Polyalkoxybenzenes from plant raw materials 4. Parsley and dill seed extracts in the synthesis of polyalkoxy-3,5-diaryl-1,2,4-oxadiazoles with antiproliferative activity

Article abstract of DOI:10.1007/s11172-010-0387-6

Polyalkoxy-5-(4-hydroxyphenyl)-3-phenyl-1,2,4-oxadiazoles were prepared from allylpolyalkoxybenzenes, the main metabolites of parsley and dill seeds. Due to the spatial arrangement of the aryl substituents provided by 1,2,4-oxadiozole fragment, these compounds can be considered as structural analogs of natural antimitotic combretastatins. The antimitotic activity of the synthesized compounds was evaluated in vivo using sea urchin embryo test.

Full text of DOI:10.1007/s11172-010-0387-6

Russian Chemical Bulletin, International Edition, Vol. 59, No. 12, pp. 2268—2275, December, 2010
2268
Polyalkoxybenzenes from plant raw materials
4*. Parsley and dill seed extracts in the synthesis of
polyalkoxyꢀ3,5ꢀdiarylꢀ1,2,4ꢀoxadiazoles with antiproliferative activity
L. D. Konyushkin,a T. I. Godovikova,a S. K. Vorontsova,a D. V. Tsyganov,b I. B. Karmanova,a M. M. Raihstat,a
S. I. Firgang,a M. A. Pokrovskii,b A. G. Pokrovskii,b M. N. Semenova,c and V. V. Semenova
aN. D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences,
47 Leninsky prosp., 119991 Moscow, Russian Federation.
Fax: +7 (499) 135 5328. Eꢀmail: vs@ioc.ac.ru
bNovosibirsk State University,
2 ul. Pirogova, 119334 Novosibirsk, Russian Federation
cN. K. Kol´tsov Institute of Developmental Biology, Russian Academy of Sciences,
26 ul. Vavilova, 119334 Moscow, Russian Federation
Polyalkoxyꢀ5ꢀ(4ꢀhydroxyphenyl)ꢀ3ꢀphenylꢀ1,2,4ꢀoxadiazoles were prepared from allylpolyꢀ
alkoxybenzenes, the main metabolites of parsley and dill seeds. Due to the spatial arrangement
of the aryl substituents provided by 1,2,4ꢀoxadiozole fragment, these compounds can be considꢀ
ered as structural analogs of natural antimitotic combretastatins. The antimitotic activity of the
synthesized compounds was evaluated in vivo using sea urchin embryo test.
Key words: apiol, dillapiol, myristicin, combretastatin, oxadiazoles, antimitotic activity, sea
urchin embryo.
Various pharmacologically active allylpolyalkoxybenꢀ
zenes 1—5 are the main metabolites of Umbelliferae
plants.2,3 The protocol for their isolation from the parsley
and dill seeds using liquid CO2 extraction followed by
highꢀefficient distillation has been developed previously.3
Allylpolyalkoxybenzenes could be successively applied in
synthesis of the analogs of the natural antimitotic comꢀ
pounds with polyalkoxybenzene fragments, e.i., comꢀ
bretastatins.1,4—6 Combretastatins Aꢀ2 (CAꢀ2, 6) and
Aꢀ4 (CAꢀ4, 7) isolated from the bark of a South African
plant Combretum caffrum7 possess a strong antimitotic
activity owing to inhibition of tubulin polimerization
and alteration of mitotic spindle structure and function.8,9
In combretastatin molecule, two aromatic rings,
the A ring and the B ring, are linked by ethylene
bridge thus arranged at a certain distance and dihedral
angle. The bridge furnishes a cisꢀconfiguration of
the stilbene template necessary for efficient interaction
with tubulin.5,6,8 Phosphorylated waterꢀsoluble derivative
CAꢀ4 prodrug currently undergoes clinical evaluation as
antitumor agent.10,11
fects and ability to overcome multiꢀdrug resistance.5,6 The
availability of raw materials and simplicity of synthetic
procedures are also important.
Modification of the B ring of combretastatins by introꢀ
duction of different substituents, e.g., NH2, F, is widely
known.5,6 Various derivatives bearing different linkers that
stabilize the cisꢀconfiguration of the stilbene template were
reported. Besides, fiveꢀmembered heterocycles, including
A numerous modern studies in medicinal chemistry
are devoted to the design of novel synthetic analogs of
combretastatin, which possess the configuration stability
and strong antimitotic effect along with minimal side efꢀ
* For Part 3, see Ref. 1.
Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 12, pp. 2212—2219, December, 2010.
1066ꢀ5285/10/5912ꢀ2268 © 2010 Springer Science+Business Media, Inc.

Polyalkoxy 3,5ꢀdiarylꢀ1,2,4ꢀoxadiazoles
Russ.Chem.Bull., Int.Ed., Vol. 59, No. 12, December, 2010
2269
readily available 1,2,4ꢀoxadiazolines12 and 1,2,4ꢀoxadiꢀ
azoles1 are regarded as bioisosteric nonꢀisomerizable and
metabolically stable substitute for the double bond in
cisꢀstilbene. Recently, combretastatin derivatives with only
two alkoxy groups in the A ring, pꢀhydroxyphenyl or
Nꢀheterocyclic moiety as the B ring, and 1,2,4ꢀoxadiazole
fragment as the bridge were synthesized.13 Among these
compounds, derivatives with a pronounced cytotoxicity
against human cancer cell lines were identified.13 Howevꢀ
er, the biological activity of the compounds bearing
more than three methoxy groups in the A ring have
been less studied
Results and Discussion
Synthetic approach to heterocyclic combretastatin anꢀ
alogs on the base of triꢀ and tetraalkoxy benzenes is given
on Scheme 1. Reaction of aldehydes 8 with iodine in nearꢀ
ly equimolar ratio in aqueous ammonia19 resulted in polyꢀ
alkoxybenzonitriles 9, which were used for the synthesis of
intermediate compounds, amidoximes 10. Target 3,5ꢀdiꢀ
arylꢀ1,2,4ꢀoxadiazoles 11—19 (Table 1) were synthesized
at heating of equimolar amounts of amidoxime 10 and
the corresponding acid in the presence of carbonyldiꢀ
imidazole.
due probably to
low availability
of the correꢀ
sponding startꢀ
ing benzaldehyꢀ
des. At the same
time, these structures are of interest owing to noticeable
antiproliferative activity exhibited by a number of natural
compounds with tetraalkoxy moiety, for example, tetraꢀ
hydroisoquinolines of Cactaceae,14 flavonoids, and isoꢀ
flavonoids.15—17
The aim of the present work is the synthesis of novel
combretastatin analogs with optimal for the antimitotic
activity number of alkoxy groups in the A and B rings,
necessary spatial orientation of which provided by
the 1,2,4ꢀoxadiazole binding block. The A ring was
designed on the base of triꢀ and tetraalkoxy benzenes
of plant origin, while pꢀhydroxyphenyl or sixꢀmembered
Nꢀheterocycles were used as the B ring. Moreover, for
a more detailed study of the structure—activity relationꢀ
ship, we synthesized compounds with 3,4ꢀmethyleneꢀ
dioxyphenyl moiety as the A ring, which presented also
in the structures of the number of secondary plant metaꢀ
bolites.18
Combretastatin analogs 20—35 bearing 3,4ꢀmethylꢀ
enedioxyphenyl, 4ꢀmethoxyphenyl, 3,4ꢀdimethoxyphenyl,
or 4ꢀpyridyl moieties as the A ring (see Table 1 and Experꢀ
imental) were synthesized similar to compounds 11—19.
