Synthesis of new arylalkoxy amido derivatives as melatoninergic ligands

Article abstract of DOI:10.1016/S0968-0896(02)00328-0

Amido derivatives 10-18 of the corresponding oxyamines were synthesised as melatoninergic ligands by the reaction of hydroxyphtalimide with the halogeno derivatives or the corresponding alcohols using Mitsunobu reaction conditions. The affinity of the com

Full text of DOI:10.1016/S0968-0896(02)00328-0

Bioorganic & Medicinal Chemistry 11 (2003) 789–800
Synthesis of New Arylalkoxy Amido Derivatives as
Melatoninergic Ligands
Cecile Pegurier,a Laurence Morellato,a Eminn Chahed,a
Jean Andrieux,a Jean-Paul Nicolas,b Jean A. Boutin,b
Caroline Bennejean,c Philippe Delagrange,d Michel Langloisa
and Monique Mathe-Allainmat,a,*
a
´
´
CNRS-BIOCIS (UPRES A 8076), Universite de Paris-Sud, Faculte de Pharmacie,
´
5 rue Jean Baptiste Clement, 92296, Chatenay-Malabry, France
ˆ
bInstitut de Recherche Servier, Centre de Croissy, 125 Chemin de ronde, 78290 Croissy sur Seine, France
c
´
ADIR, 1 rue Carle Hebert, 92415 Courbevoie, France
´
Institut de Recherche Internationale Servier, Place des Pleiades, 92415 Courbevoie, France
d
Received 15 February 2002; accepted 27 June 2002
Abstract—Amido derivatives 10–18 of the corresponding oxyamines were synthesised as melatoninergic ligands by the reaction of
hydroxyphtalimide with the halogeno derivatives or the corresponding alcohols using Mitsunobu reaction conditions. The affinity
of the compounds for chicken brain melatonin receptors and recombinant human MT1 and MT2 receptors was evaluated using
2-[125I]-iodomelatonin as the radioligand. Overall, the introduction of an oxygen atom in the amido chain was not a favourable
parameter as the compounds were less potent than the corresponding deoxy derivatives. However, nanomolar compounds were
obtained with the arylethyloxy derivatives (13c (R0=nPr), chicken brain, hMT1, hMT2, Ki values: 4.8, 3.86, 2.4 nM, respectively)
and the 2,7-dimethoxynaphthalene derivatives (17c (R0=nPr), chicken brain, hMT1, hMT2, Ki values: 0.04, 0.13, 0.1 nM, respec-
tively). The functional activity of these compounds was evaluated by the aggregation of melanophores in Xenopus laevis tadpoles
and the potency was related to the affinity of the molecules for melatonin receptors. The compounds were found to be full agonists
and compound 17a was 20-fold more potent than melatonin in this bioassay.
# 2002 Elsevier Science Ltd. All rights reserved.
Introduction
Almost all of the effects of melatonin are mediated
through G protein-coupled receptors10 which have been
cloned11 and characterised as MT1 and MT2 recep-
tors.12 Coupling to the Gi family of G-proteins appears
to be the most usual signalling pathway for these
receptors.10 Recently, considerable interest has been
focused on the search for new molecules capable of
mimicking or antagonising the response to melatonin.
These novel compounds13ꢀ15 were derived from the
indole ring, a bioisosteric naphthalene moiety such as
agomelatine or the simple phenylalkylamide framework.
Many structure–activity relationships (SAR) have been
described and the essential role of the amido function
has been emphasised in the binding of the molecule with
the receptor site. Several authors have demonstrated
that the melatoninergic activity of the molecules
depends upon the stereochemistry of the amidic moiety
with regard to that of the aromatic ring. A number of
constrained melatoninergic derivatives such as compounds
Melatonin (N-acetyl-5-methoxytryptamine) is the verte-
brate pineal gland hormone secreted during darkness.1 It is
now well established that it modulates the circadian
rhythm2 in a large number of animals and in humans. It
can be used to control diseases associated with circadian
rhythm disorders: jet-lag or delayed sleephpase
syndrome.3ꢀ5 Conversely, it has been implicated in seaso-
nal and winter depression.6 Melatonin controls the breed-
ing cycle in photoperiodic species and can be used to induce
reproduction outside the breeding season.7 Melatonin has
been also reported to have antiproliferative effects on
mammary cell lines8 and, more recently, a role for melato-
nin has been suggested in the regulation of vascular tone.9
*Corresponding author. Fax: +33-2-5112-5492; e-mail: monique.mathe@
chimie.univ_nantes.fr
0968-0896/03/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(02)00328-0

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C. Pegurier et al. / Bioorg. Med. Chem. 11 (2003) 789–800
790
1, 2, 3, 4, 5 highlighted this point.16ꢀ20 Sudgen21 sug-
gested the existence of a hydrogen bond between the
NH amidic groupand the asparagine residue of the
transmembrane helice IV of the receptor, although no
experimental data have confirmed this hypothesis. Con-
sequently, performing chemical modifications centered
around the N-acyl groupwas a good way to study the
ability of NH groupto establish a hydrogen bond with
the receptor site. We report herein the synthesis of the
N-acyl hydroxylamine derivatives 6 (Scheme 1, com-
pounds 10–18) which possess an oxygen atom on the a
position of the amido function and aromatic groups R
selected from the best melatoninergic ligands described
previously13ꢀ15 with nanomolar affinity for melatonin
receptors. The compounds were evaluated on chicken
brain melatonin receptors20 and on human MT1 and
MT2 receptors expressed in HEK-293 cells22 by their
ability to inhibit the binding of 2-[125I]-iodomelatonin.
Functional activity at melatonin receptors was eval-
uated by examining the potency of the compounds to
lighten the skin of Xenopus laevis tadpoles23 as it has
been clearly demonstrated that melatonin mediates the
aggregation of melanophores.19,24 The degree of disper-
sion of melanophores on the head and dorsal surface of
the tadpoles observed under a microscope can be asses-
sed using the melanophore index scale (1–5) of Hogben
and Slome25 and concentration–effect curves (5 con-
centrations) can be plotted.
corresponding phthalimido derivatives 8f,g with good
yields (method A). However, the conditions were drastic
and condensation of the alcohols 7a–e, h, i (X=OH)
with N-hydroxyphthalimide according to the conditions
of Mitsunobu reaction26,27 was preferred (method B). The
compounds were reacted in THF at room temperature
in the presence of DEAD and PPh3 and the yields of the
N-alkoxyphthalimides 8 were good. The O-alkylhydro-
xylamines 9 were prepared by the treatment of 8 with
hydrazine in ethanol at reflux and isolated as oils. They
were acylated with acid chlorides in the presence of
Na2CO3 in a biphasic system (CH2Cl2/H2O 1/1)
according to the process reported previously.20 Acyl
groups (R0-CO, R0=Me, Et, n-Pr, CH2Cl, cPr) were
selected according to previous results12ꢀ14 from SAR
(structure–activity relationships) obtained with ligands
for the melatonin receptors. The new compounds were
characterised by NMR spectra and microanalysis.
Results and Discussion
The binding data (Ki, nM) on chicken brain melatonin
receptors and human MT1 and MT2 receptors are
reported in Table 1. By comparison with the corre-
sponding deoxy compounds.28ꢀ30 the results show an
overall decrease in affinity for melatonin receptors. The
effect was particularly marked with the benzylhydroxyl-
amine derivatives 10, 11 and 12 which were inactive or
weakly active while the potency of 3-methoxy-
phenethyloxy derivatives such as 13c was in the nano-
molar range and nearly equipotent to the corresponding
arylpropyl derivative.29 A less marked decrease was
observed with compounds 14, 16 and 18 possessing the
tetralin, naphthalene and acenaphtene cyclic moieties,
respectively. Thus, the naphthalenic derivative 16a
(R0=Me) with the methoxy in the ortho position was
20-fold less potent than the deoxy derivative prepared
previously.28 Conversely, the naphthalenic analogues of
agomelatine 15 and of the corresponding dimethoxy
derivatives 17 were nearly equipotent to the ethylamido
derivatives. This is particularly clear with compound
17c (chicken brain, hMT1, hMT2, Ki values: 0.04, 0.13,
0.10 nM, respectively) compared to the deoxy ana-
logue31 (chicken brain, hMT1, hMT2, Ki values: 0.08,
0.02, 0.08 nM, respectively). The distance between the
methoxy groupin the melatonin-like position and the
amido group seems to be an important parameter in
determining the potency of binding with the receptor
site.32 These two groups were separated by 6 or less
bonds in compounds 10, 11, 12, 14 and 16 which were
weakly active or inactive, while a distance of 7 bonds
was present with the most active compounds such as 13,
15 and 17.
Chemistry
The presence of an oxygen atom in the chain produces
several changes in structural parameters with regard to
those of the deoxy compounds: variation of bond
angles, a different range of lower energy conformers,
introduction of additional hydrogen bonds or reduction
of the flexibility of the chain. Consequently, it was
worth examining if the presence of an oxygen atom
would modify the stereochemistry of the permissible
Compounds 6 (10–18) were synthesised according to the
chemical pathway described in the Scheme 1. Two
methods were used, according to the process already
reported, for the synthesis of the O-aryl hydroxylamine.
