Synthesis of various secosteroidal macrocycles by ring-closing metathesis

Article abstract of DOI:10.1016/j.steroids.2013.03.004

We set out to describe an efficient and versatile method for preparing secosteroidal macrocycles from cholic acid, via an oxidative ring-expansion/ring-opening sequence and a ring-closing metathesis reaction as the key steps. The characteristic ¹H and ¹³C NMR spectroscopic features of the synthesized compounds are reported.

Full text of DOI:10.1016/j.steroids.2013.03.004

Steroids 78 (2013) 651–661
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Steroids
journal homepage: www.elsevier.com/locate/steroids
Synthesis of various secosteroidal macrocycles by ring-closing metathesis
⇑
Malika Ibrahim-Ouali , Eugénie Romero
Aix Marseille Université, CNRS Institut des Sciences Moléculaires de Marseille UMR 7313, 13397 Marseille, France
a r t i c l e i n f o
a b s t r a c t
Article history:
We set out to describe an efficient and versatile method for preparing secosteroidal macrocycles from
cholic acid, via an oxidative ring-expansion/ring-opening sequence and a ring-closing metathesis reaction
as the key steps. The characteristic 1H and 13C NMR spectroscopic features of the synthesized compounds
are reported.
Received 22 November 2012
Received in revised form 5 March 2013
Accepted 23 March 2013
Available online 10 April 2013
Ó 2013 Elsevier Ltd. All rights reserved.
Keywords:
Macrocycles
Baeyer–Villiger oxidation
Ring-closing metathesis (RCM)
1. Introduction
efficient as protease inhibitors [20,21], G protein- coupled recep-
tors (GPCRs) [22–24], and protein–protein interaction inhibitors
Steroids represent an important class of natural products due to
their high ability to penetrate cells and bind to nuclear and mem-
brane receptors. The steroid system, selected by the evolutionary
process to perform some of the most fundamental biological func-
tions, has not only inspired biochemists and endocrinologists, but
has also become the basis of many important discoveries in organic
chemistry.
[25,26]. Natural products are one of the sources of bioactive mac-
rocycles such as erythromycin, rapamycin, vancomycin, cyclospor-
ine, and epothilone [27]. Natural products exhibit enormous
structural diversity [28]. However, there are several problems
associated with their use in screening experiments including diffi-
culties with purification, bioactive component identification, struc-
tural assignment, chemical modification, and analog synthesis
[29]. These difficulties have motivated medicinal chemistry
researchers to develop strategies such as diversity oriented synthe-
sis (DOS) of macrocyclic compounds with known bioactive do-
mains such as peptide motifs [30–32].
Consequently, many bioactive macrocycles with diversity and
complexity are now readily available in chemist’s showcases
[33,34].
So, it would be interesting to combine secosteroidal skeleton
with varied types of macrocycles in order to obtain a new class
of biological molecules. We report herein an efficient synthesis of
secosteroidal macrocycles from cholic acid 1 (Scheme 1). We show
that we can modify the size of the macrocycle and then lead to no-
vel interesting structures. We report here the full details of these
syntheses.
Secosteroids have attracted considerable interest because of the
broad range of biological activities of many naturally occurring
representatives, such as Vitamins D [1], anolides [2], and marine
steroids [3,4]. Apart from Vitamins D with their innumerable bio-
logical effects [5], secosteroids with cytotoxic [6–8], antihistamine
[9], and anticancer [10] activity should be mentioned as com-
pounds with great potential for drug development. The activity
of seco analogs of normal steroidal hormones in humans and high-
er animals is a matter of scientific interest as well. Some of these
compounds were prepared synthetically and showed hormonal
or antihormonal activity [11–16]. It is evident that the higher con-
formational flexibility of seco steroids in comparison with normal
steroids may result in novel, pharmaceutically useful compounds.
Otherwise, macrocyclic compounds have unique physicochem-
ical and topological properties that allow them to exhibit unusual
biological properties [17,18]. Macrocycles have the ability to
exhibit high target binding affinity, selectivity, and improved oral
bioavailability. Additionally, macrocyclization is an efficient way
of increasing cellular penetration via the decrease in polarity of
peptidic drug leads [19]. Macrocycles have been proven to be
2. Experimental section
All reactions were run under argon in oven-dried glassware. 1H
and 13C NMR spectra are recorded at 200 or 400 and 50 and
100 MHz respectively, in CDCl3 solutions. Chemical shift (d) are re-
ported in ppm with tetramethylsilane as internal standard. NMR
signals assignments were made with the aid of a combination of
2D homonuclear (1H–1H) and heteronuclear (1H–13C) correlation
⇑
Corresponding author. Tel.: +33 491288416; fax: +33 491983865.
E-mail address: malika.ibrahim@univ-amu.fr (M. Ibrahim-Ouali).
0039-128X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.steroids.2013.03.004

652
M. Ibrahim-Ouali, E. Romero / Steroids 78 (2013) 651–661
O
n
O
n
O
O
HO
O
HO
OH
H
H
HO
OH
H
HO
OH
H
HO
H
1
H
O
OH
O
n
HO
n
O
O
Scheme 1. Retrosynthetic pathway.
techniques, which included 1H–1H COSY, 1H–1H Nuclear Overhaus-
er Effect Spectroscopy (NOESY), Heteronuclear Single Quantum
Correlation (HSQC) and Heteronuclear Multiple Bond Correlation
(HMBC). IR spectra were recorded on a Perkin-Elmer 1600 spectro-
photometer. Flash chromatography was performed on silica gel
(Merk 60 F254) and TLC on silica gel. Dichloromethane was dis-
tilled from P2O5 and tetrahydrofuran (THF) over sodium/
benzophenone.
silica gel (diethyl ether/petroleum ether: 7/3), to afford 0.66 g
(70% yield) of 12-oxo steroid 4 as colorless needles. mp = 71–
72 °C; IR (neat) 3236, 1736, 1511 cmÀ1; 1H NMR (300 MHz, CDCl3):
0.85 (d, J = 6.5, 3H, H-21), 1.02 (s, 3H, H-18), 1.03 (s, 3H, H-19), 3.26
(m, 1H, H-3b), 3.28 (s, 3H, OCH3), 3.32 (s, 3H, OCH3), 3.38 (m, 1H,
H-7b), 3.66 (s, 3H, OCH3); 13C NMR (75 MHz, CDCl3): 18.4, 21.3,
21.4, 22.0, 23.7, 26.4, 27.3, 30.3, 30.9, 31.1, 31.2, 31.3, 34.4, 35.4,
35.6, 37.8, 40.4, 46.2, 51.4, 53.0, 57.1, 57.5, 58.4, 73.4, 78.4,
174.7, 214.0. HRMS (EI) for C27H44O5 [M+] calcd 448.3189 found
448.3192.
Compounds 14 and 15 were prepared according to the previ-
ously described procedure [35]. The nomenclature used for the ste-
roids is not the nomenclature used by Chemical Abstracts [36,37].
2.3. 3a,7a-Dimethoxy-13-oxa-C-homo-cholan-12-one (5)
2.1. Methyl 3a,7a-dimethoxy-12a-hydroxy-5b-cholan-24-oate (3)
To a solution of ketone 4 (0.5 g, 1.11 mmol) in dry dichloro-
methane (30 mL) containing p-toluenesulfonic acid (167 mg,
1.11 mmol) m-chloroperbenzoic acid (12 mg) was added. The solu-
tion was stirred for 24 h at room temperature. The solution was
then diluted with water and extracted with dichloromethane
(3 Ã 15 mL). The solution was washed successively with a 5% Na2S2-
O3 solution (10 mL), saturated brine (10 mL), and water (20 mL)
and was dried over anhydrous magnesium sulfate. The oily prod-
uct, obtained by evaporation of the solvent, was purified by flash
chromatography on silica gel (CH2Cl2/MeOH: 95/5) to afford
0.51 g of pure lactone 5 (98%) as an oil. 1H NMR (300 MHz, CDCl3):
0.86 (s, 3H, H-19), 1.05 (d, J = 6.4, 3H, H-21), 1.34 (s, 3H, H-18), 3.25
(m, 1H, H-3b), 3.28 (s, 3H, OCH3), 3.31 (s, 3H, OCH3), 3.32 (s, 3H,
OCH3), 3.35 (m, 1H, H-7b); 13C NMR (75 MHz, CDCl3): 14.5, 15.3,
17.7, 22.3, 24.2, 25.2, 26.1, 26.5, 27.5, 31.6, 32.8, 34.6, 35.2, 35.9,
36.3, 41.2, 42.5, 50.1, 55.7, 57.4, 58.5, 73.3, 76.9, 80.2, 86.9,
174.8, 176.5. HRMS (EI) for C27H44O6 [M+] calcd 464.3138 found
464.3143.
