Total synthesis, elucidation of absolute stereochemistry, and adjuvant activity of trihydroxy fatty acids

Article abstract of DOI:10.1016/j.tet.2006.06.088

Pinellic acid from the tuber of Pinellia ternate, an active herbal component of the traditional Japanese herbal (Kampo) medicine Sho-seiryu-to, is a C18 trihydroxy fatty acid whose absolute stereochemistry has now been determined. All stereoisomers of pinellic acid were synthesized via regioselective asymmetric dihydroxylation, regioselective inversion, and stereoselective reduction in order to determine their absolute stereochemistries and adjuvant activities. Among this series of isomers, the (9S,12S,13S)-compound, which is a natural product, exhibited the most potent adjuvant activity. Spectral data for all of the stereoisomers of the 1,2-allylic diols were compared and related to their stereochemistries.

Full text of DOI:10.1016/j.tet.2006.06.088

Tetrahedron 62 (2006) 9483–9496
Total synthesis, elucidation of absolute stereochemistry,
and adjuvant activity of trihydroxy fatty acids
Tatsuya Shirahata,a,b Toshiaki Sunazuka,a,c Kiminari Yoshida,a,b Daisuke Yamamoto,a,b
Yoshihiro Harigaya,b Isao Kuwajima,c Takayuki Nagai,a,c Hiroaki Kiyohara,a,c
a,c,
Haruki Yamadaa,c and Satoshi Omura
ꢀ
*
aKitasato Institute for Life Sciences, Kitasato University, Shirokane, Minato-ku, Tokyo 108-8641, Japan
bSchool of Pharmaceutical Science, Kitasato University, Shirokane, Minato-ku, Tokyo 108-8641, Japan
cThe Kitasato Institute, Shirokane, Minato-ku, Tokyo 108-8642, Japan
Received 10 April 2006; accepted 16 June 2006
Abstract—Pinellic acid from the tuber of Pinellia ternate, an active herbal component of the traditional Japanese herbal (Kampo) medicine
Sho-seiryu-to, is a C18 trihydroxy fatty acid whose absolute stereochemistry has now been determined. All stereoisomers of pinellic acid were
synthesized via regioselective asymmetric dihydroxylation, regioselective inversion, and stereoselective reduction in order to determine their
absolute stereochemistries and adjuvant activities. Among this series of isomers, the (9S,12S,13S)-compound, which is a natural product,
exhibited the most potent adjuvant activity. Spectral data for all of the stereoisomers of the 1,2-allylic diols were compared and related to
their stereochemistries.
Ó 2006 Published by Elsevier Ltd.
1. Introduction
has been used clinically for the treatment of cold symptoms.
In preliminary studies SSTexhibited potent antiviral activity
against influenza due to an immunostimulating activity
against nasally inoculated influenza antigen. Our research
indicated that SST had oral adjuvant activity for nasally
administered influenza vaccine.2–4 It was clear that the activ-
ity of SST was due to ingredients from Pinellia ternate, one
of the component herbs of SST. Further investigation deter-
mined that pinellic acid 1 isolated from P. ternate was the
compound responsible for the adjuvant activity (Fig. 1).
Pinellic acid 1 is an effective oral adjuvant candidate for
nasal influenza vaccine; however, P. ternate contains only a
small amount of 1 and their stereochemistry was unknown.5
Although information about the stereochemistry of these
types of fatty acids has been reported,6 there were not
absolute to overcome our problems. Herein, not only the
enantioselective total synthesis and assignment of the stereo-
chemistry of 1, but also the synthesis of stereoisomers and
their adjuvant activities, are reported.
Infection with the influenza virus is epidemic and can be
lethal for patients with respiratory diseases and those who are
elderly.1 The primary method for the treatment of influenza
is to use the influenza vaccine as a prophylaxis. Subcutaneous
injection of this vaccine is known to induce production of
serum antiviral IgG antibodies (Abs) that give a protective
effect against proliferation of the virus in lung tissue.
Because the influenza virus infects the nasal cavity first,
intranasal inoculation of the influenza vaccine has been at-
tempted in order to increase its safety and prevent antigenic
variation. However, it has been shown that vaccinations in
the nasal cavity are less effective than subcutaneous ones and
may not provide sufficient immunostimulation. In order to
overcome these problems, using adjuvants for enhancement
of the local mucosal immune response has been reported.
Several traditional Japanese herbal (Kampo) medicines have
been used for the treatment of cold-like symptoms in which
the influenza virus is known to be the causative agent. Oral
administration of the Kampo medicine, Sho-seiryu-to (SST),
O
OH
OH
13
HO
9
12
OH
Pinellic acid (1)
Keywords: Total synthesis; Adjuvant; Determination of stereochemistry.
* Corresponding author. Tel.: +81 3 3444 6101; fax: +81 3 3444 6380;
e-mail: omura-s@kitasato.or.jp
Figure 1. Structure of pinellic acid 1.
0040–4020/$ - see front matter Ó 2006 Published by Elsevier Ltd.
doi:10.1016/j.tet.2006.06.088

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T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
OH
O
O
OCO-p-Br-Ph
a
O
O
b
O
MeO
MeO
1
O
2
3
Scheme 1. Derivatization of 1. Reagents and conditions: (a) (1) TMSCHN2, benzene/MeOH (10:1), rt, 2.5 h; (2) 2,2-dimethoxypropane, PPTS, CH2Cl2, 60 ꢁC,
48 h (100% from 1); (b) p-Br-BzCl, DMAP, pyridine, rt, 10 h (68%).
2. Estimation of absolute stereochemistry of 1
synthetic route to 4 and 5 in order to synthesize all of the pos-
sible stereoisomers.
To determine the absolute stereochemistry of pinellic acid,
spectral analysis of its derivatives provided insightful infor-
mation. The CD exciton method7 was used for the estimation
of C9 stereochemistry at the allylic alcohol. The esterifica-
tion of 1 followed by dimethylacetalization gave acetonide
2 with free alcohol at C9 (Scheme 1). Both coupling constant
(J12,13¼8.0 Hz) in the 1H NMR spectrum of 2 and NOE anal-
ysis indicate a syn configuration at the C12–C13 diol
(Fig. 2).
3. Total synthesis
3.1. Synthetic strategy
The strategic disconnection is outlined in Figure 5. The most
important challenge in this synthesis is to construct the
stereochemistry of the three hydroxy groups. The syn-diol
at C12–C13 would be prepared from a diene by regioselec-
tive asymmetric dihydroxylation,9 and the C12–C13
anti-diol would be constructed via regioselective protection
of the C12 hydroxy group followed by inversion of the C13
hydroxy group. Stereoselective reduction from the cor-
responding enone would give the allylic alcohol at C9.
The corresponding p-bromobenzoate 3 was prepared with
p-bromobenzoyl chloride from 2. The coupling constant
between H9 and H10 in the H NMR spectrum of 3 was
1
7.0 Hz, indicating an antiperiplanar conformation of these
two protons. Moreover, a positive Cotton effect [lmax (D3):
244.8 (+6.97), 220.8 (+2.13), 209.1 (+5.97) (MeOH)] of 3
in the CD spectrum suggested a 9S configuration8 (Fig. 3).
3.2. Synthesis of the C18 skeleton
The synthesis of C18 skeleton 11 utilizing dithiane cou-
pling10 is shown in Scheme 2. tert-Butyl ester 7 was con-
verted from the carboxylic acid moiety in suberic acid
monomethyl ester 6 with (Boc)2O and DMAP in t-BuOH.
The diester 7 was transformed to iodide 8 in good yield by
hydrolysis of the methyl ester, followed by reduction of
the carboxylic acid,11 and iodination of the resultant primary
alcohol. The C9–C18 skeleton 10 was derived from com-
mercially available 2,4-decadienal 9. Lithiation of 10 with
n-BuLi and subsequent addition of 8 gave diene 11 in high
yield (Scheme 2).
Based on these results, the absolute configuration of 1 was
determined to be either 4 (9S,12S,13S) or 5 (9S,12R,13R)
(Fig. 4). We then attempted to establish a convergent
H
1.2%
H
10
R1
12
O
11
O
1.3%
13
H
H
3.4%
R2
4.2%
3.3. Synthesis of 4
Differential NOE
The regioselective asymmetric dihydroxylation of 11 using
AD-mix containing (DHQ)2PHAL gave C12–C13 syn-diol
(ꢀ)-1212 in disappointing yield and enantiomeric excess
(55%, 80% ee). However, the use of modified Sharpless
ligand [(DHQ)PHAL(DHQ)Me+$Iꢀ]9 for the hydroxylation
resulted in 64% yield with 95% ee. The protection of the diol
(ꢀ)-12 with excess TBSOTf followed by the deprotection of
dithioacetal (ꢀ)-13 provided enone (ꢀ)-14.
Figure 2. NOE analysis of 2.
The stereoselective reduction of enone (ꢀ)-14 to provide the
(9S)-alcohol was attempted. Diastereoselectivity was not
achieved with NaBH4 or (R)-CBS13 (diastereoselectivity
5:1). (S)-BINAL-H12,14 noticeably improved the diastereo-
selectivity due to the p-electron at C10–C11 and the bulky
O-TBS group (diastereoselectivity >20:1). The desilylation
with TBAF gave triol (ꢀ)-15 as a single isomer. Since depro-
tection of tert-butyl ester with TFA caused elimination of
the hydroxy groups at the C9 and C12 allylic positions, the
hydrolysis of the tert-butyl ester was achieved by a highly
concentrated alkaline solution to afford (ꢀ)-4, which has the
9S,12S,13S configuration (Scheme 3). Compound (ꢀ)-4 was
Figure 3. CD spectrum of 3.

