Isolation, structure elucidation and synthesis of 1-deoxyforskolin

Tetrohe&on Vol. 45, No. 3, PP. 763 lo 766, 1989 Printed

in Great Britain.

0

ISDLATIDN,

Y.

STRUCTURE

Khandelwal*,

E.R.

Hoechst

ELWIDATIW

P.K.

Jotwani,

India

AMI SYNTHESIS OF

Limited,

Inamdar,

Mulund,

N.J.

Bombay

-

Isolation

and

structure

a minor forskolin related (2), f%skohlii are described. The 9-hydroxylation Forskolin

cl),

displayed

strong

a

spasmoliticand its

forskohlii.

In

paper,

Careful

that the

confirmed

high M+ of

isolate we

the

India.

of the

2

I-deoxyforskolin

Indian

is

from

herb

Coleus

confirmed

the

Indian

intraocular

activity'.

Our

forskolin

the

Rupp

by

the

(4).

isolated

minor

describe

of

performance

m/z

394).

forskolin of

as

petroleum

liquid

The was

2

the

herb

Coleus

pressure

continued

related

isolation,

only the

its

synthesis

difference

absence

of

I-deoxyforskolin

starting

ether

forskohlii,

lowering,

interest

in

constituents

structure

extract

chromatography,

7-deacetyl-I-deoxyforskolin by

of

antihypertensive,

stimulant

R.H.

of

elucidation

broncho-

forskolin

the and

herb

and Coleus

synthesis

of

(2).

assignment

responding

to

purification

preparative

to

cyclase

us

this

diterpenoid

inotropic,

and

1988)

of

structure

1,9_dideoxyforskolin

labdane

prompted

I-deoxyforskolin

with

major positive

adenylate

analogs

174-176OC,

of

080,

elucidation

constituent,

l-DEOXYFORSKOLIN

Souza

400

(Received in UK 7 November

Abstract

de

004O-l020/89 $3.OO+.Nl 1989 Per~amonPressplc

observed the (2).

(1). from

The

of

the

roots

a

new

product

gave in

resonance

the at

Alkaline structure

1,9_dideoxyforskolin

1

R,=OH,

R,=H,R,=Ac

2 -1

R,=Rg=

Ii,

3 -9

R,=

R7=H

11,

R,=H,

-9

R,=

the (2,

spectra

4.45

ppm

hydrolysis

of

of

6

PMR

of

of

for 2

at

l-deoxyforskolin

plant, yield 2

including 0.001X,

in

lo-CEOH, O°C

which

gave

(2)

the

was

R,rAc

R~=R,=Ac

763

led cor-

further

(4).

Dedicated to Dr. E. B&tin, Managing Director, Hoechst India Limited, OD the occasion completion of twenty-five years service with Hoechst AG., West Germany.

mp

comparison

of his

et al.

Y. KHANDELWAL

164

A to

number

carbonyl

one-step ing

of

methods

group

procedure

direct

are

recorded

preferentially to

effect

oxidation

phenylseleninic

of

the

,

a

in --

situ

and

then The

were

the

with

5,

failed

at

oxidation

directed

towards

unprotected

formed

to

of

the

presumed

in --

2,

R,=R,=H

_, 6

R,=Ac,

R,=H

1,

R, = AC,

R,=SiMe,

a -3

Razz R,=Ac

ether

-9

in

obtained

50%

by

allylic

J.

at

in 6

the

4.64.

(11).

PMR

11

Alkaline

The us

in

the

of

2

with

such

as

a

involv-

thallium

all

alpha

ideally

procedures

lead

enolates

and

having

subsequent

9 not

in

THF

the

en01

at

nitrate',

tetra-scetat2, tried

first

with

our

next

failed,

oxidation.

Treatment 10

-7BOC

gave

6-acetyl-7-

ether.

R=H

6/7-hydroxy

6-acetyl-1,9_dideoxyforskolln

analog,

hexamethyldisilazane

ether is of only the II-en01 II The desired 9-hydroxylation al."

et --

II-enol

spectrum

protected

in

formation

Scherkenbeck

Se02 of

of s

ether in

of

the

at

80°C

signal

for

for 9-C!

tetrabutylammonium followed

1?_

gave

agreement

was

the

with

eventually

II-en01

the

result

achieved

by

2.

pyridine

which with

hydrolysis

by

6 hrs is

provided

absent

with

9-hydroxylated

12-C?

gave

fluoride/THF13 acetylation

the

and

appears

as

product a

singlet

6-acetyl-I-deoxyforskolin

acetic

anhydride/pyridine

gave

(2).

successful

was

The

the

used

group

using

6-acetyl-7-deacetyl-1,9_dideoxyforskolin

and

trimethylsilylchloride/lithium

confirming

which

(7) _

of

of

attempts

of

‘3 1 R=OH

with

Treatment

I-deoxyforskolin

or

THF

oxidation

Treatment lo,

4

hydroxy

derivative.

formed

R,=Ac

yield.

an aim

acid' were

enolate

however, in

the

Reagents

_, 9

R,=H,

-78°C

situ

the

hexamethyldisilazane

4,

Treatment, at

first

9-hydroxylated

analog

trimethylsilylchloride/lithium

of

With

iodosylbenzoic

of

6/7_hydroxyforskolin

our

enolates.

desired

isolation

introduction atom.

