Design and synthesis of substituted compounds containing the 1,4- benzodioxin subunit. New potential calcium antagonists

Article abstract of DOI:10.1016/S0223-5234(00)00164-1

New compounds possessing 1,4-benzodioxin or its saturated analogous heterocyclic system were synthesized and tested for calcium antagonist activity. Biological differences were seen between the different modifications applied. These compounds have been shown to be representative of a novel series of calcium channel antagonists. (C) 2000 Editions scientifiques et medicales Elsevier SAS.

Full text of DOI:10.1016/S0223-5234(00)00164-1

Eur. J. Med. Chem. 35 (2000) 663−676
© 2000 Éditions scientifiques et médicales Elsevier SAS. All rights reserved
S0223523400001641/FLA
Original article
Design and synthesis of substituted compounds containing the 1,4-benzodioxin
subunit. New potential calcium antagonists
Isabel Sáncheza, Maria Dolors Pujola*, Gerald Guillaumetb, Roy Massinghamc, André Monteild,
Georges Durengd, Eileen Winslowd
aLaboratori de Química Farmacèutica, Fac. Farmàcia, Universitat de Barcelona, Avda. Diagonal 643, 08028 Barcelona, Spain
bInstitut de chimie organique et analytique associé au CNRS, université d’Orléans, BP-6957, 45067 Orléans cedex 2, France
cUCB Pharma, Pharmaceutical Sector, Chemin du Foriest, B-1420 Braine-L’Alleud, Belgium
dLaboratoire RL-CERM, ZI La Varenne, rue Goudier, 63203 Riom cedex, France
Received 16 September 1999; revised 17 January 2000; accepted 20 January 2000
Abstract – New compounds possessing 1,4-benzodioxin or its saturated analogous heterocyclic system were synthesized and tested for
calcium antagonist activity. Biological differences were seen between the different modifications applied. These compounds have been shown
to be representative of a novel series of calcium channel antagonists. © 2000 Éditions scientifiques et médicales Elsevier SAS
calcium channel antagonists / 2,3-dihydro-1,4-benzodioxin / 1,4-benzodioxin / calmodulin antagonists
1. Introduction
mation is available concerning SARs in these calcium
antagonist agents. In general, the different results ob-
tained with calcium antagonist agents seem to provide
further support for the hypothesis that the pharmacologi-
cal activity of these structures could be due to multiple
different mechanisms of action which are differently
affected by structural changes.
In the last 10 years extensive research of calcium
antagonists has been performed. The development of
drugs that inhibit the entry of calcium ions into cells by
blocking calcium channels are one of the most important
achievements in the therapy of cardiovascular disorders
[1]. The efficacy of some compounds as calcium antago-
nists and calcium entry blockers has been shown with
several clinical studies, but adverse effects described led
us to develop new anticalcium molecules with potential
efficacy and reduced toxic effects.
These calcium antagonist agents are a group of struc-
turally varied compounds acting on several types of
calcium channels or as calmodulin inhibitors. As far as
L-type voltage-operated channels (VOC) are concerned,
preponderantly three distinct classes of selective, potent
blockers are recognized [2, 3], which are represented by
verapamil (I), nifedipine (II) and diltiazem (III) (figure
1). Some of them are also known to possess a calmodulin
antagonistic property. This suggests that there is some
structural similarity in the drug binding sites of the
calcium channel and calmodulin. Moreover, little infor-
In continuing our interest on the development of new
anticalcium agents we have prepared a series of com-
pounds containing the 1,4-benzodioxin nucleus and an
alkyl chain with a tertiary or secondary amine function.
The 1,4-benzodioxin system is considered homologous to
the 1,4-benzodioxepin, substructure of the known cal-
cium antagonist HP-406 [4], and bioisostere of the
1,4-benzoxazine [5], both nuclei of interest as antical-
cium agents. The selected amines are known substruc-
tures of intracellular calcium antagonists possessing high
activity. 1,4-Benzoxazines possessing this chain at the 2
position were of particular interest. Moreover, com-
pounds of this type have not been explored as new drugs
with potential anticalcium activity that interact with
calcium channels and/or calmodulin. This work has
allowed us to study, among other things, the influence of
* Correspondence and reprints: mdpujol@farmacia.far.ub.es

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I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
Now, as a continuation of that work, we have synthesized
12 new compounds (1–12) containing 2,3-dihydro-1,4-
benzodioxin or its unsaturated analogue as a sub-structure
(table I).
The synthetic routes used for preparing the compounds
of this series were based on the synthesis of the function-
alized heterocyclic systems followed by alkylation of the
appropriate amine with the corresponding alkylating
reagent. The synthesis of intermediates 35, 36, 37 and 38
was carried out by the Riom-CERM Laboratory, follow-
ing their own method, whereas compound 39 was pur-
chased as a commercial product (Aldrich) (figure 2).
Tertiary amines 1, 2, 3, 4, 5 and 6 were prepared as
shown in figure 3, starting from esters 13a, 13b and 14
[6–9]. Treatment of 13a with aqueous methylamine
followed by reduction of the intermediate carboxamide
15, gave the secondary amine 16. This intermediate was
then alkylated, according to standard procedures, with the
halo derivatives 37 or 38 to provide the amines 2 and 1
(43 and 53%), respectively. The reduction of esters 13a,
13b and 14 gave the key hydroxy derivatives 17–19,
which were transformed into the appropriate alkylating
intermediates. In this way, alcohols 17 and 19 were
treated with N-bromosuccinimide (NBS)/triphenyl-
phosphine (TPP) and provided the respective bromo
compounds 21 and 22 in good yield. However, several
attempts to obtain the corresponding halo derivative of
alcohol 18 failed, probably due to its lack of stability, so
we prepared the tosyl derivative 20 [10] as an alternative.
Finally, the appropriate secondary amines 35 and 39 were
alkylated with the alkyl halides described before, in the
Figure 1. Structures of the most representative calcium chan-
nel blockers.
distinct amines and the effect of the 2,3-double bond of
the heterocyclic system.
2. Chemistry
We have recently reported the synthesis and the anti-
calcium activity of some 1,4-benzoxazine derivatives [5].
Table I. Compounds containing the 1,4-benzodioxin sub-structure.
Compound
X
R1
R2
R3
R4
C2–C3
1
2
3
4
5
6
7
8
CH2
CH2
CH2
CH2
CH2
CH2
CH2
CH2
CO
CH3
CH3
CH3
CH3
CH3
D
H
H
CH3
H
CH3
CH3
C
B
A
A
A
D
A
A
H
H
H
H
OCH3
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
CH3
sat.
sat.
unsat.
sat.
sat.
sat.
unsat.
sat.
sat.
sat.
9
A
A
10
11
12
(CH2)2
CH2
CH2
H
H
H
(CH2)2–O-A
A
sat.
unsat.

