The first synthesis and antifungal activities of 9-methoxystrobilurin-type β-substituted β-Methoxyacrylate

Article abstract of DOI:10.1016/S0960-894X(02)00625-X

The first synthesis of 9-methoxystrobilurin-type β-substituted MOAs was successfully achieved. A chiral oudemansin-type β-substituted MOA was also synthesized utilizing Mukaiyama's asymmetric aldol reaction. Antifungal activities of the synthesized compounds against several representative fungi were examined by disk-diffusion assay. As a result, unique and superior antifungal properties of 9-methoxystrobilurin-type β-substituted MOAs compared with those of oudemansin-type analogue were clearly revealed.

Full text of DOI:10.1016/S0960-894X(02)00625-X

Bioorganic & Medicinal Chemistry Letters 12 (2002) 2821–2824
The First Synthesis and Antifungal Activities of
9-Methoxystrobilurin-type ꢀ-Substituted ꢀ-Methoxyacrylate
Hiromi Uchiro, Koh Nagasawa, Tomoya Kotake, Daiju Hasegawa, Aya Tomita
and Susumu Kobayashi*
Faculty of Pharmaceutical Sciences, Tokyo University of Science, 12 Ichigayafunagawara-machi, Shinjuku-ku, Tokyo 162-0826, Japan
Received 3June 2002; accepted 23July 2002
Abstract—The first synthesis of 9-methoxystrobilurin-type b-substituted MOAs was successfully achieved. A chiral oudemansin-
type b-substituted MOA was also synthesized utilizing Mukaiyama’s asymmetric aldol reaction. Antifungal activities of the syn-
thesized compounds against several representative fungi were examined by disk-diffusion assay. As a result, unique and superior
antifungal properties of 9-methoxystrobilurin-type b-substituted MOAs compared with those of oudemansin-type analogue were
clearly revealed.
# 2002 Elsevier Science Ltd. All rights reserved.
Introduction
b-Methoxyacrylate antibiotics (MOAs) represented by
strobilurins and oudemansins are highly potent anti-
fungal compounds and have, therefore, been applied to
agricultural disinfectants in many countries.1 In addi-
tion, novel 9-methoxystrobilurin-type analogues were
recently isolated as strong growth inhibitors toward
human-derived tumor cell lines.2À4 Pharmaceutical
application studies of these new types of MOAs in the
antifungal, antitumor, and antimalarial fields based on
their SAR studies are currently investigated.5À7 These
known type MOAs have an a-substituted b-methoxy-
acrylate moiety as a common pharmacophoric sub-
structure. On the other hand, several b-substituted-type
b-methoxyacrylates such as cystothiazoles8 and meli-
thiazoles9 were recently isolated from nature (Fig. 1). Both
of these new b-substituted MOAs include oudemansin-
type syn-9-methoxy-10-methyl substructures at their 9-10-
position; however, this saturated linkage seems not ideal
for their antifungal properties. Previous SAR studies
apparently indicated that the potent antifungal activities
were usually observed in the derivatives having strobilurin-
type or aromatic-type unsaturated linkage at the 9-10-
position.1,10 From this point of view, Anke and Steglich
proposed the importance of orthogonal arrangement of
the pharmacophoric MOA moiety and the molecular
plane extending from the aromatic ring to 9-10 unsatu-
rated linkage for the antifungal activity of a-substituted
Figure 1. Two types of naturally-occurred b-methoxyacrylates
(MOAs).
MOAs.11 In this paper, we would like to describe the
first synthesis of 9-methoxystrobilurin-type b-substituted
b-methoxy acrylates which have not yet been discovered
from nature. In addition, we also would like to reveal the
pharmacological superiority of the 9-methoxy strobilurin-
type b-substituted MOAs in their antifungal activities
over the oudemansin-type analogue.
Synthetic Strategy
Our synthetic strategy for 9-methoxystrobilurin-type
b-substituted b-methoxyacrylates is shown in Scheme 1.
An efficient synthetic route based on the aldol reaction of
a, b-unsaturated aldehydes 2 with a dianion generated
from 3-ketovaleric acid methyl ester 3 was designed.
The central methyl enol ether moiety on the target
*Corresponding author. Tel. +81-3-3260-8848; fax: +81-3-3260-
8848; e-mail: kobayash@ps.kagu.sut.ac.jp
0960-894X/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(02)00625-X

