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The metabolic role of the 24-ethylidenecholesterols
BIOCHEMICAL
Vol. 31, No. 4, 1968
THE METABOLIC R.T.
ROLE OF THE 24-ETHYLIDENECHOLESTEROLS*
van Aller,
H. Chikamatsu,
J.P. Department
of Chemistry,
tain
epimeric
species
Kishida which
they
(1963);
arise
Bader,
and Lederer
Knights
Lederer
(1964))
to a 24-methylenesterol.
already
that
fucosterol
of two methyl
(1964, Blondin
1965);
groups
Mercer
and Nes (in
lar
to the known
Steinberg
Goad,
by reduction reduction
of the
(1960);
Avigan
should
the
We would
compounds. and
(Castle, (1964);
Barbier
Q-sitosterol
to report
from Pinus
generally here
pinea,
the
group
algae the
it
is
addi-
and Lederer (1966);
however
Hickey (1961)).
Barbier,
with
nucleotide
and Steinberg quite
Audier,
and Goodwin
(Stokes,
Birch
of brown
(Villanueva,
could
in
of the methyl
mevalonate
A 24 -bond
like
Vetter,
via
pyridine
Hayatsu,
The manner
species
Hammam, Dennis
be present
Tsuda,
the addition
arise
from cer-
and Nes (1963);
HGgel,
In various
with
(1934);
Blondin
to involve
isolated
(1967)).
24-Ethylidenesterols
the 24-ethylidenesterols
29-isofucosterol
Baisted
does indeed
press)).
and Avigan
24-ethyl
(1965);
from methionine
(1965);
to the 24-ethylsterols
have been and Wright
and Arigoni
of methionine
tion
38677
Phipers
has been postulated Guglielmetti
de Souza,
of Mississippi,
Mississippi
(Heilbron,
(1958);
(1964);
known
University
24-ethylidenecholesterols
of plants
and Akagi
N.J.
John and W.R. Nes
University,
Several
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Castle,
also
give
in a manner and Fish If
this
in plants isolation
the biosynthesis
rise simi-
(1958); were
true,
which
form
of both of
*Inquiries regarding this paper should be addressed to Prof. W. R. Nes, Sciences, Drexel Institute of Technology, Philadelphia, Dept. of Biological Pa. 19104. The support of grant No. 5ROl-AM09100 from the National Institutes of Health is gratefully acknowledged.
842
BIOCHEMICAL
Vol. 31, No. 4, 1968
29-isofucosterol
from mevalonate
lenecholesterol,
and its
reduction.
The data
conversion
obtained
to 24-ethylidene also
Castle,
are
Blondin
(which
a category
with
a genetic
block
*24(28)
_ and
the yeasts
A7-bonds
Gas-liquid
(which
cholesterol.
m.p.
127-130°,
distinguishes acetate,
tical
with
faster
Incubation
thin
than
a radioactive layer
synthetic
from
of silica
gel
on a thin
alcohol
29-isomer (lacking
Authentic
The ketone of silica
yielded
29-isofucosterol
identified
by AgN03 -thin
843
acetate 6.0 mg. from 12.25
(1960)),
coat)
with
movements (from
latter
and g. l.c.
layer
idenmoved
procedures.
of the acetate without
(126.5-127.5’))
20) with
(as the
(as
Fucus)
acetate
Similar
Tween
which
on a AgNC3-
sample
by m.p.
350
2-C 14 -mevalonate
as the acetate
24-ketocholesterol
P layer
thin
XE-60)
an authentic
(in
28-C14 -24-methylenecholesterol
impregnated
Amax
the
was identified gel,
8 -sitosterol.
sterol,
in both outer
as did
with
fucosterol
chromatographing
precisely
137-140°,
29-isofucosterol
(silanized,
their
by the usual
m.p.
nitrate
(Dusza
(synthetic)
the
30% of 24-methyl-
129-131°,
fucosterol
and g.1.c.
Ozonization
layer
on silver
8% of total
the free
substance
of radioactivity.
movement
of
of the seeds
of 29-isofucosterol. loss
contained
have
sterol.
plates,
mixture
to be in
of which
was obtained
also
the sample
appear
investigated,
identical
29-isofucosterol. its
there
to 24-methyl alternatives).
both
spectra
sterol
for
thus
and n.m.r.
