Total synthesis of (+)-blasticidin S

Article abstract of DOI:10.1002/chem.200305358

The first total synthesis of the peptidyl nucleoside antibiotic, blasticidin S (1), has been achieved by the coupling reaction of cytosinine (3) and blastidic acid (2). A key step in the synthesis of cytosinine (3) is the sigma-tropic rearrangement of all

Full text of DOI:10.1002/chem.200305358

FULL PAPER
Total Synthesis of (+)-Blasticidin S
Yoshiyasu Ichikawa,* Keiko Hirata, Masayoshi Ohbayashi, and Minoru Isobe[a]
≈
Dedicated with respect and appreciation to Professor Otake on the occasion of his 77th birthday
Abstract: The first total synthesis of
the peptidyl nucleoside antibiotic, blas-
ticidin S (1), has been achieved by the
coupling reaction of cytosinine (3) and
blastidic acid (2). A key step in the
synthesis of cytosinine (3) is the sigma-
tropic rearrangement of allyl cyanate
24; this reaction provided efficient and
stereoselective access to 2,3-dideoxy-4-
amino-d-hex-2-enopyranose (26a). Fur-
ther elaboration of 26a gave methyl
hex-2-enopyranouronate 29, and cyto-
sine N-glycosylation of 31 using the
Vorbr¸ggen conditions for the silyl Hil-
bert Johnson reaction furnished the
differentially protected cytosinine (32)
in 11 steps from 2-acetoxy-d-glucal
(14) (4.0% overall yield). Synthesis of
the Boc-protected blastidic acid 47 in
nine steps starting from chiral carbox-
ylic acid 35 (23% overall yield) utilized
Weinreb×s protocol for the preparation
of benzyl amide 38 and Fukuyama×s
protocol for the synthesis of the secon-
dary amine 40. Assembly of the pro-
tected cytosinine (32) and blastidic acid
(47) by the BOP method in the pres-
ence of HOBt, and finally elaboration
to 1 by deprotection of the fully pro-
tected 54 established the total synthesis
of blasticidin S (1).
Keywords: amino
acids
¥
blasticidin S ¥ natural products ¥
rearrangement ¥ total synthesis
[8]
≈
Introduction
ies by Otake and co-workers, and has also been confirmed
by X-ray analysis (Scheme 1).[9] Controlled acid hydrolysis
of 1 allowed the isolation of two components, blastidic acid
(2) and cytosinine (3) as their hydrochloride salts
(Scheme 1). Blastidic acid (2) is an unusual b-amino acid
counterpart of arginine containing a modified N-methyl
guanidine group.[10] Such a b-amino acid motif is also mani-
Blasticidin S (1) is a representative peptidyl-nucleoside anti-
biotic,[1] which was first isolated from Streptomyces griseo-
[2]
chromogenes in 1958by Yonehara and co-workers.
This
antibiotic was once commercialized as a fungicide against
the virulent fungus, Piricularia oryzae, which was the cause
of the serious rice blast disease in Asia. Its biological activi-
ty results from specific inhibition of the protein biosynthesis
by interfering with the peptide bond formation in the ribo-
somal machinery.[3] Biosynthetic studies by Gould have ad-
vanced to the molecular level, whereby biosynthetic gene
clusters have been cloned and expressed.[4] Yamaguchi
found two blasticidin S resistance genes which code blastici-
din S deaminase: bsr from Bacillus cereus[5] and BSD from
Aspergillus terreus[6]. Both genes are now widely used for
genetic engineering experiments to select both prokaryotic
and eukaryotic cells that express the blasticidin S resistance
gene. The renaissance of blasticidin S is now flourishing in
the research area of molecular biology.[7]
The structure and absolute configuration of 1 have been
elucidated by chemical degradation and spectroscopic stud-
Scheme 1. Structures of blasticidin S, blastidic acid, and cytosinine.
fested in cytosinine (3). The highly functionalized structure
in 3 is characterized by a unique hexopyranosyl nucleoside
that contains a 2,3-unsaturated-4-amino pyranose attached
to cytosine.[11] The functional group richness found in blasti-
cidin S poses synthetic challenges.
[a] Prof. Dr. Y. Ichikawa, K. Hirata, M. Ohbayashi, Prof. Dr. M. Isobe
Laboratory of Organic Chemistry
School of Bioagricultural Sciences
Nagoya University, Chikusa, Nagoya 464-8601 (Japan)
Fax : (+81)52-789-4111
E-mail: ichikawa@agr.nagoya-u.ac.jp
Although the pioneering work of Kondo and Goto[12] de-
tailed the first synthesis of cytosinine in 1972, and two re-
Supporting information for this article is available on the WWW
under http://www.chemeurj.org/ or from the author.
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DOI: 10.1002/chem.200305358
¹ 2004 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

FULL PAPER
ports for the syntheses of blastidic acid appeared in 2001,[13]
the total synthesis of 1 has not yet been reported. In this
manuscript, we discuss the evolution of our strategy for the
synthesis of cytosinine in full detail, and elaborate the first
total synthesis of blasticidin S.
in Scheme 3. Synthesis of the fully protected cytosinine 8
was envisioned to arise from anomeric activation of 10 fol-
lowed by introduction of the cytosine moiety. Since the acyl
protecting group on the cytosine N4-nitrogen (PG2 in 8) is
not sufficiently robust to carry out a sequence of manipula-
tions,[17] the ester functionality was to be installed prior to
the cytosine N-glycosylation. We further reasoned that unsa-
turated aminopyranoside 11 could be elaborated by [3,3]-
sigmatropic rearrangement of allyl cyanate 12, which itself
would be derived from 2-acetoxy-d-glucal (14). Based on
this synthetic analysis, we launched a synthetic venture
toward cytosinine (3).
Synthetic analysis of cytosinine (3): We envisioned that the
suitably protected cytosinine and blastidic acid intermedi-
ates could be coupled at a late stage in the synthesis. Ac-
cordingly, our initial efforts were devoted to the synthesis of
these two components. Several years ago, we reported a syn-
thetic method for the preparation of the unsaturated amino
sugar, 2,3-dideoxy-4-amino-d-hex-2-enopyranose (7),[14] by
an allyl cyanate-to-isocyanate rearrangement[15] (Scheme 2).
Synthesis of the fully protected cytosinine 32: In our pre-
liminary investigations, we found that acid-catalyzed hydrol-
ysis of a 2,3-unsaturated 4-aminopynanoside such as 15 was
problematic (Scheme 4). Numerous attempts to obtain lactol
Scheme 4. Acid-catalyzed hydrolysis of 2,3-unsaturated-4-aminopynano-
side.
