Synthesis and antiplasmodial activity of streptocyanine/peroxide and streptocyanine/4-aminoquinoline hybrid dyes

Article abstract of DOI:10.1039/c1ob06048a

Two series of streptocyanine dyes incorporating cyclic peroxide or 4-aminoquinoline moieties are prepared and X-ray diffraction structures for three compounds are determined. All hybrid dyes show good antiplasmodial activity (0.06 to 0.66 μM) and are not

Full text of DOI:10.1039/c1ob06048a

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PAPER
Synthesis and antiplasmodial activity of streptocyanine/peroxide and
streptocyanine/4-aminoquinoline hybrid dyes†
Marie-Pierre Maether,a Virginie Bernat,a Marie Maturano,a Christiane Andre´-Barre`s,a Sonia Ladeira,b
Alexis Valentin,c Henri Viald and Corinne Payrastre*a
Received 29th June 2011, Accepted 21st July 2011
DOI: 10.1039/c1ob06048a
Two series of streptocyanine dyes incorporating cyclic peroxide or 4-aminoquinoline moieties are
prepared and X-ray diffraction structures for three compounds are determined. All hybrid dyes show
good antiplasmodial activity (0.06 to 0.66 mM) and are not or are slightly cytotoxic, except 10a.
Introduction
Malaria is one of the most serious infectious diseases in tropical
and subtropical regions. Every year, 300 million people are afflicted
leading to 880 000 deaths, especially among young children.1 An
urgent need exists to develop new classes of antimalarial drugs that
operate, or not, by novel mechanisms of action because malaria
parasites develop resistance to clinically used chemotherapeutic
Fig. 1 5C- (n = 1),5 7C- (n = 2)6 and 9C-7 chain streptocyanine dyes.
agents and prophylactic drugs such as chloroquine, mefloquine or
sulfadoxine–pyrimethamine, and recently to artemisinin.2 Nowa-
days, drug combinations are recommended to avoid the emergence
of resistant parasite strains. But the two independent drugs may
present different pharmacokinetics profiles which hamper the full
benefit of the drug combinations. A recent rational approach
involves “covalent bitherapy”. This new strategy is based on
using hybrid molecules with a dual mode of action. The two
active entities are covalently linked. These drugs allow bypassing
the development of resistance, enhancing patient compliance and
reducing drug–drug interactions.3
We previously described the synthesis of streptocyanine dyes
as a potential new class of antimalarial drugs.4 SAR studies have
shown the influence of the polymethine chain length (5C-, 7C- or
9C-) and also the great importance of the structural modifications
at the nitrogen end groups (Fig. 1). The most active compounds
displayed sub-micromolar in vitro activities against P. falciparum
and the best selectivity was obtained for 5C-streptocyanines with
morpholino end groups.
Here we report our results on the synthesis and in vitro
antiplasmodial activities of two series of new hybrid molecules.
Firstly, a cyclic peroxide moiety and secondly, a 4-aminoquinoline
moiety, were chosen to be tagged on the streptocyanines with the
aim to induce a synergic effect and/or to create molecules with
a dual mode of action. We are also interested in antiplasmodial
agents acting in a similar way to artemisinin and we focused on
the syntheses of modified bicyclic peroxide G-factors (G1, G2,
G3). These natural bicyclic peroxides are easily extracted from the
leaves of Eucalyptus grandis8 (Fig. 2). We have previously reported
the crucial role of the peroxyketal function for this activity.9 But
the most significant differences were those observed between the
natural product G3 (IC50 30 mM) and the methyl ether analogue
G3Me (IC50 0.28 mM). The latter compound was found to be
one hundred times more active than G3, indicating the crucial
role of the ether function compared to the hydroxyl one. Some
of the previously synthesised a-spiro derivatives present moderate
aLaboratoire de Synthe`se et Physico-Chimie de Mole´cules d’Inte´reˆt Bi-
ologique (UMR 5068), Universite´ de Toulouse (Universite´ Paul Sabatier),
31062 Toulouse Cedex 9, France. E-mail: payrastr@chimie.ups-tlse.fr;
Fax: (+)330561556011; Tel: (+)330561558392
bUniversite´ de Toulouse, UPS, Structure Fe´de´rative Toulousaine en Chimie
Mole´culaire, FR2599, 118 Route de Narbonne, F-31062 Toulouse, France
cUPS and IRD, Pharma-DEV, UMR 152, Universite´ de Toulouse, 118 route
de Narbonne, 31062 Toulouse cedex 9, France
dDynamique des Interactions Membranaires Normales et Pathologiques
(UMR 5235), Universite´ Montpellier; 2, cc 107, Place E. Bataillon, 34095
Montpellier cedex 5, France
† CCDC reference numbers 805391–805393. For crystallographic data in
CIF or other electronic format see DOI: 10.1039/c1ob06048a
Fig. 2 Natural G-factors and a-spirobicyclic peroxide analogues of
G3-factor.
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to good antiplasmodial activities.10 We decided to synthesise the
a-spiropiperidine analogue allowing the tag of the streptocyanine
and obtaining only one compound, avoiding the presence of two
diastereoisomers.
The 4-aminoquinoline moiety11 was introduced to the strepto-
cyanine dyes via cyclic or acyclic linkers (Fig. 3).
on the peroxyhemiketal position, using butyllithium solution then
methyl triflate at low temperature. Methylated bicyclic peroxide
5 is obtained in 65% yield after purification by silica gel column
chromatography. Then BOC is cleaved using trifluoroacetic acid
at room temperature furnishing peroxide 6 in 92% yield, which is
used in the next coupling without further purification.
The synthesis of functionalized 4-aminoquinoline 7a, b (Fig. 3)
was achieved by the nucleophilic substitution of the 4-Cl atom of
4,7-dichloroquinoline with ethylenediamine or piperazine accord-
ing to Chauhan’s procedures.11
Hybrid peroxo-streptocyanine dyes 10a, b, 11b and 12b were
synthesised by the synthetic procedures as shown in Scheme 2.
Unsymmetrical compounds 10a, b and 11b were obtained in
respective yields of 22, 28 and 48% via a hemicarboxonium salt
which is isolated (13a) or not (9b). The symmetrical hybrid peroxo-
streptocyanine dye 12b was obtained by reacting peroxide 6 with
carboxonium salt 8b in 14% yield. To obtain the compounds 10a
and 12b, Hu¨nig’s base activation of 6 was necessary to achieve the
substitution of the second ethoxy group.
Nucleophilic substitution of the ethoxy group of hemicarbox-
onium salts 13b and 14b with 7a (quino) or 7b (pipequino) gives
unsymmetrical hybrid 4-aminoquinoline–streptocyanine dyes 15b,
16b and 18b, 19b in 26, 59, 30 and 50% yield, respectively
(Scheme 3).
Fig. 3 4-Amino-7-chloroquinolines 7a and 7b.11
Results and discussion
Synthesis
Because streptocyanine dyes are synthesised by reacting various
nitrogen nucleophiles with hemicarboxonium or carboxonium
salts,5 the functionalization of peroxide and 4-aminoquinoline
moieties by a reactive primary or secondary amino group has
been necessary (Scheme 1 and Fig. 3, respectively).
The functionalized cyclic peroxide
6 (peroxo) is ob-
To introduce two 4-aminoquinoline groups, the reaction was
carried out from the carboxonium salt 8b and the symmetrical
hybrid 4-aminoquinoline–streptocyanine dyes 17b and 20b were
obtained in 30 and 70% yield respectively, depending on the acyclic
or cyclic linker used (Scheme 4).
As was previously observed for streptocyanine dyes,6,7,12 X-ray
diffraction structures of 17b (Fig. 4), 18b (Fig. 5) and 19b (Fig. 6)
reveal “all trans” 5C carbon chains, with angles of about 120◦.
The polymethine chains are almost planar and the aryl groups
are roughly perpendicular to the chains, except for 17b. The C–C
and C–N bond lengths of the conjugated system have intermediate
tained by the following procedure (Scheme 1). 1-BOC-4-
hydroxymethylpiperidine is first oxidized to aldehyde 1 in quan-
titative yield. Then, modified Knoevenagel reaction between
syncarpic acid and the iminium obtained from aldehyde 1 and
piperidine leads to Mannich base 2, which after acidic treatment
and elimination of piperidine, furnishes the enone precursor 3 in
equilibrium with the dienol form in dichloromethane solution. The
solution is kept under air, and after twelve days at room tempera-
ture 1H NMR analysis indicates that auto-oxidation is complete.
