Absolute stereochemistry and conformational analysis of achaetolide isolated from Ophiobolus sp.

Article abstract of DOI:10.1016/j.tet.2009.07.005

Achaetolide, as reported by Bodo et?al. in 1983, was isolated from a fermentation broth of Ophiobolus sp. We established the absolute stereochemistry of achaetolide to be 3S,6R,7S,9R by way of relative stereochemical assignment with ¹H NMR anal

Full text of DOI:10.1016/j.tet.2009.07.005

Tetrahedron 65 (2009) 7464–7467
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Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Absolute stereochemistry and conformational analysis of achaetolide isolated
from Ophiobolus sp.
Wilanfranco Caballero Tayone, Saori Shindo, Takanori Murakami, Masaru Hashimoto,
*
Kazuaki Tanaka, Noboru Takada
Faculty of Agriculture and Life Science, Hirosaki University, 3 Bunkyo-cho, Hirosaki 036-8561, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 3 June 2009
Received in revised form 2 July 2009
Accepted 2 July 2009
Available online 7 July 2009
Achaetolide, as reported by Bodo et al. in 1983, was isolated from a fermentation broth of Ophiobolus sp.
We established the absolute stereochemistry of achaetolide to be 3S,6R,7S,9R by way of relative ste-
reochemical assignment with 1H NMR analyses employing the corresponding acetonide, determination
of C3 and C9 stereochemistries by an extended Mosher method.
Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction
obtained consisted of pairs of signals with a 9:1 ratio, despite the
HPLC chromatograms suggesting single compound. Since the ratio
Polyhydrolxylated 10-membered macrolides, such as aspinolide
B,1 decarestrictines A–D,2–5 microcarpalide,6 pinolidoxin,7 and
herbarmins I–II,8 have been isolated from fungal sources. Since
some of these exhibit potent biological activities (cholesterol bio-
synthesis inhibition activity, antimicrofilament activity, and phy-
totoxicity, and so on), this class of lactones has received special
attention over recent years.9 They have also attracted synthetic
chemists in relation to the development of effective methodologies
constructing medium-sized rings.10–15 Within this family, the ste-
reochemistry of achaetolide (1), originally isolated from a culture
broth of Achaetomium cristalliferum by Bodo et al., remains un-
known. We succeeded in establishing the absolute stereochemistry
of 1 by way of relative stereochemical assignment with 1H NMR
analyses employing the corresponding acetonide, determination of
C3 and C9 stereochemistries by extended Mosher method,16 and
confirmation of C3 position by a combination of conversion into
in the 1H NMR spectra varied by solvents (2:1 in CD3OD, 5:3 in
acetone-d6), we concluded that the sample was pure but involved
conformational isomers in the 1H NMR spectra. Further analysis
involving 2D NMR spectra suggested a 10-membered macrolide
structure (Fig. 1), which had been reported by Bodo et al. as
achaetolide, a transpiration enhancer of cut barley leaves.17 The 1H
NMR data for the major conformer in CDCl3 accorded well with
those in the literature, although there is no mention about the
presence of conformational isomers in the 1H NMR spectra. Since
the relative and absolute stereochemistry of 1 has not been dis-
cussed, we established it as described below.
O
1
9
6
O
5
7
3
HO
OH
4
methyl L-malate with chiral GC analysis. In this paper we would also
like to discuss about conformational isomers.
OH
Figure 1. Structure of achaetolide (1).
2. Results and discussion
It was found that a sole conformer was detected in the 1H NMR
spectra when the 6-O-7-O-isopropylidene derivative 2 was mea-
sured in CDCl3/C6D6 (1:1). Isopropylidene 2 was obtained in 72%
yield by treating 1 with HClO4 in acetone (Scheme 1). This finding
allows us to discuss the relative stereochemistry of the macrolide
moiety based on the 1H NMR spectra. Although the coupling con-
stant between H6 and H7 (6.1 Hz) did not provide crucial in-
formation about the 1,3-dioxolane ring, a remarkable NOE between
these protons was observed to reveal a cis-relationship on the
dioxolane ring (Fig. 2). Large coupling constants for JH7–H
Achaetolide (1) was obtained as oil by conventional extraction/
chromatography operations from a culture broth of Ophiobolus sp.
isolated from dead stem of Achillea alpina ssp. pulchra. The ESIMS
spectrum provided
a protonated molecular ion signal at
m/z¼301.2033, thus establishing its molecular formula (C16H28O5).