I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
665
standard conditions obtaining the acid 30. The key
intermediate 3-methylsubstituted 31 was obtained by
treatment of the carboxylic acid 30 with LDA [13],
followed by the addition of iodomethane in tetrahydro-
furan. The esterification followed by reduction of 31 gave
the alcohol 33 in good yield. Finally, the compound 33
was converted in its respective tosyl derivative 34 to
provide the compound 12 by alkylation of the amine 35.
Compounds 3, 7 and 12, which have the unsaturated 2–3
positions, are of interest to determine whether there are
differences in the biological activities by comparison with
the saturated analogues. All compounds prepared were
obtained as racemic mixtures. The modest anticalcium
activity manifested by these compounds did not prompt
us to invest time in the corresponding enantioselective
synthesis.
3. Biological results and discussion
Figure 2. Structure of intermediates 35–39.
The therapeutic activity attributed to calcium antago-
nism was measured in terms of the inhibition of the
contraction induced by modification of the concentration
of free calcium. As we previously described, compounds
with extracellular activity act on the calcium channel
receptors [2, 14–17], whilst those with intracellular action
inhibit the effect of calmodulin [18–20]. The compounds
synthesized in this study were subjected to three pharma-
cological tests: caffeine (CAF), phenylephrine (PE) and
potassium (K+). Thus, responses to high concentrations of
K+ are mediated by influx of calcium into the cell, via
L-type slow calcium channels. Since part of the concen-
tration to PE is due to the mobilization of calcium from
intracellular storage sites, any compound capable of
inhibiting PE responses may have an intracellular locus
of actions. Table II shows the results obtained in the three
tests indicated for compounds 1–12.
In general, the percentages of inhibition of contractions
induced by caffeine range from 0–100% for an approxi-
mate dose of 30 µM. The most notable compounds in this
test were 3 and 12 with a benzhydryl ether as a substitu-
ent, containing, moreover, a tertiary amine in its structure.
However, the influence of the substituent of amine is not
marked for the response to the caffeine test and the
saturated or unsaturated C2–C3 was more important. The
unsaturated compound 3 is interesting in the caffeine and
K+ tests.
presence of a base (K2CO3) to give the tertiary amines 3,
4, 5 and 6 in variable yields.
Figure 4 depicts the synthetic procedure used to obtain
the secondary amines 7 and 8, which were prepared by
alkylation of the corresponding amine intermediates 25
and 26, respectively, with the halo derivative 36. Both
primary amines were prepared by reduction of cyano
derivative 23 [11], but in the case of amine 25, the
previous dehydrogenation of the positions 2–3 was re-
quired [12]. The lower yield found for preparing 7 was
attributed to the double bond of the heterocyclic system.
The 2-substituted-1,4-benzodioxin derivatives 9, 10
and 11 were synthesized starting from the ester 13a
according to the synthetic route described in figure 5.
Carboxamide 9 was directly obtained by treatment of the
ester 13a with the amine 35. Compound 10 was prepared
by alkylation of the amine intermediate 28 with the
bromoalkane 36. The transformation of the ester 13a into
the corresponding bromo derivative by reduction and
treatment with NBS, followed by treatment with NaCN
and reduction of the cyano derivative 27, gave the
primary amine 28 in four steps. The synthetic pathway for
preparing the aminoether 11 was based on the alkylation
of the aminoalcohol 30 following the Williamson reac-
tion. This key intermediate 29 was obtained in moderate
yield by treatment of the amine 16 with bromoethanol in
basic media.
Compound 6 showed greater activity according to the
results of the PE test. This compound has a piperazinyl
group as an N-substituent, whereas all other compounds
have a substituted benzhydryl group.
The 2,3-disubstituted-1,4-benzodioxin derivative 12
was prepared from the ester 13b as outlined in figure 6.
The initial step was the basic hydrolysis of the ester under
The influence of the amine substituent (A, B, C and D)
is marked for the response to the K+ test (comparing

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I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
Figure 3. i. CH3NH2/toluene; ii. LiAlH4/THF; iii. B-Cl/TEA/KI/DMF; iv. C–Cl/K2CO3/KI/DMF; v. LiAlH4/THF; vi. TsCl/TEA/
CH2Cl2; vii. NBS/TPP/DMF; viii. A–NH–(CH3)/K2CO3/KI/DMF; ix. D–NH or A–NH–(CH3)/K2CO3/KI/DMF (A, B, C, and D have
been indicated in figure 2).
compounds 1, 2 and 4 with 6). It is important to point out
that there is an inversion of activities in the tests of K+
and PE depending on the amine considered. While
benzhydryl derivatives showed a slightly higher response
for the contractions caused by a high concentration of K+
(compounds 1, 2 and 4), comparing with those induced
by PE, compound 6 with a 1-(4-fluorophenyl)piperazine
group showed the opposite. As expected from the results,
the amine radical indicated by A is the best substituent in
these series of compounds for the inhibition of contrac-
tions induced by a high concentration of K+ (compound
4, IC50(K+) = 0.28 ± 0.09 µM).

I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
667
Figure 4. i. 1.NBS/CCl4, 2. NaI/acetone; ii. LiAlH4/THF; iii. A-Br/K2CO3/KI/DMF; iv. H2/Pd–C (A has been indicated in figure 2).
By comparison of the secondary (compounds 7, 8 and
10) and the tertiary amines (compounds 1, 2, 3, 4, 5 and
6), the tertiary amine is preferable to the secondary amine
in any of the tests studied.
The influence of the methoxy group at C7 decreased
the activity according to the results of the PE test and the
K+ test (comparing compound 5 with compound 4).
The distance between the heterocyclic nucleus and the
amine of the side chain (compound 10 compared with
compound 8) reveals an activity of the same order.
Passing from X = CH2 to X = CO has great influence in
the anticalcium activity. In general the activity decreased
(comparing compound 9 with compound 4).
spectra were recorded either on a Varian Gemini-200 MHz
and/or Varian XL-300 MHz spectrometer. Chemical shifts
are reported as δ values in parts per million downfield
from tetramethylsilane as the internal standard in CDCl3.
The following abbreviations are used to denote signal
patterns: s: singlet; d: doublet; dd: double doublet; t:
triplet; q: quadruplet; ba: broad absorption; m: multiplet;
cs: complex signal. IR spectra were recorded in an FTIR
Perkin Elmer 1600 spectrometer. Reported analytical data
are within ± 0.4% of the theoretical values. Merck 60
(40–60 microns) and Merck 60 F254 silica gel were used
for column chromatography and thin layer chromatogra-
phy, respectively. The organic extracts were dried over
Na2SO4. All air and moisture sensitive reactions were
carried out in an atmosphere of inert gas (argon). Yields
were not optimized. All reagents were of commercial
quality or were purified before use. Organic solvents were
purified by standard procedures.
4. Conclusions
Although an exact relationship between structure and
calcium antagonistic activity was not seen, in general, the
kind of substituent on the amino group and the saturation
degree of C2–C3 have significant influence. While the
1-(4-fluorophenyl)piperazine shows a higher response for
the contractions caused by PE, in this series (compound 6
IC50 = 5.9 µM), the benzhydryl group entity confers the
ability to inhibit contractions induced by K+ (compound
2 IC50 = 1.5 µM).
5.1.1. 2-(N-Methylcarbamoyl)-
2,3-dihydro-1,4-benzodioxin 15
Compound 13a (10 g; 48.07 mmol) and 40% methyl-
amine (23 mL; 721 mmol) were dissolved in toluene
(20 mL). The resulting solution was stirred at 35 °C for
40 h. After removing the solvent and methylamine, the
suspension obtained was extracted with CH2Cl2, dried,
filtered and concentrated under vacuum. The residue was
purified by recrystallization from hexane/AcOEt to give
7 g (75%) of 15 as white crystals. M.p.: 112–113 °C.
1H-NMR (CDCl3, 200 MHz) δ (ppm): 2.87 and 2.90 (s,
3H, CH3N); 4.17 (dd, J1 = 11 Hz, J2 = 7 Hz, 1H, C3H);
4.53 (dd, J1 = 11 Hz, J2 = 3 Hz, 1H, C3H); 4.68 (dd, J1 =
7 Hz, J2 = 3 Hz, 1H, C2H); 6.65 (ba, 1H, NH); 6.91 (m,
5. Experimental
5.1. Chemistry
All melting points were determined in capillary tubes
on a Gallenkamp apparatus and are uncorrected. NMR

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I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
Figure 5. i. A–NH–(CH3)/K2CO3/DCC/toluene; ii. LiAlH4/THF, 2. NBS/TPP/DMF, 3. NaCN/DMSO; iii. H2/Pd–C; iv. A-Br/K2CO3/
KI/DMF; v. 1. CH3NH2/toluene, 2. LiAlH4/THF; vi. BrCH2CH2OH/K2CO3/DMF; vii. A–Br/NaH/THF (A has been indicated in figure
2).
4H, C5H, C6H, C7H, C8H). 13C-NMR (CDCl3, 50.4 MHz)
δ (ppm): 25.7 (CH3, CH3N); 65.3 (CH2, C3); 73.3 (CH,
C2); 117.1 and 117.8 (CH, C5, C8); 122.0 and 122.5 (CH,
C6, C7); 141.7 and 143.4 (C, C4a, C8a); 167.9 (C, CONH).
5.1.2. 2-(N-Methylaminomethyl)-
2,3-dihydro-1,4-benzodioxin 16
A solution of 15 (4 g; 20.7 mmol) in anhydrous THF
(20 mL) was slowly added to a suspension of LiAlH4

