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P.S. Dangate et al. / Steroids 76 (2011) 1397–1399
0.77(s, 3H, H-18), 1.06 (d, J = 6.2 Hz, 3H, H-21), 1.26 (s, 3H, H-19),
2.60 (dd, J = 12 Hz, 1H, H-8b), 3.02 (dd, J = 13, 6.6 Hz, 1H, H-6b),
3.58 (br, 1H, H-3b), 4.04 (s, 1H, H-12b).
lated computationally. Fukui indices have been used for prediction
of reactivities [7]. The Fukui function is defined as the differential
change in electron density due to an infinitesimal change in the
number of electrons. They were calculated by using software
Jaguar [8]. Methodology employed for calculation is as follows.
Conformation search was performed using Macro Model [9], to
determine lower energy conformers. Conformer with lowest
energy was selected for further calculations and was subjected to
DFT optimization using basis set B3LYP/6-31Gꢃꢃ with single point
solvation from Poisson–Boltzmann quantum theory (t-BuOH sol-
2.2.1.2.
0.29 g (7.5%); Mp: 188 °C [lit.12 189 °C]; IR (KBr): 3414.2(–OH),
2957.5, 2916(CAH), 1711(C@O) cmꢁ1 1H NMR (300 MHz, CDCl3):
3a-Hydroxy-7,12-dioxo-5b-cholane-24-oic acid 3. Yield:
;
d 0.74 (d, J = 6.2 Hz, 3H, H-21), 1.69 (s, 3H, H-19), 1.28 (s, 3H, H-
18), 3.57 (br, 1H, H-3b).
vent, e = 12.4, rprobe = 2.6 Å) and values are given in Table 1. Atomic
Fukui index f_NN HOMO indicates.
2.2.2. 3
a
,7b,12
a
-Trihydroxy-5b-cholane-24-oic acid 4
,12 -dihydroxy-7-keto-
To a solution of 2.50 g (6.1 mmol) of 3
a
a
Preferred sites of electrophilic attack and susceptibility when its
value is large [10]. Cholic acid 1 shows highest f_NN HOMO for C-7
followed by C-12 and C-3 hydroxy sites. Therefore order of reactiv-
ity of hydroxy groups would be: C-7 > C-12 > C-3. Similarly for 7b
epimer 4 highest f_NN HOMO was observed for C-12 followed by
C-7 and C-3 hydroxy sites. Therefore order of reactivity of hydroxy
groups would be: C-12 ꢄ C-7 > C-3. The oxidation of cholic acid 1
and its 7b epimer 4 using reagents such as chromic acid, bromine
in aqueous alkali was found to show very poor regioselectivity
indicating that they are not mild enough to distinguish these fine
differences in reactivity pattern of hydroxyl groups [11]. However
oxidation with NBS shows high regioselectivity and follows this
trend with cholic acid 1 but no data is available for oxidation of
7b epimer 4. IBX is a mild oxidant and it is expected to distinguish
this reactivity pattern and show high regioselectivity. Oxidation
experiments were carried out on cholic acid 1 under different con-
ditions using t-BuOH as solvent and with 1.5 equivalent of IBX un-
der reflux, as less than 1.5 equivalents were found to be insufficient
5b-cholane-24-oic acid 2 in 20 ml of anhydrous n-propanol was
added 5.09 g (220.0 mmol) of sodium metal. Reaction mixture
was refluxed and progress of reaction was monitored by TLC. Reac-
tion was completed in 3 h. The reaction mixture was cooled to
room temperature and gradually diluted with 20 ml of water, acid-
ified with dilute hydrochloric acid, and extracted with (2 ꢀ 30 ml)
of ethyl acetate. Organic layer was dried over anhydrous sodium
sulphate and concentrated under reduced pressure to get residue
which was further purified by column chromatography on silica
gel using methanol–chloroform (5:95). Yield: 2 g (80%); Mp:
144 °C [lit.12 145 °C]; IR (KBr): 1704 (C@0), 3448 (OH), 1042,
1010 cmꢁ1 1H NMR (300 MHz, CDCl3): d 0.69 (s, 3H, H-18), 0.91
;
(s, 3H, H-19), 3.51 (br m, 2H, H-3b and H-7a), 3.91(m, 1H, H-12b).
2.2.3. Oxidation of 7b epimer 4
7b Epimer 4 (4 g, 9.7 mmol) was oxidised following the general
procedure.
to take the reaction to completion. Both 3
a,12a-dihydroxy-7-
2.2.3.1.
