Assignment of the absolute configuration of blasticidin A and revision of that of aflastatin A

Article abstract of DOI:10.1016/j.tetlet.2007.02.024

The absolute configuration of blasticidin A, a strong inhibitor of aflatoxin production by Aspergillus parasiticus, was assigned by adding the data of relative configurations at its diol and pentaol moieties to previously known stereochemistry. Similarity

Full text of DOI:10.1016/j.tetlet.2007.02.024

Tetrahedron Letters 48 (2007) 2527–2531
Assignment of the absolute configuration of blasticidin A
and revision of that of aflastatin A
Shohei Sakuda,a,* Nobuaki Matsumori,b Kazuo Furihataa and Hiromichi Nagasawaa
aDepartment of Applied Biological Chemistry, The University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan
bDepartment of Chemistry, Osaka University, 1-16 Machikaneyama, Toyonaka, Osaka 560-0043, Japan
Received 25 January 2007; revised 2 February 2007; accepted 6 February 2007
Available online 9 February 2007
Abstract—The absolute configuration of blasticidin A, a strong inhibitor of aflatoxin production by Aspergillus parasiticus, was
assigned by adding the data of relative configurations at its diol and pentaol moieties to previously known stereochemistry. Simi-
larity of the NMR data of blasticidin A to those of aflastatin A allowed us to revise the stereochemistry of the diol and pentaol
moieties of aflastatin A.
Ó 2007 Elsevier Ltd. All rights reserved.
1. Introduction
C39 of AsA were determined by analysis of small frag-
ment molecules obtained by degradation experiments
of AsA.9 The absolute stereochemistry of the tetrahyd-
ropyrane ring moiety of AsA was assigned based on
the relative stereochemistry around the ring and the
absolute configuration at C33. The polyol fragment 1
was prepared from AsA and its absolute configuration
was assigned by applying acetonide and MTPA meth-
ods.10 The stereochemistry of 1 was recently confirmed
by its chiral synthesis by Evans et al.11 The long polyol
fragment 3 was also prepared from AsA. The relative
configurations from C4 to C8 and from C23 to C31 of
3 were assigned by the J-based method,12 which led to
assignment of the whole absolute configuration of AsA
(formerly proposed structure in Fig. 1).9 Recently, how-
ever, Kishi and co-workers pointed out that erythro/
threo/threo/threo is the correct relative stereochemistry
at the pentaol moiety (C25–C29) of 3 from their NMR
database study.13 On the other hand, with respect to
the stereochemistry of BcA, the absolute configurations
at C4, C6, C31–C35, C37 and all chiral centers involved
in the polyol fragment 2 were assigned by applying sim-
ilar methods used for the case of AsA.6,10 Therefore,
determination of the configurations at the diol (C8,
C9) and pentaol (C25–C29) moieties of BcA is a remain-
ing problem to complete the assignment of the absolute
stereochemistry of BcA. In this Letter, we report the
complete assignment of the absolute configuration of
BcA, and propose the revised stereochemistry of AsA,
which was deduced from similarity of the NMR data
of AsA with those of BcA.
Aflatoxins, a group of mycotoxins, show quite potent
toxicity and carcinogenicity towards mammals. Their
contamination in agricultural products is a serious prob-
lem from the viewpoint of not only food safety but also
economic loss.1,2 However, it is difficult to resolve the
problem due to lack of an effective method to control
aflatoxin production. We have been studying specific
inhibitors of aflatoxin production by Aspergillus parasit-
icus since they may be useful to prevent aflatoxin con-
tamination of foods and feeds without incurring a
rapid spread of resistant strains. We found aflastatins
A and B (AsA and AsB) and blasticidin A (BcA) from
Streptomyces metabolites as inhibitors of aflatoxin pro-
duction.3–6 They strongly inhibited aflatoxin production
of A. parasiticus by disturbing the primary metabolism
of the fungus, which may regulate a pathway leading
to expression of aflatoxin biosynthetic enzymes.7,8
These compounds have similar unique structures, which
are tetramic acid derivatives with a highly oxygenated
long alkyl chain. With respect to the stereochemistry
of AsA, we have the following information up to now.
The absolute configurations at C50, C4, C6, C33 and
Keywords: Blasticidin A; Aflastatin A; Aflatoxin; Absolute
configuration.
*
Corresponding author. Fax: +81 3 5841 8022; e-mail: asakuda@
mail.ecc.u-tokyo.ac.jp
0040-4039/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2007.02.024

