BF3·Et2O-induced stereoselective aldol reaction with benzaldehyde, and steroid sapogenins and its application to a convenient synthesis of dinorcholanic lactones

Article abstract of DOI:10.1016/j.steroids.2012.02.016

Treatment of steroid sapogenins with benzaldehyde and BF ₃·Et₂O cleanly produces E-23(23′)- benzylidenspirostanes in good yields in a reaction pathway which consists on an aldol reaction followed by a dehydration step. The obtained E-23(23′)- benzylidenspirostanes can be easily converted to dinorcholanic lactones by treatment with CrO₃ in acetic acid. The synthetic sequence to dinorcholanic lactones is compatible with the presence of double bonds and carbonyl groups in the steroid framework.

Full text of DOI:10.1016/j.steroids.2012.02.016

Steroids 77 (2012) 819–828
Contents lists available at SciVerse ScienceDirect
Steroids
journal homepage: www.elsevier.com/locate/steroids
BF3ꢀEt2O-induced stereoselective aldol reaction with benzaldehyde, and steroid
sapogenins and its application to a convenient synthesis of dinorcholanic lactones
Karen M. Ruíz-Pérez, Margarita Romero-Ávila, Verónica Tinajero-Delgado, Marcos Flores-Álamo,
⇑
Martín A. Iglesias-Arteaga
Departamento de Química Orgánica, Facultad de Química, Universidad Nacional Autónoma de México, Ciudad Universitaria, 04510 Mexico, DF, Mexico
a r t i c l e i n f o
a b s t r a c t
Treatment of steroid sapogenins with benzaldehyde and BF3ꢀEt2O cleanly produces E-23(230)-benzyliden-
spirostanes in good yields in a reaction pathway which consists on an aldol reaction followed by a dehy-
dration step. The obtained E-23(230)-benzylidenspirostanes can be easily converted to dinorcholanic
lactones by treatment with CrO3 in acetic acid. The synthetic sequence to dinorcholanic lactones is com-
patible with the presence of double bonds and carbonyl groups in the steroid framework.
Ó 2012 Elsevier Inc. All rights reserved.
Article history:
Received 29 April 2011
Received in revised form 21 February 2012
Accepted 22 February 2012
Available online 15 March 2012
Keywords:
Aldol reaction
Steroids sapogenins
E-23(230)-benzylidenspirostanes
Spiroketals
Dinorcholanic lactones
X-ray diffraction
1. Introduction
Considering that in the case of steroid sapogenins, the concen-
tration of the reactive enol ether II in the equilibrium is consider-
The chemistry of steroid sapogenins (SS) has concentrated the
attention of organic chemist for nearly a century. A wide variety
of reactions of the spiroketal side chain discovered and initially
considered mere curiosities have became useful tools for the trans-
formation of SS into bioactive compounds that include sexual and
adrenocortical hormones [1], ecdysteroids [2], plant growth stimu-
lators [3–9], and cytotoxic steroids [10–16] among many others.
The search for new reactions of the spiroketal side chain of SS stills
an interesting task from both the mechanistic and synthetic point
of views.
Meanwhile the chemical properties of the steroid nucleus of SS
are similar to those of the steroids in general, the reactivity of the
spiroketal side chain constitute the distinguishing characteristic of
SS. It is well known that the presence of Brønsted or Lewis acids
catalyzes the ketal ¡ enol ether equilibrium associated to the side
chain of SS, increasing the concentration of the exocyclic enol ether
II. The reaction of the enol ether II with electrophiles gives place to
a variety of methods for the introduction of substituents at posi-
tion C-23 that allow the transformation of the side chain paving
the way to the synthesis of a wide diversity of bioactive steroids
[17–25].
ably low and can be slightly increased by the addition of a Lewis
acid (see Scheme 1), we envisaged that in such conditions a
controlled nucleophilic addition of the in situ produced enol ether
II to a non-enolizable aldehyde could be achieved. This prompted
us to employ benzaldehyde as a model to explore the possibility
to produce aldol reactions in the side chain of readily available
SS that are commonly employed as starting materials for the
industrial production of steroids of medicinal interests.
Herein we report on the BF3ꢀEt2O-induced aldol condensation of
different steroid sapogenins with benzaldehyde that cleanly effects
the high yield and stereoselective introduction of an E-benzylidene
OA
O
26
26
20
22
O
23
20
22
23
A
O
16
16
X
I
II
+
A= Brønsted or Lewis acid, X = Electrophile (D , Br+, I+, I3+, NO+, etc)
⇑
Corresponding author. Tel./fax: +52 55 56223803.
E-mail address: martin.iglesias@servidor.unam.mx (M.A. Iglesias-Arteaga).
Scheme 1. Acid catalyzed ketal
sapogenins.
¡ enol ether of the side chain of steroid
0039-128X/$ - see front matter Ó 2012 Elsevier Inc. All rights reserved.
http://dx.doi.org/10.1016/j.steroids.2012.02.016

820
K.M. Ruíz-Pérez et al. / Steroids 77 (2012) 819–828
group at position C-23 of the spiroketal side chain. A convenient
alternative for the synthesis of diverse dinorcholanic lactones
(including those containing carbonyl groups or double bonds) by
treatment of the obtained E-23(230)-benzylidenspirostanes with
CrO3 in acetic acid is also described.