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H. Uchiro et al. / Bioorg. Med. Chem. Lett. 12 (2002) 2821–2824
use of 2 equivalents of potassium tert-butoxide and
dimethyl sulfate in dimethylformamide, and the desired
intermediate 5a was successfully obtained. The remaining
secondary hydroxyl group of 5a was then oxidized by
Dess–Martin periodinane to give the corresponding
enone 6a (55% yield for two steps). Finally, the desired
9-methoxystrobilurin-type b-substituted MOA 7a14 was
obtained in 59% yield by the addition of 2 equivalents
of potassium hexamethyldisilazide (KHMDS) and
methyl triflate in THF–HMPA (4:1) at À78 ꢀC. Several
analogues modified on the aromatic moiety 7b–e14 were
also synthesized from the corresponding a,b-unsaturated
aldehydes 2b–e. The stereochemistries of tri- and tetra-
substituted olefin moieties of 7a–e were respectively
determined by NOE measurements as shown in Scheme 2.
Scheme 1. Retrosynthesis of 9-methoxystrobilurin-type b-substituted
b-methoxyacrylates.
An optically active oudemansin-type b-substituted MOA
(+)-8 was synthesized from chiral aldol adduct 9 which
was prepared by Mukaiyama’s asymmetric aldol reac-
tion12 in 98% ee as shown in Scheme 3. The hydroxyl
group of 9 was methylated by diazomethane-boron tri-
fluoride etherate to give the intermediate ether 10. The
thiolester moiety of 10 was reduced by DIBAL to the
corresponding aldehyde 11. The second aldol reaction of
the aldehyde 11 with the lithium enolate of methyl acetate
was performed, and the resulting hydroxyl group of the
aldol adduct 12 was oxidized to give the corresponding
b-ketoester 13 (the structure is not shown). The desired
optically active b-substituted MOA (+)-814 was pre-
pared by O-methylation of a sodium enolate generated
from b-ketoester 13. The optical purity of (+)-8 was
molecule 7 could be easily constructed on enone-type
intermediate 6. The intermediate 6 would be prepared
by the direct methyl enol ether formation starting from
the aldol adduct 4 and successive oxidation of the
remaining secondary hydroxyl group of intermediate 5.
Synthesis
The aldol reaction of cinnamaldehyde 2a with a dianion
generated from methyl 3-ketovalerate 3 was first carried
out. The desired g-adduct 4a was obtained in 86% yield,
and no undesirable a-adduct was formed. The direct methyl
enol ether formation on the adduct 4a was performed by
Scheme 2. Synthesis of 9-methoxystrobilurin-type b-substituted MOAs. (i) NaH, nBuLi, THF, 0 ꢀC, 30 min; (ii) tBuOK, Me2SO4, DMF, 0 ꢀC to rt,
1.5 h; (iii) Dess–Martin periodinane, CH2Cl2, rt, 30 min; (iv) KHMDS, MeOTf, THF–HMPA (4:1), À78 ꢀC, 10 min.
Scheme 3. Synthesis of chiral oudemansin-type b-subsituted MOAs. (i) Sn(OTf)2,
,
nBu2Sn(OAc)2, CH2Cl2, À78 ꢀC, 3h; (ii) CH 2N2,
ꢀ
ꢀ
ꢀ
.
BF3 OEt2, Et2O, 0 C to rt, 1 h; (iii) DIBAL, CH2Cl2, À78 C, 30 min; (iv) AcOMe with LDA, THF, À78 C, 1 h; (v) Dess–Martin periodinane,
CH2Cl2, rt, 1 h; (vi) tBuOK, Me2SO4, DMF, 0 ꢀC, 3h.