Biological study. Sea urchin embryos are widely used
as a model for identification of compound with antiprolifꢀ
erative activity.20,21 Recently we developed a simple and
effective phenotypic sea urchin embryo assay, which serves for
both the identification of the substances arresting cell diꢀ
vision and study the mechanism of antimitotic activity.22
The effective concentrations (EC) of the substances reꢀ
sulting in the alteration of sea urchin embryo cleavage are
comparable with the IC50 for mammalian and human tuꢀ
mor cell lines.22,23 The results of sea urchin embryo assay
for the compounds synthesized are summarized in Table 1.
It was found that compound 17 with myristicin fragꢀ
ment affected sea urchin embryo cleavage at concentraꢀ
tion of 4 μmol L–1. At the same time, no antimitotic acꢀ
tivity up to 4 μmol L–1 were found for 1,2,4ꢀoxadiazole
combretastatin analogs 11 and 14, derivatives of apiol and
dillapiol, respectively, prepared by introduction of the adꢀ
ditional methoxy group in the A ring.
Nꢀheterocycle
Replacement of the isovaniline moiety in the B ring by
pꢀhydroxyphenyl group (apiol 12 derivative) was found to
Scheme 1

2270
Russ.Chem.Bull., Int.Ed., Vol. 59, No. 12, December, 2010
Konyushkin et al.
Table 1. Structure, yields, melting points, 1H NMR spectral data (DMSOꢀd6), and biological activity of combretastatin analogs 11—35
Comꢀ
pound
А
В
Yield
(%)
M.p./°C
1H NMR, δ (J/Hz)
EC*
/μmol L–1
11
18
164—168
3.87 (s, 3 H, OМе); 3.89 (s, 6 H, 2 OМе);
6.16 (s, 2 H, OCH2O); 7.16 (d, 1 H, H(5´),
Ar, J = 8.5); 7.17 (s, 1 H, H(6), Ar); 7.55
(d, 1 H, H(2´), Ar, J = 2.15); 7.63 (dd,
1 H, H(6´), Ar, J1 = 2.15, J2 = 8.5); 9.71
(s, 1 H, OH)
>4
12
13
43
18
255—256
107—108
3.87 (s, 3 H, OМе); 3.88 (s, 3 H, OМе);
6.16 (s, 2 H, OCH2O); 7.00 (d, 2 H, H(3´),
H(5´), Ar, J = 8.7); 7.19 (s, 1 H, H(6), Ar);
8.01 (d, 2 H, H(2´), H(6´), Ar, J = 7.0);
10.52 (s, 1 H, OH)
>4
>4
1.61—1.70 (m, 2 H, Ha(3´), Ha(5´)); 1.98
(d, 2 H, He(3´), He(5´), J = 11.5); 2.57—2.63
(m, 2 H, Ha(2´), Ha(6´)); 2.98 (d, 2 H, He(2´),
He(6´), J = 12.4); 3.16 (m, 1 H, H(4´)); 3.38
(s, 6 H, 2 OМе); 6.14 (s, 2 H, OCH2O); 7.08
(s, 1 H, H(6), Ar)
14
29
168—172
3.79 (s, 3 H, OМе); 3.89 (s, 3 H, OМе); 3.99
(s, 3 H, OМе); 6.13 (s, 2 H, OCH2O); 7.09 (s,
1 H, H(6), Ar); 7.16 (d, 1 H, H(5´), Ar, J = 8.5);
7.55 (d, 1 H, H(2´), Ar, J = 2.15); 7.63 (dd, 1 H,
H(6´), Ar, J1 = 2.15, J2 = 8.5); 9.71 (s, 1 H, OH)
3.79 (s, 3 H, OМе); 4.00 (s, 3 H, OМе); 6.12
(s, 2 H, OCH2O); 6.99 (d, 2 H, H(3´), H(5´),
Ar, J = 8.7); 7.10 (s, 1 H, H(6), Ar); 8.00 (d, 2 H,
H(2´), H(6´), Ar, J = 8.7); 10.51 (s, 1 H, OH)
1.62—1.68 (m, 2 H, Ha(3´), Ha(5´)); 1.98 (d, 2 H,
He(3´), He(5´), J = 12.1); 2.57—2.63 (m, 2 H,
>4
15
16
59
24
230—231
100—101
1
>4
Ha(2´), Ha(6´)); 2.95—3.00 (m, 2 H, He(2´), Hе(6´));
3.15 (m, 1 H, H(4´)); 3.75 (s, 1 H, OМе); 3.98 (s,
3 H, OМе); 6.11 (s, 2 H, OCH2O); 7.00 (s, 1 H,
H(6), Ar)
17
32
180—183
3.89 (s, 3 H, OМе); 3.94 (s, 3 H, OМе); 6.13 (s,
2 H, OCH2O); 7.16 (d, 1 H, H(5´), Ar, J = 8.5);
7.22 (d, 1 H, H(2) or H(6), Ar, J = 1.4);
4
7.34 (d, 1 H, H(2) or H(6), Ar, J = 1.4);
7.56 (d, 1 H, H(2´), Ar, J = 2.15); 7.63 (dd, 1 H,
H(6´), Ar, J1 = 2.15, J2 = 8.5); 9.71 (s, 1 H, OH)
3.93 (s, 3 H, OМе); 6.13 (s, 2 H, OCH2O); 7.00
(d, 2 H, H(3´), H(5´), Ar, J = 8.7); 7.22 (d, 1 H,
H(6), Ar, J = 0.95); 7.33 (d, 1 H, H(2), Ar,
