SYNTHESIS AND SPECTRAL STUDIES OF MODIFIED STEROIDS

• ^ . i " " " ' -

DISSERTATION SUBMITTED IN PARTIAL FULFILMENT OF THE REQUIREMENTS

FOR THE AWARD, QF THE DEGREE OF

/ lEafliter of ^IjUosiopIjp { ' • • - . . • \ w i "" •• I i'.

I C H E MISTRY "

P .// ,-'.^^: .» P.. V

. V V • • • " . . . . , . V - . — • • '•.. • • • • •••

t A i E E M F A T I M A

DEPARTMENT OF CHEMISTRY ALIGARH MUSLIM UNIVERSITY

ALIGARH (INDIA)

2008

Fed In Compute

1 9 m ^^^^

DS3866

DEPARTMENT OF CHEMISTRY ALIGARH MUSLIM UNIVERSITY, ALIGARH-202002, INDIA

Dated:0^:J.^.:?:9f^^

Certificate

This is to certify that the dissertation entitled ^^Synthesis and

Spectral Studies of Modified Steroids'\ by Ms. Tazeem Fatima is

suitable for submission for the award of the degree of Master of

Philosophy in Chemistry.

Prof. M. Mushfiq

ACKNOWLEDGEMENT

Foremost, I am thankful to the most merciful Almighty

Allah who has given me the strength to accomplish this task.

I extend my deep sense of gratitude to my supervisor

Prof. M. Mushfiq for his able guidance and ungrudged encouragement.

I am also beholden to Dr. Shamsuzzaman and Dr. Mehtab

Parveen for their kind help.

I deem it my most pleasant duty to acknowledge my family

for providing me strong support. I am highly indebted to my Uncle

Dr. S. Javed Haider for his invaluable care.

I convey my thanks to my seniors, my research colleagues

and my friends for their co-operation in giving final shape to this

dissertation. I gratefully acknowledge the facilities provided by

Department of Chemistry, A.M.U, Aligarh and financial assistance

from U.G.C.

Tazeem Fatima

CONTENTS

STEROIDAL PYRAZOLES

INTRODUCTION 1

THEORETICAL 2-36

DISCUSSION 37-51

EXPERIMENTAL 52-60

REFERENCES 61-67

I1TE0DUCTIO1

There has been considerable interest in pyrazole ring system , both

with regard to heterocyclic chemistry and pharmacological activities of several

of its derivatives.

Pyrazole and its derivatives have been extensively studied because of

ready accessibility ,diverse chemical reactivity , broad spectrum of its

biological activity such as antimicrobial' cytotoxic agents , anti-inflammator)' ^

, antitumor'*, anticonvulsant ^ antitubercular , tyrosine phosphate IB inhibitor^

and anti-HTV *etc,Looking to these multifold properties exhibited by them

,much importance has been paid to the synthesis of pyrazole derivatives.

The synthesis of steroidal pyrazole derivatives has attracted the

attention of organic chemists because of the remarkable and unusual

physiological activities and profound endocrinological interest associated with

them. Since many drugs and dyes contain pyrazole nucleus ,the fusion of

pyrazole ring to steroidal nucleus may hold promise for many potential drugs

.The unusual anabolic activities observed in humans , has attached a vital

significance with such compounds in steroids.

In view of the above description , attempts have been made to

synthesize steroidal pyrazoles. The work reported here includes a brief

introduction along with recent interesting reports for the synthesis of the

similar compounds. These compounds have been characterized on the basis of

their physical and spectral properties

The pyrazole ring system (I) consists of a doubly unsaturated five

membered ring containing two adjacent nitrogen atom .Pyrazoles are produced

synthetically through reaction of ap-unsaturated aldehydes and ketones with

hydrazine hydrate and subsequential dehydrogenation.

Knorr'"''' first synthesized a compound containing this system in 1883

by the reaction of ethylacetoacetate with phenyl hydrazine which yielded 1-

phenyl -3-methyl-5-pyrazolone (II) .His interest in quinine led to the test of

antifebrile action of this and related compounds which resulted in the

discovery of antipyrine (III), an important febrifiige.

HoC C-CHT

W N CgHs

( I ) (H) (III)

Knorr^^ introduced the name pyrazole for these compounds to denote

that nucleus was derived fi^om pyrrole by replacement of carbon by nitrogen.

He synthesized many members of this class and systematically investigated

their properties. Special attention has been given to I-phenyl pyrazole because

phenyl hydrazine was most readily available hydrazine and these continue to

be most carefully investigated derivatives.

Since many drugs and dyes contain the pyrazole nucleus , the class

has been widely studied and the field continues to be active today even

though antipyrine and related medicinal compounds are no longer in common

use.

Structurally related derivatives of pyrazoles ( I ) are pyrazoline ( IV )

and pyrazolidine ( V )

( I ) (IV) (V)

Pyrazole '*, which is unsubstituted at position - 1 exhibits tautomerism.

The tautomeric forms are in rapid equilibrium with each other and nitrogen

atoms are indistinguishable .Thus positon 3 and 5 must be equivalent in

pyrazoles exhibiting tautomerism.

Theoretical study have been done about the structure and

tautomerism of N ' unsubstituted pyrazoles in solid state''. Owing to annular

tautomerism ' , pyrazole can exist in two tautomeric forms (VI). N '

unsubstiuted3-R^and N'unsubstituted 5-R .

(VI)

If R i^ R^ four situations are possible in the solid state -(1) the

most common is that where only one tautomer is present in the crystal and

only one kind of crystal is obtained. (2) both tautomer are present in the

crystal .(3) two or eventually , more polymorphs of the same tautomer can

be isolated and (4) both tautomer crystallize in different crystal types

(desmotropy). All these situation have been found in N - unsubstituted

pyrazole compounds' " \

Another important aspect for proton transfer concerns the hydrogen

bond network formed by N'-unsubstituted pyrazoles . It has been shown by

X-Ray crystallography that certain N'-unsubstituted pyrazoles form cyclic

dimers (VII ), trimers (VIII) in the solid state depending on the ring

substitution pattern .In these cyclic structures, degenerate double , triple or

quadrapole proton transfer takes place.

The structures are shown as-

l^'^

(VII)

^ • v x , N

-R'

(VIII)

Pyrazoles monosubstituted at position at position -4 are inhibitors and

deactivators of liver alchohol dehydrogenase and one of them ,4-methyl

pyrazole (IX) has been developed into a drug for the treatment of alcoholism

and could be used as antidote in methanol and ethylene glycol

mtoxi cation 22-26

(IX)

Thus alkylation reaction of pyrazole under microwave irradiation in

absence of solvent expecting to obtain 4-substitution in addition to 1-

substituted derivative has been studied 27

Reactions were performed with pyrazole/halogen derivative in a 1/1

ration and relative amounts of products (XH) and (XIII) were determined by

mass spectroscopy in weight %.

MW C6H5CH2-X + (I)

X

(X)=Br

(XI)=C1

. - " = ^

(XIII)

Owing to its varied range of applications, pyrazoles have been easily

synthesized in excellent yield .There are several general ways for synthesizing

compounds containing pyrazole ring system.

