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Absolute configurations of 24-hydroxycholesterol and related compounds
Tetrahedron
Ld,tera
ABSOLUTE
No. 26,
pp 2203 - 2206,
CONFIGURATIONS
1975.
Pergmon
OF 24-HYDROXYCHOLESTEROL
Naoyuki Koizumi, Masuo Morisaki Laboratory
of Chemistry
Technology,
Prces.
Meguro-ku,
Printed
in Great Britain.
AND RELATED COMPOUNDS'
and Nobuo Ikekawa*
for Natural Products, Tokyo Institute of Tokyo, Japan
Atsuo Suzuki and Toru Takeshita Teijin Institute of Biomedical Institute, (Received
Hino-shi,
synthesized
received
in UK for
and la,24,25-trihydroxy'-vitamin
24-hydroxy
stereoisomers,
the stereochemistry
As a consequence
la,24-dihydroxy4-
of those studies, the D3 which is one
of vitamin D3 has been concluded
suggesting
functional
D3 has been remained
to determine
as 24R, b
6
.
activity was observed with only one of the importance of 24-hydroxylation
of the most fundamental
analog,
obscure due to the uncertainty'
of the synthetic precursor,
urgent need therefore,
24-hydroxycholesterol'.
the absolute configurations
of
It is
of 24-hydroxy-
and related compounds.
24S,25-Epoxycholesterol determined
12 play 1975)
24,25-dihydroxy3-,
However, C-24 configuration
i.e. 24-hydroxyvitamin
cholesterol
D3.
some of the biological
of vitamin D6'7.
publication
of 24-hydroxy group in 24,25_dihydroxyvitamin
of the important metabolites Furthermore,
Research
a series of C-24 epimers 1 and 1 of active
forms of vitamin D, such as 24-hydroxy*-,
stereochemistry
and Central
Tokyo, Japan
in Japan 1 Hay 1975:
We have recently
Research
previously3,
benzoate(2)
whose configuration
was refluxed with LiA1H4-A1C13(3
at C-24 has been
: 1) in ether to give
OH R2
p-;“R1 HO-' 2203
2
a,
R1 = R2 = H
b,
R1 = H; R2 = OH
c,
R
d,
R1 = R2 = OH
1
= OH; R2 = H
No. 26
2204
25-hydroxycholesterol (55 %) and 24f+hydroxycholesterol(5a)(35 %). The 24-01 5a was isolated by chromatography and converted to the corresponding dibenzoate 5b During those procedures, epimerization at C-24 did not occur as evidenced from single peak of 5b on high pressure liquid chromatography(hplc) which resolves effectively C-24 isomers10 . By a similar method, 24R,25-epoxide A3 was led to dibenzoate 6b.
Each dibenzoates z
and 6b were identified respectively, with
the known' 3f3,24c1-dibenzoate(the more polar and higher mp isomer) and 36,24E2dibenzoate(the less polar and lower mp isomer), in respect of mp, tic and hplc. Thus, it is concluded that 24E1-hydroxycholesterol(cerebrostero1)is 24Shydroxycholesterol(5a) and 24c2-isomer is 24R-hydroxycholesterol(6a). -
__ A further confirmation was obtained by the modified Horeau's method"
applied to THP ethers 5e and 6e.
These were prepared from the corresponding
dibenzoate -5b and 6b by the following sequence : (1) a partial hydrolysis with 1.1 eqiv. KOH in methanol-THF to form the monobenzoate -5c and e,
(2) treatment
5d and s, with dihydropyran in CIi2C12containing tosyl acid to the THP ether (3) saponification with KOH in methanol-THF, affording 24S1-isomer 5e mp 157158.5O and 24E2-isomer e,
mp 154.5-155.5O.As shown in Fig. the configuration
of -5e and 6e were determined as 24S-OH and 24R-OH, respectively, which are fully consistent with the above results obtained by chemical interrelations. -6e
cyclohexanol
8 (min)
0
-5e
8
(min)
0
8 (min)
Fig. Gas liquid chromatograms of the (R)-a-phenylethylamidesof excess (-)-(R)- and (+)-(S)-a-phenylbutyricacid after acylation of cyclohexanol, 6e - and -5e. Apparatus: Shimadzu, IBM-PF gas chromatograph. Column : all glass capillary column coated with OV-17, 20 m X 0.25 mm i.d., at 210°.
