C(10)-C(19) bond cleavage reaction of 19-oxygenated androst-4-ene-3,6-dione steroids under various conditions

Article abstract of DOI:10.1248/cpb.52.983

C(10)-C(19) bond cleavage reaction of 19-hydroxy- and 19-oxoandrost-4-ene- 3,6,17-triones (5, 6) was explored under various conditions. Treatment of steroids 5 and 6 with KOH in MeOH gave the A-ring aromatized product 6-oxoestrone (11) in a fair yield, respectively, in contrast, the treatment with a weak base yielded 4-methyl steroid 17 (20%) in the case of 19-alcohol 5 or 19-nor- Δ⁵⁽¹⁰⁾-steroid 9 (12-67%) along with compound 11 (6-27%) in the case of 19-aldehyde 6. Reaction of compound 6 with HCl in MeOH produced 3-methyl ethers of 6-oxoestrone and Δ⁶-estrone, compounds 12 and 14 (ca. 20% each). Thus, 6-oxosteroids 5 and 6 showed unique C(10)-C(19) bond cleavage reactions with a base or acid.

Full text of DOI:10.1248/cpb.52.983

August 2004
Notes
Chem. Pharm. Bull. 52(8) 983—985 (2004)
983
C(10)–C(19) Bond Cleavage Reaction of 19-Oxygenated Androst-4-ene-
3,6-dione Steroids under Various Conditions
Masao NAGAOKA and Mitsuteru NUMAZAWA*
Tohoku Pharmaceutical University; 4–4–1 Komatsushima, Aoba-ku, Sendai 981–8558, Japan.
Received February 16, 2004; accepted May 21, 2004
C(10)–C(19) bond cleavage reaction of 19-hydroxy- and 19-oxoandrost-4-ene-3,6,17-triones (5, 6) was ex-
plored under various conditions. Treatment of steroids 5 and 6 with KOH in MeOH gave the A-ring aromatized
product 6-oxoestrone (11) in a fair yield, respectively, in contrast, the treatment with a weak base yielded
4-methyl steroid 17 (20%) in the case of 19-alcohol 5 or 19-nor-D5(10)-steroid 9 (12—67%) along with compound
11 (6—27%) in the case of 19-aldehyde 6. Reaction of compound 6 with HCl in MeOH produced 3-methyl ethers
of 6-oxoestrone and D6-estrone, compounds 12 and 14 (ca. 20% each). Thus, 6-oxosteroids 5 and 6 showed unique
C(10)–C(19) bond cleavage reactions with a base or acid.
Key words aromatase inhibitor; 6-oxoandrostenedione; 19-oxygenated steroid; C(10)–C(19) bond cleavage
The androgen androst-4-ene-3,17-dione (androstenedione, group at C-6, compound 2, with the strong base, the aroma-
1) is converted into the estrogen estrone (10) through its 19- tized product is not formed but a 19-nor derivative, 4-ene9) or
hydroxy and 19-oxo derivatives (2, 3) by action of the en- 5(10)-ene steroid 7,10) is only produced, indicating that the C-
zyme aromatase (Fig. 1).1) 6-Oxoandrostenedione (4) that is 6 carbonyl group plays a critical role in the base-catalyzed
clinically used for treatment of estrogen-dependent breast aromatization reaction. Treatment of the 19-ol 2 with
cancer, is one of the earliest discovered suicide substrates of NaHCO3 yielded neither the 19-nor steroid 7 nor the aroma-
aromatase.2—4) The mechanism for aromatase inactivation by tized product 10.