T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
9485
O
OH
OH
O
OH
OH
13S
13R
HO
HO
9S
12S
9S
12R
OH
OH
4
5
Figure 4. Possible structures for 1.
O
OH
13R
OH
O
OH
13S
Regioselective
asymmetric
Stereoselective
reduction
9S
9
12R
12R
9
O
OH
OR
dihydroxylation
Inversion
OH
O
OH
13S
O
OH
13R
dienone
12S
12S
9R
9
9
OH
12, 13-syn-diol
OR
12, 13-anti-diol
9-alcohol
Figure 5. Synthetic strategy for all stereoisomers of 1.
O
O
O
a
b
OMe
OMe
I
HO
t-BuO
t-BuO
O
O
6
7
8
O
O
S
S
S
S
c
d
H
t-BuO
H
9
10
11
Scheme 2. Synthesis of the C18 skeleton 11. Reagents and conditions: (a) (Boc)2O, DMAP, t-BuOH, rt, 1 h (82%); (b) (1) 0.1 N NaOH in THF/MeOH/H2O
(3:1:1), rt, 28 h; (2) BH3$THF, THF, 0 ꢁC to rt, 24 h; (3) I2, PPh3, imidazole, CH2Cl2, 0 ꢁC to rt, 2 h (77% from 7); (c) 1,3-propanedithiol, BF3$OEt2, CH2Cl2,
0 ꢁC to rt, 10 h (96%); (d) n-BuLi, THF, ꢀ78 ꢁC, 1 h, then 8, ꢀ78 ꢁC, 1 h (85%).
O
OH
13S
O
a
S
S
S
S
t-BuO
t-BuO
12S
OH
(–)-12
O
11
O
OTBS
13
O
OTBS
S
S
13S
b
c
t-BuO
12
t-BuO
12S
OTBS
OTBS
(–) -13
OH
(–)-14
OH
O
OH
O
OH
13S
d
e
t-BuO
HO
9S
12S
OH
9S
OH
(–)-4
(–)-15
Scheme 3. Synthesis of 4. Reagents and conditions: (a) (DHQ)PHAL(DHQ)Me+$Iꢀ, K3[Fe(CN)6], K2CO3, K2OsO4$2H2O, methanesulfonamide, t-BuOH/H2O
(1:1), 0 ꢁC, 41 h (64%, 95% ee); (b) TBSOTf, 2,6-lutidine, ꢀ78 ꢁC, 30 min (89%); (c) Hg(ClO4)2, CaCO3, THF/H2O (5:1), rt, 30 min (83%); (d) (1) (S)-
BINAL-H, THF, ꢀ78 ꢁC, 1 h (diastereoselectivity >20:1); (2) TBAF, THF, 70 ꢁC, 3 h [76% from (ꢀ)-14]; (e) 2.0 N KOH in EtOH/H2O (5:1), rt, 46 h (82%).
identical in all respects with natural product 1 [400 MHz 1H
NMR, 100 MHz 13C NMR, IR, HRMS, optical rotation
{[a]D25 ꢀ8.0 (c 0.30, MeOH); natural:4 [a]2D8 ꢀ8.1 (c 0.32,
MeOH)}, and oral adjuvant activity] (Scheme 4).15
with 92% ee (Scheme 4). After the installation of the diol,
the sequence of five reactions was the same, yielding (+)-5.
The trihydroxy fatty acid (+)-5 was not identical to natural
1
product 1 [400 MHz H NMR, 100 MHz 13C NMR, and
optical rotation {[a]2D3 +29.8 (c 0.45, MeOH)}].
3.4. Synthesis of 5
3.5. Synthesis of (D)-4 and (L)-5
For the synthesis of (+)-5, (+)-12 with absolute configuration
12R,13R was required. Following the synthetic route for
(ꢀ)-4, asymmetric dihydroxylation using AD-mix-b con-
taining (DHQD)2PHAL of 11 gave diol (+)-12 in 75% yield
In order to investigate the oral administration of pinellic acid
analogs as adjuvants for the intranasal inoculation of influ-
enza HA vaccine, the synthesis of enantiomers of (ꢀ)-4

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T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
O
OH
13R
O
a
S
S
S
S
t-BuO
t-BuO
12R
OH
(+)-12
O
11
O
OTBS
O
OTBS
S
S
13
13R
b
c
t-BuO
12
t-BuO
12R
OTBS
OH
OTBS
(+)-13
OH
(+)-14
OH
O
O
OH
13R
d
e
t-BuO
HO
9S
9S 12R
OH
OH
(+)-16
(+)-5
Scheme 4. Synthesis of 5. Reagents and conditions: (a) (DHQD)2PHAL, K3[Fe(CN)6], K2CO3, K2OsO4$2H2O, methanesulfonamide, t-BuOH/H2O (1:1), 0 ꢁC,
73 h (75%, 92% ee); (b) TBSOTf, 2,6-lutidine, ꢀ78 ꢁC, 30 min (87%); (c) Hg(ClO4)2, CaCO3, THF/H2O (5:1), rt, 30 min (83%); (d) (1) (S)-BINAL-H, THF,
ꢀ78 ꢁC, 1 h (diastereoselectivity >20:1); (2) TBAF, THF, 70 ꢁC, 3 h [76% from (ꢀ)-14]; (e) 2.0 N KOH in EtOH/H2O (5:1), rt, 46 h (76%).
and (+)-5 containing the syn configuration at C12–C13 is
required. To construct the C9 hydroxy group, (R)-BINAL-H
would be applied to the corresponding intermediates (+)-14
and (ꢀ)-14.12 As expected, stereoselective reduction (dia-
stereoselectivity >20:1) followed by deprotection of the
TBS group gave the (9R)-alcohol. Finally, hydrolysis accord-
ing to the above procedure furnished (+)-4 and (ꢀ)-5 success-
fully (Scheme 5).
in the C12–C13 syn-diol followed by inversion of the C13
hydroxy group. The preparation of 17 (9S,12R,13S) is
shown in Scheme 6. The protecting groups were selected
carefully because only one hydroxy group (C12 or C13)
should be protected. The C12 hydroxy group is more reac-
tive than C13 due to its allylic position, therefore, the
chemoselective protection of the C12 hydroxy group in
(+)-12 was attempted. Installation of a TBS group only
on the C12 hydroxy group was problematic even at low
temperature with slow addition of reagent (C12 O-TBS:
75%, C13 O-TBS: 18%). The use of the more bulky TIPS
group successfully provided C12 O-TIPS (ꢀ)-1812 in
good yield (90%), with no formation of the C13 O-TIPS
compound.
3.6. Synthesis of 17
The C12–C13 anti-isomers were also constructed in order
to investigate structure–activity relationships. The key
step is regioselective protection of the C12 hydroxy group
O
OH
9R
OH
13R
O
OH
OH
a
b
(+)-14
HO
t-BuO
12R
OH
9R
OH
(+)-15
(+)-4
O
OH
OH
O
OH
OH
13S
c
d
(–)-14
t-BuO
HO
12S
OH
9R
9R
OH
(–)-5
(–)-16
Scheme 5. Synthesis of (+)-4 and (ꢀ)-5. Reagents and conditions: (a) (1) (R)-BINAL-H, THF, ꢀ78 ꢁC, 1 h (diastereoselectivity >20:1); (2) TBAF, THF,
70 ꢁC, 3 h [63% from (+)-14]; (b) 2.0 N KOH in EtOH/H2O (5:1), rt, 46 h (67%); (c) (1) (R)-BINAL-H, THF, ꢀ78 ꢁC, 1 h (diastereoselectivity >20:1),
(2) TBAF, THF, 70 ꢁC, 3 h [57% from (ꢀ)-14]; (d) 2.0 N KOH in EtOH/H2O (5:1), rt, 46 h (51%).
O
OH
13R
O
O
OAc
13S
S
S
a
b, c
t-BuO
t-BuO
(+)-12
12R
12R
OTIPS
OAc
OTIPS
(–)-20
(–)-18
OH
O
O
OH
OH
d
e
13S
t-BuO
HO
9S
9S
12R
OTIPS
OH
(–)-21
(+)-17
Scheme 6. Synthesis of (+)-17. Reagents and conditions: (a) TIPSOTf, 2,6-lutidine, CH2Cl2, ꢀ78 ꢁC, 8 h (90%); (b) (1) ClCH2SO2Cl, pyridine, 0 ꢁC, 2 h; (2)
CsOAc, 18-crown-6, benzene, 80 ꢁC, 20 h (83%); (c) Hg(ClO4)2, CaCO3, THF/H2O (5:1), rt, 5 min (97%); (d) (S)-BINAL-H, THF, ꢀ78 ꢁC, 90 min (99%, dr
>20:1); (e) (1) 1.0 N KOH in EtOH/H2O (4:1), rt, 5 days; (2) TBAF, THF, rt, 45 h (94%).

T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
9487
Next, we attempted inversion of the hydroxy group at C13.
Failure of the normal conditions for Mitsunobu inversion16
(DEAD, benzoic acid, PPh3) necessitated the use of the
new Mitsunobu conditions17 (TMAD, p-nitrobenzoic acid,
PBu3), which gave the (13S)-compound in 55% yield.
Unfortunately, this method presents difficulties for large-
scale synthesis. We next attempted a stepwise reaction to
construct a leaving group before inversion by nucleophilic
attack. While a methanesulfonyl group would be ideal as a
leaving group, Corey’s conditions18 using K2O are too strong
to get an inversion product. Fortunately, a small conversion
with CsOAc provided an insight in the search for another
leaving group. On the basis of this information, Nakata’s
utilizing (ꢀ)-12 as the starting material (Scheme 7). It
should be noted that the stereoselectivity of the (S)-BINAL
reduction of (+)-20 was lower than that of (ꢀ)-20 (diastereo-
selectivity 13:1). The reason for this phenomenon is
explained by the steric hindrance of the C12 O-TIPS group.
With the completion of the syntheses for the C12–C13 anti-
diols as shown in Schemes 7 and 8, all the stereoisomers
of pinellic acid have now been prepared from their cor-
responding intermediates.12
4. Stereochemistry of the allylic 1,2-diol
method,19 using
a monochloromethanesulfonyl group
(ClSO2CH2Cl, pyridine) followed by treatment with CsOAc,
gave the protected C12–C13 anti-diol (ꢀ)-19 in good yield.
The syntheses of both allylic syn- and anti-1,2-diols of pine-
llic acid have been established and it is critical for the stereo-
chemistries of the C12–C13 diols to be confirmed. The
protection of the C12–C13 diol of (+)-12 with 2-methoxy-
propene and CSA afforded (+)-24. In the 1H NMR spectrum,
an NOE between H11 and H13 resonances is observed, sug-
gesting that H12 and H13 of (+)-24 are antiperiplanar
(Scheme 9, Fig. 6).
In order to derive the alcohol group from the ketone,
deprotection of the dithioacetal furnished enone (ꢀ)-20.
The stereoselective reduction of the ketone at C9 in
(ꢀ)-20 required the reoptimization of reaction conditions
due to the presence of the bulky O-TIPS group. While (R)-
CBS reduction furnished (ꢀ)-2112 in good selectivity (dia-
stereoselectivity 16:1), (S)-BINAL-H was found to
be more efficient12 (diastereoselectivity >20:1). Finally
deprotection of the acetyl and tert-butyl groups by hydrolysis
and desilylation with TBAF gave (+)-17 (Scheme 6).
Deprotection of the OAc and O-TIPS groups in (ꢀ)-19
afforded (+)-26. This was followed by acetalization of the
C12–C13 diol to give (+)-27. In the H NMR spectrum,
1
while there is an NOE between protons H11 and H12, there
is no NOE between protons H11 and H13, indicating that
H12 and H13 of (+)-27 are synperiplanar (Scheme 10,
Fig. 7).
3.7. Synthesis of the remaining stereoisomers
Synthesis of (+)-23 with 9S,12S,13R stereocenters was
accomplished according to the following synthetic route,
These studies have established a new method to determine
the configuration of allylic 1,2-diols.
O
OH
13S
O
O
OAc
S
S
13R
a
b, c
t-BuO
t-BuO
(–)-12
12S
12S
OTIPS
OAc
OTIPS
(+)-20
OH
(+)-18
OH
O
O
OH
d
e
13R
t-BuO
HO
9S
9S
12S
OH
OTIPS
(+)-22
(+)-23
Scheme 7. Synthesis of (+)-23. Reagents and conditions: (a) TIPSOTf, 2,6-lutidine, CH2Cl2, ꢀ78 ꢁC, 8 h (79%); (b) (1) ClCH2SO2Cl, pyridine, 0 ꢁC, 1 h; (2)
CsOAc, 18-crown-6, benzene, 80 ꢁC, 20 h (75%); (c) Hg(ClO4)2, CaCO3, THF/H2O (5:1), rt, 5 min (89%); (d) (S)-BINAL-H, THF, ꢀ78 ꢁC, 1 h (99%, dr 13:1);
(e) (1) 1.0 N KOH in EtOH/H2O (4:1), rt, 5 days; (2) TBAF, THF, 45 h (98%).
O
OH
OH
13S
O
OH
OAc
a
b
HO
t-BuO
(–)-20
9R
12R
OH
9R
OTIPS
(–)-23
(–)-22
O
OH
OAc
O
OH
OH
13R
c
d
t-BuO
(+)-20
HO
9R
12S
OH
9R
OTIPS
(–)-17
(+)-21
Scheme 8. Synthesis of (ꢀ)-23 and (ꢀ)-17. Reagents and conditions: (a) (R)-BINAL-H, THF, ꢀ78 ꢁC, 1 h (82%, dr 13:1); (b) (1) 1.0 N KOH in EtOH/H2O
(4:1), rt, 5 days; (2) TBAF, THF, 45 h (94%); (c) (R)-BINAL-H, THF, ꢀ78 ꢁC, 1 h (98%, dr >20:1); (d) (1) 1.0 N KOH in EtOH/H2O (4:1), rt, 5 days; (2) TBAF,
THF, 45 h (18%).