J = 0.95); 8.02 (d, 2 H, H(2´), H(6´), Ar, J = 8.7);
10.53 (s, 1 H, OH)
1.62—1.71 (m, 2 H, Ha(3´), Ha(5´)); 1.97 (d, 2 H,
He(3´), He(5´); J = 11.8), 2.61 (т, 2 H, Ha(2´),
Ha(6´), J = 12.5); 2.99 (d, 2 H, He(2´), He(6´),
J = 12.5); 3.18 (m, 1 H, H(4´)); 3.90 (s, 3 H, OМе);
6.12 (s, 2 H, OCH2O); 7.15 (d, 1 H, H(6), Ar,
J = 1.4); 7.26 (d, 1 H, H(2), Ar, J = 1.4)
18
19
11
21
205—206
104—105
2
>4
(to be continued)

Polyalkoxy 3,5ꢀdiarylꢀ1,2,4ꢀoxadiazoles
Russ.Chem.Bull., Int.Ed., Vol. 59, No. 12, December, 2010
2271
Table 1 (continued)
Comꢀ
pound
А
В
Yield M.p./°C
(%)
1H NMR, δ (J/Hz)
EC*
/μmol L–1
20
15
204—206
6.16 (s, 2 H, OCH2O); 6.99 (d, 2 H, H(3´), H(5´),
Ar, J = 8.7); 7.12 (d, 1 H, H(5), Ar, J = 8.1); 7.50
(d, 1 H, H(2), Ar, J = 1.7); 7.64 (dd, 1 H, H(6), Ar,
J1 = 1.7, J2 = 8.1); 8.01 (d, 2 H, H(2´), H(6´), Ar,
J = 8.7); 10.54 (s, 1 H, OH)
0.5
21
35
96—97
1.61—1.70 (m, 2 H, Ha(3´), Ha(5´)); 1.96 (d, 2 H,
>4
He(3´), He(5´); J = 12.8); 2.57—2.63 (m, 2 H, Ha(2´),
Ha(6´)); 2.98 (m, 2 H, He(2´), He(6´)); 3.12—3.19
(m, 1 H, H(4´)); 6.15 (s, 2 H, OCH2O); 7.19 (d, 1 H,
H(5), Ar, J = 8.1); 7.44 (d, 1 H, H(2), Ar, J = 1.7);
7.57 (dd, 1 H, H(6), Ar, J1 = 1.7, J2 = 8.1)
22
23
31
41
207—208
199—200
6.15 (s, 2 H, OCH2O); 7.12 (d, 1 H, H(5), Ar,
J = 8.1); 7.53 (d, 1 H, H(2), Ar, J = 1.7); 7.67 (dd,
1 H, H(6), Ar, J1 = 1.7, J2 = 8.1); 8.07 (d, 2 H,
H(3´), H(5´), Py, J = 5.5); 8.90 (d, 2 H, H(2´),
H(6´), Py, J = 5.5)
5
5
6.18 (s, 2 H, OCH2O); 7.16 (d, 1 H, H(5), Ar,
J = 8.1); 7.56 (d, 1 H, H(2), Ar, J = 1.7); 7.70
(dd, 1 H, H(6), Ar, J1 = 1.7, J2 = 8.1); 8.94
(dd, 1 H, H(5´), pyrazine, J1 = 1.4, J2 = 2.4); 8.98
(d, 1 H, H(6´), pyrazine, J = 2.4); 9.49 (d, 1 H, H(3´),
pyrazine, J = 1.4)
24
25
71
41
122—123
119—120
3.89 (s, 3 H, OМе); 6.17 (s, 2 H, OCH2O); 7.13
(d, 1 H, H(5), Ar, J = 8.1), 7.31 (ddd, 1 H, H(5´),
Ar, J1 = 1.1, J2 = 2.5, J3 = 7.8); 7.53 (d, 1 H, H(2),
Ar, J = 1.7); 7.58 (t, 1 H, H(4´), Ar, J = 7.8); 7.64
(dd, 1 H, H(2´), Ar, J1 = 1.1, J2 = 2.5); 7.67 (dd,
1 H, H(6), Ar, J1 = 1.7, J2 = 8.1); 7.75 (dt, 1 H,
H(6´), Ar, J1 = 1.1, J2 = 7.8)
0>4
0>4
1.41 (t, 3 H, МеCH2O, J = 7.0); 4.23 (q, 2 H,
МеCH2O, J = 7.0); 6.17 (s, 2 H, OCH2O); 7.13
(d, 1 H, H(5), Ar, J = 8.1); 7.16 (t, 1 H, H(5´),
Ar, J = 7.7); 7.30 (d, 1 H, H(3´), Ar, J = 8.4);
7.52 (d, 1 H, H(2), Ar, J = 1.7); 7.64—7.68 (m,
2 H, H(4´), H(6), Ar); 8.06 (dd, 1 H, H(6´), Ar,
J1 = 1.8, J2 = 7.7)
26
27
28
52
35
61
192—193
207—209
150—151
3.88 (s, 3 H, OМе); 3.90 (s, 3 H, OМе); 6.16 (s,
2 H, OCH2O); 7.12 (d, 1 H, H(2), Ar, J = 8.1,);
7.22 (d, 1 H, H(5´), Ar, J = 8.4); 7.52 (d, 1 H, H(2),
Ar, J = 1.7); 7.63 (d, 1 H, H(2´), Ar, J = 2.0); 7.66
(dd, 1 H, H(6), Ar, J1 = 1.7, J2 = 8.1); 7.78 (dd,
1 H, H(6´), Ar, J1 = 2.0, J2 = 8.4)
0>4
0>4
0>4
6.16 (s, 2 H, OCH2O); 6.21 (s, 2 H, OCH2O); 7.12
(d, 1 H, H(5), Ar, J = 8.1); 7.18 (d, 1 H, H(5´),
Ar, J = 8.1); 7.51 (d, 1 H, H(2), Ar, J = 1.7); 7.63
(d, 1 H, H(2´), Ar, J = 1.7); 7.64 (dd, 1 H, H(6),
Ar, J1 = 1.7, J2 = 8.1); 7.76 (dd, 1 H, H(6´), Ar,
J1 = 1.7, J2 = 8.1)
3.78 (s, 3 H, OМе); 3.92 (s, 6 H, 2 OМе); 6.16
(s, 2 H, OCH2O); 7.12 (d, 1 H, H(5), Ar, J = 8.1);
7.42 (s, 2 H, H(2´), H(6´), Ar); 7.53 (d, 1 H,
H(2), Ar, J = 1.7); 7.67 (dd, 1 H, H(6), J1 = 1.7,
J2 = 8.1)
(to be continued)

2272
Russ.Chem.Bull., Int.Ed., Vol. 59, No. 12, December, 2010
Konyushkin et al.