2528
S. Sakuda et al. / Tetrahedron Letters 48 (2007) 2527–2531
OH
HO
OH OH OH OH OH OH
OH
OH
OH
OH OH OH OH OH
O
31
1
4
8
O
25
37
H
OH
OH
N
OH OH
OH
48
O
blasticidin A
HO
OH OH OH OH OH OH OH OH OH OH OH OH
OH
OH
OH
1
O
33
4
8
O
27
39
H
OH
N
OH
OH OH
O
5'
aflastatin A (Revised structure)
OH
HO
OH OH OH OH OH OH OH OH OH OH OH OH
OH
OH
OH
1
O
33
4
8
O
27
39
H
OH
N
OH
OH OH
O
5'
aflastatin A (Formerly proposed structure)
Figure 1. Structures of blasticidin A (BcA) and aflastatin A (AsA).
OH OH OH
OH OH OH OH
OH OH OH OH OH OH OH OH
OH
HO
HO
OH
2
1
OH
HO
OH
OH
OH OH OH OH OH OH OH OH OH OH OH OH
31
2
6
HO
O
25
37
H
OH
OH
OH OH
3
OH
O
HO
OH
OH
OH OH OH OH OH OH
OH OH OH OH OH
29
2
6
HO
23
35
H
OCH
3
OH OH
OH
4
3
2. Absolute configuration of blasticidin A
one of them shows similar JH,H values to those ob-
served in 4. The synthetic procedure of these model com-
pounds is summarized in Scheme 1. We chose 9a, 9b,
12a and 12b as precursors of 5a, 5b, 50a and 50b, respec-
tively. Compounds 9a and 12a were prepared from
To clarify the configuration at the diol and pentaol moi-
eties of BcA, we analyzed the corresponding parts of its
long polyol fragment methyl glycoside 4, which was pre-
pared from methyl glycoside of BcA as described previ-
ously.14 We assigned the absolute configuration at each
moiety as follows.
methyl
4,6-O-benzylidene-a-D-glucopyranoside
6a
according to the known procedure.15 By using the same
procedure as that used in preparation of 9a and 12a, 9b
and 12b could be synthesized from methyl 4,6-O-benzyl-
idene-a-D-galactopyranoside 6b as follows. The reaction
of 6b with 1-tosylimidazole and NaH afforded known
monotosylate 7b,16,17which was easily converted to
epoxide 8b by NaH. Compound 9b was obtained by
the reaction of 8b with methylmagnesium chloride. After
conversion of 9b to 11b, NaBH4 reduction of 11b affor-
ded 12b together with its diastereomer at C2.
2.1. Stereochemistry at the diol moiety of 4
Since the absolute configurations at C4 and C8 of 4 were
confirmed, determination of the relative stereochemistry
either from C4 to C7 or from C6 to C8 was enough for
assignment of the absolute configurations at C6 and
C7 of 4. We first tried to elucidate the relative configu-
3
rations by the J-based method.12 The values of JH,H
2,3
and
J
were obtained from DQFCOSY, HETLOC
Acid hydrolysis of 12a, followed by NaBH4 reduction,
afforded crude 50a, which was once acetylated for purifi-
cation and deacetylation of acetylated 50a afforded pure
50a. When 9a was similarly treated with acid and
NaBH4, 14a was obtained as a minor product after acet-
ylation. By the reaction, 13a was obtained as a major
product due to the much higher population of a 1,6-
anhydro form than that of a hemiacetal one in the acidic
solution during hydrolysis.18 Removal of the acetyl
groups of 14a afforded 5a. Similarly to the case of 9a,
C,H
and PFGHMBC spectra. However, any relative config-
urations involved in the C4–C8 moiety of 4 could not
be assigned due to difficulty encountered in determina-
tion of the conformations around the bonds C4–C5,
C6–C7 and C7–C8 by the obtained J values.
Therefore, we next synthesized four model stereoiso-
mers, 5a, 5b, 50a and 50b, which could cover all possible
stereochemistry for C6–C8 of 4, with expectation that

