CLIAMS:1) A process for preparation of testosterone (1) comprising:

reducing 4-androstene-3,17-dione (2) with sodium borohydride in micellar system.
2) The process according to claim 1, wherein the micellar system is obtained from water, organic solvent and anionic surfactant.
3) The process according to claim 2, wherein the organic solvent is selected from hydrocarbon such as toluene, n-hexane or n-heptane and ethers such as diethyl ether, di-isopropyl ether or 2-methyl tetrahydrofuran.
4) The process according to claim 3, wherein the organic solvent is toluene.
5) The process according to claim 2, wherein the quantity of water to 4-androstene-3,17-dione (2) is 3-12 times (volume/weight).
6) The process according to claim 5, wherein the quantity of water to 4-androstene-3,17-dione (2) is 5-10 times (volume/weight).
7) The process according to claim 4, wherein the quantity of toluene to 4-androstene-3,17-dione (2) is 3-12 times (volume/weight).
8) The process according to claim 7, wherein the quantity of toluene to 4-androstene-3,17-dione (2) is 5-10 times (volume/weight).
9) The process according to claim 2, wherein the anionic surfactant is selected from sodium lauryl sulfate, potassium lauryl sulfate, dioctyl sodium sulfosuccinate, sodium dodecyl sulfate and sodium laureth sulfate.
10) The process according to claim 9, wherein the anionic surfactant is sodium lauryl sulfate.
11) The process according to claim 2, wherein the ratio of aninoic surfactant to 4-androstene-3,17-dione (2) is in the range of about 5-40% (weight/weight).
12) The process according to claim 11, wherein the ratio of aninoic surfactant to androstene-3,17-dione (2) is 10% (weight/weight).
13) The process according to claim 1, wherein the ratio of sodium borohydride to androstene-3,17-dione (2) is in the range of about 0.5 to 4 mole equivalent.
14) The process according to claim 13, wherein mole ratio of sodium borohydride is 1 mole equivalent.
15) The process according to claim 1, wherein the reaction is carried out at temperature range of about 10-50 °C.
16) The process according to claim 15, wherein the reaction is carried out at temperature range of about 10-25°C. ,TagSPECI:FIELD OF THE INVENTION
The present invention relates to a novel process for preparation of testosterone.

BACKGROUND OF THE INVENTION
(8R,9S,10R,13S,14S,17S)-17-hydroxy-10,13-dimethyl-1,2,6,7,8,9,11,12,14,15,16,17-dodecahydrocyclopenta[a]phenanthren-3-one i.e. Testosterone [1, CAS No. 58-22-0] is a steroid hormone that belongs to androgen group and is found in all vertebrates. In mammals, testosterone is primarily secreted in testicles of male, ovaries of female and small amount in adrenal gland. Testosterone plays a key role in development of reproductive tissues and secondary sexual characteristics as well as prevention of osteoporosis.

Numbers of synthetic methods were reported in the literature for preparation of compound 1. These are divided into following categories and summarized below.

I. Protection of carbonyl function in ring A of compound 2 through appropriate reagent either through enol-ether or ketal, followed by reduction of 17-ketone:
US patent 2,294,433, US patent 2,679,502, GB patent 803178, US patent 2,588,294 and US patent 2,802,825 describe reduction by using reducing agents such as sodium metal, sodium borohydride, lithium borohydride, lithium aluminium hydride, dimethyl aluminium hydride or dimethyl aluminium then deprotection (Scheme-1).

Scheme-1
II. Selective reduction of compound 2 to compound 1:
a) GB patent 789974 describes selective reduction of compound 2 to compound 1 using sodium borohydride in methanol or tetrahydrofuran (THF).

b) The publication J.Fajkos, Coll. Czech. Chem. Commun., 1959, 24, 2284 describes selective reduction of compound 2 with lithium tri-(tert-butoxy) aluminium hydride in THF at 0°C, however, the yield for desired product was very low. Moreover, lithium tri-(tert-butoxy) aluminium hydride is pyrophoric in nature.

c) The publications Ward et al, Tetrahedron Letters, 1988, 29, 517; Norymberski & Woods J. Chem. Soc., 1955, 3426 and DD patent 298641, DD patent 294842 describe that selectivity toward compound 1 could be increased to approximately 70%, when reduction of compound 2 was carried out in methanol at -78 0C. However, formation of other over reduced products was not ruled out, thus, additional purification was necessary.

d) US patent 2,356,596 describes the preparation of testosterone by cathodic reduction of compound 2 in 5% sodium acetate solutions at 55 to 60 oC; generally such electrolytic reduction process has its own disadvantages rendering it not attractive for industrial operation.
e) D'incan et al, Tetrahedron, 1982, 38, 1755 reports selective reduction of compound 2 with various metal borohydrides and are summarized in Table 1. It is to be noted that almost in all the cases, selectivity towards the desired product i.e. compound 1 was very poor.
III. Selective oxidation of the hydroxyl group present on ring A of compound 3:
US patent 3,118,881 discloses a process for selective oxidation of compound 3 in 90% with manganese dioxide, however only with freshly prepared and activated manganese dioxide.