Table 1 (continued)
Comꢀ
pound
А
В
Yield
(%)
M.p./°C
1H NMR, δ (J/Hz)
EC*
/μmol L–1
29
30
15
189—191
3.84 (s, 3 H, OМе); 3.85 (s, 3 H, OМе); 7.00
>4
(d, 2 H, H(3´), H(5´), Ar, J = 8.7); 7.15 (d, 1 H,
H(5), Ar, J = 8.4); 7.55 (d, 1 H, H(2), Ar,
J = 2.0); 7.67 (dd, 1 H, H(6), Ar, J1 = 2.0,
J2 = 8.4); 8.03 (d, 2 H, H(2´), H(6´), Ar, J = 8.4);
10.54 (s, 1 H, OH)
1.59—1.75 (m, 2 H, Ha(3´), Ha(5´)); 1.98 (d, 2 H,
He(3´), He(5´), J = 11.8); 2.61 (t, 2 H, Ha(2´),
Ha(6´), J = 11.8); 2.99 (d, 2 H, He(2´), He(6´),
J = 11.8); 3.17 (m, 1 H, H(4´)); 3.83 (s, 6 H,
2 OМе); 7.12 (d, 1 H, H(5), Ar, J = 8.4); 7.46
(d, 1 H, H(2), Ar, J = 1.9); 7.59 (dd, 1 H, H(6),
Ar, J1 = 1.9, J2 = 8.4)
23
82—84
>4
31
32
35
65
155—157
110—112
3.85 (s, 3 H, OМе); 3.87 (s, 3 H, OМе); 3.89
(s, 3 H, OМе); 7.15 (d, 1 H, H(5), Ar, J = 8.4);
7.19 (d, 2 H, H(3´), H(5´), Ar, J = 8.9); 7.55
(d, 1 H, H(2), Ar, J = 2.0); 7.68 (dd, 1 H, H(6),
Ar, J1 = 2.0, J2 = 8.4); 8.13 (d, 2 H, H(2´), H(6´),
Ar, J = 8.9)
>4
>4
3.85 (s, 3 H, OМе); 3.87 (s, 3 H, OМе ); 3.89
(s, 3 H, OМе); 7.17 (d, 1 H, H(5), Ar, J = 8.4);
7.31 (dd, 1 H, H(4´), Ar, J1 = 2.3, J2 = 8.3);
7.57 (s, 1 H, H(2), Ar), 7.59 (т, 1 H, H(5´), Ar,
J = 8.1), 7.66 (t, 1 H, H(2´), Ar, J = 2.3),
7.70 (dd, 1 H, H(6), Ar, J1 = 1.7, J2 = 8.4);
7.77 (d, 1 H, H(6´), Ar, J = 7.7)
33
34
75
17
165—167
145—146
3.79 (s, 3 H, OМе); 3.86 (s, 3 H, OМе ); 3.88
(s, 3 H, OМе); 3.93 (s, 6 H, 2 OМе); 7.16 (d,
1 H, H(5), Ar, J = 8.4); 7.42 (s, 2 H, H(2´), H(6´),
Ar); 7.56 (d, 1 H, H(2), Ar, J = 2.0); 7.70 (dd,
1 H, H(6), Ar, J1 = 2.0, J2 = 8.4)
>4
3.87 (s, 3 H, OМе); 6,69 (t, 1 H, H(5´), Ar,
J = 8.0); 6.96 (d, 1 H, H(3´), Ar, J = 8.4);
6.98 (s, 2 H, NH2); 7.14 (d, 2 H, H(3), H(5),
Ar, J = 8.8); 7.32—7.37 (m, 1 H, H(4´), Ar);
7.84 (dd, 1 H, H(6´), Ar, J1 = 1.5, J2 = 8.0);
8.09 (d, 2 H, H(2), H(6), Ar, J = 8.8)
0>4
35
36
169—170
3.81 (s, 3 H, OМе); 3.93 (s, 6 H, 2 OМе);
7.47 (s, 2 H, H(2´), H(6´), Ar); 8.01 (d, 2 H,
H(3), H(5), Ar, J = 5.4); 8.84 (d, 2 H, H(2),
H(6), Ar, J = 5.4)
>10
* EC is an effective concentration of the compound resulting in the cleavage alteration of the sea urchin embryos.
be ineffective, while dillapiol and myristicin derivatives,
15 and 18, respectively, affect the cell cleavage at
1—2 μmol L–1. Methylenedioxy derivative 20 without
methoxy groups in the A ring was even more active, its EC
was 0.5 μmol L–1. Thus, the replacement of the pꢀOMe
group by the pꢀOH in the B ring of the oxadiazole derivaꢀ
tives increases the antimitotic effect, while in the combreꢀ
tastatins the same modification resulted in the noticeable
decrease in activity.8,24
Replacement of pꢀhydroxyphenyl moiety (compounds
12, 15, 18, and 20) by pyperidinꢀ4ꢀyl fragment (comꢀ
pounds 13, 16, 19, and 21) led to loss of activity, however,
the same modification of the B ring of the derivatives bearꢀ
ing two dialkoxy fragments in the A ring resulted in the
increase in cytotoxicity.13 Piperonylꢀcontaining derivaꢀ
tives 22 and 23 with pyridine and pyridazine as the B ring,
respectively, exhibited cleavage alteration at 5 μmol L–1
At the same time, according to the published data, replaceꢀ
.

Polyalkoxy 3,5ꢀdiarylꢀ1,2,4ꢀoxadiazoles
Russ.Chem.Bull., Int.Ed., Vol. 59, No. 12, December, 2010
2273
the amounts of THF and NH4OH were raised by 15% to the
mentionedꢀabove procedure. A mixture of aldehyde 8c (57.6 g,
320 mmol) and I2 (100.8 g, 397 mmol) in THF (345 mL) and
28% aqueous NH4OH (3200 mL) was stirred at 20 °C for 30 h.
The reaction mixture was workedꢀup as above to give an insepaꢀ
rable mixture (55 g) of nitrile 9c (88%) and unreacted aldehyde
8c (12%). This mixture was further oxidized with KMnO4 (6 g per
60 mL of H2O) in acetone (150 mL) at reflux for 4 h. Removal of
the solvent in vacuo and recrystallization of the residue from
EtOH afforded myristicin acid (4.5 g, 7%, m.p. 212 °C, cf. Ref. 27:
m.p. 211 °C) and nitrile 9c (37 g, 65%), m.p. 162—164 °C.
Found (%): C, 61.12; H, 4.05; N, 7.83. C9H7NO3. Calculatꢀ
ment of the piperidine moiety by the pyridine ring gave
inactive compounds.13 Probably, this discrepancy in the
biological activity of combretastatin derivatives at bioisoꢀ
steric replacement of the fragments is due to the difference
in the cellular mechanisms of action of the compounds.
As an example, the results obtained using the sea urchin
embryo suggested the nonꢀtubulin antiproliferative activiꢀ
ty of compound 20.