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OH
O
O
S
S
S
S
13R
a
O
O
t-BuO
t-BuO
12R
OH
(+)-12
(+)-24
Scheme 9. Synthesis of (+)-24. Reagents and conditions: (a) 2-methoxypropene, CSA, CH2Cl2, 0 ꢁC, 5 min (96%).
3.0%
H
R2
10
H
R1
H
12
10
O
R1
11
12
O
O
4.7%
11
O
4.5%
13
H
4.6%
13
R2
H
H
H
H
5.3%
2.0%
2.8%
Differential NOE
Figure 6. NOE analysis of (+)-24.
Differential NOE
Figure 7. NOE analysis of (+)-27.
5. Comparison of spectral data of all the stereoisomers
of pinellic acid
was investigated. Micewere orally administeredwithpinellic
acid analogs (1 g/mouse) using intragastric gavage followed
by the intranasal inoculation of HA vaccine (1 g/mouse).
Three weeks later, the same procedure was repeated. The
IgA and IgG antibody responses against anti-influenza virus
in the nasal cavity and serum in the vaccinated mice were
examined one week after vaccination. The results of the
adjuvant activity of all stereoisomers are shown in Figure 9.20
Comparison of the 1H NMR spectra of all synthetic stereo-
isomers of pinellic acid (Fig. 8) shows a relationship
between the stereochemistry and the pattern of the 1H
NMR resonances. Focusing on the peaks of C12–C13 diol,
the H13 proton in the syn-diol is at higher field than in the
anti-diol. Moreover, the peak patterns of H10 and H11 are
opposite in the anti- and syn-diols. The relationship between
the stereochemistry of C9 and C12 and the coupling pattern
of H10 and H11 is also interesting. When the configuration
of C9 and C12 is the same (S,S or R,R), the chemical shifts of
H10 and H11 (two doublet of doublets) are very close. When
the configuration is different, the chemical shifts of H10 and
H11 are further apart.
The antiviral IgA and IgG antibody responses, induced in the
nasal cavities of mice given pinellic acid (ꢀ)-1 with vaccine,
were enhanced 5.2- and 2-folds, respectively, compared with
control mice given the vaccine and solvent alone. Among the
C9 isomers of pinellic acid, the (9S)-compounds showed
much stronger activity compared with the (9R)-compounds.
Thus, stereochemistry at the C9 hydroxyl group is critical for
adjuvant activity. Among the (9S)-derivatives, the adjuvant
activities of the C13 (S)-compounds were stronger than
that of the C13 (R)-compounds, while the stereochemistry
of the C12 hydroxyl group was not important for adjuvant
activity. It is interesting that the adjuvant activity of the
enantiomer of natural pinellic acid is weaker than that of
the natural one.
This type of information could never have been discovered
until all the stereoisomers had been synthesized. Syntheses
of fatty acids like pinellic acid could contribute to the deter-
mination of stereochemistry of molecules of the same type
as 1.6
6. Adjuvant activity of all the stereoisomers of
pinellic acid
Also, in the data shown in Figure 9, the adjuvant activity of
pinellic acid (ꢀ)-1 from a natural source was lower than that
of the synthetic one. This result is presumably due to the
chemical purity of the available sample.
The oral administration of pinellic acid analogs as an adju-
vant for the intranasal inoculation of influenza HA vaccine
O
OAc
O
OH
13S
S
S
S
S
13S
a, b
t-BuO
t-BuO
12R
12R
OTIPS
OH
(–)-19
(+)-26
O
S
S
O
O
c
t-BuO
(+)-27
Scheme 10. Synthesis of (+)-27. Reagents and conditions: (a) KOt-Bu, t-BuOH, rt, 16 h (47%); (b) TBAF, THF, rt, 16 h (97%); (c) 2-methoxypropene, CSA,
CH2Cl2, 0 ꢁC, 20 min (98%).

T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
9489
Figure 8. 1H NMR spectra of all stereoisomers of pinellic acid.
In conclusion, we have established synthetic routes to
prepare all the stereoisomers of 1 via regioselective asym-
metric dihydroxylation, stereoselective inversion, and
stereoselective reduction. In this series, the (9S,12S,13S)-
compound has the most potent adjuvant activity. Studies
on the mechanism of adjuvant and protective effects of
Figure 9. Anti-influenza virus antibody titer (fluorescence intensity).

9490
T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
pinellic acid with nasal influenza HA vaccine against influ-
enza virus infection are currently under way.
[a]D22 ꢀ10.0 (c 0.06, CHCl3); CD (c 5.3ꢂ10ꢀ5, MeOH)
lmax (D3): 244.8 (+6.97), 220.8 (+2.13), 209.1 (+5.97); IR
(KBr) n cmꢀ1: 1724, 1633; 1H NMR (400 MHz, CDCl3) d:
7.89 (d, J¼8.9 Hz, 2H), 7.58 (d, J¼8.9 Hz, 2H), 5.84 (dd,
J¼15.2 Hz, 1H), 5.76 (dd, J¼15.2, 6.8 Hz, 1H), 5.50 (dt,
J¼7.0, 6.0 Hz, 1H), 3.99 (dd, J¼8.5, 6.8 Hz, 1H), 3.67
(m, 1H), 3.66 (s, 3H), 2.29 (t, J¼7.9 Hz, 2H), 1.21–1.79
(m, 20H), 1.41, 1.40 (s, 3H each), 0.88 (t, J¼6.2 Hz, 3H);
13C NMR (100 MHz, CDCl3) d: 137.9, 131.7 (2C), 131.1
(2C), 130.7, 81.6, 80.8, 74.7, 51.4, 34.3, 34.0, 31.9, 31.9,
29.3, 29.2 (2C), 27.3, 27.0, 25.6, 25.0, 24.9, 22.5, 14.0;
HRMS (FAB, NBA matrix), m/z: 589.2149 [M+Na]+, Calcd
for C29H43O6BrNa: 589.2141 [M+Na].
7. Experimental
7.1. General
Dry THF, toluene, ethyl ether, and CH2Cl2 were purchased
from Kanto Chemical Co. Precoated silica gel plates with
a fluorescent indicator (Merck 60 F254) were used for ana-
lytical and preparative thin-layer chromatography. Flash
column chromatography was carried out with Merck
silica gel 60 (Art. 1.09385). 1H and 13C NMR spectra were
measured on JEOL JNM-EX270 (270 MHz) or Varian VXR-
300 (300 MHz) or Varian XL-400 (400 MHz) or Varian
UNITY-400 (400 MHz). All infrared spectra were measured
on a Horiba FT-210 spectrometer. Melting points were
measured on a Yanagimoto Micro Melting Apparatus.
High- and low-resolution mass spectra were measured on
a JEOL JMS-DX300 and JEOL JMS-AX505 HA spectro-
meter. Elemental analysis data were measured on a Yanaco
CHN CORDER MT-5.
7.3. Total synthesis
7.3.1. Synthesis of C18 skeleton.
7.3.1.1. tert-Butyl-7-methoxycarbonylheptanoate (7).
To a solution of suberic acid monomethyl ester (6,
5.00 mL, 5.24 g, 27.8 mmol) in t-BuOH (56 mL) were
added (Boc)2O (9.58 mL, 41.7 mmol) and DMAP (1.02 g,
0.34 mmol). The mixture was stirred at rt for 1 h. The result-
ing mixture was treated with 0.2 N HCl (20 mL) and
extracted with CHCl3 (50 mLꢂ3). The organic layer was
washed with satd aq NaCl (50 mL), dried over Na2SO4, fil-
tered, and concentrated. The residue was purified by column
chromatography (hexane/AcOEt¼10:1) to give 7 (5.58 g,
82%) as a colorless oil. Rf¼0.41(silica gel, hexane/
AcOEt¼5:1); IR (KBr) n cmꢀ1: 1734; 1H NMR (270 MHz,
CDCl3) d: 3.62 (s, 3H), 2.26 (t, J¼7.3 Hz, 2H), 2.16 (t,
J¼7.3 Hz, 2H), 1.51–1.61 (complex m, 4H), 1.40 (s, 9H),
1.31–1.21 (complex m, 4H); 13C NMR (67.5 MHz, CDCl3)
d: 174.1, 173.0, 79.8, 51.3, 35.4, 33.9, 28.7, 28.6, 28.0
(3C), 24.8, 24.7; HRMS (FAB NBA matrix) m/z: 245.1750
[M+H]+, Calcd for C13H25O4: 245.1753 [M+H].
7.2. Estimation of absolute stereochemistry of 1
7.2.1. 12,13-O-Isopropylidene-9,12,13-trihydroxyocta-
decaenoic acid methyl ester (2). To a solution of pinellic
acid (1, 9.6 mg, 29 mmol) in benzene/MeOH (10:1)
(2.2 mL) was added TMSCHN2 (2.0 M solution in hexane,
29 mL, 58 mmol) and stirred at rt for 2.5 h, after that time
the solution was concentrated. To the solution of residue
in CH2Cl2 (0.6 mL) were added 2,2-dimethoxypropane
(14 mL, 0.12 mmol) and PPTS (7.3 mg, 29 mmol), and then
stirred at 60 ꢁC for 48 h. The solution was cooled to rt and
treated with H2O (500 mL) followed by extraction with
CHCl3 (5 mLꢂ3). The organic layer was washed with satd
aq NaCl (3 mL), dried, and evaporated, and the residue was
purified by column chromatography (hexane/AcOEt¼7:1)
to give 2 (11 mg, 100%) as a colorless oil. Rf¼0.48 (silica
gel, hexane/AcOEt¼1:1); [a]2D8 0.00 (c 0.15, MeOH); IR
(KBr) n cmꢀ1: 3452, 1741; 1H NMR (400 MHz, CDCl3) d:
5.84 (dd, J¼15.5, 5.6 Hz, 1H), 5.65 (dd, J¼15.5, 7.1 Hz,
1H), 4.16 (m, 1H), 4.00 (dd, J¼8.0, 7.1 Hz, 1H), 3.67 (m,
1H), 3.66 (s, 3H), 2.30 (t, J¼7.6 Hz, 2H), 1.63–1.24 (m,
20H), 1.412, 1.405 (s, 3H each), 0.89 (t, J¼6.3 Hz, 3H);
13C NMR (100 MHz, CDCl3) d: 177.4, 137.9, 127.4,
108.4, 81.8, 80.9, 71.8, 51.4, 37.1, 34.1, 31.9, 31.9, 29.3,
29.14, 29.05, 27.3, 27.0, 25.8, 25.2, 24.9, 22.5, 14.0;
HRMS (FAB, NBA matrix) m/z: 407.2742 [M+Na]+, Calcd
for C22H40O5Na: 407.2773 [M+Na].
7.3.1.2. tert-Butyl-8-iodooctanoate (8). To a solution of
1.5 N NaOH in MeOH/H2O/THF (3:1:1) (113 mL) was
added 7 (5.51 g, 22.6 mmol) and stirred at rt for 28 h. The
mixture was treated with 1.0 N HCl (50 mL) and extracted
with CHCl3 (50 mLꢂ3). The organic layer was washed
with satd aq NaCl (50 mL), dried over Na2SO4, filtered,
and concentrated.
The residue was dissolved in THF (41.6 mL) at 0 ꢁC. To the
mixture was added BH3$THF (1.0 M solution in THF,
20.8 mL), after that time, the solution was warmed up to rt
and stirred at rt for 12 h. The resulting mixture was treated
with satd aq NaHCO3 (50 mL) and extracted with CHCl3
(50 mLꢂ3). The organic layer was washed with satd aq NaCl
(50 mL), dried over Na2SO4, filtered, and concentrated.
7.2.2. 9-(4-Bromobenzoyloxy)-12,13-o-isopropylidene-
12,13-dihydroxyoctadecaenoic acid methyl ester (3). To
a solution of 2 (1.0 mg, 2.6 mmol) in pyridine were added
p-bromobenzoyl chloride (5.5 mg, 26 mmol) and DMAP
(0.3 mg, 26 mmol), and then stirred at rt for 10 h. The result-
ing mixture was treated with H2O (0.5 mL) and extracted
with CHCl3 (3 mLꢂ3). The organic layer was washed
with satd aq NaCl (2 mL), dried over Na2SO4, filtered, and
concentrated. The residue was purified by column chromato-
graphy (hexane/AcOEt¼5:1) to give 3 (1.0 mg, 68%) as a
colorless oil. Rf¼0.60 (silica gel, hexane/AcOEt¼1:1);
The residue was dissolved in CH2Cl2 (100 mL) at 0 ꢁC. To
the mixture were added imidazole (2.10 g, 30.9 mmol),
PPh3 (8.10 g, 30.9 mmol), and I2 (6.27 g, 24.7 mmol), after
that time, the solution was warmed up to rt and stirred at rt
for 2 h. The resulting mixture was treated with satd aq
NaHCO3 (50 mL) and extracted with CHCl3 (50 mLꢂ3).
The organic layer was washed with H2O (50 mL), 0.1 N
Na2SO3 soln (50 mL), 30% aq H2O2 (50 mL), satd aq
NaCl (50 mL), dried over Na2SO4, filtered, and concen-
trated. The residue was purified by column chromatography
(hexane/AcOEt¼50:1) to give 8 (5.53 g, 77% from 7) as

