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Scheme 1. Synthesis of 5␣-estrane-3,17-diol.
ice bath. After 18 h heating at 80 ◦C, the mixture was cooled at room
temperature and washed with hydrochloric acid (1 N, 4× 100 mL),
saturated aqueous sodium hydrogencarbonate solution (3×
100 mL), dried with brine (100 mL) and anhydrous sodium sulfate.
The brown gum obtained was purified by chromatography on a
silica gel column (5 × 30 cm, elution: hexane/diethyl ether, 90:10
to 70:30, v/v) to give 2 as a white solid (2.097 g, 84.4%).
MS (EI), m-TMS derivative, m/z (%): 450 (100), 435 (5), 313 (5),
194 (15), 182 (10), 133 (5), 105 (30), 91 (5), 77 (10). Melting point:
173.0 ◦C. TLC: Rf = 0.42 (hexane/ethyl acetate, 2:1, revelation: UV
254 nm). 1H NMR ppm: 8.01 (d, J = 5 Hz, 2H), 7.54 (t, J = 5 Hz, 1H),
7.43 (t, J = 5 Hz, 2H), 5.83 (s, 1H), 5.08 (d, J = 3.65 Hz, 1H), 0.88 (s,
3H), complex multiplets between 0.8 and 2.6 ppm. 13C NMR ppm:
199.82, 166.46, 166.01, 132.81, 130.70, 129.46, 128.35, 124.59,
82.24, 49.26, 49.22, 45.14, 40.69, 35.54, 30.08, 25.95, 24.52, 16.69.
known to improve the yield in alcohol inversion of 17-steroids [8].
Furthermore, diphenyl-(2-pyridyl)-phosphine was used instead of
the usual triphenylphosphine in order to improve the removal
of unreacted phosphine and phosphine oxide by simple washing
with dilute hydrochloric acid [9]. Other by-products and unreacted
nandrolone were removed by flash chromatography. Several exper-
iments showed that chromatographic elution with hexane/diethyl
ether or hexane/ethyl acetate mixture was equivalent. This proce-
dure afforded 2 with 84.4% yield.
Compound 2 was identified by GC/MS and after saponification,
the resultant 17␣-hydroxy-4-estren-3-one (epinandrolone) pre-
sented a single peak with a mass spectrum identical to that of
nandrolone and a retention time down-shifted of 0.41 min. This
compound is known to be a minor metabolite of nandrolone [3].
The second and last step of our procedure is a Birch reduc-
tion of the benzoate 2. This procedure was chosen because Birch
reduction of nandrolone is known to give stereoselectively 5␣-
estrane-3,17-diol, an undesired stereoisomer of our targeted
compound [10]. Benzoate ester saponification prior to the Birch
reduction was found to be unnecessary. Direct application of the
Birch procedure to 2 gave 5␣-estrane-3,17␣-diol as the sole iso-
mer. Purification by column chromatography gave the desired
2.2. 5˛-Estrane-3ˇ,17˛-diol (3)
A solution of 2 (930 mg, 2.45 mmol) in dioxane/diethyl ether
(1:1, 40 mL) was added to a solution of lithium (800 mg, 115 mmol)
in ammonia (150 mL). After 30 min ethanol (30 mL) was added
dropwise over a period of 20 min, ammonia was allowed to evapo-
rate and water (200 mL) was added. This mixture was extracted
with diethyl ether (200 mL) and ethyl acetate (200 mL). These
organic fractions were combined, dried with brine (100 mL),
sodium sulfate and evaporated under vacuum to give a light yellow
solid. This solid was purified by chromatography on a silica gel col-
umn (3 × 12 cm, dichloromethane/ethanol, 100:1, v/v) to give 3 as
a white solid (466.4 mg, 68.2%). Recrystallization from boiling ethyl
acetate gave 352 mg of white crystals (not optimized).
MS (EI), bisTMS derivative, m/z (%): 422 (3), 407 (22), 332 (65),
317 (5), 291 (4), 242 (68), 227 (13), 214 (6), 201 (32), 185 (15), 159
(7), 147 (12), 145 (12), 129 (100), 116 (19), 105 (11), 93 (15), 91
(15), 89 (13), 79 (13), 75 (63). Melting point: 204.7 ◦C. TLC: Rf = 0.30
(hexane/ethyl acetate, 1:1, revelation: phosphomolybdic acid). 1H
NMR ppm: 3.73 (d, J = 5 Hz, 1H), 3.57 (m, 1H), 0.67 (s, 3H), complex
multiplets between 0.5 and 2.3 ppm. 13C NMR ppm: 80.10, 70.55,
47.86, 47.82, 45.40, 43.48, 41.57, 41.14, 35.86, 33.62, 32.32, 31.38,
28.49, 25.53, 24.41, 17.08.
Identity and purity of this compound was established by GC/MS
data comparison with a certified reference standard. All com-
parative analytical data were identical (retention time and mass
spectrum). 1H NMR data were consistent with previously published
data [6].
In summary, a rapid, simple and stereoselective synthesis of
5␣-estrane-3,17␣-diol is described, providing horseracing labo-
ratories with an essential reference material for their antidoping
performance.
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3. Results and discussion
5␣-Estrane-3,17␣-diol was identified in post administration
urines after horse loading with nandrolone [1,2]. The absolute
configuration of the two secondary alcohols was unambiguously
confirmed by X-ray crystallography of synthetic 5␣-estrane-
to give a mixture of 3␣- and 3 isomers of estrene-5(10)-3,17␣-
diol (the yield and 3␣/3 ratio was not reported). After a tedious
chromatographic separation, the appropriate isomer was treated
with hydrogen gas in the presence of palladium catalyst to afford
5␣-estrane-3,17␣-diol [6].
Our synthesis starts from readily available and inexpensive nan-
drolone 1. The first step consists in the inversion of the 17 alcohol
into its 17␣ isomer via a classic Mitsunobu reaction [7]. In our
hands, the standard procedure using THF as a solvent failed to
give adequate yields. THF was substituted by toluene because it is
[9] Camp D, Jenkins ID. The use of a phosphine containing a basic group in the
Mitsunobu reaction. Aust J Chem 1988;41:1835–9.
[10] Bowers A, Ringold HJ, Denot E, Steroids CI. 19-Nor-dihydrotestosterone deriva-
tives. J Amer Chem Soc 1958;80:6115–8.