Scheme 2. Synthesis of 2,3-dideoxy-4-amino-d-hex-2-enopyranose by allyl
cyanate-to-isocyanate rearrangement.
17 by acid-catalyzed hydrolysis of 15 were unsuccessful, and
only 2-substituted pyrrole 16 was isolated.
To circumvent this problem, we selected p-methoxyphenyl
glycoside 18 as a starting material (Scheme 5), because such
Dehydration of allyl carbamate 4 with triphenylphosphine
(PPh3), carbon tetrabromide (CBr4), and diisopropylethyl-
amine (iPr2NEt) under modified Appel×s condition[16] pro-
vided allyl cyanate 5, which underwent [3,3]-sigmatropic re-
arrangement to afford allyl isocyanate 6. Subsequent treat-
ment of 6 with trimethylaluminum afforded unsaturated
aminopyranose 7 in 59% yield.
In this study we reasoned that our method would provide
a convenient approach to the central core of cytosinine, and
this consideration led to the retrosynthetic strategy outlined
Scheme 5. Synthesis of hex-3-enopyranoside from 2-acetoxy glucal.
a glycoside could be oxidatively hydrolyzed under mild con-
ditions.[18] As a result, synthesis of cytosinine began with a
Ferrier-type glycosylation of 2-acetoxy-tri-O-acetyl-d-glucal
(14) using p-methoxyphenol and BF3¥OEt2 as the catalyst.
The resulting p-methoxyphenyl glycoside 18 was obtained as
a crystalline solid in 49% yield.[19] Treatment of 18 with
LiAlH4 initiated a cascade reaction, which involves enol ace-
tate cleavage, b-elimination of the resulting enolate 19, and
nucleophilic hydride addition to 20 from the sterically less-
hindered b-face[20] to furnish diol 21 exclusively in 84%
yield. The primary alcohol of 21 was then selectively pro-
Scheme 3. Strategy for the synthesis of cytosinine.
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Total Synthesis of (+)-Blasticidin S
3241 3251
tected with tert-butyldimethylsilyl (TBS) chloride in the
presence of a catalytic amount of 4-dimethylaminopyridine
(DMAP) and Et3N to afford TBS ether 22 in 75% yield.[21]
With the hex-3-enopyranoside 22 in hand, the next goal
was to synthesize the 4-amino-hex-2-enopyranose using an
allyl cyanate-to-isocyanate rearrangement (Scheme 6).
because we experienced better results for the cytosine N-
glycosylation step with 26a, and deprotection of the carba-
mate could be carried out under milder conditions.[23]
With a viable route to the unsaturated amino carbohy-
drate 26a having been established, we next focused on the
transformation of 26a into the corresponding methyl hex-2-
enopyranouronate and its subsequent cytosine N-glycosyla-
tion (Scheme 7). The TBS group in 26a was removed with
tetrabutylammonium fluoride (nBu4NF) buffered with acetic
acid (AcOH) in THF to provide 27 in 79% yield.[24] Swern
oxidation (oxalyl chloride, DMSO, Et3N, CH2Cl2) of 27 gave
the rather labile aldehyde 28, which was immediately sub-
jected to sodium chlorite oxidation (NaClO2, NaH2PO4, 2-
methyl-2-butene). Subsequent treatment of the resultant car-
boxylic acid with diazomethane in methanol furnished
methyl ester 29 as a white powdery solid in 70% overall
yield for the three steps. Hydrolysis of p-methoxyphenyl gly-
coside 29 was carried out with silver(ii) bis(hydrogen dipico-
linate) in aqueous acetonitrile.[25] Unfortunately, we found
that concentration of the CH2Cl2 extracts containing the
fragile lactol 30 often resulted in a tarry oil from which pyr-
1
role-like products could be identified by H NMR spectro-
scopy. To circumvent this problem, the CH2Cl2 extracts were
immediately treated with acetic anhydride, DMAP, and pyri-
dine; this protocol successfully provided acetyl glycoside 31
as an anomeric mixture (a:b 3:1) in 71% yield.[26] Since we
had succeeded in anomeric activation, we subsequently at-
tempted the cytosine N-glycosylation of 31. After screening
several silylated cytosine derivatives and Lewis acids, the
following points were observed: 1) we selected the 4-tert-bu-
tylbenzoyl group to protect the cytosine N4-nitrogen,[27] be-
cause products that contain this liphophilic protecting group
behave better chromatographically during efforts to sepa-
rate such anomeric mixtures than the corresponding acetyl
and benzoyl counterparts; 2) we found that Lewis acids such
as SnCl4 and BF3¥OEt2 could not be used because only pyr-
Scheme 6. Allyl cyanate-to-isocyanate rearrangement for the construction
of the 4-amino-hex-2-enopyranose component in cytosinine.
Treatment of 22 with trichloroacetyl isocyanate followed by
hydrolysis with potassium carbonate (K2CO3) in aqueous
methanol provided allyl carbamate 23. Dehydration of 23
(PPh3, CBr4, Et3N) gave allyl cyanate 24, which then under-
went [3,3]-bond reorganization at 08C (60 min) to afford
allyl isocyanate 25. Since isolation of 25 using aqueous
workup would result in lower yields, isocyanate 25 was treat-
ed in situ with 2,2,2-trichloroethanol[22] to give trichloro-
ethoxy (Troc) carbamate 26a in 75% overall yield from 22
after chromatographic purification. Carbamates 26b (Cbz)
and 26c (Alloc) were also prepared in a similar manner in
64 and 80% yields, respectively, when either benzyl alcohol
or allyl alcohol was employed in the reaction with 25. After
considerable experimentation, we elected to use Troc carba-
mate 26a rather than 26b or 26c in subsequent reactions,
1
role-like compounds were detected by H NMR analysis of
the crude products.[28] Fortunately, the Vorbr¸ggen method
using trimethylsilyl triflate (TMSOTf) was found to be suc-
cessful;[29] and 3) it was found that selectivity and yield de-
pended on the solvent and amount of cytosine used.[30] b-Se-
lectivity was optimal when the reaction was carried out in
Scheme 7. Final elaboration for the synthesis of the fully protected cytosinine.
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Y. Ichikawa et al.
FULL PAPER
THF and excess use of N4-(4-tert-butylbenzoyl)cytosine (ca.
6 8equiv) resulted in reproducible and better yields.
Reaction of N4-(4-tert-butylbenzoyl)cytosine with N,O-
bis(trimethylsilyl)acetamide (THF, room temperature,
30 min) gave silylated cytosine, which was subsequently
treated with 31 in the presence of TMSOTf at 08C for 2 h.
