Biotransformation of terpenoids in mammal. I. Biotransformation of 3-carene and related compounds in rabbits


Ishida, T.; Asakawa, Y.; Okano, M.; Aratani, T.

Tetrahedron Letters 28: 2437-2440

1977


During the course of the investigation of the detoxication mechanism of terpenoids in mammals, we have found that some monoterpene hydrocarbons and a related compound to them have been stereoselectively transformed to hydroxylated products in rabbits. We report herein the biotransformation of 1-methylcyclohexene, (+)-, (-)- and (±)-2-pinenes, (-)-2(10)-pinene and (+)-3-carene.

Tetrahedron
Letters
No.
28,
pp
2437
2440,
1977.
Pergamon
Press.
Printed
in
Great
Britain.
BIOTRANSFORMATION
OF
TERPENOIDS
IN
MAMMAL.
I.
BIOTRANSFORMATION
OF
3-CARENE
AND
RELATED
COMPOUNDS
IN
RABBITS.
Takashi
ISHIDA
,
Yoshinori
ASAKAWA
,
Masayoshi
OKANO
+++
and
Takaaki
ARATANI
+14
+
Hiroshima
Institute
of
Technology,
Itsukaichi-cho,
Saeki-gun,
Hiroshima
738,
Japan
++
Institute
of
Pharmacognosy,
Tokushima-Bunri
University,
Yamashiro-cho,
Tokushima
770,
Japan
+++
Faculty
of
Integrated
Arts
and
Sciences,
Hiroshima
Universtiy,
Hiroshima
730,
Japan
(Received
in
Japan
7
May
1977;
received
in
UK
for
publication
31
may
1977)
During
the
course
of
the
investigation
of
the
detoxication
mechanism
of
terpenoids
in
mammals,
we
have
found
that
some
monoterpene
hydrocarbons
and
a
related
compound
to
them
have
been
stereoselectively
transformed
to
hydroxylated
products
in
rabbits.
We
report
herein
the
biotransformation
of
1-methylcyclohexene,
(+)-,
(-)-
and
(±)-2-pinenes,
(-)-2(10)-pinene
and
(+)-3-carene.
Each
compound
was
administered
by
a
stomach
tube
as
the
suspension
of
Tween-80
(100
ml)
to
an
unanesthetized
male
rabbit
for
four
days
after
two
days
starvation.
The
urine
was
treated
according
to
the
method
of
LUU
Bang
et
al.
1)
The
ether
extract,
after
removal
of
acidic
and
phenolic
fractions,
gave
a
neutral
fraction.
1-Methylcyclohexene
(10
g),
which
has
a
common
partial
structure
to
those
of
some
cyclic
monoterpene
hydrocarbons,
was
metabolized
and
gave
two
alcohols
(973
mg).
After
fractionation
with
column
chromatography
on
Si0
2
,
the
main
alcohol,
3-methy1-2-cyclohexen-l-
o1
2)
(6.8
%)
3)
(1)
was
isolated.
The
absolute
configuration
of
C-1
of
1
was
deduced
by
the
Horeau's
method
to
be
S.
4)
In
addition
to
1,
a
small
amount
of
2-methy1-2-cyclohexen-1-o1
5)
(2)
was
obtained
as
the
metabolite.
2-Pinenes
were
administered
and
the
main
metabolite,
verbenol
6)
(3)
was
obtained
as
shown
in
Table
1.
A
trace
of
myrtenol
7)
(4)
was
also
identified
in
each
case.
(-)-2(10)-Pinene
([ct]
D
-19.8°,
12
g)
gave
753
mg
of
neutral
metabolites.
Column
chromatography
on
Si0
2
of
this
part
separated
four
alcohols,
trans-pinocarveol
8)
(5,
0.7
%),
trans-10-pinanol
9)
(6,
2.4
%),
a-terpineol
(7,
[a]
c
,
-51.0
°
,
0.3
%)
and
1-2
.
-menthen-7,8-diol
(8,
[a]
p
-33.3°,
1.9
%).
(+)-3-Carene
([a]
0
+20.7°,
10
g)
gave
83
mg
of
neutral
metabolites,
which
gave
three
alcohols
on
SiO
2
chromatography.
The
main
alcohol
was
identified
as
m-mentha-4,6-dien-8-01
10)
(9,
11.2
%)
by
the
spectra
of
MS,
UV,
NMR,
NMDR
and
Eu(FOD)
3
experiment
and
by
the
formation
2437
2438
No.
28
Table
1.
2-pinene