1.10 (d, J = 6.9 Hz, 3H, H-21), 0.85 (s, 3H, H-19), 0.83 (s, 3H, H-
18), 0.80 (d, J = 6.6 Hz, 3H, H-27). 13C NMR (100 MHz) d ppm: d
36.7 C-1; 27.4 C-2; 73.6 C-3, 34.0 C-4, 44.6 C-5, 28.5 C-6, 32.2 C-
7, 35.0 C-8, 54.2 C-9, 35.6 C-10, 21.1 C-11, 40.2 C-12, 40.8 C-13,
56.2 C-14, 31.6 C-15, 80.4 C-16, 61.6 C-17, 16.7 C-18, 12.3 C-19,
37.3 C-20, 14.8 C-21, 110.5 C-22; 137.1 C-23⁄, 32.9 C-24, 33.1 C-
25, 66.1 C-26, 17.2 C-27, 170.7 C@O acetyl, 21.4 CH3 acetyl,
137.2 ipso⁄, 129.1 ortho, 128.1 meta, 126.5 para, 122.8 C-230. EIMS
m/z 546 M+ (25), 469(31), 468 (23), 466 (12), 456 (12), 453 (14),
428 (14), 416 (31), 415 (12), 325 (22), 315 (16), 269 (12), 256
(19), 255 (90), 254 (21), 253 (20), 252 (14), 251 (20), 239 (23),
238 (13), 227 (25), 226 (12), 215 (10), 214 (15), 213 (11), 212
(12), 211 (21), 204 (15), 203 (100), 201 (13), 199 (19), 198 (10),
197 (14), 185 (13), 183 (11), 174 (12), 173 (13), 171 (12), 165
(11), 161 (30), 159 (17), 157 (15), 155 (13), 149 (11), 147 (36),
146 (11), 145 (25), 143 (18), 142 (10), 141 (13), 135 (11), 133
(16), 131 (22), 129 (22), 128 (14), 121 (16), 119 (17), 117 (27),
115 (20), 109 (12), 107 (32) 106 (10), 105 (34), 95 (41), 93 (31)
91 (60), 81 (24), 79 (24), 77 (14), 69 (11), 55 (18). HREIMS m/z
546.3713 (calcd. for C36H50O4, 546.3704).
2. Experimental section
2.1. General experimental procedures
Reactions were monitored by TLC on ALUGRAMÒ SIL G/UV254
plates from MACHEREY–NAGEL. Chromatographic plates were
sprayed with a 1% solution of vanillin in 50% HClO4 and heated un-
til color developed. Melting points were measured on a Melt-Temp
II equipment and are uncorrected. Electronic impact Mass spectra
(70 eV) were registered in a Thermo-Electron spectrometer model
DFS (Double Focus Sector). NMR spectra were recorded in CDCl3
solutions in a Varian INOVA 400 spectrometer using the solvent
signal 7.26 ppm for 1H and 77.00 ppm for 13C as references. NMR
signals assignments were made with the aid of a combination of
2D homonuclear (1H–1H) and heteronuclear (1H–13C) correlation
techniques, which included Correlation Spectroscopy (COSY),
Nuclear Overhauser Effect Spectroscopy (NOESY), Heteronuclear
Single Quantum Correlation (HSQC) and Heteronuclear Multiple
Bond Correlation (HMBC). All 2D NMR spectra were recorded using
the standard pulse sequences and parameters recommended by
the manufacturer and processed employing the NMR processing
program MestreNova. X-ray measurements were performed on
an Oxford Diffraction Atlas (Gemini) diffractometer.
2.4. [23(230)E, 25S]-23(230)-Benzylidene-5b-spirostan-3b-ol acetate
(2b)
Yield: 475 mg (87%). Mp 197–198 °C (ethyl acetate). 1H NMR
(400 MHz, CDCl3) d ppm: 7.32 (m, 2H, H-meta), 7.21 (m, 3H, H-
ortho and H-para), 6.72 (s, 1H, H-230), 5.07 (m, 1H, H-3), 4.47
(dd, J = 16.0, 7.2 Hz, 1H, H-16), 4.05 (dd, J = 11.0, 4.2 Hz, 1H, H-26
ax.), 3.32 (d, J = 11.0 Hz, 1H, H-26 eq.), 2.72 (ddd, J = 13.6, 5.6,
1.2 Hz, 1H, H-24 eq.), 2.54–2.45 (m, 1H, H-20), 2.29 (dd, J = 13.6,
5.2 Hz, 1H, H-24 ax.), 1.92–1.84 (m, 1H, H-17) 2.04 (s, 1H, CH3 acet-
yl), 1.11 (d, J = 6.9 Hz, 3H, H-21), 0.99 (s, 3H, H-19), 0.99 (d,
J = 7.1 Hz, 3H, H-27), 0.85 (s, 3H, H-18). 13C NMR (100 MHz) d
ppm: 30.6 C-1; 25.0 C-2; 70.7 C-3, 31.6 C-4, 39.3 C-5, 26.4 C-6,
26.4 C-7, 35.2 C-8, 40.0 C-9, 35.0 C-10, 20.9 C-11, 40.4 C-12, 40.9
C-13, 56.4 C-14, 30.7 C-15, 81.0 C-16, 61.8 C-17, 16.7 C-18, 23.9
C-19, 39.8 C-20, 14.7 C-21, 111.4 C-22; 136.8 C-23⁄, 30.9 C-24,
30.3 C-25, 65.1 C-26, 18.3 C-27, 170.7 C@O acetyl, 21.5 CH3 acetyl,
137.2 ipso⁄, 129.1 ortho, 128.0 meta, 126.5 para, 122.8 C-230. EIMS
m/z 546 M+ (17), 486 (11), 469 (20), 468 (17), 429 (10), 417 (15),
416 (47), 415 (14), 395 (12), 269 (10), 256 (22), 255 (100), 253
(12), 251 (15), 239 (11), 227 (18), 211 (14), 204 (11), 203 (75),
201 (16), 199 (11), 173 (12), 161 (20), 159 (12), 149 (11), 147
(21), 145 (16), 143 (11), 133 (11), 131 (17), 129 (14), 119 (12),
117 (19), 115 (12), 107 (24), 105 (23), 95 (23), 93 (24) 91 (40),
81 (18), 79 (19), 55 (13). HREIMS m/z 546.3716, (calcd. for
2.2. General procedure for the synthesis of E-23(230)-
benzylidenspirostanes
Freshly distilled BF3ꢀEt2O (6 ml) was added to solution of the
steroid sapogenin (1 mmol) and benzaldehyde (212 mg, 2 mmol)
in dry CH2Cl2 (30 ml) and the mixture was stirred under argon at
room temperature until the starting steroid sapogenin was con-
sumed (TLC, see Table 1 for reaction times and yields). The reaction
mixture was washed with water (8 ꢁ 50 ml), dried (anh. Na2SO4)
and evaporated to produce a syrupy residue that was purified in
a chromatographic column packed with silica gel to afford the cor-
responding E-23(230)-benzylidenspirostane.