inhibitor 4 involves the two initial hydroxylations at the C-19
The bond cleavage reaction of 19-al 6 was next studied
position, producing the 19-oxo derivative 6 through the 19- under various conditions (Table 1). Treatment of compound 6
hydroxy intermediate 5, followed by an epoxidation of the with a weak base, NaHCO3 or CH3COSK, at room tempera-
4,5-double bond.5,6) In this sequence, a part of the 19-oxo in- ture gave 19-nor-D5(10)-compound 9 as the major product (56
termediate is aromatized to yield 6-oxoestrone (11).7) On the or 67%) as well as estrogen 11 (13 or 6%). In the reaction
other hand, we previously have reported that treatment of the
19-oxygenated steroids with a nucleophile, a thiol com-
pound, gives the corresponding 1,4-Michael adduct, a 4a-
thio analog, respectively, where 5(10)-ene steroid 9, the
C(10)–C(19) bond cleavage reaction product, is also obtained
from the 19-oxo steroid 6 in a low yield.8) The 1,4-addition
reaction does not occur in the reaction with corresponding 6-
deoxy steroids 2 and 3.
Taken together, we were of interest to know effect of intro-
ducing a carbonyl group at C-6 of the 19-oxygenated steroids
2 and 3 on the C(10)–C(19) bond cleavage reaction in rela-
tion to the biological aromatization. This paper describes re-
actions of 19-oxygenated 6-oxo steroids 5 and 6 under basic
and acidic conditions.
Results and Discussion
We initially examined the C(10)–C(19) bond cleavage of
19-hydroxy-6-oxo steroid 5 under a basic condition. Treat-
ment of 19-ol 5 with a strong base, KOH, in MeOH at room
temperature yielded the aromatized product 6-oxoestrone
(11) in 65% yield (Fig. 2). In contrast, the treatment with a
weak base, NaHCO3, in MeOH under reflux produced 4-
methyl-6-oxoestrone (17) in 20% yield along with the recov-
ered substrate. Although there is no evidence, the production
mechanism of compound 17 is thought as follows: migration
of the 10b-hydroxy methyl group to the 4b-position, yielding
4b-hydroxymethyl-5(10)-ene-3,6-dione intermediate (15),
followed by dehydration and a sequential isomerization of
the C-4 exocyclicmethylene double bond introduced may
give the aromatized product 17 (Fig. 2). It is known that in
the reaction of a 19-hydroxy steroid having no carbonyl Fig. 1. Structures of Steroids
* To whom correspondence should be addressed. e-mail: numazawa@tohoku-pharm.ac.jp
© 2004 Pharmaceutical Society of Japan

984
Vol. 52, No. 8
Fig. 3. Aromatization Reaction of D5(10) Steroid 9 Under a Basic or Acidic
Condition
45%) and 11 (27 or 13%) in each case. Treatment of the
D
5(10)-steroid 9 with NaOH in MeOH at room temperature af-
forded the aromatized product 11 in a good yield, suggesting
that the aromatization reaction would proceed, at least in
part, through the D5(10)-steroid. Based on the result, it is pre-
sumed that the conversion of 19-ol 5 into the aromatized
product 11 by treatment with KOH in MeOH, described
above, may proceed through 19-al 6 produced by an air-oxi-
dation of 19-ol 5 followed by the C(10)–C(19) bond cleav-
age. It has previously been reported that treatment of the 6-
deoxy derivative 3 with pyridine produces the D5(10)-deriva-
tive 7 but not the aromatized product 10.9) The 6-oxo func-
tion would be essential for the aromatization reaction of a
5(10)-en-3-one steroid.
Reaction of 19-al 6 with HCl in MeOH at room tempera-
ture gave 3-methyl ether of the aromatized product 11, com-
pound 12, as well as D6-estrone 3-methyl ether (14) in low
yields. When D5(10)-steroid 9 was treated with HCl or p-
TsOH, the aromatized product 11 was obtained in 16% or
20% yield, but the production of compounds 2 and 14 was
not detected by TLC analysis of the reaction products in each
case. These results indicate that the D5(10)-steroid 9 is aroma-
tized to produce compound 11 under not only a basic condi-
tion but also an acidic condition and this is not an intermedi-
ate in a sequence of the HCl-catalyzed production of com-
pounds 12 and 14 from 19-al 6. 6-Oxoestrone (11) was not
converted into the 3-methyl ether 12 or the D6-estrone analog
14 by the treatment with HCl in MeOH, indicating that the
production of compound 12 or 14 does not proceed through
the 6-oxo steroid 11.