Mo05.pyridine.HMPA5,

and

give

deacetyl-7-trimethylsilyl-1,9_dideoxyforskolin

@),

to 2,

KO-t-Bu/0,/(EtO)3P4,

but

attempts

efforts of

with

the

carbon

of 4

Z-(phenylsulphonylj-3-phenyloxy-sziridine7 4

describe

tertiary

conversion

ostensibly

anhydride

which

at

introduction the

converted

to

of

a

structure

of

forskolin

through

9a-OH

group

in

microbial

a It

I-deoxyforskolin.

1,9_dideoxyforskolin also

hydroxylation

provided 12

an

derivative additional

helped substrate

.

EXPERIMENTAL Melting were

points

determined

measured the

as purity

and

heating

(MeqSi

were

determined

with

solution

were carried out by LC 500 equipped with in

series.

a

with

a

Kofler

hot

stage

apparatus

and

are

uncorrected.

IR

spectra

157 spectrophotometer using KBr disks. PMR spectra were FT-90 spectrometer unless mentioned otherwise CDCl3 with a Jeol TLC plates were used to examine Precoated (silica gel 6OF254) internal standard). of the compounds. Visualization was done by spraying with anisaldehyde/H2S04 reagent All compounds were homogeneous by TLC analysis. HPLC separations the plate at IIO'C.

using

Perkin-Elmer in

Associates using a Waters a refractive index detector

Preparative and

two

prep

Liquid

chromatograph,

pak-500/silica

(35-40)

model

prep

cartridges

765

I-Deoxyforskolin Isolation The

of I-Deoxyforskolin and finely divided

dried

40->OoC (256

for

g)

was

5

was

h

on

1.6 very

with

The

to

of

0.05

12-CH),

as

eluent

g

PMR (CDC13) 3.06

=

(61”“J,

:

I?? (KB~)

cm-1

Calcd.

3$00,

for

solution

cooled

of

2940,

layer

was

The

hydroxide

of

was stirred for at a temperature

(60

washed

H,

2

(0.2

an

Rf

by

HPLC

to

lithium

a

water,

from

8.69.

5xCtl3),

(s,

LAH,

30

starting

for

C2OH32P5

:

cooled

to

C,

over

from

C,

water

rmnol)

2

in

trimethylsilane

ed

with

68.18;

brought

as

:

(~1

9-C!,

5.13

(dd, 5.97

Similarly PMR

(m,

6a-Cfi),

organic

eluant,

ml)

mixture

(s,

anhydride water

and

(d,

J6,7

5.73

(dd,

THF

dioxide (2

was

was

was residue,

ethyl

PMR

(d,

J6,7

Jcis

:

J gem

:

4JT2,9

7a-CH),

17Hz,

Jcis

=

-. _- _

was

(m,

:

(dd,

IOHz,

Acetic forskalin at

0-5’C

concentrated. mp

146-14R’C.

THF for

dry

half

THF

(6

the

ml)

was

40

stirred

dried

mixture

and over

chromatography

1.36,

1.38

The

Chloro-

decomposed

vacua,

flash

was

added.

mins.

was

in

brine,

It

reaction

base

over

hour.

was

for

added

dist.

an ml)

continued to

(8

(freshly

by

extractanhydrous

using

5.03

(dd,

(s,

Jcis

vinylic-H),

5xCti3),

ethyl

2.02

5.55

(dd,

(s,

Jgem= 0.9Hr

= 10.5Hr, J 6,7

=

4Hz,

J5,6

vinylic-E). ether14

(2)

starting

(s,

SxCb),

12-C!),

2.02,

4.8

Jtrans

=

2.04

(dd,

17Hz,

(s,

J,is

=

Jgem=

2xCOCD),

IOHZ,

2Hz,

2.05

Jgem

q

~Hz,

vinyllc-H),

5.7

vinylic-E).

at

5.29

:

mmol),

25-2R°C. The

C,

H,

on

8.63.

