Synthesis of (15E)-3β-hydroxyandrost-5-ene-15,17-dione 15-(O-carboxymethyl)oxime; The new hapten for 3β-hydroxyandrost-5-en-17-one(dehydroepiandrosterone, dhea)

Article abstract of DOI:10.1135/cccc19961647

From 17β-hydroxy-15β-[(4-methoxyphenyl)methoxy]androst-5-en-3β-yl acetate (1) by acetylation, removal of (4-methoxyphenyl)methyl group, and oxidation with Jones reagent, 15-oxoandrost-5-ene-3β,17β-diyl 3,17-diacetate (4) was prepared. (O-Carboxymethyl)hydroxylamine treatment and subsequent diazomethane methylation gave methyl ester of corresponding 15-(O-carboxymethyl)oxime derivative. Partial acid hydrolysis gave 3-acetate as a minor product, therefore the major 17-acetate was transformed in two steps into the 3-benzoate. Oxidation at position 17 and subsequent deprotection gave for both products final (15E)-3β-hydroxyandrost-5-ene-15,17-dione 15-(O-carboxymethyl)oxime (14), but for 3-acetyl derivative the whole synthesis is shorter and gave higher yield.

Full text of DOI:10.1135/cccc19961647

On Steroids
1647
SYNTHESIS OF (15E)-3β-HYDROXYANDROST-5-ENE-15,17-DIONE
15-(O-CARBOXYMETHYL)OXIME; THE NEW HAPTEN FOR
3β-HYDROXYANDROST-5-EN-17-ONE (DEHYDROEPIANDROSTERONE,
DHEA)*
1
2
Jan FAJKOS, Vladimir POUZAR and Ivan CERNY
Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic,
166 10 Prague 6, Czech Republic, e-mail: 1 pouzar@uochb.cas.cz, 2 cerny@uochb.cas.cz
Received October 15, 1996
Accepted November 12, 1996
From 17β-hydroxy-15β-[(4-methoxyphenyl)methoxy]androst-5-en-3β-yl acetate (1) by acetylation,
removal of (4-methoxyphenyl)methyl group, and oxidation with Jones reagent, 15-oxoandrost-5-ene-
3β,17β-diyl 3,17-diacetate (4) was prepared. (O-Carboxymethyl)hydroxylamine treatment and sub-
sequent diazomethane methylation gave methyl ester of corresponding 15-(O-carboxymethyl)oxime
derivative. Partial acid hydrolysis gave 3-acetate as a minor product, therefore the major 17-acetate
was transformed in two steps into the 3-benzoate. Oxidation at position 17 and subsequent deprotec-
tion gave for both products final (15E)-3β-hydroxyandrost-5-ene-15,17-dione 15-(O-carboxy-
methyl)oxime (14), but for 3-acetyl derivative the whole synthesis is shorter and gave higher yield.
Key words: Dehydroepiandrosterone; Hapten; Oxime.
Recently, we described the synthesis2 of 15-(O-carboxymethyl)oxime (CMO) deriva-
tive of testosterone, designed as hapten for testosterone immunoassays. Continuing this
study, we present now a synthesis of analogous 15-CMO derivative of dehydroepian-
drosterone (DHEA, 3β-hydroxyandrost-5-en-17-one). DHEA plays role in androgen
hormones metabolism and in last years its connection with various living processes in
human organism (obesity, aging, Alzheimer disease, cancer, AIDS, etc.) has been in-
tensively studied3.
As a starting compound we used 17β-hydroxy-15β-[(4-methoxyphenyl)methoxy]-
androst-5-en-3β-yl acetate (1), prepared previously4. Acetylation and subsequent selec-
tive removal5 of (4-methoxyphenyl)methyl (MPM) group gave derivative 3 with free
hydroxyl group at position 15, which was then oxidized into ketone giving 15-oxo
derivative 4. The key intermediate, oxime 5, was then prepared by the reaction of 4
with (O-carboxymethyl)hydroxylamine and by subsequent methylation with diazomethane.
