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o[4,4]non-1-en-4-one 3 (1 g, 5.15 mmol) and 2-(2-chloro-
ethyl)-isoindole-1,3-dione 4d (1.2 g, 6.18 mmol).
4.3.8. Synthesis of 40-(2-butyl-4-oxo-1,3-diazaspiro[4,4]
non-1-en-3-ylmethyl)-biphe-nyl-2-carboxylic acid methyl
ester (5h). The product 5h was obtained from 2-butyl-1,3-
diaza-spiro[4,4]non-1-en-4-one 3 (1 g, 5.15mmol) and 40-
bromomethyl-biphenyl-2-carboxylic acid methyl ester 4h
(1.9 g, 6.18 mmol).
IR mmax (KBr): 3099, 2932, 1734, 1578, 1495, 1465,
1365 cmꢀ1
.
1H NMR (CDCl3 400 MHz) d: 0.88–0.92 (t, 3H, –CH3),
1.2–1.4 (m, 6H, –CH2–CH2–CH2–), 1.51–1.81 (m, 8H,
–cyclopentane), 3.65–3.85 (t, 4H, –CH2–CH2–), 7.72–
8.35 (m, 4H, Ar-H).
IR mmax (KBr): 3105, 2937, 1740, 1576, 1467, 1465,
1354 cmꢀ1
.
1H NMR (CDCl3 400 MHz) d: 0.88–0.92 (t, 3H, –CH3),
1.2–1.4 (m, 6H, –CH2–CH2–CH2–), 1.51–1.81(m, 8H,
–cyclopentane), 3.92–3.93 (s, 3H, –OCH3), 4.34 (s, 2H,
–CH2-), 7.11–8.00 (m, 8H, Ar-H).
Anal. Calcd for C21H25N3O3: C, 68.64; H, 6.86; N,
11.44. Found: C, 68.63; H, 6.87; N, 11.45%.
4.3.5. Synthesis of 2-butyl-3-(6-methylbenzo[1,3]dioxole-5-
ylmethyl)-1,3-diazaspiro [4,4]non-1-en-4-one (5e). The
product 5e was obtained from 2-butyl-1,3-diaza-spiro[4,4]-
non-1-en-4-one 3 (1 g, 5.15 mmol) and 5-chloromethyl-6-
methyl-benzo[1,3]dioxole 4e (1.4 g, 6.18 mmol).
Anal. Calcd for C26H30N2O3: C, 74.61; H, 7.22; N, 6.69.
Found: C, 74.61; H, 7.23; N, 6.69%.
4.3.9. Synthesis of 40-(2-butyl-4-oxo-1,3-diazaspiro[4,4]-
non-1-en-3-ylmethyl)-biphenyl-2-carboxylic acid tert-
butyl ester (5i). The product 5i was obtained from 2-bu-
tyl-1,3-diaza-spiro[4,4]non-1-en-4-one 3 (1 g, 5.15 mmol)
and 40-bromomethyl-biphenyl-2-carboxylic acid tert-bu-
tyl ester 4i (2.1 g, 6.17 mmol).
IR mmax (KBr): 3095, 2930, 1735, 1574, 1480, 1465,
1369 cmꢀ1
.
1H NMR (CDCl3 400 MHz) d: 0.88–0.92 (t, 3H, –CH3),
1.2–1.4 (m, 6H, –CH2–CH2–CH2–), 1.51-1.81(m, 8H,
–cyclopentane), 4.34 (s, 2H, –CH2–), 5.8 (s, 2H, –CH2-),
6.46 (s, 3H, Ar-H), 5.8 (s, 3H, –CH3).
IR mmax (KBr): 3017, 2945, 1740, 1578, 1493 cmꢀ1
.
1H NMR (CDCl3 400 MHz) d: 0.88–0.92 (t, 3H, –CH3),
0.97–1.00 (s, 9H, –C(CH3)3), 1.2–1.4 (m, 6H, –CH2–
CH2–CH2–), 1.51–1.81(m, 8H, –cyclopentane), 4.34 (s,
2H, –CH2–), 7.12–7.89 (m, 8H, Ar-H).
Anal. Calcd for C20H26N2O3: C, 70.15; H, 7.65; N, 8.18.