1. Hydrazine is shown to react with ap-unsaturated carbonyl compounds to

give compounds (XIV) and(XV) ' ^

It was expected that the reaction of an aP-unsaturated carbonyl

compound with phenyl hydrazine or other substituted hydrazine would proceed

through hydrazone formation and ring closure in straight forward way to give

pyrazoline or pyrazole. This has been shown to be true with benzal

acetone^°, phenyl hydrazone is formed as an intermediate to yield pyrazoline

(XIV) along with its oxidized product-pyrazole(XV)

The reaction is shown as follows:

C6H5-CH=CH-COCH3 + C6H5NH.NH2 -H2O

C-CH3

HN' I CeHs

N

H,C-

C6H5 HC

C-CH3

N N I

CeHg

+

C-CH3

CQHS C

I

CsHs

N

(XIV) (XV)

Similar mechanism is proposed for alkynyl ketone to give pyrazole

(XVI) directly .It is interesting to note that pyrazoline (not possible as per

mechanism ) is not obtained as a product ' .

0

C6H5C=C—C—CH3 + NH2.NH2 -H2O

H2N

-CH. HC-

CfiH 6^5

-CH,

,N

H

(XVI)

2. The reaction of phenyl hydrazine and its derivatives with 1,3

dicarbonyl compounds also leads to similar results^^ and the mechanism

is shown below.

CH3COCH2CO-CH3 + C-;H<NH.NH.

H2C 1

CeHs

.CH3

N

-H,0

XH,

-H,0

HO

* //

H3C-C>S M 0 HN

CeHs

H3C- \ ^ ^

CeHg

(XVII)

M

The synthesis of pyrazolone (XVIII) from P-keto ester is analogous to

the preparation of pyrazole '*.

CH3 CO CH2 CO -OC2H5 + C6H5NH.NH2

-H,0

XH3

0 HN CeHs

-C2H5OH

H2C-

/

XH-,

^^^ -N N CeHs

(XVIII)

The reaction of aliphatic diazocompounds such as diazomethane or

diazoacetic ester with acetylenes or olefins is shown to give (I) and

(XIX) respectively ^ ' . The mechanism for the reaction ofdiazomethane

with compound containing ethylenic and acetylenic bonds to give

heterocyclic compound is suggested to proceed via 1,3 -dipolar addition

of diazomethane.

CH,

CH,

® HQCOOCH3

HC- -CH,

N

H,C-

H 2 C \ ^ N

CHC00CH3

N

N

( I )

H2C-

n2C>

•COOCH3

.N

H

(XIX)

The exploitation of simple molecule with different functionalities for

the synthesis of heterocycles is a worthwhile process in the chemistry of

heterocycles. Synthesis of some fused pyrazoles from 3,5- diaryl cyclohexenone

are reported ^ .The cyclocondensation of 6-hydroxymethylene 3,5-diaryl-2-

cyclohexenone (XX) with hydrazine hydrate in presence of acetic acid afforded

(XXI) .

NH2NH2

AcOH

A novel synthesis of pyrazole from 1,3-Dinitroalkane has been

reported''^, 1,3-Dinitroalkane can be viewed as synthetic equivalent for 1,3-

dicarbonyl compounds through Nef reaction and could be ultimately converted

into azole heterocycle (XXII).

NH,NHR'

NO2 NO2 N N

\ R'

(XXII)

H.S. Joshi et al.^' reported synthesis, antitubercular and antimicrobial

activities of some new pyrazoline derivatives .The compound (XXIII;R= 3-

bromophenyl) has been screened and found to possess antimicrobial and

antitubercular activity.

Br. /R

^^'-< yc NH2NH2

gl.CHjCOOH

(XXIII)

Synthesis of substituted pyrazoles from 1,3-diketones (XXTV) has been

studied.' ''*^ It is reported''^ that 1,3-diketones synthesized in situ from ketones

and acid chlorides ,were converted into pyrazoles (XXV) by the addition of

hydrazine .This method allows a fast and general synthesis of previously

inaccessible and synthetically demanding pyrazoles.

1) 2. leq LiHMDS toluene/ THF (5:2),0*',1 min.

R

R , 2)leq.R"C0Cl, r.t,

1 min

2eq R'

(XXIV)

^R"

34.eq NH2NH2 HjO ^.

EtOH/THF/Toluene/AcOH (10:7:5:5),reflux,5niin.

(XXV)

Pyrazole derivative(XXVI) has been prepared by palladium catalyzed

four component coupling of terminal alkyne , hydrazine , carbon monoxide

under ambient pressure and an aryl iodide''^.

R. .NH2

1.2 eq

NH'

3eq

+ Ar-I

1 mol% PdClj (PPh3)

CO(ambient pressure) THF/H20(l:l),r.t, 24 -36 hrs

R

Ph—C N

\J (XXVI)

"Ar

A general highly flexible Cu-catalyzed C-N coupling /hydroamination

reaction constitute a straightforward alternative to existing methodology for

the preparation of pyrazole(XXVII) ''"''''

11

R" ^ ^ .

•R"

+

NHBoC

NHBoC

1.2 eq

5 mol% Cul 0.2 eq MeHN-

-NHMe

l.SeqCsjCOa,

THF, 80°C,6-16hrs

(XXVII)

Recently an important mediator of imflammation , Luecattamine B

(XXVIII)was isolated '* and since then attempts to synthesize the compound

and its analogues have been observed .It was found that Luecettamine B

(XXVIII) is of near structure of pyrazolinone derivative(XXIX) '^^.

(XXVIII) (XXIX)

This encouraged synthesis of other new heterocyclic derivative including

both quinolinone and pyrazole moieties in one molecular frame work''^.

12

N2H4H2O

Dioxane

(XXX) (XXXI)

Another example is the cyclization of compound containing exocylic

enone group with thioacetamide in presence of piperidine to afford

pyrazolothiazine (XXXII).

(XXX) (xxxn)

Synthesis of new trifluoromethyl containing pyrazoie derivatives are

reported '' .The introduction of fluorine containing group into an organic

compound bring about remarkable changes in physical and chemical properties

°.Many new fluorinated material which take advantage of these useful

changes eg. drugs and agrochemicals have been designed^\

Continuing the studies directed towards synthesis of new fluorine

containing compounds, the reaction of hindered phenol containing hydrazines

with hexafluoroacetylacetone and trifluoroacetylacetone is shown".

13

t-bu

A: R=t-butyl a :R'=CF3 B: R=Me b: R'=Me

H,C COCF3

COR'

t-bu

(XXXIII)

It is reported ^ that pyrazole serve as building blocks in heterocyclic

system . It is found that 5-amino-l-(5,6 dipheyl-l,2,4-triazen-3-yI)-pyrazoIe-4-

carbonitrile (XXXVI) ,an attractive starting material for preparation of some new

polycyclic azines, has been synthesized via reaction of 5,6-diphenyl-3-

hydra2enyl-l,2,4-triazole (XXXIV) with ethoxymethylene malononitrile (XXXV)

in refluxing ethanol.