Bicyclic peroxide 4 is obtained in 68% yield after purification by
silica gel column chromatography. The peroxide is then methylated
Scheme 1 Synthesis of peroxo compound 6. Reagents and conditions: (a) SO3–Py, Et3N, DMSO; (b) syncarpic acid, piperidine; (c) NH4Cl, HCl; (d) O2;
(e) BuLi, TfOMe; (f) TFA, CH2Cl2.
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Scheme 2 Synthesis of hybrid peroxo-streptocyanine dyes 10a, 10b–12b. 8a, 8b and 13a were synthesised following previously described procedures.5
Reagents and conditions: argon, RT (a) CH3CN, iPr2NEt 2 eq., 6 2 eq., 10 d, (b) CH3CN, 6 1 eq., 24 h; (c) Et2NH or morpholine 1 eq., 24 h; (d) CH3CN,
Et2NH 1 eq., 15 min; (e) DMF, iPr2NEt 1 eq., 6 1 eq., 30 d.
Scheme 3 Synthesis of unsymmetrical hybrid 4-aminoquinoline–streptocyanine dyes 15b, 16b, 18b and 19b. Reagents and conditions: RT (a)
CH3CN/DMF, 7a 1 eq., 24 h; (b) CH3CN, 7b 1 eq., 24 h.
values between those of single and double bonds, expressing the
delocalisation of the positive charge of the 5C carbon chain.
(Vero cells), as described.16 These two cell lines were chosen for in
vitro culture ease and because one is a transformed cancerous cell
line (McF7) while the other (Vero) is not of cancerous origin, this
allowed comparisons of the two origins in the selectivity index. A
resistance index was also calculated as the ratio IC50 Fcb1/IC50 Nigerian
or IC50 FcM29/IC50 Nigerian. This ratio is 4.4 for chloroquine with the
FcB1 strain (K76T mutation at the PfCRT locus) and 8.7 for the
FcM29 strain (K76T mutation at the PfCRT locus, resistance-
associated mutations on the Pf-MDR gene) which has IC50
constantly higher than FcB1.17
All hybrid molecules exhibit good in vitro antiplasmodial activ-
ities (0.06 to 0.66 mM) similar to those of regular streptocyanines.
Moreover, they are not toxic or are slightly cytotoxic, except 10a.
Hemicarboxonium salts 13b and 14a are neither active nor toxic.
They are characterised by a less conjugated polymethine chain
Biological activity
All the biological results are summarized in Tables 1 and 2. The
antimalarial activity of the hybrid streptocyanines was evaluated
in vitro against P. falciparum on a chloroquine-sensitive (Nigerian)
strain13 and on chloroquine resistant strains (FcB1-Colombia and
FcM29),14 according to the procedures described by Desjardins
and co-workers.15 The antiplasmodial specificity of the drugs
was evaluated by comparing antiplasmodial activities toward
the mammalian MCF7 and Vero cell lines versus parasite (IC50
cells)/(IC50 P. falciparum). Two cell lines were used, the human
breast cancer cell line MCF7 and a normal cell line of simian origin
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Table 1 In vitro activity of the synthesised compounds against chloroquine-sensitive (Nigerian) and chloroquine-resistant (FcB1, FcM29) strains of
Plasmodium falciparum
IC50 P. falciparum (mM)
Resistance index
IC50 Fcb1/IC50 Nigerian
Compound
NR1R2
NR3R4
R
Nigeriana
FcB1b
FcM29b
IC50 FcM29/IC50 Nigerian
10a
10b
11b
NEt2
NEt2
peroxo
peroxo
peroxo
Me
F
F
—
0.10
0.305
0.057
0.436
0.427
0.039
0.422
0.374
—
4.27
1.40
—
4.14
1.23
12b
13b
14a
peroxo
NEt2
peroxo
OEt
OEt
F
F
Me
0.645
4.15
23.5
0.294
4.921
49.642
0.366
6.451
58.275
0.46
1.19
2.11
0.57
1.55
2.48
15b
16b
NEt2
quino
quino
F
F
0.15
0.63
0.695
0.526
—
3.942
4.60
0.83
—
6.21
17b
18b
19b
quino
NEt2
quino
pipequino
pipequino
F
F
F
0.66
0.13
0.235
1.152
0.470
0.550
1.459
0.281
0.892
1.75
3.59
2.34
2.21
2.14
3.79
20b
pipequino
pipequino
F
F
0.35
0.73
0.444
5.466
0.468
3.221
1.27
7.49
1.34
4.41
21b4
22a4,5
NEt2
NEt2
—
NEt2
NEt2
Me
F
—
—
0.13
0.15
0.008
0.03
0.420
0.248
—
0.085
0.465
0.004
0.26
3.36
1.65
—
0.68
3.10
0.5
22b
Artemisinin20
chloroquine
—
0.132
4.40
8.67
a Values reflect the mean of two experiments carried out in duplicate. b Values reflect the mean of three independent experiments, as SD was constantly
lower than 15% they were not indicated in the table.
compared to streptocyanines, with a positive global charge located
to a greater extent on the nitrogen end of the molecule than the
oxygen end. The difference in activities between hemicarboxonium
salts and streptocyanines is in agreement with the p-delocalized
lipophilic cation (DLC) hypothesis.18
Hybrid peroxo-streptocyanine dyes are ten- to one hundred-
fold more active than the cyclic peroxide 6 (IC50 5.7 mM) which is
too hydrophilic. The molecules with para-fluorophenyl groups are
less cytotoxic than the para-methylated compound 10a. The latter,
in spite of its good antiplasmodial activity (62 nM), is 47-fold
more cytotoxic toward MCF7 cells than its fluoro homologue
10b, so it has the worst selectivity index among this hybrids
series. Interestingly, unsymmetrical compounds 10a, 10b, and
11b showed increasing activity with regard to the corresponding
symmetrical streptocyanines 22a, 22b, 21b, or to the hybrid
cyanine dyes 17b and 20b. Reagents and conditions: argon, RT (a) DMF,
Scheme 4 Synthesis of symmetrical hybrid 4-aminoquinoline–strepto-
molecule 12b. The selectivity index of hybrid dyes 11b and 12b was
lower than the selectivity index of the two morpholino end groups
streptocyanine 21b. In contrast, the association of diethylamino
7a 2 eq., 6 d; (b) CH3CN, 7b 2 eq., 24 h.
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Table 2 In vitro activity of the synthesised compounds against the mammalian cells, MCF7 and Vero
IC50 (mM)
Selective toxicity
IC50 MCF7
/
IC50 Vero
IC50 Nigerian
/
IC50 MCF7
IC50 Fcb1
/
IC50 Vero
IC50 Fcb1
/
IC50 MCF7
IC50 FcM29
/
IC50 Vero/
IC50 FcM29
Compound
MCF7a
Veroa
IC50 Nigerian
10a
0.095
4.5
12
2.8
46
24
36
24
—
44
42
35
2
0.3
33
8
62
38
114
27
92
5
—
454
79
55
6
2
35
9
62
290
196
117
190
9
1.7
0.7
30
76
2
0.1
7
10
36
11
49
21
12
1
48
7
56
121
5
1
8
11
36
81
84
92
25
3
2.5
0.8
34
61
1
0.8
—
1
28
18
30
20
21
7
71
8
64
97
4
1
—
1
28
135
52
87
43
13
23
165
10b
11b
12b
22
13b
7.4
7.5
5.1
5.3
41
5.0
26.8
9.5
67
14a
50
15b
5.2
5.7
41
38
46
16b
17b
18b
19b
20b
41
21b4
22a4,5
22b
139
1.07
11
0.62
0.43
>10
3
—
72
1433
2
—
43
326
1
—
chloroquine
43
a Values reflect the mean of three experiments, SD was consistently < 15% (thus omitted for clarity); doxorubicin (positive control) IC50: 55 nM for MCF7
and 120 nM for Vero.
Fig. 5 ORTEP view of 18b. Thermal ellipsoids are shown at the 50%
probability level. Hydrogen atoms, tetrafluoroborate anion and solvent
◦
˚
molecules are omitted for clarity. Selected bond lengths (A) and angles ( ):
N1–C1 1.335(3), C1–C2 1.398(3), C2–C3 1.391(3), C3–C4 1.382(3), C4–C5
1.401(3), C5–N2 1.335(3), N1–C1–C2 122.9(2), C1–C2–C3 121.9(2),
C2–C3–C4 125.2(2), C3–C4–C5 121.1(2), C4–C5–N2 123.7(2); deviation
of the chain carbons from the mean plane: 2.68 pm.