T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
9491
a colorless oil. Rf¼0.47 (silica gel, hexane/AcOEt¼4:1); IR
(KBr) n cmꢀ1: 1730; 1H NMR (270 MHz, CDCl3) d: 3.18 (t,
J¼7.3 Hz, 2H), 2.20 (t, J¼7.6 Hz, 2H), 1.76–1.87 (complex
m, 2H), 1.53–1.60 (complex m, 2H), 1.44 (s, 9H), 1.26–1.41
(complex m, 6H); 13C NMR (67.5 MHz, CDCl3) d: 173.0,
79.8, 35.4, 33.3, 30.2, 29.0, 28.7, 28.1, 28.0 (3C), 24.9;
HRMS (EI) m/z: 326.0763 [M]+, Calcd for C12H23O2I:
326.0743 [M].
11.0 mmol), K3[Fe(CN)6] (264.4 mg, 0.803 mmol), K2CO3
(110.8 mg, 0.803 mmol), and K2OsO4$2H2O (4.0 mg,
0.011 mmol) in t-BuOH/H2O (1:1) (2.6 mL) was treated
with methanesulfonamide (25.5 mg, 0.268 mmol) at ambient
temperature. The clear yellow solution was cooled to 0 ꢁC
and 11 (117.8 mg, 0.268 mmol) was added. The solution
was stirred vigorously at 0 ꢁC for 40 h 50 min and then
quenched with solid Na2SO3 (50 mg), warmed to ambient
temperature, and stirred for further 30 min. The resultant
mixture was extracted with CHCl3 (5 mLꢂ3). The organic
layer was washed with satd aq NaCl (5 mL), dried over
Na2SO4, filtered, and concentrated. The residue was purified
by column chromatography (hexane/AcOEt¼1:1) to give
(ꢀ)-12 (81.2 mg, 64%, 95% ee) as a colorless oil. Rf¼0.38
(silica gel, hexane/AcOEt¼1:1); [a]2D4 ꢀ4.5 (c 1.08,
7.3.1.3. (E,E)-1-(1,3-Dithian)-2,4-decadiene (10). To
a solution of (E,E)-2,4-decadienal (9, 21.4 g, 25.0 mL,
141 mmol) in CH2Cl2 (140 mL) at 0 ꢁC were added 1,3-
propanedithiol (18.3 g, 17.0 mL, 169 mmol) and BF3$Et2O
(3.92 g, 3.40 mL, 27.6 mmol), and then the reaction mixture
was warmed up to rt, stirred for 12 h. The resulting mixture
was treated with satd aq NaHCO3 (200 mL) and extracted
with CHCl3 (100 mLꢂ3). The organic layer was washed
with satd aq NaCl (100 mL), dried over Na2SO4, filtered, and
concentrated. The residue was purified by column chromato-
graphy (hexane/AcOEt¼100:1) to give 10 (32.7 g, 96%) as a
colorless oil. Rf¼0.52 (silica gel, hexane/AcOEt¼5:1); IR
1
CHCl3); IR (KBr) n cmꢀ1: 3421, 1730, 1628; H NMR
(270 MHz, CDCl3) d: 5.91 (dd, J¼15.5, 6.6 Hz, 1H), 5.75
(d, J¼15.5 Hz, 1H), 4.04 (dd, J¼6.6, 5.3 Hz, 1H), 4.01–
3.00 (m, 1H), 2.87 (ddd, J¼14.2, 11.5, 2.6 Hz, 2H), 2.68–
2.63 (m, 2H), 2.35 (br s, 1H), 2.26 (br s, 1H), 2.19 (t,
J¼7.3 Hz, 2H), 2.06–2.01 (m, 2H), 1.93–1.88 (m, 2H),
1.67–1.28 (complex m, 18H), 1.44 (s, 9H), 0.89 (t,
J¼6.6 Hz, 3H); 13C NMR (67.5 MHz, CDCl3) d: 173.8,
136.5, 133.6, 80.4, 75.9, 75.1, 54.7, 42.4, 35.9, 33.5, 32.3,
29.8, 29.4, 29.3, 28.5 (3C), 27.5 (2C), 25.8, 25.6, 25.4,
24.0, 22.9, 14.4; HRMS (FAB, NaI matrix), m/z: 497.2743
[M+Na]+, Calcd for C25H46O4S2Na: 497.2735 [M+Na].
1
(KBr) n cmꢀ1: 1653; H NMR (270 MHz, CDCl3) d: 6.34
(dd, J¼15.2, 10.6 Hz, 1H), 5.99 (dd, J¼15.2, 10.6 Hz, 1H),
5.73 (dt, J¼15.2, 7.2 Hz, 1H), 5.59 (dd, J¼15.2, 7.9 Hz, 1H),
4.66 (d, J¼7.9 Hz, 1H), 2.96–2.79 (complex m, 4H), 2.23–
2.02 (complex m, 3H), 1.91–1.77 (m, 1H), 1.39–1.19 (com-
plex m, 6H), 0.87 (t, J¼6.9 Hz, 3H); 13C NMR (67.5 MHz,
CDCl3) d: 137.3, 133.9, 128.8, 126.8, 47.6, 32.8, 31.3, 30.2
(2C), 29.0, 25.1, 22.5, 14.0; HRMS (EI) m/z: 242.1169 [M]+,
Calcd for C13H22O2: 242.1163 [M].
7.3.2.2.
(12S,13S)-(E)-9-(1,3-Dithian)-12,13-di-tert-
butyldimethylsiloxy-10-octadecaenoic acid-tert-butyl ester
((L)-13). To a solution of (ꢀ)-12 (372 mg, 0.787 mmol)
in CH2Cl2 (7.9 mL) were added 2,6-lutidine (916 mL,
7.87 mmol) and TBSOTf (900 mL, 3.93 mmol) at ꢀ78 ꢁC.
The reaction mixture was stirred at ꢀ78 ꢁC for 30 min.
The resultant mixture was treated with H2O (1 mL) and
extracted with CHCl3 (5 mLꢂ3). The organic layer was
washed with satd aq NaCl (5 mL), dried over Na2SO4, fil-
tered, and concentrated. The residue was purified by column
chromatography (hexane/AcOEt¼100:1) to give (ꢀ)-13
(489 mg, 89%) as a colorless oil. Rf¼0.60 (silica gel, hex-
ane/AcOEt¼1:1); [a]D24 ꢀ24.1 (c 1.01, CHCl3); IR (KBr)
7.3.1.4. (E,E)-9-(1,3-Dithian)-10,12-octadecadienoic
acid-tert-butyl ester (11). To a solution of 10 (200 mL,
206 mg, 0.851 mmol) in THF (8.5 mL) was added n-BuLi
(1.53 M solution in hexane, 612 mL, 0.936 mmol) at ꢀ78 ꢁC
dropwise (ca. 15 min). The resulting mixture was stirred
at ꢀ78 ꢁC for 1 h followed by the addition of 8 (327 mL,
416 mg, 1.28 mmol) in one portion. The reaction mixture
was stirred at ꢀ78 ꢁC for 1 h, after that time, the solution
was treated with satd aq NH4Cl (10 mL) and extracted with
AcOEt (10 mLꢂ3). The organic layer was washed with satd
aq NaCl (10 mL), dried over Na2SO4, filtered, and concen-
trated. The residue was purified by column chromatography
(hexane/AcOEt¼100:1) to give 11 (318 mg, 85%) as a color-
less oil. Rf¼0.36 (silica gel, hexane/AcOEt¼20:1, twice); IR
(KBr) n cmꢀ1: 1730, 1695; 1H NMR (400 MHz, CDCl3) d:
6.39 (dd, J¼15.2, 10.4 Hz, 1H), 6.12 (dd, J¼14.9, 10.4 Hz,
1H), 5.76 (dt, J¼14.9, 7.2 Hz, 1H), 5.54 (d, J¼15.2 Hz, 1H),
2.88 (ddd, J¼14.0, 11.2, 2.5 Hz, 2H), 2.64 (ddd, J¼14.0,
5.2, 3.0 Hz, 2H), 2.18 (t, J¼7.2 Hz, 2H), 2.12–2.06 (m, 2H),
2.05–1.98, 1.93–1.91 (m, 1H each), 1.82–1.78 (m, 2H), 1.59–
1.52 (m, 2H), 1.47–1.36 (complex m, 4H), 1.44 (s, 9H), 1.34–
1.19 (complex m, 10H), 0.89 (t, J¼7.1 Hz, 3H); 13C NMR
(100.6 MHz, CDCl3) d: 173.2, 135.5, 133.8, 133.6, 129.0,
79.8, 54.9, 42.3, 35.5, 32.6, 31.4, 29.5, 29.0 (2C), 28.9, 28.1
(3C), 27.2 (2C), 25.5, 25.0, 23.7, 22.5, 14.0; HRMS (EI)
m/z: 440.2779 [M]+, Calcd for C25H44O2S2: 440.2783 [M].
1
n cmꢀ1: 3442, 1731, 1630; H NMR (270 MHz, CDCl3)
d: 5.98 (dd, J¼15.2, 6.6 Hz, 1H), 5.59 (d, J¼15.8 Hz, 1H),
4.24 (m, 1H), 3.59 (m, 1H), 2.98–2.87 (complex m, 1H),
2.86–2.64 (complex m, 2H), 2.18 (t, J¼7.3 Hz, 2H), 2.00–
1.87 (complex m, 2H), 1.80 (m, 2H), 1.44 (s, 9H), 1.67–
1.14 (complex m, 18H), 0.91–0.86 (complex m, 21H),
0.11–0.03 (m, 12H); 13C NMR (67.5 MHz, CDCl3) d:
173.3, 133.4, 133.0, 79.9, 75.5, 75.0, 55.0, 42.3, 35.6, 31.9,
31.1, 29.6, 29.1 (2C), 28.1 (3C), 27.1, 27.0, 26.0, 25.8
(3C), 25.7, 25.1, 23.7, 22.5, 18.2, 18.0, 14.0, ꢀ4.1, ꢀ4.6
(2C), ꢀ4.8; HRMS (FAB, NBA matrix), m/z: 701.4539
[M]+, Calcd for C37H74O4Si2S2: 702.4534 [M].
7.3.2.3. (12S,13S)-(E)-9-Oxo-12,13-di-tert-butyldimethyl-
siloxy-10-octadecaenoic acid-tert-butyl ester ((L)-14).
To a mixture of (ꢀ)-13 (494 mg, 0.704 mmol) and CaCO3
(141 mg, 1.41 mmol) in THF (14 mL) was added a solution
of Hg(ClO4)3 (638 mg, 1.41 mmol) in H2O (2.8 mL) drop-
wise. The resultant mixture was stirred at rt for 30 min,
and then diluted with ether (5 mL). This mixture was filtered
through Celite. The residue was concentrated and dissolved
in CHCl3 (5 mL). This solution was washed with satd aq
7.3.2. Synthesis of (9S,12S,13S)-(E)-9,12,13-trihydroxy-
10-octadienoic acid ((L)-4).
7.3.2.1. (12S,13S)-(E)-12,13-Dihydroxy-9-(1,3-dithian)-
10-octadecaenoic acid-tert-butyl ester ((L)-12). A well-
stirred solution of (DHQ)PHAL(DHQ)Me+$Iꢀ(10.0 mg,