It has previously been reported that the acid-catalyzed
C(10)–C(19) bond cleavage reactions of steroidal compounds
having a 1,4-dien-3-one or 2,3-dihydroxy-4-ene structure
produce the corresponding aromatized products via the
dienone-phenol and related rearrangements.16,17) Since 19-
oxygenated compounds 5 and 6 have no such structural fea-
ture, a 1,4-dien-3-one or 2,3-dihydroxy-4-ene group, re-
quired for the dienone-phenol type rearrangements, it is
likely that their aromatization reactions observed in this
study would not involve the rearrangements.
Fig. 2. Reaction of 19-Hydroxy Steroid 5 with a Base
Table 1. C(10)–C(19) Bond Cleavage of 19-Oxo Steroid 6 Under Various
Conditions
Product, %
Entry
Conditions
Aromatized,
11, 12, or 14
D
5(10) 9
Hemiacetal
13
1
2
3
4
5
6
NaHCO3–MeOH-r.t.a)
Pyridine–H2O-reflux
MeOH–H2O-reflux
CH3COSK–MeOH-r.t.
NaOH–MeOH-r.t.
HCl–MeOH-r.t.
23 (11)
37 (11)
18 (11)
6 (11)
93 (11)
56
22
55
67
—
—
—
—
—
8
—
—
18 (12), 17 (14)
a) r.t.: room temperature.
with CH3COSK, 19-hemiacetal 13 (8%) was produced in ad-
dition to the two products. The 19-hydrogen atom of com-
pound 13 was observed as a singlet peak at 4.38 ppm in the
1H-NMR spectrum. This along with the 13C-NMR spectrum
revealed that compound 13 is not a mixture of two stereoiso-
mers at the C-19 position. Semiemprical molecular orbital
PM3 calculations indicate that the 19-carbonyl group of
steroid 6 favors the over-A-ring conformation among the
possible three.11) This suggests that the nucleophile methox-
ide anion would approach the carbonyl carbon from the less
hindered over-A-ring side rather than the crowded over-C-
ring side, thereby giving streoselectively (19S)-19-hemiacetal
13, as seen in the NaBH4 reaction of 19-al 3.12,13) In this reac-
tion, the 1,4-addition product 4a-acetylthio ether was not
isolated, although the reaction with benzylmercaptan gives
the addition product 4a-benzylthio ether as well as com-
pound 9.8) When the 19-al 6 was treated with aqueous pyri-
dine or MeOH under reflux gave the two compounds 9 (12 or
The present findings indicate that an introduction of the C-
6 carbonyl group into 19-hydroxy and 19-oxo steroids 2 and
3 accelerates and/or gives rise to C(10)–C(19) bond cleavage
reactions under acidic and basic conditions, although the
exact mechanisms for the cleavage reactions are not clear.
Experimental
Melting points were measured on a Yanagimoto melting point apparatus
(Kyoto, Japan) and are uncorrected. IR spectra were recorded in KBr pellets
for the solid products or in neat forms for the oily products on a Perkin-
Elmer FT-IR 1725X spectrophotometer (Norwalk, CT, U.S.A.), and UV
spectra were determined in 95% ethanol on a Hitachi 150-20 UV spec-
trophotometer (Tokyo, Japan). 1H- and 13C-NMR spectra were obtained in

August 2004
985
CDCl3 solution with a JEOL GX 400 spectrometer (400 MHz for 1H and
100.5 MHz for 13C) (Tokyo, Japan) using tetramethylsilane as an internal
standard. High-resolution mass spectra (HR-MS) were determined with a
JEOL JMS-DX 303 spectrometer. Thin-layer chromatography (TLC) was
performed on E. Merck precoated TLC silica gel plates (silica gel 60F-254,
layer thickness 0.25 and 0.5 mm for analytical and preparation use, respec-
tively; Darmstadt, Germany). Column chromatography was conducted with
silica gel 60, 70—230 mesh (E. Merck, 70—230 mesh).