In the 1H NMR spectra in CDCl3, most resonances of the sample thus
* Corresponding author. Tel./fax: þ81 172 39 3771.
E-mail address: takada@cc.hirosaki-u.ac.jp (N. Takada).
a8
(10.2 Hz) and JH
(9.7 Hz) indicated pseudo-anti orientations
8–H9
a
0040-4020/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2009.07.005

W.C. Tayone et al. / Tetrahedron 65 (2009) 7464–7467
7465
O
C7H15
moiety. These conformers might be interconverted on the NMR
time scale.
HClO4, acetone
(72%)
K2CO3
O
1
We next investigated the absolute stereochemistry of 1. It was
found that basic treatment of 2 in methanol gave acyclic diol 3 in
97% yield. After 3 was converted to bis-(S)- and (R)-MTPA esters19
(4a: 80%, 4b: 70%, respectively) with the corresponding MTPACl,
MeOH
HO
O
(97%)
O
2
C7H15
RO
the Dd values (dS
dR, ppm) in the 1H NMR spectra (CDCl3) were
ꢀ
1) O3
2) H2O2
MeOOC
RO
MeOOC
obtained as shown in Figure 4, to which applied to the empirical
rule by Kusumi et al.16 to suggest the chiralities for C3 and C9 po-
sitions as 3S,9R. Taking the above discussions into account, our
3) TMSCHN2
(13% from 3)
HO
COOMe
O
O
5
studies disclosed the absolute stereochemistry of
3S,6R,7S,9R.
1 to be
3: R = H
(R)- or (S)-MTPACl
4a: R = (S)-MTPA
pyridine
(80%)
4b: R = (R)-MTPA
(70%)
OMTPA
+0.05
O
+0.10
-0.09
9R
C5H11
-0.16
MeO
-0.06
3S
+0.07
-0.24
O
+0.02
OMTPA
Scheme 1.
-0.02
-0.03
O
4a or 4b
+0.02 -0.04
Figure 4. Dd values (dS dR) for the MTPA esters 4a and 4b in ppm.
ꢀ
H
9R*
NOE
O
H
Hβ
O
H
C7H15
O
The stereochemistry of C3 was further confirmed by chiral GC
method. Ozonolysis of 3 followed by oxidative work-up gave malic
acid monomethyl ester, which was purified after conversion into
dimethyl malate 5 (13% from 3) by TMSCHN2.20 The chiral GC
CH3
CH3
8
7S*
H
6R*
Hα
4
3S*
H
O
5
HO
conditions (Rt-
b
DEXmÔ [0.25 mm IDꢁ30 m], at 100–200 ꢂC for
H
15 min) clearly separated the racemic dimethyl malate, with suffi-
cient reproducibility, to show a pair of peaks at tR¼9.20 and
9.25 min. The equipped MS detector provided identical mass pro-
files to confirm these GC peaks.21 On the other hand, 3 yielded
a single peak at tR¼9.25 min. Since authentic (S)-dimethyl malate
showed identical retention time (tR¼9.25 min) as 3, the absolute
chemistry of C3 position could be confirmed to be S-form, which
consisted with that suggested by the extended Mosher method.
Finally, conformation of 1 is discussed. As mentioned, 1 was
observed as a mixture of conformers in the 1H NMR spectra. Al-
though it was difficult to evaluate hydrogen bondings in solvents by
modeling calculations, the conformation search calculations with
Figure 2. Stereochemistry of the C3–C9 moiety and characteristic NOEs observed in
CDCl3/C6D6 (1:1).