* Part CCCLXXXVI in the series On Steroids; Part CCCLXXXV see ref.1
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Fajkos, Pouzar, Cerny:
1
The structure of 15-CMO derivative 5 was documented by the H NMR spectrum,
which besides two singlets of acetyl groups (δ 2.03 and 2.06) and signals of protected
CMO group (δ 3.74 s, and δ 4.55 and 4.58, AB system) contains resolved signals of
three C-ring protons. Two doublets of doublets at δ 2.95 (H-16α) and δ 2.53 (H-16β
were very similar, with geminal coupling constant amounting 18.9 Hz and about the
same coupling constants with H-17α (8.6 and 8.2 Hz), consequently H-17α was ob-
served as a triplet at δ 4.73. The configuration on the double bond C=N of CMO group
was assigned to 15E due to steric reasons.
2
OAc
OR
1
O
OR
AcO
AcO
4
1
2
R
R
H
MPM
1
2
3
MPM Ac
Ac
H
2
OR
O
N
N
1
2
1
OCH COOCH
2
3
R O
OCH COOR
2
R O
1
2
1
2
R
R
R
R
Ac Ac
Ac CH
Bz CH
5
6
7
8
9
11
12
13
14
3
3
3
H
Ac
H
Ac
H
H
H
CH
H
H
Bz Ac
Bz
H
10
MPM = (4-Methoxyphenyl)methyl; Ac = Acetyl; Bz = Benzoyl
For the further synthetic steps the derivative with protected hydroxyl group at posi-
tion 3 and with free one at position 17 was needed. Partial hydrolysis of diacetate 5 by
hydrochloric acid in methanol gave 33% of 17-acetate 6 and only 19% of desired 3-ace-
tate 7, besides 16% of diol 8 and 12% of unreacted starting diacetate 5. The structure
1
of reaction products was confirmed by H NMR spectroscopy. For distinguishing
among variously acylated compounds, protons H-3α and H-17α were suitable, resona-
ting at δ 3.52 and 4.73 for compound 6, at δ 4.60 and 3.81 for 7, and at δ 3.52 and 3.80
for diol 8, respectively.
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On Steroids
1649
The major monoacetylated product 6 was transformed into derivative suitable for
further synthesis by benzoylation at position 3 and subsequently by deacetylation of
resulted acetate benzoate 9. The deacetylation6 was carried out by hydrochloric acid in
methanol and gave 3-benzoate 10 in a 72% yield. Its 1H NMR spectrum contains multi-
plet of H-3α at δ 4.86 and triplet H-17α at δ 3.82.
Oxidation of 17β-hydroxyl group in derivatives 7 and 10 by Jones reagent afforded
ketones 11 and 12, respectively. In the case of acetyl derivative 11, the acetyl group
was hydrolyzed together with methyl ester by sodium carbonate in aqueous methanol to
give final CMO derivative 14. For the removal of benzoate protecting group in 12,
potassium hydroxide in aqueous methanol is necessary and methyl ester is split simul-
taneously as well. However, in this case the purification was more complicated due to
presence of benzoic acid. The raw product was firstly methylated by diazomethane and
then the resulting methyl ester 13 was purified by column chromatography. Mild hydro-
lysis with potassium hydrogen carbonate in aqueous methanol gave CMO derivative 14.
The overall yields of oxime 14 were 2.2% in the case of the synthesis via acetate 11
(7 steps) and 1.7% via benzoate 12 (10 steps). The structure of oxime 14 was confirmed
1
by IR and H NMR spectra. In the IR spectrum bands at 3 500–2 500 cm–1 (carboxylic
acid), 1 749 cm–1 (carboxylic acid, monomer), 1 725 cm–1 (ketone and carboxylic acid,
1
dimer), and 3 606 cm–1 (hydroxyl), were present. The H NMR spectrum displayed
characteristic signals of H-6 (δ 5.36 bd, J = 4.5 Hz), H-3α (δ 3.56 m, W = 32 Hz), and
of 15-CMO group (δ 4.65 and 4.62, AB-system). The immunological properties of this
new hapten will be reported separately.