AgN03-impregnated) synthetic
(1967)
in the dominant pinea
constituent,
29-isofucosterol
the
slightly
m.p.
24-methylene
organisms
chromatography
minor
AZ4 to 24-methylene
colorless
acetylated
as a very
the pathway:
procedures
showed layer
reduced
of Pinus
infrared
of the crude
was isolated
yielded
seeds
From thin
gel
seeds,
found
by
to 24-methylcholes-
A 5,7 -sterol)
In most
24-methy-
was converted
and Lederer
accrrmulate
from
presumably
the conversion:
step.
are
chromatography
formation
the 24-ethylidenesterol)
at a reductive
(CRC13),
with
and with
and chromatographic
-33.2’
silica
consistent
accumulate
its
to 24-ethylcholesterol
and Nes (1963)
From the ungerminated extraction
species,
similarly
to 24-ethyl
Fucus species
Wp
in this
24-Methylenecholesterol
terol.
(see
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
incubation the
acetate)
seeds
I.R., of yielded
and gas-liquid
BIOCHEMICAL
Vol. 31, No. 4, 1968
Refncubatfon
chromatography. gave
e-sitosterol
recovered
as unchanged
failed
By contrast,
labelled
highly
substance
(Russell,
van Aller
methylenecholesterol terol
was obtained,
saturated
portion
The labelled as did
to undergo
and Nes (in
identified
as is
press)).
From
layer
osmium
converted
to
tetroxide.
24-methylenecholesterol
incubation
of 28-C 14-24-
labelled
24-methylcholes-
to 29-isofucosterol
of the AgN03-thin
and when
to a chromatographically
labelled the
being
chromatographed
with
was converted
by gas-liquid
radioactivity
sample
a reaction
technique
in addition
29-isofucosterol
k-sitosterol
an authentic
29-isofucosterol by this
and purified
most of the remaining
substrate.
on AgNO -TLC) 3
3,5-cycloderivative
polar
the labelled
in 3.6% yield,
(as the acetate the
of
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
chromatography
of the side
chain
chromatogram.
References J. Avi.gan and D. Steinberg (1961)) J. Biol. Chem., 236, 2898. S. Bsder, L. Guglielmetti and D. Arigoni, (19641, Proc. Chem. Sot. (London), 16. D. J. Bafsted (1967)) private communication. A.J. Birch (1963). in T. Swain, editor, Chemical Plant Taxonomy, Academic Press, 162. M. Castle, G. Blondin and W.R. Nes (1963), J. Am, Chem. Sot., 85, 3306. M. Castle, G. Blondin and W.R. Nes (in press), J, Biol. Chem. J.P. Dusza (1960)) J. Org. Chem. , 25, 93. L.J. Goad, A.S.A. Hammam, A. Dennirand T.W. Goodwin (1966), Nature, 210, 1322. I. Heilbron, R.F. Phipers and H.R. Wright (1934)) J. Chem. Sot., 1572. M.F. H$el, N. Vetter, H. Audier, M. Barbier and E. Lederer (1964), Phytochem., 2, 7. B.A. Knights (1965)) Phytochem., 4, 857. E. Lederer (1964), Biochem. J., 93, 449. E. Lederer (1967)) Tetrahedron Letters, 12, 1049. E.J. Mercer (1965)) Bfochem. J. 2, 17P. P.T. Russell, R.T. van Aller and W.R. Nes (in press), J. Biol. Chem, D. Steinberg and J. Avigan (1960), J. Biol. Chem., 235, 3127. W.M. Stokes, F.C. Hickey and W.A. Fish (1958), J. Bm. Chem., 232, 347. K. Tsuda, R. Hayatsu, Y. Kishida and S. Akagi (1958), J. Am. Chem. Sot., 80, 921. V.R. Villanueva, M. Barbier and E. Lederer (1964), Bull. Sot. Chim. France, 1423. V.R. Villanueva, M. Barbier and E. Lederer (1965), Beitr. Biochem. Physiol. Naturstoffen Festschr., 509.
