Total synthesis of epicoccamides A and D via olefin cross-metathesis

Article abstract of DOI:10.1016/j.tetlet.2014.06.040

Epicoccamides A and D were synthesized through a route that utilizes fragment coupling via olefin cross-metathesis as a key step. The right-hand segment of the epicoccamides was synthesized by a tandem O-acylation-migration reaction, and the left-hand segments were stereoselectively synthesized through a modified version of Crich's β-selective mannosylation. The previously assigned absolute configuration of the epicoccamide D was confirmed, and that of epicoccamide A was assigned as (5S,2′S) based on the NMR and CD spectra. This Letter provides the first example of the total synthesis of epicoccamide A.

Full text of DOI:10.1016/j.tetlet.2014.06.040

Tetrahedron Letters 55 (2014) 4350–4354
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Total synthesis of epicoccamides A and D via olefin cross-metathesis
⇑
Arata Yajima , Akihiro Kawajiri, Ayaka Mori, Ryo Katsuta, Tomoo Nukada
Department of Fermentation Science, Faculty of Applied Bioscience, Tokyo University of Agriculture (NODAI), Sakuragaoka 1-1-1, Setagaya-ku, Tokyo 156-8502, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Epicoccamides A and D were synthesized through a route that utilizes fragment coupling via olefin
cross-metathesis as a key step. The right-hand segment of the epicoccamides was synthesized by a
tandem O-acylation–migration reaction, and the left-hand segments were stereoselectively synthesized
through a modified version of Crich’s b-selective mannosylation. The previously assigned absolute con-
figuration of the epicoccamide D was confirmed, and that of epicoccamide A was assigned as (5S,20S)
based on the NMR and CD spectra. This Letter provides the first example of the total synthesis of
epicoccamide A.
Received 7 May 2014
Revised 3 June 2014
Accepted 9 June 2014
Available online 16 June 2014
Keywords:
Epicoccamide
Tetramic acid
Ó 2014 Elsevier Ltd. All rights reserved.
b-Mannoside
Olefin cross-metathesis
Absolute configuration
Naturally occurring tetramic acids constitute an important class
of bioactive natural products and have attracted increasing interest
among natural product chemists and synthetic organic chemists.1
In most cases, tetramic acids are found in their 3-acylated forms,
in which various substituents or stereogenic centers are present
in the acyl group.2 Epicoccamides are naturally occurring
3-acyltetramic acids bearing b-mannosylated fatty acid chains.
Epicoccamide A (1), originally termed simply epicoccamide, was
isolated from the jellyfish-derived fungus Epicoccum purpurascens
(also known as Epicoccum nigrum) by Wright and co-workers in
2003,3 whereas epicoccamides B–D (2–4) were isolated by
Hertweck et al. in 2007 from an Epicoccum sp. growing within the
fruiting body of the tree fungus Pholiota squarrosa (Fig. 1).4
Recently, epicoccamide A was also isolated from an endophytic fun-
gus Epicoccum sp. CAFTBO associated with Theobroma cacao.5 The
structurally related, bioactive, glycosylated tetramic acids ancori-
nosides (5)6 and virgineones (6)7 have been isolated from the mar-
ine sponges Ancorina sp. and Lachnum virgineum, respectively.
Interestingly, epicoccamides B–D showed weak to moderate
antiproliferative effects on mouse fibroblast (L-929) and human
leukemia cell lines (K-562) and cytotoxicity against HeLa cells. No
significant biological activity has yet been reported for epicocca-
mide A. Based on our interest in the structure–activity relationships
of epicoccamides, we attempted to synthesize these compounds.
In the initial stages of our project, the absolute configuration of
the epicoccamides had yet to be determined. Hertweck et al.
suggested that epicoccamide D comprised a 2:3 mixture of isomers
with mixed stereochemistry at C-5 based on the HPLC analysis of
its degradation products, although the possibility of isomerization
during derivatization could not be fully excluded.4 Recently,
Loscher and Schobert reported the first total synthesis of epicocca-
mide D.8 These researchers concluded that the absolute configura-
tion of the natural epicoccamide D is 5S,20S based on comparisons
of its 13C NMR spectra and specific rotation values. These findings
led us to investigate the stereochemical assignments of epicocca-
mides A and D.