19-Hydroxyandrost-4-ene-3,6,17-trione (5) and androst-4-ene-3,6,17,19-
tetraone (6) were synthesized according to the previously reported method.6)
Reaction of 19-Hydroxy Steroid 5 with KOH or NaHCO3. (A) Reac-
tion with KOH A solution of KOH (2.7 g, 49 mmol) in 5 ml of water was
added to a solution of compound 5 (50 mg, 0.16 mmol) in 30 ml of MeOH
and the mixture was stirred at room temperature for 3.5 h. After this time,
the reaction mixture was neutralized with conc. HCl, then, condensed to a
small volume and extracted with AcOEt (50 mlꢀ2). The organic layer was
washed with saturated NaCl solution and dried (Na2SO4). After evaporation
of the solvent, the residue obtained was subjected to preparative TLC
(hexane–AcOEt; 1 : 1). The crude product was recrystallized from acetone to
give 6-oxoestrone 11 (29 mg, 65%). mp 246—248 °C (lit.15) mp 256—
257 °C). 1H-NMR d 0.93 (3H, s, 18-Me), 7.10 (1H, dd, Jꢁ8.6, 2.6 Hz, 2-H),
7.33 (1H, d, Jꢁ8.6 Hz, 1-H), 7.57 (1H, d, Jꢁ2.6 Hz, 4-H).
time, the reaction mixture was diluted with AcOEt, washed with 5%
NaHCO3 solution, NaCl solution, and dried (Na2SO4). After evaporation of
the solvent, the residue obtained was subjected to preparative TLC
(hexane–AcOEt; 1 : 1) to give two products. The less polar product was re-
crystallized from acetone to give 3-methoxyestra-1,3,5(10),6-tetraen-17-one
(14) (7.8 mg, 17%). mp 117—120 °C. 1H-NMR d 0.92 (3H, s, 18-Me), 3.80
(3H, s, 3-OMe), 6.07 (1H, dd, Jꢁ9.7, 1.6 Hz, 7-H), 6.52 (1H, dd, Jꢁ9.7,
2.3 Hz, 6-H), 6.67 (1H, d, Jꢁ2.8 Hz, 4-H), 6.75 (1H, dd, Jꢁ8.4, 2.8 Hz, 2-
H), 7.17 (1H, d, Jꢁ8.4 Hz, 1-H). 13C-NMR d 13.6, 21.5, 23.8, 31.1, 35.7,
38.2, 42.1, 48.5, 48.8, 55.3, 111.9, 112.1, 124.3, 128.6, 130.9, 131.0, 135.3,
158.3, 220.3. UV lmax (e); 226 (27000), 262 (6200) nm. IR (KBr) nmax 1736
and 1601 cmꢂ1. HR-MS Found 282.16180, Calcd for C19H22O2 282.16178.
The more polar product was recrystallized from acetone to give 3-
methoxyestra-1,3,5(10)-triene-6,17-dione (12) (8.4 mg, 18%). mp 134—
137 °C. 1H-NMR d 0.93 (3H, s, 18-Me), 3.85 (3H, s, 3-OMe), 7.13 (1H, dd,
Jꢁ8.4, 2.9 Hz, 2-H), 7.35 (1H, d, Jꢁ8.4 Hz, 1-H), 7.58 (1H, d, Jꢁ2.9 Hz, 4-
H). 13C-NMR d 13.7, 21.3, 25.2, 31.2, 35.7, 39.6, 43.0, 43.2, 47.7, 50.2,
55.5, 109.8, 121.7, 126.6, 133.3, 138.9, 158.3, 197.3, 219.6. UV lmax (e):
222 (20000), 255 (8100), 322 (2900) nm. IR (KBr) nmax 1741 and
1672 cmꢂ1. HR-MS Found; 298.15690 Calcd for C19H22O3: 298.15590.