To a stirred suspension of NaH (0.62 g, 26 mmol) in THF (10 mL)
at 0 °C under argon was added a solution of triol 2 (5 g, 11.8 mmol)
in 5 mL of THF. The reaction mixture was stirred for 15 min, and
then iodomethane (367 lL, 5.9 mmol) was added dropwise. After
24 h at room temperature, the reaction was diluted with 10 mL
of Et2O and quenched by the slow addition of 10 mL of H2O. The
combined organic extracts (3 Â 10 mL) were washed with 30 mL
of brine, dried over anhydrous MgSO4, and concentrated in vacuo.
The crude product was purified by flash chromatography on silica
gel (Et2O: 100%) to give 7 (5 g, 94%) as a white solid. mp = 80 °C; 1H
NMR (300 MHz, CDCl3): 0.59 (s, 3H, H-18), 0.83 (d, J = 6.5, 3H, H-
21), 0.85 (s, 3H, H-19), 2.94 (m, 1H, H-3b), 3.14 (s, 3H, OCH3),
3.18 (s, 3H, OCH3), 3.21 (m, 1H, H-7b), 3.26 (s, 3H, OCH3), 3.29
(m, 1H, H-12b); 13C NMR (75 MHz, CDCl3): 12.6, 17.5, 22.1, 23.0,
23.3, 26.4, 26.6, 27.5, 27.9, 28.1, 30.9, 31.2, 34.5, 35.2, 35.6, 39.8,
42.1, 42.8, 46.3, 46.4, 55.5, 55.8, 56.0, 77.3, 81.0, 82.1, 180.3. HRMS
(EI) for C27H46O5 [M+] calcd 450.3345 found 450.3348.
2.2. Methyl 3
a
,7a
-dimethoxy-12-oxo-5b-cholan-24-oate (4)
2.4. 3a,7a-Dimethoxy-11,12-seco-5b-cholan-12,13a,24-triol (6)
Alcohol 3 (1 g, 2.22 mmol) was mixed in a mortar with pyridi-
nium chlorochromate (PCC) (0.57 g, 2.66 mmol). The mixture was
transferred to a pressure-resistant tube (pyrex) and irradiated with
MW at 170 °C for 5 min. After cooling to room temperature, the
reaction mixture was filtered through a Celite pad and the filtrate
and washings (CH2Cl2, 3 Ã 10 mL) were combined and evaporated
under reduced pressure. The residue was chromatographed on
A solution of methyl ester 5 (1 g, 2.15 mmol) in dry ether
(10 mL) was added in one portion to a suspension of LiAlH4
(0.25 g, 6.46 mmol) in dry ether (20 mL) at room temperature.
After 1 h the reaction was quenched with H2O (15 mL) and EtOAc
(15 mL). The aqueous layer was acidified to pH 2 with diluted
HCl, and layers were separated. The aqueous layer was further ex-
tracted with EtOAc (3 Â 50 mL), and the combined organic layers

M. Ibrahim-Ouali, E. Romero / Steroids 78 (2013) 651–661
653
were dried over anhydrous MgSO4 and evaporated to dryness. The
2.8. General procedure for deprotection with trimethylsilyl iodide
crude product was purified by flash chromatography on silica gel
(CH2Cl2/MeOH: 9/1) to afford 0.83 g of pure triol 6 (88%) as an
To a solution of macrocycle 8 (50 mg, 0.10 mmol) in chloroform
(20 mL) was added trimethylsilyl iodide (0.1 mL). The solution was
left overnight at room temperature. Methanol (5 mL) was then
added to decompose any excess trimethylsilyl iodide. The solution
was extracted with diethyl ether (3 Â 10 mL), washed with water
(10 mL) and saturated brine (10 mL), dried over anhydrous magne-
sium sulfate. Evaporation of the solvent gave a crude product,
which was purified by chromatography on silica gel (CH2Cl2/
MeOH: 95/5), to give triol 9.
oil. IR (neat) 3280 (OH), 1220, 1070 cmÀ1 1H NMR (300 MHz,
;
CDCl3): 1.06 (d, J = 6.5, 3H, H-21), 1.16 (s, 3H, H-19), 1.50 (s, 3H,
H-18), 2.78 (m, 1H, H-3b), 2.79 (m, 1H, H-7b), 3.24 (s, 3H, OCH3),
3.25 (s, 3H, OCH3), 3.37 (m, 2H, H-24), 3.53 (m, 2H, H-12b); 13C
NMR (75 MHz, CDCl3): 19.1, 19.5, 19.7, 23.5, 24.4, 24.6, 29.9,
30.8, 31.6, 32.0, 33.8, 34.1, 35.7, 38.4, 40.8, 46.2, 46.3, 51.0, 57.1,
57.4, 59.3, 61.6, 74.9, 82.4, 82.9, 89.2. HRMS (EI) for C26H48O5
[M+] calcd 440.3502 found 440.35.09.
The above method was also used for deprotection of macrocy-
cles 13a-b.
2.5. 3a,7a-Dimethoxy-12,24-diallyloxy-11,12-seco-5b-cholan-13a-ol
(7)
2.9. Macrocycle (9)
Yield 47 mg (92%). Oil. IR (neat) 3429, 2950, 1606, 1089 cmÀ1
;
To a stirred suspension of KH (0.32 g, 6.81 mmol) in DMF
(10 mL) at 0 °C under argon was added a solution of triol 6 (1 g,
2.27 mmol) in 5 mL of DMF. The reaction mixture was stirred for
15 min, and then allyl bromide (6.43 mL, 4.99 mmol) was added
dropwise. After 48 h at room temperature, the reaction was diluted
with 10 mL of Et2O and quenched by the slow addition of 10 mL of
H2O. The solution was extracted with diethyl ether (3 Ã 10 mL) and
the combined organic extracts were washed with 30 mL of brine,
dried over anhydrous MgSO4, and concentrated in vacuo. The crude
product was purified by flash chromatography on silica gel (Et2O:
100%) to give 7 (0.68 g, 58%) as an oil. IR (neat) 2951, 1175,
1H NMR (300 MHz, CDCl3) (major isomer): 0.88 (d, J = 6.4 Hz, 3H,
H-21), 0.89 (s, 3H, H-19), 1.12 (s, 3H, H-18), 3.38 (m, 2H, H-12b
and H-24), 3.66 (m, 1H, H-7), 3.88 (m, 1H, H-3b), 4.16 (m, 4H,
OCH2–CH@), 5.82 (td, J = 6.3 and 15.6 Hz, 1H, CH@), 5.98 (td,
J = 6.2 Hz and 15.6 Hz, 1H, CH@); 13C NMR (75 MHz, CDCl3): 18.8,
19.7, 23.3, 23.4, 24.2, 25.6, 29.5, 29.9, 31.5, 32.2, 33.1, 35.7, 36.4,
38.2, 41.4, 45.6, 50.8, 53.8, 59.0, 63.4, 67.2, 69.0, 70.6, 71.7, 72.2,
74.6, 119.4, 125.8. HRMS (EI) for C28H48O5 [M+] calcd 464.3502
found 464.3507.
We give the characterization data for compounds 10-15 later in
the Experimental Section.
1096 cmÀ1 1H NMR (300 MHz, CDCl3): 1.05 (d, J = 6.4, 3H, H-21),
;
1.16 (s, 3H, H-19), 1.43 (s, 3H, H-18), 2.79 (m, 1H, H-3b), 2.82
(m, 1H, H-7b), 3.24 (s, 3H, OCH3), 3.32 (m, 2H, H-24), 3.25 (s, 3H,
OCH3), 3.38 (m, 1H, H-12b), 4.16 (m, 2H, OCH2–CH@), 4.48 (m,
2H, OCH2–CH@), 5.23 (m, 4H, CH2@), 5.78 (m, 2H, CH@); 13C
NMR (75 MHz, CDCl3): 17.9, 18.8, 24.2, 24.9, 26.1, 26.4, 28.1,
28.2, 29.3, 29.9, 31.9, 32.1, 32.1, 32.8, 33.2, 33.8, 35.7, 41.1, 45.6,
51.2, 57.0, 57.1, 71.2, 72.8, 74.5, 74.9, 82.2, 82.9, 116.3, 116.7,
131.9, 132.8. HRMS (EI) for C32H56O5 [M+] calcd 520.4128 found
520.4133.