2.3. [23(230)E,25R]-23(230)-Benzylidene-5
a-spirostan-3b-ol acetate (2a)
Yield: 502 mg (92%). Mp 215–216 °C (ethyl acetate). 1H NMR
(400 MHz, CDCl3) d ppm: 7.37–7.30 (m, 2H, H-meta), 7.27–7.19
(m, 3H, H-ortho and H-para), 6.57 (s, 1H, H-230), 4.69 (ddd,
J = 16.2, 11.2, 4.9 Hz, 1H, H-3), 4.44 (dd, J = 16.0, 7.2 Hz, 1H, H-
16), 3.61–3.49 (m, 2H, H-26 ax./eq.), 2.78 (dd, J = 13.3, 3.7 Hz, 1H,
H-24, eq.), 2.59 (dq, J = 6.9 Hz, 1H, H-20), 2.14–2.06 (m, 1H, H-24
ax.), 2.02 (s, 3H, CH3 acetyl), 1.90 (dd, J = 8.9, 7.2 Hz, 1H, H-17),
C36H50O4, 546.3704).
2.5. [23(230)E,25R]-23(230)-Benzylidenspirost-5-en-3b-ol acetate (2c)
Yield: 495 mg (91%). Mp 190–191 °C (ethyl acetate). 1H NMR
(400 MHz, CDCl3) d ppm: 7.37–7.29 (m, 2H, H-meta), 7.27–7.19
(m, 3H, H-ortho and H-para), 6.58 (s, 1H, H-230), 5.38 (d,
J = 4.9 Hz, 1H, H-6), 4.60 (ddd, J = 10.4, 8.6, 4.3 Hz, 1H, H-3), 4.46
(dd, J = 16.0, 7.2 Hz, 1H, H-16), 3.62–3.50 (m, 2H, H-26 ax./eq.),
2.78 (dd, J = 13.3, 3.9 Hz, 1H, H-24 eq.), 2.60 (m, 1H, H-20), 2.38–
2.27 (m, 2H H-4 ax./eq.), 2.15–2.06 (m, 1H, H-24 ax.), 2.03 (s, 3H,
CH3 acetyl), 1.94–1.87 (m, 1H, H-17), 1.11 (d, J = 6.9 Hz, 3H, H-
21), 1.05 (s, 3H, H-19), 0.86 (s, 3H, H-18), 0.80 (d, J = 6.6 Hz, 3H,
H-27). 13C NMR (100 MHz) d ppm: 36.9 C-1; 27.7 C-2; 73.9 C-3,
38.1 C-4, 139.7 C-5, 122.3 C-6, 30.1 C-7, 31.3 C-8, 49.9 C-9, 36.7
C-10, 20.9 C-11, 39.9 C-12, 40.5 C-13, 56.4 C-14, 31.76 C-15, 80.4
C-16, 61.5 C-17, 16.5 C-18, 19.3 C-19, 37.3 C-20, 14.9 C-21, 110.6
C-22, 137.1 C-23⁄, 32.9 C-24, 33.2 C-25, 66.1 C-26, 17.2 C-27,
170.5 C@O acetyl, 21.4 CH3 acetyl, 137.2 ipso⁄, 129.1 ortho, 128.1
meta, 126.6 para, 122.8 C-230. EIMS m/z 544 M+ (12), 467 (20),
Table 1
Results of the BF3ꢀEt2O-catalyzed reaction of steroid sapogenins with benzaldehyde.
Entry Starting material
Reaction time
(h)
Product Yielda
(%)
1
2
Tigogenin acetate (1a)
Sarsasapogenin acetate
(1b)
5.5
1.5
2a
2b
92
87
3
4
5
Diosgenin acetate (1c)
Hecogenin acetate (1d)
Hecogenin acetate (1d)
6
6
24
2c
2d
2d
91
53+b
65+b
a
Yields after purification in chromatographic column.
Recovered starting material.
b

K.M. Ruíz-Pérez et al. / Steroids 77 (2012) 819–828
821
414 (14), 282 (16), 267 (13), 254 (11), 253 (48), 227 (27), 213 (14),
204 (16), 203(100), 201 (14), 199 (15), 197 (10), 187 (11), 174 (17),
173 (11), 171 (17), 161 (15), 159 (19), 158 (15), 157 (35), 147 (18),
144 (12), 143 (30), 141 (11), 133 (19), 131 (34), 129 (26), 128 (14),
121 (16), 118 (17), 118 (11), 117 (28), 115 (17), 109 (17), 107 (19),
95 (15), 93 (21), 91 (46), 81 (22), 79 (19), 77 (10), 69 (10), 67 (15),
55 (14). HREIMS m/z 544.3570, (calcd. for C36H48O4, 544.3547).