From a structural point of view, the highly polar tetramic acid
and sugar portions of the compounds are located in the terminal
sites of the molecules and are linked together by simple long
chains. Because it is well known that 3-acyltetramic acids are bio-
synthesized through a Dieckmann-type condensation of the corre-
sponding b-ketoacid with the amino acid moiety,9 epicoccamides
and their related compounds should be biologically constructed
from the following three major elements of primary metabolites:
amino acids, fatty acids, and sugars. It is therefore reasonable to
employ these three elements as the starting materials in the total
synthesis of such natural products. We recently reported both
the total synthesis of virgineone aglycone and the stereochemical
assignment of virgineone.10 Our synthetic methodology featured
fragment coupling via olefin cross-metathesis. This methodology
enabled us to synthesize various 3-acyltetramic acid derivatives
simply by changing the starting amino acids or fatty acids. In the
present study, we focused on the total synthesis of epicoccamides
through the application of our established methodology. This
Letter describes the results of our synthetic studies of a model
compound and epicoccamides A and D and the NMR analysis
results, which confirmed Schobert’s stereochemical assignment of
⇑
Corresponding author. Tel./fax: +81 3 5477 2264.
E-mail address: ayaji@nodai.ac.jp (A. Yajima).
http://dx.doi.org/10.1016/j.tetlet.2014.06.040
0040-4039/Ó 2014 Elsevier Ltd. All rights reserved.

A. Yajima et al. / Tetrahedron Letters 55 (2014) 4350–4354
4351
Figure 1. Structures of epicoccamides and related naturally occurring 3-acyltetramic acids.
epicoccamide D and allowed the assignment of the absolute con-
figuration of the natural epicoccamide A.
decomposition of the desired product was observed when more
than 2.5 equiv of a base, such as NaHMDS, were employed. The
decomposition was suppressed to some extent when less than
2.2 equiv of NaHMDS were employed, although this reaction was
unreproducible. The optimized conditions were the same as those
reported by Yoda and co-workers for their total synthesis of peni-
cillenols.13 Although Yoda did not observe any detectable epimer-
ization of their N-methylation substrate, our substrate (13) was
prone to epimerization at C-20 under basic conditions, leading to
the isolation of 14 as an inseparable diastereomeric mixture. The
bulky side chain at C-5 may play a role in hindering deprotonation
at C-20 in Yoda’s synthesis. Unfortunately, we could not determine
the precise diastereomeric ratio of the product by 1H NMR analysis,
although 13C NMR allowed a rough estimation of a 10:1ꢀ4:1 ratio.
Next, the partially epimerized 14 was coupled with the readily
available 5-hexen-1-ol by olefin cross-metathesis using the Grubbs
second-generation catalyst,14 and the resulting double bond was
hydrogenated over catalytic Pd/C to give the model compound
(5S,20S)-15. Similarly, (5S,20RS)-15 was prepared from ( )-11. Com-
Scheme 1 shows our retrosynthetic analysis of the targeted
epicoccamides. In Schobert’s synthesis of epicoccamide D, the
researchers employed a stereo-inversion at C-2 of the b-glucoside,
asymmetric hydrogenation, and late-stage Lacy-Dieckmann
condensation to construct the b-mannoside, the C-20 stereogenic
center, and the tetramic acid portions of the compound,
respectively.8 In contrast, we employed b-selective mannosylation
for the synthesis of segment A, Evans asymmetric alkylation for the
synthesis of segment B, and tandem O-acylation–migration for the
construction of the 3-acyltetramic acid portion. Based on this con-
vergent approach, various epicoccamide stereoisomers and deriva-
tives could be synthesized by employing the appropriate segments.