Ph> ,N ^ N

Ph" "^i^^^^^^'^NH.NHj

NC

NC

Ethanol

OEt

(XXXIV) (XXXV)

(XXXVI)

14

The ethyl-4-cyano-1 -[5,6-diphenyl-1,2,4-triazen-3-yl]- lH-pyrazol-5-ylimi-

do formate(XXXVII) resulted in 70% yield via treatment of compound

(XXXVI) with triethyhlorthoformate(T.E.O.F) in acetic anhydride. Heating of

compound (XXXVII) with hydrazine hydrate in benzene - water mixture

resulted in a 61% yield of the N'-[4-cyano-l-(5,6-diphenyl-l,2,4-triazin-3-

yl)lH-pyrazole-5-yl]-imidoformic hydrazide (XXXVIII), which was transformed

into (XXXIX) with 81% yield.

(XXXVI)

T.E.O.F .r-y -CN

\

/ / ^ N N

N ^ ^ O E .

(XXXVII)

NH2NH2

Benzene/HiO

NH.NH2

Dry benzene

/ \

// \ N N w

p/ Ph

NH

,NH2

.<^

(XXXVIII) (XXXIX)

On treatment of (XXXVII) with mercaptoacetic acid in dry benzene, l-

(5,6-diphenyl-l,2,4-triazen-3-yl)-5-(2-ethoxy-4-oxo-l,3-thiazolidin-3-yl)-lH-

pyrazole-4-carbonitrile derivative (XL) was obtained in 68% yield, which on

heating in a refluxing ethanolic sodium methoxide solution afforded (XLI).

15

HS> XOOH

(xxxvn)

(XL)

MeONa

Ethanol

(XLI)

54 Literature reveals that N-substituted pyrazoles have been

obtained directly by condensation reaction of substituted hydrazines with

ethoxymethylene malononitrile .Condensation of ethoxymethylene malononitrile

with hydrazinoformic acid methyl ester (1) were performed in ethanol or

xylol at different temperatures. All synthesis gave (XLII) as sole product.

C2 HgO

H

CN

CN

(1)

(XLH)

It has been found " that ethyl-2-cyano-3,3-di(methyhhio)acrylate reacts

with 5-methyl- 1- t-butyl-4-hydrazine pyrazole (XLIII) in presence of refluxing

ethanol to give 5-amino-l-(5'methyl-l-butyl-4'-pyrazolyl-carbonyl-3-methylthio)-

lH-pyrazole-4-carboxylate (XLIV) in excellent yield.

16

Proposed mechanism for the generation of compound (XLIV) is shown

as follows:

MeS,

NHNH

(XLIII)

COOEt

MeS,

NHNH

COOEt

C ^ N

H,C

COOEt

SMe

H-,C - - ' ^ • " ^ ^ \ O' NH

-N I >-COOEt

(XLIV) SMe

Generation of (XLIV) was assumed to proceed via nucleophilic attack

of NH2 group to ethylenic bond in dithioacetal followed by elimination of 1

mole of methyl mercaptan to give an intermediate which was followed by

intramolecular attack on CN by NH and then underwent cyclization to give

final product (XLIV).

Synthesis of P-aminoethyl substituted pyrazoles has been reported

.Diketones reacts with hydrazine hydrate in presence of N-substituted amines

such as cyclohexylamine in aqeous medium at room temperature with molar

ratio of 1: 1.05: 1.1 leads smoothly to the corresponding 4-(-2-cyclohexylamino

ethyl)-3,5-dimethyl pyrazole (XLV) in high yield.

Me

=0 NH2.NH2

Me

NHR'R^ Me^ . / ^ ^ \ ,

// (

H

MR^R^

(XLV)

R'=H

R^=

Synthesis of pyrazole containing 2,4-disubstituted oxazole-5-one was

carried out by two different methods^^

In the first method (conventional method A) disubstituted pyrazole

derivative (XLVIII) was prepared by reacting l-phenyl-3p-tolyl -IH-pyrazole-

4-carbaldehyde (XLVI) with hippuric acid (XLVII) and sodium acetate in

acetic anhydride for 2.5- 4 hours.

In the second method (method B) ,the above reaction was carried out

under microwave assistance . Compared to the conventional method ,the

microwave assisted synthesis demonstrate several advantages in terms of

reaction time and overall yield.

H,C.

(XLVII)

Method A: CHjCOONa, AC2O

>-

Method B: MW, CHjCOONa.AcjO

. - - ^

(XLVIID

18

Synthesis of sterically crowded pyrazole ^ from P-diketone (XLIX) was

carried out using harsh conditions (16 hours at reflux at 110-120 in DMIF/

THF solution). These condition served quite well for pyrazole formation in

solution. The pyrazole intermediate protected as methyl ester (L) was isolated

in good yield .The crude material was passed through a short silica gel

column and then deprotected using BBrs .The final product 4-butyl-3,5-bis

(4-hydroxyphenyl)-l-phenyl-lHpyrazole(LI) was isolated in 98% yield.

0 0

MeO' OMe

(XLK)

ArNH.NH,HCl

THF/DMF(1;3)

220°C

<i:^^

MeO' OMe

BBr,

CH2CI2

(L) . ^ ^ ^ ^

(LI)

19

Ganpat L. Talesara et al. ^ reported synthesis of pyrazolo-pyrazole from

isoniazid(LII). To prepare 2-isonicotinoyl-5-methyl-2,4-dihydro-3H~pyra2ol-3-one

(LIII) , isoniazid (isonicotinic acid hydrazide) (LII) was treated with ethyl

acetoacetate in absolute alcohol . Transformation of 2-isonicotinoyI-5-methyl-

2,4-dihydro-3H-pyrazol-3-one (LIII) to its corresponding arylidine derivative

(LV) was achieved by treatment with 4-substituted benzaldehyde

(LlV).Compound (LV) when subjected to reaction with hydrazine hydrate , a

cyclocondensation afforded (LVI).

HaCv

NHNH, O OEt

(LH)

abs. ethanol

heat, 34hrs.

^ ^ ^ ^ ^

NH2NH2. H2O

(LVI)

20

Anshu Dandia et al. carried out one-pot synthesis of novel spiro

[pyrazolo benzothiazepine] using microwave irradiation . Neat synthesis of key

intermediate (LIX) was done under microwave irradiation . Isatin (LVII) and

3-methyl -l-phenyl-2-pyrazolin-5-one (LVIII) was irradiated for 6 - 8 minutes

to give (LIX) in quantitative yield . Intermediate (LIX) synthesized , in situ,

was reacted with (LX) under microwave irradiation and afforded spiro [indole-

pyrazole[4,3-c][l,5]benzothiazepines] (LXII) via formation of an intermediate

(LXI).