2.10. Methyl 3a,12a-dihydroxy-7-oxo-8-oxa-B-homo-cholan-24-oate
(16)
To a solution of ketone 15 (3 g, 7.14 mmol) in dry dichlorometh-
ane (30 mL) containing p-toluenesulfonic acid (1.11 g, 6.43 mmol)
m-chloroperbenzoic acid (1.84 g, 10.7 mmol) was added. The solu-
tion was stirred for 24 h at room temperature. The solution was
then diluted with water (20 mL) and extracted with dichlorometh-
ane (3 Ã 15 mL). The solution was washed successively with a 5%
Na2S2O3 solution, saturated brine, and water and was dried over
anhydrous magnesium sulfate. The oily product, obtained by evap-
oration of the solvent, was purified by flash chromatography on sil-
ica gel (CH2Cl2/MeOH: 95/5) to afford 2.55 g of pure lactone 5 (82%)
as an oil. 1H NMR (300 MHz, CDCl3): 0.66 (s, 3H, H-18), 0.89 (d,
J = 6.4, 3H, H-21), 0.96 (s, 3H, H-19), 3.58 (s, 3H, OCH3), 3.89 (m,
1H, H-3b), 4.18 (m, 1H, H-12b); 13C NMR (75 MHz, CDCl3): 12.5,
14.1, 17.3, 20.7, 23.1, 24.9, 27.3, 28.9, 30.8, 31.1, 34.7, 34.9, 35.8,
36.7, 41.6, 45.0, 47.1, 51.5, 53.6, 60.4, 70.4, 71.2, 80.4, 174.6,
2.6. General procedure for ring-closing metathesis reaction
A solution of the secosteroidal ester 7 in dry toluene (c = 0.15–
0.20 mM) was stirred for 5 min in 80 °C and then Grubbs II catalyst
(5 mol%) was added to the mixture. Reaction was stirred for 15 min
under 80 °C and its progress was monitored by TLC. The total
amount of catalyst (20 mol%) was added portionwise to the reac-
tion mixture with stirring within 12 of 24 h. After cooling to room
temperature, the reaction was quenched with ethyl vinyl ether and
the solvent was evaporated under reduced pressure. The crude
product was purified by chromatography on silica gel (CH2Cl2/
MeOH: 9/1), to give macrocycle 8 as an oil.
175.1. HRMS (EI) for
436.2829.
C
25H40O6 [M+] calcd 436.2825 found
2.11. Methyl 3a,12a-di(tert-butyldimethylsilyloxy)-7-oxo-8-oxa-B-
homo-cholan-24-oate (17)
The above method was also used for the RCM reactions of secos-
teroids 10a-b, 19 and 22a-b.
To a solution of lactone 16 (2 g, 4.59 mmol) in dry DMF (30 mL)
containing imidazole (1.40 g, 20.6 mmol) tert-butyldimethylsilyl
chloride (3.11 g, 20.6 mmol) was added at 0 °C. The solution was
stirred for 12 h at room temperature. The solution was then diluted
with water (10 mL) and extracted with dichloromethane
(3 Ã 15 mL). The combined organic extracts were washed with
30 mL of brine, dried over anhydrous MgSO4, and concentrated in
vacuo. The crude product was purified by flash chromatography
on silica gel (Et2O: 100%) to give 17 (2.77 g, 91%) as an oil. 1H
NMR (300 MHz, CDCl3): 0.59 (12H, s, Si(CH3)2), 0.77 (9H, s,
C(CH3)3), 0.82 (9H, s, C(CH3)3), 0.89 (3H, d, J = 6.4 Hz, H-21), 0.94
(3H, s, H-19), 2.92 (1H, m, H-8b), 3.57 (3H, s, OCH3), 3.85 (1H, m,
H-3b), 4.12 (1H, m, H-12b); 13C NMR (75 MHz, CDCl3): À4.6 (2C),
2.7. Macrocycle (8)
Yield 50 mg (65%). Oil. IR (neat) 2981, 1370, 1048 cmÀ1 1H
;
NMR (300 MHz, CDCl3) (major isomer): 1.04 (d, J = 6.4, 3H, H-21),
1.18 (s, 3H, H-19), 1.38 (s, 3H, H-18), 2.79 (m, 1H, H-3b), 2.80
(m, 1H, H-7b), 3.24 (s, 3H, OCH3), 3.26 (s, 3H, OCH3), 3.62 (m, 4H,
H-24 and H-12), 4.16 (m, 4H, OCH2–CH@), 5.92 (td, 1H, J = 6.4
and 15.2, CH@), 6.04 (td, 1H, J = 6.4 and 15.2, CH@); 13C NMR
(75 MHz, CDCl3): 18.2, 18.7, 19.4, 23.8, 24.6, 24.9, 25.0, 25.4,
25.7, 29.8, 29.9, 32.8, 35.7, 35.9, 36.4, 37.8, 42.0, 45.6, 56.8, 57.1,
60.6, 70.8, 71.1, 71.4, 73.6, 74.2, 82.5, 82.7, 122.1, 126.8. HRMS
(EI) for C30H52O5 [M+] calcd 492.3815 found 492.3821.

654
M. Ibrahim-Ouali, E. Romero / Steroids 78 (2013) 651–661
À3.4 (2C), 12.7, 15.4, 17.5, 18.1, 18.2, 23.5, 24.6 (2C), 25.8, 26.0,
27.3, 28.7, 30.6, 30.8, 31.2, 34.8, 34.9, 35.2, 35.8, 36.1, 37.3, 42.0,
42.6, 45.5, 47.1, 47.2, 51.7, 65.9, 71.4, 71.7, 79.9, 174.5, 174.8.
HRMS (EI) for C37H68O6Si2 [M+] calcd 664.4554 found 664.4559.
72.7, 129.2, 142.9. HRMS (EI) for C40H76O5Si2 [M+] calcd 692.5231
found 692.5235.
2.15. General procedure for deprotection with TBAF
To a solution of macrocycle 20 (100 mg, 0.14 mmol) in anhy-
drous THF (20 mL) was added tetra-n-butylammonium fluoride
(0.35 mL, 1 M, 2.4 equiv). The solution was left overnight at room
temperature. The solution was extracted with diethyl ether,
washed with water and saturated brine, dried over anhydrous
magnesium sulfate. Evaporation of the solvent gave a crude prod-
uct, which was purified by chromatography on silica gel (CH2Cl2/
MeOH: 9/1), to give triol 21 (60 mg, 92%) as an oil.
2.12. 3
a,12a-di(tert-butyldimethylsilyloxy)-7,8-seco-5b-cholan-
7,8 ,24-triol (18)
a
A solution of diprotected lactone 17 (2 g, 3.01 mmol) in dry
ether (30 mL) was added in one portion to a suspension of LiAlH4
(0.46 g, 12.0 mmol) in dry ether (20 mL) at 0 °C. After 12 h at room
temperature, the reaction was quenched with H2O (10 mL) and
EtOAc (10 mL). The aqueous layer was acidified to pH 2 with di-
luted HCl, and layers were separated. The aqueous layer was fur-
ther extracted with EtOAc (3 Â 50 mL), and the combined organic
layers were dried over anhydrous MgSO4 and evaporated to dry-
ness. The crude product was purified by flash chromatography on
silica gel (CH2Cl2/MeOH: 9/1) to afford 1.74 g of pure triol 18
(90%) as an oil. 1H NMR (300 MHz, CDCl3): 0.00 (12H, s, Si(CH3)2),
0.62 (s, 3H, H-18), 0.82 (18H, s, C(CH3)3), 0.88 (3H, d, J = 6.4 Hz,
H-21), 0.94 (3H, s, H-19), 3.50 (m, 1H, H-8b), 3.56 (m, 2H, H-24),
3.72 (m, 2H, H-12b and H-3b), 3.82 (m, 4H, H-24 and H-7); 13C
NMR (75 MHz, CDCl3): À4.5 (2C), À3.4 (2C), 12.9, 15.3, 18.4, 19.4
(2C), 21.0, 23.9, 25.8 (2C), 26.4, 27.2, 28.9, 29.3, 30.1, 31.7, 31.8,
32.8, 33.4, 35.9, 36.1, 37.3, 37.6, 41.4, 44.3, 47.4, 47.9, 52.1, 63.2,
65.9, 72.5, 72.9, 73.6. HRMS (EI) for C36H72O5Si2 [M+] calcd
640.4918 found 640.4923.
The above method was also used for deprotection of macrocy-
cles 24a-b.
2.16. Macrocycle 21
IR (neat) 2965, 1340, 1059 cmÀ1 1H NMR (300 MHz, CDCl3)
;
(major isomer): 0.66 (s, 3H, H-18), 0.86 (d, J = 6.4 Hz, 3H, H-21),
0.88 (s, 3H, H-19), 3.52 (m, 1H, H-8b), 3.66 (m, 1H, H-3b), 3.88
(m, 1H, H-12b), 4.28 (m, 4H, OCH2–CH@), 4.82 (m, 4H, H-7 and
H-24), 5.95 (td, J = 6.3 and 15.4 Hz, 1H, CH@), 6.12 (td, J = 6.3 Hz
and 15.4 Hz, 1H, CH@);13C NMR (75 MHz, CDCl3): 13.8, 18.1,
23.7, 24.2, 24.6, 25.7, 29.5, 29.9, 31.9, 35.4, 35.7, 36.0, 36.1, 37.3,
41.4, 47.5, 51.2, 53.8, 59.0, 63.4, 67.2, 68.0, 71.7, 71.9, 72.9, 74.0,
119.8, 126.4. HRMS (EI) for C28H48O5 [M+] calcd 464.3502 found
464.3507.