9492
T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
NaCl (5 mL), dried over Na2SO4, filtered, and concentrated.
The residue was purified by column chromatography
(hexane/AcOEt¼50:1) to give (ꢀ)-14 (357 mg, 83%) as
a colorless oil. Rf¼0.55 (silica gel, hexane/AcOEt¼6:1);
[a]D27 ꢀ49.7 (c 0.99, CHCl3); IR (KBr) n cmꢀ1: 1733,
1677, 1633; 1H NMR (270 MHz, CDCl3) d: 6.97 (dd,
J¼16.2, 3.6 Hz, 1H), 6.29 (d, J¼16.2 Hz, 1H), 4.31 (m,
1H), 3.61 (m, 1H), 2.55 (t, J¼7.3 Hz, 2H), 2.19 (t,
J¼7.6 Hz, 2H), 1.44 (s, 9H), 1.67–1.18 (complex m, 18H),
0.92–0.89 (complex m, 18-H3), 0.09–0.03 (m, 12H); 13C
NMR (67.5 MHz, CDCl3) d: 201.2, 173.7, 146.4, 129.8,
80.3, 76.2, 75.0, 40.7, 36.0, 32.2, 31.6, 29.6 (2C), 29.4,
28.6 (3C), 26.4, 26.3 (3C), 26.2 (3C), 25.5, 24.9, 23.0,
18.6, 18.4, 14.4, ꢀ3.8, ꢀ4.0, ꢀ4.3 (2C), ꢀ4.4.
(t, J¼6.3 Hz, 3H); 13C NMR (100 MHz, CD3OD) d: 177.8,
136.6, 131.1, 76.5, 75.8, 73.0, 38.3, 35.0, 33.6, 33.1, 30.5,
30.4, 30.2, 26.6, 26.5, 26.1, 23.7, 14.4; HR-FABMS m/z:
353.2305 [M+Na]+, Calcd for C18H34O5Na: 353.2304
[M+Na]; Anal. Calcd for C18H34O5$1/2H2O: C, 63.69; H,
10.39. Found: C, 63.77; H, 10.03.
7.3.3. Synthesis of (9S,12R,13R)-(E)-9,12,13-trihydroxy-
10-octadienoic acid ((D)-5).
7.3.3.1. (12R,13R)-(E)-12,13-Dihydroxy-9-(1,3-dithian)-
10-octadecaenoic acid-tert-butyl ester ((D)-12). A well-
stirred solution of AD-mix-b (1.68 g) in t-BuOH/H2O
(1:1) (1.2 mL) was treated with methanesulfonamide
(25.5 mg, 0.268 mmol) at ambient temperature. The clear
yellow solution was cooled to 0 ꢁC and 11 (528 mg,
1.20 mmol) was added. The solution was stirred vigorously
at 0 ꢁC for 73 h and then quenched with solid Na2SO3
(500 mg), warmed to ambient temperature, and stirred for
further 30 min. The resultant mixture was extracted with
CHCl3 (20 mLꢂ3). The organic layer was washed with
satd aq NaCl (20 mL), dried over Na2SO4, filtered, and con-
centrated. The residue was purified by column chromato-
graphy (hexane/AcOEt¼1:1) to give (+)-12 (424 mg, 75%)
as a colorless oil. [a]2D4 +5.2 (c 1.08, CHCl3); HRMS
(FAB, NaI matrix), m/z: 497.2740 [M+Na]+, Calcd for
C25H46O4S2Na: 497.2735 [M+Na].
7.3.2.4. (9S,12S,13S)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid-tert-butyl ester ((L)-15). To a solution of
(ꢀ)-14 (349.0 mg, 0.570 mmol) in THF (11 mL) was added
(S)-BINAL-H (0.5 M solution in THF, 7.52 mL, 3.76 mmol)
at ꢀ78 ꢁC. The reaction mixture was stirred at ꢀ78 ꢁC for
2 h 30 min. The resultant mixture was treated with 1.0 N
HCl (10 mL) and extracted with CHCl3 (20 mLꢂ3). The
organic layer was washed with 1.0 N NaOH (20 mL), satd
aq NaCl (20 mL), dried over Na2SO4, filtered, and concen-
trated. The residue was dissolved in THF (5.8 mL) at rt.
To the mixture was added TBAF (1.0 M solution in THF,
1.25 mL, 1.25 mmol) and stirred at 70 ꢁC for 3 h. This resul-
tant mixture was treated with H2O (1.0 mL) and extracted
with CHCl3 (20 mLꢂ3). The organic layer was washed
with satd aq NaCl (20 mL), dried over Na2SO4, filtered,
and concentrated. The residue was purified by column
chromatography (toluene/AcOEt¼2:1) to give (ꢀ)-15
(168.1 mg, 76%) as a colorless oil. Rf¼0.29 (silica gel, tol-
uene/AcOEt¼1:2); [a]D27 ꢀ8.8 (c 0.16, CHCl3); IR (KBr)
7.3.3.2. (12R,13R)-(E)-9-(1,3-Dithian)-12,13-di-tert-
butyldimethylsiloxy-10-octadecaenoic acid-tert-butyl ester
((D)-13). According to the synthesis of (ꢀ)-13, (+)-12
(77.9 mg) gave (+)-13 (115 mg, 87%) as a colorless oil.
[a]2D7 +22.4 (c 0.41, CHCl3); HRMS (FAB, NBA matrix),
m/z: 701.4534 [M]+, Calcd for C37H74O4Si2S2: 704.4534 [M].
1
n cmꢀ1: 3305, 1727; H NMR (270 MHz, CDCl3) d: 5.83
7.3.3.3. (12R,13R)-(E)-9-Oxo-12,13-di-tert-butyldi-
methylsiloxy-10-octadecaenoic acid-tert-butyl ester ((D)-
14). According to the synthesis of (ꢀ)-14, (+)-13 (93.8 mg)
gave (+)-14 (67.4 mg, 83%) as a colorless oil. [a]2D7 +22.4
(c 0.41, CHCl3).
(dd, J¼15.5, 5.6 Hz, 1H), 5.67 (dd, J¼15.5, 5.9 Hz, 1H),
4.15 (dd, J¼12.2, 5.9 Hz, 1H), 3.94 (t, J¼5.93 Hz, 1H),
3.47 (m, 1H), 2.34 (br s, 1H), 2.26 (br s, 1H), 2.19 (t,
J¼7.6 Hz, 2H), 1.63 (m, 2H), 1.44 (s, 9H), 1.52–1.30 (com-
plex m, 18H), 0.98 (t, J¼6.6 Hz, 3H); 13C NMR (67.5 MHz,
CDCl3) d: 173.3, 136.2, 129.7, 79.9, 75.3, 74.6, 72.0, 37.1,
35.6, 32.9, 31.8, 29.2, 29.1, 28.9, 28.1, 25.3, 25.2, 25.0,
22.6, 14.0; HRMS (FAB, NBA matrix), m/z: 409.2913
[M]+, Calcd for C22H42O5Na: 409.2930 [M].
7.3.3.4. (9S,12R,13R)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid-tert-butyl ester ((D)-16). According to the
synthesis of (ꢀ)-15, (+)-14 (26.1 mg) gave (+)-16 (12.4 mg,
76%) as a colorless oil. Rf¼0.20 (silica gel, toluene/
AcOEt¼1:2), [a]D28 +7.4 (c 0.19, CHCl3); IR (KBr) n cmꢀ1
:
1
7.3.2.5. (9S,12S,13S)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid ((L)-4). To a solution of 2.0 N KOH in
EtOH/H2O (4:1) (500 mL) was added (ꢀ)-15 (6.5 mg,
16.8 mmol) and stirred at rt for 46 h. The mixture was cooled
to 0 ꢁC, treated with 1.0 N HCl (500 mL), and extracted with
CHCl3 (2 mLꢂ3). The organic layer was washed with satd
aq NaHCO3 (5 mL), satd aq NaCl (5 mL), dried over
Na2SO4, filtered, and concentrated. The residue was purified
by column chromatography (CHCl3/MeOH¼10:1) to give
(ꢀ)-4 (4.5 mg, 82%) as a white solid. Rf¼0.24 (silica gel,
CHCl3/MeOH/AcOH¼10:1:0.1); mp 104–106 ꢁC (MeOH);
[a]2D5 ꢀ8.0 (c 0.30, MeOH), {natural; [a]2D8 ꢀ8.1 (c 0.32,
MeOH)}; IR (KBr) n cmꢀ1: 3372 (s), 1695 (m), 1637 (m);
1H NMR (400 MHz, CD3OD) d: 5.72 (dd, J¼15.5, 5.0 Hz,
1H), 5.67 (dd, J¼15.5, 5.0 Hz, 1H), 4.05 (ddd, J¼6.5, 6.0,
5.0 Hz, 1H), 3.91 (dd, J¼5.5, 5.0 Hz, 1H), 3.41 (ddd, J¼8.5,
5.5, 2.5 Hz, 1H), 2.27 (t, J¼7.5 Hz, 2H), 1.60 (dt, J¼7.5,
7.0 Hz, 2H), 1.55–1.50 (m, 4H), 1.45–1.25 (m, 14H), 0.91
3392 (s), 1732 (m); H NMR (270 MHz, CDCl3) d: 5.83
(dd, J¼15.5, 5.6 Hz, 1H), 5.67 (dd, J¼15.5, 5.9 Hz, 1H),
4.15 (dd, J¼12.2, 5.9 Hz, 1H), 3.94 (1H, t, J¼5.93 Hz,
1H), 3.47 (m, 1H), 2.34 (br s, 1H), 2.26 (br s, 1H), 2.19 (t,
J¼7.6 Hz, 2H), 1.63 (m, 2H), 1.44 (s, 9H), 1.52–1.30 (com-
plex m, 18H), 0.98 (t, J¼6.6 Hz, 3H); 13C NMR (67.5 MHz,
CDCl3) d: 173.3, 136.2, 129.7, 79.9, 75.3, 74.6, 72.0, 37.1,
35.6, 32.9, 31.8, 29.2, 29.1, 28.9 (2C), 28.1 (3C), 25.3,
25.2, 25.0, 22.6, 14.0; HRMS (FAB, NBA matrix), m/z:
409.2913 [M]+, Calcd for C22H42O5Na: 409.2930 [M].
7.3.3.5. (9S,12R,13R)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid ((D)-5). According to the synthesis of (ꢀ)-4,
(+)-16 (12.4 mg) gave (+)-5 (8.1 mg, 76%) as a white solid.
Rf¼0.23 (silica gel, CHCl3/MeOH/AcOH¼10:1:0.1); mp
68–71 ꢁC (MeOH); [a]2D3 +29.8 (c 0.45, MeOH); IR (KBr)
n cmꢀ1: 3430 (s), 1697 (m), 1632 (m); 1H NMR (400 MHz,
CD3OD) d: 5.70 (dd, J¼15.5, 5.5 Hz, 1H), 5.64 (dd,