I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
669
Figure 6. i. 1. NaOH 8%, 2. HCl 37%; ii. 1. LDA/THF, 2. CH3I/THF; iii. NaHCO3/CH3I/DMA; iv. LiAlH4/THF; v. TsCl/TEA/
CH2Cl2; vi. A–NH–(CH3)/K2CO3/KI/DMF (A has been indicated in figure 2).
Table II. Pharmacological activity of compounds 1–12.
Compound
IC50a(CAF) (µM)
IC50 (PE) (µM)
IC50 (K+) (µM)
(rabbit renal artery)
(rabbit aortic tissue)
(rabbit aortic tissue)
1
2
3
4
5
6
7
8
9
10
11
12
>30 (0 %)
>30 (1 %)
>30 (59 %)
>30 (0 %)
>30 (10 %)
>30 (0 %)
>30 (7 %)
>30 (14 %)
>30 (0 %)
>30 (0 %)
>30 (7 %)
>30 (60 %)
36.8 ± 3.5
<100 (10 %)
6.7 ± 0.6
>10 (39 %)
>10 (38 %)
–
>10 (37 %)
–
5.9
–
>10 (3 %)
–
–
–
2.2 ± 03
1.5 ± 0.1
7.9
0.28 ± 0.09
11.2
152
>100 (7 %)
>100 (2 %)
>100 (59 %)
>100 (38 %)
>100 (33 %)
–
1.52 ± 0.36
0.64
1.6
1.5
0.0086
–
Bepridil
Diltiazem
Fendiline
Flunarizine
Nifedipine
Prazosin
Rianodine
Verapamil
18.5 ± 15
64
4.2
>30 (5 %)
>10 (4 %)
>30 (23 %)
0.039 ± 0.001
>100 (32 %)
18
>10 (0 %)
0.0023
>10 (0 %)
4.4 ± 0.61
>10 (0 %)
>10 (0 %)
0.24 ± 0.06
a IC50 is defined as the concentration (10–6 M) of the tested compounds that inhibited 50% of contraction induced by caffeine, phenylephrine
or K+. Each result was obtained from five to nine preparations.

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I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
(2.4 g; 62.1 mmol) in anhydrous THF (50 mL) and stirred
at 50 °C for 24 h. Then the reaction mixture was poured
into ice-water. The precipitates were separated from the
solution by filtration. After removing the THF, the residue
was extracted with ether (3 × 40 mL). The organic layers
were washed with water, dried, filtered and concentrated
in vacuo, obtaining compound 16 as an unstable oil
(2.8 g) that was used in the subsequent reaction without
organic layers were washed several times with water,
dried, filtered and concentrated under reduced pressure.
The residue obtained was subjected to silica gel column
chromatography to give 1.55 g (60%) of a white solid
(compound 27) when hexane/AcOEt (70:30) was used as
1
eluent. M.p.: 67–69 °C. H-NMR (CDCl3, 200 MHz) δ
(ppm): 2.77 (s, 2H, CH2CN); 4.07 (dd, J1 = 11 Hz, J2 =
6 Hz, 1H, C3H); 4.31 (dd, J1 = 11 Hz, J2 = 2 Hz, 1H,
C3H); 4.49 (m, 1H, C2H); 6.89 (m, C5H, C6H, C7H,
C8H).
1
purification. H-NMR (CDCl3, 200 MHz) δ (ppm): 2.52
(s, 3H, CH3N); 2.87 (m, 2H, CH2N); 4.02 (dd, J1 =
12 Hz, J2 = 8 Hz, 1H, C3H); 4.28 (ba, 2H, C2H, C3H);
6.86 (m, 4H, C5H, C6H, C7H, C8H). 13C-NMR (CDCl3,
50.4 MHz) δ (ppm): 36.5 (CH3, CH3N); 51.9 (CH2,
CH2N); 66.5 (CH2, C3); 72.2 (CH, C2); 117.2 and 117.4
(CH, C5, C8); 121.5 and 121.6 (CH, C6, C7); 140.9 and
141.1 (C, C4a, C8a).
5.1.6. 2-Aminoethyl-2,3-dihydro-1,4-benzodioxin 28
A small amount of palladium-carbon (10%) and 37%
HCl (1 mL) were added to a solution of compound 27
(860 mg; 4.9 mmol) in 30 mL of methanol and the
suspension obtained was carried out to hydrogenation
(atmospheric pressure and room temperature). When the
consumption of H2 was finished, the catalyst was filtered
and the solvent removed. The solid obtained was dis-
solved in water and extracted with ether (3 × 20 mL). The
aqueous layer was basified with a solution of 1 N NaOH
and extracted again with CH2Cl2 (3 × 20 mL). The
organic layers were dried, filtered and concentrated in
vacuo obtaining 28 as a colourless oil with enough purity
to be directly employed in the next reaction (800 mg;
92%).1H-NMR (CDCl3, CD3OD, 200 MHz) δ (ppm):
1.71 (m, 2H, CH2CH2N); 2.70 (ba, 2H, NH2); 2.93 (m,
2H, CH2N); 3.88 (dd, J1 = 11 Hz, J2 = 8 Hz, 1H, C3H);
4.17 (m, 2H, C2H, C3H); 6.83 (m, 4H, C5H, C6H, C7H,
C8H). 13C-NMR (CDCl3, CD3OD, 50.4 MHz) δ (ppm):
33.7 (CH2, CH2CH2N); 37.6 (CH2, CH2N); 67.6 (CH2,
C3); 71.0 (CH, C2); 116.9 and 117.1 (CH, C5, C8); 121.2
and 121.4 (CH, C6, C7); 143.1 (C, C4a, C8a).
5.1.3. 2-Aminomethyl-1,4-benzodioxin 25
The cyano derivative 24 (2 g; 12.6 mmol) was slowly
added, under an argon atmosphere, to a suspension of
LiAlH4 (2.4 g; 63 mmol) in anhydrous THF (50 mL) and
the mixture was stirred at room temperature for 12 h.
After hydrolysis of LiAlH4 with small amounts of water,
the suspension was filtered and the THF removed. The
residue was diluted with water and extracted with ether
(3 × 30 mL). The organic layers were dried, filtered and
concentrated to give a dark, unstable oil whose instability
did not allow us to obtain the pure amine.
5.1.4. 2-Aminomethyl-2,3-dihydro-1,4-benzodioxin 26
Platinum oxide (IV) (124 mg; 0.54 mmol) and HCl
(37%) (2 mL) were added to a solution of 23 (4 g;
24.8 mmol) in dry methanol (100 mL). The resulting
suspension was carried out to hydrogenation (atmo-
spheric pressure and room temperature; 36 h). After
separation of the catalyst by filtration, the solvent was
removed. The resulting solid was dissolved in water and
Na2CO3 was added until basic pH. The aqueous layer was
extracted with CH2Cl2, dried, filtered and concentrated
under reduced pressure. The residue obtained was puri-
fied by silica gel column chromatography eluting with
CH2Cl2/MeOH (90:10) to give 3.4 g (83%) of 26 as a
colourless oil. 1H-NMR (CDCl3, 200 MHz) δ (ppm):
2.33 (ba, 2H, NH2); 2.96 (d, J = 6 Hz, 2H, CH2N); 4.11
(m, 3H, C2H, C3H); 6.85 (m, 4H, C5H, C6H, C7H, C8H).
5.1.7. 2-(N-Hydroxyethyl-N-
methylaminomethyl)-2,3-dihydro-1,4-benzodioxin 29
2-Bromoethanol (140 mg; 1.12 mmol) and anhydrous
K2CO3 (232 mg; 1.68 mmol) were added to a solution of
the methylamine 16 (100 mg; 0.56 mmol) in distilled
DMF (15 mL) and the mixture was stirred at 80 °C for
48 h. After removing the solvent, the residue obtained
was diluted with water and extracted with ether (3 ×
15 mL). The organic layers were dried, filtered and
concentrated under vacuum and the crude product was
subjected to silica gel column chromatography eluting
with hexane/AcOEt (50:50). The corresponding amino-
alcohol 29 (45 mg; 50%) was obtained as a yellow oil.
1H-NMR (CDCl3, 300 MHz) δ (ppm): 2.38 (s, 3H,
CH3N); 2.70 (m, 4H, CH2N); 3.00 (ba, 1H, OH); 3.60 (t,
J = 5 Hz, 2H, CH2OH); 3.98 (dd, J1 = 11 Hz, J2 = 7 Hz,
1H, C3H); 4.25 (m, 2H, C2H, C3H); 6.86 (s, 4H, C5H,
C6H, C7H, C8H).
5.1.5. 2-Cyanomethyl-2,3-dihydro-1,4-benzodioxin 27
Bromo derivative 21 (3.4 g; 14.8 mmol), obtained
from the ester 13a by reduction with LiAlH4 in tetrahy-
drofuran, was dissolved in 10 mL of DMSO and NaCN
(725 mg; 14.8 mmol) was slowly added to that solution.
After stirring at 65 °C for 3 days, the mixture was poured
into ice-water (50 mL) and extracted with ether. The