NO. 26
2205
1 a, R = R2
b, CI
d, e,
I
H
R1 = R2 = Bz 1 = IX; R2 = Bz R R1 = THP; 1 = THP; R
R2
=
Bz
R2
=
OH
R1O Prom the above analogies, the stereochemistry of the analogous dibenzoates 1 which were the synthetic precursor of la,24E1- and la,24E2-dihydroxyvitamin D3 (& and %I4
can be deduced : 24s and 24R configurations may be assigned to
the more polar(245l) and the less polar(24c2) compounds, respectively. Carbon13 nmr analysis12 of a series of C-24 epimers(Table) supported those supposition.
It can be seen that signals of C-20, -21 and -24 of 24S-OH isomers
always appeared at a lower field than those of 24R-OH isomers, presenting an useful diagnostic method for differentiation of 24-OH epimers. It has now been established that 24El-hydroxy- and 19,246'-dihydroxyvitamin D3 (& and 2)
have 24S-OH and 24E2-hydroxy- and la,24S2-dihydroxyvitamin D3
(2 and -2c) have 24R-OH.
Table
13C Chemical shift (ppm)* OBz
R(6b) -
S(5b) -
c-20
35.34
35.63
35.34
35.63
35.35
36.04
c-21
18.64
18.93
18.64
18.74
18.55
18.95
C-24
79.21
79.60
79.21
79.60
80.76
81.74
* Recorded on PS/PFT-lOO(JEOL) in deuteriochloroform with tetramethylsilane as internal standard.
No. 26
It is interesting hydroxycholesterol,
to note that cerebrosterol
while the natural
24,25-dihydroxyvitamin
OH function and a series of 24R-OH-vitamin activity
has 24R-
D analogs exert higher biological
and Footnotes
This report is ,Part 23 in the series of "Studies on Steroids". N.Awata, M.Morisaki
2.
D3(E)
than 24S-OH congeners 6,7 . References
1.
isolated from brain is 24S-
N.Ikekawa,
and N.Ikekawa,
M.Morisaki,
N.Koizumi,
Biochem. Biophys. M.Sawamura,
Part 22:
Res. Comm.,
in press.
Y.Tanaka and H.F.DeLuca,
Biochem. Biophys. Res. Comm., 62, 485 (1975). . 3.
M.Seki,
N.Koizumi,
M.Morisaki
and N.Ikekawa,
Tetrahedron
Letters,
15
(1975). 4.
M.Morisaki,
N.Koizumi,
N.Ikekawa,
T.Takeshita
and S.Ishimoto,
J. Chem.
Sot. Perkin I, in press. 5.
N.Ikekawa, w.,
6.
7.
M.MorAsaki,
Y.Kato and T.Takeshita,
N.Koizumi,
Chem. Pharm.
23, 695 (1975).
Y.Tanaka,
H.F.DeLuca,
Biophys.,
submitted.
N.Ikekawa,
Y.Tanaka,
H.Frank, H.F.DeLuca,
M.Morisaki
N.Koizumi
and
Arch. Biochem.
N.Koizumi,
and N.Ikekawa,
Biochemistry,
submitted. 8.
The configuration
of 24-hydroxycholesterol'
(W.Klyne and W.M.Stokes, data.
However
, it was suggested
J. Pharm. Sci., 2, assignment
9. 10.
in 1970
in 1954
(J.E.VanLier and L.L.Smith,
719) albeit with an undefinite
Therefore, for the previous papers 2,4 .
should be reversed.
nomenclature'
has been determined
J. Chem. Sot., 1979) based on optical rotations evidence,
we have retained
that this the original
A. Ercoli and P. de Ruggieri, J. Am. Chem. Sot., 75, 3284 (1953). Retention
times of 24S- and 24R-isomers
when analyzed with Shimadzu-DuPont
were 10.1, 9.0 min., respectively,
830 Liquid Chromatograph.
Column,
Zorbax SIL (25 cm X 2.1 mm); mobile phase, 10 % CH2C12 in hexane; pressure, 60 kg/cm2; flow rate, 0.26 ml/mini 11.
detector, W
photometer.
c. J. W. Brooks and J. D. Gilbert, J. Chem. Sot. Chem. Comm., 194 (1973); J. D. Gilbert and C. .J. W. Brooks, Anal. Letters,
12.
by _. proton nmr5, this C-24 epimers of -5b-6b and 1.