Reaction of D5(10)-Steroid 9. (A) A Basic Condition To a solution of 9
(50 mg, 0.175 mmol) in 2.5 ml of MeOH was added 1 mol/l NaOH solution
(0.17 ml, 0.174 mmol) and the reaction mixture was stirred at room tempera-
ture for 5 h. After the same workup as described above, compound 11
(25 mg, 50%) was obtained.
(B) Reaction with NaHCO3
A mixture compound 5 (50 mg,
0.16 mmol), NaHCO3 (787 mg, 9.37 mmol) and MeOH (30 ml) was heated
under reflux for 5 h. After this time, the reaction mixture was poured into
water and extracted with AcOEt. The organic layer was washed with water
and dried (Na2SO4). After evaporation of the solvent, the residue obtained
was separated by preparative TLC (hexane–AcOEt; 1 : 1). The crude product
was recrystallized from acetone to give compound 17 (9 mg, 20%). mp
167—171 °C (decomp.). 1H-NMR d 0.92 (3H, s, 18-Me), 2.53 (3H, s, 4-
Me), 6.99 (1H, d, Jꢁ8.4 Hz, 2-H), 7.17 (1H, d, Jꢁ8.4 Hz, 1-H). 13C-NMR d
13.2, 13.6, 21.4, 25.5, 31.2, 35.8, 38.5, 43.0, 45.0, 47.6, 50.6, 119.5, 123.2,
126.0, 132.7, 139.8, 152.9, 199.6, 219.8. UV lmax (e); 260 (6800) and 330
(2400) nm. IR nmax; 3400, 1728 and 1676 cmꢂ1. HR-MS Found: 298.1567;
Calcd for C19H22O3: 298.1565. The starting material 5 was also recovered in
20% yield.
(B) An Acidic Condition 1) Reaction with p-TsOH: p-TsOH monohy-
drate (21 mg, 0.11 mmol) was added to a solution of compound 9 (50 mg,
0.175 mmol) in 2.5 ml of acetone and the reaction mixture was stirred at
room temperature for 2 d. After the same workup as described above, com-
pound 11 (10 mg, 20%) was obtained. 2) Reaction with HCl: HCl gas was
bubbled for 1 min in a solution of compound 9 (50 mg, 0.175 mmol) in 15 ml
of MeOH, and the reaction mixture was allowed to stand at room tempera-
ture for 2 d. After the same workup as described above, compound 11 (8 mg,
16%) was obtained but the production of the other aromatized products 12
and 14 was not detected by the TLC analysis.
References
Reaction of 19-Oxo Steroid 6 under Various Conditions. (A) A Basic
Condition Compound 6 (50 mg, 0.16 mmol) in MeOH (15 ml) or pyridine
(7.5 ml) was treated with a base for an appropriate time. The reaction mix-
ture was diluted with AcOEt, washed with saturated NaCl solution and dried
(Na2SO4). After evaporation of the solvent, the product was purified by
preparative TLC (hexane–AcOEt; 1 : 1). 1) Reaction with NaOH: 1 mol/l
NaOH solution (1.57 ml) was added to a solution of compound 6 in MeOH
and the mixture was stirred at room temperature for 2 h. 6-Oxoestrone (11)
(43 mg, 93%). 1H-NMR d 0.93 (3H, s, 18-Me), 7.10 (1H, dd, Jꢁ8.6, 2.6 Hz,
2-H), 7.33 (1H, d, Jꢁ8.6 Hz, 1-H), 7.60 (1H, d, Jꢁ2.6 Hz, 4-H). 2) Reaction
with NaHCO3: 13.2 mg of NaHCO3 (0.16 mmol) was added to a solution of
compound 6 in MeOH and the reaction mixture was stirred at room temper-
ature for 3.75 h. Compound 11 (5.6 mg, 13%). Estr-5(10)-ene-3,6,17-trione
(9) (25 mg, 56%), mp 166—168 °C (from acetone) (lit.14) mp 167—169 °C).