Scheme 2 shows our synthesis of model compound 15, which
does not include the mannose moiety. Because Wright et al.
reported the NMR data of the aglycone of epicoccamide A, which
was prepared by the hydrolysis of the natural product, we assumed
that a comparison of the NMR spectra of 15 with the reported spec-
tra would enable us to assign the absolute configuration of the nat-
ural product. First, the optically active carboxylic acid segment B
was prepared from commercially available 7 via the Evans asym-
metric alkylation of 9.11 The diastereomeric purity of 10 was esti-
mated to be better than 98:2 by 1H NMR analysis. Removal of the
chiral auxiliary gave (S)-11 (=B). Tandem O-acylation–migration
between 11 and the known compound 12,12 followed by the
deprotection of the Boc group, afforded 13 without epimerization
of the two stereogenic centers. In the next N-methylation step,
13
paring the 1H and C NMR spectra of the synthetic (20S)- and
(20RS)-15, we concluded that the natural epicoccamide D is not a
diastereoisomeric mixture and that Hertweck’s suggestion is incor-
1
rect. The H NMR spectra of (20RS)-15 recorded in CD3OD or CDCl3
showed two pairs of doublets corresponding to the methyl group
at C-20, whereas only the doublet of the major isomer was observed
1
in the H NMR spectrum of (20S)-15. However, it was difficult to
determine the relative configurations of the natural epicoccamides
by 1H NMR due to the relatively small differences between the
Scheme 1. Retrosynthetic analysis of epicoccamides.

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A. Yajima et al. / Tetrahedron Letters 55 (2014) 4350–4354
Scheme 2. Synthesis of model compounds. Reagents, conditions, and yields: (a) (i) PivCl, LiCl, Et3N, THF, ꢁ20 °C; (ii) nBuLi, (S)-8, THF, ꢁ78 °C then mixed anhydride, 94%; (b)
NaHMDS, MeI, THF, ꢁ78 °C, 96%, ds >98:2; (c) LiOH, H2O2, THF, H2O, 74%; (d) DCC, 12, DMAP, CH2Cl2; (e) TFA, CH2Cl2, 77% in two steps; (f) NaHMDS, MeI, THF, ꢁ78 °C to
ꢁ40 °C, 83%; (g) Grubbs second-generation catalyst, 5-hexene-1-ol, CH2Cl2, reflux; (h) H2, Pd/C, MeOH, 28% in two steps.
diastereomers. In contrast, the 13C NMR spectra clearly showed
significant differences between the two diastereomers when the
spectra were recorded in CDCl3. Fortunately, Hertweck reported
the 13C NMR spectra of epicoccamides B–D recorded in CDCl3.4
Because the spectrum of the synthetic (5S,20S)-15 was in good
agreement with the reported data, we assigned the absolute con-
figurations of epicoccamides B–D as 5S,20S, in agreement with
Schobert’s work. Because Wright and co-workers only reported
NMR data recorded in CD3OD, it was impossible to assign the
the synthesis of 15, it could be avoided by stopping the reaction
after a relatively short time. Because it was unlikely that the sugar
moiety of 20 decomposed under such conditions, the decomposi-
tion likely occurred at the 3-acyltetramic acid moiety. To overcome
these issues, we attempted the protection of the 3-acyltetramic
acid moiety as a cyclic borate according to the methods reported
by Loscher and Schobert.8 As the model compound, 14 was treated
with BF3 etherate to afford 21, and the cross-metathesis of 14 and
21 gave the coupled product 22 in moderate yield. Unfortunately,
the hydrogenation and concomitant removal of the protecting
groups were unsuccessful and gave only a complex mixture under
the reported conditions (H2, Pd/C, MeOH).
Jones and co-workers reported that a 3-acyltetramic acid deriv-
ative is relatively unstable under hydrogenation conditions (H2, Pd/
C, EtOH).17 However, in our synthesis of virgineone aglycone,
decomposition of the product was not observed (H2, Pd(OH)2/C,
TFA, t-BuOH).10 The instability of the 3-acyltetramic acid deriva-
tives under hydrogenation conditions may depend on the steric
hindrance of the substituents at C-5. In an effort to avoid decompo-
sition, we studied the effects of a number of hydrogenation condi-
tions on the tetramic acid segments 14 and 21 (Table 1). Although
we could not find the ideal, decomposition-free conditions for the
hydrogenation of 14, we found that Pearlman’s catalyst in ethyl
acetate was effective for the hydrogenation of 14 with only slight
decomposition of the product 23 (entry 4). The hydrogenation of
21 resulted in decomposition under hydrogenation conditions
(entry 5). Although the cyclic borate ester moiety of 21 was readily
hydrolyzed in MeOH, the terminal olefin was isomerized to give
the internal olefin mixtures 24 (entry 6).
absolute configuration of epicoccamide
A at this stage. We
assumed, however, that it should be the same as the configurations
of epicoccamides B–D because these compounds are derived from
a common biogenetic origin.