EXPERIMENTAL
Melting points were determined on a Boetius micro melting point apparatus (Germany). Optical ro-
tations were measured at 25 °C on a Perkin–Elmer 141 MC polarimeter; [α]D values are given in
10–1 deg cm2 g–1. Infrared spectra (wavenumbers in cm–1) were recorded on a Bruker IFS 88 spectro-
1
meter in chloroform. H NMR spectra were taken on a Varian UNITY-200 (200 MHz) spectrometer at
23 °C in deuteriochloroform with tetramethylsilane as an internal standard. Chemical shifts are given
in ppm (δ-scale), coupling constants (J) and width of multiplets (W) in Hz. The purity of the pro-
ducts and reaction course were checked by thin-layer chromatography (TLC) performed on silica
gel G (ICN Biochemicals) developed in benzene–ether mixture (9 : 1 to 1 : 1) followed by spraying
with concentrated sulfuric acid and heating. Column chromatography was performed on silica gel
(60–120 µm, Service Laboratories, this Institute). Prior to evaporation on a rotary evaporator in
vacuo (bath temperature 50 °C), solutions in organic solvents were dried over magnesium sulfate.
Analytical samples were dried over phosphorus pentoxide at 40 °C/26 Pa for 12 h.
15β-[(4-Methoxyphenyl)methoxy]androst-5-ene-3β,17β-diyl 3,17-Diacetate (2)
The hydroxy derivative4 1 (5.0 g, 10.7 mmol) in pyridine (50 ml) was treated with acetic anhydride
(40 ml) and then allowed to stand for 18 h at room temperature. The excess reagent was decomposed
with ice and water and the product was extracted with ethyl acetate. The extract was sequentially
washed with 5% hydrochloric acid, water, 5% sodium hydrogen carbonate solution, dried and the
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Fajkos, Pouzar, Cerny:
solvent was removed in vacuo. The oily product was purified by column chromatography over silica
gel (300 g) in benzene–ether 3 : 1 to yield 4.6 g (84%) of the oily diacetate 2, [α]D –67° (c 1.7,
chloroform). IR spectrum: 1 727 (C=O); 1 252 (C–O, acetate); 1 033 (C–O). 1H NMR spectrum:
7.20 bd, 2 H, J ≈ 8 (H-2 and H-6 of 4-CH3OC6H4CH2O); 6.86 bd, 2 H, J ≈ 8 (H-3 and H-5 of
4-CH3OC6H4CH2O); 5.36 bd, 1 H, J ≈ 4.5 (H-6); 4.59 m, 1 H, W = 32 (H-3α); 4.56 t, 1 H, J = 8.5
(H-17α); 4.44 and 4.18 AB-system, 2 H, J(AB) = 11.4 (OCH2); 4.12 t, 1 H, J = 5.2 (H-15α); 3.80 s,
3 H (CH3O); 2.53, 1 H, W = 31 (H-16β); 2.06 s, 3 H (CH3COO); 2.04 s, 3 H (CH3COO); 1.04 s,
6 H (3 × H-19 and 3 × H-18). For C31H42O6 (510.7) calculated: 72.91% C, 8.29% H; found:
72.86% C, 8.16% H.
15β-Hydroxyandrost-5-ene-3β,17β-diyl 3,17-Diacetate (3)
The derivative 2 (4.5 g, 8.8 mmol) was dissolved in dichloromethane (350 ml) and water (3 ml) was
added. Then 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (11.0 g, 58 mmol) was added in one portion
under vigorous stirring. After 20 min the reaction mixture was diluted with chloroform, washed with
5% solution of potassium hydrogen carbonate, water, dried, and the solid residue was chromato-
graphed on a silica gel column (280 g) in benzene–ether (20 : 1). Crystallization from methanol afforded
2.76 g (80%) of the hydroxy derivative 3, m.p. 229–230 °C, [α]D –82° (c 1.9, chloroform). IR spectrum:
1
3 619, 3 500 (O–H); 1727 (C=O); 1 254 (C–O, acetate); 1 034 (C–O). H NMR spectrum: 5.39 bd,
1 H, J ≈ 4.5 (H-6); 4.60 m, 1 H, W = 32 (H-3α); 4.53 t, 1 H, J = 8.7 (H-17α); 4.27 m, 1 H, W = 19
(H-15α); 2.69, 1 H, W = 31 (H-16β); 2.06 s, 3 H (CH3COO); 2.03 s, 3 H (CH3COO); 1.06 s, 6 H (3 × H-19
and 3 × H-18). For C23H34O5 (390.5) calculated: 70.74% C, 8.78% H; found: 70.62% C, 8.63% H.
