Bortolini et al.
orange color was obtained. After the mixture was allowed to
stand for 5 min, several drops of isopropyl alcohol were added
to destroy the oxidant in excess. The reaction mixture was
filtrated over Celite, concentrated under reduced pressure,
diluted with 20 mL of water and extracted with ethyl acetate.
The organic layer was dried over anhydrous Na2SO4 and the
solvent was removed in a vacuum to give the expected carbonyl
derivative in nearly quantitative yield. 3a : white crystal; mp
138-140 °C (lit.5 mp 140 °C); 1H NMR16,17 δ 0.69 (s, 3H), 0.92
(d, J ) 5.2 Hz, 3H), 1.01 (s, 3H), 3.62 (s, 3H). 6a : white crystal;
mp 154-155 °C (lit.18 mp 153-154 °C); 1H NMR16,17 δ 0.69 (s,
3H), 0.92 (d, J ) 5.7 Hz, 3H), 1.27 (s, 3H), 3.65 (s, 3H). 6b:
ethyl acetate (30 mL) in the presence of succinic anidride (6
g, 60 mmol), triethylamine (1.27 mL, 12.7 mmol), and DMAP
(0.15 g, 1.27 mmol). The reaction mixture, monitored by TLC
(ethyl acetate/cyclohexane/HAc 50/50/1), was refluxed for 40
or 10 h, respectively, depending on the bile acid precursor.
After completion, 20 mL of ethyl acetate and 15 mL of water
were added to the mixture, and the organic layer was washed
with acid water (15 mL, pH ca. 1), dried on anydrous Na2SO4,
and evaporated to obtain the bis-hemisuccinate. The bis-hemi-
succinates were used for the subsequent reaction without fur-
ther purification. The identity of both intermediates could be
1
validated by H NMR: (R-isomer) δ 0.65 (s, 3H), 0.95 (s, 3H),
white crystal; mp 184-186 °C (lit.5 mp 187 °C); 1H NMR16,17
δ
0.97 (d, J ) 6.5 Hz, 3H), 4.60 (m, 1H), 4.92 (m, 1H); (â-isomer)
(CD3OD) δ 0.70 (s, 3H), 0.92 (d, J ) 6.5 Hz, 3H), 0.98 (s, 3H),
4.65 (m, 1H), 4.78 (m, 1H). The C(3) hydrolysis of both products
was obtained by dissolving the bis-hemisuccinates in MeOH/
NaOH 5% (1:1 v/v), following the reaction by TLC (ethyl ace-
tate/cyclohexane/HAc 50:50:1). After 1 h, the solution was acid-
ified with dilute H2SO4. Addition of 50 mL of H2O caused the
precipitation of the 3-hydroxy derivative that was recovered
by filtration. The 7-hemisuccinates were used for the subse-
quent reaction without further purification. The identity of
0.87 (d, J ) 6 Hz, 3H), 1.05 (s, 3H), 1.10 (s, 3H), 3.60 (s, 3H).
1
7: white crystal; mp 237-240 °C (lit.19 mp 238-240 °C); H
NMR16,17 δ 0.85 (d, J ) 5.2 H, 3H), 1.06 (s, 3H), 1.40 (s, 3H),
3.61 (s, 3H).
7-Keto- (4a ), 12-k eto- (5a ), a n d 7,12-d ik eto-5â-ch ola n -
24-oic a cid (6c) were prepared starting from the 3-keto-7-
hydroxy 3b, 3-keto-12-hydroxy 3f, and 3-keto-7,12-dihydroxy
derivative 3g, respectively. Four grams of 85% KOH and 5
mL of 80% hydrazinium hydroxide were added to a solution
containing 10 mmol of 3b, 3f, or 3g, 35 mL of methylcellosolve
(ethylene glycol monomethyl ether), and 3 mL of H2O. The
solution was heated at 110 °C for 4 h and then distilled to
obtain a temperature of the reaction mixture of about 135 °C.
This temperature was maintained for an additional 8 h. The
reaction mixture was cooled to 20 °C, diluted with 12 mL of
water, and acidified to pH 2 using H2SO4 (40%).20 The product
was collected by filtration and submitted to the oxidation step
using J ones’ protocol described for compound 3a ; see the
previous paragraph. 4a : overall yield 70%; white crystal; mp
149-151 °C (methanol/water) (lit.21 mp 150-151 °C); 1H
NMR16,17 δ 0.65 (s, 3H), 0.90 (d, J ) 5.2 Hz, 3H), 1.18 (s, 3H),
3.65 (s, 3H). 5a : overall yield 75%; white crystal; mp 185-
187 °C (ethyl acetate/n-hexane) (lit.22 mp 187-189 °C); 1H
NMR16,17 δ 0.82 (d, J ) 6 Hz, 3H), 0.99 (s, 6H), 3.62 (s, 3H).
