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D. Rotili et al. / European Journal of Medicinal Chemistry 89 (2015) 156e171
(0.27 mL, 2 mmol) in 2 mL of a mixture DMSO/acetone (1:1 v/v),
and the resulting solution was stirred at 50 ꢀC for 1 h. The pH was
maintained at around 7.0 by dropwise addition of 1 M KOH. The
excess of 6-mercaptohexan-1-ol was removed by the addition of 3-
bromopyruvic acid (250.4 mg, 1.5 mmol), and the obtained solid
was filtered off, washed on the filter with distilled H2O, and then
dried over anhydrous Na2SO4. The desired product was a light
(10 mL), and the resulting mixture was stirred at room temperature
for 3 h. After filtration of the morpholinium salt, the resulting so-
lution was treated with O-(tetrahydro-2H-pyran-2-yl)hydroxyl-
amine (493 mg, 4.21 mmol) for 2 h at room temperature. The
removal of the solvent in vacuum provided a white solid, that was
dissolved in a mixture of H2O (10 mL) and CHCl3 (10 mL). The
aqueous phase was basified with 2 N KOH and extracted with CHCl3
(4 ꢃ 10 mL). The combined organic phases were dried over Na2SO4,
filtered and evaporated under reduced pressure to provide a white
solid that was treated with a 1 M solution of sodium methoxide in
dry methanol (10 mL). After stirring at room temperature for 4 h,
the reaction mixture was acidified with 5% (w/w) citric acid, and
afterward the aqueous phase was extracted with CHCl3 (4 ꢃ 15 mL).
The organic phase was dried over Na2SO4, filtered and evaporated
under reduced pressure providing a crude solid, which was tritu-
rated with petroleum ether, thus obtaining the desired 4-
mercapto-N-((tetrahydro-2H-pyran-2-yl)oxy)benzamide 45. 1H
yellow solid. 1H NMR (DMSO)
d 1.2e1.5 (m, 6H, SCH2CH2CH2CH2),
1.6 (t, 2H, CH2CH2OH), 3.2 (t, 2H, SCH2), 3.4 (t, 2H, CH2OH), 7.4 (d,
1H, CH benzoxadiazole ring), 7.7 (d, 1H, CH benzoxadiazole ring).
MS-ESI, m/z: 333 [M þ H]þ.
4.1.5. Synthesis of 6-((7-nitrobenzo[c][1,2,5]oxadiazol-4-yl)amino)
hexan-1-ol (40)
Pyridine (0.54 mL, 6.8 mmol) and 6-aminohexan-1-ol
(588.3 mg, 5.0 mmol) were added to a solution of 4-chloro-7-
nitrobenzo[c][1,2,5]oxadiazole (575 mg, 2.89 mmol) in 42 mL of a
mixture EtOH:H2O (0.3:1). The reaction mixture was stirred at
room temperature for 6 h, and then the aqueous phase was
extracted with CHCl3 (5 ꢃ 20 mL). The combined organic phases
were dried over Na2SO4, filtered and evaporated under reduced
pressure to provide a crude solid that was finally purified by col-
umn chromatography on SiO2 gel eluting with a mixture CHCl3/
NMR (CDCl3)
d 1.63e1.69 (m, 4H, CH2 tetrahydropyran ring),
1.87e1.89 (m, 2H, CH2 tetrahydropyran ring), 3.63e3.70 (m, 2H, CH2
tetrahydropyran ring), 4.15e4.19 (s br, 1H, SH), 5.37e5.39 (t, 1H,
OCHO), 7.49e7.51 (d, 2H, CH benzene ring), 7.63 (d, 2H, CH benzene
ring). MS-ESI, m/z: 254 [M þ H]þ.
MeOH (100:1). 1H NMR (DMSO)
d
1.35e1.70 (m, 8H,
4.1.8. Synthesis of ethyl 4-(mercaptomethyl)benzoate (51)
NCH2CH2CH2CH2CH2CH2O), 3.38e3.40 (t, 2H, NCH2CH2), 3.46 (m,
2H, CH2CH2OH), 4.33e4.35 (t, 1H, CH2OH), 6.41 (d, 1H, CH ben-
zoxadiazole ring), 8.51 (d, 1H, CH benzoxadiazole ring), 9.55 (s, 1H,
NH). MS-ESI, m/z: 281 [M þ H]þ.