(LVII) (Lvni)

Heat

MW

(LIX)

01 SH

•'-^f^'^NH^

(LX) (LXI)

(LXII)

21

STEROIDAL PYRAZOLE

The wide range of application of pyrazoles have resulted in

tremendous amount of interest in the synthesis of steroidal pyrazoles

derivative which is our field of concern .Several papers have appeared ^ "'

dealing with the preparation of steroidal pyrazoles. Here, the synthesis of

steroidal heterocycles containing pyrazole ring fused to steroidal nucleus is

discussed .

R.O. Clinton et al. synthesized steroidal [ 3,2-c] pyrazoles . The

reaction was carried out using 2-hydroxymethylene-17amethylandrostan-17P-

-ol-3-one (LXIII) which was condensed with phenylhydrazine and a single

pyrazole (LXIV)was obtained.

+ C6H5NH.NH2

(LXIV)

Ralph Hirschmann et al. " reported synthesis of phenyl pyrazoles. Effort

was made with 4 ,5a-dihydrocortisol (LXV) which was converted into the

bismethylene dioxy (BMD) derivative (LXVI) and then allowed to react with

ethyl formate to give (LXVII) .Thus condensation of (LXVII) with

22

phenylhydrazine gave a mixture which was separated by chromatography into

two isomeric pyrazoles (LXVIII) and (LXIX) showing absorption maxima at

269 mn (log s 4.30) and at 251m^ (log 8 4.04) respectively.

CH2OH

(LXV) (LXVI)

(Lxvn)

C6H5-N

(LXVffl)

(LXK)

23

B.Green et al7'has reported the synthesis of steroidal [16a, 17a-d]-2'-

pyrazolines [16,17-d]-pyrazoles. In the reaction reported , the addition of

diphenylnitrilimine to 3p-acetoxy-17-cyanoandrostan-5,I6-diene(LXX) took place

to yield 3p-acetoxy-17p-cyanoandrost-5-eno-[16a,17a-d]-r,3'-diphenyl-2'-

pyrazoline (LXXI). Pyrolysis of (LXXI) at 290° led to the elimination of HCN

and formation of 3P-acetoxyandrost-5-eno-[16,17,d]-diphenylpyrazole(LXXII).

CN

CH3C00

© © + C6H5-N-N=C-C6H5

(LXX)

CHgCOO'

CfiK 6^5 -HCN

(LXXI)

CH3COO

CeHs

(LXXII)

24

Klaus Peseke* et al. ^ synthesized new pyrazole derivative . With the

aim to obtain pyrazolo annelated androstenalones ,3P-acetoxy-16-

[bis(methylthio) methylene]-5-androst-5-en-17-one (LXIII) was allowed to react

with 80% hydrazine hydrate in boiling methanol affording 5'methyl thio-

pyrazolo[4'3':16,17]androst-5-en-3p-ol (LXXIV)in 70% yield.

O

SMe

AcO

SMe NH2NH2

MeOH, reflux,-5hrs

(LXXIII) N NH

SMe

(LXXIV)

A facile synthesis of steroidal pyrazoles from testosterone has been

reported '^ . 17P- Hydroxy-2-ethoxy methylene androst-4-en-3-one (LXXV) on

treating with hydrazine hydrate in presence of ethanol and acetic acid

afforded (LXXVI) in 85% yield.

25

(LXXV)

N,H. 2^4

EtOH.AcOH

(LXXVI)

Similarly 17P-Hydroxy-2-bis(methylthio) methylene androst-4-en-3-one

(LXXVII) was treated with hydrazine hydrate in presence of ethanol and

acetic acid and (LXXVIII) was obtained in 64% yield.

H3CS

(LXXVII)

N2H4

EtOH.AcOH

(LXXVIII)

26

Synthesis of steroidal pyrazoline '^ was carried out by reacting cholest-

4-en-3-one (LXXIX) with hydrazine hydrate which gave steroidal pyrazoline

(LXXX) in quantitative yield.

98^17

(LXXIX)

N2H4.H20

Gl acetic acid

CBH 8^17

(LXXX)

A similar synthesis of steroidal pyrazoline was carried out by using

3p-acetoxycholest-5-en-7-one (LXXXI) and 3p-chlorocholest-5-en-7-one

(LXXXII) to yield similar product (LXXXIII).

NH2NH2H2O

(LXXXI) AcO

(LXXXII) CI

Gl . acetic acid, reflux

CRH 8rl17

(LXXXIII)

27

For the characterization of the compounds obtained during the study ,

their spectral properties are used. For a ready reference , the spectral data of

some interesting compounds along with their references are given in table as

follows:

28

U

H ;x4

(U

Pi

ON

U •a

• S V TO

1/5 o\

U- 00 'O -^ E»

00 >-i m >/ cs 00 t~-' i r i •^ m

s t ^ y—i

r-l 00

u o

CO

ON

00

.2

" -s -

-O T3 ^--'

fS (N VO Tf —1 00

r^ ir^ o en r^ fn (N

E p

00

o o ON \ 0

n o

2^

r^ 00 o O v- ON

o

o

o u

2 -I

29

00 00

pi

u

ir i vTi m

. *\

6? o

>—• CO

^ m m • ^ "

O

a ^ a

tf ^ ' c«

X i r-ON o 1—I

g 1

ou <n »-H

r-

/— O

o 1

o (S

— ( / }

o o\ m

. .

a>

IS •4-»

o ffi ;^

*- M

i n 1-H

ir>

> s ^

M

• < * •

VO

m

u

(/ i t—H

(N CS

u p

w /-=; O ^ ' e o S C'O - s S ^ oo ~ m

/~\ S O T1-'

#s ^>w/

O O --i TJ- 0> 00 c s ^ >n m >-< '—'

0\ i - ^

T3 C cd

O m cs r-H

en I

• ^ Tj- 0 0 - ^ <N VO m CN f T ) f—( r-H f -H

O CM

O u

30

U 12

» ^

in

a

CO I

X o O

• ^ OJ

N H .

Q . X

c c

M E ^ m CO CO CO to CN od t^ »;j

. a TO 0) O) c 03 o O

CM

Q X

« X TO < — ' <U • -

| s o cvi

. . f f i

4) CJ

a> o

s

K II U

-a ,-v

3

-^ ^- ""• O O fO o .

ffi in 6 2 6^

CO " ^ * -

ON O fO 1—I i n >—I

in •^ M -—'

E o P

e o

o m o" m

Q ^,

O

O

u ^ V

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37

STEROIDAL PYRAZOLE

The diverse chemical reactivity and broad spectrum biological activity of

pyrazoles has attracted the attention of organic chemists towards synthesis of

steroidal pyrazoles.

Inspired by the results obtained by earlier workers , we took up the

study of synthesis of steroidal pyrazoles from aP-unsaturated ketones

belonging to cholestane series. In the light of earlier observation , easily

accessible ketones such as :3P-Acetoxycholest-4-en-6-one(LXXXIV); Cholest-4

- en -3 ,6 -dione (LXXXV ) ; 6p- bromocholest-4-en-3-one ( LXXXVI) ; 3(3 -

acetoxycholest-5- ene -7 one (LXXXI) ;3p-chlorocholest -5-en-7-one ( LXXXII)

were subjected to the reaction of hydrazines / phenyl hydrazine in presence of

acetic acid and the compounds have been characterized on the basis of their

physical and spectral properties.