2.13. 3
5b-cholan-8
a
,12
a
a
-di(tert-butyldimethylsilyloxy)-7,24-diallyloxy-7,8-seco-
-ol (19)
2.17. General procedure for esterification
A solution of diprotected pentahydroxysecocholane 6 or 18
(0.78 mmol), 3-butenoic acid (0.14 g, 1.63 mmol), N,N-dicyclohex-
ylcarbodiimide (0.14 g, 0.70 mmol) and 4-dimethyl-aminopyridine
(86 mg, 0.70 mmol) in dichloromethane (5 mL) was stirred at room
temperature until the reaction was completed (about 12 h). The
N,N-dicyclohexyl urea was filtered off and the filtrate was washed
with water, 5% acetic acid solution and again water, dried over
magnesium sulfate and the solvent was evaporated to afford 3-
butenoate 10a or 22a.
To a stirred suspension of KH (112 mg, 2.34 mmol) in DMF
(10 mL) at 0 °C under argon was added a solution of triol 18
(0.5 g, 0.78 mmol) in 5 mL of DMF. The reaction mixture was stir-
red for 15 min, and then allyl bromide (152 lL, 1.72 mmol) was
added dropwise. After 48 h at room temperature, the reaction
was diluted with 10 mL of Et2O and quenched by the slow addition
of 10 mL of H2O. The combined organic extracts (3 Ã 10 mL) were
washed with 30 mL of brine, dried over anhydrous MgSO4, and
concentrated in vacuo. The crude product was purified by flash
chromatography on silica gel (Et2O: 100%) to give 18 (315 mg,
56%) as an oil. 1H NMR (300 MHz, CDCl3): 0.02 (12H, s, Si(CH3)2),
0.65 (s, 3H, H-18), 0.83 (18H, s, C(CH3)3), 0.89 (3H, d, J = 6.4 Hz,
H-21), 0.94 (3H, s, H-19), 3.40 (m, 1H, H-3b), 3.46 (m, 1H, H-8b),
3.56 (m, 2H, H-24), 3.88 (m, 3H, H-12b and H-7), 4.20 (m, 2H,
OCH2–CH@), 4.62 (m, 2H, OCH2–CH@), 5.20 (m, 4H, CH2@), 5.87
(m, 2H, CH@); 13C NMR (75 MHz, CDCl3): À4.4 (2C), À3.5 (2C),
13.4, 17.9, 18.3 (2C), 24.6, 24.9, 26.1 (2C), 27.5, 29.6, 30.4, 31.0,
31.8, 31.9, 32.1, 33.5, 35.2, 36.0, 36.2, 37.4, 37.5, 38.2, 41.5, 42.6,
47.3, 47.6, 47.7, 63.5, 68.4, 70.9, 72.1, 72.7, 74.5, 79.1, 116.5,
The above method was also used for preparation of esters 10b-c
from 6 and 22b-c from 18.
2.18. Secosteroid 10a
Yield 350 mg (78%). Oil. IR (neat) 3280, 1220, 1070 cmÀ1 1H
;
NMR (300 MHz, CDCl3): 0.96 (s, 3H, H-18), 1.02 (d, J = 6.4 Hz, 3H,
H-21), 1.06 (s, 3H, H-19), 2.88 (m, 1H, H-7b), 2.92 (m, 1H, H-3b),
2.96 (m, 4H, CH2–C@O), 3.22 (s, 3H, OCH3), 3.24 (s, 3H, OCH3),
4.16 (m, 4H, H-12 and H-24), 5.22 (m, 4H, CH2@), 6.01 (m, 2H,
CH@); 13C NMR (75 MHz, CDCl3): 18.6, 20.8, 23.3, 24.3, 24.7,
26.8, 31.9, 32.2, 32.5, 32.8, 33.6, 35.6, 35.7, 37.9, 38.4, 41.0, 41.2,
45.3, 47.5, 51.4, 57.2, 57.6, 64.2, 64.6, 72.5, 75.4, 81.9, 82.6,
116.9, 117.6, 128.6, 129.1, 168.4, 169.0. HRMS (EI) for C34H56O7
[M+] calcd 576.4026 found 576.4031.
116.8, 134.7, 134.9. HRMS (EI) for
720.5544 found 720.5549.
C
42H80O5Si2 [M+] calcd
2.14. Macrocycle (20)
Yield 95 mg (90%). Oil. IR (neat) 3455, 2976, 1606 cmÀ1
;
1H
2.19. Secosteroid 10b
NMR (300 MHz, CDCl3) (major isomer): 0.00 (12H, s, Si(CH3)2),
0.64 (s, 3H, H-18), 0.83 (18H, s, C(CH3)3), 0.83 (3H, s, H-19), 0.88
(3H, d, J = 6.4 Hz, H-21), 3.57 (m, 1H, H-8b), 3.72 (m, 1H, H-3b),
3.90 (m, 1H, H-12b), 4.36 (m, 4H, OCH2–CH@), 4.88 (m, 4H, H-24
and H-7), 6.12 (td, J = 6.4 Hz, J = 15.3 Hz, 1H, CH@), 6.17 (td,
J = 6.2 Hz, J = 15.3 Hz, 1H, CH@); 13C NMR (75 MHz, CDCl3): À4.4
(2C), À3.6 (2C), 12.7, 13.1, 15.4, 17.9, 18.3 (2C), 24.6, 24.7 (2C),
25.3, 26.0, 27.6, 28.3, 29.7, 30.4, 31.5, 31.9, 32.4, 33.0, 35.2, 36.0,
36.2, 37.5, 41.9, 44.6, 47.2, 47.6, 63.7, 66.0, 71.2, 72.3, 72.2, 72.3,
Yield 360 mg (76%). Oil. IR (neat) 3418, 2961, 1145 cmÀ1 1H
;
NMR (300 MHz, CDCl3): 0.98 (d, J = 6.4 Hz, 3H, H-21), 1.02 (s, 3H,
H-18), 1.07 (s, 3H, H-19), 2.28 (m, 4H, CH2–C@O), 2.34 (m, 4H,
CH2–C@), 2.81 (m, 1H, H-7b), 2.88 (m, 1H, H-3b), 3.23 (s, 3H,
OCH3), 3.24 (s, 3H, OCH3), 4.13 (m, 4H, H-12 and H-24), 5.17 (m,
4H, CH2@), 5.98 (m, 2H, CH@); 13C NMR (75 MHz, CDCl3): 18.9,
20.3, 23.7, 23.9, 24.4, 28.8, 31.1, 32.6, 32.7, 33.2, 34.0, 34.3, 34.7,
35.5, 35.9, 37.6, 38.1, 41.2, 41.7, 44.9, 47.2, 51.3, 57.2 , 57.4, 64.8,

M. Ibrahim-Ouali, E. Romero / Steroids 78 (2013) 651–661
655
65.1, 72.3, 75.7, 81.8, 82.3, 117.4, 118.1, 129.6, 129.9, 172.1, 172.9.
HRMS (EI) for C36H60O7 [M+] calcd 604.4339 found 604.4344.
H-7), 5.01 (m, 4H, CH2@), 5.80 (m, 2H, CH@); 13C NMR (75 MHz,
CDCl3): À4.8 (2C), À3.6 (2C), 12.8, 14.2, 17.5, 18.2, 20.2 (2C),
21.0, 22.7, 24.3 (2C), 25.5, 25.9, 26.3, 28.5, 28.9, 32.6, 33.9, 35.0,
35.3, 36.0, 37.2, 44.6, 47.2, 47.5, 51.6, 53.4, 53.8, 60.5, 68.3, 69.3,
72.2, 72.5, 73.8, 79.4, 115.4, 115.5, 136.3, 136.4, 1724.2, 173.4.
HRMS (EI) for C44H84O7Si2 [M+] calcd 804.5756 found 804.5761.
2.20. Secosteroid 10c
Yield 360 mg (72%). Oil. IR (neat) 3280, 1220, 1070 cmÀ1 1H
;
NMR (300 MHz, CDCl3): 1.00 (d, J = 6.4 Hz, 3H, H-21), 1.02 (s, 3H,
H-18), 1.06 (s, 3H, H-19), 2.16 (m, 4H, CH2–C@), 2.28 (m, 4H,
CH2–C@O), 2.82 (m, 1H, H-7b), 2.92 (m, 1H, H-3b), 3.23 (s, 3H,
OCH3), 3.25 (s, 3H, OCH3), 4.15 (m, 4H, H-12 and H-24), 5.21 (m,
4H, CH2@), 5.92 (m, 2H, CH@); 13C NMR (75 MHz, CDCl3): 18.7,
20.6, 23.3, 24.1, 24.4, 24.7, 25.6, 28.3, 31.3, 32.7, 33.0, 33.4, 34.6,
34.1, 34.9, 35.7, 36.0, 37.3, 38.4, 40.9, 41.6, 44.3, 47.8, 51.6, 57.4 ,
57.7, 65.1, 65.6, 71.9, 75.3, 81.6, 82.9, 118.3, 118.7, 129.1, 129.8,
172.7, 173.0. HRMS (EI) for C38H64O7 [M+] calcd 632.4652 found
632.4657.