4.6 Hz, 1H, H-16), 4.66 (tt, J = 11.4, 4.9 Hz, 1H, H-3), 2.56 (dq,
J = 7.6, 1.0 Hz, 1H, H-20), 2.24 (ddd, J = 10.5, 7.8, 6.7 Hz, 1H), 2.01
(s, 3H, CH3-acetyl), 1.29 (d, J = 7.6 Hz, 3H, H-21), 0.82 (s, 3H, H-
19), 0.73 (s, 3H, H-18). 13C NMR (100 MHz) d ppm: C-1 36.7, C-2
28.2, C-3 73.5, C-4 36.1, C-5 44.6, C-6 28.3, C-7 32.0, C-8 35.5,
C-9 54.6, C-10 36.0, C-11 20.5, C-12 38.3, C-13 41.7, C-14 54.2, C-
15 33.0, C-16 82.7, C-17 59.0, C-18 13.8, C-19 12–2, C-20 36.1, C-
21 17.9, C-22 181.3 CH3 acetyl 21.4, C@O acetyl 170.6.
2.6. [23(230)E,25R]-23(230)-Benzyliden-3b-acetoxy-5
one (2d)
a-spirostan-12-
2.9. 3b-Acetoxy-16-hydroxy-5b-dinorcholanic acid-(22 ? 16) lactone
(3b)
Yield: 364 mg (65%). Mp 260–262 °C (ethyl acetate). 1H NMR
(400 MHz, CDCl3) d ppm: 7.33 (m, 2H, H-meta), 7.23 (m, 3H, H-
ortho and H-para), 6.54 (s, 1H, H-230), 4.68 (ddd, J = 15.9, 11.0,
4.8 Hz, 1H, H-3), 4.37 (dd, J = 17.6, 8.8 Hz, 1H, H-16), 3.56–3.51
(m, 2H, H-26 ax./eq.), 2.79 (dd, J = 13.3, 4.0 Hz, 1H, H-24 eq.),
2.67 (dd, J = 8.9, 7.2 Hz, 1H, H-17), 2.50–2.38 (m, 1H, H-20 and
H-11 ax.), 2.25 (dd, J = 14.3, 5.0 Hz, 1H, H-11 eq.), 2.02 (s, 3H, CH3
acetyl), 1.20 (d, J = 6.9 Hz, 3H, H-21), 1.11 (s, 3H, H-18), 0.93 (s,
3H, H-19), 0.80 (d, J = 6.6 Hz, 3H, H-27). 13C NMR (100 MHz) d
ppm: 36.2 C-1, 27.2 C-2, 73.1 C-3, 33.8 C-4, 44.4 C-5, 28.1 C-6,
31.5 C-7, 34.2 C-8, 55.3 C-9, 36.1 C-10, 37.7 C-11, 213.2 C-12,
55.3 C-13, 55.6 C-14, 31.0 C-15, 78.9 C-16, 52.9 C-17, 16.2 C-18,
11.9 C-19, 37.8 C-20, 13.6 C-21, 110.6 C-22; 137.0 C-23, 32.9 C-
24, 33.1 C-25, 66.1 C-26, 17.2 C-27, 170.6 C@O acetyl, 21.4 CH3
acetyl, 137.0 ipso, 129.1 ortho, 128.1 meta, 126.6 para, 122.9 C-
230. EIMS m/z 561 MH+, (16), 484 (23), 471 (11), 432 (12), 431
(38), 430 (15), 255 (14), 239 (16), 231 (11), 227 (26), 226 (17),
211 (24), 204 (15), 203 (100), 201 (15), 199 (13), 197 (17), 187
(13), 185 (15), 174 (15), 173 (14), 172 (12), 172 (19), 170 (10),
169 (13), 161 (14), 159 (14), 157 (19), 156 (11), 155 (17), 147
(29), 145 (41), 144 (13), 143 (24), 142 (11), 142 (13), 135 (18),
134 (10), 133 (16), 132 (11), 131 (63), 129 (27), 121 (27), 119
(23), 118 (12), 117 (44), 116 (10), 115 (22), 109 (29), 108 (16),
107 (33), 105 (44), 95 (24), 93 (37), 91 (77), 81(24), 77 (18), 69
(11), 55 (19). HREIMS m/z 561.3585 MH+ (calcd. for C36H49O5,
561.3575).
Yield: 270 mg (70%). Mp. 180 °C (methanol), Lit. 184.5–185.5 °C
[27] 1H NMR (400 MHz, CDCl3) d ppm: 5.05 (m, J = 3 Hz, 1H, H-3),
4.93 (td, J = 8, 5 Hz, 1H, H-16), 2.56 (qd, J = 8, 1 Hz, 1H, H-20), 2.25
(ddd, J = 13, 8, 6 Hz, 1H), 2.03 (s, 3H, CH3 acetyl), 1.30 (d, J = 8 Hz,
3H, H-21), 0.97 (s, 3H, H-19), 0.73 (s, 3H, H-18) 13C NMR
(100 MHz) d ppm: C-1 30.5, C-2 25.0, C-3 70.5, C-4 30.7, C-5
37.2, C-6 26.3, C-7 26.2, C-8 35.0, C-9 40.1, C-10 35.0, C-11 20.4,
C-12 38.5, C-13 41.8, C-14 54.7, C-15 33.0, C-16 82.8, C-17 59.1,
C-18 13.8, C-19 23.8, C-20 36.0, C-21 18.0, C-22 181.3, CH3 acetyl
21.5, C@O acetyl 170.7.