Moving from methylenedioxy derivatives to dimethoxy
ones caused the loss of the activity (compounds 29—33).
Similar change in the structure of the natural compounds,
αꢀconidendrin, podophyllotoxin, and sikkimotoxin, yieldꢀ
ed 10ꢀfold decrease in cytotoxicity.25
According to the preliminary tests, 1,2,4ꢀoxadiazole
combretastatin derivatives 11, 14, and 17, pyrazine derivꢀ
ative 23, and compound 35 showed cytotoxicity against
U937, MT4, and CEM13 human leukemia cells.
In summary, allylpolyalkoxybenzenes extracted from
parsley and dill seeds are easily available starting material
for the synthesis of biologically active compounds posꢀ
sessing antiproliferative activity, which could be used later
in design of novel antitumor agents.
1
ed (%): C, 61.02; H, 3.98; N, 7.91. H NMR (CDCl3), δ: 3.92
(s, 3 H, OMe); 6.08 (s, 2 H, OCH2O); 6.80 (d, 1 H, H(2), Ar,
J = 1.4 Hz); 6.86 (d, 1 H, H(6), Ar, J = 1.4 Hz). MS, m/z (Irel (%)):
177 [M]+ (100), 176 (77), 162 (15), 132 (42), 104 (26).
Synthesis of amidoximes 10a—d (general procedure). To
a solution of nitrile 9 (10 mmol) and NH2OH•HCl (1.4 g,
20 mmol) in EtOH (20 mL), a solution of NaHCO3 (1.68 g,
20 mmol) in water (5 mL) was added at room temperature with
stirring. The reaction mixture was refluxed for 12 h, the solvent
was removed in vacuo. Recrystallization of the residue from
aqueous EtOH afforded amidoximes 10a—d in 65—75% yields
(85—90% purity), which were used on the next step without
further purification.
2,5ꢀDimethoxyꢀ3,4ꢀmethylenedioxybenzamidoxime (10a).
M.p. 150—152 °C. Found (%): C, 50.18; H, 5.16; N, 11.50.
C10H12N2O5. Calculated (%): C, 50.00; H, 5.04; N, 11.66.
1H NMR (DMSOꢀd6), δ: 3.77 (s, 3 H, OMe); 3.80 (s, 3 H,
OMe); 5.60 (s, 2 H, NH2); 6.04 (s, 2 H, OCH2O); 6.66 (s, 1 H,
H(6), Ar); 9.40 (s, 1 H, OH). MS, m/z (Irel (%)): 240 [M]+ (13),
223 (16), 209 (6), 208 (32), 207 (52), 193 (30), 192 (46), 68 (49),
66 (84), 59 (100).
2,3ꢀDimethoxyꢀ4,5ꢀmethylenedioxybenzamidoxime (10b).
M.p. 147—148 °C. Found (%): C, 50.12; H, 5.12; N, 11.46.
C10H12N2O5. Calculated (%): C, 50.00; H, 5.04; N, 11.66.
1H NMR (DMSOꢀd6), δ: 3.69 (s, 3 H, OMe); 3.83 (s, 3 H,
OMe); 5.60 (s, 2 H, NH2); 6.01 (s, 2 H, OCH2O); 6.62 (s, 1 H,
H(6), Ar); 9.41 (s, 1 H, OH). MS, m/z (Irel (%)): 240 [M]+ (13),
223 (17), 208 (28), 207 (56), 193 (18), 192 (38), 68 (34), 66 (72),
59 (100).
3ꢀMethoxyꢀ4,5ꢀmethylenedioxybenzamidoxime (10c). M.p.
170—171 °C. Found (%): C, 51.58; H, 4.92; N, 13.19. C9H10N2O4.
Calculated (%): C, 51.43; H, 4.80; N, 13.33. 1H NMR (DMSOꢀd6),
δ: 3.85 (s, 3 H, OMe); 5.75 (s, 2 H, NH2); 6.01 (s, 2 H, OCH2O);
6.89 (s, 1 H, Ar); 6.98 (s, 1 H, Ar); 9.52 (s, 1 H, OH). MS, m/z
(Irel (%)): 210 [M]+ (31), 193 (35), 179 (8), 178 (22), 77 (53),
65 (90), 63 (100).
3,4ꢀMethylenedioxybenzamidoxime (10d). M.p. 150—152 °C
(cf. Ref. 28: m.p. 151 °C). 1H NMR (DMSOꢀd6), δ: 5.71 (s, 2 H,
NH2); 6.03 (s, 2 H, OCH2O); 6.90 (d, 1 H, H(5), Ar, J = 8.5 Hz);
7.18 (s, 1 H, H(2), Ar); 7.19 (dd, 1 H, H(6), Ar, J = 1.6 Hz,
J = 8.5 Hz); 9.50 (s, 1 H, OH). MS, m/z (Irel (%)): 180 [M]+
(26), 163 (32), 149 (7), 148 (19), 77 (50), 65 (91), 63 (100).
3,4ꢀDimethoxybenzamidoxime (10e). To a solution of 3,4ꢀdiꢀ
methoxybenzonitrile (5.0 g, 30.6 mmol) and NH2OH•HCl
(4.26 g, 61.3 mmol) in EtOH (150 mL), a solution of NaOH
(2.45 g, 61.3 mmol) in water (20 mL) was added at room temperꢀ
ature with stirring. The reaction mixture was refluxed for 14 h,
the solvent was removed in vacuo, the residue was washed with
water (2×20 mL), and recrystallized from 50% aqueous EtOH
Experimental
1
H and 13C NMR spectra were run on a Bruker DRXꢀ500
instrument at 500.13 and 125.76 MHz, respectively, in CDCl3 or
DMSOꢀd6 with Me4Si as internal standard. Mass spectra were
recorded on a Finnigan MAT INCOS 50 quadrupole mass specꢀ
trometer (direct inlet, ionization potential 70 eV). Elemental
analysis was carried out on an automated Perkin—Elmer 2400
CHN microanalyzer. TLC was performed on Merck 60 F254
plates. Preparative chromatography was carried out on silica gel
Acros 0.035—0.070 mm, 60 A. Melting points were determined
at Boetius apparatus and uncorrected.
Synthesis of polyalkoxybenzonitriles 9 (general procedure).
A solution of aldehyde 8 (see Ref. 3) (200 mmol) in THF
(300 mL) was added to a solution of I2 (56.85 g, 224 mmol) in
28% aqueous NH4OH (1800 mL). The resulting mixture was
stirred at 20 °C for 12 h and extracted with CHCl3 (3×300 mL).
The combined organic layers were washed with water (300 mL),
10% aqueous Na2S2O3 (300 mL), and brine, dried with Na2SO4,
and the solvent was removed in vacuo. The residue was recrystalꢀ
lized from EtOH.