After workup, a 7:3 mixture of 32 and 33 was obtained in
55 63% yield in which the desired b-isomer predominat-
ed.[31] Careful separation of the mixture by silica-gel chro-
matography and repeated recrystallization afforded the fully
protected cytosinine 32 as a glassy wax. The stereochemistry
Scheme 9. Deprotection of nosyl group leading to a spontaneous cycliza-
tion to lactam.
reliable procedure on a gram-scale was realized by applying
the Weinreb protocol.[35] Treatment of 36 with dimethylalu-
minum benzylamide (PhCH2NHAlMe2) in benzene at room
temperature consistently provided 38 in yields of 72 83%.
The requisite N-methyl amino substituent was then intro-
duced by Fukuyama×s procedure.[36] Thus, substitution of al-
cohol 38 with N-methyl 2-nitrobenzenesulfonamide under
Mitsunobu conditions furnished o-nitrobenzenesulfonamide
39 in 83% yield. Deprotection of the o-nitrobenzenesulfonyl
group in 39 with thiophenol and cesium carbonate in aceto-
nitrile, followed by treatment of the resultant N-methyl-
amine 40 with N,N-di-(tert-butoxycarbonyl)-S-methyliso-
thiourea and mercuric chloride[37] afforded bis(Boc)-protect-
ed N-methyl guanidine 41 in 89% overall yield for the two
steps.
1
of 32 was determined on the basis of H NMR coupling con-
stants. The diagnostic coupling constant between H-4 and
H-5 for the b-isomer 32 is 9.0 Hz, whereas the a-isomer 33,
in which the cytosine moiety adopts in the pseudo-equatori-
al position, has a coupling constant of J4,5 =5.5 Hz.
Synthesis of blastidic acid: Retrosynthetic analysis of blasti-
dic acid (2) revealed that this b-amino acid possesses a
hidden symmetry. Therefore, the synthesis began with the
preparation of chiral carboxylic acid 35 by enantioselective
hydrolysis of meso-diester 34[32] using pig liver esterase[33]
(Scheme 8).
With the requisite functional groups for blastidic acid now
in place, we turned to the hydrolysis of the benzyl amide in
41. This involved a two-step procedure; amide activation
followed by hydrolysis[38] (Scheme 10). Initial attempts to se-
Scheme 10. Synthesis of the Boc-protected blastidic acid.
Scheme 8. Synthesis of blastidic acid statrting from meso-diester.
lectively introduce the tert-butoxycarbonyl (Boc) group onto
the nitrogen of the benzylamide in 41 [(Boc)2O, DMAP,
THF] failed, and only a mixture of N-Boc imides was isolat-
ed. Moreover, NMR analysis of the products was complicat-
ed by the existence of rotamers and broadening of the spec-
tra. The following observations for the N-Boc carboxylation
of model compounds 48 and 49 suggested a solution for this
step (Figure 1).
Firstly, selective N-Boc carboxylation of the benzyl amide
in 48 failed, and a mixture of products was isolated. Second-
ly, the nitrogen in the bis(Boc)-protected N-methyl guani-
dine 49 also underwent Boc-carboxylation to furnish tris-
(Boc)-protected N-methyl guanidine 50. Thirdly, the com-
Although diborane reduction of carboxylic acid 35 yielded
alcohol 36, sometimes small amounts of lactone 37 were
formed during workup. Moreover, preliminary experiments
revealed that methyl ester 42 rapidly cyclized to form
lactam 43 upon removal of the sulfonyl group (Scheme 9).
To avoid these problems, methyl ester 36 was transformed
into benzyl amide 38.[34] In practice, the crude product 36
was treated with benzylamine in refluxing benzene. This
gave 38 in 74% yield on a 200 mg scale, while on larger
scales the yield decreased to less than 50%. As a result of
this problem another method was briefly investigated and a
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Total Synthesis of (+)-Blasticidin S
3241 3251
under vacuum for several hours, and the resultant trifluoro-
acetate salt was treated with 3n HCl. To our surprise,
1H NMR analysis indicated that the resultant product was a
mixture of blastidic acid and a by-product, which was subse-
quently identified as pseudoblastidone (52).[40] To circum-
vent the formation of 52, the crude trifluoroacetate salt of
blastidic acid was immediately treated with 3n HCl and
Figure 1. Model compounds for N-Boc carboxylation.
1
petitive carboxylation of 48 and 49 revealed that bis(Boc)-
protected N-methyl guanidine 49 is acylated more rapidly
then purified by ion-exchange chromatography. The H and
13C NMR spectra for our synthetic 51 were found to be iden-
tical to those obtained for a sample that had been prepared
from natural blasticidin S.[41]
1
than 48. Finally, H NMR analysis of 50 showed the pres-
ence of two rotamers in a 3:1 ratio. Therefore, benzylamide
41 was subjected to exhaustive acylation with di-tert-butyl
dicarbonate (6.0 equiv) and DMAP (1.5 equiv) in THF to
furnish the penta-N-Boc imide 44 in 67% yield. Unfortu-
nately, saponification of 44 (LiOH, THF, H2O) gave only
complex mixtures; these were presumed to arise from base-
catalyzed b-elimination of the imide group at C-3. This un-
desired elimination could be suppressed by lowering the
leaving group ability. Therefore, the Cbz group in 44 was re-
moved by catalytic hydrogenolysis to give the Boc-carba-
mate 45 in 77% yield, and methanolysis of 45 with tetrame-
thylguanidine in methanol then cleanly afforded methyl
ester 46 in 87% yield.[39] Finally, hydrolysis of methyl ester
46 with lithium hydroxide in aqueous THF furnished the
Boc-protected blastidic acid 47 in 97% yield.
Since NMR analysis of 47 was complicated with extensive
broadening, we attempted to prepare blastidic acid dihydro-
chloride 51 in order to compare it with an authentic sample
prepared from natural blasticidin S (Scheme 11). Boc-Pro-
tected blastidic acid 47 was thereby treated with trifluoro-
acetic acid (TFA) in CH2Cl2 at room temperature for 2 h.
The reaction mixture was then concentrated and dried
Total synthesis of blasticidin S: With the fully protected cy-
tosinine 32 and blastidic acid 47 in hand, our efforts turned
toward coupling these two components (Scheme 12). Initial-
ly, we focused on removing the Troc group in 32; this
proved to be more difficult than expected. Attempts to de-
protect 32 with zinc (washed with aqueous HCl)[42] and
acetic acid in THF afforded non-reproducible results, be-
cause deprotection was also accompanied by hydrolysis of
the N4-cytosine tert-butylbenzoyl group. We then tried to
use zinc that had been activated with TMSCl.[43] Unfortu-
nately, hydrolysis of the sensitive tert-butylbenzoyl group as
well as mono-dechlorination of the Troc group occurred.