2.10. 3b-Acetoxy-16-hydroxy-dinorchol-5-enic acid-(22 ? 16)
lactone (vespertilin acetate) (3c)
Yield: 247 mg (64%). Mp. 211–213 °C (ethyl acetate/hexane),
Lit. 218–219.5 °C [28]. 1H NMR (400 MHz, CDCl3) d ppm: 5.35 (d,
J = 5.2 Hz, 1H, H-5), 4.94 (td, J = 7.8, 4.7 Hz, 1H, H-16), 4.66–4.50
(m, 1H, H-3), 2.57 (qd, J = 7.6, 1.2 Hz, 1H, H-2), 2.02 (s, 3H, CH3
acetyl), 1.30 (d, J = 7.6 Hz, 1H, H-21), 1.02 (s, 1H, H-19), 0.76 (s,
1H, H-18). 13C NMR (100 MHz) d ppm: 36.9 C-1, 27.6 C-2, 73.7 C-
3, 38.0 C-4, 139.8 C-5, 121.9 C-6, 31.8 C-7, 31.2 C-8, 49.9 C-9,
36.6 C-10, 20.3 C-11, 38.1 C-12, 41.4 C-13, 54.6 C-14, 33.0 C-15,
82.6 C-16, 58.9 C-17, 13.7 C-18, 19.3 C-19, 36.0 C-20, 18.0 C-21,
181.2 C-22, 170.4, C@O acetyl, 21.4 CH3 acetyl.
2.7. General procedure for the synthesis of of dinorcholanic lactones
2.11. 3b-Acetoxy-12-oxo-16-hydroxy-5b-dinorcholanic acid-(22 ? 16)
lactone (3d)
A
solution of the E-23(230)-benzylidenspirostane (2a–d)
(1 mmol) in an acetic (35 ml) was warmed until total dissolution
of the solid, followed by addition of CrO3 (545 mg dissolved in
6 ml of 4/1 AcOH/H2O mixture). The reaction mixture was stirred
until the starting material was consumed (TLC, aprox. 30 min)
poured into water and extracted with ethyl acetate (2 ꢁ 50 ml).
The organic layer was washed with water (2 ꢁ 20 ml), aqueous
(10%) NaHCO3 (5 ꢁ 20 ml), water (2 ꢁ 20 ml), brine (20 ml), dried
(anh. Na2SO4) and evaporated to produce a yellowish syrup. Crys-
tallization from the indicated solvent (see below) afforded the pure
lactone (See Table 2 for yields).
Yield: 258 mg (64%) Mp. 222–224 °C (acetone). 1H NMR
(400 MHz, CDCl3) d ppm: 4.88 (1H, td, J = 7.7, 4.7 Hz, 1H, H-16),
4.66 (1H, tt, J = 11.1, 4.9 Hz, 1H, H-3), 2.63–2.52 (1H, m), 2.01 (s,
3H, CH3 acetyl), 1.38 (3H, d, J = 7.5 Hz, 3H, H-21), 1.05 (s, 3H, H-
18), 0.91 (s, 3H, H-19). 13C NMR (100 MHz) d ppm: 36.2 C-1, 27.1
C-2, 73.9 C-3, 33.7 C-4, 44.4 C-5, 27.9 C-6, 31.3 C-7, 34.1 C-8,
55.5 C-9, 36.1 C-10, 37.2 C-11, 212.3 C-12, 55.7 C-13, 50.8 C-14,
32.4 C-15, 80.8 C-16, 53.9 C-17, 13.8 C-18, 11.8 C-19, 36.7 C-20,
17.2 C-21, 180.7 C-22, 170.6, C@O acetyl, 21.3 CH3 acetyl. EIMS
m/z 402 M+ (5), 343 (24), 342 (100), 329 (10), 327 (15), 300 (18),
283 (14), 246 (24), 179 (27), 173 (20), 161 (14), 159 (11), 149
(12), 148 (20), 147 (63), 145 (18), 135 (14), 131 (11), 123 (20),
122 (12), 121 (29), 119 (22), 109 (24), 108 (26), 107 (42), 106
(13), 105 (54), 95 (24), 94 (10), 93 (38), 91 (31), 86 (34), 84 (55),
81 (26), 79 (29), 77 (22), 69 (12), 67 (22). HREIMS m/z 402.2423
(calcd. for C24H34O5, 402.2401).
2.8. 3b-Acetoxy-16-hydroxy-5a-dinorcholanic acid-(22 ? 16) lactone.
(3a)
Yield: 270 mg (70%). Mp. 222–224 °C, (methanol), Lit. 222–
224 °C [26]. 1H NMR (400 MHz, CDCl3) d ppm: 4.92 (td, J = 7.8,
Table 2
Yields of dinorcholanic lactones (3a–d).
Entry
Starting material
Product
Yield⁄ (%)
1
2
3
4
(23(230)E)-23(230)-Benzylidentigogenin acetate (2a)
(23(230)E)-23(230)-Benzylidensarsasapogenin acetate (2b)
(23(230)E)-23(230)-Benzyliden diosgenin acetate (2c)
(23(230)E)-23(230)-Benzyliden hecogenin acetate (2d)
3a
3b
3c
3d
70
70
64
64
The reactions were conducted in 1–3 mmol amounts with similar results.

822
K.M. Ruíz-Pérez et al. / Steroids 77 (2012) 819–828
as well to the field effects associated to the presence of the C-
3. Crystal structure determination
23–C-230 double bond and the aromatic ring (see Fig. 1). The strong
NOE effect observed between H-20 and H-230 indicates proximity
between these nuclei only possible in the E-isomer (see Fig. 1
and Supplementary information file). The other 1H and 13C signals
corresponding to the steroid skeleton are in good agreement with
previous reported data [31–33].
Colorless crystals of 2a, 2b and 3b grown on slow evaporation of
ethyl acetate solutions were mounted under paratone oil on glass
fibers and placed under a cold nitrogen stream at 130 K on a Oxford
Diffraction Gemini ‘‘A’’ diffractometer with a CCD area detector,
with radiation source of kCuK = 1.5418 Å using graphite-mono-
a
chromatized radiation. CrysAlisPro and CrysAlis RED software
packages were used for data collection and data integration. [29]
The structures of both molecules were solved and refined using
SHELXLS and SHELXL-97 [30]. All non-hydrogen atoms were re-
fined anisotropically. The hydrogen atoms were found in difference
Fourier maps and placed at geometrically calculated positions and
isotropically refined using the riding model. Crystal data of com-
pounds 2a, 2b and 3d are shown in Table 3.