3.5 g (90%); Mp: 137 °C [lit.12 136 °C]; IR (KBr): 1740, 1670
(C@O) cmꢁ1 1H NMR (300 MHz, CDCl3): d 1.08 (s, 6H, H-18 and
H-19), 3.58 (br m, 2H, H-3b and H-7 ).
3a,7b-Dihydroxy-12-oxo-5b-cholane-24-oic acid 5. Yield:
oxo-5b-cholan-24-oic acid 2 and 3 -hydroxy-7, 12-dioxo-5b-cho-
a
lan-24-oic acid 3 were isolated in the yields of 90% and 7.5%,
respectively (Scheme 1). Thus showing very high regioselectivity
towards 7a-hydroxy group.
;
a
Reduction of compound 2 was performed following the litera-
ture procedure with sodium in anhydrous n-propanol to obtain
80% yield of 7b epimer 4 and 10% of cholic acid 1 after isolation
by column chromatography. When 7b epimer 4 was subjected
oxidation under the same condition almost exclusively 3a,7b-dihy-
droxy-12-oxo-5b-cholane-24-oic acid 5 was obtained 90%, with no
2.2.2.2.
reduction of 3
3
a
,7b-Dihydroxy-5b-cholane-24-oic acid 6. Wolf–Kishner
a,7b,12-oxo-5b-cholane-24-oic acid 5 was carried
out by using reported literature procedure and isolated after
chromatography:
To a suspension of 0.5 g (1.23 mmol) 3
5b-cholane-24-oic acid and 99% hydrazine hydrate 0.21 g
a,7b-dihydroxy-12-oxo-
5
detectable amount of diketone 3 (Scheme 2). Again showing very
(4.31 mmol) with 6 ml of triethylene glycol, was added potassium
hydroxide 0.24 g (4.18 mmol). The reaction mixture was heated to
110 °C for 3 h and distilled until the temperature of the reaction
was raised to 135 °C and maintained this temperature till the reac-
tion gets completed. Then reaction mixture was cooled to room
temperature and acidified with dilute hydrochloric acid to get white
residue after filtration. Which was further purified by column chro-
matography on silica gel using methanol–chloroform (3:97) Yield:
0.39 g (82%); Mp 196 °C [lit.12 198 °C]; IR (KBr) 3518, 3552 (OAH),
high regioselectivity but towards 12a hydroxy group. These results
are consistent with the reactivity as predicted by atomic Fukui
indices. Lastly dihydroxy-ketone 5 was subjected to Wolf–Kishner
reduction by known process [12]e and isolated ursodeoxycholic
acid 6 in 82% yields.
Many procedures are reported for synthesis of ursodeoxycholic
acid 6 from cholic acid 1 [12] and all the chemical synthesis routes
require at least one protection–deprotection step to overcome the
problem of chemoselectivity with various oxidising agents
2937(CAH), 1716, 1654 (C@O) cmꢁ1 1H NMR (CDCl3): d 0.76 (s,
.
employed, however, recently
a protection-group-free chemo-
3H, H-18), 1.00 (d, J = 6.2 Hz, 3H, H-21), 1.00 (s, 3H, H-19), 3.55
enzymatic route is disclosed by Giovannini et al. [12e] in which
they obtained 70% overall yield of ursodeoxycholic acid 6. With
(br, 2H, H-3b and H-7
a
). ½a 2D5
ꢂ
= +59 (c = 0.2 ethanol).
Table 1
3. Results and discussion
Calculated values of atomic Fukui indices for cholic acid 1 and 7b epimer 4.
Cholic acid 1 and its 7b epimer 4 have three hydroxyl groups
present at different positions and study of their oxidation would
give some insights into the chemistry of IBX and outcome may also
have some practical utility. Before investigations of oxidation some
theoretical calculation were performed to understand the prepon-
derance of hydroxyl group towards oxidation. To visualise reactiv-
ity pattern of hydroxyl groups and scope for regioselective
oxidation in substrates for cholic acid 1 and 7b epimer 4, DFT
(density functional theory) based atomic Fukui indices were calcu-
Substrate
Atoms (hydroxy sites)
fꢀk
f_NN HOMO for reactive
sites (t-BuOH)
Cholic acid 1
7b Epimer 4
O3
O7
O12
0.0048
0.2093
0.0728
O3
O7
O12
0.0021
0.0168
0.3264