H. Uchiro et al. / Bioorg. Med. Chem. Lett. 12 (2002) 2821–2824
2823
Scheme 4. Synthesis of racemic oudemansin-type and epi-oudemansin-type-b-substituted MOAs. (i) BuOK, Me2SO4, DMF, 0 ꢀC, 3h; (ii) CH N2,
t
2
ꢀ
.
BF3 OEt2, Et2O, 0 C to rt, 3h.
Table 1. Antifungal activities of b-substituted b-methoxyacrylates
transformed to the desired b-methoxyacrylates (Æ)-8 and
(Æ)-1414 via methyl enol ether formation and sequential
9-O-methylation.
Diameter of inhibition zone (mm)a,b
Compd
Conc
(mg/disk)
Pc
Af
Fs
Ca
Sc
Nystatin
(positive control)
10
10
16
—
16
18
Antifungal Activity
Antifungal activities of the synthesized compounds
toward several pathogenic or non-pathogenic fungi were
examined by disk-diffusion assay (Table 1). 9-Methoxy-
strobilurin-type b-substituted MOAs bearing unsub-
stituted benzene ring 7a exhibited strong antifungal
activity similar to that of 9-methoxystrobirulin A (1)
which is the corresponding a-substituted MOA. In addi-
tion, 7a also showed fungicide activity against drug-resis-
tant Fusarium solani although it was still in an incomplete
manner.13 The optically active and racemic oudemansin-
type analogues (+)-8 and (Æ)-8 were apparently less
effective than 7a, and no significant effect between
molecular chirality and their antifungal property was
observed. The racemic epi-oudemansin-type analogue
was almost ineffective to all fungi tested. On the other
hand, aromatic-modified compounds 7b–d were inferior
to 7a except for a thiophene analogue 7e. These results
are in sharp contrast to those of previously reported 9-
methoxystrobilurin-type a-substituted MOAs.6
1
10
1
0.1
33
2328
19
38
16
—
—
42i
i
Æ
32i
27i
—
44i
39i
13i
— 2
3
7a
10
1
0.1
42
17
—
36
17
—
30i
22i
—
46i
30i
—
7b
10
1
0.1
33
18
—
12
8
—
—
—
—
Æ
—
—
18i
12i
—
7c
10
1
0.1
—
—
—
—
—
—
—
—
—
—
—
—
Æ
—
—
7d
10
1
0.1
1316
—
—
27
10
—
i
19i
—
14i
8i
—
21i
15i
—
7e
10
1
0.1
40
21
Æ
29
20
—
—
—
—
32i
16i
—
49i
31i
—
(+)-8
(Æ)-8
(Æ)-14
10
1
0.1
28
Æ
23—
—
—
20
—
—
i
—
—
43i
25i
—
—
10
1
0.1
26
Æ
23—
—
—
14
—
—
i
—
—
39i
Æ
Conclusion
—
—
In conclusion, the first synthesis of 9-methoxystrobilurin-
type b-substituted MOAs was successfully achieved and
their unique and superior antifungal properties were also
revealed. Further extensive SAR studies of 9-methoxy-
strobilurin-type b-substituted MOAs to develop a more
effective antifungal compound, and synthetic studies of
9-methoxystrobilurin-type analogues of natural b-sub-
stituted MOAs are now in progress.
10
1
0.1
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
Pc, Penicilium ctrinum R-3703; Af, Aspergillus fumigatus R-1301;
Fs, Fusarium solani R-2800; Ca, Candida albicans IFO 1594;
Sc, Saccharomyces cerevisae IAM 4861.
aThe diameter of each inhibition zones (mean value of two samples)
were measured after 48 hr incubation.
b—, not effective, Æ, slightly effective, i, incomplete inhibition.
References and Notes
determined to be 97%ee by chiral HPLC analysis (col-
umn: Daicel Chiralcel OD; eluent: hexane/2-propa-
nol=50/1).
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Racemic oudemansin-type and epi-oudemansin-type
b-substituted MOAs (Æ)-8 and (Æ)-14 were also synthe-
sized as shown in Scheme 4. The diastereomers of racemic
aldol adduct syn-4a and anti-4a were separated by column
chromatography, and each diastereomer was respectively
4. Nicholas, G. M.; Blunt, J. W.; Cole, A. L. J.; Munro,
M. H. G. Tetrahedron Lett. 1997, 38, 7465.