4.1. X-ray diffraction studies of benzylidene spirostanes
We were pleased to find that compounds 2a and 2b produced
suitable crystals for X-ray diffraction studies that confirmed the
proposed structures and the E-geometry of the introduced double
bond. Steroid sapogenins 2a and 2b contain the (25R)-5a- and
(25S)-5b-spirostane frameworks, respectively. Puckering parame-
ters for both compounds were calculated as described by Cremer
and Pople [34].
4. Results and discussion
The presence of the b-equatorial acetoxy group attached to C-3
in 2a does not disturb the chair conformation of ring A [puckering
parameters Q = 0.575(2) Å, h = 5.1(2) and / = 308(3)°, if the calcula-
tion starts from C-1 to C-10 and proceeds in a counterclockwise
direction]. The acetoxy group is arranged in such a way that the
Treatment of the (25R)-5a-spirostan-3b-ol acetate (1a) (namely
tigogenin acetate) with BF3ꢀEt2O and two equivalents of benzalde-
hyde in dry CH2Cl2 produced the stereoselective introduction of an
E-benzylidene group at C-23 in excellent yield (See Table 1). To test
the generality of the reaction, the procedure was applied to other
steroid sapogenins. When sarsasapogenin, diosgenin and hecoge-
nin acetates (1b–d) were subjected to the same treatment, similar
results were obtained, indicating that the procedure is applicable
to 25S-spirostans as 1b and that is also compatible with the pres-
ence of a double bond at C-5 in 1c, or a carbonyl group at C-12 in
1d (see Scheme 2 and Table 1). The reactions were carried out in 1–
5 mmol amounts with similar results.
The presence of the benzylidene moiety in the side chain is
characterized by the 1H and 13C signals associated with the phenyl
group. In addition the 13C signals of the sp2 carbons C-23 and C-230
as well as the signal of olefinic proton attached to C-230 corrobo-
rates the produced transformation. The observed downfield shift
of the signal of H-24 eq. can be attributed to allylic compression
oxygen atom of the carbonyl nearly eclipses the a-axial hydrogen
bonded to C-3. Ring B assumes an almost perfect chair conforma-
tion [puckering parameters Q = 0.579(2) Å, h = 3.5(2) and / =
291(3)°, if the calculation starts from C-5 to C-10 and proceeds in
a counterclockwise direction]. Ring C also assumes an almost per-
fect chair conformation [puckering parameters Q = 0.5736(19) Å,
h = 6.54(18) and / = 258.3(16)° if the calculation starts from C-8
to C-14 and proceeds in a clockwise direction]. The A/B and B/C
rings junctions are trans. Ring D has an envelope on C-14 confor-
mation [puckering parameters q2 = 0.4653(19) Å, /2 = 208.2(2)°],
while ring E shows twisted O-1-exo, C-22-endo conformation
[puckering parameters q2 = 0.3709(18) Å, /2 = 197.7(3)°]. The tet-
rahydrofuran ring E is cis-fused to the cyclopentane ring D. In spite
the presence of the benzylidene group on C-23, the F ring assumes
Table 3
Crystal data and structure refinement for 2a, 2b and 3d.
2a
2b
3d
Empirical formula
Formula weight
Temperature (K)
Wavelength (Å)
Crystal system
Space group
Unit cell dimensions a (Å)
b (Å)
c (Å)
C
36H50O4
C36H50O4
546.76
130(2)
1.54184
Monoclinic
C 2
23.381(3)
7.2007(5)
C24H34O5
402.51
130(2)
546.79
130(2)
1.54184
Monoclinic
P 21
14.8327(4)
5.98150(10)
17.5061(5)
0.71073
Orthorhombic
P 212121
6.4021(2)
18.1058(6)
18.7844(5)
20.4260(19)
b (°)
103.233(3)
1511.93(6)
117.497(13)
3050.4(5)
Volume (Å3)
2177.40(11)
Z
2
4
4
Density (calculated) (Mg/m3)
Absorption coefficient (mmꢂ1
F(000)
1.201
0.593
596
1.191
0.588
1192
1.228
0.084
872
)
Crystal size (mm3)
Theta range for data collection (°)
Index ranges
0.33 ꢁ 0.23 ꢁ 0.19
0.36 ꢁ 0.19 ꢁ 0.065
0.36 ꢁ 0.23 ꢁ 0.05
3.53–68.10
3.81–68.10
3.44–26.06
ꢂ176h616, ꢂ56k67, ꢂ216l621
9563
ꢂ286h627, ꢂ86k64, ꢂ246l624
9702
ꢂ76h67, ꢂ226k619, ꢂ236l623
15880
Reflections collected
Independent reflections
Completeness to theta = 68.10° (%)
Absorption correction
Refinement method
4187 [R(int) = 0.0210]
99.9
3681 [R(int) = 0.0385]
100.0
4283 [R(int) = 0.0279]
99.7
Analytical
Analytical
Analytical
Full-matrix least-squares on F2
4187/7/370
Full-matrix least-squares on F2
3681/1/366
Full-matrix least-squares on F2
4283/0/276
Data/restraints/parameters
Goodness-of-fit on F2
1.036
1.044
1.057
Final R indices [I > 2
R indices (all data)
r
(I)]
R1 = 0.0364, wR2 = 0.0937
R1 = 0.0386, wR2 = 0.0958
0.27(19)
0.280 and ꢂ0.216
R1 = 0.0392, wR2 = 0.0956
R1 = 0.0470, wR2 = 0.1010
0.0(2)
0.168 and ꢂ0.236
R1 = 0.0398, wR2 = 0.0877
R1 = 0.0464, wR2 = 0.0915
0.1(10)
0.184 and ꢂ0.197
Absolute structure parameter
Largest diff. peak and hole (e Åꢂ3
)

K.M. Ruíz-Pérez et al. / Steroids 77 (2012) 819–828
823
R2
25
R2
27
21
17
26
O
O
R3
R3
R1
R1
18
23
20
22
24
11
23´
12
19
O
13
O
1
4
9
16
O
2
14
.