844
Vol. 31, No. 4, 1968
THE METABOLIC R.T.
ROLE OF THE 24-ETHYLIDENECHOLESTEROLS*
van Aller,
H. Chikamatsu,
J.P. Department
of Chemistry,
tain
epimeric
species
Kishida which
they
(1963);
arise
Bader,
and Lederer
Knights
Lederer
(1964))
to a 24-methylenesterol.
already
that
fucosterol
of two methyl
(1964, Blondin
1965);
groups
Mercer
and Nes (in
lar
to the known
Steinberg
Goad,
by reduction reduction
of the
(1960);
Avigan
should
the
We would
compounds. and
(Castle, (1964);
Barbier
Q-sitosterol
to report
from Pinus
generally here
pinea,
the
group
algae the
it
is
addi-
and Lederer (1966);
however
Hickey (1961)).
Barbier,
with
nucleotide
and Steinberg quite
Audier,
and Goodwin
(Stokes,
Birch
of brown
(Villanueva,
could
in
of the methyl
mevalonate
A 24 -bond
like
Vetter,
via
pyridine
Hayatsu,
The manner
species
Hammam, Dennis
be present
Tsuda,
the addition
arise
from cer-
and Nes (1963);
HGgel,
In various
with
(1934);
Blondin
to involve
isolated
(1967)).
24-Ethylidenesterols
the 24-ethylidenesterols
29-isofucosterol
Baisted
does indeed
press)).
and Avigan
24-ethyl
(1965);
from methionine
(1965);
to the 24-ethylsterols
have been and Wright
and Arigoni
of methionine
tion
38677
Phipers
has been postulated Guglielmetti
de Souza,
of Mississippi,
Mississippi
(Heilbron,
(1958);
(1964);
known
University
24-ethylidenecholesterols
of plants
and Akagi
N.J.
John and W.R. Nes
University,
Several
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
Castle,
also
give
in a manner and Fish If
this
in plants isolation
the biosynthesis
rise simi-
(1958); were
true,
which
form
of both of
*Inquiries regarding this paper should be addressed to Prof. W. R. Nes, Sciences, Drexel Institute of Technology, Philadelphia, Dept. of Biological Pa. 19104. The support of grant No. 5ROl-AM09100 from the National Institutes of Health is gratefully acknowledged.
842
BIOCHEMICAL
Vol. 31, No. 4, 1968
29-isofucosterol
from mevalonate
lenecholesterol,
and its
reduction.
The data
conversion
obtained
to 24-ethylidene also
Castle,
are
Blondin
(which
a category
with
a genetic
block
*24(28)
_ and
the yeasts
A7-bonds
Gas-liquid
(which
cholesterol.
m.p.
127-130°,
distinguishes acetate,
tical
with
faster
Incubation
thin
than
a radioactive layer
synthetic
from
of silica
gel
on a thin
alcohol
29-isomer (lacking
Authentic
The ketone of silica
yielded
29-isofucosterol
identified
by AgN03 -thin
843
acetate 6.0 mg. from 12.25
(1960)),
coat)
with
movements (from
latter
and g. l.c.
layer
idenmoved
procedures.
of the acetate without
(126.5-127.5’))
20) with
(as the
(as
Fucus)
acetate
Similar
Tween
which
on a AgNC3-
sample
by m.p.
350
2-C 14 -mevalonate
as the acetate
24-ketocholesterol
P layer
thin
XE-60)
an authentic
(in
28-C14 -24-methylenecholesterol
impregnated
Amax
the
was identified gel,
8 -sitosterol.
sterol,
in both outer
as did
with
fucosterol
chromatographing
precisely
137-140°,
29-isofucosterol
(silanized,
their
by the usual
m.p.
nitrate
(Dusza
(synthetic)
the
30% of 24-methyl-
129-131°,
fucosterol
and g.1.c.
Ozonization
layer
on silver
8% of total
the free
substance
of radioactivity.
movement
of
of the seeds
of 29-isofucosterol. loss
contained
have
sterol.
plates,
mixture
to be in
of which
was obtained
also
the sample
appear
investigated,
identical
29-isofucosterol. its
there
to 24-methyl alternatives).
both
spectra
sterol
for
thus
and n.m.r.