We then synthesized epicoccamides A and D with the 5S,20S
configuration, as shown in Schemes 3 and 4. First, the glycosyl
donor 18 was prepared from the known compound 1615 by protec-
tion of the 2- and 3-hydroxy groups as NAP (2-naphtylmethyl) and
benzyl ethers. The b-mannosides were prepared according to a
slightly modified Crich’s condition16 followed by deprotection of
the NAP group with DDQ to give 19a and 19b as mixtures of insep-
arable anomeric isomers (ꢀ10:1). By switching the chain lengths of
the glycosyl acceptors, the segments for the synthesis of epicocca-
mides A and D were easily prepared. The couplings of 19a or 19b
with the 3-acyltetramic acid segment 14 were successfully carried
out by olefin cross-metathesis using Grubbs second-generation
catalyst, yielding 20a and 20b. Unfortunately, the final remaining
tasks, which comprised the deprotection of the benzylidene acetal
and benzyl groups, were troublesome. Under catalytic hydrogena-
tion conditions, with or without TFA, the reactions gave complex
mixtures. Prior to deprotection of the benzylidene acetal and
benzyl groups of 20, decomposition of the starting material and
product was indicated by a TLC analysis of the reaction mixture.
Although decomposition was noted in the hydrogenation step in
Employing the established procedure, we sought to finalize the
total synthesis of epicoccamides (Scheme 4). The benzylidene
acetal groups of the cross-coupling products 20a and 20b
were hydrolyzed with TFA, and this step was followed by the

A. Yajima et al. / Tetrahedron Letters 55 (2014) 4350–4354
4353
Scheme 3. b-Selective mannosylation and olefin cross-metathesis. Reagents, conditions, and yields: (a) NaOH aq., NAPBr, Bu4NHSO4, CH2Cl2, reflux, 57%; (b) NaH, BnBr, DMF,
0 °C to rt, 95%; (c) 1-benzenesulfinyl piperidine (BSP), 2,4,6-tri-t-butylpyrimidine (TTBP), Tf2O, MS3A, 1-hexene, 7-octen-1-ol or 9-decen-1-ol, CH2Cl2, ꢁ78 °C to rt; (d) DDQ,
CH2Cl2, 0 °C to rt; 86% in two steps for 19a or 51% in two steps for 19b; (e) 14, Grubbs second-generation catalyst, toluene, reflux; (f) BF3OEt2, CH2Cl2, reflux, 75%; (g) 19b,
Grubbs second-generation catalyst, toluene, reflux, 47%.
Scheme 4. Completion of the total synthesis of epicoccamides A and D. Reagents, conditions, and yields: (a) TFA, CH2Cl2, H2O; (b) Pd(OH)2/C, H2, EtOAc, 20% in three steps for
1 or 16% in three steps for 4.
Table 1
time resulted in decomposition of the products under the reaction
conditions.
Investigation of the hydrogenation reaction with model compounds
We then compared the 1H and 13C NMR spectra of the synthetic
compounds with the reported spectra of epicoccamides A and D. As
described above, the absolute configuration of the natural epicoc-
camide D was determined by Loscher and Schobert as 5S,20S based
on the specific rotation values and partial 13C NMR data recorded
in CDCl3.8 We confirmed this assignment with the aid of the 13C
NMR spectra of 15 and the synthetic epicoccamide D and found
that the 1H and 13C NMR spectra of the synthetic 4 recorded in
CDCl3 were identical with those of the natural product. Loscher
and Schobert reported the NMR spectra of their synthetic com-
pound recorded in CD3OD. However, it was very difficult to verify
the consistency of our NMR spectra with their spectra because the
use of CD3OD as the solvent in our study led to slight variations in
the data across a number of trials. In fact, clear differences were
immediately apparent between our 1H NMR spectrum and the
Entry
Substrate
Conditions
Result (yield%)
1
2
3
4
5
6
14
14
14
14
21
21
H2, Pd/C, MeOH
H2, Pd/C, EtOAc
Ar, Pd/C, EtOAc
H2, Pd(OH)2/C, EtOAc
H2, Pd/C, MeOH
Ar, MeOH
Decomposition
Decomposition
No reaction
23 (quant.)