T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
9493
J¼15.5, 6.0 Hz, 1H), 4.03 (ddd, J¼6.5, 6.0, 5.5 Hz, 1H), 3.87
(dd, J¼6.0, 5.5 Hz, 1H), 3.40 (ddd, J¼7.0, 5.5, 2.0 Hz, 1H),
2.27 (t, J¼7.5 Hz, 2H), 1.60 (dt, J¼7.5, 7.0 Hz, 2H), 1.55–
1.50 (m, 4H), 1.44–1.25 (m, 14H), 0.91 (t, J¼6.3 Hz, 3H);
13C NMR (100 MHz, CD3OD) d: 178.2, 136.7, 131.3, 76.7,
75.7, 73.2, 38.3, 36.0, 33.8, 33.1, 30.5, 30.4, 30.2, 26.5,
26.5, 26.2, 23.7, 14.4; HR-FABMS m/z: 353.2309
[M+Na]+, Calcd for C18H34O5Na: 353.2304 [M+Na].
(67.5 MHz, CDCl3) d: 173.2, 135.7, 133.7, 79.9, 77.3,
75.5, 54.2, 42.2, 35.5, 32.6, 31.9, 29.7, 29.6, 29.1, 29.0,
28.0 (3C), 27.0, 26.9, 25.7, 25.5, 23.9, 22.6, 18.1 (6C),
14.0, 12.5 (3C); HRMS (FAB, NaI matrix) m/z: 653.4061
[M+Na]+, Calcd for C34H66O4SiS2Na: 653.4070 [M+Na].
7.3.5.2. (12R,13S)-(E)-13-Acetoxy-9-(1,3-dithian)-12-
triisopropylsiloxy-10-octadecaenoic acid-tert-butyl ester
((L)-19). To a solution of (ꢀ)-18 (13.0 mg, 0.021 mmol)
in pyridine (0.5 mL) was added ClCH2SO2Cl (3.9 mL,
0.030 mmol) at 0 ꢁC. The resultant mixture was stirred at
0 ꢁC for 2 h, treated with H2O (0.5 mL), and successfully
extracted with CHCl3 (5 mLꢂ3). The combined organic
layer was washed with satd aq NaCl (5 mL), dried over
Na2SO4, and concentrated.
7.3.4. Syntheses of (9R,12R,13R)-(E)-9,12,13-trihydroxy-
10-octadienoic acid ((D)-4) and (9R,12S,13S)-tri-
hydroxy-10-octadienoic acid ((L)-5).
7.3.4.1. (9R,12R,13R)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid-tert-butyl ester ((D)-15). According to
the synthesis of (ꢀ)-15, the reduction of (+)-14 (26.1 mg)
using (R)-BINAL-H gave (+)-15 (10.8 mg, 63%) as a color-
less oil. [a]2D7 +6.6 (c 0.21, CHCl3); HRMS (FAB, NBA
matrix), m/z: 409.2908 [M]+, Calcd for C22H42O5Na:
409.2930 [M].
To a solution of the residue of previous reaction in benzene
was added CsOAc (19.8 mg, 0.10 mmol) and 18-crown-6
(4.1 mg, 0.021 mmol) at rt. The resultant mixture was
warmed, refluxed for 20 h, and then cooled to rt again to treat
with H2O (500 mL) and extracted with CHCl3 (5 mLꢂ3).
The organic layer was washed with satd aq NaCl (5 mL),
dried over Na2SO4, filtered, and concentrated. The residue
was purified by column chromatography (hexane/AcOEt¼
50:1) to give (ꢀ)-19 (11.5 mg, 83% from (ꢀ)-18) as a
colorless oil. Rf¼0.50 (silica gel, hexane/AcOEt¼8:1);
[a]2D4 ꢀ21.8 (c 0.87, CHCl3); IR (KBr) n cmꢀ1: 1734 (s),
1635 (m); 1H NMR (270 MHz, CDCl3) d: 5.89 (dd,
J¼15.5, 6.3 Hz, 1H), 5.69 (d, J¼15.5 Hz, 1H), 4.95–4.89
(m, 1H), 4.47 (dd, J¼6.3, 2.6 Hz, 1H), 2.93–2.79 (m, 2H),
2.69–2.61 (m, 2H), 2.18 (t, J¼7.3 Hz, 2H), 2.05 (s, 3H),
2.02–1.87 (m, 2H), 1.83–1.66 (m, 2H), 1.47–1.15 (complex
m, 18H), 1.43 (s, 9H), 1.10–0.95 (m, 21H), 0.87 (t,
J¼6.9 Hz, 3H); 13C NMR (67.5 MHz, CDCl3) d: 173.2,
170.8, 135.0, 133.1, 79.9, 77.4, 77.2, 54.3, 42.2, 35.5,
31.7, 29.6, 29.1, 29.0 (2C), 28.1 (3C), 27.0, 26.9, 25.5,
25.3, 25.0, 23.8, 22.4, 21.2, 18.0 (6C), 14.0, 12.5 (3C);
HRMS (FAB, NaI matrix), m/z: 695.4162 [M+Na]+, Calcd
for C36H68O5SiS2Na: 695.4175 [M+Na].
7.3.4.2. (9R,12R,13R)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid ((D)-4). According to the synthesis of
(ꢀ)-4, the deprotection of (+)-15 (10.8 mg) gave (+)-4
(4.8 mg, 67%) as a white solid. Mp 98–104 ꢁC (MeOH);
[a]2D8 +12.9 (c 0.48, MeOH); HR-FABMS m/z: 353.2307
[M+Na]+, Calcd for C18H34O5Na: 353.2304 [M+Na].
7.3.4.3. (9R,12S,13S)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid-tert-butyl ester ((L)-16). According to the
synthesis of (ꢀ)-15, the reduction of (ꢀ)-14 (10.8 mg) using
(R)-BINAL-H gave (ꢀ)-16 (8.9 mg, 57%) as a colorless oil.
[a]2D7 ꢀ9.9 (c 0.99, CHCl3); HRMS (FAB, NBA matrix), m/z:
409.2910 [M]+, Calcd for C22H42O5Na: 409.2930 [M].
7.3.4.4. (9R,12S,13S)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid ((L)-5). According to the synthesis of (+)-
5, the deprotection of (ꢀ)-16 (8.9 mg) gave (ꢀ)-5 (3.7 mg,
51%) as a white solid. Mp 69–74 ꢁC (MeOH); [a]D22 ꢀ24.0
(c 0.30, MeOH); HR-FABMS m/z: 353.2307 [M+Na], Calcd
for C18H34O5Na: 353.2304.
7.3.5.3. (12R,13S)-(E)-13-Acetoxy-9-oxo-12-triisopro-
pylsiloxy-10-octadecaenoic acid-tert-butyl ester ((L)-
20). To a mixture of (ꢀ)-19 (304 mg, 0.452 mmol) and
CaCO3 (90.4 mg, 0.904 mmol) in THF (4.5 mL) was added
a solution of Hg(ClO4)3 (410 mg, 0.904 mmol) in H2O
(900 mL) dropwise. The resultant mixture was stirred at rt
for 5 min, and then diluted with ether (2 mL). This mixture
was filtered through Celite. The residue was concentrated
and dissolved in CHCl3 (15 mL). This solution was washed
with satd aq NaCl (5 mL), dried over Na2SO4, filtered, and
concentrated. The residue was purified by column chromato-
graphy (hexane/AcOEt¼10:1) to give (ꢀ)-20 (250 mg,
97%) as a colorless oil. Rf¼0.55 (silica gel, hexane/AcOEt¼
6:1); [a]2D4 ꢀ22.0 (c 0.98, CHCl3); IR (KBr) n cmꢀ1: 1735
(m), 1680 (m), 1633 (m); 1H NMR (270 MHz, CDCl3)
d: 6.71 (dd, J¼15.8, 5.9 Hz, 1H), 6.24 (d, J¼15.8 Hz, 1H),
4.93 (m, 1H), 4.48 (dd, J¼5.9, 3.6 Hz, 1H), 2.55 (t,
J¼7.6 Hz, 2H), 2.19 (t, J¼7.3 Hz, 2H), 2.04 (s, 3H), 1.73–
1.17 (complex m, 18H), 1.44 (s, 9H), 1.12–0.98 (m, 21H),
0.87 (t, J¼6.3 Hz, 3H); 13C NMR (67.5 MHz, CDCl3) d:
200.3, 173.2, 170.6, 144.6, 130.5, 79.8, 76.4, 74.2, 40.2,
35.5, 31.5, 31.4, 29.0 (2C), 28.9, 28.9, 28.0 (3C), 25.2,
24.9, 24.1, 22.4, 21.0, 17.9 (6C), 13.9, 12.3 (3C); HRMS
7.3.5. Synthesis of (9S,12R,13S)-(E)-9,12,13-trihydroxy-
10-octadienoic acid ((D)-17).
7.3.5.1. (12R,13R)-(E)-9-(1,3-Dithian)-13-hydroxy-12-
triisopropylsiloxy-10-octadecaenoic acid tert-butyl ester
((L)-18). To a mixture of (+)-12 (326 mg, 0.689 mmol)
and 2,6-lutidine (160 mL, 7.87 mmol) in CH2Cl2 (14 mL)
was added TIPSOTf (194 mL, 0.723 mmol) dropwise
over 20 min at ꢀ78 ꢁC. The reaction mixture was stirred
at ꢀ78 ꢁC for 8 h. The resultant mixture was treated with
H2O (1 mL) and extracted with CHCl3 (10 mLꢂ3). The
organic layer was washed with satd aq NaCl (5 mL), dried
over Na2SO4, filtered, and concentrated. The residue was
purified by column chromatography (hexane/AcOEt¼50:1)
to give (ꢀ)-18 (391 mg, 90%) as a colorless oil. Rf¼0.44
(silica gel, hexane/AcOEt¼5:1); [a]2D4 ꢀ4.8 (c 1.01,
CHCl3); IR (KBr) n cmꢀ1: 3442 (s), 1731 (m), 1630 (m);
1H NMR (270 MHz, CDCl3) d: 5.91 (dd, J¼15.5, 7.6 Hz,
1H), 5.68 (d, J¼15.5 Hz, 1H), 4.16 (dd, J¼7.6, 6.9 Hz,
1H), 4.01–3.00 (m, 1H), 2.92–2.77 (m, 2H), 2.69–2.63 (m,
2H), 2.18 (t, J¼7.3 Hz, 2H), 2.11–1.87 (m, 2H), 1.83–1.67
(m, 2H), 1.67–1.58 (complex m, 18H), 1.42 (s, 9H), 1.15–
1.02 (m, 21H), 0.89 (t, J¼6.6 Hz, 3H); 13C NMR