2.25. Secosteroid 22c
Yield 480 mg (74%). Oil. IR (neat) 3429, 1330, 1042 cmÀ1 1H
;
NMR (300 MHz, CDCl3): 0.00 (12H, s, Si(CH3)2), 0.65 (s, 3H, H-18),
0.83 (18H, s, C(CH3)3), 0.88 (3H, s, H-19), 0.93 (3H, d, J = 6.4 Hz,
H-21), 2.04 (4H, m, CH2–C@), 2.27 (m, 4H, CH2–C@O), 3.53 (m,
1H, H-8b), 3.77 (m, 1H, H-12b), 3.87 (m, 1H, H-3b), 4.03 (m, 4H,
H-24 and H-7), 5.01 (m, 4H, CH2@), 5.74 (m, 2H, CH@); 13C NMR
(75 MHz, CDCl3): À4.5 (2C), À3.6 (2C), 12.9, 17.7, 18.2 (2C), 24.2,
24.5 (2C), 24.6, 25.4, 25.9, 27.2, 27.7, 29.5, 29.7, 30.3, 31.4, 31.8,
33.1, 33.7, 34.0, 34.4, 35.0, 35.9, 36.1, 37.4, 37.5, 38.4, 39.2, 41.7,
42.4, 42.9, 44.8, 47.5, 47.7, 64.8, 64.9, 70.3, 72.2, 72.6, 115.3,
115.4, 137.8, 173.7, 173.8. HRMS (EI) for C48H88O7Si2 [M+] calcd
832.6069 found 832.6074.
2.21. Macrocycle 11a
Yield 60 mg (56%). Oil. IR (neat) 2951, 1732, 1370, 1150 cmÀ1
;
1H NMR (300 MHz, CDCl3) (major isomer): 0.98 (s, 3H, H-18),
1.04 (d, J = 6.4 Hz, 3H, H-21), 1.07 (s, 3H, H-19), 2.89 (m, 1H, H-
7b), 2.92 (m, 4H, CH2–C@O), 2.94 (m, 1H, H-3b), 3.23 (s, 3H,
OCH3), 3.24 (s, 3H, OCH3), 4.18 (m, 4H, H-12 and H-24), 5.92 (td,
J = 6.3 and 15.1 Hz, 1H, CH@), 6.09 (td, J = 6.3 Hz and 15.3 Hz, 1H,
CH@); 13C NMR (75 MHz, CDCl3): 18.4, 20.6, 23.1, 24.6, 24.9,
27.1, 31.6, 32.4, 32.7, 33.1, 33.8, 34.9, 36.3, 37.1, 38.3, 41.6, 42.0,
45.4, 47.1, 52.3, 56.9, 57.2, 61.3, 67.2, 72.8, 75.7, 82.4, 82.9,
119.9, 126.7, 171.9, 172.6. HRMS (EI) for C32H52O7 [M+] calcd
548.3713 found 548.3716.
2.26. General procedure for the reduction of 11a-b and 23a-b
A flask equipped with a magnetic stirring bar, an argon outlet
and a condenser was charged with NaBH4 (90 mg, 0.40 mmol)
and anhydr. THF (7 mL) – diglyme (3 mL) under argon. The solution
was cooled at 0 °C and then a solution composed of boron trifluor-
ide etherate (0.42 g, 3 mmol), macrocycle 11a-b or 23a-b
(0.18 mmol) and anhydr. THF (5 mL) was added. After completion
of the reaction (TLC), it was quenched by addition of 2 N hydro-
chloric acid (1 mL) and water (10 mL), the product was extracted
with ether (3 Â 20 mL). The extracts were dried over MgSO4, fil-
tered and then concentrated under vacuum. The residue was chro-
matographed on silica gel (Et2O-petroleum ether 1:1).
2.22. Macrocycle 11b
Yield 55 mg (48%). Oil. IR (neat) 2965, 1728, 1609, 1502 cmÀ1
;
1H NMR (300 MHz, CDCl3) (major isomer): 1.01 (d, J = 6.4 Hz, 3H,
H-21), 1.04 (s, 3H, H-18), 1.12 (s, 3H, H-19), 2.35 (m, 4H, CH2–
C@O), 2.44 (m, 4H, CH2–C@), 2.83 (m, 1H, H-7b), 2.90 (m, 1H, H-
3b), 3.24 (s, 3H, OCH3), 3.25 (s, 3H, OCH3), 4.07 (m, 4H, H-12 and
H-24), 5.86 (td, J = 6.4 and 15.3 Hz, 1H, CH@), 6.01 (td, J = 6.3 Hz
and 15.3 Hz, 1H, CH@); 13C NMR (75 MHz, CDCl3): 18.9, 21.1,
23.4, 24.9, 25.1, 27.8, 31.4, 32.1, 32.9, 33.6, 34.2, 34.9, 35.4, 36.1,
36.8, 37.4, 38.0, 42.2, 42.6, 45.9, 47.4, 52.7, 57.0, 57.7, 61.8, 68.2,
72.8, 74.9, 82.1, 82.7, 119.4, 126.1, 172.8, 173.0. HRMS (EI) for
2.27. Macrocycle 12a
Yield 52 mg (56%). Oil. IR (neat) 3280, 1220, 1070 cmÀ1 1H
;
NMR (300 MHz, CDCl3) (major isomer): 1.02 (s, 3H, H-18), 1.06
(d, J = 6.4 Hz, 3H, H-21), 1.14 (s, 3H, H-19), 2.26 (m, 4H, CH2–C@),
2.87 (m, 1H, H-7b), 2.92 (m, 1H, H-3b), 3.24 (s, 3H, OCH3), 3.25
(s, 3H, OCH3), 3.42 (m, 4H, H-12 and H-24), 5.88 (td, J = 6.4 and
15.3 Hz, 1H, CH@), 6.02 (td, J = 6.3 Hz and 15.3 Hz, 1H, CH@); 13C
NMR (75 MHz, CDCl3): 18.9, 20.4, 23.6, 24.8, 25.1, 28.4, 31.8,
32.4, 32.6, 32.9, 33.4, 34.6, 36.5, 37.3, 39.1, 41.9, 42.6, 46.2, 47.3,
52.1, 56.4, 57.5, 66.8, 67.4, 70.2, 71.1, 72.4, 75.2, 82.1, 82.6,
119.8, 126.3. HRMS (EI) for C32H56O5 [M+] calcd 520.4128 found
520.4133.
C
34H56O7 [M+] calcd 576.4026 found 576.4031.
2.23. Secosteroid 22a
Yield 500 mg (82%). Oil. IR (neat) 3424, 1338, 1130 cmÀ1 1H
;
NMR (300 MHz, CDCl3): 0.00 (12H, s, Si(CH3)2), 0.66 (s, 3H, H-18),
0.85 (18H, s, C(CH3)3), 0.97 (3H, d, J = 6.4 Hz, H-21), 1.02 (3H, s,
H-19), 3.08 (m, 1H, H-8b), 3.44 (m, 4H, CH2–C@O), 3.55 (m, 2H,
H-12 and H-3b), 3.90 (m, 4H, H-24 and H-7b), 5.18 (m, 4H,
CH2@), 5.83 (m, 2H, CH@); 13C NMR (75 MHz, CDCl3): À4.9 (2C),
À3.6 (2C), 12.8, 17.5, 18.0, 20.9 (2C), 23.5, 25.8 (2C), 26.0, 27.6,
29.2, 30.3, 31.2, 31.5, 35.1, 37.5, 37.6, 39.0, 41.4, 41.6, 42.5, 42.9,
44.1, 46.3, 47.5, 47.8, 47.9, 49.6, 50.5, 62.2, 63.2, 72.6, 72.7, 73.4,
118.3 (2C), 130.5 (2C), 174.2, 174.4. HRMS (EI) for C44H80O7Si2
[M+] calcd 776.5443 found 776.5449.