Fig. 4 ORTEP view of 17b. Thermal ellipsoids are shown at the 50%
probability level. Hydrogen atoms, tetrafluoroborate anion and solvent
˚
molecules are omitted for clarity. Selected bond lengths (A) and angles
(◦): N1–C1 1.3389(19), C1–C2 1.398(2), C2–C3 1.392(2), C3–C4 1.389(2),
C4–C5 1.401(2), C5–N2 1.3382(19), N1–C1–C2 122.39(14), C1–C2–C3
124.53(14), C2–C3–C4 121.55(14), C3–C4–C5 125.86(14), C4–C5–N2
120.63(13); deviation of the chain carbons from the mean plane: 7.6 pm.
The antiplasmodial activities of the hybrid 4-aminoquinoline–
streptocyanine dyes range from 0.13 to 0.66 mM, i.e., between the
streptocyanines 22b and 21b activities. These values are close to
those obtained with other promising antimalarial compounds. In
the regular 5C-, 7C-, 9C-streptocyanines series,4 compounds with
secondary amino groups at the end of the polymethine chain have
shown a better activity than molecules with primary amino groups.
In the same way, pipequino hybrid dyes are more active than the
ones with quino moieties (20b/17b IC50 0.35/0.66 mM, 19b/16b
0.23/0.63 mM). Moreover, their cytotoxicities are lower, except for
20b, leading to better selective toxicity profiles. Hybrid molecules
with quino moieties, 17b and 16b, have similar activities to the
one with two morpholino end groups, 21b, but are more cytotoxic
against MCF7 as well as Vero cells. Although hybrid molecules
with pipequino moieties, 20b and 19b, are respectively 2- and
and peroxo groups has been beneficial because the hybrid molecule
10b has retained the good antiplasmodial activity of 22b with less
cytotoxicity. So, 10b exhibits high selectivity indices of 44 and 454,
respectively, to MCF7 and Vero cells. For most of the tested drugs
(except 10b, 15b, and 21b), the resistance ratio was lower than 4
or 8 for FcB1 or FcM29, respectively. This indicated the absence
of a chloroquine-like resistance pathway for these molecules. The
resistance index of 12b is less than one, which is a characteristic
of artemisinin-like compounds.19,20 It seems that 12b present an
artemisinin-like action mode, whereas the peroxo moiety of 10b,
and to a lesser extent of 11b, doesn’t seem to be responsible for the
observed antiplasmodial activity.
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on Bruker AC 200, Bruker AC 250, AM 300 WB, AM 400 WB
or AVANCE 500 spectrometers. Chemical shifts are given in ppm
from TMS. Mass spectra were obtained on a Perkin-Elmer SCIEX
API 365 apparatus with electrospray (positive mode, CH3CN), a
Nermag R10–10 apparatus for DCI and a GC TOF Waters and
Waters Q/TOF Ultima apparatus for high resolution electrospray.
Elemental analysis was performed by the microanalysis service
from the Laboratoire de Chimie de Coordination (LCC) in
Toulouse.
Preparation of tetrafluoroborate carboxonium salts 8a, b
Fig. 6 ORTEP view of 19b. Thermal ellipsoids are shown at the 50%
These compounds were synthesised following previously described
procedures.5
probability level. Hydrogen atoms, tetrafluoroborate anion and solvent
◦
˚
molecules are omitted for clarity. Selected bond lengths (A) and angles ( ):
N1–C1 1.337(3), C1–C2 1.415(3), C2–C3 1.388(3), C3–C4 1.402(3), C4–C5
1.398(3), C5–N2 1.350(3), N1–C1–C2 123.3(2), C1–C2–C3 121.3(2),
C2–C3–C4 124.9(2), C3–C4–C5 121.3(2), C4–C5–N2 123.1(2); deviation
of the chain carbons from the mean plane: 3.68 pm.
Preparation of tetrafluoroborate hemicarboxonium salts 13a, b and
14a–, b
These compounds were synthesised following previously described
procedures.5 One equivalent of amine (morpholine, diethylamine)
was added dropwise onto carboxonium salt in solution in dry
acetonitrile at room temperature, under argon atmosphere. After
15 min stirring, the solvent was removed under reduced pressure
and the solid was washed with pentane and dried. The crude
streptocyanine was crystallized in 100% ethanol and dried under
vacuum at 40 ◦C. Only characterizations for new compounds are
described below.
3-fold more active than 21b, their selectivity indices are of the same
order for 19b and weaker for 20b. Symmetrical hybrid compounds
17b and 20b are respectively 4- and 2-fold less active than 22b,
but absence of cytotoxicity on MCF7 cells gives them selectivity
indices 20- and 10-fold higher. Against Vero cells, only 20b shows a
better selectivity (117). As observed in peroxo-streptocyanine dyes,
unsymmetrical hybrid compounds with diethylamino groups, 15b
and 18b, are more interesting than the corresponding symmetrical
dyes 22b because of the decrease in MCF7 cells cytotoxicity
afforded by the 4-aminoquinoline moiety. Against Vero cells, 15b
is twice as cytotoxic as 22b, but 18b is four-fold less cytotoxic and
this compound shows the best selectivity index (289) of the series
of 4-aminoquinoline–streptocyanines synthesised.
1-Ethoxy-5-diethylamino-1,5-bis(4-fluorophenyl)-penta-1,3-di-
enylium tetrafluoroborate 13b. 13b was obtained as a yellow
powder (1.65 g, 69%). (Found: C, 60.26, H, 6.02, N, 3.09.
◦
C23H26BF6NO requires C, 60.41, H, 5.73, N, 3.06%); mp 146 C
3
(dec); dH(300 MHz; CD3CN) 1.20 (3H, t, J 7.2, NCH2CH3),
3
3
1.44 (3H, t, J 7.1, OCH2CH3), 1.48 (3H, t, J 7.2, NCH2CH3),
Conclusion
3.48 (2H, q, 3J 7.2, NCH2CH3), 3.95 (2H, q, 3J 7.2, NCH2CH3),
3
We have designed and synthesised two novel series of strepto-
cyanine dyes incorporating cyclic peroxide or 4-aminoquinoline
moieties. The resulting hybrid compounds take advantage of
the preservation of the good antiplasmodial activities of the
streptocyanines while decreasing their cytotoxicity. If two active
moieties are introduced, the corresponding symmetrical hybrid
streptocyanines (12b, 17b, 20b) have submicromolar activities
but stay less selective than the two morpholino end group
streptocyanine (21b). The latter was found to be the most selective
compound (92 and 189 to MCF7 and Vero cells, respectively) in
previous RSA studies. However, with our synthetic method, we
can easily obtain unsymmetrical compounds to act separately on
activity and cytotoxicity. So, the unsymmetrical hybrids, which
combine the active but toxic diethylamino group with peroxide
or aminoquinoline moieties, have better selectivity index (44 and
454 for 10b, 38 and 289 for 18b against MCF7 and Vero cells,
respectively).
4.22 (2H, q, J 7.2, OCH2CH3), 6.23 (1H, part X of ABX syst.,
3J 11.7, H2), 6.65 (1H, part B of ABX syst., J 11.7 and 14.1,
3
H3), 6.84 (1H, part A of ABX syst., 3JAB 14.1, H4) and 7.00–7.30
(8H, m, Harom); dC(75 MHz; CD3CN) 12.3, 12.9 (NCH2CH3), 13.5
(OCH2CH3), 47.3, 50.8 (NCH2CH3), 66.6 (OCH2CH3), 103.6 (C2),
2
115.3, 116.2 (2d, JCF 22.2, CHarom), 116.5 (C4), 127.2, 129.9 (2d,
4JCF 3.3; Carom–C1–5), 130.4, 131.8 (2d, 3JCF 8.9, CHarom), 160.6 (C3),
163.8, 164.0 (2d, 1JCF 248.3, F–Carom), 173.1 and 174.7 (C1–5); m/z
(ESI+; CH3CN) 370.3 (M+, 100%).
1-Ethoxy-5-morpholino-1,5-bis(4-methylphenyl)-penta-1,3-di-
enylium tetrafluoroborate 14a. 14a was obtained as a yellow
powder (1.50 g, 70%). (Found: C, 64.93, H, 6.66, N, 3.00.
C25H30BF4NO2 requires C, 64.81, H, 6.53, N, 3.02%); mp 192 ◦C;
3
dH(300 MHz; CD3CN) 1.44 (3H, t, J 7.1, OCH2CH3), 2.37 et
2.42 (6H, 2 s, CH3–C6H4), 3.62 (2H, m, NCH2CH2), 3.77 (2H, m,
NCH2CH2), 3.99 (2H, m, OCH2CH2), 4.10 (2H, m, OCH2CH2),
4.24 (2H, q, 3J 7.1, OCH2CH3), 6.23 (1H, m, H3), 6.91 (2H,
m, H2–4), 7.16–7.43 (8H, m, Harom) and dC(75 MHz; CD3CN)
13.5 (OCH2CH3), 20.4 (CH3–Carom), 51.1, 53.8 (NCH2CH2), 65.8,
66.4 (OCH2CH2), 66.5 (OCH2CH3), 103.4, 115.5 (C2–4), 127.9,
130.6 (Carom–C1–5), 128.6, 128.9, 129.5, 129.8 (CHarom), 141.8, 142.2
(Carom–CH3), 162.0 (C3), and 175.0, 175.5 (C1–5); m/z (ESI+;
CH3CN) 375.5 (M+, 100%).
