CA2188981C - Isomerisation of equilin - Google Patents

Abstract

The invention relates to a method of isomerisation of equilin or a derivative thereof to delta (8, 9)-dehydro estrone [delta (8, 9)DHE], characterized in that equilin or a derivative thereof is treated with a lithium salt of ethylenediamine or with lithium amide in dimethylsulfoxide.

Description

~I 8~i98~
ISOMERISATION OF EQUILIN
The invention relates to a method of isomerisation of equilin or derivatives thereof, more particularly to the isomerisation of equilin to 3-hydroxy-estra-1,3,5(10),8(9)-tetra-en-17-one Zo [delta (8, 9) -dehydro estrone~ delta (8, 9) DHE; delta 8 estrone: 8,9 dehydro estrone; CAS no. 61612-83-7].
The sulfate of the delta(8,9) derivative of equilin [delta (8, 9) DHES] is present in minor amounts i5 of about 3-4o in natural conjugated estrogen compositions, for instance in the commercially available product Premariri which is being used in hormone replacement therapy. It has been suggested in SCRIP no. 2049 (1995), p. 15 that minor amounts 20 of delta(8,9)DHES could have a significant contribution to the effect of conjugated estrogens.
It has further been suggested that delta(8,9)DHES, which has a relatively low affinity to the estrogen receptor, has a high functional activity, which may 2s play a role in the reported LDL-cholesterol-reducing properties and cardiovascular effects of conjugated estrogens, in particular of Premariri . Data reveal that delta(8,9)DHES contributes about 18% of Premarin's circulating estrogens. It is therefore of 3o importance to obtain an easy method of production for delta(8,9)DHE, which can easily be converted by methods known in the art to the sulfate of delta (8, 9) DHES.
r 35 Apart from cumbersome total synthesis, J.C.
Jacquesy et al., Chem. Abstr. 76 (1972), 154000f ' ~ _2 2 ~ 88981 disclosed isomerisation of equilin in hyperacidic media. Conversion to delta(8,9)DHE was achieved by using hydrogen fluoride or hydrogen fluoride/antimony fluoride at -30 °C. It is evident s that such dangerous reaction conditions are completely unsuitable and unacceptable for large scale production of delta(8,9)DHE. Moreover, in US
patent 5,395,831, wherein the method of Jacquesy is applied, it has been disclosed that said hydrogen Zo fluoride method does not provide pure delta (8, 9) DHE, but in addition thereto 10 0 of the unwanted delta(9,11)-isomer. Methods of production which are commercially acceptable, whether or not through isomerisation of equilin, thus have not been 15 disclosed.
The present invention offers the first easy and inexpensive method of production of delta(8,9)DHE, through isomerisation of equilin or a derivative ao thereof to said derivative, characterized in that equilin or a derivative thereof is treated with a lithium salt of ethylenediamine or with lithium amide in dimethylsulfoxide.
The general formula of equiline and said 25 derivatives is indicated in Formula I:
~3 I

wherein R1 is H, alkyl, acyl or silyl{alkyl)3~
R2 is H and R~~ is OH, 0-acyl, 0-alkyl or O-silyl(alkyl)3 or R3 is H and Rz is OH, 0-acyl, 0-alkyl 30 or 0-silyl{alkyl); or R2 and R~ together represent 0;
or R2 and R3 together represent acetal or cyclic :r acetal. R1 might also represent a substituted alkyl such as e.g. methoxy ethoxy methyl.
According to the isomerization method of the present invention derivatives of the general formula II can be prepared:
R
~3 wherein R1, RZ and R3 have the previously defined meaning.
According to one aspect of the present invention, there is provided a method of isomerisation of equilin or a derivative thereof according to the general formula I:
R
~3 I

wherein R1 is H, alkyl, acyl or silyl(alkyl)3;
Rz is H and R3 is OH, O-acyl, 0-alkyl or O-silyl(alkyl)3;
or R3 is H and Rz is OH, O-acyl, 0-alkyl or 0-silyl(alkyl)3 or R2 and R3 together represent 0;
or R2 and R3 together represent acetal or cyclic acetal, to delta(8,9)-dehydro estrone or a derivative thereof according to the general formula II:

-3a-R~
~3 II

wherein R1, RZ and R3 are as defined above, characterized in that equilin or the derivative thereof is treated with a lithium salt of ethylene-diamine or with lithium amide in dimethylsulfoxide.
In a preferred embodiment the isomerisation is performed using lithium salts of ethylenediamine, since this method results into the production of very pure delta(8,9)DHE. Such lithium salts can be prepared by treatment of ethylenediamine with lithium or with alkyllithium, preferably with methyllithium. (Co)solvents like tetrahydrofuran, dimethylsulfoxide, and the like may be added. Usually mixtures of delta(8,9)DHE and equilin or derivatives thereof are obtained when (co)solvents are added. Lithium amide in dimethylsulfoxide (DMSO) also provides mixtures of delta(8,9)DHE and equiline or derivatives thereof, which can be converted as such into their sulfates, to be used in the manufacture of pharmaceutical compositions containing conjugated estrogens.
The term alkyl, as used in the definition of formula I, means a branched or unbranched alkyl group having preferably 1-6 carbon atoms, like hexyl, isobutyl, tertiary butyl, propyl, isopropyl, ethyl, and, preferably, methyl.
The term acyl means an acyl group derived from an alkylcarboxylic acid, the alkyl moiety having the meaning given previously, or derived from formic acid. Acetals are derived from alcohols having preferably 1-6 carbon atoms.
s When C3-esters of equilin are used, common esters such as esters of aliphatic carboxylic acids with 1-6 carbon atoms or simple aromatic carboxylic acids are preferred. Examples are esters of formic acid, acetic acid, propionic acid, benzoic acid and the io like. Esters of acetic acid and benzoic acid are preferred. Under the reaction conditions applied, the esters are usually simultaneously saponified and the free delta(8,9)DHE or a mixture thereof with equilin is obtained. If necessary, the reaction is product can further be saponified using methods generally known in the art.
If in the compound of formula I, R1 is silyl(alkyl)3 the derivatives will also be hydrolyzed during the reaction resulting in compounds according zo to formula II wherein R1 is H. Similarly, if Rz or R3 is O-acyl or silyl(alkyl)3, compounds can be isolated wherein R2 or R3 is OH.
Preferably, the isomerisation is performed at a temperature of between about 0 and 90 °C, and with 2s more preference at about 30 °C if equiline or a derivative thereof is treated with a lithium salt of ethylenediamine or about 65 °C if equiline or a derivative thereof is treated with lithium amide in dimethylsulfoxide.
The conditions of isomerisation according to this invention are unobvious. The most straightforward method of isomerisation appears to be treatment of equilin under acidic conditions, for instance with 3s acetic acid, hydrochloric acid, trifluorosulfonic acid, boron trifluoroetherate, or combinations thereof in solvents such as methanol, ethanol, _5_ 21 X8981 tetrahydrofuran or toluene. However, under none of these conditions suitable results were obtained, since no reaction at all occurred or an intractable mixture of compounds was obtained.
Isomerisation under catalytic conditions (for instance palladium/carbon/benzyl alcohol) neither led to suitable results. The same holds for isomerisation under most of the alkaline conditions.
Obvious methods such as isomerisation with io butyllithium/potassium tert-butoxide, sodium amide, potassium tert-butoxide or sodium hydride in the usual solvents, lithium amide in dimethylformamide, sodium or potassium in ethylenediamine, and lithium in various amines, among which diisobutylamine, pentylamine, dimethylethylenediamine, piperazine, and piperidine, were virtually completely unsuccessful. Under the best conditions only 2 to 8%
of desired material was obtained in an intractable mixture of various isomers, unknown reaction 2o products, and starting material. Surprisingly only isomerisation using lithium in ethylenediamine, which afforded 950 of delta(8,9)DHE and lithium amide in DMSO, which afforded a mixture of about 550 of delta(8,9)DHE and 450 of starting material (equilin) which mixture can be used as such, appears to be successful. It is believed that these unique conditions provide a rare, if not the only possibility to obtain directly in a commercially available manner delta(8,9)DHE from equilin.
The following examples are illustrative for the invention and should in no way be interpreted as limiting the scope of the invention.
F