for H7/Ha8 and Ha8/H9. Interestingly the Hb8 appeared as a doublet
(J¼15.8 Hz) coupled only with Ha8, which suggests perpendicular
relationships for H7/Hb8 and Hb8/H9. The NOESY spectrum pro-
vided correlation peaks H5/Ha8, H6/H7, H6/H9, and H7/H9, con-
firming the stereochemical relationship for the C6–C9 moiety. The
coupling constants for H3/H4 and H5/H6 (8.3 and 9.0 Hz, re-
spectively) and strong NOESY signals H3/H5 and H4/H6 disclosed
the C3–C6 plane. Combining these results revealed a conformation
as depicted in Figure 2 and the relative stereochemistry of C3–C9
HF-631G provided two characteristic conformers of model B within
*
2.0 kcal/mol from that with lowest steric energy as shown in Fig-
ure 5. These were conformational isomers caused by flipping the
C3–C6 plane.22 Figure 6 shows an olefinic proton region in the 1H
NMR spectrum of 1 in CDCl3 disclosing that 1 consists pre-
dominantly of two conformers with a 9:1 ratio. The signal profiles
differ between the major and minor conformers, which indicate
that the circumstances around the C4]C5 double bonds for these
conformers are dissimilar to each other. These discussions are
consistent with those based on the molecular modeling
calculations.
moiety to be 3S
*
,6R
*
,7S
*
,9R .
*
However, conformation around C1–C3 did not seem to be fixed
because the coupling constants for Ha2/H3 and Hb2/H3 were both
8.0 Hz. A conformational search of the model A with semiempirical
AM118 found six stable conformers within 2.0 kcal/mol from the
global minimum conformation. As shown in Figure 3, all of these
conformers took a similar conformation for C3–C9 and satisfied the
coupling constants and NOEs discussed above. These conformers
results in conformational change of C3OH group and O–C1–C2
Figure 3. Six stable conformers of model A within 2.0 kcal/mol obtained by confor-
Figure 5. Two characteristic stable conformers of model B within 2.0 kcal/mol ob-
mational search with AM1.
tained by conformational search with HF-631G .
*

7466
W.C. Tayone et al. / Tetrahedron 65 (2009) 7464–7467
8.5 Hz, H-9), 4.92 (1Hꢁb, m, H-9), 5.41 (1Hꢁb, dd, J¼8.8, 16.4 Hz, H-
4), 5.57 (1Hꢁb, dd, J¼7.3, 16.4 Hz, H-5), 5.69 (1Hꢁa, ddd, J¼1.4, 2.8,
15.8 Hz, H-5), 5.97 (1Hꢁa, dd, J¼3.4, 15.8 Hz, H-4); 1H NMR (ace-
tone-d6, 500 MHz, a¼0.63, b¼0.37)
d
0.86 (3H, t, J¼7.1 Hz, H-16),
1.27 (10H, m), 1.37 (1Hꢁb, d, J¼15.8 Hz, H-8
J¼15.7 Hz, H-8 ), 1.49 (2Hꢁa, m, H-10), 1.52 (1Hꢁb, m, H-10
(1Hꢁb, m, H-10 ), 2.22 (1Hꢁb, dd, J¼8.8, 13.4 Hz, H-2
(1Hꢁa, ddd, J¼8.6, 9.8, 15.7 Hz, H-8 ), 2.34 (1Hꢁb, ddd, J¼5.8, 11.3,
15.8 Hz, H-8 ), 2.49 (1Hꢁa,
), 2.44 (1Hꢁa, dd, J¼3.5, 11.7 Hz, H-2
dd, J¼3.7, 11.7 Hz, H-2 ), 2.93 (1Hꢁb, dd, J¼7.7, 13.4 Hz, H-2 ), 3.30
b
), 1.40 (1Hꢁa, d,
), 1.66
), 2.34
b
b
b
a
a
a
b
a
a
(1Hꢁb, br s, –OH), 3.46 (1Hꢁb, m, H-7), 3.65 (1Hꢁa, br d, J¼9.8 Hz,
H-7), 3.75 (1Hꢁb, br s, –OH), 3.77 (1Hꢁa, br s, –OH), 3.83 (2Hꢁa, br
s, –OH), 4.03 (1Hꢁb, br s, –OH), 4.25 (1Hꢁb, dd, J¼3.3, 7.5 Hz, H-6),
4.43 (1Hꢁa, br s, H-6), 4.45 (1Hꢁb, m, H-3),4.66 (1Hꢁa, dt, J¼8.6,
8.4 Hz, H-9), 4.67 (1Hꢁa, m, H-3), 4.92 (1Hꢁb, ddd, J¼4.8, 8.7,
11.3 Hz, H-9), 5.41 (1Hꢁb, dd, J¼8.8, 16.4 Hz, H-4), 5.54 (1Hꢁb, dd,
J¼7.5, 16.4 Hz, H-5), 5.68 (1Hꢁa, dd, J¼2.7, 15.7 Hz, H-5), 5.99
(1Hꢁa, dd, J¼5.6, 15.7 Hz, H-4); HRESIMS m/z found 301.2033
[MþH]þ, calcd for C16H29O5 301.2015, found 283.1914
[MþHꢀH2O]þ, calcd for C15H27O4: 283.1909. The 1H and 13C NMR
spectral data in CDCl3 regarding the major conformer accorded well
with those reported by Bodo et al.