Found: C, 70.13; H, 7.65; N, 8.19%.
4.3.6. Synthesis of 2-butyl-3-[2-(4-chlorophenyl)-2-oxoeth-
yl]-1,3-diazaspiro[4,4]non-1-en-4-one (5f). The product 5f
was obtained from 2-butyl-1,3-diaza-spiro[4,4]non-1-en-
4-one 3 (1 g, 5.15 mmol) and 2-Bromo-1-(4-chloro-
phenyl)ethanone 4f (1.4 g, 6.18 mmol).
Anal. Calcd for C29H36N2O3: C, 75.62; H, 7.88; N, 6.08.
Found: C, 75.61; H, 7.87; N, 6.09%.
4.3.10. Synthesis of 3-(4-bromobenzyl)-2-butyl-1,3-diaza-
spiro[4,4]non-1-en-4-one (5j). The product 5j was ob-
tained from 2-butyl-1,3-diaza-spiro[4,4]non-1-en-4-one
3 (1 g, 5.15 mmol) and 1-bromo-4-bromomethyl-ben-
zene 4j (1.5 g, 6.18 mmol).
IR mmax (KBr): 3115, 2940, 1743, 1572, 1488, 1455, 1345 cmꢀ1.
1H NMR (CDCl3 400 MHz) d: 0.88–0.92 (t, 3H, –CH3),
1.2–1.4 (m, 6H, –CH2–CH2–CH2–), 1.51–1.81 (m, 8H,
–cyclopentane), 4.34 (s, 2H, –CH2–), 7.65 (dd, 2H, Ar-
H), 7.88 (dd, 2H, Ar-H).
IR mmax (KBr): 3098, 2941, 1740, 1579, 1478, 1462,
1345 cmꢀ1
.
Anal. Calcd for C19H23ClN2O2: C, 65.79; H, 6.68; N,
8.08. Found: C, 65.78; H, 6.69; N, 8.10%.
1H NMR (CDCl3 400 MHz) d: 0.88–0.92 (t, 3H, –CH3),
1.2–1.4 (m, 6H, –CH2–CH2–CH2–), 1.51–1.81 (m, 8H,
–cyclopentane), 4.34 (s, 2H, –CH2–), 7.25 (dd, 2H, Ar-
H), 7.46 (dd, 2H, Ar-H).
4.3.7. Synthesis of 2-butyl-3-[20-(1-trityl-tetrazol-5-
yl)biphenyl-4-ylmethyl]-1,3-diaza spiro[4,4]non-1-en-4-
one (5g). The product 5g was obtained from 2-butyl-1,
3-diaza-spiro[4,4]non-1-en-4-one 3 (1 g, 5.15 mmol) and
5-(40-bromomethyl-biphenyl-2-yl)-1-trityl-1H-tetrazole 4g
(3.4 g, 6.18 mmol).
Anal. Calcd for C18H23BrN2O: C, 59.51; H, 6.38; N,
7.71. Found: C, 59.51; H, 6.39; N, 7.70%.
5. Biology: in vitro acetylcholinesterase assay
IR mmax (KBr): 3114, 2945, 1740, 1570, 1486, 1451, 1340 cmꢀ1.
The cholinesterase assay method of Ellman et al.17 was
used to determine the in vitro cholinesterase activity.
The activity was measured by increase in absorbance
at 412 nm due to the yellow color produced from the
reaction of acetylthiocholine iodide with the dith-
iobisnitrobenzoate (DTNB) ion. AChE was obtained
from the brain of Wistar rats by homogenizing under
a Teflon blender for 10 min in 0.1 M KH2PO4 buffer,
1H NMR (CDCl3 400 MHz) d: 0.88–0.92 (t, 3H, –CH3),
1.2–1.4 (m, 6H, –CH2–CH2–CH2–), 1.51–1.81(m, 8H,
–cyclopentane), 4.34 (s, 2H, –CH2–), 6.79–8.05 (m,
24H, Ar-H).
Anal. Calcd for C44H42N6O: C, 78.78; H, 6.31; N, 12.53.
Found: C, 78.79; H, 6.32, N, 12.52%.