2,5ꢀDimethoxyꢀ3,4ꢀmethylenedioxybenzonitrile (9a). Yield
89%, m.p. 134—135 °C (cf. Ref. 26: m.p. 135.5 °C). Found (%):
C, 58.06; H, 4.47; N, 6.69. C10H9NO4. Calculated (%): C, 57.97;
H, 4.38; N, 6.76. 1H NMR (CDCl3), δ: 3.86 (s, 3 H, OMe); 4.07
(s, 3 H, OMe); 6.07 (s, 2 H, OCH2O); 6.71 (s, 1 H, Ar). MS, m/z
(Irel (%)): 207 [M]+ (100), 192 (95), 162 (17), 134 (20).
2,3ꢀDimethoxyꢀ4,5ꢀmethylenedioxybenzonitrile (9b). Yield
91%, m.p. 92—94 °C. Found (%): C, 57.89; H, 4.44; N, 6.70.
C10H9NO4. Calculated (%): C, 57.97; H, 4.38; N, 6.76. 1H NMR
(CDCl3), δ: 3.95 (s, 3 H, OMe); 4.03 (s, 3 H, OMe); 6.03 (s, 2 H,
OCH2O); 6.64 (s, 1 H, Ar). MS, m/z (Irel (%)): 207 [M]+ (100),
192 (93), 162 (13), 134 (15).
3ꢀMethoxyꢀ4,5ꢀmethylenedioxybenzonitrile (9c). When the
reaction was carried out with poor soluble myristicin aldehyde,

2274
Russ.Chem.Bull., Int.Ed., Vol. 59, No. 12, December, 2010
Konyushkin et al.
(50 mL) to give amidoxime 10e (4.3 g, 71.5%), m.p. 142—143 °C
(cf. Ref. 29: m.p. 137 °C). 1H NMR (DMSOꢀd6), δ: 3.76 (s, 6 H,
2 OMe); 5.73 (s, 2 H, NH2); 6.94 (d, 1 H, H(5), Ar, J = 8.4 Hz);
7.23 (dd, 1 H, H(6), Ar, J = 2.0 Hz, J = 8.4 Hz); 7.25 (d, 1 H, H(2),
Ar, J = 2.0 Hz); 9.46 (s, 1 H, OH). MS, m/z (Irel (%)): 196 [M]+
(59), 179 (55), 165 (12), 164 (34), 79 (72), 77 (47), 51 (100).
Synthesis of 3,5ꢀdiarylꢀ1,2,4ꢀoxadiazoles 11—35 (general
procedure). Carbonyldiimidazole (0.58 g, 3.6 mmol) was added
to a stirred suspension of crude amidoxime 10 (3 mmol) in anhyꢀ
drous MeCN (5 mL) and stirring was continued at room temperꢀ
ature for 1 h until the amidoxime was dissolved completely.
Then the corresponding carboxylic acid (3 mmol) was added
and stirring was continued for 12 h. The solvent was removed
in vacuo, the solid residue was dissolved in anhydrous DMF
(5 mL), and the solution was stirred at 120—125 °C for 3 h. The
solvent was removed in vacuo, the target product was isolated by
column chromatography (silica gel, elution with ethyl acetate—hexꢀ
anes, 1 : 4). Yields, melting points, and 1H NMR spectral data of
the synthesized combretastatin analogs are given in Table 1.
3ꢀ(2,5ꢀDimethoxyꢀ3,4ꢀmethylenedioxyphenyl)ꢀ5ꢀ(3ꢀhydrꢀ
oxyꢀ4ꢀmethoxyphenyl)ꢀ1,2,4ꢀoxadiazole (11). Found (%): C, 58.12;
H, 4.44; N, 7.63. C18H16N2O7. Calculated (%): C, 58.07;
H, 4.33; N, 7.52. MS, m/z (Irel (%)): 372 [M]+ (13), 266 (17),
193 (26), 178 (17), 151 (100).
3ꢀ(2,5ꢀDimethoxyꢀ3,4ꢀmethylenedioxyphenyl)ꢀ5ꢀ(4ꢀhydroxyꢀ
phenyl)ꢀ1,2,4ꢀoxadiazole (12). Found (%): C, 59.59; H, 4.07;
N, 8.28. C17H14N2O6. Calculated (%): C, 59.65; H, 4.12; N, 8.18.
MS, m/z (Irel (%)): 342 [M]+ (5), 313 (0.5), 297 (1), 221 (5),
193 (32), 178 (11), 121 (100), 119 (10), 109 (10), 93 (25), 77 (10).
3ꢀ(2,5ꢀDimethoxyꢀ3,4ꢀmethylenedioxyphenyl)ꢀ5ꢀ(piperidinꢀ
4ꢀyl)ꢀ1,2,4ꢀoxadiazole (13). Found (%): C, 57.80; H, 5.84; N, 12.51.
C16H19N3O5. Calculated (%): C, 57.65; H, 5.75; N, 12.61. MS,
m/z (Irel (%)): 333 [M]+ (2), 251 (5), 226 (11), 225 (100), 223
(14), 208 (7), 207 (8), 109 (11), 83 (24), 82 (54), 68 (17), 55 (5).
3ꢀ(2,3ꢀDimethoxyꢀ4,5ꢀmethylenedioxyphenyl)ꢀ5ꢀ(3ꢀhydrꢀ
oxyꢀ4ꢀmethoxyphenyl)ꢀ1,2,4ꢀoxadiazole (14). Found (%): C, 58.01;
H, 4.25; N, 7.49. C18H16N2O7. Calculated (%): C, 58.07;
H, 4.33; N, 7.52. MS, m/z (Irel (%)): 372 [M]+ (41), 266 (29),
193 (82), 178 (17), 151 (100).
C
15H17N3O4. Calculated (%): C, 59.40; H, 5.65; N, 13.85. MS,
m/z (Irel (%)): 303 [M]+ (4), 195 (63), 193 (12), 192 (6), 178 (6),
109 (30), 84 (6), 83 (40), 82 (100), 76 (6), 68 (26), 55 (96).
5ꢀ(4ꢀHydroxyphenyl)ꢀ3ꢀ(3,4ꢀmethylenedioxyphenyl)ꢀ1,2,4ꢀoxaꢀ
diazole (20). Found (%): C, 63.74; H, 3.51; N, 9.71. C15H10N2O4.
Calculated (%): C, 63.83; H, 3.57; N, 9.92. MS, m/z (Irel (%)):
282 [M]+ (38), 163 (50), 162 (13), 121 (43), 119 (10), 106 (11),
105 (23), 93 (61), 77 (100).