2.28. Macrocycle 12b
Yield 48 mg (48%). Oil. IR (neat) 2981, 1370, 1048 cmÀ1 1H
;
NMR (300 MHz, CDCl3) (major isomer): 1.01 (d, J = 6.4 Hz, 3H, H-
21), 1.04 (s, 3H, H-18), 1.12 (s, 3H, H-19), 2.35 (m, 4H, CH2–
C@O), 2.44 (m, 4H, CH2–C@), 2.83 (m, 1H, H-7b), 2.90 (m, 1H, H-
3b), 3.24 (s, 3H, OCH3), 3.25 (s, 3H, OCH3), 4.07 (m, 4H, H-12 and
H-24), 5.86 (td, J = 6.4 and 15.3 Hz, 1H, CH@), 6.01 (td, J = 6.3 Hz
and 15.3 Hz, 1H, CH@); 13C NMR (75 MHz, CDCl3): 18.7, 21.3,
23.6, 24.5, 24.9, 27.6, 29.6, 31.7, 32.4, 33.1, 34.3, 34.7, 35.1, 36.7,
36.9, 37.1, 37.8, 41.9, 42.3, 44.7, 47.2, 51.9, 57.3, 57.9, 62.3, 69.0,
70.4, 71.6, 73.0, 74.3, 81.9, 82.5, 121.1, 126.3. HRMS (EI) for
2.24. Secosteroid 22b
Yield 470 mg (75%). Oil. IR (neat) 3280, 1356, 1048 cmÀ1 1H
;
NMR (300 MHz, CDCl3): 0.00 (12H, s, Si(CH3)2), 0.64 (s, 3H, H-18),
0.83 (18H, s, C(CH 3)3), 0.89 (3H, s, H-19), 0.94 (3H, d, J = 6.4 Hz,
H-21), 2.40 (m, 8H, CH2–C@O and CH2–C@), 3.42 (m, 1H, H-8b),
3.66 (m, 1H, H-12b), 3.89 (m, 1H, H-3b), 4.08 (m, 4H, H-24 and
C
34H60O5 [M+] calcd 548.4441 found 548.4446.

656
M. Ibrahim-Ouali, E. Romero / Steroids 78 (2013) 651–661
2.29. Macrocycle 13a
31.8, 32.0, 33.3, 35.6, 36.3, 36.7, 37.2, 38.4, 39.6, 41.3, 41.7, 46.6,
47.1, 51.2, 52.3, 66.2, 68.1, 70.6, 70.9, 72.4, 73.1, 76.4, 127.4,
129.9. HRMS (EI) for C42H80O5Si2 [M+] calcd 720.5544 found
720.5549.
Yield 45 mg (92%). Oil. IR (neat) 2984, 1374, 1038 cmÀ1 1H
;
NMR (300 MHz, CDCl3) (major isomer): 1.01 (s, 3H, H-18), 1.06
(d, J = 6.4 Hz, 3H, H-21), 1.16 (s, 3H, H-19), 2.24 (m, 4H, CH2–C@),
3.12 (m, 1H, H-7b), 3.16 (m, 1H, H-3b), 3.52 (m, 4H, H-12 and H-
24), 5.89 (td, J = 6.1 and 15.4 Hz, 1H, CH@), 6.04 (td, J = 6.3 Hz
and 15.3 Hz, 1H, CH@); 13C NMR (75 MHz, CDCl3): 18.7, 20.6,
23.3, 24.1, 25.3, 28.6, 31.7, 32.6, 32.9, 33.1, 33.7, 34.2, 36.7, 37.8,
39.2, 41.6, 42.1, 44.9, 46.9, 52.4, 62.3, 65.1, 66.5, 68.9, 71.3, 77.8,
72.3, 73.9, 119.6, 126.1. HRMS (EI) for C30H52O5 [M+] calcd
492.3815 found 492.3819.
2.34. Macrocycle 24b
Yield 35 mg (46%). Oil. IR (neat) 3435, 2990, 1058 cmÀ1 1H
;
NMR (300 MHz, CDCl3) (major isomer): 0.00 (12H, s, Si(CH3)2),
0.66 (s, 3H, H-18), 0.89 (18H, s, C(CH3)3), 0.98 (3H, d, J = 6.4 Hz,
H-21), 1.17 (3H, s, H-19), 2.31 (m, 8H, (CH2)2–C@), 3.41 (m, 1H,
H-8b), 3.48 (m, 2H, H-12b and H-3b), 3.67 (m, 4H, CH2–O), 3.89
(m, 4H, H-24 and H-7), 5.42 (td, J = 6.1 and 15.4 Hz, 1H, CH@),
5.87 (td, J = 6.2 Hz and 15.3 Hz, 1H, CH@); 13C NMR (75 MHz,
CDCl3): À4.9 (2C), À3.6 (2C), 12.4, 18.7, 19.1 (2C), 23.1, 24.5 (2C),
25.6, 25.9, 26.4, 26.8, 27.1, 29.2, 31.4, 32.0, 34.1, 34.3, 34.9, 35.1,
35.8, 36.6, 37.4, 37.5, 38.6, 39.8, 41.3, 43.2, 45.9, 46.2, 47.4, 54.1,
64.2, 67.9, 69.9, 70.5, 71.1, 72.8, 73.2, 128.1, 129.6. HRMS (EI) for
2.30. Macrocycle 13b
Yield 40 mg (90%). Oil. IR (neat) 3429, 2950, 1606, 1089 cmÀ1
;
1H NMR (300 MHz, CDCl3) (major isomer): 0.99 (s, 3H, H-18),
1.02 (d, J = 6.4 Hz, 3H, H-21), 1.10 (s, 3H, H-19), 2.14 (m, 4H,
CH2–C@), 3.22 (m, 1H, H-7b), 3.31 (m, 1H, H-3b), 4.02 (m, 8H,
CH2-O, H-12 and H-24), 5.72 (td, J = 6.1 and 15.4 Hz, 1H, CH@),
5.97 (td, J = 6.2 Hz and 15.3 Hz, 1H, CH@); 13C NMR (75 MHz,
CDCl3): 18.4, 21.6, 23.9, 24.1, 24.7, 27.3, 29.2, 31.4, 32.0, 32.7,
33.9, 34.6, 35.2, 36.9, 37.1, 37.4, 37.9, 41.6, 43.0, 44.2, 47.1, 52.1,
63.4, 68.8, 70.6, 71.9, 73.1, 73.9, 80.8, 81.7, 119.6, 125.9. HRMS
(EI) for C32H56O5 [M+] calcd 520.4128 found 520.4134.
C
44H84O5Si2 [M+] calcd 748.5857 found 748.5861.
2.35. Macrocycle 25a
Yield 28 mg (95%). Oil. IR (neat) 3410, 2940, 1608 cmÀ1 1H
;
NMR (300 MHz, CDCl3) (major isomer): 0.64 (s, 3H, H-18), 1.04
(3H, d, J = 6.4 Hz, H-21), 1.12 (3H, s, H-19), 3.32 (m, 2H, H-12b
and H-3b), 3.42 (m, 1H, H-8b), 3.48 (m, 4H, CH2–O), 3.72 (m, 4H,
H-24 and H-7), 5.67 (td, J = 6.4 and 15.3 Hz, 1H, CH@), 5.89 (td,
J = 6.3 Hz and 15.4 Hz, 1H, CH@); 13C NMR (75 MHz, CDCl3): 14.1,
17.9, 23.6, 24.4, 27.1, 27.8, 29.1, 31.0, 31.6, 31.7, 32.4, 33.7, 34.8,
36.2, 36.9, 37.6, 38.9, 45.7, 47.8, 51.6, 52.1, 65.8, 67.9, 70.4, 71.2,
72.6, 73.4, 75.9, 126.9, 129.1. HRMS (EI) for C30H52O5 [M+] calcd
492.3815 found 492.3821.
2.31. Macrocycle 23a
Yield 80 mg (54%). Oil. IR (neat) 3280, 1730, 1220, 1070 cmÀ1
;
1H NMR (300 MHz, CDCl3) (major isomer): 0.00 (12H, s, Si(CH3)2),
0.65 (s, 3H, H-18), 0.83 (18H, s, C(CH3)3), 0.94 (3H, d, J = 6.4 Hz,
H-21), 1.02 (3H, s, H-19), 3.38 (m, 1H, H-8b), 3.44 (m, 4H, CH2–
C@O), 3.55 (m, 2H, H-12b and H-3b), 3.90 (m, 4H, H-24 and H-7),
5.76 (td, J = 6.4 and 15.2 Hz, 1H, CH@), 6.02 (td, J = 6.1 Hz and
15.3 Hz, 1H, CH@); 13C NMR (75 MHz, CDCl3): À4.4 (2C), À3.5
(2C), 12.6, 15.3, 17.9, 18.3 (2C), 24.3, 24.7 (2C), 25.1, 26.2, 27.4,
29.8, 30.6, 31.2, 31.6, 31.9, 33.2, 35.4, 36.1, 36.3, 36.6, 37.8, 41.3,
42.1, 46.2, 47.0, 47.3, 50.7, 52.6, 65.4, 66.2, 72.5, 72.9, 76.5,
127.6, 133.9, 174.4, 174.6. HRMS (EI) for C42H76O7Si2 [M+] calcd
748.5130 found 748.5135.