N-Hydroxyphthalimide was reacted in DMF25 at 100 ꢁC
with the arylmethyl chloride 7f, g (X=Cl) to give the

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C. Pegurier et al. / Bioorg. Med. Chem. 11 (2003) 789–800
791
Scheme 1. (a) 100 ꢁC, DMF; (b) THF, DEAD, PPh3, rt; (c) NH2NH2, EtOH, Á; (d) R0COCl, CH2Cl2/H2O, Na2CO3.
conformers. Conformational search (Alchemy 2000) was
performed on 17a (R0=Me) and calculations made
around the three rotable bonds C(12)–C(13), C(13)–
O(14) and O(14)–N(15) (Fig. 1). A low energy conformer,
similar to the putative active conformer previously
reported by us for the deoxy compound,33 was identified
among a number of permissible conformers. Thus, 17a
can be superimposed (Fig. 1) on the folded putative
active conformer of melatonin determined indepen-
dently by Sicsic33 and Marot34 by the CoMFA method
and the potent melatoninergic ligands derived from phe-
nalene derivatives,20 suggesting that the presence of an
oxygen atom exerts a weak influence on the general
structure of the pharmacophore. Consequently, electro-
nic rather than steric factors appear to be implicated in
the dropin activity for almost all the compounds studied.
of the skin of Xenopus laevis similarly to melatonin. As
previously observed, potency was related to the affinity
calculated from the receptor binding assays. Almost all
the compounds (Table 1) were less potent than melato-
nin. However, as in the other series previously studied,
the potency of the 2,7-dimethoxynaphthalene derivative
17a was clearly emphasized (23-fold more potent than
melatonin) but to a lesser extent than in the series of
corresponding deoxy compounds which were more than
100-fold more potent than melatonin.19
In summary, introduction of an oxygen atom in the
amido chain of melatoninergic ligands does not modify
the pharmacological profile of the compounds since,
similarly to the parent compounds, they were full ago-
nists in the functional assay measuring the lightening of
the skin of Xenopus laevis tadpoles. On the other hand,
the presence of an oxygen rather than a carbon atom
seems less favourable for binding to the receptor site as
almost all the compounds were less potent than the
corresponding deoxy derivatives. However, a potent
compound, 17a, was obtained, equipotent to melatonin
in the binding assay and clearly superior in the melano-
phore aggregation assay.
Furthermore, results obtained for the affinities of the
different compounds for MT1 and MT2 receptors indi-
cate that the oxygen atom does not influence selectivity
for these receptors.
However, the introduction of an oxygen atom in the
amido chain could modify the flexibility or acidic char-
acter of the NH groupand, therefore, influence the
ability of the molecules to activate the receptor. The
pharmacological profiles of these compounds were
evaluated on the dermal melanocytes of X. laevis tad-
poles. Melatonin, naphthalenic20 and indolic24 com-
pounds have been shown to be potent agonists in this
bioassay.
Experimental
1H and 13C spectra were recorded on a BRUCKER AC
200 spectrometer (200 and 50.3 MHz respectively) with
tetramethylsilane as the internal standard. Chemical
shifts are reported in parts per million (ppm) in d units.
1H NMR multiplicity data are denoted by s (singlet), d
(doublet), dd (doublet of doublet), triplet (triplet), q
(quadruplet), quin (quintuplet), sx (sextuplet), m (mul-
tiplet) and br (broad). Coupling constants are in Hertz.
Elemental analyses were performed at the Microanalysis
service in the Chatenay-Malabry Pharmacy Faculty,
(France). IR spectra were recorded on a Perkin–Elmer
PE 481 spectrometer. Column chromatography was
performed using SDS Silica 60-A (35–70 mm) as the sta-
tionary phase. 3-Methoxy benzyl alcohol, 2,5-dime-
thoxy benzyl alcohol, 3-methoxyphenylethanol and
Several of the compounds synthesized (10, 12–17a) were
studied in the functional assay, and their EC50 values
are reported in Table 2. Variations in the EC50 values of
melatonin have been observed between batches of tad-
poles at different times of the year; consequently the
potency of the molecules was calculated as the EC50
(melatonin)/EC50 (compound) ratio determined in the
same experiment.
All tested compounds were characterised as full agonists
(100% of effect) and produced dose-related lightening

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C. Pegurier et al. / Bioorg. Med. Chem. 11 (2003) 789–800
792
Table 1. Inhibition of 2-[125I]-iodomelatonin binding by compounds 10–18
Compd
R0
hMT1 receptorsa Ki (nM)
hMT2 receptorsa Ki (nM)
Chicken brain receptorsb, Ki (nM)
10a
10b
10c
10d
10e
11a
11b
11c
11d
11e
12a
12b
12c
12d
12e
13a
13b
13c
13d
13e
14a
14b
14c
14d
14e
15a
15c
15e
16a
16b
16e
17a
17b
17c
17d
17e
18a
18b
18c
18d
18e
Melatonin
Me
Et
nPr
ClCH2
cPr
Me
Et
nPr
ClCH2
cPr
Me
Et
nPr
ClCH2
cPr
Me
Et
nPr
ClCH2
cPr
Me
Et
nPr
ClCH2
cPr
Me
nPr
cPr
Me
Et
cPr
Me
>1000
>1000
>1000
>1000
>1000
69.4ꢂ6.9
>1000
510
640
>1000
>1000
>1000
185ꢂ16.4
46.3ꢂ10
>1000
520
>1000
879ꢂ375
>1000
>1000
>1000
>1000
>1000
>1000
>1000
95.8ꢂ8.2
>1000
170ꢂ50
>1000
>1000
>1000
>1000
940ꢂ180
270ꢂ65
210ꢂ50
160ꢂ32
>1000
201ꢂ34
172ꢂ76.1
104ꢂ15.5
>1000
186ꢂ37
89.7ꢂ8
63.7ꢂ5.4
>1000
7.51ꢂ2.1
2.1ꢂ0.4
3.86ꢂ0.08
1.22ꢂ0.02
1.84ꢂ0.5
27.1ꢂ7.8
10.6ꢂ3.5
14ꢂ4
11.7ꢂ2
2.6ꢂ0.6
2.4ꢂ0.04
1.3ꢂ0.05
4.43ꢂ0.03
9.43ꢂ0.6
2.32ꢂ0.6
1.42ꢂ0.01
0.93ꢂ0.05
96.2ꢂ14.8
NT
55ꢂ17
20ꢂ4
4.8ꢂ1.2
4.2ꢂ0.7
6.6ꢂ1.5
40ꢂ11
17ꢂ4
8.5ꢂ1.5
8.4ꢂ1.7
69ꢂ14
5.57ꢂ1.2
471ꢂ89
NT
1.7ꢂ0.4
2ꢂ0.5
0.41ꢂ0.01
0.23ꢂ0.02
NT
NT
NT
NT
1.5ꢂ0.3
55ꢂ3.2
75
NT
NT
NT
NT
>1000
0. 24ꢂ0.07
0. 117ꢂ0.04
0. 133ꢂ0.6
0.034ꢂ0.01
0. 28ꢂ0.08
286ꢂ26
109ꢂ11.5
132ꢂ22.5
23.5ꢂ2.5
43.9ꢂ2.5
0.54ꢂ0.1
0. 28ꢂ0.07
0. 125ꢂ0.6
0.01ꢂ0.03
0.059.4ꢂ0.02
0.51ꢂ0.13
35.1ꢂ13.1
10.4ꢂ5.5
6.13ꢂ2
5.4ꢂ0.1
67ꢂ16
0.26ꢂ0.07
0.08ꢂ0.013
0.042ꢂ0.012
0.02ꢂ0.006
0.34ꢂ0.1
>1000
370ꢂ65
350ꢂ220
130ꢂ60
500ꢂ220
0.56ꢂ0.03
Et
nPr
ClCH2
cPr
Me
Et
nPr
ClCH2
cPr
0.34ꢂ0.1
a2-[125I]-iodomelatonin was used as radioligand and binding assays were carried out using membranes prepared from HEK 293 cells expressing
human MT1 or MT2 receptors. Membrane aliquots were incubated for 2 h at 37 ꢁC with 0.025 nM and 0.2 nM 2-[125I]-iodomelatonin for hMT1 and
hMT2 receptors, respectively.
bBinding assays were carried out using membranes prepared from chicken brain. Membrane aliquots were incubated with 0.05 nM 2-[125I]-iodome-
latonin for 1 h at 25 ꢁC. a,bNon-specific binding was defined with 10 mM 2-iodomelatonin. Each binding assay was performed in triplicate. Experi-
ments were performed twice.
Figure 1. Minimum energy folded conformer of 17a, melatonin and phenalene derivative 3. The conformers of 17a and 3 have been selected by
conformational search (Alchemy 2000, version 3.2) and by their structural similarity to the CoMFA model of the melatonin pharmacophore
described previously.33 The fit of the three conformers on carbon atom C(5), nitrogen atom N(15) and oxygen atoms O(20) and O(39) gave a rms
value of 0.2.

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793
Table 2. Biological data for compounds 10a, 12a, 13a, 14a, 15a, 16a
and 17a for X. laevis melanophore aggregation
16.1 mmol), N-hydroxyphthalimide (2.62 g, 16.1 mmol)
and triphenylphosphine (4.22 g, 17.7 mmol) were dis-
solved in 80 mL of anhydrous THF. Diethyl azodi-
carboxylate (DEAD, 2.79 mL, 17.7 mmol) was added
dropwise while stirring to the solution and the mixture
was stirred for 25 h at room temperature. The solvent
was evaporated under vacuum and the compound iso-
lated as a yellow solid. It was purified by column chro-
matography (silica gel) and eluted with CH2Cl2.ꢁ2.26 g
(yield: 88%) of white solid was obtained, mp121 C. 1H
NMR (200 MHz, CDCl3) d (ppm) 3.82 (s, 3H), 5.19 (s,
2H), 6.87–6.94 (m, 1H), 7.07–7.12 (m, 2H), 7.28 (t,
J=7.8 Hz, 1H), 7.70–7.83 (m, 4H). 13C NMR
(200 MHz, CDCl3) d (ppm) 55.34, 79.77, 114.65, 115.46,
121.98, 129.57, 129.09, 135.40, 159.52, 163.56. Anal.
(C16H13NO4) calcd C 67.84H 4.62 N 4.94; F. C 67.71H
4.73 N 4.93.
Compd
EC50 (nM) for
melanophore aggregation
Potency with regard
to melatonin (=1ꢁ)
10a
12a
13a
14a
15a
16a
17a
Melatonin
Agomelatine
7.6 [4.9–11.70]
6.9 [3.21–14.9]
0.068 [0.034–0.13]
0.34 [0.16–0.73]
0.008 [0.0034–0.019]
0.42 [0.19–0.92]
0.00017 [0.00007–0.0004]
0.0039 [0.0022–0.0071]
0.00031 [0.00014–0.00065]
0.0005
0.0005
0.057
0.011
0.48
0.009
23
1
12.5
X. laevis tadpoles were placed in groups of 5 in 100 mL beakers. The
compounds under test (5 concentrations) were dissolved in a final
volume of 5 mL and added to the beaker. The degree of the melano-
phore response was determined by examination of the head and body
surface using the melanophore index (1–5) of Hogben and Slome.25
EC50 values were determined using the PRISM software package and
are the result of 2 separate experiments. The mean value is given with
the range of values in brackets. The potency of the molecules was cal-
culated as the EC50 (melatonin)/EC50 (compound) ratio determined in
the same experiment.