98^17

AcO 6 (LXXXIV) (LXXXV)

AcO

(LXXXVI) (LXXXI)

(LXXXII)

38

(a) Reaction of 3B-Acetoxvcholest-4- en-6 -one (LXXXIV) with phenyl

hydrazine ;

The ketone (LXXXIV) on treatment with phenylhydrazine in presence

of glacial acetic acid after usual work-up and column chromatography over

silica gel afforded a single compound, m.p.226° C.

AcO

Pa' ir

6 (LXXXIV) (LXXXVII)

AcO

(LXXXVIII) (LXXXIX)

Charactrization of the compound . m.p. 226° as 3-oxo cholest -4- eno [ 4 , 6 -

c. d1 -2'- phenyl pvrazole (LXXXVII);

The compound was analyzed for C 33 H 46 N2 O . Its mass spectrum gave

molecular peak ion at m/z 486( C33 H46 N2 0 ) . The examination of i.r

spectrum of this compound revealed the absence of absorption band at 1725

and 1230 cm " , typical of acetate group ^^{-Clh COO ) . The absence of these

characteristic bands in its i.r spectrum completely ruled out the possibility of

expected compound (LXXXIX) .However the presence of two nitrogen atoms

in the molecule indicated formation of pyrazole moiety . The sharp band at

1680 cm " clearly indicated presence of an aP - unsaturated carbonyl

compound in the molecule . Further , there were prominent bands 1600, 750

and 690 cm" , characteristic of monosubstituted benzene ring observed in

39

phenylpyrazoline type derivatives .The other significant band appeared at

1490 cm " , characteristic of (C=N) vibration . ^'^^ These spectral data and

molecular composition of the compound , m.p. 226 ° are compatible with tv/o

isomeric structures ( LXXXVII) and ( LXXXVIII ) discarding expected

structures (LXXXIX).

SCHEME -lA

C«H

Ph (LXXXVIIa)

Oxidation

(LXXXVII)

40

In this mechanism, it is proposed that aerial oxidation involving free

radical mechanism leads to formation of (LXXXVII) .The hydroxyl

intermediate (LXXXVIIa) being an allylic alchohol may undergo ready

oxidation that leads to ap- unsaturated ketone (more stable).

An alternative mechanism may also be suggested for the formation of

(LXXXVII) from (LXXXIV) as shown in scheme 1 (B).

SCHEME H B)

C«H

AcO

41

H,0 ®

Oxidation

HNN

Ph

H N -I Ph

-N

(LXXXVII)

In this mechanistic speculation the desired compound (LXXXVII) is

obtained through bis hydrazone formation.

Thus the compound, m.p 226° ,on the basis of its molecular composition

, spectral evidences , mechanistic speculation and general consideration could

be best characterized as 3-oxo-Cholest-4-eno[4,6-c,d]-

2'-phenylpyrazole(LXXXVII).

(b) Reaction of cholest-4-en-3 ,6-dione (LXXXV) and phenylhydrazine;

Cholest-3-oxo-4-enor4,6-c,d1-2'-phenvlpvrazole (LXXXVII):

CRH B'^U

0 (LXXXV)

(Lxxxvn)

42

The diketone ( LXXXV ) was subjected to phenylhyrazine reaction.

After usual work-up and column chromatography , the compound m.p. 226°

was obtained as the sole product of the reaction . This compound was found

to be identical with pyrazole (LXXXVII) , m.p. 226° , in all respects (1.1.

c , m.p. , i.r , nmr and mass).

The formation of the compound (LXXXVII) from ketone (LXXXV)

may be shown according to the proposed mechanism ( Scheme - 2 ) .

SCHEME-2

98^17

O (LXXXV)

1 .Hydrolysis

2.Oxidation

43

(c) Reaction of 6B-Broinocholest-4- ene-3 -one ( LXXXVI ) with

phenylhydrazine:

The ketone (LXXXVI) on treatment with phenyl hydrazine in presence

of acetic acid under similar conditions described above furnished after usual

work-up and column chromatography two compounds m.p. 226" and 195°.

Both the compounds obtained from this reaction showed negative Beilstein

test .

98^17

(LXXXVII)

(XC)

N I I NH

(XCI)

NNHPh

(XCII)

Interestingly , the reaction of bromoketone with phenyl hydrazine too

furnished , as one of its product, the same compounds , m.p. 226° ( identical

in all respects) as obtained from the ketones (LXXXIV) and (LXXXV). It

again offered room for mechanistic consideration .

The formation of ( LXXXVII) from (LXXXVI) may be proposed

according to Scheme 3 ( A ) :

SCHEME 3 f A)

44

98^17

HNN

Oxidation

(XC)

e H.O

N N

Ph (LXXXVII)

45

In this scheme , at the first stage the formation of phenyl hydrazone

takes place. After this the other molecule of phenylhyrdazine attacks at C - 6

in the fashion of the nucleophilic displacement and thus bromide ion is

liberated . Intramolecular reaction and oxidation results in the formation of

pyrazole (LXXXVII).

Alternatively , the following mechanism could also be taken into

account to support the structure (LXXXVII ) as proposed according to

Scheme 3 ( B ) :

SCHEME 3rB)

HNI

Ph Br

N ^ H

•N Br I Ph

H3O ©

^

H Ph—N Br

I NH2

(LXXXVII)

46

Characterization of the compound m.p. 195° as Cholest-4-eno r4.6-c, d 1 -V-

phenyl pvrazole-3-one phenvihvdrazone (XCV.

The compound m.p. 195° analyzed for C39 H52 N4 .This composition

was further supported by its mass spectrum giving molecular ion peak at m/z

576 (C39 H52 N4) . The composition C39 H52 N4 can lead to several possible

structures (XC; XCI; XCII) formulated for the compound , m p. 195° giving

negative Beilstien test (Brabsent).

The examination of i.r. spectrum of the compound , m.p. 195° contrary

to the first product of the reaction , revealed no absorption band for the

carbonyl group . The band observed at 3240 cm"' can be ascribed to NH

vibration which indicated presence of phenyl hydrazone moiety as shown in

all three possible structures ( XC; XCI; XCH ) .The bands of 1605 , 750 and

695 cm ' can be attributed to aromatic grouping . Two prominent absorption

bands at 1590 and 1480 cm'' can be assigned to C=N vibration .The i.r. data

are in agreement with the three proposed structures ( XC; XCI; XCII) .A

distinction can be made on the basis of its n.m.r spectrum . The n.m.r

spectrum of the compound , m.p. 195°C exhibited a signal between 5 7.1-7.5

as multiplet integrating for ten protons (2 Ce H5 ) .A singlet at 6 12.7 may

be ascribed to NH proton of phenyl hydrazone moiety present in the

molecule.