S. Sakuda et al. / Tetrahedron Letters 48 (2007) 2527–2531
2529
6
O
1
OCH3
OH
O
1'
ph
ph
O
O
O
O
OCH3
OH
OCH3
OTs
OCH3
O
O
O
OH
6a
c
a
b
ph
O
ph
O
ph
O
O
CH3
9a, 9b
OH
7a, 7b
8a, 8b
O
OCH3
OH
O
O
OH
6b
OH OH
1
OH
OH
HO
O
OAc OAc
O
1
OH CH3
5a
1
g
OAc
d, e, f
AcO
+
AcO
OAc
OAc CH3
OH OH
CH3
13a, 13b
14a, 14b
HO
OH CH3
5b
OH OH
1
OH
HO
OH CH3
O
O
O
OCH3
O
OCH3
OH
OCH3
O
O
O
5'a
O
j, e, f, g
e
i
h
9a, 9b
ph
O
ph
O
OH OH
ph
O
CH3
11a, 11b
CH3
OH
CH3
10a, 10b
HO
12a, 12b
OH CH3
5'b
Scheme 1. Reagents and conditions: (a) 1-tosylimidazole, NaH, DMF; (b) NaH; (c) CH3MgCl; (d) 1 M HCl:1,4-dioxane (1:1), 100 °C, 5 h; (e)
NaBH4; (f) (CH3CO)2O, pyridine; (g) MeONa; (h) (CF3CO)2O, DMSO; (i) Et3N; (j) 1 N H2SO4, 110 °C, 1.5 h.
3
Table 1. JH,H Values in 5a, 50a, 5b, 50b and 4a
A
C
C
C
21
22
24
Compound
3JH-2,H-3
3JH-3,H-4
3JH-4,H-5
C
C
20
21
23
HO
22b
3
H
23
5a
50a
5b
50b
4
7.5
8.0
5.0
b
3.7
2.4
4.3
8.0
4.2d
2.3
4.0
4.3
1.8
e
H
H
22b
22a
G
H
G
H
22a
H
OH
G
21
4.2c
C
A
A
a Spectra were obtained in pyridine-d5. J Values were confirmed by
conventional decoupling experiments.
b Cannot be determined due to signal overlappings.
2
J
= 3.0 Hz
H22a,C24
J
= -2.3 Hz
H22a,C21
c 3
A
J
.
A
H-6,H-7
C
C
C
25
C
d 3
24
J
.
H-7,H-8
e Not determined.
HO
H
27
26
HO
HO
H
26
25
G
26
28
C
25
27
G
acid hydrolysis of 9b followed by NaBH4 reduction and
acetylation afforded 13b and 14b as major and minor
products, respectively. The stereochemistry of 14b was
unambiguously confirmed by that of 13b. Compound
50b was obtained from 12b as a major product.
HO
H
H
A
C
A
2
J
= 0 Hz
= -1 Hz
2
H26,C27
H27,C26
J
J
= 0 Hz
= -1 Hz
H25,C26
H26,C25
2
J
2
3
3
1
Figure 2. Dominant conformations for C21–C22, C22–C23, C25–C26
and C26–C27 of 4 deduced from the J-based method. 3JH,H Values are
listed in Table 2. A, anti. G, gauche. The typical J values used for the
method in general are 9–12 Hz (3JH,H), 6–8 Hz (3JC,H) and 0 to ꢀ2 Hz
(2JC,H) for anti relationship, 7–10 Hz (3JH,H), 5–7 Hz (3JC,H) and 0–
+2 Hz (2JC,H) for anti relationship in the case of diol system, 2–4 Hz
(3JH,H), 1–3 Hz (3JC,H) and ꢀ5 to ꢀ7 Hz (2JC,H) for gauche relation-
ship, and 0–3 Hz (3JH,H), 1–3 Hz (3JC,H) and ꢀ4 to ꢀ6 Hz (2JC,H) for
gauche relationship in the case of diol system.12
The JH-2,H-3 and JH-3,H-4 values observed in the H
NMR spectra of 5a, 5b, 50a and 50b are listed in Table
1. Among them, only the values observed in 5b
(3JH-2,H-3 = 5.0 Hz, 3JH-3,H-4 = 4.3 Hz) were comparable
to those in the C6–C8 moiety of 4 (3JH-6,H-7 = 3JH-7,H-8
=
4.2 Hz), indicating that the relative configurations for
both C6–C7 and C7–C8 were assigned as threo. Based
on the absolute configuration at C8, the absolute stereo-
chemistry at C6 and C7 of 4 was determined.
C29 of 4 were elucidated by the J-based method. Each
configuration for C21–C23, C25–C26 and C26–C27
was assigned as syn, threo and threo, respectively, from
each dominant conformation of C21–C22, C22–C23,
C25–C26 and C26–C27 deduced from the typical J val-
ues (Fig. 2), but the configurations for C23–C24, C24–
2.2. Stereochemistry at the pentaol moiety of 4
To assign the absolute stereochemistry at the pentaol
moiety based on the absolute configurations at C21
and C29 of 4, the relative configurations from C21 to