After considerable frustration, we were very gratified to
find that a cadmium/lead (Cd/Pb) couple reported by Ciufo-
lini[44] was a mild and efficient method for this deprotection.
Treatment of 32 with a large excess (40 equiv) of the Cd/Pb
couple at room temperature for 30 min afforded the desired
amine 53 (93% based on consumed 32) together with recov-
ered starting material 32 (13% recovered) to set the stage
for the coupling reaction. After a number of coupling meth-
ods for amide bond construction were surveyed, an in situ
activation method using the BOP reagent[45] proved to be
most effective. In particular, attempts to condense 53 with
protected blastidic acid 47 using BOP and diisopropylethyl-
amine (iPr2NEt) gave rise to the desired coupled product
54, albeit in only 32% yield. However, addition of 1-hy-
droxy-benzotriazole (HOBt)[46] improved the yields up to
73%.
All that remained to complete the total synthesis was to
remove the six protecting groups present in 54. However,
we were particularly concerned about the possibility of the
blastidic acid moiety undergoing base-catalyzed cyclization
Scheme 11. Preparation of blastidic acid dihydrochloride.
Scheme 12. Total synthesis of blasticidin S.
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Y. Ichikawa et al.
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≈
tamers are listed in parentheses. High-resolution mass spectra (HRMS)
are reported in m/z. Elemental analyses were performed by the Analyti-
cal Laboratory at the Graduate School of Bioagricultural Sciences,
Nagoya University. Reactions were run under an atmosphere of nitrogen
if the reactions were sensitive to moisture or oxygen. Dichloromethane
was dried over molecular sieves (3 ä), while acetonitrile was stored over
molecular sieves (4 ä). Pyridine and triethylamine were stored over an-
hydrous KOH. All other commercially available reagents were used as
received.
during the deprotection sequence. Indeed, Otake reported
that treatment of blasticidin S (1) with base afforded cyto-
mycin (55)[8] (Scheme 13).
Mol scale preparation of 2-acetoxy-d-glucal (14): A 5 L three-necked,
round-bottomed flask equipped with a mechanical stirrer and a dropping
funnel was charged with acetic anhydride (2.0 L) and perchloric acid
(HClO4, 30%, 7.0 mL). d-Glucose (500 g, 2.78mol) was added to this sol-
ution portionwise over 2 h. After stirring at room temperature overnight,
the reaction mixture was cooled to ꢀ108C and PBr3 (1.30 kg, 13.7 mol)
was added through a dropping funnel. Mechanically efficient stirring was
necessary during the addition because, otherwise, pentaacetyl glucopyra-
noside began to crystallize upon cooling. Water (133 mL, 5.54 mol) was
added dropwise to this solution as the temperature was maintained be-
tween ꢀ5 to ꢀ108C. The cooling bath was removed, the mixture was al-
lowed to stand for 4 d at room temperature, and was then poured into ice
water (ca. 7.0 L). The resultant brown mixture was stirred vigorously
with a mechanical stirrer as the product gradually solidified. The crude
product was filtered and then washed with water. The resultant brown
solid was dissolved in CH2Cl2 (ca. 800 mL) and the solution was washed
with saturated aqueous NaHCO3, dried (Na2SO4), and concentrated to
give a viscous oil. This material was dissolved in diethyl ether and then
concentrated, and this procedure was repeated until crystallization began
to furnish white crystals (974.8g, 85%). The product was then used in
the next reaction without further purification.
Scheme 13. Base-catalyzed cyclization of blasticidin S to afford cytomy-
cin.
Therefore, deprotection of 54 began with the base-cata-
lyzed hydrolysis of the tert-butylbenzoyl protecting group
and methyl ester substituent. Treatment of 54 with Et3N in
MeOH promoted cleavage of the labile tert-butylbenzoyl
group, while addition of water to the reaction mixture re-
sulted in the hydrolysis of the methyl ester group. After the
reaction mixture was concentrated, the four Boc protecting
groups in the resultant carboxylate were removed by se-
quential treatment with TFA in CH2Cl2 and then 3n HCl.
The resulting blasticidin S hydrochloride was purified on
Amberlite IRA-410 (OHꢀ form) to afford blasticidin S (1)
as the free base in 85% yield.[47] Our synthetic material was
found to be identical (1H NMR, 13C NMR, IR, [a]D, TLC)
with an authentic sample of natural blasticidin S.[48]
DBU (305 mL, 2.04 mol) was added through a dropping funnel to a solu-
tion of a-bromo tetra-O-acetyl-glucopyranoside (778g, 1.89 mol) dis-
solved in DMF (1.70 L) at ꢀ108C. After stirring at ꢀ108C for 60 min,
the resultant brown reaction mixture was poured into ice water (ca.
10 L). Crystallization began with the aid of seeding and vigorous scratch-
ing. The crude product was filtered and then dried in air to afford the
Conclusion
crude solid (533 g). Recrystallization from
a mixture of methanol
(500 mL) and water (400 mL) afforded 14 (359 g, 57%) as white crystals.
An allyl cyanate-to-isocyanate rearrangement has been suc-
cessfully employed for the construction of an unsaturated
amino sugar moiety in cytosinine. Furthermore, synthesis of
the protected cytosinine 32 was achieved in 11 steps starting
from 2-acetoxy-tri-O-acetyl-d-glucal (14) in 4.0% overall
yield.[49] The Boc-protected blastidic acid (47) was synthe-
sized in nine steps from chiral carboxylic acid 35 (overall
yield 23%). The two components 53 and 47 were coupled
using the BOP method in the presence of HOBt, and subse-
quent global deprotection of 54 completed the first total
synthesis of blasticidin S (1).
p-Methoxyphenyl 2,4,6-triacetyl-3-deoxy-a-d-erythro-hex-2-enopyrano-
side (18): BF3¥OEt2 (1.20 mL, 9.5 mmol) was added to a solution of 2-ace-
toxy-d-glucal (14; 100.0 g, 303 mmol) and 4-methoxyphenol (40.0 g,
322 mmol) in benzene (1.20 L) under an atmosphere of nitrogen. After
stirring at room temperature overnight, the reaction mixture was poured
into saturated aqueous NaHCO3. The separated organic layer was
washed with brine, dried (Na2SO4), and concentrated under reduced pres-
sure. The resultant orange viscous oil (113.7 g) was recrystallized from
methanol to afford 18 (56.9 g, 49%) as white crystals. M.p. 658C; [a]D21
=
+150.8( c=1.02 in CHCl3); 1H NMR (300 MHz, CDCl3): d=2.03 (s,
3H), 2.10 (s, 3H), 2.19 (s, 3H), 3.77 (s, 3H), 4.18 4.38 (m, 3H), 5.52
(ddd, J=10.0, 2.0, 1.0 Hz, 1H), 5.56 (brs, 1H), 5.86 (d, J=2.0 Hz, 1H),
6.83 (m, 2H), 7.03 ppm (m, 2H); 13C NMR (75 MHz, CDCl3): d=20.5,
20.76, 20.80, 55.5, 62.3, 65.1, 67.8, 93.7, 114.5, 116.0, 118.7, 145.7, 151.0,
155.5, 168.2, 170.1, 170.7 ppm; IR (KBr): n˜max =2955, 2839, 1748, 1508,
1373, 1214, 1034 cmꢀ1; elemental analysis calcd (%) for C19H22O9: C
57.86, H 5.62; found: C 57.78, H 5.62.