2.36. Macrocycle 25b
Yield 25 mg (96%). Oil. IR (neat) 3386, 1070, 820 cmÀ1; 1H NMR
(300 MHz, CDCl3) (major isomer): 0.67 (s, 3H, H-18), 0.99 (3H, d,
J = 6.4 Hz, H-21), 1.11 (3H, s, H-19), 2.16 (m, 8H, (CH2)2–C@),
3.21 (m, 2H, H-12b and H-3b), 3.39 (m, 4H, CH2–O), 3.45 (m, 1H,
H-8b), 3.67 (m, 4H, H-24 and H-7), 5.48 (td, J = 6.2 and 15.4 Hz,
1H, CH@), 5.89 (td, J = 6.2 Hz and 15.3 Hz, 1H, CH@); 13C NMR
(75 MHz, CDCl3): 12.9, 18.4, 23.3, 24.8, 25.9, 27.1, 29.3, 30.7,
32.4, 33.7, 34.1, 34.9, 35.5, 35.9, 36.2, 37.1, 38.0, 38.9, 42.2, 45.6,
46.3, 47.1, 53.9, 64.6, 68.5, 69.1, 70.7, 71.4, 72.6, 73.7, 127.2,
2.32. Macrocycle 23b
Yield 78 mg (50%). Oil. IR (neat) 3448, 1732, 1607, 1041 cmÀ1
;
1H NMR (300 MHz, CDCl3) (major isomer): 0.00 (12H, s, Si(CH3)2),
0.64 (s, 3H, H-18), 0.83 (18H, s, C(CH3)3), 0.92 (3H, d, J = 6.4 Hz,
H-21), 1.22 (3H, s, H-19), 2.31 (m, 8H, (CH2)2–C@), 3.36 (m, 1H,
H-8b), 3.42 (m, 4H, CH2–C@O), 3.52 (m, 2H, H-12b and H-3b),
3.86 (m, 4H, H-24 and H-7), 5.48 (td, J = 6.4 and 15.4 Hz, 1H,
CH@), 5.93 (td, J = 6.2 Hz and 15.3 Hz, 1H, CH@); 13C NMR
(75 MHz, CDCl3): À4.6 (2C), À3.6 (2C), 12.8, 18.1, 18.6 (2C), 22.6,
24.3, 25.8 (2C), 26.0, 26.2, 26.7, 26.9, 29.6, 31.3, 31.9, 34.1, 34.7,
34.9, 35.5, 35.6, 35.9, 37.2, 37.4, 38.0, 40.9, 41.6, 44.5, 46.3, 47.2,
47.5, 53.5, 64.8, 68.0, 70.5, 72.5, 72.7, 128.4, 129.4, 172.1, 173.2.
HRMS (EI) for C44H80O7Si2 [M+] calcd 776.5443 found 776.5448.
129.8. HRMS (EI) for
520.4132.
C
32H56O5 [M+] calcd 520.4128 found
3. Results and discussion
Olefin metathesis has become one of the most powerful and
attractive tools for the formation of carbon-carbon double bonds
and is widely used in organic synthesis. The spectacular improve-
ments in this reaction achieved over the last two decades are well
known to most chemists. An increasing number of papers devoted
to applications of olefin metathesis in the synthesis of natural
products are observed [38–44]. Although ring-closing metathesis
(RCM) is known to be a powerful tool for the preparation of mac-
rocycles [45–49], in the field of steroid chemistry, only few synthe-
ses have been reported based on metathesis reactions [50].
For some years, we have been interested to develop new syn-
thetic approaches to prepare secosteroidal molecules. We recently
described a simple preparation of 12,13- (Scheme 2) and 7,8-secos-
teroids (Scheme 4) possessing a macrocycle using a ring-closing
metathesis reaction as the key step [51]. Indeed, this combination
2.33. Macrocycle 24a
Yield 40 mg (52%). Oil. IR (neat) 3420, 1264, 905 cmÀ1; 1H NMR
(300 MHz, CDCl3) (major isomer): 0.00 (12H, s, Si(CH3)2), 0.66 (s,
3H, H-18), 0.85 (18H, s, C(CH3)3), 1.02 (3H, d, J = 6.4 Hz, H-21),
1.08 (3H, s, H-19), 3.34 (m, 1H, H-8b), 3.43 (m, 2H, H-12b and H-
3b), 3.66 (m, 4H, CH2–O), 3.90 (m, 4H, H-24 and H-7), 5.72 (td,
J = 6.3 and 15.2 Hz, 1H, CH@), 5.96 (td, J = 6.3 Hz and 15.4 Hz, 1H,
CH@); 13C NMR (75 MHz, CDCl3): À4.6 (2C), À3.5 (2C), 12.2, 14.9,
18.1, 18.8 (2C), 24.1, 24.9 (2C), 25.4, 26.0, 27.5, 28.9, 30.1, 31.4,

M. Ibrahim-Ouali, E. Romero / Steroids 78 (2013) 651–661
657
Scheme 2. Synthesis of a 12,13-secosteroidal macrocycle 9 from cholic acid through 8 steps sequence.
of secocholanic skeleton with varied types of macrocycles, pro-
duces high levels of skeletal diversity and complexity.
Diallylation of primary alcohols at C-24 and C-12 produced the cor-
responding diallyl adduct 7 in moderate yield (58%).
Our strategy is based on a sequential ring-expansion/ring-open-
ing and on a ring closing metathesis reaction [51]. In the two pre-
viously described syntheses (Schemes 2 and 4), we prepared the
RCM substrates from cholic acid, a commercial bile acid both inex-
pensive and readily available. In the case of 12,13-secosteroidal
macrocycles matching a cis A/B ring junction, the key reactions
are depicted in Scheme 2. Simple esterification of cholic acid 1
led to methyl cholate [52], which was methylated with methyl io-
The key step of the synthesis, the RCM reaction of 7 was accom-
plished efficiently with the use of a catalytic amount of Grubbs-II
catalyst (20 mol%) in toluene at 80 °C providing secosteroid 8 in
good yield (65%). However, the reaction was not stereoselective,
a mixture of geometric isomers 8 was obtained with the E isomer.
The structure of this macrocycle was completely characterized by
NMR (400 MHz) and mass spectroscopy methods (see experimen-
tal part). A 8:2 ratio of isomers was obtained with the trans
(J = 15.2 Hz) isomer prevailing over the cis (J = 10.6 Hz) isomer.
Finally, removal of methoxy groups of 8 was carried out with
trimethylsilyl iodide [33] to afford the desired compound 9 in
92% yield.
dide-sodium hydride in THF affording methyl 3a,7a-dimethoxy
cholate 3 in a yield of 94%. Simultaneous protection of the second-
ary hydroxyl groups at C-3 and C-7 was needed prior to the reduc-
tive opening of the lactone ring. Microwave (MW) [53] irradiation
of 3 with pyridinium chlorochromate furnished ketone 4 very
quickly, in a few minutes, and in 70% yield. Baeyer-Villiger oxida-
tion of ketocholane 4 led to lactone 5 as the single regioisomer,
as a result of a higher migration aptitude of the quaternary C-13
compared to the secondary C-11. Next, the simultaneous reduction
of both the lactone moiety on ring C and the ester function at C-24
afforded the diprotected pentahydroxysecocholane 6 in 88% yield.
Encouraged by this result and to show that this ring enlarge-
ment is generally applicable, different types of olefins were sub-
jected to metathesis reaction (Scheme
3 and Table 1). We
present now results of our further study on metathesis of secoster-
oidal esters with various sized acid chain (C4, C5 and C6). Dienes
10a-c were prepared in high yields by reaction of steroid 6 with
alkenoic acids in the presence of DCC and DMAP. All esters

658
M. Ibrahim-Ouali, E. Romero / Steroids 78 (2013) 651–661
O
n
O
n
OH
O
O
OH
HO
HO
a
b
H
H
H
H
MeO
OMe
MeO
OMe
H
H
6
10a-c
O
n
n
O
O
n
n
O
O
O
HO
HO
a : n = 1
b : n = 2
c : n = 3
c
H
H
OR
H
H
OMe
RO
MeO
H
H
11a,b
12a,b R = Me
13a,b R = H
d
Reaction conditions : (a) CH2=CH(CH2)nCOOH, DCC, DMAP, 0 °C to r.t., 12 h;
(b) Grubbs-II catalyst (20 mol%), see table 1; (c) NaBH4, BF3.Et2O, THF-diglyme,
0 °C, 4 h; (d) ISi(CH3)3, CHCl3, r.t., 24 h.
Scheme 3. Synthesis of 12,13-secosteroidal macrocycles 13 from diprotected secosteroid 6.
Table 1
RCM did not occur and the remaining material was mainly the
unreacted starting diene 10c.
Metathesis reactions via Scheme 3.