60.28, J6,7

eluant

1.24,

q

Jgem

4Hz,

7a-Cl),

2Hz,

flash

acetic

into dried

ice over

chromatography

from

H,

and

poured

water,

by

68.89; (9. 80°C%d

washed

lo,

ethyl

acetate:

8.39%

1.7 (m,

water,

by

: (s,

dried

flash

mmol) The over

in drv residue

anhydrous

chromatography

using

25%. RxCti3),

vinylic-l_l),

5.68

0.083 a. 0.1 concentrated.

with

yield

1.36,

q

with

crystallised

purification

gave

1.28,

lOHz,

=

C,

errolether 6 h at was

on

washed

mmol)

was

(IO)

to

layer

0.58

purification was

Found

for

residue

as

0.96, Jcis

(d,

organic

The

(12:BB)

(dd,

was added was heated

g, mixture

was

which

i,

(0.1

reaction

layer

residue

gave

PTSA

The

organic

The

68.55;

The

acetate.

5.28

2.0,

5.26

6a-CH),

(dd,

5.84

2.08

(s,

Jtrans

(dd,

2xCOCti3),

= 18Hz,

Jtrans

=

Jgem

=

IBHz,

(II) THF

(270

=

5.32

(dd,

: 6 3.01

Jtrans

(dd, (1.0 mg)

and on

ISHz,

= IRHz,

mmol)

was

added

temperature. The

residue

was

concentrated. (15:85)

as

eluant.

1.43,

1.62

(s,

1.3, jem

•_‘~~:’

J,.:tz

a

stirred

was

The

solution

with

residue

ethyl

was

:

Yield 5~cTi3),

of

for

continued

extracted

and

J

to

Stirring

ether 0.9,

(d, q

Jtrans

I-Deoxyforskolin

poured

0.03

room

sulphate

CDCl3)

12-C!),

anhydride (50

mg, at

concentrated.

sodium

MHz,

5.88

and

ml)

acetate:petroleum

IRHz,

of

578

(2

was

over

ethyl

6a-CH),

Synthesis

and mg;

THF

in

and

by 1.27,

1.49

g,

h

0.21 mmol) mixture

4.92

dry

dried

PMR

added

17.5Hz,

5.08

2.0

10

concentrated.

mixture

using

(d, Jgem vinylic+),

in

water

ether14

fluoride

in

reaction

layer

vacua

organic

znylic-H).

mmol)

graphy

in

9.22%.

Excess of 2 h. was concentrated

0.9Hz,

=

2.2H2,

concentrated.

ether

Tetrabutylammonium the

ice

mixture

The

161

anhydrous refluxed

7bCfi),

I q

(2, for

C74H3606

6-Acetyl-I-deoxyforskGlinT4 0.03

was

0.99,

1.36,

(d,

(3:17) as eluant 1.6 g, mp 125-127°C.

ethyl and

12-CH), IOHz,

ZHz,

and

concentrated

: H,

mmol)

was

with

= ~Hz,

Jtrans

0.94,-1.26,

and

(0.021 a. The reaction

(COC13)

(s,

vinylic-p),

ml)

0.92,

dichloromethane.

acetate:petroleum

2Hz,

stirred reaction

sulphate

68.56;

mmol)

purified

17.5Hz,

stirred

for

with

4.64

was

6-Acetyl-1,9-dideoxy-II-enol-trimethylsilyl

:

sulphate

sulphate

a

The

dichloromethane. Yield

C,

stirring

washed

10.5Hz,

= 4Hz,

with

ml).

extracted

sodium

18Hz

7

vrnylrc-l).

(8)

Calcd.

Selenium

to

ml).

sodium

was

for mixture

6-Acetyl-I-deoxyforskolin-II-enol-trimethylsilyl pyridine

added (4

with

3.74

3.07

(3

temperature reaction

:

4.63

Jtrans

using ethyl acetste:hexane petroleum ether, yield Anal.

Jgem= Jgem

2Hz,

7.

1,9_dideoxyforskoiin

sodium

was

mmol)-in

and

q

:

0.27,

4.92

extracted

anhydrous

=

(8).

: 6

ml)

g,

The

3.75

prepared

9-CH),

(15

dry

3xSiCH3),

Jtrans (dd, J,_s

= 2.2Hz,

of

(d,

lO.RHz,

8.5%

solution

Found,

(0.79 ml, mixture

layer

gave

60.15

6-Acetyl-1,9_dideoxyforskolin A

in

at room ml) and the

12-C%),

was

(CDC13)

T2

(CHC13). 2.5

-

plates

acetate:petroleum

Jgem

H,

ether.

3.41

hexane,

temperature

mmol)

6-Acetyl-1,9_dideoxyforskolin

459 (d, vinyli&),

67.2;

anhydrous

9.15.

(1.16

concentrated.

(CDC13)

2.62

TLC

acetate:di-

11. H,

in

room

3.23

The

and (3:7)

vinylic-H), I 3.5Hz,-6W?j), from

ml,

to

by stirring acid (0.2

acetate.