2 - [(2,5 - Dimethoxyphenyl)methoxy] - 1H - isoindole -
1,3(2H)-dione (8b). It was prepared according to the
previous method described for 8a. The compound was
crystallised in an EtOAc/hexaꢁne mixture to give white
1
crystals (yield: 75%), mp130 C. H NMR (200 MHz,
acenapht-1-ol were commercially available. 7-Methoxy-
1,2,3,4-tetrahydronapht-1-ol was prepared by the
reduction of the corresponding ketone with AlLiH4.
(2,7-dimethoxynaphth-1-yl) methanol and 1-chloro-
methyl-2-methoxynaphthalene, 1-chloromethyl-7-meth-
oxynaphthalene were prepared according to the process
already reported.28
CDCl3) d (ppm) 3.71–3.75 (2s, 6H), 5.25 (s, 2H), 6.86
(dd, J=9.4 Hz, J=3.0 Hz, 1H), 7.05 (sd, J=2.9 Hz,
1H), 7.71–7.78 (m, 4H). 13C NMR (200 MHz, CDCl3) d
(ppm) 55.83, 56.19, 74.50, 111.95, 115.94, 117.08,
123.35,134.30, 129.02,130.00, 152.46,153.46, 163.46.
Anal. (C17H15NO5) calcd C 65.17 H 4.82 N 4.47; F. C
65.12 H 4.75 N 4.43.
2-[1-(3-Methoxyphenyl)ethoxy]-1H-isoindole-1,3(2H)-
dione (8c). It was prepared according to the previous
method described for 8a. The compound was isolated as
a colourless oil (yield: 92%). 1H NMR (200 MHz,
CDCl3) d (ppm) 1.69 (d, J=6.5 Hz, 3H), 3.81 (s, 3H),
5.49 (q, J=6.6 Hz, 1H), 6.82–7.10 (m, 3H), 7.22 (t,
J=7.9 Hz, 1H), 7.67–7.73 (m, 4H). 13C NMR
(200 MHz, CDCl3) d (ppm) 20.79, 55.31, 85.04, 112.52,
114.92, 119.83, 129.35, 123.38, 134.32, 128.98, 140.81,
159.61, 163.82. Anal. (C17H15NO4) calcd C 68.68 H
5.08 N 4.71; F. C 68.54 H 5.06 N 4.67.
Preparation of N-alkoxyphtalimides 8a–i—general
procedure. Method A
2 - [(2 - Methoxy - 1 - naphthyl)methoxy] - 1H - isoindole -
1,3(2H)-dione (8g). Sodium acetate (0.1 M) was added
to a solution of N-hydroxyphthalimide (0.1 M) in 60 mL
of DMSO which turned dark red. 1-Chloromethyl-2-
methoxynaphthalene (0.1 M) in 20 mL of DMSO was
added slowly to the mixture and the solution was
warmed with stirring at 100 ꢁC until decolouration of
the solution had occurred. The mixture was extracted
with chloroform (500 mL) and the organic solution was
washed twice with saturated NaCl solution and dried
over Na2SO4. The organic solvents were evaporated
under reduced pressure and the solid residue was pur-
ified by column chromatography (silica gel) and eluted
with CH2Cl2. The compound was isolated as a white
solid (yield: 80%) mp174 ꢁC. 1H NMR (200 MHz,
CDCl3) d (ppm) 3.90 (s, 3H), 5.80 (s, 2H), 7.20–8.50 (m,
10H).
2-[(3-Methoxyphenethyl)oxy]-1H-isoindole-1,3(2H)-dione
(8d). It was prepared according to the previous method
described for 8a. The compound was crystallised after
chromatography in an EtOAc/hexane mixture to give a
white powder (yield: 83%), mp 85 ꢁC. 1H NMR
(200 MHz, CDCl3) d (ppm) 3.13 (t, J=7.3 Hz, 2H), 3.80
(s, 3H), 4.43 (t, J=7.3 Hz, 2H), 6.71–6.90 (m, 3H), 7.20
(t, J=8.2 Hz, 1H), 7.71–7.8 ((m, 4H).). 13C NMR
(200 MHz, CDCl3) d (ppm) 34.66, 55.21, 78.39, 112.29,
114.40, 121.16, 129.54, 123.54,134.50, 128.96, 138.41,
159.80, 163.68. Anal. (C17H15NO4) calcd C 68.68 H
5.08 N 4.71; F. C 68.59 H 5.03 N 4.65.
2 - [(7 - Methoxy - 1 - naphthyl)methoxy] - 1H - isoindole -
1,3(2H)-dione) (8f). It was prepared according to the
previous method described for 8g. The compound was
isolated as a white solid (yield: 90%) mp202 ꢁC. H
1
NMR (200 MHz, CDCl3) d (ppm) 4.05 (s, 3H), 5.50 (s,
2H), 7.10–8.10 (m, 10H).
2-[(7-Methoxy-1,2,3,4-tetrahydro-1-naphthalenyl)me-
thoxy]-1H-isoindole-1,3(2H)-dione (8e). It was prepared
according to the previous method described for 8a. The
compound was crystallised in an EtOAc/hexane mixture
to give a white powder (yield: 69%), mp 88 ꢁC. 1H
NMR (200 MHz, CDCl3) d (ppm) 1.74–2.92 (m, 6H),
3.77 (s, 3H), 5.26 (t, J=3.7 Hz), 6.83 (dd, J=8.4 Hz,
Method B
2-[(3-Methoxyphenyl)methoxy]-1H-isoindole-1,3(2H)-
dione (8a). 3-Methoxybenzylic alcohol (2 mL,

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C. Pegurier et al. / Bioorg. Med. Chem. 11 (2003) 789–800
794
J=2.9 Hz, 1H), 7.04 (d, J=8.4 Hz, 1H), 7.27 (sd,
J=2.7 Hz, 1H), 7.71–7.84 (m, 4H). 13C NMR
(200 MHz, CDCl3) d (ppm) 18. 13, 27. 55, 28. 25, 55.34,
83.69, 114.88, 116.46, 129.96, 123.44,134.47, 129.09,
130.94, 132.76, 157.72, 164.32. Anal. (C19H17NO4) calcd
C 70.58 H 5.34 N 4.33; F. C 70.37 H 5.25 N 4.28.
for 9a. The compound was isolated as a colourless oil
(yield: 95%). 1H NMR (200 MHz, CDCl3) d (ppm) 1.41
(d, J=6.5 Hz, 3H), 3.82 (s, 3H), 4.64 (q, J=6.5 Hz, 1H,
br s, 2H), 6.80–6.94 (m, 3H), 7.28 (t, J=7.5 Hz, 1H).
13C NMR (200 MHz, CDCl3) d (ppm) 221.93, 55.23,
82.90, 111.69, 113.21, 118.67, 129.65, 144.77, 159.91.
2-[(2,7-Dimethoxy-1-naphthyloxy]-1H-isoindole-1,3(2H)-
dione (8h). It was prepared according to the previous
method described for 8a. The compound was crystallised
in an EtOAc/hexane mixture to give a light yellow solid
(yield: 90%). 1H NMR (200 MHz, CDCl3) d (ppm) 3.97–
4.11 (2s, 6H), 5.74 (s, 2H), 7.02 (dd, J=8.9 Hz, J=2.5 Hz,
1H), 7.10 (d, J=9.0 Hz, 1H), 7.64–7.89 (m, 6H). 13C
NMR (200 MHz, CDCl3) d (ppm) 55.76, 56.97, 70.53,
101.97, 110.44, 117.05, 128.78, 131.72, 113.56, 124.79,
145.90, 123.43,134.38, 129.40, 157.68, 159.33, 163.13.
1-[2-(Aminooxy)ethyl]-3-methoxybenzene (9d). It was
prepared according to the previous method described
for 9a. The compound was isolated as a colourless oil
(yield: 89%). 1H NMR (200 MHz, CDCl3) d (ppm))
2.88 (t, J=6.9 Hz, 2H), 3.79 (s, 3H), 3.89 (t, J=6.9 Hz,
2H), 4.95 (br s, 2H), 6.75–6.84 (m, 3H), 7.22 (t,
J=8.4 Hz, 1H). 13C NMR (200 MHz, CDCl3) d (ppm)
35.11, 55.16, 76.38, 111.57, 114.72, 121.28, 129.40,
140.51, 159.71. Anal. (C9H15NO2) calcd C 64.65 H
7.84 N 8.38; F. C 63.73 H 7.50 N 8.31.
2 - [1,2 - Dihydro - 1 - acenaphthylenyloxy) - 1H - isoindole -
1,3(2H)-dione) (8i). It was prepared according to the
previous method described for 8a. The compound was
crystallised in an EtOAc/hexane mixture to give light
yellow solid (yield: 81%), mp158 ꢁC. 1H NMR
(200 MHz, CDCl3) d (ppm) 3.74 (dd, J=17.0 Hz,
J=6.6 Hz, 1H), 3.69 (d, J=17.2 Hz, 1H), 6.24 (sd,
J=6.5 Hz, 1H), 7.35 (d, J=6.8 Hz, 1H), 7.47–7.58 (m,
2H), 7.65–7.88 (m, 7H). 13C NMR (200 MHz, CDCl3) d
(ppm) 38.11, 87.33, 120.13, 123.67, 122.90, 126.29,
127.78, 128.21, 123.58, 134.51, 129.41, 132.75, 139.39,
142.22, 142.75, 163.69. Anal. (C20H13NO3) calcd C
76.18 H 4.15 N 4.44; F. C 76.09 H 4.26 N 4.40.
1-(Aminooxy)-7-methoxy-1,2,3,4-tetrahydronaphthalene
(9e). It was prepared according to the previous method
described for 9a. The compound was isolated as a col-
1
ourless oil (yield: 83%). H NMR (200 MHz, CDCl3) d
(ppm) 1.61–2.86 (m, 6H), 3.79 (s, 3H), 4.61 (t,
J=3.7 Hz, br s, 2H), 6.79 (dd, J=8.4 Hz, J=2.7 Hz,
1H), 6.97 (ds, J=2.7 Hz, 1H), 7.03 (d, J=8.4 Hz, 1H).