This downfield shift of NH proton to such an extent is attributable to

the fact that there is no vicinal coupling . The other notable features of the

spectrum were found to show signals at 5 2.9 and 2.5 . The multiplet at 6

2.9 may be ascribed to C2-H2 and doublet at 5 2.25 ( J 10 Hz) t o C ? -

methylene protons .Methyl protons were seen at 5 1.2 ,0 .9 , 0.8 and 0.73 .The

structure (XCII )can be discarded on the basis of n.m.r. spectrum where a

singlet would have appeared in the region 5 2.4 -2.6 for Ce - P H. Now the

choice is narrowed down between two possible structures ( X C ) and (XCI)

because the i.r. and n.m.r. value are in good agreement with these two

structures. Mechanism wise both the structures (XC) and (XCI) are possible.

47

The formation of structure ( X C ) from ( L X X X V I ) can be shown

according to Scheme 4 :

SCHEME 4

98^17

Br (LXXXVI)

H,0 0

p\r I" NH2

H Br

"J H H

N -I Ph

, ^ N—H

Aerial oxidation •

(LXXXVII)

PhNHNH,

48

The formation of (XCI) from (LXXXVI) can be shown according

to mechanistic pathway as shown in Scheme 5

SCHEME-5

PsHi?

Br (LXXXVI)

N

I 1 Ph

rVtU NH I Ph

N ^

N I Ph

N I NH I Ph

(XCI)

49

The compound (LXXXVII) which has been obtained from this

reaction as one of the product , can conveniently lead to (XC )and vice -

versa.

It is reasonable to prefer the structure ( XC ) over ( XCI) for the

compound , m.p. 195° in the light of spectral data , mechanistic and general

consideration with certain limitations.In view of this discussion , the

compound , m.p. 195 °C can be characterized as cholest -4 -eno [ 4 , 6 c,d ]- 2 -

phenylpyrazole-3-one phenylhydrazone (XC).

Further to substantiate the structure ( XC ) , the compound (LXXXVII)

was subjected to the reaction of phenyl hydrazine under milder conditions.

This reaction, after usual work-up , afforded a compound ,m.p. 195° C

which was found identical in all respects (t. l .c . , m.p. and spectra with the

one obtained from 6P-bromocholest-4-en-3-one( LXXXVI) wth phenyl hydrazine

as one of its product.

(LXXXVII)

This chemical transformation confirms the structure ( XC ) for

compound , m.p. 195°C and thus the ( XCI) is disfavoured.

50

(d) Reaction of the 3B- Acetoivcholest -5-en-7-one ( LXXXD and 36-

ChIorocholest-5- ene -7-one ( LXXXn ) with hydrazine hydrate;

Cholest-3-en-r 5.7-c,d 1 pyrazoline fLXXXnr>:

The reaction of (LXXXI) and (LXXXII) with hydrazine hydrate in

glacial acetic acid , after usual work - upand column chromatography ,

afforded identical product (LXXXIII) with negative Beilstien test.

CflH 8^17

R

(LXXXI) AcO

(LXXXII) CI

NH2NH2.H2O

1

Gl.acetic acid, reflux

(LXXXni)

Characterization of the compound , m.p. 205° as ; Cholest-3-en- \ 5 ,7 -c,d 1

pyrazoline (LXXXni):

The i.r. spectrum of the compound (LXXXIII),m.p. 205° C , showed

characteristic absorption band at 3448 cm "'(NH of pyrazoline ring ) and 1590

cm ' (C=N). Its NMR spectrum exhibited one proton singlet at 5 6.4 for

NH proton , another singlet for two (pyrazoline ring methylene ) protons at 6

2.6, a one proton doublet at 5 6.1 (C4-H)and multiplet for one proton at 5

5.9(C3-H). The spectral data confirms the structure (LXXXin).

The formation of (LXXXIII) from (LXXXI) and (LXXXII) can be

rationalized on the basis of Michael addition as shown in Scheme - 6

SCHEME-6

51

98^17

R (LXXXI) AcO

(Lxxxn) CI

NH2NH2.H20

Gl.acetic acid, reflux

52

All melting points are uncorrected. I.R. spectra were measured with a Perkin-

Elmer 237 spectrophotometer. N.M.R spectra were run in CDC^on a Varian

A60 instrument with TMS as internal standard. Thin layer chromatographic

plates were coated with silica gel. Light petroleum refers to the fraction of

b.p. 60-80°. N.M.R values are given in ppm (s, singlet ; d, doublet ; t,triplet

;dd, doublet of doublet ; mc, multiplet centered at ). I.R. values are given in

cm'' (s, strong ; m, medium ; w, weak ; br, broad ).

3B-Acetoxvcholest-5-ene.

A mixture of cholesterol (50gm , 0.129 mole), pyridine (75ml),and acetic

anhydride (50ml) was heated on a steam bath for 2 hours .The resulting

brown solution was poured onto crushed ice water mixture with stirring . A

light brown solid was obtained, which was filtered under suction ,washed with

water until free from pyridine and air dried .The crude product on

recrystallization from acetone gave the pure acetate (45gm,0.105 mole), m.p.

114-115 (lit. *^m.p. 116°).

3B-Acetory-6-nitrochoIest-5-ene:

3p-AcetoxychoIest-5-ene (5 gm,0.0116 mole) was covered with nitric

acid (125ml;d, 1.52) and sodium nitrite (5gm) was gradually added over a

period of 1 hour with continuous strirring . Slight external cooling was also

affected during the course of reaction and stirring was continued for

additional 2 hours when a yellow spongy mass was separated on the surface

of the mixture. The mixture was diluted with cold water (100ml) when green

coloured solution was obtained . The whole mass was extracted with ether.

The ethereal layer was washed with water , sodium bicarbonate solution (10

%) until washing was pink and again with water and dried over anhydrous

sodium sulphate. Removal of the solvent provided the nitro compound as an

oil and crystallized from methanol (3.5 gm, 0.00738 mole) m.p. 104 (lit. * m.p.

102-104°).

53

3B-Acetoxv-Sa-cholestan-6-one;

3P-Acetoxy-6-nitrocholest-5-ene ( 3 gm, 0.0063 mole ) was dissolved in

glacial acetic acid (125 ml) by warming the mixture and zinc dust (6 gm)

was added in small portion with shaking . The suspension was heated

under reflux for 4 hours and water (6 ml) was added now and then during

the course of the reaction . The hot solution was fihered, cooled to room

temperature and diluted with a large excess of ice cold water . The

precipitate thus obtained was taken in ether and ethereal solution was

washed with sodium bicarbonate solution(10%) and water and dried over

anhydrous sodium sulphate . Evaporation of solvent gave ketone as an oil

which was crystallized from methanol (2. Igm, 0.0047 mole) m.p. 128 (lit.

m.p. 127 - 128° ) .

3B-Acetoxv-5-bromo-5a-cholestan-6-one;

To a cooled solution of 3p -Acetoxy -5a cholestan-6-one (2 gm , 0.0045

mole) in acetic acid (5ml) and ether (18 ml) , bromine solution ( 1.1 gm of

bromine in 15ml of acetic acid) was added gradually with shaking. Few drops

of hydrobromic acid were added to catalyse the reaction. The bromo

compound that precipitate out was filtered and recrystallised from chloroform-

ether (1.2 gm, 0.0023 mole),m.p. 162 - 164° ( lit. ' m.p. 162° ).