Decomposition
24 (quant.)
concomitant hydrogenolysis of a benzyl group and hydrogenation
of a double bond with Pearlman’s catalyst in ethyl acetate to give
epicoccamides A and D in 15–20% isolated yields through the three
transformations from 19a and 19b. Again, a prolonged reaction
reported spectrum: we observed
a Me–N singlet signal at
2.95 ppm, whereas the reported spectrum showed small doublet
signals at approximately 2.8 ppm. This inconsistency was noted
by Loscher and Schobert.18 Wright and co-workers reported the

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A. Yajima et al. / Tetrahedron Letters 55 (2014) 4350–4354
NMR spectroscopic data of natural epicoccamide A recorded in
CD3OD. The 1H spectrum of our synthetic epicoccamide A showed
relatively a good agreement with the reported data and with a
spectrum kindly provided by Professor König.3 However, slight dif-
ferences between the synthetic and natural epicoccamide A were
observed in their 13C NMR spectra. Broad signals may be expected
for the tetramic acid portion of the compound due to the complex
tautomerism observed in the NMR spectrum of 3-acyltetramic acid
derivatives.19 The differences observed in this study may be caused
by a complex range of variables, such as pH, concentration, and
temperature of the samples. We also measured the 1H and 13C
NMR spectra of epicoccamide A in DMSO-d6 and found that these
spectra were relatively suitable for stereochemical assignment
because they did not exhibit substantial pH or concentration
dependency. Thus, we propose DMSO-d6 as a good choice for the
measurement of the NMR spectra of 3-acyltetramic acid deriva-
tives. Our synthetic epicoccamide D exhibited relatively similar
D. Structure–activity relationship studies of the epicoccamide
derivatives are currently underway.
Acknowledgments
We thank Professor G. M. König and Dr. S. Kehraus (University
of Bonn) for providing copies of the NMR spectra of epicoccamide
A. We also thank Professor R. Schobert and Dr. S. Loscher for the
useful information regarding the NMR spectra of epicoccamide D.
Supplementary data
Supplementary data (experimental procedures and 1H and 13C
NMR and CD spectra of the synthetic compounds) associated with
this article can be found, in the online version, at http://dx.doi.org/
10.1016/j.tetlet.2014.06.040.
[
[[
[[
a
]
a
a
values [[
a
]
30 = ꢁ54.2 (c 0.20, MeOH)] as the reported natural
D
D
References and notes
]
]
25 = ꢁ40.4 (c 0.20, MeOH)]4 and synthetic epicoccamide D
D
24 = ꢁ39 (c 0.20, MeOH)].8 Although our synthetic epicocca-
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D
mide A exhibited considerably smaller [
(c 0.10, EtOH)] than the natural epicoccamide A [[
a]
values [[
a]
25 = ꢁ40.6
D
D
a
]
D
20 = ꢁ10.3 (c
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and those of the natural epicoccamide A matched. Thus, we con-
cluded that the absolute configuration of the natural epicoccamide
A is 5S,20S. The clear difference of the [
a]D values between the syn-
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pH of the samples.
In summary, we achieved the total synthesis of epicoccamides A
and D using olefin cross-metathesis as the key segment-coupling
reaction. This Letter constitutes the first example of the total syn-
thesis of epicoccamide A. Although the stereochemical purities of
the synthesized compounds were not optimal, our established
methodology could be applied to the synthesis of related 3-acyltet-
ramic acid natural products, such as virgineones and ancorino-
sides. Although the stereoisomers generated by N-methylation
and b-selective mannosylation reactions may be eliminated
through HPLC or related chromatographic techniques, the develop-
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we concluded that the absolute configuration of the natural
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epicoccamide
A
is 5S,20S, identical to that of epicoccamide