13C NMR (200 MHz, CDCl3) d (ppm) 18.96, 27.14,
28.58, 55.37, 79.81, 113.91, 114.65, 129.93, 130.33,
139.18, 157.68. Anal. (C11H15NO2) calcd C 68.37 H
7.82 N 7.25; F. C 68.17 H 7.80 N 7.21.
1-[(Aminooxy)methyl]-7-methoxynaphthalene (9f). It was
prepared according to the previous method described
for 9a. The compound was isolated as a colourless oil.
1H NMR (200 MHz, CDCl3) d (ppm), 3.95 (s, 3H), 5.25
(s, 2H), 5.30 (s, 2H), 7.05–8.20 (m, 6H).
Preparation of O-alkylhydroxylamines—general procedure
1 - [(Aminooxy)methyl] - 3- methoxybenzene (9a). The
phthalimide derivative 8a (1.75 g, 6.2 mmol) was dis-
solved in 12 mL of EtOH. Hydrazine (0.3 mL, 6.2 mmol)
was added slowly and the mixture was refluxed for 1 h.
The mixture was cooled and poured into a solution
(20 mL) of Na2CO3 (3%). The mixture was extracted
with diethyl ether and the organic solution was dried
over MgSO4. The organic solvent was evaporated under
vacuum and the hydroxylamine was obtained as a col-
ourless oil (0.84 g, yield: 88%). 1H NMR (200 MHz,
CDCl3) d (ppm) 3.77 (s, 3H); 4.62 (s, 2H); 5.06 (br s,
2H), 6.81–6.92 (m, 3H); 7.24 (t, J=7.9 Hz, 1H); 7.70–
7.83 (m, 4H). 13C NMR (200 MHz, CDCl3) d (ppm)
55.67, 78.83, 114.59, 114.67, 121.52, 130.50, 140.27,
161.33. Anal. (C8H11NO2) calcd C 62.73 H 7.24 N 9.14;
F. C 62.54 H 7.28 N 8.97.
1-[(Aminooxy)methyl]-2-methoxynaphthalene (9g). It
was prepared according to the previous method descri-
bed for 9a. The compound was isolated as a colourless
1
oil. H NMR (200 MHz, CDCl3) d (ppm), 3.90 (s, 3H),
5.25 (s, 2H), 5.30 (s, 2H), 7.05–8.20 (m, 6H).
1-[(Aminooxy)methyl]-2,7-dimethoxynaphthalene hydro-
chloride (9h). It was prepared according to the previous
method described for 9a. The compound was isolated as
a colourless oil. The hydrochloride was prepared by ꢁthe
addition of HCl in diethyl ether (yield: 60%) mp178 C.
1H NMR (200 MHz, CD3OD) d (ppm) 3.94–3.98 (2s,
6H), 5.57 (s, 2H), 7.01 (dd, J=8.8 Hz, J=2.5 Hz, 1H),
7.24 (d, J=9.1 Hz, 1H), 7.32 (ds, J=2.5 Hz, 1H) 7.71 (d,
J=8.8 Hz, 1H), 7.88 (d, J=9.2 Hz, 1H). 13C NMR
(200 MHz, CD3OD) d (ppm) 56.06, 56.99, 68.80, 102.50,
111.27, 117.50, 131.33, 133.46, 113.09, 125.99, 136.60,
159.13, 160.79. Anal. (C13H16NO3Cl) calcd C 57.81 H
5.98 N 5.19; F. C 57.70 H 5.99 N 5.20.
1-[(Aminooxy)methyl]-2,5-dimethoxybenzene (9b). It was
prepared according to the previous method described
for 9a. The compound was isolated as a colourless oil
(yield: 85%). 1H NMR (200 MHz, CDCl3) d (ppm) 2.03
(br s, 2H), 3.77–3.79 (2s, 6H), 4.73 (s, 2H), 6.80–6.94
(m, 3H). 13C NMR (200 MHz, CDCl3) d (ppm) 55.79,
56.16, 72.96, 111.76, 113.80, 115.78, 126.62, 151.92,
153.58. Anal. (C9H13NO3) calcd C 59.00 H 7.15 N 7.64;
F. C 58.71 H 7.18 N 7.61.
1-(Aminooxy)-1,2-dihydroacenaphthylene (9i). It was
prepared according to the previous method described
for 9a. The compound was isolated as a colourless oil
1
(yield: 83%). H NMR (200 MHz, CDCl3) d (ppm) 3.40
(dd, J=16.5 Hz, J=2.5 Hz, 1H), 3.63 (dd, J=16.9 Hz,
J=7.0 Hz, 1H), 7.30 (d, J=6.4 Hz, 1H), 7.42–7.65 (m, 4H),
7.75 (d, J=8.0 Hz, 1H). 13C NMR (200 MHz, CDCl3) d
1-[1 - (Aminooxy)ethyl]-3-methoxybenzene (9c). It was
prepared according to the previous method described

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795
(ppm) 38.15, 86.07, 120.62, 122.20, 123.59, 126.12, 128.66,
128.96, 132.28, 139.54, 142.33, 142.77. Anal. (C12H11NO)
calcd C 77.81 H 5.98 N 7.56; F. C 77.57 H 6.21 N 7.40.
method described for 10a with cyclopropylcarbonyl
chloride. The compound was isolated as a white solid
ꢁ
1
(yield: 81%) mp87 C. H NMR (200 MHz, CD3OD) d
(ppm) 0.73–0.88 (m, 4H), 1.28–1.46 (m, 1H), 3.79 (s,
3H), 4.79 (s, 2H), 6.87–6.98 (m, 3H); 7.26 (t, J=7.8 Hz,
1H). 13C NMR (200 MHz, CD3OD) d (ppm) 7.09,
12.13, 55.72, 79.00, 115.33, 115.51, 122.37, 130.52,
138.58, 161.30, 173.81. Anal. (C12H15NO3) calcd C
65.15 H 6.83 N 6.33; F. C 65.01 H 6.94 N 6.27.
Preparation of the N-acyl-O-alkylhydroxylamines—
general procedure
N-[(3-Methoxy-1-phenyl)methoxy]acetamide (10a). The
amine 9a (0.3 g, 1.9 mmol) was dissolved in a biphasic
system CH2Cl2/H2O (10 mL/10 mL) in the presence of
Na2CO3 (1.41 g, 13.3 mmol). Acetyl chloride (135 mL,
13.3 mmol) was added slowly while stirring at 0 ꢁC. The
mixture was stirred at room temperature for 3 h. The
organic phase was separated, washed with a 1 N HCl
solution and a NaCl saturated solution. The organic
solution was dried over MgSO4 and evaporated under
vacuum. The residue was purified by column chroma-
tography (silica gel) and eluted with CH2Cl2 and a
CH2Cl2/MeOH mixture (97/3) to give a yellow oil
N-[(2,5-Dimethoxy-1-phenyl)methoxy]acetamide (11a). It
was prepared according to the previous method descri-
bed for 10a with acetyl chloride. The compound was
isolated as a white solid (yield: 59%) mp59 ꢁC. 1H
NMR (200 MHz, CD3OD) d (ppm) 1.82 (s, 3H), 3.74–
3.76 (2s, 6H), 4.85 (s, 2H), 6.81–6.98 (m, 3H). 13C NMR
(200 MHz, CD3OD) d (ppm) 19.48, 56.19, 56.64, 73.60,
113.02, 115.71, 117.61, 126.06, 153.58, 155.00, 169.97.
Anal. (C11H15NO4) calcd C 58.66 H 6.71 N 6.22; F. C
58.68 H 6.73 N 6.11.
1
(0.13 g, yield: 34%). H NMR (200 MHz, CD3OD) d
(ppm) 1.82 (s, 3H), 3.77 (s, 3H), 4.79 (s, 2H), 6.86–6.98
(m, 3H), 7.26 (t, J=7.8 Hz, 1H). 13C NMR (200 MHz,
CD3OD) d (ppm) 19.46, 55.72, 78.91, 115.35, 115.48,
122.34, 130.52, 138.52, 170.12. Anal. (C10H13NO3) calcd
C 60.96 H 6.70 N 7.11; F. C 60.90 H 6.97 N 6.80.
N - [(2,5 - Dimethoxy - 1 - phenyl)methoxy]propionamide
(11b). It was prepared according to the previous
method described for 10a with propionyl chloride. The
compound was isolated as a white solid (yield: 61%) mp
74 C. H NMR (200 MHz, CD3OD) d (ppm) 1.09 (t,
J=7.6 Hz, 3H), 2.05 (q, J=7.6 Hz, 2H), 3.74–3.76 (2s,
6H), 4.85 (s, 2H), 6.81–6.97 (m, 3H). 13C NMR
(200 MHz, CD3OD) d (ppm) 10.29, 27.4, 56.21, 56.65,
73.55, 113.03, 115.73, 117.69, 126.09, 153. 61, 154.98,
173.80. Anal. (C12H17NO4) calcd C 60.24 H 7.16 N 5.85;
F. C 60.13 H 7.18 N 5.83.
ꢁ
1
N-[(3-Methoxy-1-phenyl)methoxy]propionamide (10b). It
was prepared according to the previous method descri-
bed for 10a with propionyl chloride. The compound was
isolated as a yellow oil (yield: 42%). 1H NMR
(200 MHz, CD3OD) d (ppm) 1.09 (t, J=7.6 Hz, 3H),
2.05 (q, J=7.6 Hz, 2H), 3.79 (s, 3H), 4.79 (s, 2H), 6.86–
6.97 (m, 3H), 7.26 (t, J=7.8 Hz, 1H). 13C NMR
(200 MHz, CD3OD) d (ppm) 10.23, 27.11, 55.73, 78.87,
115.35, 115.50, 122.42, 130.50, 138.49, 161.30, 173.85.