3ꢀ(3,4ꢀMethylenedioxyphenyl)ꢀ5ꢀ(piperidinꢀ4ꢀyl)ꢀ1,2,4ꢀoxaꢀ
diazole (21). Found (%): C, 61.63; H, 5.65; N, 15.21. C14H15N3O3.
Calculated (%): C, 61.53; H, 5.53; N, 15.38. MS, m/z (Irel (%)):
273 [M]+ (3), 165 (27), 164 (26), 163 (11), 146 (9), 109 (17),
84 (7), 83 (38.5), 82 (100), 81 (59), 77 (13), 68 (40), 55 (95).
3ꢀ(3,4ꢀMethylenedioxyphenyl)ꢀ5ꢀ(pyridinꢀ4ꢀyl)ꢀ1,2,4ꢀoxadiꢀ
azole (22). Found (%): C, 63.04; H, 3.48; N, 15.62. C14H9N3O3.
Calculated (%): C, 62.92; H, 3.39; N, 15.72. MS, m/z (Irel (%)):
267 [M]+ (100), 266 (12), 163 (66), 162 (22), 146 (9), 133 (21),
106 (20), 78 (34), 77 (25).
3ꢀ(3,4ꢀMethylenedioxyphenyl)ꢀ5ꢀ(pyrazinꢀ2ꢀyl)ꢀ1,2,4ꢀoxadiꢀ
azole (23). Found (%): C, 58.12; H, 2.94; N, 21.04. C13H8N4O3.
Calculated (%): C, 58.21; H, 3.01; N, 20.89. MS, m/z (Irel (%)):
268 [M]+ (100), 267 (16), 239 (1), 163 (51), 162 (25), 147 (6),
146 (16), 133 (7), 107 (6), 105 (6), 79 (39), 77 (31).
5ꢀ(3ꢀMethoxyphenyl)ꢀ3ꢀ(3,4ꢀmethylenedioxyphenyl)ꢀ1,2,4ꢀ
oxadiazole (24). Found (%): C, 64.75; H, 3.96; N, 9.60.
C16H12N2O4. Calculated (%): C, 64.86; H, 4.08; N, 9.45. MS,
m/z (Irel (%)): 296 [M]+ (100), 163 (87), 162 (27), 147 (18),
135 (50), 133 (12), 107 (25), 77 (65).
5ꢀ(2ꢀEthoxyphenyl)ꢀ3ꢀ(3,4ꢀmethylenedioxyphenyl)ꢀ1,2,4ꢀ
oxadiazole (25). Found (%): C, 65.95; H, 4.72; N, 8.84.
C17H14N2O4. Calculated (%): C, 65.80; H, 4.55; N, 9.03. MS,
m/z (Irel (%)): 310 [M]+ (41), 295 (3), 265 (6), 163 (38), 162 (58),
149 (8), 147 (59), 133 (92), 121 (100), 119 (48), 105 (24), 77 (48).
5ꢀ(3,4ꢀDimethoxyphenyl)ꢀ3ꢀ(3,4ꢀmethylenedioxyphenyl)ꢀ
1,2,4ꢀoxadiazole (26). Found (%): C, 62.45; H, 4.22; N, 8.72.
C17H14N2O5. Calculated (%): C, 62.57; H, 4.32; N, 8.58. MS, m/z
(Irel (%)): 326 [M]+ (37), 165 (100), 163 (29), 137 (8), 121 (6), 77 (29).
3,5ꢀBis(3,4ꢀmethylenedioxyphenyl)ꢀ1,2,4ꢀoxadiazole (27).
Found (%): C, 62.12; H, 3.33; N, 8.87. C16H10N2O5. Calculated (%):
C, 61.94; H, 3.25; N, 9.03. MS, m/z (Irel (%)): 310 [M]+ (43),
163 (30), 162 (13), 149 (100), 147 (8), 146 (19), 121 (18), 77 (27).
3ꢀ(3,4ꢀMethylenedioxyphenyl)ꢀ5ꢀ(3,4,5ꢀtrimethoxyphenyl)ꢀ
1,2,4ꢀoxadiazole (28). Found (%): C, 60.80; H, 4.72; N, 7.76.
C18H16N2O6. Calculated (%): C, 60.67; H, 4.53; N, 7.86. MS,
m/z (Irel (%)): 356 [M]+ (28), 341 (1), 195 (100), 178 (13), 163
(10), 161 (8), 152 (14), 122 (6), 77(12).
3ꢀ(3,4ꢀDimethoxyphenyl)ꢀ5ꢀ(4ꢀhydroxyphenyl)ꢀ1,2,4ꢀoxadiꢀ
azole (29). Found (%): C, 64.36; H, 4.67; N, 9.51. C16H14N2O4.
Calculated (%): C, 64.42; H, 4.73; N, 9.39. MS, m/z (Irel (%)):
298 [M]+ (17), 179 (21), 163 (9), 136 (22), 121 (90), 119 (37),
105 (22), 93 (46), 77 (35).
3ꢀ(3,4ꢀDimethoxyphenyl)ꢀ5ꢀ(piperidinꢀ4ꢀyl)ꢀ1,2,4ꢀoxadiꢀ
azole (30). Found (%): C, 62.43; H, 6.74; N, 14.36. C15H19N3O3.
Calculated (%): C, 62.27; H, 6.62; N, 14.52. MS, m/z (Irel (%)):
289 [M]+ (1), 181 (83), 179 (11), 163 (8), 109 (28), 84 (8),
83 (50), 82 (100), 77 (12), 68 (33), 55 (99).
3ꢀ(2,3ꢀDimethoxyꢀ4,5ꢀmethylenedioxyphenyl)ꢀ5ꢀ(4ꢀhydroxyꢀ
phenyl)ꢀ1,2,4ꢀoxadiazole (15). Found (%): C, 59.55; H, 4.01;
N, 8.02. C17H14N2O6. Calculated (%): C, 59.65; H, 4.12; N, 8.18.
MS, m/z (Irel (%)): 342 [M]+ (38.5), 327 (3), 297 (10), 223 (10),
221 (15), 207 (6), 193 (54), 121 (100), 119 (11), 93 (47), 77 (18).
3ꢀ(2,3ꢀDimethoxyꢀ4,5ꢀmethylenedioxyphenyl)ꢀ5ꢀ(piperidinꢀ
4ꢀyl)ꢀ1,2,4ꢀoxadiazole (16). Found (%): C, 57.74; H, 5.78;
N, 12.46. C16H19N3O5. Calculated (%): C, 57.65; H, 5.75; N, 12.61.
MS, m/z (Irel (%)): 333 [M]+ (2), 226 (11), 225 (90), 223 (12),
208 (12), 207 (11), 109 (17), 83 (47), 82 (100), 68 (34), 55 (96).