9494
T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
(FAB, NaI matrix); m/z: 605.4202 [M+Na]+, Calcd for
C33H62O6SiS2Na: 605.4213 [M+Na].
(166 mg) gave (+)-18 (154 mg, 79% based on recovered
(ꢀ)-12) as a colorless oil. [a]2D9 +5.9 (c 0.37, CHCl3);
HRMS (FAB, NaI matrix), m/z: 653.4053 [M+Na]+, Calcd
for C34H66O4SiS2Na: 653.4070 [M+Na].
7.3.5.4. (9S,12R,13S)-(E)-13-Acetoxy-9-hydroxy-12-
triisopropylsiloxy-10-octadecaenoic acid-tert-butyl ester
((L)-21). To a solution of (ꢀ)-20 (18.7 mg, 0.033 mmol)
in THF (300 mL) was added (S)-BINAL-H (0.5 M solution
in THF, 215 mL, 0.107 mmol) at ꢀ78 ꢁC. The reaction mix-
ture was stirred at ꢀ78 ꢁC for 1 h 30 min. The resultant mix-
ture was treated with 1.0 N HCl (1 mL) and extracted with
CHCl3 (5 mLꢂ3). The organic layer was washed with
1.0 N NaOH (5 mL), satd aq NaCl (5 mL), dried over
Na2SO4, filtered, and concentrated. The residue was purified
by column chromatography (hexane/AcOEt¼10:1) to give
(ꢀ)-21 (18.6 mg, 99%) as a colorless oil. Rf¼0.44 (silica
gel, hexane/AcOEt¼4:1); [a]2D5 ꢀ18.9 (c 1.40, CHCl3); IR
7.3.6.2. (12S,13R)-(E)-13-Acetoxy-9-(10,3-dithian)-12-
triisopropylsiloxy-10-octadecaenoic acid-tert-butyl ester
((D)-19). According to the synthesis of (ꢀ)-19, (+)-18
(154 mg) gave (+)-19 (124 mg, 75%) as a colorless oil.
[a]2D5 +23.6 (c 1.10, CHCl3); HRMS (FAB, NaI matrix),
m/z: 695.4148 [M+Na]+, Calcd for C36H68O5SiS2Na:
695.4175 [M+Na].
7.3.6.3. (12S,13R)-(E)-13-Acetoxy-9-oxo-12-triisopro-
pylsiloxy-10-octadecaenoic acid-tert-butyl ester ((D)-
20). According to the synthesis of (ꢀ)-20, (+)-19 (114 mg)
gave (+)-20 (89 mg, 89%) as a colorless oil. [a]2D5 +22.6
(c 0.83, CHCl3); HRMS (FAB, NaI matrix), m/z: 605.4201
[M+Na]+, Calcd for C33H62O6SiS2Na: 605.4213 [M+Na].
1
(KBr) n cmꢀ1: 1733 (m), 1630 (m); H NMR (270 MHz,
CDCl3) d: 5.69 (dd, J¼15.8, 5.6 Hz, 1H), 5.62 (dd,
J¼15.8, 5.9 Hz, 1H), 4.93 (m, 1H), 4.29 (dd, J¼5.9,
3.0 Hz, 1H), 4.11–4.07 (m, 1H), 2.19 (t, J¼7.3 Hz, 2H),
2.03 (s, 3H), 1.79–1.20 (complex m, 20H), 1.44 (s, 9H),
1.10–0.98 (m, 21H), 0.87 (t, J¼6.3 Hz, 3H); 13C NMR
(67.5 MHz, CDCl3) d: 173.2, 170.8, 135.2, 130.2, 79.9,
77.1, 74.8, 72.2, 37.1, 35.5, 31.5, 29.3, 29.2, 29.0, 28.9,
28.1 (3C), 25.3, 25.2, 25.0, 22.5, 21.2, 18.0 (6C), 14.0, 12.4
(3C); HRMS (FAB, NaI matrix), m/z: 607.4372 [M+Na]+,
Calcd for C33H64O6SiNa: 607.4370 [M+Na].
7.3.6.4. (9S,12S,13R)-(E)-13-Acetoxy-9-hydroxy-12-
triisopropylsiloxy-10-octadecaenoic acid-tert-butyl ester
((D)-22). According to the synthesis of (ꢀ)-21, the reduc-
tion of (+)-20 (78.0 mg) using (S)-BINAL-H gave (+)-22
(70.3 mg, 99% based on recovered (+)-20) as a colorless
oil. Rf¼0.43 (silica gel, hexane/AcOEt¼4:1); [a]2D5 +25.8
(c, CHCl3); IR (KBr) n cmꢀ1: 3439 (s), 1734 (m), 1640
(m); 1H NMR (270 MHz, CDCl3) d: 5.71 (m, 2H), 4.86 (m,
1H), 4.28 (dd, J¼5.3, 4.0 Hz, 1H), 4.13–4.07 (m, 1H), 2.19
(t, J¼7.3 Hz, 2H), 2.04 (s, 3H), 1.79–1.20 (complex m,
18H), 1.44 (s, 9H), 1.10–0.98 (m, 21H), 0.87 (t, J¼6.3 Hz,
3H); 13C NMR (67.5 MHz, CDCl3) d: 173.3, 170.9, 135.2,
130.5, 79.9, 77.2, 74.8, 72.2, 37.1, 35.5, 31.7, 29.4, 29.3,
29.0 (2C), 28.0 (3C), 25.3, 25.0, 21.2, 21.2, 18.0 (6C),
14.0, 12.4 (3C); HRMS (FAB, NBA matrix), m/z: 607.4364
[M+Na]+, Calcd for C33H64O6SiNa: 607.4370 [M+Na].
7.3.5.5. (9S,12R,13S)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid ((D)-17). To a solution of 1.0 N KOH in
EtOH/H2O (4:1) (500 mL) was added (ꢀ)-20 (17.2 mg,
29.8 mmol) and stirred at rt for 120 h. The mixture was
cooled to 0 ꢁC and treated with 1.0 N HCl (500 mL) and
extracted with CHCl3 (5 mLꢂ3). The organic layer was
washed with satd aq NaHCO3 (5 mL), satd aq NaCl
(5 mL), dried over Na2SO4, filtered, and concentrated.
To a solution of the residue of previous reaction in THF
(10 mL) at 0 ꢁC was added TBAF (1.0 M solution in THF,
30 mL, 29.8 mmol). The resultant mixture was warmed to rt
and stirred for 45 h before being treated with satd aq
NH4Cl (500 mL) and extracted with AcOEt (5 mLꢂ3). The
organic layer was washed with satd aq NaCl (5 mL), dried
over Na2SO4, filtered, and concentrated. The residue was
purified by column chromatography (AcOEt) to give (+)-17
(9.3 mg, 94%) as a white solid. Rf¼0.23 (silica gel, CHCl3/
MeOH/AcOH¼10:1:0.1); mp 67–70 ꢁC (MeOH); [a]D25
+7.8 (c 0.18, MeOH); IR (KBr) n cmꢀ1: 3421 (s), 1699
7.3.6.5. (9S,12S,13R)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid ((D)-23). According to the synthesis of
(+)-17, the deprotection of (+)-22 (45.9 mg) gave (+)-23
(25.7 mg, 98%) as a white solid. Rf¼0.24 (silica gel,
CHCl3/MeOH/AcOH¼10:1:0.1); mp 91–94 ꢁC (MeOH);
[a]2D5 +6.7 (c 0.14, MeOH); IR (KBr) n cmꢀ1: 3420 (s),
1
1701 (m), 1637 (m); H NMR (400 MHz, CD3OD) d: 5.73
(dd, J¼15.9, 5.0 Hz, 1H), 5.68 (dd, J¼15.9, 5.5 Hz, 1H),
4.05 (ddd, J¼6.0, 5.5, 5.0 Hz, 1H), 3.93 (dd, J¼5.0,
4.5 Hz, 1H), 3.47 (ddd, J¼8.5, 4.5, 2.1 Hz, 1H), 2.27 (t,
J¼7.5 Hz, 2H), 1.60 (dt, J¼7.5, 7.0 Hz, 2H), 1.55–1.50
(m, 2H), 1.45–1.25 (m, 16H), 0.91 (t, J¼6.3 Hz, 3H);
13C NMR (100 MHz, CD3OD) d: 177.7, 136.5, 130.9,
76.5, 75.7, 73.0, 38.3, 34.9, 33.5, 33.1, 30.5, 30.3, 30.2,
26.7, 26.4, 26.1, 23.7, 14.4; HR-FABMS m/z: 353.2336
[M+Na]+, Calcd for C18H34O5Na: 353.2304 [M+Na].
1
(m), 1637 (m); H NMR (400 MHz, CD3OD) d: 5.72 (dd,
J¼15.8, 5.5 Hz, 1H), 5.66 (dd, J¼15.8, 6.0 Hz, 1H), 4.04
(ddd, J¼6.5, 6.0, 5.0 Hz, 1H), 3.91 (dd, J¼5.5, 4.5 Hz,
1H), 3.49 (ddd, J¼7.5, 4.5, 2.0 Hz, 1H), 2.27 (t, J¼7.5 Hz,
2H), 1.60 (dt, J¼7.6, 6.9 Hz, 2H), 1.55–1.50 (m, 4H),
1.45–1.25 (m, 14H), 0.91 (t, J¼6.3 Hz, 3H); 13C NMR
(100 MHz, CD3OD) d: 177.8, 136.7, 130.9, 76.6, 75.7,
73.3, 38.4, 35.1, 33.7, 33.1, 30.6, 30.4, 30.2, 26.7, 26.5,
26.1, 23.7, 14.4; HR-FABMS m/z: 353.2307 [M+Na], Calcd
for C18H34O5Na: 353.2304 [M+Na].
7.3.6.6. (9R,12R,13S)-(E)-13-Acetoxy-9-hydroxy-12-
triisopropylsiloxy-10-octadecaenoic acid-tert-butyl ester
((L)-22). According to the synthesis of (+)-22, the reduction
of (ꢀ)-20 (67.8 mg) using (R)-BINAL-H gave (ꢀ)-22
(55.4 mg, 82%) as a colorless oil.
7.3.6. Syntheses of all the stereoisomers of pinellic acid.
7.3.6.1. (12S,13S)-(E)-9-(1,3-Dithian)-13-hydroxy-12-
triisopropylsiloxy-10-octadecaenoic acid-tert-butyl ester
((D)-18). According to the synthesis of (ꢀ)-18, (ꢀ)-12
7.3.6.7. (9R,12R,13S)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid ((L)-23). According to the synthesis of
(+)-17, the deprotection of (ꢀ)-22 (26.5 mg) gave (ꢀ)-23