weight
of
[a]
D'
administered
weight
neutral
metabolite
verbenol
*
yield
-30.9°,
8g
1.423
g
-31.2°,
17.6
%
+22.0°,
11)
10
g
0.318
g
t
0.0°,
11)
2.1
%
±
0.0°,
10
g
1.199
g
-11.3°,
8.1
%
*
Yield
:
verbenol
/
2-pinene
of
an
adduct
with
maleic
anhydride.
This
alcohol
has
not
been
found
as
a
natural
product.
The
minor
alcohol
was
determined
as
the
structure
10
(1.6
%)
on
the
basis
of
IR,
NMR
and
MS
spectral
data
and
also
proved
by
the
chemical
correlation.
The
spectra
of
IR
and
NMR
of
10
showing
the
absorption
at
3350
cm
and
a
singlet
of
two
protons
at
3.33
ppm,
respectively
suggest
this
alcohol
to
be
primary.
This
was
further
supported
by
the
spectra
of
its
acetate
at
1720
cm
-1
and
a
sharp
singlet
at
3.80
ppm.
The
NMR
spectrum
of
10
also
posseses
two
methine
protons
(1.00
-
0.73
ppm)
on
the
cyclopropane
ring.
The
chemical
shifts
of
C-8
and
C-9
methyl
groups
of
3-carene
are
0.80
and
1.07
ppm,
respectively,
12)
while
that
of
one
tertiary
methyl
group
of
10
resonanced
at
0.88
ppm,
indicating
the
maintenance
of
C-8
methyl
group
and
the
location
of
hydroxyl
group
to
be
at
C-9.
Thus,
the
structure
of
this
alcohol
was
established
to
be
3-caren-9-ol.
3-Caren-9-ol
has
neither
been
prepared
by
in
vitro
oxidation
of
3-caren
nor
found
in
nature.
The
third
alcohol
was
found
in
a
trace
amount
and
identified
as
m-cymen-8-ol
13)
(11).
In
the
case
of
1-methylcyclohexene
and
2-pinenes,
an
endo-cyclic
allylic
oxidation
occurred
selectively
rather
than
exo-cyclic
one.
The
metabolism
of
2-pinenes
suggests
that
rather
(-)-2-pinene
is
well
transformed
to
(-)-verbenol
than
(+)-2-pinene.
In
3-carene
we
could
not
detect
the
allylic
oxidation
products,
3-caren-2-ol
or
3-caren-5-ol
but
it
may
be
reasonable
that
the
rearrangement
of
either
of
these
alcohols
gave
m-mentha-4,6-dien-8-o1
14)
,
which
again
might
aromatize
to
m-cymen-8-ol.
The
formation
of
3-caren-9-ol
shows
the
first
example
of
the
stereoselective
hydroxylation
of
the
gem-dimethyl
group
on
the
cyclopropane
ring.
The
similar
biooxidation
of
992-dimethY1
group
has
been
known
in
the
metabolites
of
(+)-camphor
15)
and
retinoic
acid
15)
in
mammals.
Recently,
Renwick
et
a1.
17)
and
Southwe1l
18)
reported
the
metabolites
of
pinenes
in
a
bark
beetle
and
in
koala,
respectively.
On
the
basis
of
the
comparison
of
these
metabolites
with
our
results,
it
is
considered
that
the
detoxication
processes
of
pinenes
in
insect
and
in
each
mammal
are
considerably
different.
Moreover
it
is
interesting
that
3-methyl-2-cyclo-
hexen-l-ol
and
verbenol
have
been
found
as
pheromones
of
bark
beetles
2)
'
19)
and
we
are
currently
investigating
this
point.
Ackowledgement
:
The
authors
wish
to
express
their
thanks
to
Dr.
Yoshio
Hirose,
The
Institute
of
Food
Chemistry
for
the
MS
measurement
and
to
Yasuhara
Perfumary
Co.
Ltd.
for
the
administered
monoterpene
hydrocarbons.
H
OH
H
OH
S
HO
OH
1
2
OMB
OMNI;
CH
2
OH
OH
3
4
S
OH
of•
8
8
OEM
4
OH
S
S
CH
2
OH
9
10
II
OH
8
Alcohol
1
:
MS
M
112(C
7
H
12
0);
NMR(90MHz)
5.47(HC-2),
4.12(HCOH), 2.36(OH),
1.68(H
3
C-7);
IR(neat)
3350-1160-1140-10750-1060-
1030-990-855-810;
M
D
+1.64°;
Acetate
:
MS
re
154(C
9
H
14
0
2
);
NMR
5.37(HC-2),
5.17(HC-0Ac),
2.00(0Ac),
1.67(H
3
C-7).
Alcohol
2
:
NMR(90MHz)
5.56(HC-3),
3.96(HCOH),
1.70(H
3
C-7).
Alcohol
3
:
NMR
5.35(HC-3),
4.28(HCOH),
1.72(H
3
C-10),
1.33
&
0.87(H
3
C-8
&
H
3
C-9);
Acetate
;
NMR
5.35(HC-3),
2.03(0Ac),
1.73
(H
3
C-10),
1.34
&
0.93(H
3
C-8
&
H
3
C-9).
Alcohol
5
:
NMR
5.00
&
4.80(H
2
C-10),
4.45(HC-3),
1.32
&
0.67(H
3
C-8
&
H
3
C-9);
M
D
+28.2°.
Alcohol
6
:
MS
139(e-15);
NMR
3.50(H
2
C-10),
1.26
&
0.95(H
3
C-8
&
H
3
C-9);
[4
1
;
-21.4°.
Alcohol
8
:
7-Acetate-8-ol
:
NMR
5.75(HC-2),
4.48(H
2
C-7),
2.08(0Ac),
1.20(H
3
C-9
&
H
3
C-10).
Alcohol
9
:
MS
M
+
152(C
10
H
16
0);
UV
XmaxEt0H
264nm(log
c
2.95);
M
0
-116.1°;
NMR
(90MHz)
5.84,
5.64
&
5.57(HC-4,
HC
-5
&
HC
-6),1.80
(H
3
C-7),
1.20(H
3
C-8
&
H
3
C-9);
Eu(F0D)
3
(52.0mmol,
Ad-ppm)
HC
-3(2.57),
H
2
C-2(1.59
&
1.35),
H
3
C-9
&
H
3
C-10(2.08,
2.01).
Alcohol
10
:
NMR
5.27(HC-4),
3.33(H
2
C-9),
2.17(OH),
1.62(H
3
C-10),
0.88(H
3
C-8);
IR(CHC1
3
)
3350-1430-1370-1240-1020-820-780.
Acetate
:
MS
M
+
194.
(C12H102);
NMR(90MHz)
5.22(HC-4),
3.80(H
2
C-9),
2.04(0Ac),
1.60(H
3
C-10),
0.84(H
3
C-8).
Alcohol
11
:
NMR
7.40-6.35(aromatic
4H),
2.35(H
3
C-7),
1.55(H
3
C-8
&
H
3
C-9).
`O
2440
No.
28
Refernces
and
Notes
1)
LUU
Bang,
and
G.
Ourisson,
Tetrahedron
Letters,
1975,
1881.
LUU
Bang,
G.
Ourisson
and
P.
Teissire,
Tetrahedron
Letters,
1975,
4307.
LUU
Bang,
G.
Ourisson
and
P.
Teissire,
Tetrahedron
Letters,
1975,
2211.
By
this
method
neither
isomerization
nor
rearrangement
of
the
administered
compounds
occurred
during
the
incubation
and
extraction
process.
2)
J.P.
Vite,
G.B.
Pitman,
A.F.
Fentiman,Jr.
and
G.W.
Kinzer,
Naturwissenschaften,
59,
469
(1972).
3)
Percent
in
parentheses
means
the
yield
of
the
metabolite
to
the
administered
compound.
4)
A.
Horeau
and
H.
Kagan,
Tetrahedron,
20,
2431
(1964).
The
regenerated
a-phenylbutyric
acid
showed
[a]p
5
-1.53°
(c
2.74,
C0
6
).
5)
K.
Arata,
S.
Akutagawa
and
K.
Tanabe,
Bull.
Chem.
Soc.
Jpn,
48,
1097
(1975).
6)
K.
Mori,
Agr.
Biol.
Chem.,
40,
415
(1976).
7)
J.A.A.
Renwick,
P.R.
Huges
and
Tanletin
DeJ.
TY.,
J.
Insect
Physiol.,
19
1735
(1973).
8)
The
authors
are
indebted
to
Dr.
V.S.
Joshi,
National
Chemical
Laboratory,
Poona,
India
for
the
NMR
and
IR
spectra
of
(+)-trans-pinocarveol.
9)
B.M.
Mitzer,
V.J.Mancini,
S.
Lemberg
and
T.
Theimer,
Applied
Spectroscopy,
22,
34
(1968).
N.
Nakagawa
and
S.
Saito,
Tetrahedron
Letters,
1967,
1003.
10)
K.
Gollnick,
G.
Schade
and
G.
Schroeter,
Tetrahedron
22,
139
(1966).
11)
It
might
be
considered
that
the
low
value
of
[a]
cl
of
(+)-2-pinene
was
due
to
the
coexistence
of
(-)-2-pinene
and
the
negligible
[a]
p
of
verbenol
metabolized
from
(+)-2-pinene
was
due
to
the
(-)-verbenol
from
(-)-2-pinene.
12)
S.P.
Acharya,
Tetrahedron
Letters,
1966,
4117.
13)
W.D.P.
Burns,
M.S.
Carson,
W.
Cocker
and
P.V.R.
Shannon,
J.
Chem.
Soc.
(C),
1968,
3073.
14)
D.A.
Baines
and
W.
Cocker,
J.
Chem.
Soc.
Perkin
1,
1975,
2232.
15)
Y.
Asahina
and
M.
Ishidate,
Ber.
Disch.
Chem.
Ges.,
68B,
947
(1935).
16)
R.
Hanni,
F.
Bigler,
W.
Meister
and
G.
Fungert,
Hely.
Chim.
Acta,
59,
2221
(1976).
17)
J.A.A.
Renwick,
P.R.
Huges
and
I.S.
Krull,
Science,
191,
199
(1976).
18)
I.A.
Southwell,
Tetrahedron
Letters,
1975,
1885.
19)
J.A.A.
Renwick,
Contrib.
Boyce
Thompson
Inst.,
23,
355
(1967).