3B-Acetoxvcholest-4-en-6-one(LXXXIV>;

A solution of 3P-Acetoxy cholest-5-bromo-5a-cholestan-6-one ( 2 gm

,0.0038 mole) and pyridine (20 ml) was heated under reflux for 8 hours under

anhydrous condition .The reaction mixture was poured into ice cold water,

acidified with dilute hydrochloric acid and extracted with ether.The ethereal

layer was washed successively with water, dilute hydrochloric acid sodium

bicarbonate ( 10 % ) and water and dried over anhydrous sodium sulphate .

removal of solvent provided an oil which was crystallized from methanol to

54

give ketone (LXXXIV ) , (1.5 gm, 0.0034 mole), m.p. 106-108° (lit.*' m.p.

110°).

Reaction of 3B-Acetoxvcholest-4-en-6-one ( LXXXIV ) phenyl hydrazine: 3-

oxo-cholest-4-eno [ 4,6-c.d 1-2' - phenyl pyrazole (LXXXVID;

A solution of 3p- acetoxy cholest-4-ene-6-one ( 1 gm, 0.0026 mole) in

benzene was treated with phenyl hydrazine ( 1 m l ) and acetic acid ( 2 m l ) .

The reaction mixture was heated under reflux for 4 - 6 hours . Then benzene

was removed under reduced pressure and the residue was extracted with

ether. The ethereal layer was washed with water , sodium bicarbonate solution

( 10 % ) and finally with water and dried over anhydrous sodium sulphate .

Removal of solvent gave a semisolid material which was chromatographed

over silica gel (20 g m ) and fraction of 25ml were collected. Elution with

light petroleum : ether ( 4: 1 ) fiirnished 3-oxo-choIest-4-eno[4,6-c,d]-2'- phenyl

pyrazole (LXXXVII) when recrystallized from chloroform and methanol (380

mgm, 0.00078 m o l e ) , m.p. 226°) .

Dmax 1680 ( C=C-C-0 ) , 1600 ( C=C ), 1490 (C=N),

750 and 690 cm "

Analysis Found C, 81.12 ; H, 9.20 ; N , 2.90

C33 H46 N2 0 requires C , 81.48 ; H , 9.46 ; N , 2.88 %

3B, 5 , 6B - Trihydroxy - 5a - cholestane;

A mixture of cholesterol (20 gm, 0.0518 mole) and formic acid (20 ml

; 88 % ) was heated on a water bath at 70 - 80° for 5 minutes and then

allowed to attain room temperature. Hydrogen peroxide ( 2 0 ml ; 30 % ) was

added to the mixture and it was kept at the room temperature for 12 hours

with occasional shaking . Boiling water (300 ml) was added to the mixture

with stirring and the reaction mixture was allowed to attain room temperature

when a white solid separated which was filtered under suction and air dried .

The solid was dissolved in methanol( 600 ml ) and the solution heated with

sodium hydroxide solution (20 ml; 25% ) for 10 minutesf on a steam bath . It

was acidified with hydrochloric acid and diluted with boiling water (300 ml

) . The triol obtained on cooling was collected by filtration under reduced

pressure and recrystallized from methanol (18 gm, 0.0426 mole) m.p. 23S)°

(lit^^m.p. 237-239°).

5-Hvdroxv - Sg- cholestane-3,6- dione;

A Suspension of 3P, 5, 6p-trihydroxy -5a-cholestane ( 5 gm , 0.0118

mole) in acetone (200 ml) was was cooled in an ice bath . Jone's reagent

was added gradually with stirring over a period of 30 minutes . Water (200

ml) was added to the reaction mixture and the precipitate thus obtained was

collected by filtration under suction. The crude product was subjected to

column chromatography over silica gel (100 gm).Elution with chloroform gave

dione which was crystallized from methanol, (3.2 gm, 0.0076 mole) m.p. 255

(lit^^ m.p. 232-235°).

V max 3345 cm - ( O H ) , 1720 cm' ' ( C = 0 ) .

Cholest-4-ene-3.6-dione (LXXXV);

A mixture of 5-hydroxy-5a-cholestane-3,6- dione (2 gm , 0.0048 mole ),

dioxane ( 140 m l ) and sulphuric acid ( 2 ml ) was heated under reflux for 1

hour .The solvent was removed under reduced pressure and the residue was

diluted with water and extracted with ether. The ethereal solution was washed

successively with water , sodium bicarbonate solution ( 10%) and water and

dried over anhydrous sodium sulphate . Removal of dessicant and the solvent

provided desired dione (LXXXV) recrystallized from light petroleum ( 1.4 gm ,

0.0035 mole) m.p. 122° ( l i t " , m.p. 122-123°) .

Reaction of choIest-4-ene-3,6-dione (LXXXV) with phenvlhvdrazine; 3-oxo-

cholest-4-eno r4,6-c.d 1 -2'-phenvl pvrazolef LXXXVID;

A solution of ketone ( LXXXV ) (1.0 gm , 0.0025 mole) in benzene

(20 ml) was treated with treated with phenyl hydrazine (1.0ml) and acetic

acid (2 ml) .The reaction mixture was heated under reflux for 8 hours.The

benzene was removed under reduced pressure and the residue was extracted

with ether. The ethereal layer after washing by usual procedure provided a

semi solid material which was chromatographed over silica gel ( 20 gm ).

Elution with light petroleum ether (35:1) provided the compound (LXXXVII)

which was recrystallized fi^om methanol, (320 mgm, 0.00658 mole ) ,m.p. 226

. It was found identical in all respects (t.l.c ,co-t.l.c, m.p and spectral data)

with one obtained from the reaction of 3P- acetoxy cholest-4-ene-6- one

(LXXXIV) with phenyl hydrazine.

3p-Hvdroxv- 5a ,6B-dibroniocholestane-

To a solution of cholesterol (5 gm, 0.0129 mole) in ether (30 ml) was

added bromine solution (0.9 ml of bromine in 20 ml of glacial acetic acid

containing 0.2 gm of anhydrous sodium acetate ) with stirring. The solution

turned yellow and promptly set to stiff paste of the dibromide. The mixture

was cooled in an ice bath and stirred with glass rod to ensure complete

crystallization . The product was then collected by filtration under suction

and washed with cold acetic acid until the filterate was completely colourless (

6.9 gm, 0.0126 mole), m.p. 112° (lit. ^ m.p. 113°).

5a , 6B Dibromo cholestan-3-one -

The moist dibromide ( 6.9 gm , 0.0126 mole) was suspended in acetone

( 150 ml ) in a three necked round bottom flask fitted with stirring and

dropping fijnnel . The suspension was stirred for 5 minutes and jone's reagent

(10 ml) was then added in drops from dropping frinnel in 15 minutes . The

temperature of the reaction mixture, during oxidation was maintained between

57

0 - 5 by external cooling .After the addition is complete , stirring was

continued for ISminutesand cold water (200 ml) was added. The product

was collected on a buchner funnel and washed thoroughly with water and

methanol and air dried ,( 5 gm, 0.0091 mole) m.p. 73° (decomposition), (lit. *

m.p. 73 -15° ).

