102
B. Kaboudin et al. / Bioorganic Chemistry 37 (2009) 101–105
Acetylcholinesterease from electric eel and butyrylcholinesterase
(BChE) from horse serum, actetylcholine chloride, and gelatin were
from Sigma and were used without further treatment. Acetylcho-
linesterease and butyrylcholinesterase activities were measured
using Ellman method [20].
2.5.2. S-pyridin-2-ylmethyl-O,O0-diethyl phosphorothioate (2b)
1H NMR (CDCl3 – 250 MHz): d = 1.17 (6H, t, J = 7.0 Hz), 3.92–4.07
(6H, m), 7.10 (1H, m), 7.30 (1H, m), 7.55 (1H, m), 8.45 (1H, m); 31P
NMR (CDCl3–H3PO4): d = 26.61; 13C NMR (CDCl3 – 62.9 MHz):
d = 15.8 (d, Jpc = 7.4 Hz), 36.4 (d, Jpc = 3.7 Hz), 63.5 (d, Jpc = 5.8 Hz),
122.3 (d, Jpc = 8.6 Hz), 123.9, 136.6 (d, Jpc = 14.4 Hz), 149.5 (d,
Jpc = 5.8 Hz), 156.9 (d, Jpc = 4.8 Hz). Anal. Calcd for C10H16NO3PS: C,
45.97; H, 6.18; N, 5.36. Found: C, 45.78; H, 6.20; N, 5.23.
2.2. Synthesis of ammonium O,O0-diethylthiophosphate using of a
mixture of ammonium hydrogen carbonate, sulfur, and
diethylphosphite [17]
2.5.3. S-quinolin-2-ylmethyl-O,O0-diethyl phosphorothioate (2c)
1H NMR (CDCl3 – 250 MHz): d = 1.26 (6H, t, J = 7.0 Hz), 3.98–4.22
(4H, m), 4.33 (2H, d, J = 14.5 Hz), 7.48–7.56 (2H, m), 7.66–7.80 (2H,
m), 8.02–8.14 (2H, m); 31P NMR (CDCl3–H3PO4): d = 26.68; 13C NMR
(CDCl3 – 26.9 MHz): d = 15.9 (d, Jpc = 7.4 Hz), 37.0 (d, Jpc = 3.8 Hz),
63.7 (d, Jpc = 5.9 Hz), 121.1, 126.6, 127.1, 127.5, 128.9, 129.8, 137.0,
147.6, 157.3 (d, Jpc = 5.1 Hz). Anal. Calcd for C14H18NO3PS: C, 54.00;
H, 5.83; N, 4.50. Found: C, 53.86; H, 5.75; N, 4.38.
Ammonium hydrogen carbonate (0.1 mol, 7.9 g) was added to a
mixture of sulfur (0.1 mol, 3.2 g) and diethylphosphite (0.1 mol,
13.8 g) in EtOAc/Et2O (300 mL, 1:1). Reaction mixture was stirred
for 24 h at room temperature. Evaporation of the solvent under re-
duced pressure gave the pure ammonium O,O0-diethylthiophos-
phate as solid in a quantitative yield that could be recrystallized
in ethanol.
2.3. Synthesis of ammonium O,O0-diethyldithiophosphate using of a
mixture of ammonium hydrogen carbonate, phosphorus pentoxide,
and ethanol [18]
2.5.4. S-[6-(diethoxy-thiophosphorylsulfanylmethyl)-pyridin-2-
ylmethyl]-O,O0-diethyl phosphorothioate (2d)
1H NMR (CDCl3 – 250 MHz): d = 1.30 (12H, t, J = 7.1 Hz), 4.05–
4.17 (12H, m), 7.32 (2H, d, J = 7.7 Hz), 7.64 (1H, t, J = 7.7 Hz); 31P
NMR (CDCl3–H3PO4): d = 27.18; 13C NMR (CDCl3 – 62.9 MHz):
d = 16.3 (d, Jpc = 7.3 Hz), 36.7 (d, Jpc = 1.9 Hz), 64.0 (d, Jpc = 6.0 Hz),
122.3, 137.9, 157.5 (d, Jpc = 5.2 Hz). Anal. Calcd for C15H27NO6P2S2:
C, 40.63; H, 6.14; N, 3.16. Found: C, 40.90; H, 6.33; N, 3.29.
Ammonium hydrogen carbonate (0.1 mol, 7.9 g) was added to a
mixture of phosphorus pentoxide (50 mmol, 11.1 g) in EtOH
(100 mL) at 60 °C. Reaction mixture was stirred for 1 h at 60 °C.
Evaporation of the solvent under reduced pressure gave the pure
ammonium O,O0-diethyldithiophosphate as solid in quantitative
yield that could be recrystallized in ethanol.
2.5.5. S-[2-(1,3-dioxo-1,3-dihidro-isoindol-2-yl)-ethyl]-O,O0-diethyl
phosphorothioate (2e)
2.4. General procedure for the synthesis of phosphorothioates (2) [17]
1H NMR (CDCl3 – 250 MHz): d = 1.21 (6H, t, J = 7.0 Hz), 3.03 (2H,
m), 3.85 (2H, m), 4.02 (4H, m), 7.59–7.72 (4H, m); 31P NMR
(CDCl3–H3PO4): d = 26.62; 13C NMR (CDCl3 – 62.9 MHz): d = 15.9
(d, Jpc = 7.2 Hz), 28.6 (d, Jpc = 4.0 Hz), 38.1 (d, Jpc = 4.3 Hz), 63.7 (d,
Jpc = 6.1 Hz), 123.2, 131.7, 134.0, 167.7. Anal. Calcd for C14H18NO5PS:
C, 48.97; H, 5.29; N, 4.08. Found: C, 48.81; H, 5.12; N, 4.19.