T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
9495
(14.0 mg, 94%) as a white solid. Mp 88–93 ꢁC (MeOH); [a]D30
ꢀ5.3 (c 0.15, MeOH); HR-FABMS m/z: 353.2307 [M+Na]+,
Calcd for C18H34O5Na: 353.2304 [M+Na].
3H); 13C NMR (67.5 MHz, CDCl3) d: 173.3, 135.7, 133.6,
79.9, 76.2, 75.2, 54.6, 42.4, 35.6, 32.6, 32.0, 29.7, 29.1,
29.0, 28.1 (3C), 27.1, 27.0, 25.6, 25.5, 25.1, 23.7, 22.5,
18.1 (6C), 14.1, 12.4 (3C); HRMS (FAB, NBA matrix), m/z:
630.4178 [M]+, Calcd for C34H66O4S2Si: 630.4172 [M].
7.3.6.8. (9R,12S,13R)-(E)-13-Acetoxy-9-hydroxy-12-
triisopropylsiloxy-10-octadecaenoic acid-tert-butyl ester
((D)-21). According to the synthesis of (ꢀ)-21, the reduc-
tion of (+)-20 (32.9 mg) using (R)-BINAL-H gave (+)-21
(79.4 mg, 98%) as a colorless oil.
7.4.3. (12R,13S)-(E)-9-(1,3-Dithian)-12,13-dihydroxy-10-
octadecaenoic acid-tert-butyl ester ((D)-26). To a solution
of (ꢀ)-25 (4.3 mg, 6.8 mmol) in THF (500 mL) was added
TBAF (1.0 M solution in THF, 6.8 mL, 6.8 mmol) at rt. The
resultant mixture was stirred at rt for 16 h, treated with
H2O (500 mL), and then extracted with CHCl3 (3 mLꢂ3).
The combined organic layer was washed with satd aq NaCl
(3 mL), dried over Na2SO4, filtered, and concentrated. The
residue was purified by column chromatography (hexane/
AcOEt¼1:1) to give (+)-26 (3.1 mg, 97%) as a colorless
oil. Rf¼0.38 (silica gel, hexane/AcOEt¼1:1); [a]2D4 +0.4
(c 0.52, CHCl3); IR (KBr) n cmꢀ1: 3428 (s), 1731 (m),
1630 (m); 1H NMR (270 MHz, CDCl3) d: 5.98 (dd,
J¼15.2, 6.9 Hz, 1H), 5.72 (d, J¼15.2 Hz, 1H), 4.23 (d,
J¼6.9, 3.6 Hz, 1H), 3.77–3.71 (m, 1H), 2.94–2.81 (m, 2H),
2.68–2.32 (m, 2H), 2.19 (t, J¼7.3 Hz, 2H), 1.84–1.78 (m,
2H), 1.67–1.28 (complex m, 20H), 1.44 (s, 9H), 0.89
(t, J¼6.6 Hz, 3H); 13C NMR (67.5 MHz, CDCl3) d: 173.3,
136.4, 131.3, 80.0, 75.1, 74.2, 54.7, 42.0, 35.5, 32.4,
31.8, 29.4, 29.0, 28.9, 28.1 (3C), 27.2 (2C), 25.5 (2C),
24.9, 23.7, 22.5, 14.0; HRMS (FAB, NBA matrix), m/z:
497.2744 [M+Na]+, Calcd for C25H46O4S2Na: 497.2735
[M+Na].
7.3.6.9. (9R,12S,13R)-(E)-9,12,13-Trihydroxy-10-octa-
decaenoic acid ((L)-17). According to the synthesis of
(+)-17, the deprotection of (+)-21 (32.9 mg) gave (ꢀ)-17
(7.5 mg, 18%) as a white solid. Mp 65–74 ꢁC (MeOH);
[a]3D0 ꢀ7.1 (c 0.14, MeOH); HR-FABMS m/z: 353.2307
[M+Na]+, Calcd for C18H34O5Na: 353.2304 [M+Na].
7.4. Stereochemistry on allylic 1,2-diol
7.4.1. (12R,13R)-(E)-9-(1,3-Dithian)-12,13-isopropyl-
idenedioxy-10-octadecaenoic acid-tert-butyl ester ((D)-
24). To a solution of (+)-12 (47.3 mg, 99.7 mmol) in
CH2Cl2 (1.0 mL) were added CSA (2.3 mg, 9.97 mmol)
and 2-methoxypropene (147 mL, 150 mmol) at 0 ꢁC. The re-
sultant mixture was stirred at 0 ꢁC for 5 min, treated with
H2O (1 mL), and then extracted with CHCl3 (5 mLꢂ3).
The organic layer was washed with satd aq NaCl (5 mL),
dried over Na2SO4, filtered, and concentrated. The residue
was purified by column chromatography (hexane/
AcOEt¼20:1) to give (+)-24 (49.0 mg, 96%) as a colorless
oil. Rf¼0.38 (silica gel, hexane/AcOEt¼1:1); [a]2D2 +27.2
(c 1.50, CHCl3); IR (KBr) n cmꢀ1: 3461 (s), 1730 (m),
1630 (m); 1H NMR (400 MHz, CDCl3) d: 5.86 (dd,
J¼15.0, 7.2 Hz, 1H), 5.74 (d, J¼15.0 Hz, 1H), 4.11 (dd,
J¼8.0, 7.2 Hz, 1H), 3.69 (ddd, J¼8.0, 6.5, 5.0 Hz, 1H),
2.91–2.84 (m, 2H), 2.67–2.61 (m, 2H), 2.18 (t, J¼7.2 Hz,
2H), 2.06–2.00 (m, 1H), 1.88–1.82 (m, 1H), 1.79 (ddd,
J¼11.0, 6.0, 3.5 Hz, 2H), 1.59–1.21 (complex m, 18H),
1.44 (s, 9H), 1.41 (s, 6H), 0.88 (t, J¼6.6 Hz, 3H); 13C
NMR (100 MHz, CDCl3) d: 173.2, 137.2, 131.3, 108.6,
81.8, 80.9, 79.9, 54.7, 42.4, 35.6, 32.0, 31.9, 29.6, 29.1,
29.0, 28.1 (3C), 27.3, 27.2 (2C), 27.0, 25.8, 25.5, 25.0,
23.6, 22.5, 14.0; HRMS (FAB, NAI matrix), m/z: 514.3145
[M]+, Calcd for C28H50O2: 514.3151 [M].
7.4.4. (12R,13S)-(E)-9-(1,3-Dithian)-12,13-isopropylene-
dioxy-10-octadecaenoic acid-tert-butyl ester ((D)-27).
To a solution of (+)-26 (17.1 mg, 37.3 mmol) in CH2Cl2
(0.7 mL) were added CSA (0.9 mg, 3.73 mmol) and 2-meth-
oxypropene (5.3 mL, 56.0 mmol) at 0 ꢁC. The resultant mix-
ture was stirred at 0 ꢁC for 20 min, treated with H2O (1 mL),
and then extracted with CHCl3 (2 mLꢂ3). The organic layer
was washed with satd aq NaCl (2 mL), dried over Na2SO4,
filtered, and concentrated. The residue was purified by
column chromatography (hexane/AcOEt¼20:1) to give
(+)-27 (18.2 mg, 98%) as a colorless oil. Rf¼0.38 (silica
gel, hexane/AcOEt¼1:1); [a]2D5 0.0 (c 0.87, CHCl3); IR
1
(KBr) n cmꢀ1: 3446 (s), 1730 (m), 1628 (m); H NMR
(400 MHz, CDCl3) d: 5.88 (dd, J¼15.0, 8.0 Hz, 1H), 5.67
(d, J¼15.0 Hz, 1H), 4.60 (dd, J¼8.0, 6.0 Hz, 1H), 4.15
(ddd, J¼8.0, 6.0, 5.0 Hz, 1H), 2.94–2.84 (m, 2H),
2.67–2.60 (m, 2H), 2.18 (t, J¼7.2 Hz, 2H), 2.06–2.00,
1.88–1.82 (m, 1H each), 1.80–1.75 (m, 2H), 1.58–1.23 (m,
18H), 1.49 (s, 3H), 1.43 (s, 9H), 1.37 (s, 3H), 0.88 (t,
J¼6.6 Hz, 3H); 13C NMR (100 MHz, CDCl3) d: 173.2,
136.7, 130.5, 108.1, 79.9, 78.9, 78.4, 54.5, 42.3, 35.6,
31.9, 30.7, 29.6, 29.1, 29.0, 28.4, 28.1 (3C), 27.1, 27.1,
25.9, 25.7, 25.5, 25.0, 23.7, 22.6, 14.0.
7.4.2. (12R,13S)-(E)-9-(1,3-Dithian)-13-hydroxy-12-tri-
isopropylsiloxy-10-octadecaenoic acid-tert-butyl ester
((L)-25). To a solution of (ꢀ)-19 (26.3 mg, 39.1 mmol) in
t-BuOH (800 mL) was added KOt-Bu (17.5 mg, 157 mmol)
at rt. The resultant mixture was stirred at rt for 16 h, treated
with 1.0 N HCl (1 mL), and then extracted with CHCl3
(3 mLꢂ5). The combined organic layer was washed with
satd aq NaCl (5 mL), dried over Na2SO4, filtered, and con-
centrated. The residue was purified by column chromato-
graphy (hexane/AcOEt¼40:1) to give (ꢀ)-25 (11.5 mg,
47%) as a colorless oil. Rf¼0.44 (silica gel, hexane/AcOEt¼
5:1); [a]2D3 ꢀ12.5 (c 0.72, CHCl3); IR (KBr) n cmꢀ1: 3434
Acknowledgements
1
(s), 1724 (m), 1625 (m); H NMR (270 MHz, CDCl3) d:
This work was supported by the Ministry of Education,
Science, Sports, and Culture, Japan and the Japan Keirin
Association, a Grant of the 21st Century COE Program.
We acknowledge and thank Ms. Noriko Sato for NMR
measurements and Ms. Chikako Sakabe and Ms. Akiko
Nakagawa for mass spectrometric analysis.
5.93 (dd, J¼15.5, 7.3 Hz, 1H), 5.64 (d, J¼15.5 Hz, 1H),
4.29 (dd, J¼7.3, 3.3 Hz, 1H), 3.77–3.67 (m, 1H), 2.97–
2.84 (m, 2H), 2.66–2.57 (m, 2H), 2.18 (t, J¼7.6 Hz, 2H),
2.01 (br s, 1H), 1.82–1.76 (m, 2H), 1.67–1.58 (complex m,
20H), 1.44 (s, 9H), 1.14–1.07 (m, 21H), 0.87 (t, J¼6.6 Hz,

9496
T. Shirahata et al. / Tetrahedron 62 (2006) 9483–9496
(for C9). These results confirmed that the dihydroxylation
References and notes
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O
OMTPA
S –0.17–0.02
S
t-BuO
+0.03
–0.31 –0.12
OTIPS
=
–
δ(–) δ(+)
δ
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(+)/(–)-MTPA-(–)-18
O
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OTIPS
δ=δ(–)
–
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9S
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OTIPS
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