Thus it is evident form prior art that there is need for novel, cost effective, green and industrial feasible process for synthesis of compound 1. This invention provides it.
SUMMARY OF THE INVENTION
The present invention provides a novel process for synthesis of testosterone (1) in micellar aggregates as shown in Scheme-2.

Scheme-2

BRIEF DESCRIPTION OF THE DRAWING
Figure -1: Reduction of ketone using sodium borohydride in micellar aggregates.

Table 1: Reduction of compound 2 with various borohydrides under different reaction conditions
Reducing agent Solvent/Reaction Temp Reaction Time (hours) % yield
(Compound 6) (Compound 7) (Compound 8) (Compound 1) (Compound 3)
LiBH4 Diethyl ether/Reflux 2 - - - - 100
Zn (BH4)2 Diethyl ether/ Reflux 1 - - - 40 40
NaBH4 Ethanol/200C 0.25 - - - 20 -
NaBH4 Ethanol/200C 2 - - - 70 30
NaBH4 THF/600C 24 40 30 30 - -
nBu4NBH4 THF/600C 24 10 27 63 - -
nBu4NBH4 + 2.5 eq.TMEDA THF/600C 24 - 30 70 - -
nBu4NBH4 Methanol/200C 2 - - - 90 10
TMEDA: Tetramethylethylenediamine

DETAILED DESCRIPTION OF THE INVENTION
The publications Usha & Sunil, Ind. Eng. Chem. Res. 2007, 46, 1923-1927; Mariano et al J.Org.Chem, 2004, 69, 8224-8230; Mariano et al J.Org.Chem, 2004, 69, 8231-8238 describe reduction of carbonyl compounds to corresponding hydroxyl in micellar aggregates with sodium borohydride.
The publications Yongmin and Peipei, Tetrahedron Asymmetry, 1996, 7, 3055-3058; Debapratim et al, Organic Letters, 2004, 22, 4133-4136 and Sangita et al, Langmuir 2005, 21, 10398-10404; describe stereo-selective reduction of ketone in presence of chiral surfactant, however, enantiomeric excess for corresponding hydroxyl compound was never significant.
The publications Mariano et al J.Org.Chem, 2004, 69, 8224-8230 and Mariano et al J.Org.Chem, 2004, 69, 8231-8238 describe that reduction of carbonyl function to corresponding hydroxyl compound in micellar aggregates takes place at the interface i.e. pseudo phase. It was also mentioned that, sodium borohydride was insoluble in organic solvent (hydrocarbon) and ketone substrate has very poor solubility in water. Moreover, anion BH4- has the same charge of the anionic surfactant. Considering all these aspects, reaction can only occur at the interface of the aggregates i.e. in pseudo phase (Figure 1).
Although, regio-selective reduction employing of borohydride reduction in micellar aggregates was reported, however, regio-selectivity was very poor.
The publication Usha & Sunil, Ind. Eng. Chem. Res. 2007, 46, 1923-1927 describe reduction of isophorone in reverse micellar aggregates with sodium borohydride (one mole equivalent with respect to isophorone), gave mixture of 1,2 and 1,4 reduced product as shown in Scheme 3.
Scheme 3
The publication Nikles and Sukenik Tetrahedron Letters 23, 1982, 4211-4214 also reports the reduction of enone such as 3-methyl-2-cyclohexene-one, cyclopropylcarbinyl enone, carvone and 4-phenyl3-butene-2-one in micellar aggregates. Further it states that in micellar aggregates 1, 4 reduction was more preferred over 1, 2 reduction (Scheme 4). In the above reactions 1,2-reduction product is allylic alcohol while rest of two products are 1,4-products.
Scheme 4
It could be summarized that overall regio-selectivity in micellar aggregates reduction was very poor.
In our patent application PCT/IB2013/056805 and IN/1097/MUM/2014, it had been demonstrated high degree of chiral induction in sodium borohydride reduction in micellar aggregates.
Surprisingly present inventors found that reduction of compound 2 with sodium borohydride in achiral micellar systems gave high degree of regio-selectivity as well as stereo-selectivity for obtaining compound 1.
One could rationalize that regio-selectivity was due to preferable orientation of molecule in pseudo phase. As mentioned hereinbefore, that reduction occurs at interface i.e. pseudo phase and probably orientation of molecule is such that ring A may not be getting exposed to pseudo phase, which resulted in high degree of regio-selectivity.
In the present case it has been also presumed that compound 2 as referred in Figure 1 adopts a conformation through interaction in pseudo phase so that borohydride anion could predominately approach carbonyl group only from one direction. This results in predominately formation of one isomer. In short, the transition state of the resulting isomer has considerable lower energy due to some interaction in the pseudo phase.