Although C-24 epimers of g were distinguished technique could not differentiate
639 (1973).
Ld,tera
ABSOLUTE
No. 26,
pp 2203 - 2206,
CONFIGURATIONS
1975.
Pergmon
OF 24-HYDROXYCHOLESTEROL
Naoyuki Koizumi, Masuo Morisaki Laboratory
of Chemistry
Technology,
Prces.
Meguro-ku,
Printed
in Great Britain.
AND RELATED COMPOUNDS'
and Nobuo Ikekawa*
for Natural Products, Tokyo Institute of Tokyo, Japan
Atsuo Suzuki and Toru Takeshita Teijin Institute of Biomedical Institute, (Received
Hino-shi,
synthesized
received
in UK for
and la,24,25-trihydroxy'-vitamin
24-hydroxy
stereoisomers,
the stereochemistry
As a consequence
la,24-dihydroxy4-
of those studies, the D3 which is one
of vitamin D3 has been concluded
suggesting
functional
D3 has been remained
to determine
as 24R, b
6
.
activity was observed with only one of the importance of 24-hydroxylation
of the most fundamental
analog,
obscure due to the uncertainty'
of the synthetic precursor,
urgent need therefore,
24-hydroxycholesterol'.
the absolute configurations
of
It is
of 24-hydroxy-
and related compounds.
24S,25-Epoxycholesterol determined
12 play 1975)
24,25-dihydroxy3-,
However, C-24 configuration
i.e. 24-hydroxyvitamin
cholesterol
D3.
some of the biological
of vitamin D6'7.
publication
of 24-hydroxy group in 24,25_dihydroxyvitamin
of the important metabolites Furthermore,
Research
a series of C-24 epimers 1 and 1 of active
forms of vitamin D, such as 24-hydroxy*-,
stereochemistry
and Central
Tokyo, Japan
in Japan 1 Hay 1975:
We have recently
Research
previously3,
benzoate(2)
whose configuration
was refluxed with LiA1H4-A1C13(3
at C-24 has been
: 1) in ether to give
OH R2
p-;“R1 HO-' 2203
2
a,
R1 = R2 = H
b,
R1 = H; R2 = OH
c,
R
d,
R1 = R2 = OH
1
= OH; R2 = H
No. 26
2204
25-hydroxycholesterol (55 %) and 24f+hydroxycholesterol(5a)(35 %). The 24-01 5a was isolated by chromatography and converted to the corresponding dibenzoate 5b During those procedures, epimerization at C-24 did not occur as evidenced from single peak of 5b on high pressure liquid chromatography(hplc) which resolves effectively C-24 isomers10 . By a similar method, 24R,25-epoxide A3 was led to dibenzoate 6b.
Each dibenzoates z
and 6b were identified respectively, with
the known' 3f3,24c1-dibenzoate(the more polar and higher mp isomer) and 36,24E2dibenzoate(the less polar and lower mp isomer), in respect of mp, tic and hplc. Thus, it is concluded that 24E1-hydroxycholesterol(cerebrostero1)is 24Shydroxycholesterol(5a) and 24c2-isomer is 24R-hydroxycholesterol(6a). -
__ A further confirmation was obtained by the modified Horeau's method"
applied to THP ethers 5e and 6e.
These were prepared from the corresponding
dibenzoate -5b and 6b by the following sequence : (1) a partial hydrolysis with 1.1 eqiv. KOH in methanol-THF to form the monobenzoate -5c and e,
(2) treatment
5d and s, with dihydropyran in CIi2C12containing tosyl acid to the THP ether (3) saponification with KOH in methanol-THF, affording 24S1-isomer 5e mp 157158.5O and 24E2-isomer e,
mp 154.5-155.5O.As shown in Fig. the configuration
of -5e and 6e were determined as 24S-OH and 24R-OH, respectively, which are fully consistent with the above results obtained by chemical interrelations. -6e
cyclohexanol
8 (min)
0
-5e
8
(min)
0
8 (min)
Fig. Gas liquid chromatograms of the (R)-a-phenylethylamidesof excess (-)-(R)- and (+)-(S)-a-phenylbutyricacid after acylation of cyclohexanol, 6e - and -5e. Apparatus: Shimadzu, IBM-PF gas chromatograph. Column : all glass capillary column coated with OV-17, 20 m X 0.25 mm i.d., at 210°.