Ammonium O,O0-diethyl phosphorothioate (5 mmol, 0.935 g)
was dissolved in acetonitrile (10 mL). Alkyl halide (5 mmol) was
added to reaction mixture and stirred for 24 h at reflux. Evapora-
tion of solvent and washing with sodium hydrogen carbonate
(50 ꢀ 3 mL) gave crude product S-alkyl O,O0-diethyl phosphoro-
thioate. Chromatography through a plug of silica gel with EtOAc/
n-hexane (1:1) and evaporation of the solvent under reduced pres-
sure gave the pure products as oils in 34–80% yields. All products
gave satisfactory spectral data in accord with the assigned
structures.
2.5.6. S-pyridin-4-ylmethyl-O,O0-diethyl phosphorodithioate (3a)
1H NMR (CDCl3 – 250 MHz): d = 1.30 (6H, t, J = 7.1 Hz), 3.98–
4.19 (6H, m), 7.50 (2H, d, J = 5.7 Hz), 8.60 (2H, d, J = 5.7 Hz); 31P
NMR (CDCl3–H3PO4): d = 92.87; 13C NMR (CDCl3 – 62.9 MHz):
d = 15.7 (d, Jpc = 8.2 Hz), 36.5 (d, Jpc = 3.1 Hz), 64.4 (d, Jpc = 6.3 Hz),
124.9, 141.5, 147.2. Anal. Calcd for C10H16NO2PS2: C, 43.31; H,
5.82; N, 5.05. Found: C, 43.15; H, 5.68; N, 5.15.
2.5. General procedure for the synthesis of phosphorodithioates (3)
[18]
2.5.7. S-pyridin-2-ylmethyl-O,O0-diethyl phosphorodithioate (3b)
1H NMR (CDCl3 – 250 MHz): d = 1.30 (6H, t, J = 7.25 Hz), 3.95–
4.35 (6H, m), 7.30 (1H, m), 7.43 (1H, m), 7.69 (1H, m), 8.57 (1H,
Solvent-free reaction method, which is operationally simple,
was used to synthesize phosphorodithioates. Ammonium O,O0-
diethyl phosphorodithioate (5 mmol, 0.935 g) was added to alu-
mina (Al2O3, acidic, 2.5 g). Alkyl halide (5 mmol) was added to
reaction mixture and grinded for 1–2 h. The mixture was
washed with chloroform (100 mL) and neutralized with sodium
hydrogen carbonate (50 ꢀ 3 mL). Solvent was evaporated to give
crude product alkyl O,O0-diethyl phosphorodithioate. Chroma-
m); 31P NMR (CDCl3–H3PO4): d = 93.44; 13C NMR (CDCl3
–
62.9 MHz): d = 15.7 (d, Jpc = 8.2 Hz), 38.9 (d, Jpc = 3.8 Hz), 64.1 (d,
Jpc = 6.3 Hz), 122.5, 123.6, 137.2, 149.0, 156. 7 (d, Jpc = 5.0 Hz). Anal.
Calcd for C10H16NO2PS2: C, 43.31; H, 5.82; N, 5.05. Found: C, 43.25;
H, 5.62; N, 5.10.
tography through
a
plug of silica gel with EtOAc/n-hexane
2.5.8. S-quinolin-2-ylmethyl-O,O0-diethyl phosphorodithioate (3c)
1H NMR (CDCl3 – 250 MHz): d = 1.28 (6H, t, J = 7.2 Hz), 4.00–
4.20 (4H, m), 4.45 (2H, d, J = 16.0 Hz), 7.54–7.62 (2H, m), 7.72–
7.85 (2H, m), 8.13–8.21 (2H, m); 31P NMR (CDCl3–H3PO4):
d = 93.32; 13C NMR (CDCl3 – TMS – 62.9 MHz): d = 15.8 (d,
Jpc = 8.2 Hz), 38.1, 64.3 (d, Jpc = 6.3 Hz), 121.5, 129.4, 130.9, 134.2,
144.3, 156.9 (d, Jpc = 5.03 Hz). Anal. Calcd for C14H18NO2PS2: C,
51.37; H, 5.55; N, 4.28. Found: C, 51.20; H, 5.73; N, 4.35.
(1:9) and evaporation of the solvent under reduced pressure
gave the pure products as oils in 70–85% yields. All products
gave satisfactory spectral data in accord with the assigned
structures.
2.5.1. S-pyridin-4-ylmethyl-O,O0-diethyl phosphorothioate (2a)
1H NMR (CDCl3 – 250 MHz): d = 1.30 (6H, t, J = 7.1 Hz), 3.99–
4.13 (6H, m), 7.34 (2H, d, J = 5.9 Hz), 8.59 (2H, d, J = 5.9 Hz); 31P
NMR (CDCl3–H3PO4): d = 26.21; 13C NMR (CDCl3 – 62.9 MHz):
d = 16.3 (d, Jpc = 7.29 Hz), 33.4 (d, Jpc = 3.8 Hz), 64.2 (d, Jpc = 6.0 Hz),
124.2, 147.3 (d, Jpc = 4.3 Hz), 150.3. Anal. Calcd for C10H16NO3PS: C,
45.97; H, 6.18; N, 5.36. Found: C, 45.83; H, 6.10; N, 5.25.
2.5.9. S-[6-(diethoxy-thiophosphorylsulfanylmethyl)-pyridin-2-
ylmethyl]-O,O0-diethyl phosphorodithioate (3d)
1H NMR (CDCl3 – 250 MHz): d = 1.30 (12H, t, J = 7.1 Hz), 4.00–
4.30 (12H, m), 7.29 (2H, d, J = 7.7 Hz), 7.61 (1H, t, J = 7.72 Hz), 31P