Figure 6. Olefinic proton region in the 1H NMR spectrum of 1 in CDCl3 (Small signals
due to the third isomer were also seen at 6.05 ppm.).
We succeeded in revealing the absolute stereochemistry of
achaetolide (1) by converting it into the isopropylidene 2. We pri-
marily revealed that 1 exists as a mixture of conformers in the 1H
NMR spectra. Thus far, we have found no potent biological activi-
ties; further biological investigations of 1 are presently under way
in our laboratories.
3.3. Achaetolide C6,C7-O-acetonide (2)
To a stirred solution of 1 (5.3 mg, 16 mmol) in acetone (1 mL) was
added perchloric acid (1 drop). The reaction mixture was stirred at
room temperature for 20 min. After neutralization with satd
NH4Claq (10 mL), the reaction mixture was extracted with EtOAc
(10 mLꢁ3). The combined EtOAc layer was washed with brine
(5 mL), dried with Na2SO4, and concentrated in vacuo. The residue
3. Experimental
3.1. General
was purified by silica gel column chromatography (hexane/
The 1H (500 MHz) and 13C (125 MHz) NMR spectra were recor-
24
EtOAc¼3:1) to give 2 (4.5 mg, 72%) as a colorless oil. [
a
]
ꢀ33 (c
D
ded in CDCl3
(dH 7.24 ppm), CD3OD (dH 3.30 ppm), acetone-d6
(dH
0.35, CHCl3); IR (film) nmax¼3444 (O–H), 3000 (C–H), 2920 (C–H),
2.04 ppm), or CDCl3/C6D6 (1:1)
(dH 7.16 ppm for C6HD5, dC
2870 (C–H), 1732 (O–C]O) cmꢀ1
500 MHz)
s, acetonide–Me), 1.30 (1H, m, H-10
1.44 (1H, d, J¼15.8 Hz, H-8 ), 1.51 (1H, m, H-10
J¼9.7, 10.2, 15.8 Hz, H-8 ), 2.16 (1H, dd, J¼8.2, 13.1 Hz, H-2
(1H, dd, J¼7.8, 13.1 Hz, H-2
;
1H NMR (CDCl3/C6D6¼1:1,
128.0 ppm for C6D6) using a JEOL JNM-ECA500 spectrometer. The
EI-GC/MS spectrum was measured with a Shimadzu GC–MS-
QP2010 spectrometer using an Rt-bDEXmÔ (Shimadzu GLC Inc.)
chiral capillary column (0.25 mm IDꢁ30 m). The HRESI/MS spec-
trum was obtained by a HITACHI NanoFrontier LD spectrometer.