PMR COCb),

g) (4,

(7)

sol

was

sulphate

acetate:hexane

M

1,9_dideoxyforskolin

(0.41

ethyl

1.6

and

followed of acetic

sodium

at layer

(55.6

gel

19.2O

COCHJ),

ISHz,

methanol

acetate:petroleum

hexamethyldisilazane The reaction atmosphere.

N2

mixture

at -7R°C addition

L)

residue

residue

ethyl -

Jcis

=

The with water. and extracted

of

under

-7ROC

reaction

ml,

solution

ml)

(dd,

a

22

The

hexane:ethyl

from

Jtrans

x

ether

silica

on

[a165

Found

in

0.51

dried

ethyl

prepared

(2.13

stirred

give

using

2.16

(3

vacua.

petroleum

1,9_dideoxyforskolin

value

crystsllised

6-Acetyl-7-deacetyl-7-trimethylsilyl-1,9-dideoxyforskolin~~ KRutyl

to crude

mp 174-176°C

(dd,

mmol

g,

diluted methanol

was

C&cd.

1.5

mg,

with

crystallised

Similarly, Anal.

in the

1280.

66.98;

30 mins at O°C and < 40°C to remove

was

5.34

benzene

and

gave

6a-Ctj),5.02

(m,

1760,

C,

I-deoxyforskolin

separated,

residue

;700, :

(s,

L)

(3)

sodium

solution

1.52

4.34

2

x

concentrated

was

roots)

7a-CH),

C22H3406

7-Deacetyl-I-deoxyforskolin A

12-Ctj),

concentrated

gel15 with

which

with

and 2

purified

of

1.5,

4.5Hz

q

2,

weight

1.36,

lElHz, 7

were

further

provide

dry

extracted

60-80°,

layers

was

1.22,

J

5.22 Anal.

of

was

column of silica have an impurity

to

(0.001%

kq) filtered

(bp

ether

a to

material

:60.96,

(d,

vinyiic-H),

on found The

(5 were

ether

petroleum

2.

(7:l:Z)

:

yield

material extgacts

petroleum

combined

that

chloromethane

(2) &nt

combined

usual purification The crude 4 was

9). close

ether,

the

triturated

decanted.

which

and.

1.96,

by

(14.0 an

acetate.

purified 90X,

s

half

hour

The flash

mg, and

organic chromate

mp 136’C. 2.02

(s,

~:,~~~6_H*j975,‘~p’(dJ,c~6,~

lo”,‘,;,

2xCOCH3),

2.44

J~$cE~,1~k9

lOHz,‘vinylic-!i_T.’

(2) ml)

was-added

pyridine crushed

(IO ice,

to

an

ml). extracted

The

ice-cooled

stirred

reaction with

mixture

dichloromethane.

solution was

stirred The

of

7-deacetyl-l-deoxyfor

organic

another layer

3 was

hours washed

Y. KHANDELWALet al.

766 with 10% HCl followed resldue was crystallised mix.

mp

175'C PMR

IZ-CH),

[aID

vinyiic-H), 5.84 (dd: J,is IR

~10.8

(KBr)

Anal.

cm-l

Calcd.

1.22,

18

Hz,

1.5,

1.52

4.34

(m,

60-CH),

5.34

(dd:

Hz,-7wCH),

Jtrans=

3500, for

1.36,

IZ-CH),

= 4.5

HZ,

sodium sulphate ether (1:l) yield

and

concentrated. The 75%, mp 174-176°C,

:

(CHC13).

:6 0.96, .I (JFJ,,,

(d, 5.22

water, dried over anhydrous from ethyl acetate:petroleum

- 18"

(CDC13)

3.06

by

IB HZ,

2940,

C22H3406

5xCy3),

2.16

5.02

(s,

(dd,

Jtrans

18

q

COCl3),

J,is

=

Hz,

2.5

10.8

Jgem

(d,

HZ,

:

Jgem=18

Jgem

2 Hz,

Hz,

= 2 HZ,

vlnylic-i),

vinylic-H).

1700, : C,

(s,

1360,

66.98;

12%. H,

8.69.

Found

C,

67.34;

H,

8.58%.

References 1.

N.

J.

de

Science 2.

A.

3.

K.

Soura J.

Yamakawa, II,

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J.

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G.

W.

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F.

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D.

T.

1981,

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Satoh,

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in

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Khandelwal,

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Welzel,

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5949.

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W.

Aretz,

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E.

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14.

Analytical

15.

S.

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D.

Corey Bhat,

Etittger and

8.

sample B.

S.

6. of

and

K.

Sauber,

Snider,

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H.

_, 9

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Dornauer

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N.

J.

compounds

Eajwa,

Am.

de

1972, not

1986,

367,

213.

9'1, 2549.

prepared.

Souza,

Tet.

Lett.,

1977,

1669.

1986,