Fan, S. A. Van Arman, S. Kincaid and A. D. Hamilton, J. Am.
Chem. Soc., 1993, 115, 369; P. J. Smith, M. V. Reddington and
C. S. Wilcox, Tetrahedron Lett., 1992, 33, 6085.
4 G. J. Pernia, J. D. Kilburn, J. W. Essex, R. J. Mortishire-Smith and
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5 Enantioselective recognition of amino acids: A. P. Davis and
L. J. Lawless, Chem. Commun., 1999, 9 and refs therein.
6 For related receptors, designed to bind to both the syn and anti lone
pairs of a carboxylate, see: B. C. Hamann, N. R. Branda and
J. Rebek Jr., Tetrahedron Lett., 1993, 34, 6837; J. S. Albert and
A. D. Hamilton, Tetrahedron Lett., 1993, 34, 6483; C. Raposo,
M. Crego, L. Mussons, C. Caballero and J. R. Moran, Tetrahedron
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Y. Umezawa, Tetrahedron Lett., 1995, 36, 6483.
9 Data were collected on a Nonius Kappa CCD area detector
diffractometer with a rotating anode following standard procedures.
Crystal data for 6: C23H32N6O2S, Mr = 456.61, T = 150(2) K,
¯
triclinic, space group P1, a = 9.4566(3), b = 10.7149(4), c =
13.2326(4) Å, α = 89.975(2), β = 79.123(2), γ = 65.1932(12)Њ,
V = 1190.56(7) Å3, ρcalc = 1.274 g cmϪ3, µ = 0.168 mmϪ1, Z = 2,
reflections collected: 18090, independent reflections: 4737
(Rint = 0.043), final R indices [I > 2σI]: R1 = 0.0429, wR2 = 0.0976,
R indices (all data): R1 = 0.0529. wR2 = 0.1023. CCDC reference
number 160088. See http://www.rsc.org/suppdata/p1/b1/b102298a/
for crystallographic files in .cif or other electronic format.
10 The binding constants were calculated by fitting the data to a 1 : 1
binding isotherm using NMRTit HG software, kindly provided by
Professor C. A. Hunter, University of Sheffield. See: A. P. Bisson,
C. A. Hunter, J. C. Morales and K. Young, Chem. Eur. J., 1998,
4, 845.
11 K. Kavallieratos, C. M. Bertao and R. H. Crabtree, J. Org. Chem.,
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12 D. M. Kneeland, K. Ariga, V. M. Lynch, C.-Y. Huang and
E. V. Anslyn, J. Am. Chem. Soc., 1993, 115, 10042.
13 G. A. Olah, S. C. Narang, B. Gupta and R. Malhotra, J. Org. Chem.,
1979, 44, 1247.
14 P. Scrimin, U. Tonellato, D. Milani and R. Fornasier, J. Chem. Soc.,
Perkin Trans. 2, 1986, 233.
7 The conformational preferences of pyridine-2,6-dicarboxamides
and isophthalamides have been discussed in detail: C. A. Hunter and
D. H. Purvis, Angew. Chem., Int. Ed. Engl., 1992, 31, 792.
J. Chem. Soc., Perkin Trans. 1, 2001, 1258–1263
1263