Experimental Section
p-Methoxyphenyl 3,4-dideoxy-a-d-erythro-hex-3-enopyranoside (21): A
solution of lithium aluminum hydride (150 mg, 3.94 mmol) in THF
(8.8 mL) was cooled to 08C under an atmosphere of nitrogen. Glycoside
18 (500 mg, 1.32 mmol) in THF (1.0 mL) was then added dropwise to this
solution over 1 h. After stirring at 08C for 1 h, EtOAc (0.10 mL), water
(0.15 mL), hexane (6.0 mL), 15% aqueous NaOH (0.15 mL), and water
(0.45 mL) were sequentially added, and the reaction was stirred at room
temperature overnight. The reaction mixture was then filtered though a
pad of Super Cell, and the filter cake was washed with EtOAc and a 5:1
mixture of CH2Cl2 and methanol. Concentration of the filtrate under re-
duced pressure gave a crude solid (447 mg). This was purified by silica-
gel chromatography (2:1 diethyl ether/hexane followed by diethyl ether
as eluent) to afford 21 (277 mg, 84%) as white crystals. M.p. 1408C;
[a]2D1 =+93.8( c=0.96 in MeOH); 1H NMR (300 MHz, CDCl3): d=1.86
(brs, 1H), 2.34 (d, J=12.0 Hz, 1H), 3.58 3.78 (m, 2H), 3.80 (s, 3H),
4.31 4.45 (m, 2H), 5.61 (d, J=4.0 Hz, 1H), 5.78(dt, J=11.0, 1.5 Hz,
General: Melting points were recorded on a micro melting-point appara-
tus and are not corrected. Optical rotations are given in units of
10ꢀ1 degcm2 gꢀ1. Infrared spectra are reported in wave number (cmꢀ1).
1H NMR chemical shifts (d) are reported in parts per million (ppm) rela-
tive to tetramethylsilane (TMS) (d 0.00 in CDCl3), CHD2OD (d 3.31 in
CD3OD), tBuOH (d 1.24 in D2O), or [D4]3-(trimethylsilyl)propionic-
2,2,3,3 acid sodium salt (TSP) (d 0.00 in 1n DCl) as internal standards.
Data are reported as follows: chemical shift, integration, multiplicity (s=
singlet, d=doublet, t=triplet, q=quartet, qn=quintet, sext=sextet,
br=broadened, m=multiplet), coupling constants (J, given in Hz).
13C NMR chemical shifts (d) are recorded in parts per million (ppm) rela-
tive to CDCl3 (d 77.0), CD3OD (d 49.0), or 1,4-dioxane (d 67.4 in D2O or
1n DCl) as internal standards. The tri-Boc-N-methyl guanidine deriva-
tives exit as a mixture of rotamers on the NMR time scale. In instances
where two rotamers were observed, the carbon signals for the minor ro-
3246
¹ 2004 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
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Chem. Eur. J. 2004, 10, 3241 3251

Total Synthesis of (+)-Blasticidin S
3241 3251
1H), 5.92 (ddd, J=11.0, 4.0, 1.5 Hz, 1H), 6.85 (m, 2H), 7.06 ppm (m,
2H); 13C NMR (75 MHz, CDCl3): d=55.6, 64.2, 64.7, 69.7, 96.9, 114.7,
118.1, 126.7, 128.7, 151.0, 155.4 ppm; IR (KBr): n˜max =3336, 3228, 2950,
1509, 1457, 1228, 1038, 933, 828 cmꢀ1; elemental analysis calcd (%) for
C13H16O5: C 61.90, H 6.39; found: C 61.88, H 6.33.
and concentrated under reduced pressure. The crude residue was purified
by silica-gel chromatography (1:1 diethyl ether/hexane followed by dieth-
yl ether as eluent) to furnish 27 (197 mg, 79%) as a low-melting solid.
M.p. 388C; [a]2D6 =+71.9 (c=0.99 in CHCl3); 1H NMR (300 MHz,
CDCl3): d=2.60 (t, J=7.0 Hz, 1H), 3.73 3.88 (m, 3H), 3.78 (s, 3H), 4.53
(brt, J=10.0 Hz, 1H), 4.69 (d, J=12.0 Hz, 1H), 4.84 (d, J=12.0 Hz, 1H),
5.08(d, J=9.5 Hz, 1H), 5.63 (brs, 1H), 5.97 6.08(m, 2H), 6.83 (m, 2H),
7.03 ppm (m, 2H); 13C NMR (75 MHz, CDCl3): d=45.2, 55.5, 61.6, 71.5,
74.6, 93.8, 95.3, 114.6, 118.4, 127.1, 131.1, 151.0, 155.1, 155.2 ppm; IR
(KBr): n˜max =3738, 2944, 1735, 1560, 1508, 1216, 1102 cmꢀ1; elemental an-
alyis calcd (%) for C16H18Cl3NO6: C 45.04, H 4.25, N 3.28; found: C
45.04, H 4.26, N 3.30.
p-Methoxyphenyl 6-O-(tert-butyldimethylsilyl)-3,4-dideoxy-a-d-erythro-
hex-3-enopyranoside (22): tert-Butyldimethylsilyl chloride (10.0 g,
66.3 mmol) was added to a solution of diol 21 (25.0 g, 99 mmol), triethyl-
amine (23.0 mL, 165 mmol), and 4-dimethylaminopyridine (0.80 g,
6.6 mmol) in CH2Cl2 (2.00 L). After stirring at room temperature over-
night, additional triethylamine (8.0 mL, 57 mmol), 4-dimethylaminopyri-
dine (0.25 g, 2.0 mmol), and tert-butyldimethylsilyl chloride (5.0 g,
33.3 mmol) were added, and stirring was continued overnight. The reac-
tion mixture was concentrated to one-third of its volume and was then di-
luted with hexane. The resultant solution was washed with water, 1m
aqueous KHSO4, saturated aqueous NaHCO3, brine, and dried (Na2SO4).