Experimental
All experiments were performed under dry conditions (argon
atmosphere, anhydrous solvents) to avoid degradation of the
carboxonium salt. Melting points were determined with a Bu¨chi
capillary apparatus. 1H NMR and 13C NMR spectra were recorded
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1-Ethoxy-5-morpholino-1,5-bis(4-fluorophenyl)-penta-1,3-di-
enylium tetrafluoroborate 14b. 14b was obtained as a yellow
powder (1.60 g, 70%). (Found: C, 58.61, H, 5.44, N, 2.91.
C23H24BF6NO2 requires C, 58.62, H, 5.13, N, 2.97%); mp 221 ◦C
79.7 (C), 154.6 (C O) and 202.9 (CH O); m/z (ESI+, MeOH)
236 ([MNa]+, 100%).
4-(3,5,6,7,8,8a-Hexahydro-8a-hydroxy-6,6,8,8-tetramethyl-5,7-
dioxobenzo[c][1,2]dioxin-3-yl)piperidine-1-tert-butyl carboxylate 4.
Under argon, at room temperature, aldehyde 1 (0.38 mg, 1.8 mmol,
1 eq.) dissolved in anhydrous dichloromethane (9 mL) is added to
piperidine (0.09 mL, 0.9 mmol, 0.5 eq.). Piperidine (0.09 mL,
0.9 mmol, 0.5 eq.) is added to a suspension of syncarpic acid
(0.32 g, 1.8 mmol, 1 eq.) in dichloromethane (9 mL). After 45 min,
the solution of syncarpic acid is poured into the solution of
iminium. After 30 min stirring, the mixture is concentrated under
vacuum. Mannich base 2 is obtained as a white powder. The
Mannich base is then solubilised in dichloromethane and treated
with a saturated solution of NH4Cl in HCl 1 M. The biphasic
mixture is stirred for 15 min; the organic phase is recovered, dried
on MgSO4, filtered and concentrated. Enone 3 is obtained as
a yellow oil. It is then dissolved in dichloromethane, and kept
under air for thirteen days. After evaporation, the crude mixture
is purified by silica gel column chromatography using petroleum
ether/ethyl acetate: 8/2 as eluent. Endo-peroxide 4 (0.49◦g, 1.2
mmol) is obtained in 68% yield as a white solid. mp 152 C; Rf
(PE/AcOEt) = 0.46; dH(300 MHz; CDCl3) 1.03 (s, 3H, CH3 K/L),
1.32 (s, 3H, CH3 K/L), 1.35 (s, 3H, CH3M/N), 1.37 (s, 3H, CH3M/N),
1.45 (s, 9H, 3CH3), 1.61–2.04 (m, 4H, ABX syst., CH2O), 3.00–
4.10 (m, 5H, ABX syst., CH2P and OH) and 7.06 (s, 1H, CHE);
dC(75 MHz; CDCl3) 15.1–21.0 (CH3 K/L), 24.1–26.6 (CH3M/N), 28.4
(C(CH3)3), 30.5–32.5 (CH2O), 38.7 (CH2P), 51.7 (CJ), 55.0 (CH),
78.7 (CD), 80.1 (C(CH3)3), 97.7 (CA), 132.8 (CF), 140.9 (CHE),
154.5 (NC O), 198.1 (CG) and 210.3 (CI); IR (KBr, n): 3394 (OH),
2978 to 2877 (CH2 and CH3), 1731 (C O), 1691 (a,b-unsaturated
3
(dec); dH(250 MHz; CD3CN) 1.45 (3H, t, J 7.0, OCH2CH3),
3.60 (2H, m, NCH2CH2), 3.77 (2H, m, NCH2CH2), 4.00 (2H,
3
m, OCH2CH2), 4.11 (2H, m, OCH2CH2), 4.25 (2H, q, J 7.0,
OCH2CH3), 6.26 (1H, part X of ABX syst., 3J 11.4, H2), 6.83 (1H,
3
part B of ABX syst., J 11.4 and 14.1, H3), 6.95 (1H, part A of
ABX syst., 3J 14.1, H4), 7.13–7.41 (8H, m, Harom) and dC(75 MHz;
CD3CN) 13.5 (OCH2CH3), 51.3, 54.0 (NCH2CH2), 65.8, 66.3
(OCH2CH2), 66.7 (OCH2CH3), 103.8 (C2), 115.3, 116.5 (2d, 2JCF
22.2, CHarom), 116.1 (C4), 126.9, 129.9 (C1–5-Carom), 131.3, 131.9 (2d,
3JCF 9.0, CHarom), 161.4 (C3), 164.1, 164.2 (2d, 1JCF 248.8, F–Carom
and 173.6, 174.3 (C1–5); m/z (ESI+; CH3CN) 384.4 (M+, 100%).
)
Preparation of tetrafluoroborate streptocyanine dyes 21a, b and
22a, b
These compounds were synthesised following previously described
procedures.4 Two equivalents of amine (morpholine, diethylamine)
were added dropwise onto carboxonium salt in solution in dry
acetonitrile at room temperature, under argon atmosphere. After
stirring for 24 h, the solvent was removed under reduced pressure
and the solid was washed with pentane and dried. The crude
streptocyanine was crystallized from 100% ethanol and dried
under vacuum at 40 ◦C. Only characterization for the new
compound is described below.
1,5-Bis(diethylamino)-1,5-bis(4-fluorophenyl)-penta-1,3-di-
enylium tetrafluoroborate 22b. 22b was obtained as a yellow
powder (1.60 g, 50%). (Found: C, 61.67, H, 6.92, N, 5.55.
C
O), 1669 (NC O), 1636 (C C), 1276 (C–N) and 1099 (C–O
◦
C25H31BF6N2 requires C, 62.00, H, 6.45, N, 5.78%); mp 258 C;
peroxide); HRMS (DCI/CH4, CH2Cl2, negative mode) 409.2101
(M+. C21H31NO7 requires 409.2101).
dH(300 MHz; CD3CN) 1.05 (6H, t, 3J 7.2, NCH2CH3), 1.36 (6H, t,
3J 7.2, NCH2CH3), 3.21 (4H, q, 3J 7.2, NCH2CH3), 3.67 (4H, q, 3J
7.2, NCH2CH3), 5.87 (1H, part X of AXX¢ syst., 3J 13.8 and 12.0,
H3), 6.18 (2H, part AA¢ of AA¢X syst., 3J 13.0, H2–4) and 7.06–7.11
(8H, m, Harom); dC(75 MHz; CD3CN) 11.6, 13.2 (NCH2CH3), 44.9,
48.1 (NCH2CH3), 105.2 (C2–4), 115.5 (d, 2JCF 22.4, CHarom), 129.1
(d, 4JCF 3.5, C–C1–5), 130.5 (d, 3JCF 8.9, CHarom), 161.4 (C3), 163.1
4-(3,5,6,7,8,8a-Hexahydro-8a-methoxy-6,6,8,8-tetramethyl-5,7-
dioxobenzo[c][1,2]dioxin-3-yl)piperidine-1-tert-butyl carboxylate 5.