NO. 26
2205
1 a, R = R2
b, CI
d, e,
I
H
R1 = R2 = Bz 1 = IX; R2 = Bz R R1 = THP; 1 = THP; R
R2
=
Bz
R2
=
OH
R1O Prom the above analogies, the stereochemistry of the analogous dibenzoates 1 which were the synthetic precursor of la,24E1- and la,24E2-dihydroxyvitamin D3 (& and %I4
can be deduced : 24s and 24R configurations may be assigned to
the more polar(245l) and the less polar(24c2) compounds, respectively. Carbon13 nmr analysis12 of a series of C-24 epimers(Table) supported those supposition.
It can be seen that signals of C-20, -21 and -24 of 24S-OH isomers
always appeared at a lower field than those of 24R-OH isomers, presenting an useful diagnostic method for differentiation of 24-OH epimers. It has now been established that 24El-hydroxy- and 19,246'-dihydroxyvitamin D3 (& and 2)
have 24S-OH and 24E2-hydroxy- and la,24S2-dihydroxyvitamin D3
(2 and -2c) have 24R-OH.
Table
13C Chemical shift (ppm)* OBz
R(6b) -
S(5b) -
c-20
35.34
35.63
35.34
35.63
35.35
36.04
c-21
18.64
18.93
18.64
18.74
18.55
18.95
C-24
79.21
79.60
79.21
79.60
80.76
81.74
* Recorded on PS/PFT-lOO(JEOL) in deuteriochloroform with tetramethylsilane as internal standard.
No. 26
It is interesting hydroxycholesterol,
to note that cerebrosterol
while the natural
24,25-dihydroxyvitamin
OH function and a series of 24R-OH-vitamin activity
has 24R-
D analogs exert higher biological
and Footnotes
This report is ,Part 23 in the series of "Studies on Steroids". N.Awata, M.Morisaki
2.
D3(E)
than 24S-OH congeners 6,7 . References
1.
isolated from brain is 24S-
N.Ikekawa,
and N.Ikekawa,
M.Morisaki,
N.Koizumi,
Biochem. Biophys. M.Sawamura,
Part 22:
Res. Comm.,
in press.
Y.Tanaka and H.F.DeLuca,
Biochem. Biophys. Res. Comm., 62, 485 (1975). . 3.
M.Seki,
N.Koizumi,
M.Morisaki
and N.Ikekawa,
Tetrahedron
Letters,
15
(1975). 4.
M.Morisaki,
N.Koizumi,
N.Ikekawa,
T.Takeshita
and S.Ishimoto,
J. Chem.
Sot. Perkin I, in press. 5.
N.Ikekawa, w.,
6.
7.
M.MorAsaki,
Y.Kato and T.Takeshita,
N.Koizumi,
Chem. Pharm.
23, 695 (1975).
Y.Tanaka,
H.F.DeLuca,
Biophys.,
submitted.
N.Ikekawa,
Y.Tanaka,
H.Frank, H.F.DeLuca,
M.Morisaki
N.Koizumi
and
Arch. Biochem.
N.Koizumi,
and N.Ikekawa,
Biochemistry,
submitted. 8.
The configuration
of 24-hydroxycholesterol'
(W.Klyne and W.M.Stokes, data.
However
, it was suggested
J. Pharm. Sci., 2, assignment
9. 10.
in 1970
in 1954
(J.E.VanLier and L.L.Smith,
719) albeit with an undefinite
Therefore, for the previous papers 2,4 .
should be reversed.
nomenclature'
has been determined
J. Chem. Sot., 1979) based on optical rotations evidence,
we have retained
that this the original
A. Ercoli and P. de Ruggieri, J. Am. Chem. Sot., 75, 3284 (1953). Retention
times of 24S- and 24R-isomers
when analyzed with Shimadzu-DuPont
were 10.1, 9.0 min., respectively,
830 Liquid Chromatograph.
Column,
Zorbax SIL (25 cm X 2.1 mm); mobile phase, 10 % CH2C12 in hexane; pressure, 60 kg/cm2; flow rate, 0.26 ml/mini 11.
detector, W
photometer.
c. J. W. Brooks and J. D. Gilbert, J. Chem. Sot. Chem. Comm., 194 (1973); J. D. Gilbert and C. .J. W. Brooks, Anal. Letters,
12.
by _. proton nmr5, this C-24 epimers of -5b-6b and 1.
Although C-24 epimers of g were distinguished technique could not differentiate
639 (1973).