15-Oxoandrost-5-ene-3β,17β-diyl 3,17-Diacetate (4)
A solution of hydroxy derivative 3 (1.2 g, 3.1 mmol) in acetone (120 ml) was treated with excess
Jones reagent. After 5 min at room temperature the excess reagent was destroyed with methanol,
water was added (50 ml), and acetone was distilled off under reduced pressure. The crystalline pro-
duct was collected by suction, dissolved in ethyl acetate, and the solution was washed with water,
dried, and the solvent was distilled off in vacuo. The residue was crystallized from methanol to yield
970 mg (81%) of the ketone 4, m.p. 184–186 °C, [α]D –36° (2.6, chloroform). IR spectrum: 1 734
(C=O); 1 251 (C–O, acetate). 1H NMR spectrum: 5.39 bd, 1 H, J ≈ 4.5 (H-6); 4.96 t, 1 H, J = 8.4
(H-17α); 4.60 m, 1 H, W = 32 (H-3α); 2.09 s, 3 H (CH3COO); 2.03 s, 3 H (CH3COO); 1.03 s, 3 H
(3 × H-19); 0.89 s, 3 H (3 × H-18). For C23H32O5 (388.5) calculated: 71.11% C, 8.30% H; found:
70.92% C, 8.19% H.
(15E)-15-Oxoandrost-5-ene-3β,17β-diyl 3,17-Diacetate 15-(O-Carboxymethyl)oxime
Methyl Ester (5)
The ketone 4 (8.5 g, 21.8 mmol) in pyridine (80 ml) was treated with (O-carboxymethyl)hydroxyl-
amine hemihydrochloride (6.0 g, 65.2 mmol) and the mixture was allowed to stand for 18 h at room
temperature. After pouring on ice containing concentrated hydrochloric acid (115 ml), the product
was extracted with ethyl acetate. The extract was washed with a saturated sodium chloride solution
to neutrality, dried, and the solvent was removed in vacuo. The residue was dissolved in methanol
(50 ml) and ether (200 ml) and treated with excess diazomethane in ethereal solution. The excess
diazomethane was removed with acetic acid, the reaction mixture was diluted with ethyl acetate, and
the solution was washed with 5% sodium hydrogen carbonate. The oily residue after evaporation of
the solvents was purified by column chromatography over silica gel in benzene–ether (5 : 1) to yield
7.2 g (69%) of the oily oxime 5, [α]D –44° (c 1.6, chloroform). IR spectrum: 1 754 (C=O,
Collect. Czech. Chem. Commun. (Vol. 61) (1996)

On Steroids
1651
COOCH3); 1 730 (C=O, acetate); 1 252 (C–O, acetate); 1 068, 1 047, 1 034 (C–O). 1H NMR spec-
trum: 5.37 bd, 1 H, J ≈ 4.5 (H-6); 4.73 t, 1 H, J = 8.4 (H-17α); 4.60 m, 1 H, W = 32 (H-3α); 4.58
and 4.55 AB-system, 2 H, J(AB) = 16.3 (OCH2COO); 3.74 s, 3 H (COOCH3); 2.95 dd, 1 H, J =
18.9, J′ = 8.6 (H-16α); 2.53 dd, 1 H, J = 18.9, J′ = 8.2 (H-16β); 2.06 s, 3 H (CH3COO); 2.03 s, 3 H
(CH3COO); 1.04 s, 3 H (3 × H-19); 0.84 s, 3 H (3 × H-18). For C26H37NO7 (475.6) calculated:
65.66% C, 7.84% H, 2.95% N; found: 65.49% C, 7.76% H, 2.92% N.