Concentration under reduced pressure gave a viscous oil. This was dis-
solved in hexane, and upon cooling to ꢀ208C, 22 (23.88 g) crystallized as
a low-melting, white solid, which was filtered. The mother liquors were
concentrated and then purified by recrystallization to provide additional
product (3.18g, total yield of 75%). M.p. 52 8C; [a]2D4 =+58.0 (c=1.03 in
CHCl3); 1H NMR (300 MHz, CDCl3): d=0.05 (s, 6H), 0.88 (s, 9H), 2.36
(brd, J=12.0 Hz, 1H), 3.62 (dd, J=10.5, 6.0 Hz, 1H), 3.70 (dd, J=10.5,
5.5 Hz, 1H), 3.77 (s, 3H), 4.24 (m, 1H), 4.35 (m, 1H), 5.56 (d, J=4.5 Hz,
1H), 5.80 5.92 (m, 2H), 6.82 (m, 2H), 7.07 ppm (m, 2H); 13C NMR
(75 MHz, CDCl3): d=ꢀ5.50, ꢀ5.46, 18.1, 25.7, 55.6, 64.3, 65.2, 69.7, 96.9,
114.6, 118.0, 127.3, 127.9, 151.2, 155.2 ppm; IR (KBr): n˜max =2929, 2858,
1509, 1473, 1219, 1096 cmꢀ1; elemental analysis calcd (%) for C19H30O5Si:
C 62.26, H 8.25; found: C 62.23, H 8.40.
Methyl 1-O-p-methoxyphenyl-2,3,4-trideoxy-4-(2,2,2-trichloroethoxycar-
bonylamino)-a-d-erythro-hex-2-enopyranosyluronate
(29):
DMSO
(0.62 mL, 8.7 mmol) in CH2Cl2 (6.0 mL) was added to a solution of
oxalyl chloride (0.56 mL, 6.42 mmol) in CH2Cl2 (18mL) at ꢀ788C under
an atmosphere of nitrogen. After stirring at ꢀ788C for 10 min, alcohol 27
(1.37 g, 3.21 mmol) in CH2Cl2 (6.0 mL) was added and stirring was con-
tinued for 1 h. Triethylamine (2.24 mL, 16.1 mmol) was introduced and
the reaction mixture was stirred for 10 min at ꢀ788C, and was then al-
lowed to warm to 08C. The resultant reaction mixture was poured into
cold saturated aqueous NH4Cl and the aqueous layer was extracted with
diethyl ether. The combined organic layers were washed with brine, dried
(Na2SO4), and then concentrated under reduced pressure to give the
crude aldehyde 28 as a labile dark-brown liquid. This material was em-
ployed immediately without further purification in subsequent reactions.
Sodium chlorite (85%, 0.64 g, 7.1 mmol) was added portionwise to a sol-
ution of aldehyde 28, 2-methyl-2-butene (2 drops), NaH2PO4 monobasic
dihydrate (8.0 g) in a mixture of tert-butyl alcohol (32 mL) and water
(8.0 mL). After stirring at room temperature for 1 h, the reaction mixture
was poured into saturated aqueous NH4Cl and the aqueous layer was ex-
tracted with diethyl ether. The combined organic layers were dried
(Na2SO4) and then concentrated under reduced pressure. Since the resul-
tant crude carboxylic acid was not stable upon storage, the residue was
immediately dissolved in methanol (32 mL) and was then treated at 08C
with a diethyl ether solution of diazomethane until the yellow color per-
sisted. The excess diazomethane was quenched by dropwise addition of
acetic acid until the color dissipated, and the resultant reaction mixture
was concentrated under reduced pressure to give a yellow oil. Purifica-
tion of the crude product by silica-gel chromatography (1:2 diethyl ether/
hexane) provided 29 (1.02 g, 70% for three steps) as white crystals. M.p.
p-Methoxyphenyl 6-O-(tert-butyldimethylsilyl)-2,3,4-trideoxy-4-(2,2,2-tri-
chloroethoxycarbonylamino)-a-d-erythro-hex-2-enopyranoside (26a): Tri-
chloroacetyl isocyanate (10.5 mL, 92.4 mmol) was added to a solution of
22 (28.2 g, 77.0 mmol) in CH2Cl2 (300 mL) at 08C. After stirring at 08C
for 1 h, the reaction mixture was concentrated and the resultant residue
was dissolved in methanol (100 mL). Water (100 mL) and potassium car-
bonate (16.1 g, 231 mmol) were added at 08C, and the cooling bath was
removed. After stirring at room temperature for 2.5 h, the reaction mix-
ture was concentrated under reduced pressure to remove methanol. The
resultant aqueous phase was extracted with CH2Cl2 and the combined or-
ganic layers were dried (Na2SO4) and concentrated to afford carbamate
23 (25.6 g), which was used for the next reaction without further purifica-
tion.
1
438C; [a]2D6 =+57.1 (c=1.43 in CHCl3); H NMR (300 MHz, CDCl3): d=
3.76 (s, 3H), 3.77 (s, 3H), 4.49 (d, J=9.0 Hz, 1H), 4.62 (brt, J=9.0 Hz,
1H), 4.73 (brs, 2H), 5.25 (brd, J=9.0 Hz, 1H), 5.72 (brs, 1H), 6.02 (m,
2H), 6.83 (m, 2H), 7.03 ppm (m, 2H); 13C NMR (75 MHz, CDCl3): d=
47.1, 52.6, 55.4. 70.3, 74.5, 93.4, 95.2, 114.5, 118.3, 126.8, 130.4, 150.8,
154.2, 155.2, 169.3 ppm; IR (KBr): n˜max =3343, 2955, 2837, 1745, 1508,
1440, 1216, 1036, 994, 828, 725 cmꢀ1; elemental analysis calcd (%) for
C17H18Cl3NO7: C 44.91, H 3.99, N 3.08; found: C 45.08, H 4.07, N 3.14.