AgN03-impregnated) synthetic
(1967)
in the dominant pinea
constituent,
29-isofucosterol
the
slightly
m.p.
24-methylene
organisms
chromatography
minor
AZ4 to 24-methylene
colorless
acetylated
as a very
the pathway:
procedures
showed layer
reduced
of Pinus
infrared
of the crude
was isolated
yielded
seeds
From thin
gel
seeds,
found
by
to 24-methylcholes-
A 5,7 -sterol)
In most
24-methy-
was converted
and Lederer
accrrmulate
from
presumably
the conversion:
step.
are
chromatography
formation
the 24-ethylidenesterol)
at a reductive
(CRC13),
with
and with
and chromatographic
-33.2’
silica
consistent
accumulate
its
to 24-ethylcholesterol
and Nes (1963)
From the ungerminated extraction
species,
similarly
to 24-ethyl
Fucus species
Wp
in this
24-Methylenecholesterol
terol.
(see
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
incubation the
acetate)
seeds
I.R., of yielded
and gas-liquid
BIOCHEMICAL
Vol. 31, No. 4, 1968
Refncubatfon
chromatography. gave
e-sitosterol
recovered
as unchanged
failed
By contrast,
labelled
highly
substance
(Russell,
van Aller
methylenecholesterol terol
was obtained,
saturated
portion
The labelled as did
to undergo
and Nes (in
identified
as is
press)).
From
layer
osmium
converted
to
tetroxide.
24-methylenecholesterol
incubation
of 28-C 14-24-
labelled
24-methylcholes-
to 29-isofucosterol
of the AgN03-thin
and when
to a chromatographically
labelled the
being
chromatographed
with
was converted
by gas-liquid
radioactivity
sample
a reaction
technique
in addition
29-isofucosterol
k-sitosterol
an authentic
29-isofucosterol by this
and purified
most of the remaining
substrate.
on AgNO -TLC) 3
3,5-cycloderivative
polar
the labelled
in 3.6% yield,
(as the acetate the
of
AND BIOPHYSICAL RESEARCH COMMUNICATIONS
chromatography
of the side
chain
chromatogram.
References J. Avi.gan and D. Steinberg (1961)) J. Biol. Chem., 236, 2898. S. Bsder, L. Guglielmetti and D. Arigoni, (19641, Proc. Chem. Sot. (London), 16. D. J. Bafsted (1967)) private communication. A.J. Birch (1963). in T. Swain, editor, Chemical Plant Taxonomy, Academic Press, 162. M. Castle, G. Blondin and W.R. Nes (1963), J. Am, Chem. Sot., 85, 3306. M. Castle, G. Blondin and W.R. Nes (in press), J, Biol. Chem. J.P. Dusza (1960)) J. Org. Chem. , 25, 93. L.J. Goad, A.S.A. Hammam, A. Dennirand T.W. Goodwin (1966), Nature, 210, 1322. I. Heilbron, R.F. Phipers and H.R. Wright (1934)) J. Chem. Sot., 1572. M.F. H$el, N. Vetter, H. Audier, M. Barbier and E. Lederer (1964), Phytochem., 2, 7. B.A. Knights (1965)) Phytochem., 4, 857. E. Lederer (1964), Biochem. J., 93, 449. E. Lederer (1967)) Tetrahedron Letters, 12, 1049. E.J. Mercer (1965)) Bfochem. J. 2, 17P. P.T. Russell, R.T. van Aller and W.R. Nes (in press), J. Biol. Chem, D. Steinberg and J. Avigan (1960), J. Biol. Chem., 235, 3127. W.M. Stokes, F.C. Hickey and W.A. Fish (1958), J. Bm. Chem., 232, 347. K. Tsuda, R. Hayatsu, Y. Kishida and S. Akagi (1958), J. Am. Chem. Sot., 80, 921. V.R. Villanueva, M. Barbier and E. Lederer (1964), Bull. Sot. Chim. France, 1423. V.R. Villanueva, M. Barbier and E. Lederer (1965), Beitr. Biochem. Physiol. Naturstoffen Festschr., 509.
844