(15E)-3β-Hydroxy-15-oxoandrost-5-en-17β-yl 17-Acetate 15-(O-Carboxymethyl)oxime
Methyl Ester (6)
The diacetate 5 (10.5 g, 22 mmol) in methanol (200 ml) was treated with 1% methanolic hydro-
chloric acid (400 ml) and the mixture was allowed to stand at 18 °C for 3 h. After neutralization with
5% sodium hydrogen carbonate solution, methanol was distilled off under reduced pressure. The pro-
duct was taken into ethyl acetate, the solution was washed with water, dried and the solvent was
removed in vacuo. The residue consisted of four products: The lipophilic starting diacetate 5, the
3-hydroxy derivative 6, the 17-hydroxy derivative 7 and the diol 8. It was chromatographed on silica
gel (500 g) in benzene–ether (19 : 1). Fractions with the most lipophilic component afforded 1.3 g
(12%) of the starting diacetate 5. Further elution with the same solvent mixture yielded fractions with
the monoacetate 6. Crystallization from methanol gave 3.1 g (33%) of 6, m.p. 64 °C, [α]D –33° (c 1.7,
chloroform). IR spectrum: 3 609 (O–H); 1 754 (C=O, COOCH3); 1 733 (C=O, acetate); 1 254 (C–O,
acetate); 1 095, 1 062 (C–O). 1H NMR spectrum: 5.34 bd, 1 H, J ≈ 4.5 (H-6); 4.73 t, 1 H, J = 8.5
(H-17α); 4.58 and 4.54 AB-system, 2 H, J(AB) = 16.4 (OCH2COO); 3.74 s, 3 H (COOCH3); 3.52 m,
1 H, W = 32 (H-3α); 2.94 dd, 1 H, J = 18.9, J′ = 8.5 (H-16α); 2.53 dd, 1 H, J = 18.9, J′ = 8.2
(H-16β); 2.06 s, 3 H (CH3COO); 1.03 s, 3 H (3 × H-19); 0.84 s, 3 H (3 × H-18). For C24H35NO6
(433.5) calculated: 66.49% C, 8.14% H, 3.23% N; found: 66.38% C, 8.19% H, 3.14% N.
(15E)-17β-Hydroxy-15-oxoandrost-5-en-3β-yl 3-Acetate 15-(O-Carboxymethyl)oxime
Methyl Ester (7)
Continuing the chromatography from the foregoing experiment by the elution with the same solvent
mixture yielded fractions containing the hydroxy derivative 7. Evaporation left 1.8 g (19%) of an oil,
[α]D –68° (c 1.6, chloroform). IR spectrum: 3 612, 3 501 (O–H); 1 758 (C=O, COOCH3); 1 727
1
(C=O, acetate); 1 255 (C–O, acetate); 1 031 (O–H). H NMR spectrum: 5.37 bd, 1 H, J ≈ 4.5 (H-6);
4.60 m, 1 H, W = 32 (H-3α); 4.59 s, 2 H (OCH2COO); 3.81 t, 1 H, J = 8.7 (H-17α); 3.74 s, 3 H
(COOCH3); 2.83 dd, 1 H, J = 18.9, J′ = 8.6 (H-16α); 2.46 dd, 1 H, J = 18.9, J′ = 8.6 (H-16β); 2.03 s,
3 H (CH3COO); 1.05 s, 3 H (3 × H-19); 0.80 s, 3 H (3 × H-18). For C24H35NO6 (433.5) calculated:
66.49% C, 8.14% H, 3.23% N; found: 66.27% C, 8.10% H, 3.05% N.
(15E)-3β,17β-Dihydroxyandrost-5-en-15-one 15-(O-Carboxymethyl)oxime Methyl Ester (8)
Further elution (see above) with benzene–ether (3 : 1) yielded fractions with the diol 8. Crystal-
lization from methanol–water (7 : 3) gave 1.4 g (16%) of 8, m.p. 98–99 °C, [α]D –59° (c 1.3, chloro-
form). IR spectrum: 3 610, 3 466 (C–O); 1 756 (C=O); 1 092, 1 069 (C–O). 1H NMR spectrum:
5.34 bd, 1 H, J ≈ 4.5 (H-6); 4.56 s, 2 H (OCH2COO); 3.80 bt, 1 H, J ≈ 8.5 (H-17α); 3.74 s, 3 H
(COOCH3); 3.52 m, 1 H, W = 32 (H-3α); 2.82 dd, 1 H, J = 18.9, J′ = 8.5 (H-16α); 2.46 dd, 1 H, J = 18.9,
J′ = 8.7 (H-16β); 1.03 s, 3 H (3 × H-19); 0.80 s, 3 H (3 × H-18). For C22H33NO5 (391.5) calculated:
67.49% C, 8.50% H, 3.58% N; found: 67.31% C, 8.29% H, 3.57% N.