Under argon, endo-peroxide 4 (1.25 g, 3.0 mmol, 1 eq.) is dissolved
in anhydrous tetrahydrofuran (100 mL). At -78 ◦C, butyllithium
solution (1.3 M in hexane) (2.10 mL, 3.3 mmol, 1.1 eq.) is slowly
added. After 15 min stirring, methyl triflate (0.38 mL, 3.3 mmol,
1.1 eq.) is added. The mixture is stirred for 4 h at -78 ◦C and then
hydrolysed with saturated NH4Cl solution. The aqueous phase
is extracted with dichloromethane, the organic phases gathered,
washed, dried over MgSO4, filtered and evaporated. Methylated
endo-peroxide 5 is obtained after purification by silica gel column
chromatography (PE/AcOEt 9/1) as a white solid (0.80 g, 1.9
mmol) in 65% yield. Rf (PE/AcOEt: 8/2) = 0.39; dH(300 MHz;
C6D6) 0.71 (s, 3H, CH3 K/L), 1.08 (m, ABX syst., 1H, CH2O), 1.17–
1.31 (m, ABX syst., 2H, CH2O), 1.34 (s, 3H, CH3M/N), 1.39 (s, 3H,
CH3M/N), 1.47 (s, 9H, 3CH3), 1.49 (s, 3H, CH3 K/L), 1.59 (m, ABX
syst., 1H, CH2O), 2.87 (m, ABX syst., 1H, CH2P), 3.10 (m, ABX
syst., 1H, CH2P), 3.50–4.14 (m, ABX syst., 2H, CH2P) and 6.93
(s, 1H, CHE); dC(75 MHz; C6D6) 16.1–21.6 (CH3 K/L), 24.9–26.2
(CH3M/N), 28.4 (CH3), 30.4–32.3 (CH2O), 39.2 (CH2P), 53.2 (CJ),
54.5 (OCH3), 54.8 (CH), 77.9 (CD), 79.4 (C(CH3)3), 101.1 (CA),
129.7 (CF), 143.4 (CHE), 154.4 (NC–O), 197.8 (CG) and 209.1 (CI);
IR (KBr, n): 2973 to 2840 (CH2 and CH3), 1726 (C–O), 1692 (a,b-
unsaturated C O and carbamoyl), 1634 (C C), 1279 (C–N) and
1102 (C–O peroxide); m/z (ESI+, CH2Cl2/MeOH) 446 ([MNa]+,
1
(d, JCF 242.8, F–Carom) and 168.3 (C1–5); m/z (ESI+; CH3CN)
397.3 (M+, 100%).
Preparation of hybrid molecules with peroxide moiety 10a, b, 11b
and 12b
4-Formylpiperidine-1-tert-butyl carboxylate 1. Under argon,
1-BOC-4-hydroxymethylpiperidine (0.40 g, 1.9 mmol, 1eq.) is
dissolved in anhydrous dichloromethane (1.6 mL mmol-1). DMSO
(2 mL mmol-1), triethylamine (1.29 mL, 9.3 mmol, 5 eq.) and SO3–
pyridine complex (1.48 g, 9.3 mmol, 5 eq.) are successively added.
After 30 min stirring at room temperature, the mixture is diluted
in ethyl ether then treated with a saturated NaCl solution. The
organic phase is dried on magnesium, filtered then concentrated.
Aldehyde 1 (0.40 mg, 1.9 mmol) is obtained as a yellow oil in a
quantitative yield. dH(300 MHz; CDCl3) 1.44 (s, 9H, 3CH3), 1.55
(m, 2H, ABX syst., CH2–CHCHO), 1.87 (m, 2H, ABX syst., CH2–
CHCHO), 2.39 (m, 1H, CHCHO), 2.91 (m, 2H, ABX syst., CH2–
N) and 3.99 (m, 2H, ABX syst., CH2–N); dC(75 MHz; CD3Cl3)
25.1 (CH2–CHCHO), 28.4 (CH3), 42.8 (CH2–N), 48.0 (CHCHO),
7406 | Org. Biomol. Chem., 2011, 9, 7400–7410
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100%); HRMS (ESI+, MeOH) 446.2155 ([MNa]+. C22H33NO7Na
requires 446.2192).
was then added to the mixture (0.04 cm3, 0.20 mmol). After
24 h stirring, diethylamine (0.03 cm3, 0.20 mmol) was added to
the mixture. 24 h later, the solvent is removed under reduced
pressure. Water is added and extraction with dichloromethane
performed, followed by washing the organic phases, drying on
MgSO4 and evaporation to furnish a solid which was precipitated
in ethyl acetate/petroleum ether (4/1). 10b was obtained as an
orange powder (0.04 g, 28%). dH(300 MHz; CDCl3) 0.90–1.17
3-Piperidin-4-yl-8,8a-dihydro-8a-methoxy-6,6,8,8-tetramethyl-
benzo[c][1,2]dioxine 5,7(3H,6H)-dione 6. Under argon at room
temperature, trifluoroacetic acid (0.66 mL, 8.8 mmol, 20 eq.) is
added to endo-peroxide 5 (0.19 g, 0.4 mmol, 1 eq.) dissolved in
anhydrous dichloromethane (22 mL). After 24 h, the mixture is
neutralized with saturated NaHCO3 solution. The aqueous phase
is extracted with dichloromethane, the organic phases gathered,
washed, dried over MgSO4, filtered and evaporated. Amine 6
(0.13 g, 0.4 mmol) is obtained as a yellow solid in 92% yield. mp
-1
(6H, CH3 K L + N–CH2–CH3), 1.20–1.40 (9H, CH3 K/L/M/N), 1.44
(3H, t, 3J 6.3, N–CH2–CH3), 1.50–2.60 (4H, m, CH2O), 3.20–4.50
3
(11H, CH2P + N–CH2–CH3 + OCH3), 5.99 (1H, t, J 12.9, H3),
6.50, 6.56 (2H, 2d, 3J 12.9, H2–4), 6.95–7.25 (8H, m, Harom) and 7.31
(1H, s, CHE); dC(75 MHz; CDCl3) 12.5, 14.2 (N–CH2–CH3), 15.6,
21.7, 24.8, 24.9, 25.9 (CK/L/M/N), 31.2, 32.2 (CO), 43.8, 46.6 (CP),
45.6, 48.3 (N–CH2–CH3), 53.1 (CJ ), 54.8 (CH), 55.1 (O–CH3), 77.6
◦
55 C; Rf (CH2Cl2/MeOH saturated with NH4OH 19/1) = 0.18;
dH(300 MHz; C6D6) 0.75 (s, 3H, CH3 K/L), 1.10–1.39 (m, ABX
syst., 2H, CH2O), 1.37 (s, 3H, CH3M/N), 1.42 (s, 3H, CH3M/N), 1.53
(s, 3H, CH3 K/L), 1.40–1.71 (m, ABX syst., 2H, CH2O), 2.38–2.83
(m, 4H, ABX syst., CH2P), 3.21 (s, 3H, OCH3) and 7.14 (s, 1H,
CHE); dC(75 MHz; C6D6) 16.1–21.7 (CH3 K/L), 25.0, 26.2 (CH3M/N),
31.4, 32.8 (CH2O), 41.2, 41.3 (CH2P), 53.2 (CJ), 54.5 (OCH3), 54.8
(CH), 78.1 (CD), 101.1 (CA), 129.5 (CF), 143.9 (CHE), 198.0 (CG)
and 209.3 (CI); IR (KBr, n): 2973 to 2873 (CH2 and CH3) and 1718
(C O), 1690 (a,b-unsaturated C O), 1636 (C C), 1278 (C–N),
1102 (C–O peroxide); m/z (DCI/CH4, CH2Cl2, positive mode):
324 ([M + 1]+, 100%); HRMS (DCI/CH4, CH2Cl2, positive mode)
324.1811 ([M + 1]+. C17H26NO5 requires 324.1806).
2
(CD), 100.9 (CA), 107.2, 108.5 (C2–4), 115.8, 115.9 (2d, JCF 21.7,
4
CHarom), 128.6, 128.8 (2d, JCF 3.4, C1–5-Carom), 130.0 (CF), 130.2
1
(CHarom), 142.4 (CE), 162.7 (C3), 163.5, 164.9 (2d, JCF 250.7, F–
Carom), 167.2, 168.8 (C1–5), 198.2 (CG) and 210.0 (CI); dF(300 MHz;
CD3CN) -152.7, -152.6 (BF4), -109.7, -109.2 (Farom); HRMS
(ESI+, MeOH) 647.3287 (M+. C38H45N2O5F2 requires 647.3297).
1,5-Bis(4-fluorophenyl)-1-morpholino-5-3-piperidin-4-yl-8,8a-
dihydro-8a-methoxy-6,6,8,8-tetramethylbenzo-[c][1,2]-dioxine-5,7-
(3H,6H)-dione-penta-1,3-dienylium tetrafluoroborate 11b. A so-
lution of 6 (0.07 g, 0.20 mmol) in dry DMF (10 cm3) was added
dropwise to a solution of 8b (0.09 g, 0.20 mmol) in dry DMF
(40 cm3) at room temperature. After 6 h stirring, morpholine
(0.02 cm3, 0.20 mmol) was added to the mixture. 24 h later, the
solvent was removed under reduced pressure, and the crude solid
was purified by silica gel chromatography (dichloromethane/ethyl
acetate/acetonitrile 4/1/2). 11b was obtained as an orange powder
(0.07 g, 48%). TLC Rf (CH2Cl2/AcOEt/CH3CN 4/1/2) 0.31;
1,5-Bis(4-methylphenyl)-1-diethylamino-5-3-piperidin-4-yl-8,8a-
dihydro-8a-methoxy-6,6,8,8-tetramethylbenzo[c][1,2]-dioxine-5,7-
(3H,6H)-dione-penta-1,3-dienylium tetrafluoroborate 10a.