Carbon tetrabromide (18.8 g, 56.6 mmol) in CH2Cl2 (30 mL) was added
to a solution of carbamate 23 (8.27 g, 20.2 mol), triphenylphosphine
(13.3 g, 50.5 mmol), and triethylamine (7.04 mL, 50.5 mmol) in CH2Cl2
(170 mL) at ꢀ408C under an atmosphere of nitrogen. The reaction mix-
ture was gradually warmed to 08C over 30 min and was then stirred at
08C for 1 h. 2,2,2-Trichloroethanol (11.6 mL, 0.12 mol) was introduced,
and stirring was continued at 08C for 2 h and then at room temperature
overnight. The reaction mixture was washed with 1n HCl and aqueous
saturated NaHCO3, dried (Na2SO4), then concentrated under reduced
pressure. The residue was purified by silica-gel chromatography (stepwise
gradient of 15 30% diethyl ether/hexane as eluent) to furnish 26a
(10.1 g, 75% for the two steps). M.p. 778C; [a]2D6 =+108.5 (c=1.01 in
CHCl3); 1H NMR (300 MHz, CDCl3): d=0.025 (s, 3H), 0.035 (s, 3H),
0.86 (s, 9H), 3.70 3.84 (m, 2H), 3.77 (s, 3H), 3.93 (m, 1H), 4.32 (t, J=
9.0 Hz, 1H), 4.71 (d, J=12.0 Hz, 1H), 4.79 (d, J=12.0 Hz, 1H), 5.28
(brs, 1H), 5.52 (brs, 1H), 5.92 6.04 (m, 2H), 6.82 (m, 2H), 7.06 ppm (m,
2H); 13C NMR (75 MHz, CDCl3): d=ꢀ5.64, ꢀ5.56, 18.2, 25.8, 46.6, 55.6,
63.5, 71.5, 74.5, 76.2, 94.0, 95.4, 114.5, 119.0, 126.6, 131.6, 151.3, 154.4,
155.2 ppm; IR (KBr): n˜max =3804, 3746, 1718, 1735, 1719, 1541, 1508,
1216 cmꢀ1; elemental analysis calcd (%) for C22H32Cl3NO6Si: C 48.85, H
5.96, N 2.59; found: C 48.84, H 5.99, N 2.60.
Methyl 1-O-acetyl-2,3,4-trideoxy-4-(2,2,2-trichloroethoxycarbonylamino)-
d-erythro-hex-2-enopyranosyluronate (31): Silver(ii) bis(hydrogen dipico-
linate) (3.70 g, 8.00 mmol) was added to
a solution of 29 (1.25 g,
2.77 mmol) in a mixture of acetonitrile (15.0 mL) and water (5.0 mL) at
08C. The resultant suspension was vigorously stirred at 08C for 15 min.
The reaction mixture was diluted with CH2Cl2 (30 mL) to precipitate a
white slurry, and the organic layer was decanted and then washed with
water (40 mL). The aqueous layer was extracted with CH2Cl2 (about
40 mL) and the combined organic layers were washed with saturated
aqueous NaHCO3, dried (Na2SO4), and filtered. Pyridine (4.0 mL), acetic
anhydride (3.0 mL), and 4-dimethylaminopyridine (0.50 g, 4.10 mmol)
were added to the filtrate. After standing at room temperature overnight,
the resultant solution was concentrated under reduced pressure. The
crude residue was purified by silica-gel chromatography (1:3 EtOAc/
hexane) to afford acetyl glycoside 31 (760 mg, 71%) as an inseparable
mixture of two anomers 3:1.
p-Methoxyphenyl 2,3,4-trideoxy-4-(2,2,2-trichloroethoxycarbonylamino)-
a-d-erythro-hex-2-enopyranoside (27):
A solution of 26a (318mg,
0.59 mmol) and acetic acid (0.20 mL, 3.53 mmol) in THF (6.0 mL) cooled
to 08C was treated with tetrabutylammonium fluoride (1.0m solution in
THF, 0.70 mL, 0.70 mmol). After stirring at 08C for 1 h, the reaction mix-
ture was poured into water and the aqueous layer was extracted with di-
ethyl ether. The combined organic layers were washed with saturated
aqueous NH4Cl, saturated aqueous NaHCO3, and brine, dried (Na2SO4),
Fully protected cytosinine 32, its epimer 33, and the pyrrole by-prod-
uct[31]: N,O-Bis(trimethylsilyl)acetamide (2.00 mL, 7.54 mmol) was slowly
added to a slurry of N4-tert-butylbenzoylcytosine (912 mg, 3.35 mmol) in
THF (10.0 mL) under an atmosphere of nitrogen at room temperature.
After stirring at room temperature for 30 min a homogeneous solution
resulted. Trimethysilyl trifluoromethanesulfonate (TMSOTf, 0.14 mL,
3247
Chem. Eur. J. 2004, 10, 3241 3251
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¹ 2004 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

Y. Ichikawa et al.
FULL PAPER
0.78mmol) was then added and stirring was continued for 30 min. The
resultant solution was cooled to 08C, and a solution of 31 (159 mg,
0.41 mmol) in THF (1.25 mL) was added. After stirring at 08C for 2.0 h,
the reaction mixture was diluted with CH2Cl2 and then poured into aque-
ous NaHCO3. The resultant precipitate was filtered through Super Cell
and the aqueous layer was extracted with CH2Cl2. The combined extracts
were dried (Na2SO4) and concentrated. The crude residue (379 mg) was
purified by silica-gel chromatography (1:7 and 1:1 of EtOAc/hexane fol-
lowed by EtOAc) to afford a mixture of the two anomers, 32 and 33
(156 mg, 63%, 32:33=7:3, as determined by 1H NMR analysis). The re-
sultant mixture was roughly separated by silica-gel chromatography
(100:1 CH2Cl2/MeOH), and 32 was further recrystallized from a mixture
of EtOAc and hexane (three times) to afford pure 32 as pale-yellow crys-
tals. Further purification of 33 was performed by preparative thin-layer
chromatography (95:5 CH2Cl2/MeOH).
by silica-gel chromatography (1:3 EtOAc/hexane) to yield 34 as a color-
less oil (30.29 g, 72%).
Benzyl-(3S)-3-benzyloxycarbonylamino-5-hydroxypentanamide (38):
A
diborane solution (1.08m solution in THF, 3.50 mL, 3.70 mmol) was
added to a solution of carboxylic acid 35 (1.00 g, 3.70 mmol) in THF
(30.0 mL) at ꢀ108C under an atmosphere of nitrogen. After stirring at
ꢀ108C for 1 h the mixture was allowed to warm to room temperature,
and stirring was continued at room temperature for 3 h. The resultant re-
action mixture was quenched by the addition of MeOH, and was then
concentrated under reduced pressure. The resultant residue was purified
by silica-gel short column chromatography (eluted with EtOAc) to afford
crude alcohol 36 (899 mg) as a colorless oil which was used in the next
step without further purification. An analytically pure sample was pre-
pared in another run and purified by chromatography to give 36 as a col-
orless oil. [a]2D7 =ꢀ34.0 (c=0.75 in CHCl3); 1H NMR (300 MHz, CDCl3):
d=1.64 (m, 1H), 1.68(brs, 1H), 1.80 (m, 1H), 2.56 (dd, J=16.0, 5.0 Hz,
1H), 2.66 (dd, J=16.0, 5.0 Hz, 1H), 2.90 3.00 (brs, 1H), 3.60 3.72 (brs,
1H), 3.68(s, 3H), 4.22 (m, 1H), 5.10 (d, J=12.0 Hz, 1H), 5.13 (d, J=
12.0 Hz, 1H), 5.62 (brd, J=9.0 Hz, 1H), 7.30 7.39 ppm (m, 5H);
13C NMR (75 MHz, CDCl3): d=37.0, 38.6, 44.8, 51.7, 58.6, 66.9, 128.1,
128.2, 128.6, 136.3, 157.0, 172.2 ppm; IR (KBr): n˜max =3355, 1718, 1540,
1259, 1061 cmꢀ1; elemental analysis calcd (%) for C14H19NO5: C 59.78, H
6.81, N 4.98; found: C 59.78, H 6.83, N 4.86.