Anal. (C11H15NO3) calcd C 63.14 H 7.22 N 6.69; F. C
62.95 H 7.31 N 6.54.
N-[(2,5-Dimethoxy-1-phenyl)methoxy]butyramide (11c).
It was prepared according to the previous method
described for 10a with butyryl chloride. The compound
was isolated as a white solid (yield: 66%) mp71 ꢁC. H
1
NMR (200 MHz, CD3OD) d (ppm) 0.90 (t, J=7.3 Hz,
3H), 2.05 (sx, J=7.3 Hz, 2H), 3.74–3.77 (2s, 6H), 4.86
(s, 2H), 6.81–6.98 (m, 3H). 13C NMR (200 MHz,
CD3OD) d (ppm) 13.88, 20.10, 35.72, 56.19, 56.62,
73.58, 112.99, 115.72, 117.63, 126.08, 153. 62, 154.98,
172.64. Anal. (C13H19NO4) calcd C 61.64 H 7.56 N 5.53
F. C 61.64 H 7.61 N 5.45.
N-[(3-Methoxy-1-phenyl)methoxy]butyramide (10c). It
was prepared according to the previous method descri-
bed for 10a with butyryl chloride. The compound was
isolated as a colourless oil (yield: 86%). 1H NMR
(200 MHz, CD3OD) d (ppm) 0.89 (t, J=7.4 Hz, 3H),
1.59 (sx, J=7.3 Hz, 2H), 2.01 (t, J=7.3 Hz, 2H), 3.79 (s,
3H), 4.80 (s, 2H), 6.86–6.98 (m, 3H), 7.25 (t, J=7.8 Hz,
1H). 13C NMR (200 MHz, CD3OD) d (ppm) 13.85,
20.09, 35.70, 55.73, 78.90, 115.38, 115.47, 122.42,
130.51, 138.52, 161.30, 172.82. Anal. (C12H17NO3) calcd
C 64.55 H 7.67 N 6.27; F. C 64.64 H 7.80 N 6.20.
N - [(2,5 - Dimethoxy - 1 - phenyl)methoxy]chloracetamide
(11e). It was prepared according to the previous
method described for 10a with chloroacetyl chloride.
The compound was isolated as a white solid (yield:
ꢁ
30%) mp72 C. 1H NMR (200 MHz, CD3OD) d (ppm),
N-[(3-Methoxy-1-phenyl)methoxy]chloracetamide (10d).
It was prepared according to the previous method
described for 10a with chloracetyl chloride. The com-
pound was isolated as a white solid (yield: 86%) mp
3.74–3.77 (2s, 6H), 3.91 (s, 2H), 4.89 (s, 2H), 6.87–6.97
(m, 3H). 13C NMR (200 MHz, CD3OD) d (ppm) 40.89,
56.22, 56.66, 73.68, 113.08, 116.01, 117.79, 125.70, 153.
68, 154.99, 166.75. Anal. (C11H14NO4Cl) calcd C
50.88 H 5.43 N 5.39; F. C 50.84 H 5.46 N 5.19.
88 ꢁC. H NMR (200 MHz, CD3OD) d (ppm) 3.78 (s,
1
3H); 3.92 (s, 2H), 4.83 (s, 2H), 6.86–6.98 (m, 3H), 7.26
(t, J=7.8 Hz, 1H). 13C NMR (200 MHz, CD3OD) d
(ppm) 40.90, 55.78, 79.00, 115.38, 115.57, 122.53,
130.59, 138.12, 161.31, 166.37. Anal. (C10H12NO3Cl)
calcd C 52.30 H 5.27 N 6.10; F. C 52.28 H 5.34 N 6.03.
N - [(2,5 - Dimethoxy - 1 - phenyl)methoxy]cyclopropylcar-
boxamide (11f). It was prepared according to the
previous method described for 10a with cyclo-
propylcarbonyl chloride. The compound was isolated
as a white solid (yield: 84%) mp83 ꢁC. 1H NMR
(200 MHz, CD3OD) d (ppm) 0.67–0.90 (m, 4H), 1.33–
1.43 (m, 1H), 3.74–3.77 (2s, 6H), 4.85 (s, 2H), 6.82–6.97
N - [(3- Methoxy - 1 - phenyl)methoxy]cyclopropylcarbox-
amide (10e). It was prepared according to the previous

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C. Pegurier et al. / Bioorg. Med. Chem. 11 (2003) 789–800
796
(m, 3H). 13C NMR (200 MHz, CD3OD) d (ppm) 7.05,
12.15, 56.19, 56.64, 73.64, 113.01, 115.69, 117.60,
126.57, 154.61, 155.00, 174.54. Anal. (C13H17NO4) calcd
C 62.14 H 6.82 N 5.57; F. C 62.17 H 6.88 N 5.57.
114.87, 120.21, 130.54, 161.37, 173.89. Anal.
(C13H17NO3) calcd C 66.36 H 7.28 N 5.95; F. C 66.23 H
7.39 N 5.94.
N-[(3-Methoxyphenethyl)oxy]acetamide (13a). It was
prepared according to the previous method described
for 10a with acetyl chloride. The compound was iso-
lated as a colourless oil (yield: 70%). 1H NMR
(200 MHz, CD3OD) d (ppm) 1.84 (s, 3H), 2.90 (t,
J=7.0 Hz, 2H), 3.75 (s, 3H), 4.01 (t, J=7.0 Hz, 2H),
6.71–6.81 (m, 3H), 7.17 (t, J=8.2 Hz, 1H). 13C NMR
(200 MHz, CD3OD) d (ppm) 19.48, 35.52, 55.64, 77.95,
112.95, 115.64, 122.27, 130.46, 140.94, 161.33, 170.16.
Anal. (C11H15NO3) calcd C 63.14 H 7.22 N 6.69; F. C
62.89 H 7.35 N 6.57.
N-[1-(3-Methoxyphenyl)ethoxy]acetamide (12a). It was
prepared according to the previous method described
for 10a with acetyl chloride. The compound was iso-
lated as a white foam (yield: 50%) mp93 ꢁC. H NMR
1
(200 MHz, CD3OD) d (ppm) 1.48 (d, J=6.5 Hz, 3H),
1.75 (s, 3H), 3.79 (s, 6H), 4.89 (q, J=6.5 Hz, 1H), 6.83–
6.94 (m, 3H), 7.25 (t, J=7.8 Hz, 1H). 13C NMR
(200 MHz, CD3OD) d (ppm) 19.39, 21.50, 55.78, 84. 47,
113.30, 114.97, 120.26, 130.61, 144.61, 161.42, 174.33.
Anal. (C11H15NO3) calcd C 63.14 H 7.22 N 6.69; F. C
63.16 H 7.27 N 6.60.
N-[(3-Methoxyphenethyl)oxy]propionamide (13b). It was
prepared according to the previous method described
for 10a with propionyl chloride. The compound was
isolated as a colourless oil (yield: 85%). 1H NMR
(200 MHz, CD3OD) d (ppm) 1.11 (t, J=7.6 Hz), 2.08 (q,
J=7.6 Hz), 2.90 (t, J=7.0 Hz, 2H), 3.75 (s, 3H), 4.01 (t,
J=7.1 Hz, 2H), 6.71–6.82 (m, 3H), 7.17 (t, J=8.1 Hz,
1H). 13C NMR (200 MHz, CD3OD) d (ppm) 10.24,
27.18, 35.61, 55.69, 77.98, 112.99, 115.70, 122.32,
130.51, 141.03, 161.37, 173.98. Anal. (C12H17NO3) calcd
C 64.55 H 7.67 N 6.27; F. C 64.60 H 7.70 N 6.20.
N-[1-(3-Methoxyphenyl)ethoxy]propionamide (12b). It
was prepared according to the previous method descri-
bed for 10a with propionyl chloride. The compound was
isolated as a white foam (yield: 72%) mp84 ꢁC. 1H
NMR (200 MHz, CD3OD) d (ppm) 1.03 (t, J=7.6 Hz),
1.48 (d, J=6.5 Hz, 3H), 1.98 (q, J=7.6 Hz, 2H), 3.79 (s,
6H), 4.89 (q, J=6.6 Hz, 1H), 6.83–6.94 (m, 3H), 7.25 (t,
J=7.8 Hz, 1H). 13C NMR (200 MHz, CD3OD) d (ppm)
10.35, 21.36, 27.07, 55.72, 84. 30, 113.33, 114.93, 120.29,
130.53, 144.26, 161.35, 173.78. Anal. (C12H17NO3) calcd
C 64.55 H 7.67 N 6.27; F. C 64.42 H 7.76 N 6.12.
N-[2-(3-Methoxyphenethyl)oxy]butyramide (13c). It was
prepared according to the previous method described
for 10a with butyryl chloride. The compound was
isolated as a colourless oil (yield: 88%). 1H NMR
(200 MHz, CD3OD) d (ppm) 0.98 (t, J=7.4 Hz), 1.62
(sx, J=7.3 Hz, 2H), 2.04 (t, J=7.3 Hz, 2H), 2.90 (t,
J=7.0 Hz, 2H), 3.75 (s, 3H), 4.01 (t, J=7.1 Hz, 2H),
6.71–6.82 (m, 3H), 7.16 (t, J=8.1 Hz, 1H). 13C NMR
(200 MHz, CD3OD) d (ppm) 13.90, 20.07, 35.57, 35.73,
55.64, 77.98, 112.94, 115.65, 122.27, 130.47, 161.33,
172.90. Anal. (C13H19NO3) calcd C 65.80 H 8.07 N 5.90;
F. C 65.67 H 8.10 N 5.80.
N-[1-(3-Methoxyphenyl)ethoxy]butyramide (12c). It was
prepared according to the previous method described
for 10a with butyryl chloride. The compound was iso-
lated as a white foam (yield: 81%) mp82 ꢁC. H NMR
1
(200 MHz, CD3OD) d (ppm) 0.82 (t, J=7.3 Hz, 3H),
1.46–1.59 (m, 5H), 1.95 (t, J=7.2 Hz, 2H), 3.78 (s, 3H),
4.90 (q, J=6.5 Hz, 1H), 6.83–6.94 (m, 3H), 7.24 (t,
J=7.8 Hz, 1H). 13C NMR (200 MHz, CD3OD) d (ppm)
13.79, 35.64, 55.73, 84. 37, 113.32, 114.97, 120.30,
130.53, 144.79, 161.84, 174.61. Anal. (C13H19NO3) calcd
C 65.80 H 8.07 N 5.92; F. C 65.64 H 8.08 N 5.85.