1H-NMR d 0.93 (3H, s, 18-Me), 2.98 and 3.25 (1H each, d, Jꢁ21.1 Hz, 4-
H). 3) Reaction with CH3COSK: CH3COSK (22.5 mg, 0.197 mmol) was
added to a solution of compound 6 and the mixture was stirred at room tem-
perature for 3 h. Compound 11 (2.6 mg, 6%), mp 235—237 °C (from ace-
1) For a review see: Covey D. F., “Steroid Biosynthesis Inhibitors: Phar-
maceutical and Agrochemical Aspects,” ed. by Berg D., Plemel M.,
Ellis Horwood Ltd., Chichester, 1988, pp. 534—571.
2) Schwarzel W. C., Kruggel W., Brodie H. J., Endocrinology, 92, 866—
880 (1973).
3) Covey D. F., Hood W. F., Endocrinology, 108, 1597—1599 (1981).
4) Numazawa M., Tsuji M., Mutsumi A., J. Steroid Biochem., 28, 337—
344 (1987).
5) Numazawa M., Midzuhasi K., Nagaoka M., Biochem. Pharmacol., 47,
717—726 (1994).
6) Numazawa M., Mutsumi A., Tachibana M., Biochem. Pharmacol., 52,
1253—1259 (1996).
7) Numazawa M., Sugiyama T., Nagaoka M., Biol. Pharm. Bull., 21,
289—292 (1998).
8) Numazawa M., Tachibana M., Steroids, 58, 423—428 (1993).
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85 (1960).
10) Barber G. W., Ehrenstein M., J. Org. Chem., 20, 1253—1259 (1955).
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722—726 (2004).
tone) and compound
9 (30.6 mg, 67%), mp 166—168 °C. (19-S)19-
Methoxy-19-hydroxyandrost-4-ene-3,6,17-trione (13) (4.3 mg, 8%); mp
98—101 °C. 1H-NMR d 0.95 (3H, s, 18-Me), 3.81 (3H, s, 19-OMe), 4.38
(1H, s, 19-CH), 6.08 (1H, d, Jꢁ0.7 Hz, 4-H). 13C-NMR d 13.8, 21.6, 21.8,
29.8, 31.4, 32.8, 35.1, 35.7, 38.1, 47.6, 50.2, 50.9, 52.8, 53.3, 71.7, 129.8,
157.8, 174.2, 198.8, 219.9. UV lmax (e): 237 (7400) nm. IR (KBr) nmax; 3391,
1727 and 1684 cmꢂ1. HR-MS; 346.1826; Calcd for C20H26O5: 346.1872. 4)
Reaction with MeOH or pyridine: 2.5 ml of water was added to a solution of
compound 6 in MeOH or pyridine and the mixture was heated under reflux
for 9 h, giving compound 11 (12 mg, 27% or 6.0 mg, 13%) and compound 9
(5.5 mg, 12% or 20.5 mg, 45%).
14) Numazawa M., Oshibe M., Matsuzaki H., Steroids, 58, 423—428
(1993).
15) Yoshizawa I., Kamiya E., Itoh S., Yakugaku Zasshi, 101, 285—290
(1981).
16) Kirk P. N., Hartshorn M. P., “Steroid Reaction Mechanisms,” Chap. 5,
Elsevier Publishing Company, Amsterdam, 1968, pp. 277—290.
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25, 2343—2349 (1977).
(B) An Acidic Condition HCl gas was bubbled for 1 min in a solution
of compound 6 (50 mg, 0.16 mmol) in 15 ml of MeOH, and the resulting re-
action mixture was allowed to stand at room temperature for 2 d. After this