Measurements of IR spectra were performed with a HORIBA FT-720
spectrometer. The optically rotation value were measured using
a HORIBA SEP-700 spectrometer. Chemicals used in these experi-
ments were obtained from Wako Pure Chemical Industries Ltd. and
Nacalai Tesque Inc.
d
0.81 (3H, t, J¼6.9 Hz, H-16),1.08–1.16 (10H, m),1.20 (3H,
b
), 1.34 (3H, s, acetonide–Me),
), 2.11 (1H, ddd,
), 2.64
b
a
a
b
a
), 3.92 (1H, dd, J¼6.0, 10.2 Hz, H-7), 4.23
(1H, ddd, J¼7.8, 8.2, 8.3 Hz, H-3), 4.39 (1H, dd, J¼6.0, 9.0 Hz, H-6),
4.64 (1H, ddd, J¼5.5, 9.0, 9.7 Hz, H-9), 5.42 (1H, dd, J¼8.3, 16.0 Hz,
H-4), 5.56 (1H, dd, J¼9.0, 16.0 Hz, H-5); 13C NMR (CDCl3/C6D6¼1:1,
125 MHz)
d 14.0 (q, C-16), 22.6 (t, C-15), 25.3 (q, acetonide–Me),
25.6 (t, C-14), 28.0 (q, acetonide–Me), 29.2 (t, C-13), 29.3 (t, C-12),
31.8 (t, C-11), 35.7 (t, C-10), 38.0 (t, C-8), 43.8 (t, C-2), 70.0 (d, C-3),
74.9 (d, C-9), 77.3 (d, C-6), 80.8 (d, C-7), 108.6 (s, acetal), 129.6 (d, C-
5), 133.5 (d, C-4), 170.0 (s, C-1); HRESIMS m/z found 341.2317
[MþH]þ, calcd for C19H33O6 341.2328.
3.2. Isolation of achaetolide (1)
Ophiobolus sp. collected at Rishiri Island (Hokkaido) in 2007 was
cultured in a potato/sucrose medium [600 mL, prepared from
a potato extract (from 120 g potato), 12 g of sucrose, and water] at
25 ꢂC for 40 days on a rotary shaker (100 rpm). After filtration, the
culture broth was extracted with ethyl acetate (600 mLꢁ2) and
concentrated in vacuo. The residue was subjected to silica gel col-
3.4. Methyl (3S,6R,7S,9R,E)-C6,C7-O-isopropilydene-3,6,7,9-
tetrahydroxydodec-4-enoate (3)
A solution of 2 (6.6 mg, 18
mmol) in MeOH (2 mL) was stirred
umn chromatography (hexane/ethyl acetate¼1:5) to afford achae-
with K2CO3 (9.8 mg, 97 mol) at room temperature. After stirring
m
23
tolide (1) 42.1 mg. [
a]
ꢀ27 (c 0.52, MeOH); IR (film) nmax¼3448
for 9 h, the suspension was poured into satd NH4Claq (3 mL) and
extracted with EtOAC (20 mL). The EtOAc layers were combined,
washed with brine (3 mL), dried with Na2SO4, and concentrated in
D
(O–H), 3255 (O–H), 2950 (C–H), 2870 (C–H), 1709 (O–C]O) cmꢀ1
;
1H NMR (CD3OD, 500 MHz, a¼0.67, b¼0.33)
d
0.89 (3H, t, J¼7.1 Hz,
), 1.41 (1Hꢁa, d,
), 1.70
), 2.37
), 2.49 (1Hꢁa, dd, J¼3.4,
), 2.52 (1Hꢁa, dd, J¼3.8, 11.8 Hz, H-2 ), 2.93 (1Hꢁb,
H-16), 1.28 (10H, m), 1.38 (1Hꢁb, d, J¼14.6 Hz, H-8
b
vacuo. Silica gel column chromatography of the residue (hexane/
23
J¼15.3 Hz, H-8
(1Hꢁb, m, H-10