5ꢀ(3ꢀHydroxyꢀ4ꢀmethoxyphenyl)ꢀ3ꢀ(3ꢀmethoxyꢀ4,5ꢀmethylꢀ
enedioxyphenyl)ꢀ1,2,4ꢀoxadiazole (17). Found (%): C, 59.58;
H, 4.23; N, 8.07. C17H14N2O6. Calculated (%): C, 59.65; H, 4.12;
N, 8.18. MS, m/z (Irel (%)): 342 [M]+ (88), 193 (26), 171 (8),
151 (100).
5ꢀ(4ꢀHydroxyphenyl)ꢀ3ꢀ(3ꢀmethoxyꢀ4,5ꢀmethylenedioxyꢀ
phenyl)ꢀ1,2,4ꢀoxadiazole (18). Found (%): C, 61.75; H, 3.96;
N, 8.90. C16H12N2O5. Calculated (%): C, 61.54; H, 3.87; N, 8.97.
MS, m/z (Irel (%)): 312 [M]+ (79), 297 (1), 193 (89), 177 (6), 156
(17), 148 (30), 121 (100), 119 (18), 93 (41), 77 (18).
3ꢀ(3,4ꢀDimethoxyphenyl)ꢀ5ꢀ(4ꢀmethoxyphenyl)ꢀ1,2,4ꢀoxadiꢀ
azole (31). Found (%): C, 65.29; H, 5.09; N, 8.84. C17H16N2O4.
Calculated (%): C, 65.38; H, 5.16; N, 8.97. MS, m/z (Irel (%)):
312 [M]+ (10), 283 (1), 179 (27), 164 (8), 163 (6), 136 (32),
135 (100), 133 (16), 119 (14), 107 (18), 104 (10), 92 (28), 77 (54).
3ꢀ(3ꢀMethoxyꢀ4,5ꢀmethylenedioxyphenyl)ꢀ5ꢀ(piperidinꢀ4ꢀ
yl)ꢀ1,2,4ꢀoxadiazole (19). Found (%): C, 59.43; H, 5.74; N, 14.01.

Polyalkoxy 3,5ꢀdiarylꢀ1,2,4ꢀoxadiazoles
Russ.Chem.Bull., Int.Ed., Vol. 59, No. 12, December, 2010
2275
3ꢀ(3,4ꢀDimethoxyphenyl)ꢀ5ꢀ(3ꢀmethoxyphenyl)ꢀ1,2,4ꢀoxadiꢀ
azole (32). Found (%): C, 65.48; H, 5.17; N, 9.11. C17H16N2O4.
Calculated (%): C, 65.38; H, 5.16; N, 8.97. MS, m/z (Irel (%)):
312 [M]+ (8), 179 (9), 136 (23), 135 (61), 134 (15), 107 (54),
103 (23), 92 (60), 77 (100).
4. D. V. Tsyganov, A. P. Yakubov, L. D. Konyushkin, S. I.
Firgang, V. V. Semenov, Izv. Akad. Nauk, Ser. Khim., 2007,
2375 [Russ. Chem. Bull., Int. Ed., 2007, 56, 2460].
5. N.ꢀH. Nam, Curr. Med. Chem., 2003, 10, 1697.
6. G. C. Tron, T. Pyrali, G. Sorba, F. Pagliai, S. Busacca, A. A.
Genazzani, J. Med. Chem., 2006, 49, 3033.
3ꢀ(3,4ꢀDimethoxyphenyl)ꢀ5ꢀ(3,4,5ꢀtrimethoxyphenyl)ꢀ1,2,4ꢀ
oxadiazole (33). Found (%): C, 61.36; H, 5.49; N, 7.64.
C19H20N2O6. Calculated (%): C, 61.28; H, 5.41; N, 7.52. MS,
m/z (Irel (%)): 372 [M]+ (13), 196 (95), 195 (100), 193 (6),
186 (36), 179 (37), 177 (68), 167 (18), 163 (16), 152 (56),
137 (38), 136 (40), 135 (23), 134 (37), 109 (36), 93 (45), 77 (88).
5ꢀ(2ꢀAminophenyl)ꢀ3ꢀ(4ꢀmethoxyphenyl)ꢀ1,2,4ꢀoxadiazole
(34). Found (%): C, 67.48; H, 5.04; N, 15.87. C15H13N3O2.
Calculated (%): C, 67.41; H, 4.90; N, 15.72. MS, m/z (Irel (%)):
267 [M]+ (6), 133 (5), 120 (100), 118 (12), 106 (7), 92 (29), 77 (8).
3ꢀ(Pyridinꢀ4ꢀyl)ꢀ5ꢀ(3,4,5ꢀtrimethoxyphenyl)ꢀ1,2,4ꢀoxadiꢀ
azole (35). Found (%): C, 61.44; H, 4.88; N, 13.36. C16H15N3O4.
Calculated (%): C, 61.34; H, 4.83; N, 13.41. MS, m/z (Irel (%)):
313 [M]+ (100), 298 (52), 270 (10), 195 (44), 152 (45), 151 (45),
137 (35), 136 (15), 135 (30), 120 (27), 109 (16), 92 (15), 78 (42).
Antimitotic activity of the compounds synthesized tested on the
sea urchin embryo.22 The study was carried out in the biological
laboratory of N. K. Kol´tsov Institute of Developmental Biology,
Russian Academy of Sciences on Cyprus. Adult sea urchin Paraꢀ
centrotus lividus L. (Echinidae, Echinodermata) were collected on
the coast and kept in the aerated seawater tank. The gametes were
obtained by intracoelomic injection of 0.5 M KCl (1—2 mL).
Thus obtained eggs were washed with seawater filtered through
a nylon filter and fertilized with a few drops of diluted sperm.
Embryos (600—2000 eggs per 1 mL) were incubated in the filꢀ
tered seawater at room temperature (18—23 °C) in sixꢀwell plates.
The stock solutions of the tested compounds were prepared in
DMSO or 95% EtOH, the maximum concentrations of compounds
depend on their solubility. Solubility of compounds in the solvents
and seawater were monitored by microscopic examination.
The embryos were treated with solutions of compounds 11—35
after 10—15 min of fertilization. An egg suspension (5 mL) was
placed to each well and the corresponding volume of solution of
the tested compound was added to achieve the desired final conꢀ
centration. The maximum solvent concentration of the solvent did
not exceed the highest tolerated concentration (1% for EtOH and
0.05% for DMSO). In the tests, twoꢀfold decreasing compound
concentrations were used. Antimitotic activity was evaluated by the
effective (threshold) concentration (EC) resulting cleavage alteraꢀ
tion. The embryos development were monitored with a Biolam
light microscope (LOMO, St.Petersburg, Russian Federation).
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Received September 17, 2010;
in revised form November 25, 2010