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Fajkos, Pouzar, Cerny:
(15E)-15-Oxoandrost-5-ene-3β,17β-diyl 17-Acetate 3-Benzoate 15-(O-Carboxymethyl)oxime
Methyl Ester (9)
The hydroxy derivative 6 (1.15 g, 2.67 mmol) was dissolved in pyridine (10 ml) and treated with
benzoyl chloride (1.5 ml, 12.9 mmol). After 18 h at room temperature, the excess reagent was de-
composed with ice and water, and the crystalline product was collected by suction. The product was
dissolved in ethyl acetate, the solution was washed with 5% hydrochloric acid, water, 5% sodium
hydrogen carbonate, dried, and the solvent was removed in vacuo. Crystallization from a chloroform–
methanol mixture afforded 1.15 g (71%) of the benzoate 9, m.p. 175–176 °C [α]D –16° (c 1.9, chlo-
roform). IR spectrum: 1 756 (C=O, COOCH3); 1 733 (C=O, acetate); 1 713 (C=O, benzoate); 1 276
1
(C–O, benzoate); 1 250 (C–O, acetate), 1 034 (C–O). H NMR spectrum: 8.04 m, 2 H (H-2 and H-6
of C6H5COO); 7.50 m, 3 H (H-3, H-4 and H-5 of C6H5COO); 5.42 bd, 1 H, J ≈ 4.5 (H-6); 4.86 m,
1 H, W = 32 (H-3α); 4.74 t, 1 H, J = 8.4 (H-17α); 4.59 and 4.54 AB-system, 2 H, J(AB) = 16.5
(OCH2COO); 3.75 s, 3 H (COOCH3); 2.95 dd, 1 H, J = 18.8, J′ = 8.7 (H-16α); 2.55 dd, 1 H, J = 18.8,
J′ = 7.8 (H-16β); 2.07 s, 3 H (CH3COO); 1.09 s, 3 H (3 × H-19); 0.86 s, 3 H (3 × H-18). For
C31H39NO7 (537.7) calculated: 69.25% C, 7.31% H, 2.61% N; found: 69.17% C, 7.14% H, 2.56% N.
(15E)-17β-Hydroxy-15-oxoandrost-5-en-3β-yl 3-Benzoate 15-(O-Carboxymethyl)oxime
Methyl Ester (10)
The acetate 9 (1.3 g, 2.4 mmol) was dissolved in chloroform (10 ml) and treated with 1% methanolic
hydrochloric acid (10 ml). After 18 h at 45 °C, the reaction mixture was neutralized with 5% sodium
hydrogen carbonate solution, solvents were distilled off under reduced pressure, and the product was
extracted with ethyl acetate. The solution was washed with water, dried, and the solvent was
removed. The residue gave on crystallization from ethanol 850 mg (72%) of the monoester 10, m.p.
164 °C, [α]D –35° (c 1.4, chloroform). IR spectrum: 3 612 (O–H); 1 756 (C=O, COOCH3); 1 711
(C=O, benzoate); 1 277 (C–O, benzoate); 1 257 (C–O, COOCH3); 1 095, 1 025 (C–O). 1H NMR
spectrum: 8.03 m, 2 H (H-2 and H-6 of C6H5COO); 7.49 m, 3 H (H-3, H-4 and H-5 of C6H5COO);
5.42 bd, 1 H, J ≈ 4.5 (H-6); 4.86 m, 1 H, W = 32 (H-3α); 4.56 s, 2 H (OCH2COO); 3.82 bt, 1 H, J ≈ 9
(H-17α); 3.74 s, 3 H (COOCH3); 2.84 dd, 1 H, J = 18.6, J′ = 8.6 (H-16α); 2.48 dd, 1 H, J = 18.6,
J′ = 9.5 (H-16β); 1.09 s, 3 H (3 × H-19); 0.81 s, 3 H (3 × H-18). For C29H37NO6 (495.6) calculated:
70.28% C, 7.52% H, 2.83% N; found: 70.16% C, 7.44% H, 2.76% N.