(1Hꢁa, m, H-8 ), 2.37 (1Hꢁb, m, H-8
11.8 Hz, H-2
dd, J¼7.6, 13.5 Hz, H-2
a
), 1.52 (2Hꢁa, m, H-10), 1.55 (1Hꢁb, m, H-10
b
b
EtOAc¼1:1) gave 3 (7.5 mg, 97%) as a colorless oil. [
a]
ꢀ2.7 (c 0.68,
D
a
), 2.26 (1Hꢁb, dd, J¼8.7, 13.5 Hz, H-2
MeOH); IR (film) nmax¼3448 (O–H), 2945 (C–H), 2880 (C–H), 1739
(O–C]O) cmꢀ1 1H NMR (CDCl3, 500 MHz)
0.85 (3H, t, J¼6.8 Hz,
H-16), 1.35 (3H, s, acetonide–Me), 1.30–1.20 (10H, m), 1.39 (1H, ddd,
J¼3.3, 5.3, 14.2 Hz, H-8 ), 1.40–1.45 (2H, m, H-10), 1.46 (3H, s, ace-
tonide–Me), 1.60 (1H, ddd, J¼2.9, 10.0, 14.2 Hz, H-8 ), 1.91 (1H, br s,
9-OH), 2.53 (1H, dd, J¼5.0, 16.2 Hz, H-2 ), 2.56 (1H, dd, J¼5.0,
b
a
;
d
b
a
a), 3.44 (1Hꢁb, m, H-7), 3.64 (1Hꢁa, d,
b
J¼9.9 Hz, H-7), 4.28 (1Hꢁb, dd, J¼3.2, 7.3 Hz, H-6), 4.44 (1Hꢁa, m,
H-6), 4.44 (1Hꢁb, m, H-3), 4.68 (1H, m, H-3), 4.72 (1H, dt, J¼6.4,
a
b

W.C. Tayone et al. / Tetrahedron 65 (2009) 7464–7467
7467
16.2 Hz, H-2
a
), 2.87 (1H, br s, 3-OH), 3.69 (3H, s, ester–Me), 3.77
dimethyl malate (5) (0.2 mg, 13% from 3) as a colorless oil. The
methanolic solution of 5 (2 L, 0.1 mg/mL) was injected into the
GC–MS instrument (splitter ratio: 100; column: Rt- DEXmÔ
(1H, m, H-9), 4.42 (1H, ddd, J¼3.3, 6.5, 10.0 Hz, H-7), 4.55 (1H, dd,
J¼6.5, 7.0 Hz, H-6), 4.56 (1H, m, H-3), 5.71 (1H, ddd, J¼1.0, 7.0,
15.5 Hz, H-5), 5.76 (1H, dd, J¼5.0, 15.5 Hz, H-4); 13C NMR (CDCl3,
m
b
(0.25 mm IDꢁ30 m, Shimadzu GLC LTD.); temperature: 100–200 ꢂC
for 15 min, gas flow: 1.33 mL/min). (2R)- and (2S)-Dimethyl malate
(5) were eluted from the column after 9.2 and 9.3 min, respectively.
125 MHz)
d 14.1 (q, C-16), 22.6 (t, C-15), 25.6 (q, acetonide–Me),
25.7 (t, C-14), 28.2 (q, acetonide–Me), 29.2 (t, C-13), 29.6 (t, C-12),
31.8 (t, C-11), 37.4 (t, C-8), 37.8 (t, C-10), 41.0 (t, C-2), 51.9 (q, ester–
Me), 68.0 (d, C-12), 69.1 (d, C-9), 75.1 (d, C-7), 78.5 (d, C-6), 108.3 (s,
acetal–C), 127.3 (d, C-5), 134.6 (d, C-4), 172.5 (s, C-1); HRESIMS m/z
found: 373.2578 [MþH]þ, calcd for C20H37O6 373.2590.
1H NMR (CDCl3, 400 MHz)
dd, J¼4.7, 16.5 Hz), 3.15 (1H, br d, J¼5.1 Hz), 3.70 (3H, s), 3.79 (3H, s),
d
2.79 (1H, dd, J¼5.5, 16.5 Hz), 2.84 (1H,
4.49 (1H, ddd, J¼4.7, 5.1, 5.5 Hz); EIMS m/z 103 [MꢀCO2CH3]þ.
Acknowledgements
3.5. Methyl (3S,6R,7S,9R,E)-C6,C7-O-isopropilydene-3,6,7,9-
tetrahydroxydodec-4-enoate C2,C9-O-bis-(S)-MTPA ester (4a)
Part of this work was supported by a Grant for Priority Research
designated by the President of Hirosaki University.