I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
671
5.1.8. (3-Methyl-1,4-benzodioxin-2-yl)carboxylic acid 31
5.1.10.2. 2-Hydroxymethyl-1,4-benzodioxin 18
A solution of 30 (2 g; 11.23 mmol) in 10 mL of
anhydrous THF was cooled at –78 °C. Then, a solution of
LDA (2 M in hexane) (22.5 mL; 44.92 mmol of base)
was slowly added, under an argon atmosphere, and the
mixture was stirred at –78 °C for 2.5 h. After, iodomethane
was added (7 mL; 112.3 mmol) and the mixture was
stirred until room temperature was reached. Then, a
saturated solution of NH4Cl was added (20 mL) and the
suspension obtained was extracted with ether (3 × 30 mL).
The organic layers were dried and filtered. After remov-
ing the solvent, a yellow oil (1.5 g; 70%) was obtained
but the compound of methylation was not purified be-
cause of its high polarity.
Compound 18 was obtained from the ester 13b follow-
ing the general method for reduction of esters (91%
yield).
5.1.10.3. 2-Hydroxymethyl-7-
methoxy-2,3-dihydro-1,4-benzodioxin 19
Compound 19 was synthesized from the ester 14 via
the general procedure for reduction of esters (85% yield).
1H-NMR (CDCl3, 200 MHz) δ (ppm): 2.40 (s, 1H, OH);
3.72 (s, 3H, CH3O); 3.85 (d, J = 4 Hz, 2H, CH2OH); 4.05
(dd, J1 = 12 Hz, J2 = 6 Hz, 1H, C3H); 4.21 (m, 2H, C2H,
C3H); 6.45 (m, 2H, C6H, C8H); 6.80 (d, J = 8 Hz, 1H,
C5H).
5.1.9. 3-Methyl-2-methoxycarbonyl-1,4-benzodioxin 32
The oil containing the carboxylic acid obtained in the
previous reaction was dissolved in 15 mL of dimethyl-
acetamide and NaHCO3 (3.35 g; 39.3 mmol) and methyl
iodide (2.2 mL; 33.7 mmol) were added. The mixture was
stirred at room temperature for 36 h, then it was washed
with brine and extracted with CH2Cl2 (3 × 25 mL). The
organic layers were dried, filtered and concentrated under
reduced pressure. The crude product was distilled
(20 mm Hg; 60 °C) in order to remove the excess of
DMA, giving yellow crystals of a high degree of purity
(2.3 g; 100%). M.p.: 74–76 °C. IR (ν, cm–1) 1 700, 1 650,
1 250. 1H-NMR (CDCl3, 300 MHz) δ (ppm): 2.31 (s, 3H,
CH3C); 3.85 (s, 3H, CH3O); 6.61 (m, 2H, C5H, C8H);
6.77 (m, 2H, C6H, C7H). 13C-NMR (CDCl3, 50.4 MHz)
δ (ppm): 16.2 (CH3, CH3C); 51.8 (CH3, CH3O); 115.8
and 116.1 (CH, C5, C8); 123.9 and 125.0 (CH, C6H,
C7H); 137.4 (C, C2); 141.2 and 142.1 (C, C4a, C8a); 148.2
(C, C3); 168.9 (C, COOMe).
5.1.10.4. 2-Hydroxymethyl-3-methyl-1,4-benzodioxin 33
Alcohol 33 was synthesized from the ester 32 in 84%
yield following the general procedure for reduction of
esters to alcohols. M.p.: 90–92 °C. IR (ν, cm–1):
1
3 500–2 900, 1 710, 1 270. H-NMR (CDCl3, 300 MHz)
δ (ppm): 1.72 (t, 1H, OH); 1.84 (s, 3H, CH3C); 4.15 (d,
J = 6 Hz, 2H, CH2OH); 6.70 (m, 4H, C5H, C6H, C7H,
C8H). 13C-NMR (CDCl3, 50.4 MHz) δ (ppm): 13.7 (CH3,
CH3C); 56.6 (CH2, CH2OH); 115.9 (CH, C5, C8); 123.5
(CH, C6, C7); 131.1 and 132.6 (C, C2, C3); 142.0 (C, C4a,
C8a).
5.1.11. Bromo derivatives from alcohols.
General procedure
N-bromosuccinimide (2 mmol) and triphenylphosphine
(2 mmol) were added to a solution of the appropriate
alcohol (1 mmol) in 10 mL of dry DMF. The mixture was
stirred at 40 °C for 6 h. After removing the excess of
DMF, the suspension obtained was poured into water and
extracted with ether. The organic layers were dried,
filtered and concentrated under reduced pressure. Purifi-
cation of the crude mixture by silica gel column chroma-
tography, eluting with hexane/AcOEt (70:30) gave the
corresponding bromide compound as a colourless oil.
5.1.10. Reduction of esters to alcohols.
General procedure
A solution of the suitable ester (1 mmol) in anhydrous
THF (10 mL) was slowly added, under an argon atmo-
sphere, to a suspension of LiAlH4 (2 mmol) in anhydrous
THF (50 mL). The mixture was stirred at room tempera-
ture for 30 min. After hydrolysis with small amounts of
water, the suspension obtained was filtered and the THF
removed. The residue was diluted with water and ex-
tracted with ether. The organic layers were dried, filtered
and concentrated to give white crystals pure enough to be
employed in the next reaction.
5.1.11.1. 2-Bromomethyl-2,3-dihydro-1,4-benzodioxin 21
Compound 21 was obtained from the alcohol 17 via the
general procedure for bromination of alcohols (96%
1
yield). H-NMR (CDCl3, 300 MHz) δ (ppm): 3.53 (dd,
J1 = 6 Hz, J2 = 5 Hz, 2H, CH2Br); 4.17 (dd, J1 = 11 Hz,
J2 = 6 Hz, 1H, C3H); 4.32 (m, 2H, C2H, C3H); 6.87 (m,
4H, C5H, C6H, C7H, C8H). 13C-NMR (CDCl3, 75.5 MHz)
δ (ppm): 28.8 (CH2, CH2Br); 65.5 (CH2, C3); 71.9 (CH,
C2); 117.1 and 117.2 (CH, C5, C8); 121.6 and 121.8 (CH,
C6, C7); 142.3 and 142.8 (C, C4a, C8a).
5.1.10.1. 2-Hydroxymethyl-2,3-
dihydro-1,4-benzodioxin 17
Compound 17 (98% yield) was obtained from ester
13a following the general procedure for reduction of
esters to alcohols.