N-[1-(3-Methoxyphenyl)ethoxy]chloracetamide (12d). It
was prepared according to the previous method descri-
bed for 10a with chloroacetyl chloride. The comꢁpound
was isolated as a white foam (yield: 48%) mp100 C. 1H
NMR (200 MHz, CD3OD) d (ppm) 1.50 (d, J=6.5 Hz,
3H), 3.79 (s, 3H), 3.84 (s, 2H), 4.95 (q, J=6.5 Hz, 1H),
6.83–6.96 (m, 3H), 7.26 (t, J=7.8 Hz, 1H). 13C NMR
(200 MHz, CD3OD) d (ppm) 21.23, 40.72, 55.67, 84. 54,
113.27, 115.07, 120.22, 130.55, 143.80, 161.34, 166.31.
Anal. (C11H14NO3Cl) calcd C 54.22 H 5.79 N 5.75; F. C
54.31 H 5.83 N 5.75.
N-[(3-Methoxyphenethyl)oxy]chloracetamide (13d). It
was prepared according to the previous method descri-
bed for 10a with chloracetyl chloride. The compound
was isolated as a white solid after recrystallisation in an
AcOEt/hexane mixture (yield: 59%) mp62 ꢁC. 1H
NMR (200 MHz, CD3OD) d (ppm) 2.92 (t, J=7.0 Hz,
2H), 3.75 (s, 3H), 3.95 (s, 2H), 4.01 (t, J=7.1 Hz, 2H),
6.72–6.83 (m, 3H), 7.17 (t, J=7.9 Hz, 1H). 13C NMR
(200 MHz, CD3OD) d (ppm) 35.52, 40.97, 55.71, 77.99,
113.04, 115.68, 122.31, 130.54, 161.32, 166.40. Anal.
(C11H14NO3Cl) calcd C 54.22 H 5.79 N 5.75; F. C
54.36 H 5.98 N 5.64.
N-[1-(3-Methoxyphenyl)ethoxy]cyclopropylcarboxamide
(12e). It was prepared according to the previous
method described for 10a with propionyl chloride. The
compound was isolated as a white foam after recrys-
tallisation in an AcOEt/Hexane mixture (yield: 86%)
N - [(3- Methoxyphenethyl)oxy]cyclopropylcarboxamide
(13e). It was prepared according to the previous
method described for 10a with cyclopropanecarbonyl
chloride. The compound was isolated as a white solid
after recrystallisation in an AcOEt/hexane mixture
ꢁ
mp126 C. 1H NMR (200 MHz, CD3OD) d (ppm) 0.66–
(yield: 69%) mp86 ꢁC. H NMR (200 MHz, CD3OD) d
1
0.86 (m, 4H), 1.28–1.38 (m, 1H), 1.48 (d, J=6.5 Hz,
3H), 3.79 (s, 3H), 4.88 (q, J=6.5 Hz, 1H), 6.83–6.95 (m,
3H), 7.26 (t, J=7.8 Hz, 1H). 13C NMR (200 MHz,
CD3OD) d (ppm) 12.03, 21.42, 55.71, 84. 42, 113.27,
(ppm) 0.72–0.91 (m, 4H), 1.39–1.42 (m, 1H), 2.90 (t,
J=7.0 Hz, 2H), 3.75 (s, 3H), 4.01 (t, J=7.1 Hz, 2H),
6.71–6.82 (m, 3H), 7.17 (t, J=7.9 Hz, 1H). 13C NMR

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C. Pegurier et al. / Bioorg. Med. Chem. 11 (2003) 789–800
797
(200 MHz, CD3OD) d (ppm) 7.26, 12.29, 35.61, 55.69,
78.09, 112.96, 115.70, 122.33, 130.51, 141.01, 161.32,
174.10. Anal. (C11H17NO3) calcd C 66.36 H 7.28 N 5.95;
F. C 66.24 H 7.38 N 5.92.
N-[(7-Methoxy-1,2,3,4-tetrahydro-1-naphthalenyl)oxy]-
cyclopropylcarboxamide (14e). It was prepared accord-
ing to the previous method described for 10a with
cyclopropylcarbonyl chloride. The compound was iso-
lated as a white solid after recrystallisation in an
AcOEt/hexane mixture (yield: 68%) mp160 ꢁC. 1H
NMR (200 MHz, CD3OD) d (ppm) 0.76–0.91 (m, 4H),
1.32–1.51(m, 1H), 1.70–2.25 (m, 4H), 2.54–2.79 (m, 2H),
3.77( s, 3H), 4.80 (t, J=3.7 Hz, 2H), 6.79 (dd,
J=8.4 Hz, J=2.8 Hz, 1H), 7.00 (d, J=8.45 Hz, 1H),
7.13 (d, J=2.3 Hz, 1H). 13C NMR (200 MHz, CD3OD)
d (ppm) 7.03, 12.25, 19.61, 28.14, 29.38, 35.90, 55.76,
82.21, 116.09, 116.47, 130.78, 131.58, 135.65, 159.14,
174.36. Anal. (C15H19NO3) calcd C 68.94 H 7.31 N 5.36;
F. C 68.75 H 7.38 N 5.32.
N-[(7-Methoxy-1,2,3,4-tetrahydro-1-naphthalenyl)oxy]-
acetamide (14a). It was prepared according to the pre-
vious method described for 10a with acetyl chloride.
The compound was isolated as a white solid after
recrystallisation in an AcOEt/hexane mixture (yield:
53%) mp70 ꢁC. H NMR (200 MHz, CD3OD) d (ppm)
1
1.69–2.77 (m, 6H), 1.88 (s, 3H), 3.76 (s, 3H), 4.80 (t,
J=3.7 Hz, br s, 2H), 6.79 (dd, J=8.4 Hz, J=2.7 Hz,
1H), 6.99 (d, J=8.4.7 Hz, 1H), 7.03 (d, J=2.6 Hz, 1H).
13C NMR (200 MHz, CD3OD) d (ppm) 20.63, 29.17,
30.38, 56.78, 83.26, 117.11, 117.40, 131.79, 136.51,
160.13, 171.25. Anal. (C13H17NO3) Calc. C 66.36 H
7.28 N 5.95; F. C 66.19 H 7.36 N 5.88.
N-[7-Methoxy-1-naphthyl)methoxy]acetamide (15a). It
was prepared according to the previous method descri-
bed for 10a with acetyl chloride. The compound was
isolated as a white solid after recrystallisation in an
Et2O/CH2Cl2 mixture (yield: 90%) mp125 ꢁC. 1H
NMR (200 MHz, CDCl3) d (ppm) 1.85 (s, 3H), 3.95 (2s,
3H), 5.30 (s, 2H), 7.10–7.80 (m, 6H), 8.45 (s, 1H). Anal.
(C14H15NO3) calcd C 68.57 H 6.17 N 5.72; F. C 68.48 H
6.29 N 5.68.
N-[(7-Methoxy-1,2,3,4-tetrahydro-1-naphthalenyl)oxy]-
propionamide (14b). It was prepared according to the
previous method described for 10a with propionyl
chloride. The compound was isolated as a white solid
after recrystallisation in an AcOEt/hexane mixture
(yield: 77%) mp113 ꢁC. H NMR (200 MHz, CD3OD)
1
d (ppm) 1.15 (t, J=7.6 Hz, 3H), 1.70–2.24 (m, 4H), 2.13
(q, J=7.6 Hz, 2H), 2.53–2.79 (m, 2H), 3.77( s, 3H), 4.81
(t, J=3.7 Hz, 2H), 6.79 (dd, J=8.4 Hz, J=2.8 Hz, 1H),
7.00 (d, J=8.5 Hz, 1H), 7.13 (d, J=2.5 Hz, 1H). 13C
NMR (200 MHz, CD3OD) d (ppm) 19.60, 27.26, 28.15,
29.36, 55.74, 82.19, 116.10, 116.38, 130.75, 131.55,
135.59, 159.10, 173.07. Anal. (C14H19NO3) calcd C
67.45 H 7.68 N 5.62; F. C 67.24 H 7.76 N 5.73.
N-[7-Methoxy-1-naphthyl)methoxy]butyramide (15c). It
was prepared according to the previous method descri-
bed for 10a with butyryl chloride. The compound was
isolated as a white solid after recrystallisation in an
Et2O/CH2Cl2 mixture (yield: 90%) mp98 ꢁC. H NMR
1
(200 MHz, CDCl3) d (ppm) 0.95 (t, J=7 Hz, 3H), 1.75
(m, 2H), 2.10 (m, 2H), 4.00 (s, 3H), 7.15–7.85 (m, 6H),
8.20 (s, 1H). Anal. (C16H19NO3) calcd C 70.33 H 7.01 N
5.13; F. C 70.33 H 7.10 N 5.08.
N-[(7-Methoxy-1,2,3,4-tetrahydro-1-naphthalenyl)oxy]-
butyramide (14c). It was prepared according to the pre-
vious method described for 10a with butyryl chloride. The
compound was isolated as a white solid after recrystallisa-
N-[7-Methoxy-1-naphthyl)methoxy]cyclopropylcarboxa-
mide (15e). It was prepared according to the previous
method described for 10a with cyclopropylcarbonyl
chloride. The compound was isolated as a white solid
after recrystallisation in cyclohexane (yield: 90%) mp
ꢁ
tion in an AcOEt/Hexane mixture (yield: 32%) mp82 C.