15
BF3 Et2O
10
8
7
+
only E-isomer
3
5
H
CH2Cl2
CH3COO
CH3COO
6
R
2
R
1
a) R= 5α-H, R1 = H,H; R2= H; R3= CH3 b) R= 5β-H, R1= H,H; R2= CH3; R3= H
c) No R Δ5; R1= H,H; R2= H; R3= CH3 d) R= 5α-H, R1= O; R2= H; R3= CH3
Scheme 2. BF3ꢀEt2O-catalyzed reaction of steroid sapogenins with benzaldehyde.
Fig. 1. 1H signals and selected and correlations of the side chain of compound 2a.
an almost perfect chair conformation [puckering parameters
Q = 0.5464(19) Å, h = 180.0(2) and / = 114(6)°; if the calculation
starts from O-2 to C-26 and proceeds in a counterclockwise direc-
tion]. Finally the benzylidene moiety attached to C-23 is essentially
planar (r.m.s. deviation = 0.0065 Å).
For 2b the presence of the b-equatorial acetoxy group attached
to C-3 does not disturb the ring A which has a well defined inverted
chair conformation [puckering parameters Q = 0.532(3) Å,
h = 175.7(3) and / = 171(4)°, if the calculation starts from C-1 to
C-10 and proceeds in a clockwise direction]. The acetoxy group is
arranged in such a way that the carbonylic oxygen nearly eclipses
Finally, as described for 2a, the benzylidene group introduced in
the side chain is essentially planar (r.m.s. deviation = 0.0039 Å).
The absolute configuration of the chiral centers of the steroid
framework in 2a and 2b are as those from the starting materials
1a and 1b. The most salient stereochemical features around the
modified side chain are: 2a (C-20: S, C-25: R) and 2b (C-20: S, C-
25: S). Fig. 2 shows the crystal structures of 2a and 2b with the
ellipsoids drawn at 30% of probability level [35,36].
4.2. Possible reaction mechanism
the axial hydrogen atom at the 3
a
-position. Ring B assumes an
The reaction can be explained through a pathway that com-
prises a nucleophilic attack of the enol ether II to benzaldehyde
producing the carbenium ion III. Considering that the resulting
intermediary III bears two new chiral centers at C-23 and C-230,
the exclusive occurrence of the product as the E-isomer may be ex-
plained by a pathway in which the loss of a proton from C-23 leads
to the enol ether IV that, after placing the phenyl group away from
the tetrahydofuran ring, undergoes (a) departure of the oxygen
attached to C-230, (b) migration of the C-22–C-23 double bond to
C-23–C-230 and (c) F ring closing, all in concerted mode (see
Scheme 3).
An alternative pathway in which the oxonium III undergoes F
ring closure to V followed by elimination of H-23 and the oxygen
attached to C-230 leading to a mixture of the E- and Z-isomers,
should be also considered. The fact that only the E-isomer was ob-
tained can be then rationalized on the basis of Z?E isomerization
that may occur through a pathway that involves acid-catalyzed F
almost perfect chair conformation [puckering parameters
Q = 0.558(3) Å, h = 0.0(3) and / = 311(32)°, if the calculation starts
from C-5 to C-10 and proceeds in a clockwise direction]. Ring C also
assumes an almost perfect chair conformation [puckering parame-
ters Q = 0.576(3) Å, h = 3.0(3) and / = 310(5)° if the calculation
starts from C-8 to C-14 and proceeds in a counterclockwise direc-
tion]. The A/B and B/C rings junctions are cis and trans, respectively.
Ring D has an envelope on C-14 conformation [puckering parame-
ters q2 = 0.469(3) Å, /2 = 209.1(4)°], while ring E shows twisted
O-1-exo, C-22-endo conformation [puckering parameters
q2 = 0.392(3) Å, /2 = 198.4(4)°]. The tetrahydrofuran ring E is cis-
fused to the cyclopentane ring D. The presence of the benzylidene
attached to C-23 does not disturb the F ring which assumes a very
similar to
a chair conformation [puckering parameters Q =
0.557(3) Å, h = 175.7(3) and / = 197(4)°; if the calculation starts
from O-2 to C-26 and proceeds in a counterclockwise direction].

824
K.M. Ruíz-Pérez et al. / Steroids 77 (2012) 819–828
Fig. 2. Crystal structures of 2a (upper) and 2b (lower).
ring opening to the oxonium Z-VI, which as the benzylic carbocat-
ion VII adopts a conformation with the phenyl group away from
the tetrahydrofuran ring to produce the oxonium E-VI that under-
goes F ring closing to afford the observed E-isomer (see Scheme 4).
Interestingly, the reaction time for the 25S-sapogenin 1b is con-
siderably shorter compared to those of the 25R-spirostans 1a, 1c,
and 1d (see Table 1). Provided that the reaction rate should depend
on the concentration of the reactive enol II (see Scheme 3), the ob-
served difference in reactivity may be an indication that in the 25S
series (1b), F ring opening leading to II is more favored because it
produces the release of the strain associated with the presence of
the axial methyl group attached to C-25.
The longer reaction needed to achieve acceptable yields of 2d
from 1d is hard to rationalize. Although it could be attributed to
the fact that coordination of BF3 to the carbonyl function at C-12
may decrease the amount of the Lewis acid available to produce
both the F ring opening and the coordination to the aldehyde nec-
essary for the reaction, the fact that even employing a large excess
of BF3ꢀEt2O no product of aldol reaction at C-11 was isolated dis-
cards this assumption.
O
BF3
O
BF3
BF3
H
O
O
23
O
O
BF3
*
23
.
*
BF3 Et2O
O
23´
H
O
H
I
II
III
- H+-23
O
BF3
O
c
O
BF3
a
O
O
b
E
H
IV
Scheme 3. Possible mechanism.