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I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
5.1.11.2. 2-Bromomethyl-7-
methoxy-2,3-dihydro-1,4-benzodioxin 22
5.1.13.1. 2-(N-Diphenylmethoxyethyl-N-
methylaminomethyl)-2,3-dihydro-1,4-benzodioxin 1
Compound 1 (53% yield) was prepared via the general
procedure B, starting from diphenylmethoxyethyl chlo-
ride 38 (1.65 g; 6.7 mmol); the amine 16 (600 mg;
3.35 mmol) and K2CO3 (1.4 g; 10.05 mmol). Anal. calcd.
for C25H27NO3: C, 77.09% H, 6.99% N, 3.59%. Found:
C, 77.14% H, 6.97% N, 3.59%. IR (ν, cm–1): 3 100,
The bromo derivative 22 was prepared from the
alcohol 19 following the general procedure of bromina-
1
tion (70% yield). H-NMR (CDCl3, 200 MHz) δ (ppm):
3.52 (d, J = 5 Hz, 2H, CH2Br); 3.73 (s, 3H, CH3O); 4.13
(m, 1H, C3H); 4.27 (m, 2H, C2H, C3H); 6.45 (m, 2H,
C6H, C8H); 6.79 (d, J = 9 Hz, 1H, C5H).
1
2 868, 1 465, 1 269, 1 098. H-NMR (CDCl3, 200 MHz)
δ (ppm): 2.35 (s, 3H, CH3N); 2.69 (m, 4H, CH2NCH2);
3.55 (t, J = 5 Hz, 2H, CH2O); 3.86 (dd, J1 = 11 Hz, J2 =
7 Hz, 1H, C3H); 4.18 (m, 2H, C2H, C3H); 5.30 (s, 1H,
CHAr); 6.84 (m, 4H, C5H, C6H, C7H, C8H); 7.31 (m,
10H, C2′H, C3′H, C4′H, C5′H, C6′H). 13C-NMR (CDCl3,
50.4 MHz) δ (ppm): 43.6 (CH3, CH3N); 57.4 and 57.6
(CH2, CH2N); 66.4 and 67.1 (CH2, CH2O, C3); 71.3 (CH,
C2); 83.8 (CH, CHAr); 116.9 and 117.3 (CH, C5, C8);
121.1 and 121.3 (CH, C6, C7); 126.8 (CH, C2′, C6′); 127.3
(CH, C4′); 128.3 (CH, C3′, C5′); 142.2 (C, C4a, C8a); 143.2
(C, C1′).
5.1.12. Tosyl derivatives from alcohols.
General procedure
A solution of the corresponding alcohol (1 mmol) in
5 mL of dry CH2Cl2 was cooled at 0 °C and tosyl chloride
(1.3 mmol) and triethylamine (1.3 mmol) were added.
The mixture was stirred at 0 °C for 10 min. Next, the
reaction continued, keeping the mixture at –20 °C for
12 h without stirring. When the room temperature was
recovered, the mixture was acidified with a solution of
1 N HCl and extracted with CH2Cl2. The organic layer
was dried, filtered and the solvent removed to give a
crude product which was used in the next reaction
without purification because of its instability.
5.1.13.2. 2-{N-[4,4-Bis(p-fluorophenyl)butyl]-
N-methylaminomethyl}-2,3-dihydro-1,4-benzodioxin 2
Compound 2 (43% yield) was obtained following the
general method A, from the amine 16 (2 g; 11.17 mmol);
5.1.12.1. Tosyl derivative
of 2-hydroxymethyl-1,4-benzodioxin 20
Compound 20 was prepared from the alcohol 18
following the Sharpless procedure.
4,4-bis(p-fluorophenyl)butylchloride
37
(3.4 g;
12.3 mmol) and TEA (4.6 mL; 33.51 mmol). Anal. calcd.
for C26H27F2NO2: C, 73.74% H, 6.42% N, 3.31%.
Found: C, 73.75% H, 6.44% N 3.29%. IR (ν, cm–1):
2 943, 2 803, 1 506, 1 265, 1 158. 1H-NMR (CDCl3,
200 MHz) δ (ppm): 1.42 (m, 2H, CH2CH2CH2); 2.00 (q,
J = 8 Hz, 2H, CH2CH2CH); 2.31 (s, 3H, CH3N); 2.50 (t,
J = 7 Hz, 2H, CH2N); 2.62 (t, J = 6 Hz, 2H, CH2N); 3.85
(t, J = 8 Hz, 1H, CHAr); 3.94 (dd, J1 = 12 Hz, J2 = 7 Hz,
1H, C3H); 4.25 (m, 2H, C2H, C3′′H); 6.84 (s, 4H, C5H,
C6H, C7H, C8H); 6.95 (m, 4H, C3H, C5′′H); 7.15 (m, 4H,
C2′′H, C6′′H). 13C-NMR (CDCl3, 50.4 MHz) δ (ppm):
25.2 (CH2, CH2CH2CH2); 33.3 (CH2, CH2CH); 42.8
(CH3, CH3N); 49.5 (CH, CHAr); 57.4 and 57.9 (CH2,
CH2NCH2); 66.6 (CH2, C3); 71.1 (CH, C2); 115.2 and
115.6 (CH, J = 21 Hz, C3′′,C5′′); 117.2 and 117.5 (CH, C5,
C8); 121.5 and 121.7 (CH, C6, C7); 129.1 and 129.2 (CH,
J = 8 Hz, C2′′, C6′′); 140.5 (C, C4a, C8a); 143.4 (C, C1′′);
159.1 and 164.0 (C, J = 246 Hz, C4′′).
5.1.12.2. Tosyl derivative
of 2-hydroxymethyl-3-methyl-1,4-benzodioxin 34
Tosylate 34 is obtained from hydroxyderivative 33 via
the general procedure.
5.1.13. Alkylation of the corresponding amines
with the suitable alkylating agent. General procedures
Method A:
Distilled TEA and a catalytic amount of KI were added
to a solution of the corresponding amine and the alkylat-
ing agent in 15 mL of dry DMF. The mixture was stirred,
under argon atmosphere, at 80 °C for 48 h. After remov-
ing the solvent, the residue was diluted with water and
extracted with ether. The organic layers were dried,
filtered and concentrated under reduced pressure. Purifi-
cation of the crude product by silica gel column chroma-
tography, eluting with hexane/AcOEt (70:30) allowed us
to obtain the corresponding alkylated compound.
Method B:
5.1.13.3. 2-[N-Bis(p-fluorophenyl)
methoxyethyl-N-methylaminomethyl]-1,4-benzodioxin 3
The preparation of compound 3 (10% global yield) was
carried out by the same procedure as described in method
B, taking as starting compounds the impurified tosyl
derivative 20 (7.6 mmol theoretically); N-methyl-bis(p-
fluorophenyl)methoxyethylamine 35 (4.12 g; 14.9 mmol)
The general procedure B was carried out in the same
way as described in method A, but using anhydrous
K2CO3 as a base and keeping the mixture reaction stirring
at room temperature under argon atmosphere for 5 days.