1H NMR (200MHz, CD3OD) d (ppm) 0.95 (t, J=7.2 Hz,
2H), 1.6 (sx, J=7.4 Hz, 2H), 1.56–2.26 (m, 4H), 2.09 (t,
J=7.2 Hz, 2H), 2.52–2.81 (m, 2H), 3.77( s, 3H), 4.81 (t,
J=3.7 Hz, 2H), 6.79 (dd, J=8.4 Hz, J=2.8 Hz, 1H), 6.99
(d, J=8.45 Hz, 1H), 7.13 (d, J=2.6Hz, 1H). 13C NMR
(200MHz, CD3OD) d (ppm) 19.62, 20.17, 28.16, 29.38,
35.90, 55.76, 82.23, 116.10, 116.408, 130.76, 131.55, 135.59,
159.11, 173.06. Anal. (C15H21NO3) calc. C 68.42 H 8.04N
5.32; F. C 68.23 H 7.98 N 5.52.
ꢁ
1
125 C. H NMR (200 MHz, CDCl3) d (ppm) 0.65–1.20
(m, 5H), 3.95 (s, 3H), 5.30 (s, 2H), 7.15–7.80 (m, 5H),
8.40 (s, 1H). Anal. (C16H17NO3) calcd C 70.83 H
76.32 N 5.16; F. C 70.76 H 6.47 N 5.11.
N-[2-Methoxy-1-naphthyl)methoxy]acetamide (16a). It
was prepared according to the previous method descri-
bed for 10a with acetyl chloride. The compound was
isolated as a white solid after recrystallisation in an
Et2O/CH2Cl2 mixture (yield: 39%) mp112 ꢁC. 1H
NMR (200 MHz, CDCl3) d (ppm) 1.9 (s, 3H), 3.95 (s,
3H), 5.45 (s, 2H), 7.10–8.40 (m, 6H), 8.50 (s, 1H). Anal.
(C14H15NO3) calcd C 68.57 H 6.17 N 5.72; F. C 68.41 H
6.27 N 5.73.
N-[(7-Methoxy-1,2,3,4-tetrahydro-1-naphthalenyl)oxy]-
chloracetamide (14d). It was prepared according to the
previous method described for 10a with chloroacetyl
chloride. The compound was isolated as a white solid
after recrystallisation in an AcOEt/Hexane mixture
(yield: 21%) mp91 ꢁC. H NMR (200 MHz, CD3OD) d
1
(ppm) 1.70–2.27 (m, 4H), 2.61–2.79 (m, 2H), 3.77( s,
3H), 3.97 (s, 2H), 4.84 (t, J=3.7 Hz, 2H), 6.80 (dd,
J=8.4 Hz, J=2.8 Hz, 1H), 7.00 (d, J=8.4 Hz, 1H), 7.12
(d, J=2.4 Hz, 1H). 13C NMR (200 MHz, CD3OD) d
(ppm) 19.55, 28.15, 29.33, 41.05, 55.79, 82.50, 116.07,
116.58, 130.83, 131.61, 135.19, 159.13, 166.58. Anal.
(C13H161NO3Cl) calcd C 57.89 H 5.98 N 5.19; F. C
58.08 H 6.10 N 5.03.
N-[2-Methoxy-1-naphthyl)methoxy]propionamide (16b).
It was prepared according to the previous method
described for 10a with propionyl chloride. The com-
pound was isolated as a white solid after recrystallisa-
tion in Et2O (yield: 39%) mp: 118 ꢁC. 1H NMR
(200 MHz, CDCl3) d (ppm) 1.15 (t, 3H), 1.9–2.5 (m,
2H), 3.90 (s, 3H), 5.50 (s, 2H), 7.10–8.40 (m, 6H), 8.80

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C. Pegurier et al. / Bioorg. Med. Chem. 11 (2003) 789–800
798
(s, 1H). Anal. (C15H17NO3) calcd C 69.47 H 6.62 N
5.40; F. C 69.25 H 6.71 N 5.34.
The compound was isolated as a beige solid after
recrystallisation in an AcOEt/hexane mixture (yield:
67%) mp122 ꢁC. 1H NMR (200 MHz, CD3OD) d
(ppm), 3.92–3.94 (2s, 6H), 3.96 (s, 2H), 5.41 (s, 2H),
6.96 (dd, J=8.9 Hz, J=2.4 Hz, 1H), 7.18 (d, J=9.0 Hz,
1H), 7.65 (d, J=8.9 Hz, 1H), 7.73 (d, J=2.0 Hz, 1H),
7.79 (d, J=9.0 Hz, 1H). 13C NMR (200 MHz, CD3OD)
d (ppm) 56.11, 57.05, 69.25, 103.40, 111.45,115.62,
117.71, 125.94, 130.76, 132.26, 137.11, 158.67, 160.43,
166.45. Anal. (C15H16NO4Cl) calcd C 58.17 H 5.21 N
4.52; F. C 58.23 H 5.23 N 4.48.
N-[2-Methoxy-1-naphthyl)methoxy]cyclopropylcarbox-
amide (16e). It was prepared according to the previous
method described for 10a with cyclopropylcarbonyl
chloride. The compound was isolated as a white solid
after recrystallisation in Et2O (yield: 39%) mp145 ꢁC.
1H NMR (200 MHz, CDCl3) d (ppm) 0.51–1.30 (m,
5H), 3.95 (s, 3H), 5.50 (s, 2H), 7.10–8.20 (m, 6H), 8.40
(s, 1H). Anal. (C16H17NO3) calcd C 70.82 H 6.32 N
5.16; F. C 70.98 H 6.52 N 5.11.
N-[2,7-Dimethoxy-1-naphthyl)methoxy]cyclopropylcar-
boxamide (17e). It was prepared according to the pre-
N-[2,7-Dimethoxy-1-naphthyl)methoxy]acetamide (17a).
It was prepared according to the previous method
described for 10a with acetyl chloride. The compound
was isolated as a beige solid after recrystallisation in an
AcOEt/hexane mixture (yield: 39%) mp143 ꢁC. 1H
NMR (200 MHz, CD3OD) d (ppm) 1.85 (s, 3H), 3.93–
3.95 (2s, 6H), 5.38 (s, 2H), 6.96 (dd, J=8.9 Hz,
J=2.4 Hz, 1H), 7.19 (d, J=9.0 Hz, 1H), 7.66 (d,
J=8.8 Hz, 1H), 7.73 (d, J=2.3 Hz, 1H), 7.80 (d,
J=9.0 Hz, 1H). 13C NMR (200 MHz, CD3OD) d (ppm)
19.59, 56.07, 57.09, 62.73, 103.42, 111.52, 117.66,
130.74, 132.12, 125.96, 136.60, 137.25, 158.61, 160.35,
170.13. Anal. (C15H17NO4) calcd C 65.44 H 6.22 N 5.09;
F. C 65.10 H 6.28 N 4.80.
vious
method
described
for
10a
with
cyclopropylcarbonyl chloride. The compound was iso-
lated as a beige solid after recrystallisation in an AcOEt/
hexane mixture (yield: 67%) mp149 ꢁC. 1H NMR
(200 MHz, CD3OD) d (ppm) 0.72–0.79 (m, 2H), 1.32–
1.42 (m, 1H), 3.91–3.93 (2s, 6H), 5.36 (s, 2H), 6.96 (dd,
J=9.0 Hz, J=2.4 Hz, 1H), 7.16 (d, J=9.0 Hz, 1H),
7.62–7.67 (m, 2H), 7.77 (d, J=9.0 Hz, 1H). 13C NMR
(200 MHz, CD3OD) d (ppm) 5.09, 7.00, 12.27, 56.01,
57.09, 69.14, 103.45, 111.56, 116.14, 117.56, 125.99,
130.76, 132.07, 137.06, 158.78, 160.32, 173.99. Anal.
(C17H19NO4) calcd C 67.76 H 6.35 N 4.65; F. C 67.65 H
6.42 N 4.62.
N - [2,7 - Dimethoxy - 1 - naphthyl)methoxy]propionamide
(17b). It was prepared according to the previous
method described for 10a with propionyl chloride. The
compound was isolated as a beige solid after recrys-
tallisation in an AcOEt/hexane mixture (yield: 60%) mp
N-[1,2-Dihydro-1-acenaphthylenyloxy)acetamide (18a).
It was prepared according to the previous method
described for 10a with acetyl chloride. The compound
was isolated as a white foam after recrystallisation in an
AcOEt/hexane mixture (yield: 71%) mp125 ꢁC. 1H
NMR (200 MHz, CD3OD) d (ppm) 1.87 (s, 3H), 3.45 (d,
J=17 Hz, 1H), 3.66 (dd, J=17.7 Hz, J=6.8 Hz, 1H),
5.89 (d, J=5.1 Hz), 7.30 (d, J=6.8 Hz, 1H), 7.43–7.67
(m, 4H), 7.78 (d, J=8.0 Hz, 1H). 13C NMR (200 MHz,
CD3OD) d (ppm) 19.59, 38.01, 87.44, 120.88, 123.26,
123.81, 126.75, 128.86, 129.25, 132.77, 139.49, 142.26,
142.70, 170.69. Anal. (C14H13NO2) calcd C 73.99 H
5.76 N 6.16; F. C 73.81 H 5.92 N 6.08.
121 ꢁC. H NMR (200 MHz, CD3OD) d (ppm) 1.12 (t,
1
J=7.6 Hz, 3H), 2.07 (q, J=7.6 Hz, 2H), 3.92–3.95 (2s,
6H), 5.37 (s, 2H), 6.97 (dd, J=8.9 Hz, J=2.4 Hz, 1H),
7.18 (d, J=9.0 Hz, 1H), 7.65 (d, J=9.0 Hz, 1H), 7.70 (d,
J=2.2 Hz, 1H), 7.79 (d, J=9.0 Hz, 1H). 13C NMR
(200 MHz, CD3OD) d (ppm) 10.33, 27.29, 56.05, 57.04,
69.04, 103.49, 111.49, 116.02, 117.61, 130.76, 132.10,
125.97, 137.12, 158.61, 160.33, 174.52. Anal.