K.M. Ruíz-Pérez et al. / Steroids 77 (2012) 819–828
825
H
O
BF3
O
H
O
O
BF3
*
23
F-ring closing
O
*
23´
F3BO
H
V
III
OH
elimination
O
OH
23
23
23´
23´
O
O
- H+
O
+ H+
VIIa
Z-VI
Z (not observed)
rotation arround
C-23-C-23´ bond
+
OH
O
OH
23
23
- H+
O
23´
23´
O
O
+ H+
E
VIIb
E-VI
Scheme 4. Alternative mechanism including Z ? E isomerization.
After several unsuccessful attempts at trapping the cation VII by
addition of nucleophiles in different stages of the reaction, we
envisaged that the double bond placed between positions C-22
and C-23, in VII could be cleaved with and adequate oxidant lead-
ing to dinorcholanic lactones. Accordingly, we were pleased to find
that treatment of acetic acid solutions of the E-23(230)-benzyliden-
spirostanes 2a–d with CrO3 for 30 min produced moderate yields
of the corresponding dinorcholanic lactones 3a–d (see Scheme 5
and 6).
The NMR of the obtained dinorcholanic lactones are in good
agreement with the previously reported data for this kind of
compounds [26,41] (see Section 2 and Supplementary information
file). In addition, X-ray studies of the dinorcholanic lactone 3d
confirmed the obtained structure.
as described by Cremer and Pople [34]. As described for 2a and
2b the presence of the b-equatorial acetoxy group attached to C-
3 in 3d does not disturb the chair conformation of Ring A
[puckering parameters Q = 0.580(2) Å, h = 0.0(2) and / = 198(6)°,
if the calculation starts from C-1 to C-10 and proceeds in
counterclockwise direction]. Ring B assumes a very similar to chair
conformation [puckering parameters Q = 0.585(2) Å, h = 2.8(2) and
/ = 326(4)°, if the calculation starts from C-5 to C-10 and proceeds
in a counterclockwise direction]. Ring C shows a slightly flattened
chair conformation [puckering parameters Q = 0.565(2) Å,
h = 12.8(2) and / = 292.1(8)°], if the calculation starts from C-8 to
C-14 and proceeds in a clockwise direction]. While ring D has a
twisted conformation on C-13-C-14 [puckering parameters
q2 = 0.4352(17) Å, /2 = 198.1(2)°], the lactone ring E shows an
envelope conformation on C-17 [puckering parameters
q2 = 0.2197(18) Å, /2 = 79.4(4)°]. The A/B, B/C and C/D rings
junctions are trans, and the lactone ring E is cis-fused to the cyclo-
pentane ring D. Fig. 3 shows the crystal structure of 3b with the
ellipsoids drawn at 30% of probability level [35,36].
4.3. X-ray studies of dinorcholanic lactone 3d
Puckering parameters for compound 3d, which contains the 5
a-
dinorcholanic acid-(22 ? 16) lactone framework, were calculated
R2
25
27
21
17
21
O
26
R3
R1
R1
18
23
18
20
22
20
22
O
24
17
13
11
11
23´
12
12
O
O
19
19
13
1
4
1
4
9
16
9
16
2
2
14
14
15
15
10
8
7
10
8
7
CrO3
AcOH
3
3
5
5
CH3COO
CH3COO
6
6
R
R
2
3
a) R= 5α-H, R1 = H,H; R2= H; R3= CH3
b) R= 5β-H, R1= H,H; R2= CH3; R3= H
c) No R Δ 5; R1= H,H; R2= H; R3= CH3
d) R= 5α-H, R1= O; R2= H ; R3= CH3
a) R= 5α-H, R1 = H,H
b) R= 5β-H, R1= H,H
c) No R Δ5; R1= H,H
d) R= 5α-H, R1=O
Scheme 5. Synthesis of dinorcholanic lactones.

826
K.M. Ruíz-Pérez et al. / Steroids 77 (2012) 819–828
oxidation
HO
O
HO
O
AcOH
Z-VI
VIIb
O
O
CrO3
O
O
CrO3
not observed
HO
O
O
O
VIIIb
23-oxosapogenin
dinorcholanic lactone
O
O
or
CrO3
OAc
CrO3
O
COOH
over-oxidation
products
O
O
not observed
not observed
O
VIIIa
Scheme 6. Possible oxidative cleavage pathway.
Fig. 3. Crystal structure of dinorcholnic lactone 3d.
4.4. Possible reaction mechanism
O
O
The oxidative cleavage of the side chain can be explained in
terms of the acid catalyzed F ring opening to the intermediate cat-
ion Z-VI. Cleavage of the C-22–C-23 double bond in the resonance
form VIIa leads to the observed dinorcholanic lactones. The fact
that no 23-oxosapogenin due to cleavage of the benzylidene dou-
ble bond was isolated indicates that the presence of the benzyli-
dene moiety favors the fast F ring opening to the intermediate
COOH
O
CrO3/H2O
AcOH
O
Scheme 7. CrO3 oxidation of steroid sapogenins bearing an unchanged side chain.
a) X= NH seven steps [37]
X
O
F
incompatible with
O
E
O
b, c , d and e
E
O
F
O
b) X= O H NO3/Et2O [38]
D
D
hecogenin
c) X= O I 2/H2SO4/CH3COOOH [39]
E
O
C
D
d) peracids
X= O
[40-42]
[22]
B
BF3.Et2O/CH3COOH
O
A
e) [26]
AcO
incompatible
with b, c, and d
O
diosgenin
N
O
O
OH
O
NH2OH/CH3COONa
EtOH reflux.
Scheme 8. Synthetic methodologies for the preparation of dinorcholanic lactones [22,26,37–42].

K.M. Ruíz-Pérez et al. / Steroids 77 (2012) 819–828
827
cation Z-VI in which the benzylic resonance form VIIb undergoes
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