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S. Sakuda et al. / Tetrahedron Letters 48 (2007) 2527–2531
Table 2. NMR data for the diol and pentaol moieties of 3 and 4a
Carbon
4
dC
dH
Coupled protons
3JH,H (Hz)
27.9(27.8)b
2.15(2.14)
H-4,H-5a
H-4,H-5b
H-5a,H-6
H-5b,H-6
H-6,H-7
H-7,H-8
H-21,H-22a(H-23,H-24a)
H-21,H-22b(H-23,H-24b)
H-22a,H-23(H-24a,H-25)
H-22b,H-23(H-24b,H-25)
H-23,H-24(H-25,H-26)
H-24,H-25(H-26,H-27)
H-25,H-26(H-27,H-28)
H-26,H-27(H-28,H-29)
H-27,H-28a(H-29,H-30a)
H-27,H-28b(H-29,H-30b)
H-28a,H-29(H-30a,H-31)
H-28b,H-29(H-30a,H-31)
7.3(7.5)
6.0(5.0)
4.4(4.5)
9.2(9.0)
4.2(4.0)
4.2(4.0c)
3.0(3.0)
8.8(8.0)
3.0(3.0)
8.8(8.0)
7.6(7.5)
<2(3.0)
4.4(5.0)
3.4(3.0)
6.5(8.0)
6.5(6.0)
3.5(3.5)
9.2(10.0)
5
42.2(42.3)
1.79(1.84)(Ha)
1.67(1.75)(Hb)
4.26(4.29)
4.02(4.00)
4.63(4.67)
6
7
69.9(69.5)
77.4(77.9)
71.2(71.1)
21(23)
22(24)
42.1(42.1)
2.58(2.59)(Ha)
2.16(2.14)(Hb)
4.68(4.67)
4.33(4.34)
4.97(4.96)
23(25)
24(26)
25(27)
26(28)
27(29)
72.2(72.2)
76.2(76.2)
71.5(71.5)
75.4(75.4)
70.8(70.9)
4.50(4.49)
4.93(4.92)
28(30)
29(31)
37.6(37.5)
72.8(72.8)
3.18(3.17)(Ha)
2.49(2.47)(Hb)
4.21(4.24)
a Spectra were obtained in pyridine-d5.
b The values in parentheses indicate the chemical shifts in 3 cited from Ref. 9.
c This value is corrected from the one (8.0 Hz) reported previously.
C25 and C27–C29 could not be assigned because ambig-
uous medium J values were obtained around the bonds
of C23–C24, C24–C25 and C27–C28. On the other
hand, the NMR database method clearly showed that
the relative configuration from C23 to C27 was
erythro/threo/threo/threo from the profile of the observed
In conclusion, we have assigned the absolute configura-
tion of blasticidin A and revised that of aflastatin A.
They have very similar stereochemistry through the
overall structures.
3
3JH,H values (3JH-23,H-24 = 7.6 Hz, JH-24,H-25 = <2 Hz,
Acknowledgements
3
3JH-25,H-26 = 4.4 Hz, JH-26,H-27 = 3.4 Hz),13 which con-
firmed the configurations for C25–C26 and C26–C27
assigned by the J-based method and determined the
stereochemistry for C23–C24 and C24–C25. This
relative stereochemistry from C21 to C27 afforded the
absolute configuration at the pentaol moiety of 4 based
on the configuration at C21. From the results obtained,
the absolute configuration of BcA was completely
assigned as shown in Figure 1.
We are grateful to Professor Yasuhiro Uozumi and Dr.
Hiroaki Sasagawa of the Institute for Molecular Science
for measurement of 920 MHz NMR spectra and Dr.
Shinji Nagata of the University of Tokyo for measure-
ment of MS spectra.
Supplementary data
Experimental section including spectral data of this arti-
cle. Supplementary data associated with this article can
be found, in the online version, at doi:10.1016/
j.tetlet.2007.02.024.
3. Revision of the absolute configuration at diol and
pentaol moieties of aflastatin A
3
Table 2 summaries d and JH,H values around the diol
and pentaol moieties of 3 and 4. The values observed
in the moiety from C23 to C31 of 3 coincided well with
those in the corresponding C21–C29 moiety in 4, indi-
cating that both moieties should have the same relative
stereochemistry. The NMR data at C4–C7 of 3 were
also very similar to those at C4–C7 of 4, which showed
that 3 and 4 have the same relative configuration from
C4 to C7. From these observations, we propose the
revised absolute stereochemistry of AsA which is
shown in Figure 1.
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