Anhydrous K2CO3 (681.4 mg, 4.93 mmol) and potassium ethyl
xanthogenate (790 mg, 4.93 mmol) were added to a solution of
ethyl 4-(bromomethyl)benzoate (0.4 g, 1.64 mmol) in dry CH3CN
(8.5 mL), and the resulting mixture was stirred at room tempera-
ture for 20 h. Then the reaction was quenched with H2O (10 mL)
and extracted with AcOEt (6 ꢃ 10 mL). The combined organic
phases were washed with brine (10 mL), dried over Na2SO4, filtered
and evaporated under reduced pressure to obtain a crude product
that was purified by column chromatography on SiO2 gel eluting
with a mixture AcOEt/n-hexane (1:20) to give the ethyl 4-
(((ethoxycarbonothioyl)thio)methyl)benzoate as an oily interme-
diate. EtONa (1.48 mL of a 1 M solution in EtOH) was added to a
solution of ethyl 4-(((ethoxycarbonothioyl)thio)methyl)benzoate
(0.2 g, 0.74 mmol) in dry EtOH (3 mL), and the resulting mixture
was left under stirring at room temperature for 6 h. At the end of
the reaction the solvent was evaporated under vacuum, the crude
product taken into H2O (5 mL), made acidic with 5% (w/w) citric
acid, and then extracted with AcOEt (4 ꢃ 10 mL). The combined
organic phases were dried over anhydrous Na2SO4, filtered and
evaporated under vacuum to give a crude that was purified by
column chromatography on SiO2 gel eluting with a mixture AcOEt/
n-hexane (1:20) to give the desired ethyl 4-(mercaptomethyl)
4.1.6. Synthesis of N-hydroxy-4-mercaptobenzamide (42)
Ethyl chloroformate (0.65 mL, 6.8 mmol) was added at 0e5 ꢀC to
a solution of 4-mercaptobenzoic acid (500 mg, 3.24 mmol) and N-
methylmorpholine (0.75 mL, 6.8 mmol) in dimethoxyethane
(10 mL), and the resulting mixture was stirred at room temperature
for 3 h. After filtration of the morpholinium salt, the resulting so-
lution was treated with O-(2-methoxy-2-propyl)hydroxylamine
(1.08 mL,14.58 mmol) for 2 h at room temperature. The solvent was
removed in vacuo, and the crude solid was dissolved in a mixture of
H2O (10 mL) and CHCl3 (10 mL), the aqueous phase was basified
with 2 N KOH and extracted with CHCl3 (4 ꢃ 10 mL). The combined
organic phases were dried over Na2SO4, filtered and evaporated
under reduced pressure to provide a solid that was treated with a
1 M solution of sodium methoxide in dry methanol (10 mL). After
stirring at room temperature for 5 h, the reaction mixture was
acidified with 5% (w/w) citric acid and the aqueous phase was
extracted with CHCl3 (4 ꢃ 15 mL). The organic phase was dried over
Na2SO4, filtered and evaporated under reduced pressure providing
the O-protected hydroxamate intermediate 4-mercapto-N-((2-
methoxypropan-2-yl)oxy)benzamide (49). The O-protected
hydroxamate 49 was finally dissolved in MeOH (5 mL) and treated
with Amberlyst 15 ion exchange resin (0.65 g) for 2 h at room
temperature. Afterward, the reaction mixture was filtered, and the
filtrate was concentrated in a vacuum to give the crude N-hydroxy-
4-mercaptobenzamide that was purified by trituration with diethyl
ether and used in the next step without further purification. 1H
benzoate 51. 1H NMR (CDCl3)
d 1.41 (t, 3H, COOCH2CH3), 1.80 (t, 1H,
SH), 3.79 (d, 2H, CH2SH), 4.39 (q, 2H, COOCH2CH3), 7.41 (d, 2H, CH
benzene ring), 8.02 (d, 2H, CH benzene ring). MS-ESI, m/z: 197
[M þ H]þ.
4.1.9. Synthesis of 4-(mercaptomethyl)benzoic acid (52)
2 N KOH solution (2 mL) was added to a solution of ethyl 4-
(((ethoxycarbonothioyl)thio)methyl)benzoate
(250
mg,
NMR (DMSO)
d
5.72 (s, 1H, SH), 7.34e7.36 (d, 2H, CH benzene ring),
0.93 mmol) in 4 mL of an EtOH:H2O mixture (1:1), and the reaction
was stirred at room temperature for 7 h. After the end of the re-
action, the EtOH was evaporated at reduced pressure and the
resulting water was acidified with 5% (w/w) citric acid to give a
solid in suspension which was filtered off to provide the crude 4-
(mercaptomethyl)benzoic acid 52, that was finally purified by col-
umn chromatography on SiO2 gel eluting with a mixture AcOEt/n-
7.61e7.63 (d, 2H, CH benzene ring), 8.95 (s, 1H, NH), 11.12 (s, 1H,
OH). MS-ESI, m/z: 170 [M þ H]þ.
4.1.7. General procedure for the synthesis of 4-mercapto-N-
substituted benzamides (41 and 43e50). Example: 4-mercapto-N-
((tetrahydro-2H-pyran-2-yl)oxy)benzamide (45)
Ethyl chloroformate (0.65 mL, 6.8 mmol) was added at 0e5 ꢀC to
a solution of 4-mercaptobenzoic acid (500 mg, 3.24 mmol) and N-
methylmorpholine (0.75 mL, 6.8 mmol) in dimethoxyethane
hexane (2:1). 1H NMR (DMSO)
d 2.96 (t,1H, SH), 3.79 (d, 2H, CH2SH),
7.46 (d, 2H, CH benzene ring), 7.88 (d, 2H, CH benzene ring), 12.84
(s, 1H, COOH). MS-ESI, m/z: 169 [M þ H]þ.