I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
673
and K2CO3 (2.05 g; 14.88 mmol). M.p.: 100–102 °C.
Anal. calcd. for C25H23F2NO3: C, 70.91% H, 5.47% N,
3.31%. Found: C, 70.87% H, 5.42% N, 3.28%. IR (ν,
cm–1): 3 000, 2 863, 1 510, 1 264, 1 153. 1H-NMR
(CDCl3, 200 MHz) δ (ppm): 2.40 (s, 3H, CH3N); 2.78
(m, 4H, CH2NCH2); 3.58 (m, 2H, CH2O); 5.32 (s, 1H,
CHAr); 5.49 (s, 1H, C3H); 6.88 (m, 8H, C5H, C6H, C7H,
C8H, C3′H, C5′H); 7.30 (m, 4H, C2′H, C6′H). 13C-NMR
(CDCl3, 50.4 MHz) δ (ppm): 43.3 (CH3, CH3N); 55.9
(CH2, CH2N); 57.7 (CH2, CH2N); 66.0 (CH2, CH2O);
82.7 (CH, CHAr); 92.7 (CH, C3); 115.0 and 115.4 (CH,
J = 22 Hz, C3′, C5′); 116.7 and 117.7 (CH, C5, C8); 121.6
and 121.8 (CH, C6, C7); 128.2 and 128.4 (CH, J = 8 Hz,
C2′, C6′); 137.9 (C, C2, C1′); 141.0 (C, C4a, C8a); 159.6
and 164.5 (C, J = 247 Hz, C4′).
1H, C5H); 6.99 (m, 4H, C3′H, C5′H); 7.26 (m, 4H, C2′H,
C6′H). 13C-NMR (CDCl3, 50.4 MHz) δ (ppm): 43.8
(CH3, CH3N); 55.6 (CH3, CH3O); 57.6 (CH2, CH2N);
57.8 (CH2, CH2N); 66.4 (CH2, CH2O); 67.2 (CH2, C3);
71.7 (CH, C2); 82.6 (CH, CHAr); 102.6 (CH, C8); 107.1
(CH, C6); 115.0 and 115.4 (CH, J = 21 Hz, C3′, C5′);
117.1 (CH, C5); 128.4 and 128.5 (CH, J = 8 Hz, C2′, C6′);
137.9 (C, C7); 138.0 (C, C1′); 143.0 (C4a, C8a); 159.9 and
164.7 (C, J = 242 Hz, C4′).
5.1.13.6. 2-[4-(p-Fluorophenyl)-1-
piperazinylmethyl]-2,3-dihydro-1,4-benzodioxin 6
Compound 6 (65% yield) was synthesized via the
general procedure B, taking as depart products 4-(p-
fluorophenyl)piperazine 39 (3 g; 16.25 mmol); the halo
derivative 21 (1.5 g; 6.5 mmol) and K2CO3 (2.7 g;
19.5 mmol). M.p. (HCl): 228–230 °C. Anal. calcd. for
C19H21FN2O2: C, 69.49% H, 6.44% N, 8.53%. Found: C,
69.53% H, 6.42% N 8.54%. IR (ν, cm–1): 2 948, 2 827,
1 500, 1 262. 1H-NMR (CDCl3, 200 MHz) δ (ppm): 2.65
(m, 6H, CH2NCH2, CH2N); 3.11 (m, 4H, CH2N–Ar);
4.01 (dd, J1 = 11 Hz, J2= 7 Hz, 1H, C3H); 4.33 (m, 2H,
C2H, C3H); 6.97 (m, 8H, C5H, C6H, C7H, C8H, C2′′H,
C3′′H, C5′′H, C6′′H). 13C-NMR (CDCl3, 50.4 MHz) δ
(ppm): 50.1 (CH2, CH2N); 53.8 (CH2, CH2N); 58.4 (CH2,
CH2N); 66.8 (CH2, C3); 71.1 (CH, C2); 115.2 and 115.7
(CH, J = 22 Hz, C3′′, C5′′); 117.1 and 117.4 (CH, C5, C8);
117.6 and 117.8 (CH, J = 7 Hz, C2′′, C6′′); 121.4 and
121.5 (CH, C6, C7); 142.5 and 142.8 (C, C4a, C8a); 148.9
(C, C1′′); 154.3 and 159.2 (C, J = 245 Hz, C4′′).
5.1.13.4. 2-[N-Bis(p-fluorophenyl)methoxyethyl-
N-methylaminomethyl]-2,3-dihydro-1,4-benzodioxin 4
The preparation of compound 4 (26% yield) was
carried out by the procedure of method A, starting from
the halo derivative 21 (2.2 g; 9.6 mmol); N-methyl-bis(p-
fluorophenyl)methoxyethylamine 35 (2.9 g; 10.56 mmol)
and TEA(3 g; 28.8 mmol). Anal. calcd. for C25H25F2NO3:
C, 70.57% H, 5.92% N, 3.29%. Found: C, 70.52% H,
5.90% N, 3.32%. IR (ν, cm–1): 2 900, 2 867, 1 494,
1 265, 1 093. 1H-NMR (CDCl3, 200 MHz) δ (ppm): 2.41
(s, 3H, CH3N); 2.78 (m, 4H, CH2NCH2); 3.54 (t, J =
6 Hz, 2H, CH2O); 3.98 (dd, J1 = 10 Hz, J2 = 6 Hz, 1H,
C3H); 4.25 (m, 2H, C2H, C3H); 5.31 (m, 1H, CHAr); 6.84
(m, 4H, C5H, C6H, C7H, C8H); 7.03 (m, 4H, C3′H, C5′H);
7.28 (m, 4H, C2′H, C6′H). 13C-NMR (CDCl3, 50.4 MHz)
δ (ppm): 43.2 (CH3, CH3N); 56.9 and 57.3 (CH2,
CH2NCH2); 66.0 and 66.7 (CH2, CH2O, C3); 70.9 (CH,
C2); 82.0 (CH, CHAr); 114.6 and 115.0 (CH, J = 21 Hz,
C3′, C5′); 116.6 and 116.9 (CH, C5, C8); 120.8 and 121.0
(CH, C6, C7); 127.9 and 128.1 (CH, J = 8 Hz, C2′, C6′);
137.4 (C, C1′); 144.0 (C, C4a, C8a); 159.8 and 164.2 (C,
J = 235 Hz, C4′).
5.1.13.7. 2-[N-Bis(p-fluorophenyl)-
methoxyethylaminomethyl]-1,4-benzodioxin 7
Compound 7 (4% overall yield) was synthesized via
the general method (B), starting from the impure amine
25 (12.6 mmol theoretically); the halo derivative 36
(8.2 g; 25.2 mmol) and K2CO3 (5.2 g; 37.8 mmol). Anal.
calcd. for C24H21F2NO3: C, 70.40% H, 5.17% N, 3.42%.
Found: C, 70.35% H, 5.16% N, 3.45%. 1H-NMR (CDCl3,
200 MHz) δ (ppm): 2.88 (t, J = 6 Hz, 2H, CH2NCH2CH2);
3.12 (s, 1H, NH); 3.50 (t, J = 6 Hz, 2H, CH2O); 5.29 (s,
3H, CHAr, C–CH2NCH2); 5.80 (s, 1H, C3H); 6.61 (m,
2H, C5H, C8H); 6.83 (m, 2H, C6H, C7H); 6.92 (m, 4H,
C3′H, C5′H); 7.24 (m, 4H, C2′H, C6′H). 13C-NMR (CDCl3,
50.4 MHz) δ (ppm): 53.6 (CH2, CH2N); 53.7 (CH2,
CH2N); 67.6 (CH2, CH2O); 82.5 (CH, CHAr); 115.1 and
115.5 (CH, J = 21 Hz, C3′, C5′); 116.0 and 116.3 (CH, C5,
C8); 123.9; 124.0 and 124.6 (C3, C6, C7); 128.5 and 128.7
(CH, J = 8 Hz, C2′, C6′); 135.1 (C, C2); 137.9 (C, C1′);
141.1 and 141.2 (C, C4a, C8a); 159.8 and 164,7 (C, J =
245 Hz, C4′).
5.1.13.5. 2-[N-Bis(p-fluorophenyl)methoxyethyl-N-methyl-
aminomethyl]-7-methoxy-2,3-dihydro-1,4-benzodioxin
5
Compound 5 (50% yield) was obtained via the general
method B, taking as starting compounds the amine 35
(1.07 g; 3.86 mmol); the halo derivative 22 (0.5 g;
1.93 mmol) and K2CO3 (801 mg; 5.8 mmol). Anal. calcd.
for C26H27F2NO4: C, 68.56% H, 5.97% N, 3.07%.
Found: C, 68.57% H, 5.94% N, 3.05%. IR (ν, cm–1):
2 950, 2 800, 1 450, 1 259, 1 158. 1H-NMR (CDCl3,
200 MHz) δ (ppm): 2.37 (s, 3H, CH3N); 2.76 (m, 4H,
CH2N); 3.52 (t, J = 4 Hz, 2H, CH2O); 3.71 (s, 3H,
CH3O); 3.90 (m, 2H, C3H); 4.20 (m, 1H, C2H); 5.31 (s,
1H, CHAr); 6.45 (m, 2H, C6H, C8H); 6.78 (d, J = 9 Hz,