_6 X188981 Example 1 Lithium ( 13 g) was added portionwise to 920 ml of ethylenediamine under an atmosphere of nitrogen at s 95 °C and the mixture was stirred for 30 min at 100 °C. The reaction mixture was cooled to 23 °C, after which 100 g of equilin were added at a temperature of <_ 30 °C. The mixture was stirred for another 2 h at 30 °C. The suspension was poured into 2.5 1 of io ice water and at a temperature 5 25 °C acetic acid was added until pH 7. The aqueous layer was extracted three times with 2.5 1 of ethyl acetate.
The organic layer was washed with water, 5 g of active carbon (Norit~) were added and the suspension i5 was stirred at 21 °C for 30 min. The suspension was filtered over dicalite and the filtrate was evaporated under vacuo until a volume of about 500 ml. The suspension was stirred for 1 h at 0 °C, after which the crystalline material was filtered 20 off, washed with ethyl acetate and dried under vacuo at 40 °C, to obtain 81 g of delta(8,9)-dehydro estrone, having a purity of about 950.
The contents of delta(8,9)DHE and equiline were determined using 1H-NMR spectroscopy, characteristic 25 peaks of which are 0.90 ppm (C18) for delta(8,9)DHE
and 5.53 ppm (C7) and 0.79 ppm (C18) for equilin.
Example 2 so Lithium amide (5 g) was added to a mixture of 5 g of equilin in 150 ml of DMSO. The mixture was heated to 65 °C and stirred for 70 min. The reaction mixture was poured into 500 ml of water and acidified to pH 6.5 using 4N hydrochloric acid. The crystals were filtered off, washed with water and , dried under vacuo at 40 °C to obtain 5 g of a 4:5 mixture of equilin and delta(8,9)-dehydro estrone.
Example 3 A 6% solution of methyllithium-lithiumbromide complex in diethylether (23.5 ml) was added during approximately 15 minutes to 46 rtl ethylenediamine under an atmoshere of nitrogen at a temperature of to approximately 25°C. The temperature of the mixture was raised to 55°C and diethylether was distilled off. Subsequently the reaction mixture was stirred for 1 h at 55°C. The mixture was cooled to 20°C and 2.5 g of equilin was added. The mixture was stirred i5 for another 90 minutes at 30°C.
The suspension was poured into ice water and the mixture was extracted with ethyl acetate. After evaporation of the ethyl acetate extract until a volume of 20 ml was reached and cooling to 0°C, 2 g zo of crystalline delta 8-estrone was isolated.
Example 4 Lithium (1,1 g) was added portionwise to 80 ml of 2s ethylenediamine under an atmosphere of nitrogen at 100 °C and the mixture was stirred for 30 min at 100 °C. The reaction mixture was cooled to 23 °C , after which 4 g of 17(3-dihydroequilin were added at a temperature of <_ 30 °C. The mixture was stirred for 3o another 4 h at 30 °C. The suspension was poured into 250 ml of ice water and at a temperature of <_ 25 °C
acetic acid was added until pH 7. The suspension was cooled to 5 °C and the crystals were filtered off.
The crystals were suspended in 150 ml of water and _8 ~ 18881 100 ml of ethyl acetate were added. The layers were separated and the ethyl acetate solution was evaporated under vacuo until a volume of 20 ml. The suspension was stirred at -15 °C for 1 h, after s which the crystals were filtered off, washed with ethyl acetate and dried under vacuo at 40 °C, to obtain 2.5 g of 8,9-dehydro-17(3-estradiol, having a purity of > 95 0.
to Example 5 A solution of methyllithium-lithiumbromide (20 ml, 2.1 M) in diethylether was added in 10 min to 40 ml of ethylenediamine under an atmosphere of is nitrogen at 36 °C. The temperature of the mixture was raised to 55 °C and diethylether was distilled off. The mixture was stirred for 1 h at 55 °C. The reaction mixture was cooled to 3 °C , after which 2 g of equilin-3-methylether were added at a 2o temperature of <_ 10 °C. The mixture was stirred for another 2 h at 12 °C, after which 200 ml of ice water were added. To the mixture acetic acid was added until pH 8. The suspension was stirred for 1 h at 15 °C, after' which the crystals were filtered zs off, washed with water and dried under vacuo at 45 °C, to obtain 2.0 g of 8,9-dehydro-estrone-3-methylether, having a purity of approx. 80 0.
Example 6 According to the procedure described in example 5, 17(3-dihydroequilin 3,17-diacetate, was treated with methyllithium/ethylenediamine at 30 °C to give ~quantitavely 8,9-dehydro-17(3-estradiol having a 3s purity of approx. 90 0.

' -9- Z 18~9~ 1 Example 7 According to the procedure described in example 4, equilin-17-neopentylacetal was treated with lithium/ethylenediamine at 20 °C to give in a yield of 90 % 8,9-dehydro-estron-17-neopentylacetal having a purity of approx. 90 0.
to Example 8 According to the procedure described in example 4, 17~i-dihydroequilin-3,17-di(trimethylsilylether~
was treated with lithium/ethylenediamine to give i5 quantitavely 8,9-dehydro-17(3-estradiol having a purity of approx. 90 0.

Claims (7)

1. A method of isomerisation of equilin or a derivative thereof according to general formula I:
wherein R1 is H, alkyl, acyl or silyl(alkyl)3;
R2 is H and R3 is OH, O-acyl, O-alkyl or O-silyl(alkyl)3;
or R3 is H and R2 is OH, O-acyl, O-alkyl or O-silyl(alkyl)3 or R2 and R3 together represent O;
or R2 and R3 together represent acetal or cyclic acetal, to delta(8,9)-dehydro estrone or a derivative thereof according to general formula II:
wherein R1, R2 and R3 are as defined above, wherein equilin or a derivative thereof is treated with a lithium salt of ethylene-diamine or with lithium amide in dimethylsulfoxide.

2. The method according to claim 1, wherein R1 is acyl and R2 and R3 together represent O.

3. The method according to claim 1 or 2, wherein equiline or a derivative thereof is treated with a lithium salt of ethylenediamine.

4. The method according to claim 1 or 2, wherein equiline or a derivative thereof is treated with lithium amide in dimethylsulfoxide.

5. The method according to any one of claims 1-4, wherein the reaction temperature is between about 0 and 90°C.

6. The method according to claim 3, wherein the reaction temperature is about 30°C.

7. The method according to claim 4, wherein the reaction temperature is about 65°C.