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Med Chem Res (2011) 20:1102–1110
Mass spectrum: 442 (4; (M ? i-Bu-1)?); 386 (100;
(M ? 1)?); 368 (17; (M-OH)?); 342 (19); 269 (21; (M-
succinoyloxy ? 1)?).
3-Hydroxy-17-glutaroyloxy-16,17-secoestra-1,3,5(10)-
triene-16-nitrile (4) and 3-Hydroxy-17-succinoyloxy-
16,17-secoestra-1,3,5(10)-triene-16-nitrile (5)
Anal. Calcd. for C22H27NO5: C, 68.37; H, 7.30; N, 3.62.
Found: C, 68.66; H, 7.32; N, 3.56.
To the solution of the corresponding benzyl ether 2 or 3
(2 mmol) in methylene chloride–methanol mixture (2:1,
10 ml), 10% Pd/C (0.38 g) was added. The suspensions were
stirred at room temperature for 1.5 and 18 h under an
atmosphere of hydrogen. After the removal of the catalyst,
the solvent was evaporated to dryness and the products were
purified by crystallization from toluene, i.e., methylene
chloride. Analytically pure 3-hydroxy-17-glutaroyloxy-16,17-
secoestra-1,3,5(10)-trien-16-nitrile (4) and 3-hydroxy-17-
succinoyloxy-16,17-secoestra-1,3,5(10)-trien-16-nitrile (5)
were obtained in yields 50.33 and 60.46% (mp 113–115 and
129–131°C), respectively.
Biological tests
All experiments were approved by the Local Ethical
Committee of the University of Novi Sad and were per-
formed and conducted in accordance with the principles
and procedures of the NIH Guide for Care and Use of
Laboratory Animals.
Uterotrophic and antiuterotrophic assay
Compound 4 IR spectrum: 3450, 2930, 2220, 1750, 1720,
1630, 1510, 1190, 950, 890, and 810.
1H-NMR spectrum (DMSO): 0.86 (s, 3H, CH3, H-18);
The estrogenic and antiestrogenic effects of compounds 2,
3, 4, and 5 were tested on experimental animals using the
uterotrophic and antiuterotrophic methods (Emmens,
1950).
2.35 (m, 6H, 3CH2 from glutaroyloxy group); 3.81 (d, 1H,
H-17a, Jgem = 11.59 Hz); 4.03 (d, 1H, H-17b, Jgem
=
11.59 Hz); 6.45 (d, 1H, H-4, J4,2 = 2.26 Hz); 6.53 (dd, 1H,
H-2, J2,1 = 8.45 Hz, J2,4 = 2.23 Hz); 7.08 (d, 1H, H-1,
J1,2 = 8.50 Hz); 9.06 (s, 1H, OH, C-3); 12.16 (s, 1H,
COOH, C-17).
13C-NMR-spectrum (DMSO-d6): 15.05 (CH2, C-15);
15.59 (CH3, C-18); 19.96 (C-30 from the side chain); 25.64;
26.62; 29.44; 32.67 (C-20 from the side chain); 32.73 (C-40
from the side chain); 37.29; 40.16; 40.50; 41.31; 42.20;
70.51 (C-17); 112.95; 114.61; 120.57 (C:N); 126.27;
129.84; 136.88; 155.09 (C-3); 172.44 (COOR); 174.00
(COOH).
Immature Wistar strain female rats (21–23 days old,
raised under controlled environmental conditions—tem-
perature 22 2°C and 14 h light/10 h dark, with food and
water ad libitum) were randomly divided into groups of six
to eight animals each. The animals were treated by sub-
cutaneous injection once a day for 3 days with 0.1 ml of a
solution of the test compound in olive oil, either solely or
in combination with estradiol benzoate (EB). The control
group obtained the vehicle only. The total administered
amount of tested compounds was 5 mg/kg of body weight
(b.w.), except in case of tamoxifen, used as a comparator,
and compound 5, where doses were 5 mg/kg b.w., i.e.,
25 mg/kg b.w., whereas the EB dose was 30 lg/kg b.w.
The animals were sacrificed on the fourth day. The uteri
were removed, dissected free of adhering fat and blotted
dry after expulsion of uterine fluid and the wet weights
were recorded.
Mass spectrum: 400 (100; (M ? 1)?); 382 (81; (M-
OH)?); 2.68 (18; (M-glutaroyloxy ? 1)?).
Anal. Calcd. for C23H29NO5: C, 69.15; H, 7.32; N, 3.50.
Found: C, 69.43; H, 7.35; N, 3.37.
Compound 5 IR spectrum: 3450, 2930, 2220, 1730, 1710,
1610, 1510, 1230, 1160, 1040, 1000, 950,750, and 700.
1H-NMR spectrum (DMSO-d6): 0.86 (s, 3H, CH3, H-18);
2.50 (2dd, 2H, H-15, Jgem = 26.03 Hz, J15a,14 = 4.62 Hz);
2.68 (s, 4H, 2CH2 from succinyloxy group); 3.81 (d, 1H,
H-17a, Jgem = 11.59 Hz); 4.13 (d, 1H, H-17b); 6.45 (d, 1H,
H-4, J4,2 = 2.45 Hz); 6.53 (dd, 1H, H-2, J2,1 = 8.38 Hz,
J2,4 = 2.50 Hz); 7.07 (d, 1H, H-1, J1,2 = 8.53 Hz); 9.06 (s,
1H, OH, C-3); 12.24 (s,1H, COOH).
13C-NMR-spectrum (DMSO): 15.03 (CH2, C-15); 15.60
(CH3, C-18); 25.67; 26.60; 28.77 (CH2 from succinyloxy
group); 28.95 (CH2 from succinyloxy group); 29.43; 35.27;
37.39; 40.51; 41.00; 42.09; 70.52 (C-17); 112.95; 114.62;
120.49 (C:N); 126.25; 129.90; 136.89; 155.08 (C-3);
171.88 (COOR); 173.44 (COOH).
The differences of uteri weights of treated and control
animals served for the calculation of the agonistic and
antagonistic effects (Wakeling et al., 1984). Percentage of
agonistic and antagonistic activity in immature rat uterine
weight assays were calculated from the ratio of values
recorded in the treated and control animals, thus:
% agonism = ðC ꢁ AÞ ꢂ 100=ðB ꢁ AÞ
% antagonism = ðB ꢁ DÞ ꢂ 100=ðB ꢁ AÞ
In these equations A, B, C, and D are uterine wet
weights, corrected for differences in body weights (mg/
100 g body weight) for vehicle alone, EB, test compound
alone, or test compound plus EB, respectively.
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