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I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
5.1.13.8. 2-[N-Bis(p-fluorophenyl)methoxy-
C1′, rotamers A and B); 142.8 and 143.3 (C, C4a, C8a);
159.8 and 159.9 (C, J = 247 Hz, C4′); 166.6 and 167.4
(C, CONR, rotamers A and B).
ethylaminomethyl]-2,3-dihydro-1,4-benzodioxin 8
The preparation of compound 8 (52% yield) was
carried out following the general procedure (B), starting
from the amine 26 (450 mg; 2.72 mmol); the bromo
derivative 36 (1.8 g; 5.44 mmol) and K2CO3 (1.1 g;
8 mmol). Anal. calcd. for C24H23F2NO3: C, 70.06% H,
5.63% N, 3.40%. Found: C, 70.02% H, 5.67% N, 3.37%.
IR (ν, cm–1): 3 068, 3 019, 2 866, 1 502, 1 262, 1 100.
1H-NMR (CDCl3, 200 MHz) δ (ppm): 2.87 (m, 4H,
CH2NCH2); 3.46 (t, J = 6 Hz, 2H, CH2O); 3.91 (dd, J1 =
11 Hz, J2 = 7 Hz, 1H, C3H); 4.18 (m, 2H, C3H, C2H);
5.25 (s, 1H, CHAr); 6.83 (s, 4H, C5H, C6H, C7H, C8H);
7.00 (m, 4H, C3′H, C5′H); 7.23 (m, 4H, C2′H, C6′H).
13C-NMR (CD3OD, 50.4 MHz) δ (ppm): 56.7 and 56.9
(CH2, CH2NCH2); 67.6 and 68.9 (CH2O, C3); 73.5 (CH,
C2); 83.7 (CH, CHAr); 116.0 and 116.4 (CH, J = 22 Hz,
C3′, C5′); 118.2 and 118.5 (CH, C5, C8); 122.4 and 122.6
(CH, C6, C7); 129.3 and 130.1 (CH, J = 8 Hz, C2′, C6′);
139.9 (C, C1′); 144.1 and 144.8 (C, C4a, C8a); 161.4 and
166.3 (C, J = 246 Hz, C4′).
5.1.13.10. 2-[N-Bis(p-fluorophenyl)methoxy-
ethylaminoethyl]-2,3-dihydro-1,4-benzodioxin 10
Compound 10 (32% yield) was obtained following the
general method B, taking as starting compounds the
amine 28 (540 mg; 3.01 mmol); the bromo derivative 36
(2 g; 6.02 mmol) and K2CO3 (1.24 g; 9 mmol). Anal.
calcd. for C25H25F2NO3: C, 70.57% H, 5.92% N, 3.29%.
Found: C, 70.61% H, 5.96% N, 3.31%. IR (ν, cm–1):
2 928, 2 855, 1 507, 1 236, 1 158. 1H-NMR (CDCl3,
200 MHz) δ (ppm): 1.62 (m, 2H, CH2CH2N); 2.69 (m,
4H, CH2NCH2); 3.73 (dd, J1 = 11 Hz, J2 = 8 Hz, 1H,
C3H); 3.82 (t, J = 6 Hz, 2H, CH2O); 4.05 (m, 2H, C2H,
C3H); 5.16 (s, 1H, CHAr); 6.76 (m, 4H, C5H, C6H, C7H,
C8H); 6.86 (m, 4H, C3′H, C5′H); 7.11 (m, 4H, C2′H,
C6′H). 13C-NMR (CDCl3, 50.4 MHz) δ (ppm): 24.9
(CH2, CH2CH2N); 45.3 (CH2, CH2N); 54.3 (CH2, CH2N);
67.7 (CH2, CH2O, C3); 71.5 (CH, C2); 82.6 (CH, CHAr);
115.0 and 115.4 (C, J = 22 Hz, C3′, C5′); 117.0 and 117.2
(CH, C5, C8); 121.2 and 121.3 (CH, C6, C7); 128.4 and
128.5 (CH, J = 8 Hz, C2′, C6′); 138.2 (C, C1′); 142.2 (C,
C4a, C8a); 159.9 and 164.8 (C, J = 247 Hz, C4′).
5.1.13.9. 2-[N-Bis(p-fluorophenyl)methoxyethyl-
N-methylcarbamoyl]-2,3-dihydro-1,4-benzodioxin 9
Anhydrous K2CO3 (1.1 g; 8.1 mmol) and a catalytic
amount of DCC were added to a solution of the amine 35
(1.5 g; 5.4 mmol) and the ester 13a (1.7 g; 8.1 mmol) in
dry toluene (100 mL). The mixture was refluxed for 48 h,
then the solvent was removed. The suspension obtained
was poured into ice-water and extracted with CH2Cl2 (3 ×
25 mL). The organic layers were dried, filtered and
concentrated under reduced pressure to give a white solid
(hexane/AcOEt) identified as the compound 9 (1.36 g;
58% yield). M.p.: 102–103 °C. Anal. calcd. for
C25H23F2NO4: C, 68.33% H, 5.27% N, 3.19%. Found: C,
68.35% H, 5.24% N, 3.25%. 1H-NMR (CDCl3, 200 MHz)
δ (ppm): 2.96 and 3.29 (s, 3H, CH3N rotamers A and B);
3.61 (cs, 4H, CH2N, CH2O); 4.25 (dd, J1 = 11 Hz, J2 =
8 Hz, 1H, C3H); 4.37 (dd, J1 = 11 Hz, J2 = 2 Hz, 1H,
C3H); 4.85 (dd, J1 = 8 Hz, J2 = 2 Hz, 1H, C2H); 5.06
(dd, J1 = 8 Hz, J2 = 3 Hz, 1H, C2H); 5.32 (s, 1H,
CHAr); 6.89 (m, 4H, C5H, C6H, C7H, C8H); 6.96 (m, 4H,
C3′H, C5′H); 7.25 (m, 4H, C2′H, C6′H). 13C-NMR (CDCl3,
50.4 MHz) δ (ppm): 33.8 and 36.8 (CH3, CH3N rotamers
A and B); 48.3 and 49.3 (CH2, CH2N rotamers A and B);
64.9 and 65.1 (CH2, CH2O rotamers A and B); 65.9 and
66.6 (CH2, C3 rotamers A and B); 70.3 and 70.6 (CH, C2
rotamers A and B); 82.4 and 82.9 (CH, CHAr rotamers A
and B); 115.1 and 115.6 (CH, J = 22 Hz, C3′, C5′ rotamer
A); 115.3 and 115.6 (CH, J = 22 Hz, C3′, C5′, rotamer B);
117.3 (CH, C5, C8); 121.8 and 122.2 (CH, C6, C7); 128.4
and 128.5 (CH, J = 8 Hz, C2′, C6′); 137.2 and 137.6 (C,
5.1.13.11. 2-[N-7,7-Bis(p-fluorophenyl)-3,6-dioxaheptyl]-
N-methylaminomethyl]-2,3-dihydro-1,4-benzodioxin 11
A solution of aminoalcohol 29 (800 mg; 3.6 mmol) and
the halo derivative 36 (3 g; 8.9 mmol) in 20 mL of
anhydrous THF were slowly added to a suspension of
NaH (215 mg; 4.5 mmol; 50% wealth) and refluxed
under argon. When the room temperature was recovered,
the mixture was poured into water and extracted with
ether (3 × 20 mL). The organic layers were dried, filtered
and the solvent removed under reduced pressure. The
aminoether 11 (300 mg; 18%) was separated from the
residue obtained by silica gel column chromatography,
using hexane/AcOEt (70:30) as an eluent. IR (ν, cm–1):
3 100, 2 866, 1 508, 1 263, 1 232. Anal. calcd. for
C27H29F2NO4: C, 69.07% H, 6.22% N, 2.98%. Found: C,
69.09% H, 6.25% N, 2.94%. 1H-NMR (CDCl3, 200 MHz)
δ (ppm): 2.40 (s, CH3N); 2.72 (m, 4H, CH2NCH2); 3.59
(m, 6H, CH2O); 3.94 (dd, J1 = 12 Hz, J2 = 7 Hz, 1H,
C3H); 4.28 (m, 2H, C2H, C3H); 5.32 (s, 1H, CHAr); 6.85
(s, 4H, C5H, C6H, C7H, C8H); 6.99 (m, 4H, C3′′H, C5′′H);
7.27 (m, 4H, C2′′H, C6′′H). 13C-NMR (CDCl3, 50.4 MHz)
δ (ppm): 43.8 (CH3, CH3N); 57.4 and 57.8 (CH2,
CH2NCH2); 66.6; 66.8; 68.5 and 70.4 (CH2, CH2O); 71.4
(CH, C2); 82.5 (CH, CHAr); 115.1 and 115.5 (CH, J =
22 Hz, C3′′, C5′′); 117.1 and 117.4 (CH, C5, C8); 121.4
and 121.6 (CH, C6, C7); 128.6 and 128.8 (CH, J = 8 Hz,