In an embodiment of the present invention provides a novel process for preparation of testosterone (1) comprising: reducing 4-androstene-3,17-dione (2) with sodium borohydride in micellar system.
Micellar system is obtained from water, organic solvent and anionic surfactant. The organic solvent selected from hydrocarbon such as toluene, n-hexane or n-heptane and ether such as diethyl ether, di-isopropyl ether or 2-methyl THF; preferably toluene.
The quantity of water to 4-androstene-3,17-dione (2) is 3-12 times (volume/weight); preferably 5-10 times (volume/weight).
The quantity of toluene to 4-androstene-3,17-dione (2) is 3-12 times (volume/ weight); preferably 5-10 times (volume/weight).
Anionic surfactant is selected from sodium lauryl sulfate, potassium lauryl sulfate ammonium lauryl sulfate, dioctyl sodium sulfosuccinate, sodium dodecyl sulfate and sodium laureth sulfate; preferably sodium lauryl sulfate
Ratio of anionic surfactant to substrate is varied from 5-40% (weight/weight). It has been observed that 10% (weight/weight) of surfactant with respect to substrate gave the best results.
To effect the concentration of sodium borohydride the reaction is carried by varying the mole ratio between 0.5 to 4 mole with respect to compound 2; preferably 1 mole.
The reaction is carried out at temperature of about 10-50°C, preferably at 20-30°C.
Remarkably, with anionic surfactant such as sodium lauryl sulfate as amphiphilizing, regio-selectivity as well as stereo-selectivity for compound 1 is found to be constantly >95%.
The crude testosterone obtained by according to the present invention further purified by crystallisation from mixture of water/ethanol.
The isolated testosterone may be dried using different techniques of drying like tray drying and rotatory drying techniques with or without application of vacuum and/or under inert condition.
The present invention is further illustrated by the following example which should not be construed to limit the scope of the invention.
EXPERIMENTAL DETAILS
Example-1; Preparation of testosterone (1):

In a multi neck reactor, equipped with thermo-pocket and overhead stirrer, 4-androstene-3,17-dione (50 g, 175 mmol) was added to a solution of deionized water (250 ml), toluene (250 ml) and sodium lauryl sulphate (5 g, 10% weight/weight) at room temperature. Reaction mixture was cooled to 15°C and sodium borohydride (6.6 g, 175 mmol) was added in two successive portions and stirred at 15-25oC for 60 hours. The reaction was quenched by treating with isopropyl alcohol (75 ml), 2N HCl (85 ml) and sodium chloride (90 gm), and stirred further for 1 hour. The organic phase was separated and the aqueous phase was extracted with toluene (200 ml x 2). The combined organic phases were successively treated with 2% aqueous sodium bicarbonate solution (100 ml) and water (150 ml). The organic phase was evaporated to give crude product (53 g). HPLC Purity: 96.5%.
Example-1 (a): Preparation of pure testosterone (1)
Crude product (50 g) was suspended in mixture of water (150 ml) and ethanol (250 ml), the resulting reaction mixture was heated to 60 0C and stirred for 1 hour to solublize the crude product. Reaction mixture was gradually cooled to 20 0C and stirred further for 2 hours at 20 0C to precipate out pure testosterone, it was filtered and dried under vaccum at 500C for 24 hours. Yield: 38.5 g (77 % white crystalline solid), HPLC Purity: 99.25%.
FTIR (KBr): 3510, 3276, 2948, 2924, 2848, 1667, 1651, 1347, 1231, 1057 cm-1.
MS (EI): C19H28O2 ; Exact Mass: 288.21 Observed Mass: 289.2 (M+1).
Melting point: 154.2 °C

Various experiments for preparation of testosterone by equimolar quantity of 4-androstene-3,17-dione (2) and NaBH4:
Example No Compound 2 NaBH4 Sodium Lauryl Sulphate Yield* (%) HPLC Purity (%)
g Mole g Mole (g) % weight wrt compound-2
2 15 0.052 1.98 0.052 6 40 50 99.4
3 20 0.07 2.64 0.07 2 10 70 99.84
4 20 0.07 2.64 0.07 2 10 75 99.6
5 50 0.175 6.6 0.175 5 10 78 99.36
6 70 0.244 9.2 0.244 7 10 75 99.43
7 70 0.244 9.2 0.244 7 10 75 99.3

g: Gram; w/w: weight/weight
(*obtained after crystallisation)
Chromatographic System:
Instrument : HPLC equipped with Pump, UV detector and Recorder.
Column : Altima C18 C8 (4.6 x 250 mm), 5µm
Wavelength : UV at 254 nm
Flow rate : 1.5 mL/min
Injection volume : 20 mL
Column oven temp : 40 °C
Auto sampler temp : 25 °C
Mobile phase-A: Prepare a homogeneous mixture of HPLC grade water and methanol (45:55 v/v). Mixwell and sonicate to degas.
Mobile phase-B: Use HPLC grade methanol.
Diluent preparation: Use HPLC grade methanol as a diluent.
Gradient Programme:

Time (In mins) Mobile phase-A (%) Mobile phase-B (%)
0.01 100 0
4 100 0
24 60 40
53 0 100
55 0 100
56 100 0
70 Stop

Retention time of 4-androstene-3, 17-dione:18.6 min
Retention time of testosterone: 21.5 min
TLC, Rf (Acetic acid: Ethanol: Dioxane: Dichloromethane; 1:2:10:90 ) = 0.8