Entry
Olefin
n
Cata
Condition
11: yield (%)
Otherwise, the reaction was not stereoselective. All metathesis
products were obtained as mixtures of geometric E/Z-isomers with
the E isomer predominating. The structure of these new secosteroi-
dal macrocycles 11a-b was completely characterized by NMR
(400 MHz) and mass spectroscopic methods (see experimental
part).
In the following step, in order to obtain ether-linked macrocy-
cles as for compound 9 (n = 0, see Scheme 2), we reduced esters
11a and 11b to the corresponding ethers 12a and 12b, was carried
out using sodium borohydride and boron trifluoride, in diglyme-
tetrahydrofuran at 0 °C during 4 h. Thus, in the two cases, reduc-
tion according to the Pettit and Piatak procedure [54], afforded
the expected macrocycle derivatives 12a and 12b in satisfactory
yields : 56% for 12a and 48% for 12b.
Finally, removal of methoxy groups of macrocycles 12a and 12b
was carried out with trimethylsilyl iodide [55,56] to afford the de-
sired compounds 13a and 13b in good yields. The problem of for-
mation of isomers can be, of course, overcome by hydrogenation of
the newly formed double bond.
As a continuation of our synthetic and stereochemical studies
on secosteroidal systems containing a macrocycle, a similar strat-
egy was used to obtain 7,8-secosteroidal macrocycles (Scheme 4).
The regioselective oxidation of the hydroxyl group at C-7 was
performed with NBS [57]. The resulting 7-keto derivative 14
was then converted into cholate 15 in good yield (95% yield over
two steps).
1
2
3
4
5
6
7
8
9
10a
10a
10a
10b
10b
10b
10c
10c
10c
1
1
1
2
2
2
3
3
3
A
B
B
A
B
B
A
B
B
CH2Cl2, rt, 24 h
CH2Cl2, rt, 24 h
18
29
56
11
23
48
–
MePh,
D, 4 h
CH2Cl2, rt, 24 h
CH2Cl2, rt, 24 h
MePh,
D, 4 h
CH2Cl2, rt, 24 h
CH2Cl2, rt, 24 h
–
–
MePh,
D
, 4 h
a
A = RuCl2(PCy3)2 = CHCHCMe2;
B = (IMES)(PCy)3Cl2Ru = CHPh;
IMES = 1,3-
dimesityl-4,5-dihydroimidazol-2-ylidene.
obtained were found to be stable and were subjected to RCM reac-
tions. According to the literature, synthesis of unsaturated lactones
through RCM has been accomplished using either Grubbs, first or
second generation, or Hoveyda catalyst.
We decided to test two commercially available metathesis pro-
moters: Grubbs first-generation (GI) and Grubbs second-genera-
tion GII (Table 1). In all cases, the Grubbs catalyst of second
generation was more efficient than the first generation. We carried
out the first experiments under relatively mild conditions: 20 mol%
of catalyst in dry and degassed dichloromethane at room temper-
ature. Using these experimental conditions, we observed fair yields
for 11a and 11b. The yields of the cyclization products of 10a and
10b were improved by increasing the temperature (Table 1, entries
3 and 6). Although many successful applications of RCM to the syn-
thesis of macrocycles with n = 1 and n = 2, in the case of n = 3, the
As expected, the Baeyer–Villiger oxidation of ketocholane 15
furnished exclusively regioisomer 16, as a result of the favored

M. Ibrahim-Ouali, E. Romero / Steroids 78 (2013) 651–661
659
O
O
HO
HO
H
OR
OH
H
a
c
H
O
H
H
H
HO
HO
OH
H
H
1
14 R = H
15 R = CH3
b
O
OMe
RO
TBSO
OH
H
H
H
H
e
f
H
O
OH
OH
RO
TBSO
H
O
16 R = H
17 R = TBS
18
d
RO
TBSO
H
H
H
H
O
g
O
OH
OH
RO
TBSO
O
O
19
20 R = TBS
21 R = H
h
Reaction conditions : (a) NBS, H2O, NaHCO3, 12 h at r.t. then 2h at 80-85 °C, 95%; (b) MeOH, PTSA, Δ, 2h, 100%;
(c) m-CPBA, PTSA, CH2Cl2, r.t., 24 h, 82%; (d) TBDMSCl, imidazole, DMF, r.t., 12 h, 91%; (e) LiAlH4,THF,
0 °C to r.t., 12 h, 90%; (f) allyl bromide, KH, DMF, 0 °C to r.t., 12 h, 56%; (g) Grubbs-II catalyst (20 mol%), CH2Cl2,
r.t., 12 h, 90%; (h) TBAF, THF, r.t.,12 h, 92%.
Scheme 4. Synthesis of a 7,8-secosteroidal macrocycle 21 from cholic acid through 8 steps sequence.
migration of the tertiary C-8 compared to the secondary C-6.
Consecutively, a reaction sequence consisted in TBDMS protection
of OH functionalities, followed by the reductive opening of the lac-
tone ring with simultaneous reduction of the ester function at C-24
and finally regioselective diallylation reaction of primary alcohols
at C-24 and C-7 led to the diallyl secocholane 19 in 38% overall
yield (over four steps). The RCM was promoted by the second gen-
eration Grubbs’ catalyst and the reaction was very efficient. Using
the same conditions as described above for 9, we obtained the
macrocyclic product 20 in 90% yield. Here too, this latter 20 was
completely characterized by NMR and mass spectroscopic methods
(see experimental part). The E/Z 9:1 ratio observed in the 1H NMR
spectrum of the crude substance was strongly in favor of the E-iso-
mer (J = 15.6 Hz) with small amounts of sterically less favored Z-
double bond (J = 10.4 Hz).
ogy is the possibility for the expansion of the ring size by adjusting
the length of the acid source used for the synthesis of RCM precur-
sors 22.
Thus, three alkenoates (compounds 22a-c) were prepared in
good yields by reaction of diprotected pentadihydroxysecocholane
18 with alkenoic acids in presence of DCC and DMAP. Here too, es-
ters 22a-c were found to be stable and were subjected to RCM
reactions using Grubbs first-generation (GI) or second-generation
(GII) as catalyst (see Table 2).
It is worth pointing out similar results with the work we de-
scribed above for the synthesis of 12,13-secosteroidal macrocycles.
Indeed, the cyclization of 22 was observed for n = 1 and n = 2 but
without any success for n = 3. Here too, the remaining material
was mainly the unreacted starting diene 22c. It is interesting to
note that the yields and the isomer ratio observed for 23a and
23b are exactly or quite the same as those reported for the
12,13-secosteroidal series. Otherwise, in some experiments, the
reaction temperature was raised to 40 °C without significant
change of reaction course. Only the use of the catalyst of second
generation (Grubbs II) at 80 °C in dry and degassed toluene proved
effective in the RCM products formation 23a or 23b (Table 2,
The 3- and 12-TBS ethers on the A and C rings of 20 were depro-
tected in a standard manner to afford the corresponding secoster-
oidal alcohol 21 in good yield.
We have then turned our attention toward the synthesis of new
macrocycles by introducing ester functionalities on the secosteroi-
dal alcohol 18 (Scheme 5). An interesting feature of this methodol-

660
M. Ibrahim-Ouali, E. Romero / Steroids 78 (2013) 651–661
TBSO
H
TBSO
OH
H
H
H
O
a
b
OH
OH
OH
O
n
TBSO
TBSO
O
18
O
n
22a-c
RO
H
TBSO
H
H
H
O
O
a : n = 1
b : n = 2
c : n = 3
c
OH
OH
n
O
n
RO
TBSO
n
O
O
n
O
24a,b R = TBS
25a,b R = H
23a,b
d
Reaction conditions : (a) CH2=CH(CH2)nCOOH, DCC, DMAP, 0 °C to r.t., 12 h;
(b) Grubbs-II catalyst (20 mol%), see table 2; (c) NaBH4, BF3.Et2O, THF-diglyme,
0 °C, 4 h; (d) TBAF, THF, r.t.,12 h.
Scheme 5. Synthesis of 7,8-secosteroidal macrocycles 25 from diprotected secosteroid 18.
Table 2
to check the biological activity of these steroids and application
of this strategy to obtain new structures are being conducted by
our group.
Metathesis reactions via Scheme 5.
Entry
Olefin
n
Cata
Condition
23: yield (%)
1
2
3
4
5
6
7
8
9
22a
22a
22a
22b
22b
22b
22c
22c
22c
1
1
1
2
2
2
3
3
3
A
B
B
A
B
B
A
B
B
CH2Cl2, rt, 24 h
CH2Cl2, rt, 24 h
12
26
54
9
24
50
–
Acknowledgements
MePh,
D, 4 h
CH2Cl2, rt, 24 h
CH2Cl2, rt, 24 h
This work has been financially supported by the CNRS and the
Ministère de l’Enseignement Supérieur et de la recherche.
MePh,
D, 4 h
CH2Cl2, rt, 24 h
CH2Cl2, rt, 24 h
–
–
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D
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