(15E)-15,17-Dioxoandrost-5-en-3β-yl 3-Acetate 15-(O-Carboxymethyl)oxime Methyl Ester (11)
The hydroxy derivative 7 (1.85 g, 4.3 mmol) in acetone (40 ml) was treated with excess Jones rea-
gent. After 10 min at room temperature, methanol (3 ml) was added to destroy the oxidizing agent,
the reaction mixture was diluted with water (15 ml), and the solvents were distilled off under reduced
pressure. The crystalline product was collected by suction, dissolved in ethyl acetate, and the solution
was washed with water, dried, and the solvent was distilled off. Crystallization from methanol gave
1.42 g (77%) of the ketone 11, m.p. 169–170 °C, [α]D –53° (c 2.1, chloroform). IR spectrum: 1 751
1
(C=O, COOCH3, ketone); 1 728 (C=O, acetate); 1 254 (C–O, acetate); 1 102, 1 031 (C–O). H NMR
spectrum: 5.39 bd, 1 H, J ≈ 4.5 (H-6); 4.64 and 4.58 AB-system, 2 H, J(AB) = 16.2 (OCH2COO);
4.60 m, 1 H, W = 32 (H-3α); 3.75 s, 3 H (COOCH3); 3.59 d, 1 H, J = 22.3 (H-16α); 2.78 d, 1 H, J = 22.3
(H-16β); 2.03 s, 3 H (CH3COO); 1.06 s, 3 H (3 × H-19); 0.98 s, 3 H (3 × H-18). For C24H33NO6
(431.5) calculated: 66.80% C, 7.71% H, 3.25% N; found: 66.63% C, 7.56% H, 3.17% N.
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1653
(15E)-15,17-Dioxoandrost-5-en-3β-yl 3-Benzoate 15-(O-Carboxymethyl)oxime Methyl Ester (12)
The hydroxy derivative 10 (1.5 g, 3 mmol) was oxidized with Jones reagent in acetone (40 ml) as
described in the previous experiment. Similar work-up and crystallization from ethanol afforded 1.03 g
(68%) of the ketone 12, m.p. 187–188 °C, [α]D –17° (c 1.9, chloroform). IR spectrum: 1 752 (C=O,
COOCH3); 1 711 (C=O, benzoate and ketone); 1 277 (C–O, benzoate); 1 099, 1 026 (C–O).
1H NMR spectrum: 8.04 m, 2 H (H-2 and H-6 of C6H5COO); 7.49 m, 3 H (H-3, H-4 and H-5 of
C6H5COO); 5.43 bd, 1 H, J ≈ 4.5 (H-6); 4.86 m, 1 H, W = 32 (H-3α); 4.65 and 4.59 AB-system,
2 H, J(AB) = 16.5 (OCH2COO); 3.75 s, 3 H (COOCH3); 3.61 d, 1 H, J = 22.3 (H-16α); 2.80 d, 1 H,
J = 22.3 (H-16β); 1.11 s, 3 H (3 × H-19); 1.00 s, 3 H (3 × H-18). For C29H35NO6 (493.6) calculated:
70.57% C, 7.15% H, 2.84% N; found: 70.44% C, 7.02% H, 2.80% N.