6B-Bromocholest-4-en-3-one(LXXXVD

5a, 6p- Dibromocholest-3-one ( 5 gm , 0.0091 mole) was dissolved in

absolute methanol ( 100ml) by warming on water bath. Anhydrous potassium

acetate(2.5 gm) was added to the above solution and the mixture was heated

under reflux for 2 hours. The resuhing yellow coloured solution was poured

into crushed ice-water mixture and the white precipitate thus obtained was

taken in ether , washed several times with water and dried over anhydrous

sodium sulphate. Evaporation of the solvent gave 6P- bromo choles-4-ene -3-

one(LIV) as an oil which was crysyallized from methanol, (2.2gm , 0.0047

mole) m.p. 132° (lit.*^ m.p. 132°).

Reaction of 6B-Bromocholest-4-en-3-one ( LXXXVI) with phenyl hydrazine:

Cholest -4-eno f 4,6-c,dl -2*phenvl pyrazole-3-one phenyl hydrazone ( XC )

and 3-oxo-cholest-4-eno [4, 6 -c,d 1- 2'-phenyl pvrazolef LXXXVn);

The ketone (LXXXVI) (2 gm, 0.0043 mole) was dissolved in benzene

(30 ml) and to this was added phenyl hydrazine ( 2.5 ml) and acetic acid

(2 ml) The reaction mixture was then heated under reflux for 8 hours. Benzene

was removed by distillation under reduced . The residue was extracted with

ether .The ethereal layer was washed water sodium bicarbonate solution (5 %)

and water, dried over anhydrous sodium sulphate . Removal of solvent gave

oily material which was chromatographed over silica gel (50gm)and fraction

of 30 ml were collected. Elution with light petroleum ether (50: 1) furnished

the compound (XC) which on recrystallization from methanol afforde orange

colour crystals, (400 mgm ,0.00069 mole) m.p. 195°.

u max 3240 ( NH ) , 1605 ( C=C ) , 1590 (C=N ), 750 and 695 cm '

M^ 576(C39H52N4)

Analysis Found : C , 81.15 ; H , 9.20 ; N , 9.40,

C 39H 52N4 Requires: C , 81.25 ; H, 9.03 ; N , 9.72%

Further elution with light petroleum -ether ( 4 : 1 ) afforded the pyrazole

recrystallized from methanol - chloroform , (300 mgm , 0.000617 mole) , m.p.

226°.

3B-Acetoxv-cholest-5-en-7-one ( LXXXI);

To a solution of 3P - acetoxy cholest -5 - en ( 8 gm , 0.01869 mole ) in

CCI4 (150 ml), acetic acid (30 ml ) acetic anhydride (10 m l ) , was added 0°

C to a solution of t-butyl -chromate ( from t - butyl alchohol ( 60 ml )

chromium trioxide(20 gms), acetic acid (84 ml) and acetic anhydride (10 ml).

The reaction mixture was heated under reflux for 4 hours and the reaction

mixture was diluted with cold water . The organic layer was washed with

water and dried over anhydrous sodium sulphate . Evaporation of solvent

under reduced pressure furnished an oil which was crystallized from methanol

to give the ketone (3.5 gm, 0.0079 mole), m.p. 156°.

Reaction of 3B-Acetoxvcholest -5-en-7-one (LXXXD with hydrazine hydrate;

Cholest-3-en-r5,7-c,d 1-pyrazolinef LXXXIID;

A mixture of 3P-Acetoxycholest-5 en-7-one ( LV ) ( 1.15 gm ,0.0026

mole) and hydrazine hydrate ( 3 ml) in acetic acid ( 20 ml) was refluxed for 5

hours, the progress of the reaction was monitored by T . L . C. After the

completion of the reaction, reaction mixture was cooled and poured in ice -

cold water . The solution was extracted with chloroform, organic layer was

washed with water, sodium bicarbonate solution ( 5 % ) and dried over

anhydrous sodium sulphate. The solution was concentrated under reduced

59

pressure and residue was crystallized from chloroform methanol , which

provided compound ( LXXXIII) as yellow needles (0.816gm, 0.0021 mole).

m. p. 205°

V max : 3448 ( N H ) , 1654 (C=C), 1590 (C=N) cm-

Analysis Found : C, 81.76; H,11.6; N, 7.06 %

C 27H 44 N2 Requires : C, 81.75 ; H, 11.18 ; N, 7.06 %

3B -Chlorocholest-5-ene;

Freshly prepared thionyl chloride ( 75 ml ) was added gradually to

cholesterol ( 100 gm , 259.06 mmol) at room temperature . A vigorous reaction

ensured with the evolution of gaseous product .When the reaction slackened,

the mixture was gently heated and poured in ice cold water with constant

stirring . The yellow solid thus obtained was filtered under suction washed

thoroughly with cold water and air dried. Recrystallization from acetone gave

3P - chloro cholest -5 - ene ( 94 gm , 0.2323 mole) m.p. 95° ( lit. ° ,m.p. 95 -

96°)

3B-Chlorocholest-5-en-7- one (LXXXH);

A solution of t - butyl chromate [ t - butyl alchohol ( 60 ml ) , CraOs

(20gm), acetic acid (84ml) acetic anhydride (10ml)] was added at O C to

the solution of 3P-chIorocholest-5-ene (8gm, 0.0197 mole) in carbon tetra­

chloride (150 ml ) , acetic acid (30ml) and acetic anhydride (10ml). The

mixture was heated under reflux for 3 hours and diluted with water . The

organic layer was washed successively with water, sodium bicarbonate solution

( 5 % ), water and dried over anhydrous sodium sulphate . Evaporation of

solvent under reduced pressure furnished an oil which was crystallized from

methanol to give ketone (3 . 5 gm , 0.00836 mole ) m. p. 145° .(lit ^\ m.p. 144-

145° C).

60

Reaction of 3B-Chlorochoiest -5-en-7-one ( LXXXn ) with hydrazine

hydrate ; Cholest-3-en-r 5,7-c.d 1 pvrazoline (LXXXnP;

A mixture of 3P-Chlorocholest-5 -en-7-one( LXXXII ) (1.11 gm,0.0026

mole) and hydrazine hydrate ( 3 ml) in acetic acid ( 20 ml ) was refluxed for

7 hours , the progress of the reaction was monitored by T . L. C. After

completion of the reaction, reaction mixture was cooled and poured in ice cold

water , The solution was extracted with chloroform , organic layer was

washed with water, sodium bicarbonate ( 5%) and passed through anhydrous

sodium sulphate. The solution was concentrated under reduced pressure and

residue was crystallized from chloroform -methanol which provided compound

( LXXXIII), (0.826 gm , 0.002 mole ) and m.p. 205° C, identical to the same

product reported earlier in the reaction of 3P-acetoxycholest-5-en-7-one(

LXXXI) with hydrazine hydrate.

61

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