6c: overall yield 65%; white crystal; mp 119-121 °C (ethyl
1
both intermediates could be validated by H NMR: (R-isomer)
(CDCl3/CD3OD 1:1 v/v) δ 0.60 (s, 3H), 0.88 (s, 3H), 0.90 (d, J
) 6.5 Hz), 3.42 (m, 1H), 4.85 (m, 1H); (â-isomer) δ 0.65 (s, 3H),
0.92 (d, J ) 6.5 Hz, 3H), 0.95 (s, 3H), 3.58 (m, 1H), 4.75 (m,
1H). The further J ones’ oxidation of the C(3) hydroxy function
was carried out as described for compound 3a , see previous
paragraph. 3d : final yield 90%; mp 85-87 °C; 1H NMR δ 0.70
(s, 3H), 0.95 (d, J ) 6.5 Hz, 3H), 1.05 (s, 3H), 5.05 (m, 1H). 3e:
final yield 92%; mp 92-95 °C; 1H NMR δ 0.73 (s, 3H), 0.94 (d,
J ) 6.5 Hz, 3H), 1.08 (s, 3H), 4.85 (m, 1H).
3-Keto-7â-h yd r oxy-5â-ch ola n -24-oic Acid (3c). A 4.9 g
(10 mmol) portion of the 3-keto-7â-hemisuccinyloxy 3e was
dissolved in 30 mL of NaOH (20%) and the reaction mixture
refluxed for 1 h. The solution was cooled and acidified with
dilute H2SO4, and the precipitate was collected by filtration.
The crude product was purified by crystallization (ethyl ace-
tate): yield 95%; mp73 °C (ethyl acetate);26 1H NMR (CD3OD)26
δ 0.75 (s, 3H), 0.96 (d, J ) 6.5 Hz, 3H), 1.05 (s, 3H), 3.50 (m,
1H).
3-Keto-7r-h yd r oxy- (3b),27 3-k eto-12r-h yd r oxy- (3f),27
3-k eto-7,12r-d ih yd r oxy- (3g),27 3-k eto-7,12r-d ih yd r oxy-
a ceta te- (3h ),27 3,12-d ik eto-7r-h yd r oxy-5â-ch ola n -24-oic
a cid (6e),27 3,7r-d ih yd r oxy-12-k eto-5â-ch ola n -24-oic a cid
(5c),28 3r-h yd r oxy-7-k eto-5â-ch ola n -24-oic a cid (4b),29 3,-
12r-d ih yd r oxy-7-k eto-5â-ch ola n -24-oic a cid (4c),29 3r-
h yd r oxy-12-k eto-5â-ch ola n -24-oic a cid (5b),30 a n d 3r-
h yd r oxy-7,12-d ik et o-5â-ch ola n -24-oic a cid (6f)30 were
prepared and characterized according to recently described
procedures.27-30
1
acetate/n-hexane) (lit.23 mp 120-122 °C); H NMR16,17 δ 0.83
(d, J ) 6 Hz, 3H), 1.02 (s, 3H), 1.27 (s, 3H) 3.62 (s, 3H).
3,7-Dik eto-12r-h yd r oxy-5â-ch ola n -24-oic a cid (6d ) was
obtained from the 3-keto-7,12-dihydroxy derivative 3g by
oxidation with NBS.24 A 2.03 g (5 mmol) portion of 3g was
dissolved, upon warming, in 75 mL of tap water containing
2.5 g of sodium bicarbonate. The solution was cooled to 25 °C,
treated with 1.11 g (6.25 mmol) of N-bromosuccinimide, and
shaken occasionally until complete dissolution of the reagent.
The yellow solution was allowed to stand at 25 °C for about
24 h, heated on the steam bath for 1 h, cooled to 0 °C, and
acidified with 10% aqueous HCl. The keto acid 6d separate
as a white solid that was collected by filtration and crystallized
from ethyl acetate: yield 1.5 g (75%); mp 168-170 °C (lit.25
mp 168-170 °C); 1H NMR (CD3OD)25 δ 0.75 (s, 3H), 1.00 (d, J
) 6 Hz, 3H), 1.30 (s, 3H), 4.05 (m, 1H).
Ack n ow led gm en t. We acknowledge financial sup-
port by MURST-Cofin 2000 and the University of
Ferrara.
3-Keto-7r-su ccin yloxy-5â-ch ola n -24-oic a cid (3d ) and
3-k et o-7â-su ccin yloxy-5â-ch ola n -24-oic a cid (3e) were
prepared from the commercially available chenodeoxycholic or
ursodeoxycholic acid, respectively. A 5 g (12.7 mmol) portion
of chenodeoxycholic or ursodeoxycholic acid was suspended in
Su p p or tin g In for m a tion Ava ila ble: MM2 total and
relative energies. This material is available free of charge via
the Internet at http://pubs.acs.org.
J O020146B
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