(C16H19NO4) calcd C 66.42 H 6.62 N 4.84; F. C 66.38 H
6.67 N 4.76.
N - [1,2 - Dihydro - 1 - acenaphthylenyloxy)propionamide
(18b). It was prepared according to the previous
method described for 10a with propionyl chloride. The
compound was isolated as a white foam after recrys-
tallisation in an AcOEt/hexane mixture (yield: 81%) mp
N - [2,7 - Dimethoxy - 1 - naphthyl)methoxy]butyramide
(17c). It was prepared according to the previous
method described for 10a with butyryl chloride. The
compound was isolated as a white solid after recrys-
tallisation in an AcOEt/hexane mixture (yield: 61%) mp
137 ꢁC H NMR (200 MHz, CD3OD) d (ppm) 1.08 (t,
1
J=7.6 Hz, 3H), 2.08 (q, J=7.6 Hz), 3.44 (d, J=17.8 Hz,
1H), 3.44 (d, J=17.8 Hz, 1H), 3.60 (dd, J=18.2 Hz,
J=6.6 Hz, 1H), 5.84 (d, J=4.9 Hz, 1H), 7.27 (d,
J=6.7 Hz, 1H), 7.40–7.64 (m, 4H), 7.76 (d, J=8.0 Hz,
1H). 13C NMR (200 MHz, CD3OD) d (ppm) 10.27,
27.22, 38.03, 87.42, 120.87, 123.29, 123.80, 126.74,
128.84, 129.24, 132.74, 139.51, 142.27, 142.73, 174.48.
Anal. (C15H15NO2) calcd C 74.67 H 6.27 N 5.80; F. C
74.53 H 6.41 N 5.75.
86 ꢁC. H NMR (200 MHz, CD3OD) d (ppm), 0.93 (t,
1
J=7.3 Hz, 3H), 1.62 (sx, J=7.3 Hz, 2H), 2.03 (t,
J=7.3 Hz, 2H), 3.88–3.94 (2s, 6H), 5.34 (s, 2H), 6.95
(dd, J=8.9 Hz, J=2.3 Hz, 1H), 7.13 (d, J=9.0 Hz, 1H),
7.62 (d, J=9.0 Hz, 1H), 7.68 (d, J=2.1 Hz, 1H), 7.75 (d,
J=9.0 Hz, 1H). 13C NMR (200 MHz, CD3OD) d (ppm)
13.96, 20.18, 35.93, 56.07, 57.02, 69.17, 103.52, 111.45,
115.99, 117.61, 125.95, 130.75, 132.09, 125.97, 137.10,
158.55, 160.32, 172.91. Anal. (C17H21NO4) calcd C
67.31 H 6.98 N 4.62; F. C 67.02 H 7.00 N 4.55.
N-[1,2-Dihydro-1-acenaphthylenyloxy)butyramide (18c).
It was prepared according to the previous method
described for 10a with butyryl chloride. The compound
was isolated as a white solid after recrystallisation in an
AcOEt/hexane mixture (yield: 63%) mp105 ꢁC. 1H
N-[2,7-Dimethoxy-1-naphthyl)methoxy]chloracetamide
(17d). It was prepared according to the previous
method described for 10a with chloroacetyl chloride.

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C. Pegurier et al. / Bioorg. Med. Chem. 11 (2003) 789–800
799
NMR (200 MHz, CD3OD) d (ppm) 0.88 (t, J=7.4 Hz,
3H), 1.59 (sx, J=7.3 Hz, 2H), 2.05 (t, J=7.3 Hz, 2H),
3.46 (d, J=17.8 Hz, 1H), 3.446 (d, J=17.8 Hz, 1H), 3.65
(dd, J=18.3 Hz, J=6.7 Hz, 1H), 5.89 (d, J=4.9 Hz,
1H), 7.30 (d, J=6.4 Hz, 1H), 7.43–7.66 (m, 4H), 7.78 (d,
J=8.0 Hz, 1H). 13C NMR (200 MHz, CD3OD) d (ppm)
13.89, 20.12, 35.81, 38.03, 87.42, 120.87, 123.29, 123.79,
126.74, 128.85, 129.23, 132.46, 139.21, 142.33, 143.01,
173.32. Anal. (C16H17NO2) calc. C 75.27 H 6.71 N 5.48;
F. C 75.25 H 6.80 N 5.40.
with buffer, dried, and counted on
a
g-counter
(Crystal-Packard). Non-specific binding was defined
with 10 mM 2-iodomelatonin and represented 10% of
the total binding. Ki values were determined using the
Cheng–Prussof equation.
Melatonin receptor binding assay with human MT1 and
MT2 receptors. Competitive inhibition curves were per-
formed according to the methods already described22
with the receptors expressed in HEK-293 cells with
[
125I]iodomelatonin as the radioligand. Membranes
N - [1,2 - dihydro - 1 - acenaphthylenyloxy)chloracetamide
(18d). It was prepared according to the previous
method described for 10a with chloroacetyl chloride.
The compound was isolated as a white solid after
recrystallisation in an AcOEt/hexane mixture (yield:
72%) mp: 132 ꢁC. 1H NMR (200 MHz, CD3OD) d
(ppm) 3.47 (d, J=17.8 Hz, 1H), 3.68 (dd, J=178.2 Hz,
J=6.7 Hz, 1H), 3.3.98 (s, 2H), 5.93 (d, J=6.1 Hz, 1H),
7.32 (d, J=6.9 Hz, 1H), 7.44–7.68 (m, 4H), 7.80 (d,
J=8.1 Hz, 1H). 13C NMR (200 MHz, CD3OD) d (ppm)
38.01, 40.96, 87.62, 120.95, 123.38, 123.85, 126.85,
128.84, 129.28, 132.81, 140.01, 142.62, 145.01, 162.50.
Anal. (C14H12NO2Cl) calc. C 64.25 H 4.62 N 5.35; F. C
64.17 H 4.77 N 5.26.
obtained from HEK-293 cells expressing hMT1 or
hMT2 receptors were incubated in Tris–HCl buffer
(50 mM, pH 7.4) with 0.025 nM or 0.2 nM of
2-[125I]iodomelatonin, respectively and seven con-
centrations of the compound under test. Each bind-
ing assay was performed in triplicate. The incubation
(37 ꢁC, 2 h) was stopped by immediate vacuum fil-
tration through glass fibre filters (GF/B Unifilters)
using a Packard cell harvester as above. The filters
were washed, dried, and counted in a Packard g
counter (TopCount). Non-specific binding was
defined with 10 mM melatonin. Ki values were calcu-
lated as above.
Melanophore contraction in X. laevis tadpoles. The X.
laevis tadpoles (stage 41) used in this study were
obtained from the Laboratoire de Biologie Cellulaire et
Reproduction CNRS (Rennes, France). They were
maintained in an aquarium in the laboratory at 22 ꢁC
under natural illumination for 8 days and fed daily with
powdered fish food. 18 h before the bioassay, tadpoles
of uniform stage, size, and color, were selected, removed
from the aquarium, and placed in groups of 5 in 100 mL
beakers on a dark background and filled with 45 mL of
pool water. They were illuminated with artificial light
(60 W) for 3 h before the experiments which were per-
formed at midday. The compound under test was
dissolved in a DMF and water mixture in a final volume
of 5 mL and added to the liquid in the beaker (45 mL) to
achieve the final selected concentration. After 15 min,
the experiment was terminated by the addition of a 37%
formaldehyde solution. The degree of the melanophore
response in each tadpole was determined by examination
of the melanophore configuration under a microscope
(Leitz, magnification ꢃ4) and evaluated according to
the melanophore index scale (1–5) of Hogben and
Slome.25 The data are the results of the sum of the
determinations of the melanophore index on the body
and the dorsal surface of the tadpole. EC50 values for
the compounds were determined from the concentra-
tion-response curves obtained with 5 concentrations in
the range of 0.1–100ꢃthe Ki values for chicken brain
receptors. The mean of the control data (animals treated
with vehicle) represented 100%. The results (n=2) were
calculated using the PRISM program (Graphpad).
N - [1,2 - Dihydro - 1 - acenaphthylenyloxy)cyclopropylcar-
boxamide (18e). It was prepared according to the pre-
vious
method
described
for
10a
with
cyclopropanecarbonyl chloride. The compound was
isolated as a white powder after recrystallisation in an
AcOEt/hexane mixture (yield: 59%) mp: 126 ꢁC. 1H
NMR (200 MHz, CD3OD) d (ppm) 0.77–0.89 (m, 4H),
3.47 (d, J=17.7 Hz, 1H), 3.67 (dd, J=18.3 Hz,
J=6.5 Hz, 1H), 5.88 (d, J=6.3 Hz, 1H), 7.32 (d,
J=6.6 Hz, 1H), 7.44–7.67 (m, 4H), 7.79 (d, J=8.0 Hz,
1H). 13C NMR (200 MHz, CD3OD) d (ppm) 7.18,
12.19, 37.89, 87.38, 120.80, 123.19, 123.70, 126.62,
128.74, 129.14, 132.59, 139.37, 142.14, 142.61, 174.64.
Anal. (C16H15NO2) calcd C 75.87 H 5.97 N 5.53; F. C
75.67 H 6.14 N 5.45.
Melatonin receptor binding assay in chicken brain mem-
branes. Chickens (Redbrook, male or female, 4 months
(3–4 kg); Cellubio, France) were decapitated at 12 am.
The brains were quickly removed and stored at ꢀ80 ꢁC.
The brains were homogenized (Polytron) in 10 vol of
ice-cold Tris–HCl buffer (50 mM, pH 7.4) and washed
twice by centrifugation (44 000 g, 25 min, 4 ꢁC). The
resulting pellet was resuspended in 10 vol of the same
buffer to a final concentration of 5 or 6 mg of pro-
tein/mL as determined by the method of Lowry.
Membrane aliquots (30 mL) were incubated in a total
volume of 0.25 mL Tris–HCl buffer (50 mM, pH 7.4)
with 0.05 nM 2-[125I]iodomelatonin and seven con-
centrations of the compound under test. Each bind-
ing assay was performed in triplicate. The incubation
(25 ꢁC, 60 min) was stopped by the addition of 3 mL
of ice-cold buffer and immediate vacuum filtration
through glass fiber filters (GF/B Whatman strips)
presoaked in 0.1% poly(ethyleneimine) using a Bran-
del cell harvester. The filters were washed (3ꢃ4 mL)
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