6
(0.04 g, 0.13 mmol) was added to 13a (0.05 g, 0.13 mmol) in DMF
(5 cm3) at room temperature. Diisopropylethylamine (0.02 cm3,
0.13 mmol) was then added to the mixture. After 30 days stirring,
the solvent was removed at reduced pressure. Water was added
and extraction with dichloromethane performed, followed by
washing the organic phases, drying on MgSO4 and evaporation
to furnish a solid. This was washed with pentane and precipitated
in ethyl acetate/petroleum ether (4/1) to give 10a as an orange
powder (0.03 g, 32%).
-1
dH(500 MHz; CDCl3) 0.94, 1.03 (3H, 2 s, CH3 K L ), 1.12–1.38
(9H, massif, CH3 K/L/M/N), 1.60–2.50 (4H, m, CH2O), 3.30–4.50
(15H, 2 N–CH2 + 2 O–CH2 + OCH3 + CH2P), 6.15 (1H, part
A of AMX syst., 3JAX 12.5 and 3JAM 13.0, H3), 6.65 (1H, part M of
AMX syst., 3JAM 13.0, H2 or H4), 6.71 (1H, part X of AMX syst.,
3JAX 12.5, H2 or H4), 7.00–7.25 (8H, m, Harom) and 7.28 (1H, s,
CHE); dC(125 MHz; CDCl3) 15.6, 21.6, 21.9, 24.8, 25.9 (CK/L/M/N),
30.3, 32.3 (CO), 44.2, 46.8 (CP), 49.1, 51.5 (N–CH2), 53.1 (CJ ),
54.8 (CH), 55.1 (O–CH3), 66.5, 66.9 (O–CH2), 77.5 (CD), 101.0
(CA), 108.4, 108.7 (C2–4), 116.1 (d, 2JCF 21.7, CHarom), 128.2, 128.5
dH(300 MHz; CDCl3) 0.90–1.15 (6H, CH3 K/L + N–CH2–CH3),
3
1.23–1.38 (9H, CH3 K/L/M/N), 1.44 (3H, t, J 6.6, N–CH2–CH3),
1.70–2.30 (4H, m, CH2O), 2.35 (6H, s, CH3–C6H4), 3.23 (2H, q,
3J 6.6, N–CH2–CH3), 3.30–4.50 (9H, m, N–CH2–CH3 + OCH3 +
CH2P), 6.09 (1H, t, 3J 12.6, H3), 6.55, 6.60 (2H, 2d, 3J 12.6, H2–4),
6.70–7.15 (8H, m, Harom) and 7.25 (1H, s, CHE); dC(125 MHz;
CDCl3) 11.7, 13.2 (N–CH2–CH3), 15.1, 21.2, 21.3, 24.3, 25.3
(CH3), 20.4 (2 s, CH3–C6H4), 29.6, 30.6, 31.3, 32.3 (CO), 43.3, 43.5,
46.3, 46.4 (CP), 45.0, 46.4 (N–CH2–CH3), 52.8 (CJ), 54.4 (CH), 54.6
(O–CH3), 77.9 (CD), 100.9 (CA), 104.7, 106.3 (C2–4), 128.0–129.1
(massif, CHarom), 129.2, 129.8 (C1–5–Carom), 129.9 (CF), 139.9, 140.4
(CH3–Carom), 143.1 (CE), 162.5 (C3), 168.8, 170.3 (C1–5), 198.0 (CG)
and 210.0 (CI); dF(300 MHz; CDCl3) -152.99, -153.05 (BF4); m/z
(ESI+; MeOH) 639.6 (M+, 100%); HRMS (DCI/NH3, MeOH,
positive mode) 639.3768 (M+. C40H51N2O5 requires 639.3798).
4
(2d, JCF 3.7, C1–5-Carom), 130.0 (CF), 130.6, 131.0 (m, CHarom),
142.1 (CE), 163.1 (C3), 163.6 (d, 1JCF 257.0, F–Carom), 168.1 (C1–5),
198.0 (CG) and 209.9 (CI); dF(300 MHz; CD3CN) –151.88, -151.83
(BF4), -111.9, -111.8 (Farom); HRMS (ESI+, MeCN) 661.3088
(M+. C38H43N2O6F2 requires 661.3089).
1,5-Bis(4-fluorophenyl)-1, 5-bis(3-piperidin-4-yl-8,8a-dihydro-8a-
methoxy-6,6,8,8-tetramethylbenzo-[c][1,2]-dioxine 5,7 (3H,6H)-
dione-penta-1,3-dienylium tetrafluoroborate 12b. A solution of 6
(0.14 g, 0.40 mmol) in dry CH3CN (10 cm3) was added dropwise to
a solution of 8b (0.09 g, 0.20 mmol) in dry CH3CN (40 cm3) at room
temperature. Diisopropylethylamine (0.08 cm3, 0.40 mmol) was
then added to the mixture. After 10 days at room temperature, the
solvent was removed under reduced pressure. Water was added and
extraction with dichloromethane performed, followed by washing
the organic phases, drying on MgSO4 and evaporation to furnish
a solid. This was washed with diethyl ether and precipitated in
1,5-Bis(4-fluorophenyl)-1-diethylamino-5-3-piperidin-4-yl-8,8a-
dihydro-8a-methoxy-6,6,8,8-tetramethylbenzo-[c][1,2]-dioxine-5,7-
(3H,6H)-dione-penta-1,3-dienylium tetrafluoroborate 10b. A so-
lution of 6 (0.07 g, 0.20 mmol) in dry CH3CN (5 cm3) was
added dropwise to a solution of 8b (0.09 g, 0.20 mmol) in dry
CH3CN (35 cm3) at room temperature. Diisopropylethylamine
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ethyl acetate/petroleum ether (4/1) to give 12b as a yellow powder
(0.026 g, 14%). dH(300 MHz; CDCl3) 0.90–1.50 (24H, CH3 K/L/M/N),
1.50–2.60 (8H, m, CH2O), 3.20–4.60 (14H, CH2P + OCH3), 6.13
(1H, part A of AX2 syst., 3JAX 12.6, H3), 6.68 (2H, part X of AX2
syst., 3JAX 12.6, H2–4) and 7.05 (10H, m, Harom + CHE); dC(125 MHz;
CDCl3) 15.6, 21.8, 24.9, 25.9 (CK/L/M/N), 26.9, 29.2, 31.3, 32.4
(CO), 39.8, 39.9, 44.2, 46.7 (CP), 53.1 (CJ ), 54.8 (CH), 55.0 (O–
CH3), 75.7 (CD), 101.0 (CA), 108.6 (C2–4), 116.1 (CHarom), 128.4
(C1–5-Carom), 130.4 (CF), 130.8 (m, CHarom), 142.0 (CE), 163.4 (C3),
(CG), 148.2 (CE), 151.1 (CC), 151.2 (CA), 161.4 (C3), 163.6, 164.2
(2d, JCF 249.1, F–Carom) and 167.9, 169.6 (C1–5); dF(300 MHz;
1
MeOD) -154.4, -154.3 (BF4), -111.4, -110.4 (Farom); m/z (ESI+;
MeOH) 559.5 (M+,100%), 561.3 (38 [M+2]+).
5-(7-Chloro-4-(piperazin-1-yl)quinoline)-1-diethylamino-1,5-
bis(4-fluorophenyl)penta-1,3-dienylium tetrafluoroborate 18b.
(Found: C, 60.54, H, 5.47, N, 8.26. C34H34BClF6N4·1H2O
◦
requires C, 60.33, H, 5.36, N, 8.28%); mp 206 C; dH(300 MHz;
3
3
CD3CN) 1.08 (3H, t, J 7.0, N–CH2–CH3), 1.38 (3H, t, J 7.0,
1
163.6 (d, JCF 251.4, F–Carom), 168.0 (C1–5), 198.1 (CG) and 209.7
N–CH2–CH3), 3.22–3.52 (8H, m, N–CH2–CH3, N–CH2–CH2),
(CI); dF(300 MHz; CD3CN) -151.7, -151.6 (BF4), -108.5 (Farom);
HRMS (ESI+, MeOH) 897.4136 (M+. C51H59N2O10 F2 requires
897.4138).