A solution of 3 (0.5 mg, 1.3 mmol) in pyridine (0.2 mL) was stir-
red with (R)-methoxytrifluorophenylacetyl (MTPA) chloride
(5.0 mg) at room temperature for 2 days. After quenching with satd
NaHCO3aq (2 mL), the reaction mixture was extracted with EtOAc
(5 mLꢁ3). The combined EtOAc layer was washed with satd
NH4Claq (2 mL) and brine (2 mL), dried with Na2SO4, and concen-
trated in vacuo. Silica gel column chromatography of the residue
Supplementary data
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.tet.2009.07.005.
(0.3 g, hexane/EtOAc¼5:1) gave ester 4a (0.8 mg, 80%) as a colorless
References and notes
16
oil. [
a]
ꢀ90 (c 0.06, CHCl3); IR (film) nmax¼2931 (C–H), 2854 (C–
D
H), 1747 (O–C]O) cmꢀ1
J¼7.2 Hz, H-16), 1.16–1.31 (10H, m), 1.26 (3H, s), 1.43 (3H, s), 1.47–
1.63 (4H, m), 2.59 (1H, dd, J¼4.9, 16.3 Hz, H-2 ), 2.75 (1H, dd, J¼8.9,
16.3 Hz, H-2 ), 3.47 (3H, s), 3.55 (3H, s), 3.57 (3H, s), 3.88 (1H, m, H-
; d 0.84 (3H, t,
1H NMR (CDCl3, 500 MHz)
1. Fuchser, J.; Zeeck, A. Liebigs Ann./Recueil 1997, 87.
2. Grabley, S.; Granzer, E.; Hu¨tter, K.; Ludwig, D.; Mayer, M.; Thiericke, R.; Till, G.;
Wink, J.; Hoechst, A. G.; Phillips, S.; Zeeck, A. J. Antibiot. 1992, 45, 56.
3. Go¨hrt, A.; Zeeck, A.; Hu¨ tter, K.; Thiericke, R.; Kirsch, R.; Kluge, H. J. Antibiot.
1992, 45, 66.
4. Grabley, S.; Hammann, P.; Hu¨ tter, K.; Kirsch, R.; Kluge, H.; Thiericke, R.;
Hoechst, A. G.; Mayer, M.; Zeeck, A. J. Antibiot. 1992, 45, 1176.
5. Ayer, W. A.; Sun, M.; Browne, L. M.; Brinen, L. S.; Clardy, J. J. Nat. Prod. 1992,
55, 649.
b
a
7), 4.28 (1H, dd, J¼4.9, 6.5 Hz, H-6), 5.18 (1H, m, H-9), 5.73 (1H, dd,
J¼6.6, 15.6 Hz, H-4), 5.76 (1H, dd, J¼4.9, 15.6 Hz, H-5), 5.86 (1H, m,
H-3), 7.34–7.39 (6H, m), 7.44–7.47 (2H, m), 7.52–7.54 (2H, m);
HRESIMS m/z found 827.3198 [MþH]þ, calcd for C40H50F6NaO10
827.3206.
6. Ratnayake, A. S.; Yoshida, W. Y.; Mooberry, S. L.; Hemscheidt, T. Org. Lett. 2001,
3, 3479.
7. Fu¨ rstner, A.; Radkowski, K. Chem. Commun. 2001, 671.
8. Fu¨rstner, A.; Radkowski, K.; Wirtz, C.; Goddard, R.; Lehmann, C. W.; Mynott, R. J.
Am. Chem. Soc. 2002, 124, 7061.
9. Dra¨ger, G.; Kirschning, A.; Thiericke, R.; Zerlin, M. Nat. Prod. Rep. 1996, 13, 365.
10. Rousseau, G. Tetrahedron 1995, 51, 2777.
3.6. Methyl (3S,6R,7S,9R,E)-C6,C7-O-isopropilydene-3,6,7,9-
tetrahydroxydodec-4-enoate C2,C9-O-bis-(R)-MTPA ester (4b)