(15E)-3β-Hydroxyandrost-5-ene-15,17-dione 15-(O-Carboxymethyl)oxime Methyl Ester (13)
The benzoate 12 (2.3 g, 4.7 mmol) in methanol (40 ml) was treated with 5% potassium hydroxide
solution in 80% methanol (20 ml) and set aside for 18 h. The alkali was removed with 2% hydro-
chloric acid and methanol was distilled off in vacuo. The residue was extracted with ethyl acetate,
the extract was washed with cold saturated sodium chloride solution to neutrality, dried, and the sol-
vent was distilled off. To remove the benzoic acid the residue was dissolved in methanol (10 ml) and
treated with excess diazomethane in ether. After 10 min the excess diazomethane was destroyed with
acetic acid, the reaction mixture was diluted with ether, washed with 5% sodium hydrogen carbonate
solution, and solvents were removed. The residue was chromatographed over silica gel (100 g) in
benzene. After elution of methyl benzoate the product was eluted with benzene–ether (20 : 1). The
yield of the oily ester 13 was 1.28 g (70%), [α]D –39° (c 1.2, chloroform). IR spectrum: 3 608, 3 489
1
(O–H); 1 752 (C=O, COOCH3); 1 706 (C=O, ketone); 1 104, 1 045 (C–O). H NMR spectrum: 5.37 bd,
1 H, J ≈ 4.5 (H-6); 4.64 and 4.58 AB-system, 2 H, J(AB) = 16.4 (OCH2COO); 3.75 s, 3 H
(COOCH3); 3.60 d, 1 H, J = 22.4 (H-16α); 3.54 m, 1 H, W = 32 (H-3α); 2.79 d, 1 H, J = 22.4
(H-16β); 1.05 s, 3 H (3 × H-19); 0.99 s, 3 H (3 × H-18). For C22H31NO5 (389.5) calculated:
67.84% C, 8.02% H, 3.60% N; found: 67.74% C, 7.88% H, 3.55% N.
(15E)-3β-Hydroxyandrost-5-ene-15,17-dione 15-(O-Carboxymethyl)oxime (14)
A) The acetate methyl ester 11 (1.2 g, 2.7 mmol) in methanol (70 ml) was treated with a solution
of sodium carbonate (350 mg, 3.3 mmol) in water (15 ml) and allowed to stand under nitrogen for
12 h at room temperature. Methanol was distilled off under reduced pressure and the residue was
acidified with 2% hydrochloric acid. The product was extracted with ethyl acetate, the extract was
washed with a cold saturated sodium chloride solution to neutrality, dried, and the solvent was
removed in vacuo. The yield of the oily oxime 14 was 420 mg (40%), [α]D –36° (c 1.0, chloroform).
IR spectrum: 3 606 (O–H); 3 500–2 500 (O–H, acid); 1 749 (C=O, acid monomer); 1 725 (C=O,
1
ketone and acid dimer); 1 045 (C–O). H NMR spectrum: 5.36 bd, 1 H, J ≈ 4.5 (H-6); 4.65 and 4.62
AB-system, 2 H, J(AB) = 17.1 (OCH2COO); 3.58 d, 1 H, J = 22.3 (H-16α); 3.56 m, 1 H, W = 32
(H-3α); 2.80 d, 1 H, J = 22.3 (H-16β); 1.05 s, 3 H (3 × H-19); 0.98 s, 3 H (3 × H-18). For
C21H29NO5 (375.5) calculated: 67.18% C, 7.79% H, 3.73% N; found: 67.06% C, 7.61% H, 3.65% N.
B) A solution of the methyl ester 13 (420 mg, 1.1 mmol) in methanol (40 ml) was treated with a
solution of potassium hydrogen carbonate (500 mg) in water (10 ml) and allowed to stand for 7 days at
50 °C. Methanol was removed in vacuo, the residue was acidified with 2% hydrochloric acid, and the
product was extracted with ethyl acetate. The extract was washed with a cold saturated sodium
chloride solution to neutrality, dried, and the solvent was removed. The yield of the oily oxime 14 was
230 mg (57%) with properties identical in all respects with the product prepared by procedure A).
Collect. Czech. Chem. Commun. (Vol. 61) (1996)

1654
Fajkos, Pouzar, Cerny:
The authors are indebted Dr L. Bednarova for taking and interpretation of IR spectra and to Mrs M.
Snopkova for measurements of the 1H NMR spectra. The elemental analyses were carried out in the
Analytical Laboratory (Dr V. Pechanec, Head). This work was supported in part from Grant No.
203/96/0329 of the Grant Agency of the Czech Republic.
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Collect. Czech. Chem. Commun. (Vol. 61) (1996)