3
3.73 (2H, q, J 7.0, N–CH2–CH3), 4.03 (2H, m, N–CH2–CH2),
6.05 (1H, part A of AMX syst., 3JAM 12.9 and 3JAX 12.6, H3), 6.28
3
(1H, part M of AMX syst., JMA 12.9, H4 or H2), 6.30 (1H, part
3
3
X of AMX syst., JXA 12.6, H2 or H4), 6.98 (1H, d, J 4.8, HB),
Preparation of hybrid molecules with quinoline moiety 15b–20b
3
4
7.15 (8H, m, Harom), 7.53 (1H, dd, J 9.0, J 2.1, HH), 8.05 (1H,
4
3
3
d, J 2.1, HF), 8.09 (1H, d, J 9.0, HI) and 8.75 (1H, d, J 4.8,
HC); dC(75 MHz; CD3CN) 11.4, 12.8 (N–CH2–CH3), 44.9, 48.2
(N–CH2–CH3), 47.4, 50.2, 50.6, 51.4 (N–CH2–CH2), 104.6, 106.6
(C2–4), 109.3 (CB), 115.2, 115.4 (2d, 2JCF 22.2, CHarom), 121.3 (CJ ),
Compounds 7a and 7b were synthesised following previously
described procedures.11
1 equivalent of 7a in solution in DMF, or 7b in solution in
CH3CN (around 2 cm3), was added to 1 equivalent of 13b or 14b
in solution in CH3CN (around 30 cm3) at room temperature. After
24 h stirring, the solvent was removed under reduced pressure,
and the solid was washed with diethyl ether and dried. The crude
streptocyanine◦was crystallized from 100% MeOH and dried under
vacuum at 40 C. 15b was obtained as a yellow powder (0.03 g,
26%), 16b as an orange powder (0.12 g, 59%), 18b as yellow crystals
(0.04 g, 30%) and 19b as yellow crystals (0.14 g, 50%).
4
125.4, 125.8 (CI–H), 128.2 (CF), 128.4, 128.6 (2d, JCF 3.6, Carom),
3
130.1, 131.0 (d, JCF 8.7, CHarom), 134.1 (CG), 149.8 (CE), 151.9
(CC), 155.4 (CA), 161.7 (C3), 162.8, 163.1 (2d, 1JCF 247.0, F–Carom
)
and 167.4, 169.1 (C1–5); dF(300 MHz; CD3CN) -151.7, -151.6
(BF4), -112.5, -112.1 (Farom); m/z (ESI+; MeOH) 571.3 ([M]+,
100%), 573.3 (40, [M+2]+).
5-(7-Chloro-4-(piperazin-1-yl)quinoline)-1,5-bis(4-fluorophenyl)-
1-morpholino-penta-1,3-dienylium
tetrafluoroborate
19b.
5-(N -(7-Chloroquinolin-4-yl)ethane-1,2-diamine)-1-diethyl-
(Found: C, 59.52, H, 4.80, N, 8.05. C34H32BClF6N4O·0.75H2O
amino-1,5-bis(4-fluorophenyl)penta-1,3-dienylium
tetrafluoro-
◦
requires C, 59.49, H, 4.92, N, 8.16%); mp 208 C; dH(300 MHz;
borate 15b. (Found: C, 57.77, H, 5.24, N, 8.68.
C32H32BClF6N4·1.8H2O requires C, 57.77, H, 5.39, N, 8.42%);
mp 160 ◦C; dH(300 MHz; MeOD) 1.12 (3H, m, N–CH2–CH3),
1.36 (3H, m, N–CH2–CH3), 3.25 (2H, m, N–CH2–CH3), 3.68
(2H, m, N–CH2–CH3), 3.85 (4H, m, NH–CH2), 5.79, 6.00 (2H,
CD3CN) 3.20–4.20 (16H, m, O–CH2, N–CH2), 6.26 (3H, m,
3
ABX syst., H2–3–4), 6.98 (1H, d, J 4.8, HB), 7.17 (8H, m, Harom),
3
4
4
7.54 (1H, dd, J 9.0, J 2.4, HH), 8.05 (1H, d, J 2.4, HF), 8.10
3
3
(1H, d, J 9.0, HI) and 8.75 (1H, d, J 4.8, HC); dC(75 MHz;
3
3
3
CD3CN) around 51.2 (broad signal, N–CH2), around 66.0 (broad
2d, J 12.9, H2–4), 6.19 (1H, t, J 12.9, H3), 6.77 (1H, d, J 6.0,
2
3
4
signal, O–CH2), 106.1, 106.3 (C2–4), 109.7 (CB), 115.8 (d, JCF
HB), 7.15 (8H, m, Harom), 7.47 (1H, dd, J 9.0 and J 1.8, HH),
7.83 (1H, d, 4J 1.8, HF), 8.03 (1H, d, 3J 9.0, HI) and 8.45 (1H, d,
3J 6.0, HC); dC(75 MHz; MeOD) 11.2, 12.9 (N–CH2–CH3), 41.4,
42.1 (NH–CH2), 41.2, 44.8 (N–CH2–CH3), 98.6 (CB), 102.4, 105.8
(C2–4), 115.1, 115.4 (2d, 2JCF 28.2, CHarom), 117.5 (CJ ), 123.3 (CI),
22.2, CHarom), 121.7 (CJ ), 125.7 (CI), 126.2 (CH), 128.6 (CF), 128.5,
128.8 (2d, 4JCF 3.4, Carom), 131.2, 131.3 (2d, 3JCF 8.6, CHarom), 134.5
(CG), 150.2 (CE), 152.2 (CC), 155.7 (CA), 162.7 (C3), 163.5 (d, 1JCF
247.6, F–Carom) and 168.7, 169.0 (C1–5); dF(300 MHz; CD3CN)
-151.7, -151.6 (BF4), -111.8 (Farom); m/z (ESI+; MeOH) 585.3
([M]+, 100%), 587.3 (44, [M+2]+).
4
125.1 (CH), 125.9 (CF), 128.8, 130.1 (2d, JCF 3.4, Carom), 130.3,
3
131.0 (d, JCF 8.6, CHarom), 135.4 (CG), 147.8 (CE), 150.7 (CC),
151.5 (CA), 161.2 (C3), 163.3, 164.0 (2d, 1JCF 248.7, F–Carom) and
168.4, 168.6 (C1–5); dF(300 MHz; MeOD) -154.2, -154.1 (BF4),
-112.1, -110.9 (Farom); m/z (ESI+; MeOH) 545.5 (M+, 100%),
547.5 (37, [M+2]+).
1,5-Bis(N-(7-chloroquinolin-4-yl)ethane-1,2-diamine)-1,5-bis(4-
fluorophenyl)penta-1,3-dienylium
tetrafluoroborate
17b. 7a
(0.21 g, 0.95 mmol) in solution in dry DMF (20 cm3) was
added dropwise to 8b (0.20 g, 0.47 mmol) in solution in dry
DMF (20 cm3) at room temperature. After 6 days stirring, the
solvent was removed under reduced pressure, and the solid was
washed with dichloromethane and dried. The crude product was
crystallized from 100% MeOH and dried under vacuum at 40 ◦C.
17b was obtained as yellow crystals (0.11 g, 30%). (Found: C,
56.66, H, 4.58, N, 10.18. C39H33BCl2F6N6·2.5H2O requires C,
56.68, H, 4.63, N, 10.17%); mp 142 ◦C; dH(300 MHz; MeOD)
3.82–3.88 (8H, m, N–CH2–CH2), 5.48 (2H, part A of A2X syst.,
5-(N-(7-Chloroquinolin-4-yl)ethane-1,2-diamine)-1,5-bis(4-fluo-
rophenyl)-1-morpholino-penta-1,3-dienylium tetrafluoroborate 16b.