2824
H. Uchiro et al. / Bioorg. Med. Chem. Lett. 12 (2002) 2821–2824
5. (a) Uchiro, H.; Nagasawa, K.; Aiba, Y.; Kobayashi, S.
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zones became gradually obscure and were lost after 96 h.
14. Physical data of synthesized compounds: 7a: 1H NMR
(CDCl3, 500 MHz) d 1.99 (s, 3H), 3.63 (s, 3H), 3.73 (s, 3H),
3.74 (s, 3H), 5.22 (s, 1H), 6.53 (d, 1H, J=15.9 Hz), 6.78 (d,
1H, J=15.9 Hz), 7.22 (t, 1H, J=7.3Hz), 7.30 (dd, 2H, J=7.3,
7.9 Hz), 7.37(d, 1H, J=7.9 Hz); EI–MS 288 (M+); 7b: 1H
NMR (CDCl3, 300 MHz) d 1.98 (s, 3H), 3.63 (s, 3H), 3.73 (s,
3H), 3.73 (s, 3H), 3.88 (s, 3H), 3.89 (s, 3H), 5.21 (s, 1H), 6.39
(d, 1H, J=15.8 Hz), 6.73(d, 1H, J=15.8 Hz), 6.81 (d, 1H,
J=8.3Hz), 6.88 (dd, 1H, J=1.8, 8.3Hz; EI–MS 348 (M +);
6. Uchiro, H.; Nagasawa, K.; Sawa, T.; Hasegawa, D.;
Kotake, T.; Sugiura, Y.; Kobayashi, S.; Otoguro, K.; Omura,
S. Bioorg. Med. Chem. Lett. In press.
-
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gami, Y.; Fudou, R.; Yoshimura, T.; Yamanaka, S. J. Anti-
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Akita, H. Tetrahedron Lett. 2002, 43, 643. (b) Williams, D. R.;
Patnaik, S.; Clark, M. P. J. Org. Chem. 2001, 66, 8463. (c)
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3184.
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B.; Forche, E.; Steinmetz, H.; Hofle, G.; Reichenbach, H. J.
Antibiot. 1999, 52, 721. (b) Bohlendorf, B.; Herrmann, M.; Hecht,
H.-J.; Sasse, F.; Forche, E.; Kunze, B.; Reichenbach, H.; Hofle,
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1
7c: H NMR (CDCl3, 300 MHz) d 2.03 (s, 3H), 3.65 (s, 3H),
3.72 (s, 3H), 3.84 (s, 3H), 5.22 (s, 1H), 6.59 (d, 1H, J=15.9
Hz), 7.52 (d, 1H, J=15.9 Hz), 7.34–8.17 (m, 7H); EI–MS 338
(M+); 7d: 1H NMR (CDCl3, 300 MHz) d 2.01 (s, 3H), 3.63 (s,
3H), 3.76 (s, 3H), 3.77 (s, 3H), 5.25 (s, 2H), 6.65 (d, 1H,
J=15.8 Hz), 6.95 (d, 1H, J=15.9 Hz), 7.36–7.81 (m, 7H); EI–
MS 338 (M+); 7e: 1H NMR (CDCl3, 500 MHz) d 1.97 (s, 3H),
3.63 (s, 3H), 3.72 (s, 3H), 3.73 (s, 3H), 5.21 (s, 1H), 6.34 (d,
1H, J=15.3Hz), 6.89 (d, 1H, J=15.3Hz), 6.96 (dd, 1H,
J=3.1, 4.9 Hz), 6.99 (d, 1H, J=3.1 Hz), 7.16 (d, 1H, J=4.9
1
Hz); EI–MS 294 (M+); (+)-8: H NMR (CDCl3, 500 MHz) d
1.21 (d, 3H, J=7.0 Hz), 3.31 (s, 3H), 3.57 (s, 3H), 3.66 (s, 3H),
3.74 (dd, 1H, J=8.2, 8.2 Hz), 4.20 (dq, 1H, J=7.0, 8.2 Hz),
4.94 (s, 1H), 6.07 (dd, 1H, J=8.2, 15.9 Hz), 6.47 (d, 1H, J=15.9
Hz), 7.23(t, 1H, J=7.2 Hz), 7.30 (dd, 1H, J=7.2, 7.3Hz), 7.34
(d, 1H, J=7.3Hz); EI–MS 290 (M +); [a]D26=+239 (c 1.0,
1
CHCl3); (Æ)-14: H NMR (CDCl3, 300 MHz) d 1.04 (d, 3H,
J=7.0 Hz), 3.26 (s, 3H), 3.66 (s, 3H), 3.68 (s, 3H), 3.83 (dd, 1H,
J=8.5, 9.2 Hz), 4.21 (dq, 1H, J=7.0, 9.2 Hz), 5.08 (s, 1H), 6.05
(dd, 1H, J=8.5, 15.9 Hz), 6.56 (d, 1H, J=15.9 Hz), 6.88 (d,
1H, J=15.8 Hz), 7.25 (t, 1H, J=7.3Hz), 7.33(dd, 1H, J=7.3,
7.3Hz), 7.40 (d, 1H, J=7.3Hz); EI–MS 290 (M +).
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Grabley, S., Thiericke, R., Eds., Springer: Berlin, 1999; Chap-
ter 18, p 320.