I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
675
C2′′, C6′′); 137.8 (C, C1′′); 143.2 and 143.3 (C, C4a, C8a);
159.9 and 164.8 (C, J = 247 Hz, C4′′).
Krebs solution and maintained at 37 °C. Each ring is set
at an optimal pre-load of 10 g (determined from pre-
load–response curves and the pre-calibrated pen re-
corder). The aortic ring is maintained under 10 g applied
tension for 60–90 min with adjustments for any sponta-
neous relaxation. Following washing with fresh Krebs
solution the preparation is contracted two or three times
by addition of acetylcholine chloride (AchCl, 1 µmol/L).
If AchCl produces more than 50% relaxation of the
phenylephrine induced contraction the endothelium is
considered functionally intact, otherwise the preparation
is discarded and replaced.
5.1.13.12. 2-[N-Bis(p-fluorophenyl)methoxyethyl-
N-methylaminomethyl]-3-methyl-1,4-benzodioxin 12
Compound 12 (6% global yield) was prepared follow-
ing the general procedure (B), starting from the unpuri-
fied tosyl derivative 34 (3.8 mmol theoretically); the
amine 35 (3.15 mmol; 11.4 mmol) and K2CO3 (1.5 g;
11.4 mmol). Anal. calcd. for C26H25F2NO3: C, 71.38%
H, 5.76% N, 3.20%. Found: C, 71.35% H, 5.72% N,
3.24%. IR (ν, cm–1): 2 900, 2 868, 1 502, 1 498, 1 150.
1H-NMR (CDCl3, 200 MHz) δ (ppm): 1.49 (s, 3H,
CH3C); 2.38 (s, 3H, CH3N); 2.76 (m, 4H, CH2N2CH);
3.51 (t, J = 7 Hz, 2H, CH2O); 5.29 (s, 1H, CHAr); 6.80
and 7.01 (cs, 8H, C5H, C6H, C7H, C8H, C3′H, C5′H); 7.39
(m, 4H, C2′H, C6′H). 13C-NMR (CDCl3, 50.4 MHz) δ
(ppm): 24.1 (CH3, CH3C); 43.3 (CH3, CH3N); 56.8 and
57.8 (CH2, CH2NCH2); 66.1 (CH2, CH2O); 82.8 (CH,
CHAr); 115.1 and 115.5 (CH, J = 20 Hz, C3′, C5′); 116.7
and 117. 2 (CH, C5, C8); 121.5 and 121.8 (CH, C6, C7);
128.6 and 128.4 (CH, J = 8 Hz, C2′, C6′); 131.2 and 132.8
(C, C2, C3); 137.6 (C, J = 3 Hz, C1′); 140.0 and 140.9 (C,
C4a, C8a); 160.1 and 165.0 (C, J = 247 Hz, C4′).
5.2.1.3. Procedure
The acceptable aortic rings are contracted with either
phenylephrine (0.3 µmol/L) or KCl (40 µmol/L) until
stable tone is attained. The preparation is washed with
fresh Krebs solution, allowed to relax to baseline values
and again stimulated with the same agonist. The proce-
dure is repeated until stable control responses are ob-
tained. Following the last relaxation to baseline values
the first concentration of test compound (usually 0.1 µmol/
L) is added to the preparation and another five contrac-
tions are provoked by the previous agonist. Each reduc-
tion in contraction is expressed as a percentage of the
control response. With an equilibration time of approxi-
mately 90 min this procedure is repeated with 1 µmol/L
and 10 µmol/L dose levels of the test compound. Each
test compound is tested in at least three different tissue
preparations.
5.2. Pharmacological methods
5.2.1. Assay method [21]
for phenylephrine or K+ induced contractions
5.2.1.1. Test medium
An aortic ring preparation is placed in an organ bath
containing a Krebs solution of the following composition:
NaCl (118.1 mmol/L); KCl (4.7 mmol/L); MgSO4·7H2O
(0.6 mmol/L); KH2PO4 (1.8 mmol/L); D-glucose
(11.1 mmol/L); NaHCO3 (25.0 mmol/L); CaCl2·2H2O
(25.0 mmol/L); Ultrapure water (Milli-Q quality) to 1 L.
The pH is approximately 7.4 after 30 min gassing with
5% CO2 in O2.
5.2.1.4. Evaluation of responses
The percentage inhibition of contractions is plotted
against the logarithm of concentration of test compound.
The concentration of test compound which inhibits con-
tractions to phenylephrine or to high K+ by 50% is termed
the IC50.
5.2.2. Assay method [22]
for caffeine-induced contractions
5.2.1.2. Aortic ring preparation
The principle of the test is to assess intracellularly
mediated vasodilation using caffeine-induced contraction
in isolated renal arteries.
A male rabbit (New Zealand white rabbits, weighting
2.2–3.5 kg) is stunned by a blow to the neck or by a
pentobarbital sodium injection and exsanguinated. The
thorax is opened and the descending aorta dissected free,
put into a Petri dish and covered with gassed Krebs
solution. The vessel is carefully cleaned of fat and
connective tissue and then cut into rings of 3–5 mm
width. Each ring is placed onto stainless steel hooks, care
being taken to avoid damage to the intimal surface and
then mounted in an organ bath, with one hook being
attached to the bath and the other to the tension trans-
ducer. The organ bath is previously filled with gassed
5.2.2.1. Procedure
Left renal artery rings, approximately 2.5–3 mm long
were obtained from male New Zealand white rabbits and
mounted between stainless steel hooks in an organ bath
filled with Krebs solution at 37 °C gassed with 95%
oxygen/5% CO2. Each ring is set at an optimal pre-load
of 3 g. Tissues are contracted with caffeine (10 mM) and
test substances added 20 min before challenge with
agonist.

676
I. Sánchez et al. / Eur. J. Med. Chem. 35 (2000) 663–676
[7] Coudert G., Guillaumet G., Loubinoux B., Tetrahedron Lett. (1978)
5.2.3. IC50 determination
1059–1062.
IC50 values defined the concentration (µM) of the test
compounds that inhibited the 50% of contraction induced
by potassium, phenylephrine or caffeine and were ob-
tained using linear regression analysis.
[8] Khouili M., Guillaumet G., Coudert G., Pujol M.D., Il Farmaco 51
(1996) 175–184.
[9] Khouili M., Guillaumet G., Coudert G., Pujol M.D., Il Farmaco 51
(1996) 185–188.
[10] Klunder J.M., Ko S.Y., Sharpless K.B., J. Org. Chem. 51 (1986)
3710–3712.
[11] Caputo J.F., Martin A.R., Mallick S.K., J. Org. Chem. 39 (1974)
Acknowledgements
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[12] Guillaumet G., Coudert G., Ponchant M., Beaucourt J.P., Pichat L.,
J. Label. Compd. Radiopharm. 21 (1984) 161–172.
We thank F. Palomar for her collaboration in the
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[13] Adam W., Peters E.M., Peters K., Platsch H., Schmidt E., Von
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[14] Vo D., Matowe W.C., Ramesh M., Iqbal N., Wolowyk M.W.,
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