11. Still, W. C.; Galynker, I. Tetrahedron 1981, 23, 3981.
12. Shirahama, H.; Matsumoto, T. J. Synth. Org. Chem. Jpn. 1985, 43, 205.
13. Andrus, M. B.; Shih, T.-L. J. Org. Chem. 1996, 61, 8780.
In a similar manner as that described for bis-(S)-MTPA ester, 3
(0.7 mg, 2
give 4b (1.1 mg, 70%). [
(C–H), 2854 (C–H), 1747 (O–C]O) cmꢀ1; 1H NMR (CDCl3, 500 MHz)
0.84 (3H, t, J¼7.3 Hz, H-16), 1.14–1.29 (10H, m), 1.30 (3H, s), 1.41
(3H, s), 1.48–1.62 (4H, m), 2.61 (1H, dd, J¼4.7, 16.4 Hz, H-2 ), 2.78
), 3.48 (3H, s), 3.52 (3H, s), 3.64 (3H, s),
mmol) was treated with (S)-MTPA chloride (5.0 mg) to
14. Pilli, R. A.; Victor, R. M.; de Meijere, A. J. Org. Chem. 2000, 65, 5910.
15. Davoli, P.; Spaggiari, A.; Castagnetti, L.; Plati, F. Org. Biomol. Chem. 2004, 2, 38.
16. Otani, I.; Kusumi, T.; Kashman, Y.; Kakisawa, H. J. Am. Chem. Soc. 1991, 113, 4092.
17. Bodo, B.; Molho, L.; Davoust, D.; Molho, D. Phytochemistry 1983, 22, 447.
18. Spartan 04 version 1.0.3, Wavefunction, 18401 Von Karman Avenue, Suite 370,
Irvine, CA 92612, USA.
15
a
]
ꢀ70 (c 0.09, CHCl3), IR (film) nmax¼2931
D
d
b
19. Dales, J. A.; Mosher, H. S. J. Am. Chem. Soc. 1973, 95, 512.
20. Hashimoto, N.; Aoyama, T.; Shioiri, T. Chem. Pharm. Bull. 1981, 29, 1475.
21. The MS conditions provided m/z¼103 [Mꢀ(CH3O)]þ but not its molecular ion
signal m/z¼162.
(1H, dd, J¼9.2, 16.4 Hz, H-2
a
4.12 (1H, m, H-7), 4.44 (1H, dd, J¼5.6, 5.7 Hz, H-6), 5.20 (1H, m,
H-9), 5.66 (1H, dd, J¼6.8, 15.5 Hz, H-4), 5.71 (1H, dd, J¼5.6, 15.5 Hz,
H-5), 5.85 (1H, ddd, J¼4.7, 6.8, 9.2 Hz, H-3), 7.31–7.41 (6H, m), 7.43–
7.46 (2H, m), 7.51–7.54 (2H, m); HRESIMS m/z found 827.3220
[MþH]þ, calcd for C40H50F6NaO10 827.3206.
22. Molecular modeling calculation provided the expected conformation of tran-
sition state, as shown below, through flipping the C3–C6 plane, whose acti-
vation energy was 16.2 kcal/mol. It is well-known that two N-methyl protons in
DMF are observed as two distinct signals in 1H NMR spectrum, resulting from
restriction of C–N bond rotation in amide moiety. This energy barrier was es-
timated to be 19.2 kcal/mol. Therefore, flipping of the C3–C6 plane may account
for the observation of conformational mixture in 1H NMR spectrum of 1.
3.7. Chiral GC–MS analysis of dimethyl malate (5) derived
from achaetolide
A solution of 3 (3.5 mg, 9.4 mmol) in MeOH (1 mL) was stirred at
ꢀ78 ꢂC. A steam of ozone was passed through for 20 min. After
warming to room temperature, 90% HCOOHaq (1 mL) and 34%
H2O2aq (0.5 mL) were added. The resulting solution was stood
overnight and then concentrated in vacuo to give the crude mate-
rials, which were diluted again with benzene (700
(250 L). After the addition of trimethylsilyldiazometane (2.0 M in
diethyl ether, 50 L) at room temperature, the resulting solution
mL) and MeOH
m
m
was stood for 1 h and then concentrated in vacuo. Silica gel column
chromatography of the residue (0.3 g, hexane/EtOAc¼1:2) gave