(Found: C, 58.39, H, 4.73, N, 8.29. C32H30BClF6N4O·0.75H2O re-
quires C, 58.28, H, 4.80, N, 8.50%); mp 204 ◦C (dec); dH(300 MHz;
CD3CN) 3.30–3.90 (12H, m, O–CH2, N–CH2 et NH–CH2), 5.73,
5.89 (2H, 2d, 3J 12.9, H2–4), 6.32 (2H, m, H3 + NH), 6.66 (1H, d,
3
4
3J 5.4, HB), 7.15 (8H, m, Harom), 7.44 (1H, dd, J 9.0 and J 2.1,
3
4
HH), 7.82 (1H, d, J 9.0, HI), 7.88 (1H, d, J 2.1, HF) and 8.53
3
3
(1H, d, J 5.4, HC); dC(75 MHz; MeOD) 41.3, 42.2 (NH–CH2),
3JAX 12.9, H2–4), 6.49 (1H, part. X of A2X syst., JAX 12.9, H3),
3
3
66.1 (O–CH2), 98.7 (CB), 104.0, 105.2 (C2–4), 115.2, 115.7 (2d, 2JCF
22.3, CHarom), 117.6 (CJ ), 123.3 (CI), 125.0 (CH), 126.2 (CF), 128.5,
129.9 (2d, 4JCF 3.3, Carom), 131.0, 131.1 (d, 3JCF 9.4, CHarom), 135.1
6.86 (2H, d, J 6.0, HB), 7.17 (8H, m, Harom), 7.51 (2H, d, J 9.0,
3
HH), 7.85 (2H, m, HF), 8.07 (2H, d, J 9.0, HI) and 8.51 (2H,
d, 3J 6.0, HC); dC(75 MHz; D6DMSO) 41.7, 42.5 (NH–CH2),
7408 | Org. Biomol. Chem., 2011, 9, 7400–7410
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2
99.4 (CB), 103.1 (C2–4), 116.0 (d, JCF 21.9, CHarom), 117.2 (CJ ),
CCDC 805391 for 17b, 805392 for 18b and 805393 for 19b
contain the supplementary crystallographic data for this paper.†
124.4 (CI), 125.1 (CH), 126.0 (CF), 130.2 (d, 4JCF 3.0, Carom), 132.1
(d, JCF 8.9, CHarom), 136.0 (CG), 145.0 (CE), 148.8 (CC), 152.8
(CA), 159.8 (C3), 163.9 (d, JCF 246.5, F–Carom) and 168.0 (C1–5);
3
1
Drug inhibition of in vitro cultured P. falciparum parasite
dF(300 MHz; MeOD) -153.94, -153.89 (BF4), -110.3 (Farom); m/z
(ESI+; MeOH) 693.3 ([M]+, 100%), 695.2 (68, [M+2]+), 697.1 (14,
[M + 4]+).
The three strains of P. falciparum (Nigerian, chloroquino-sensitive,
and FcB1 and FcM29, both being chloroquino-resistant) were
asexually cultured in human blood in complete medium (RPMI
1640 supplemented with 25 mM Hepes, pH 7.4) and 10% AB+
human serum.24 Drug effects were measured in microtiter plates
on suspensions of asynchronous P. falciparum infected red blood
cells (1.5% final haematocrit, 0.6% parasitemia) according to
Desjardins et al.15,25 Drugs (previously dissolved in DMSO) were
diluted in culture medium so that the final DMSO concentration
never exceeded 0.25%. After 48 h incubation at 37 ◦C, parasite
viability was assayed by the incorporation of (3H)Hypoxanthine
(0.5 mCi/well, 22.2 kBq) in parasitic nucleic acids for 18 h.
Cells were then lysed using an automatic cell harvester and the
parasite radioactive nucleic acids were retained on glass fiber
filters (Filtermate collector, Packard Biosciences). The filters were
counted for radioactivity (TopCount NXT, Packard Biosciences).
The radioactivity background was obtained by incubation of non-
infected erythrocytes under the same conditions, and deducted.
Analyses of dose–effect curves were performed with the Graphpad
Prism analytical software. The results are expressed as IC50,
corresponding to the drug concentration leading to 50% parasite
growth inhibition. Values are the means of at least two independent
experiments (different cell cultures, different drug dilution stocks),
each performed in duplicate.
1,5-Bis(7-chloro-4-(piperazin-1-yl)quinoline)-1,5-bis(4-fluoro-
phenyl)penta-1,3-dienylium tetrafluoroborate 20b. 7b (0.12 g, 0.50
mmol) in solution in dry CH3CN (20 cm3) was added dropwise to
8b (0.11 g, 0.25 mmol) in solution in dry CH3CN (20 cm3) at room
temperature. After 24 h stirring, the solvent was removed under
reduced pressure and the solid was washed with pentane and dried.
The crude product was crystallized from 100% MeOH and dried
under vacuum at 40 ◦C. 20b was obtained as yellow crystals (0.15 g,
70%). (Found: C, 58.04, H, 4.49, N, 9.66. C43H37BCl2F6N6·3H2O
◦
requires C, 58.19, H, 4.88, N, 9.47%); mp 150 C; dH(300 MHz;
CD3CN) 3.00–4.20 (16H, m, N–CH2), 6.28 (3H, m, syst. AA¢X,
H2–3–4), 6.98 (2H, d, 3J 5.2, HB), 7.20 (8H, m, Harom), 7.54 (2H, dd,
4
4
3
3J 9.0, J 2.1, HH), 8.05 (2H, d, J 2.1, HF), 8.10 (2H, d, J 9.0,
3
HI) and 8.73 (2H, d, J 5.2, HC); dC(75 MHz; CD3CN) around
50.8 (broad signal, NH–CH2), 106.4 (C2–4), 109.3 (CB), 115.9 (d,
2JCF 22.3, CHarom), 121.3 (CJ ), 125.9 (CI), 126.2 (CH), 127.9 (CF),
4
3
128.7 (d, JCF 3.6, Carom), 131.3 (d, JCF 8.9, CHarom), 134.9 (CG),
149.4 (CE), 151.4 (CC), 156.0 (CA), 162.8 (C3), 163.6 (d, 1JCF 247.6,
F–Carom) and 169.0 (C1–5); dF(300 MHz; CD3CN) -151.7, -151.6
(BF4), -111.7 (Farom); m/z (ESI+; MeOH) 745.3 ([M]+, 100%),
747.3 (80, [M + 2]+), 748.3 (18, [M+4]+).
For all the structures, data were collected at low temperature
on a Bruker-AXS APEX II diffractometer or on a Bruker
APEX II Quazar with ImS, using Mo-Ka radiation (l = 0.7103
In vitro cytotoxicity assay
Cytotoxicity was estimated, on one hand, on human MCF7 breast
cancer cells and, on the other hand, on Vero cells (monkey kidney
normal cells). The cells were cultured in the same conditions (same
culture medium) as those used for P. falciparum, except that the
10% human serum was replaced by 5% foetal calf serum (Cambrex,
Belgium). Cells were distributed in 96-well plates (20 ¥ 103
cells/well and 5 ¥ 103 cells/well for MCF7 and Vero, respectively)
and the streptocyanines were added at various concentrations
in triplicate. Cell growth was estimated by [3H]-Hypoxanthine
(9250 Bq/well added 24 h before the end of the experiment)
incorporation after 48 h incubation.15 The [3H]-hypoxanthine
incorporation, in the presence of streptocyanine, was compared
with that of control cultures without extract (positive control being
doxorubicin (Sigma)). IC50 was determined graphically on growth
inhibition vs. concentration curves. The experiments were repeated
three times.
˚
A). The selected crystals were mounted on a glass fiber using
perfluoropolyether oil and cooled rapidly to 193(2) K or 100(2) K
in a stream of cold N2. Semi-empirical absorption corrections were
employed.21 Structures were solved by direct methods (SHELXS–
97)22 and refined using the least-squares method on F2.23
Crystallographic data for 17b
C39H33BCl2F4N6 2(CH3OH), M = 845.51, T = 193(2) K, mono-
˚
˚
clinic, space group P21/c, a =◦10.6578(2) A, b = 32.0622(6) A, c =
3
˚
˚
12.1186(3) A, b = 100.604(1) , V = 4070.36(15) A , Z = 4, rc =
1.3080 Mg m-3, m = 0.231 mm-1, 134094 reflections, 13147 unique
(R(int) = 0.0368), R1 [I > 2s(I)] = 0.0526, wR2 [all data] = 0.1581.
Crystallographic data for 18b
C34H34BClF6N4O0.5, M = 666.91, T = 193(2) K, monoclinic, space
˚
˚
group P21/c, a =◦17.3524(3) A, b = 11.5299(2) A, c = 18.2276(3)
Acknowledgements
3
-3
˚
˚
A, b = 112.482(1) , V = 3369.66(10) A , Z = 4, rc = 1.315 Mg m ,
m = 0.178 mm-1, 49681 reflections, 7401 unique (R(int) = 0.0366),
R1 [I > 2s(I)] = 0.0683, wR2 [all data] = 0.2135.
We thank the Ministe`re de la Recherche (France) and the Univer-
site´ Paul Sabatier (Toulouse) for financial support and M. May-
nadier and B. Moukarzel for in vitro assay.
Crystallographic data for 19b. C34H32BClF6N4O (0.5 CH3OH,
1 H20), M = 706.93, T = 100(2) K, monoclinic, space group
˚
˚
˚
C2/c, a = ◦21.3858(4) A, b = 11.0501(2) A, c = 30.9188(7) A, b =
Notes and references
3
-3
˚
108.770(1) , V = 6918.0(2) A , Z = 8, rc = 1.357 Mg m , m =
0.182 mm-1, 30548 reflections, 7013 unique (R(int) = 0.0723), R1
[I > 2s(I)] = 0.0532, wR2 [all data] = 0.1333.
1 World Malaria Report 2010, World Health Organisation,
Geneva
(Switzerland),
http://www.who.int/malaria//world_
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©