Compound 32: m.p. 1358C; [a]2D7 =+102.6 (c=0.28in CHCl 3); 1H NMR
(300 MHz, CD3OD): d=1.36 (s, 9H), 3.72 (s, 3H), 4.43 (d, J=9.0 Hz,
1H), 4.62 (dq, J=9.0, 2.5 Hz, 1H), 4.79 (m, 2H), 5.95 (ddd, J=10.5, 2.5,
2.0 Hz, 1H), 6.18(dt, J=10.5, 2.0 Hz, 1H), 6.66 (brs, 1H), 7.58(d, J=
8.5 Hz, 2H), 7.63 (d, J=7.5 Hz, 1H), 7.92 (d, J=8.5 Hz, 1H), 8.23 ppm
(d, J=7.5 Hz, 1H); 13C NMR (75 MHz, CD3OD): d=31.5, 36.0, 48.0,
53.3, 75.6, 77.4, 81.3, 97.1, 99.4, 126.9, 127.2, 129.3, 131.7, 134.5, 147.3,
156.5, 157.9, 158.3, 165.5, 170.4 ppm; IR (KBr): n˜max =3421, 2959, 1734,
1699, 1670, 1486, 1254 cmꢀ1
; elemental analysis calcd (%) for
Benzylamine (0.50 mL, 4.1 mmol) was slowly added to a solution of tri-
methylaluminum (1.0m solution in hexane, 1.70 mL, 1.70 mmol) in ben-
zene (25.0 mL) at ꢀ108C under an atmosphere of nitrogen. After being
stirred at ꢀ108C for 20 min, the mixture was allowed to warm to room
temperature and was then stirred for a further 45 min. Methyl ester 36
(899 mg, crude) in benzene (1.0 mL) was then added to this solution at
08C. After being stirred at 08C for 20 min, the reaction mixture was al-
lowed to warm to room temperature and was stirred for a further 2 h.
The reaction was then quenched by the addition of MeOH. The resultant
mixture was filtered through a pad of Super Cell and the filtrate was con-
centrated under reduced pressure to give a residue. This was purified by
silica-gel column chromatography (eluted with 95:5 CH2Cl2/MeOH) to
afford benzylamide 38 (1.10 g, 83% for two steps) as pale-yellow crystals.
M.p. 1278C); [a]2D4 =ꢀ16.4 (c=1.02 in MeOH); 1H NMR (300 MHz,
CD3OD): d=1.73 (m, 2H), 2.45 (d, J=7.0 Hz, 2H), 3.59 (d, J=6.0 Hz,
1H), 3.61 (d, J=6.0 Hz, 1H), 4.12 (brqn, J=7.0 Hz, 1H), 4.31 (d, J=
15.0 Hz, 1H), 4.35 (d, J=15.0 Hz, 1H), 5.05 (s, 2H), 7.18 7.35 ppm (m,
10H); 13C NMR (75 MHz, CD3OD): d=38.5, 42.5, 44.1, 47.6, 59.7, 67.4,
128.3, 128.7, 128.9, 129.1, 129.55, 129.62, 138.4, 139.9, 158.4, 173.3 ppm;
IR (KBr): n˜max =3309, 1691, 1645, 1542, 1272 cmꢀ1; elemental analysis
calcd (%) for C20H24N2O4: C 67.40, H 6.79, N 7.86; found: C 67.40, H
6.68, N 7.97.
C25H27ClN4O7: C 49.89, H 4.52, N 9.31; found: C 49.78, H 4.64, N 9.03.
Compound 33: m.p. 1378C; [a]2D6 =ꢀ51.9 (c=0.27 in CHCl3); 1H NMR
(300 MHz, CD3OD): d=1.35 (s, 9H), 3.75 (s, 3H), 4.48(m, 1H), 4.51 (d,
J=5.5 Hz, 1H), 4.80 (m, 2H), 6.02 (ddd, J=10.5, 2.5, 2.0 Hz, 1H), 6.30
(ddd, J=10.0, 3.5, 2.0 Hz, 1H), 6.77 (brs, 1H), 7.54 7.60 (m, 3H), 7.91
(m, 2H), 8.25 ppm (d, J=7.5 Hz, 1H); 13C NMR (75 MHz, CD3OD): d=
31.5, 36.0, 47.5, 53.2, 74.1, 75.6, 79.4, 97.1, 98.6, 126.9, 129.3, 131.7, 132.4,
148.1, 156.4, 158.2, 158.3, 165.5, 170.8 ppm; IR (KBr): n˜max =3423, 1741,
1654, 1256 cmꢀ1; elemental analysis calcd (%) for C25H27ClN4O7: C 49.89,
H 4.52, N 9.31; found: C 49.88, H 4.70, N 9.10.
N,O-Bis(trimethylsilyl)acetamide (1.50 mL, 6.08mmol) was slowly added
to a slurry of N4-tert-butylbenzoylcytosine (1.33 g, 4.88 mmol) in THF
(12.0 mL) under an atmosphere of nitrogen at room temperature. After
stirring at room temperature for 30 min a homogeneous solution resulted.
Trimethysilyl trifluoromethanesulfonate (TMSOTf, 0.18mL, 0.99 mmol)
was then added and stirring was continued for a further 30 min. The re-
sultant solution was cooled to 08C and
a solution of 31 (327 mg,
0.96 mmol) in THF (2.0 mL) was added. After stirring at 08C for 3.5 h,
the reaction mixture was diluted with CH2Cl2 and then poured into aque-
ous NaHCO3. The resultant precipitate was filtered through Super Cell
and the aqueous layer was extracted with CH2Cl2. The combined extracts
were then dried (Na2SO4) and concentrated. The crude residue (579 mg)
was purified by silica-gel chromatography (1:7 and 1:1 of EtOAc/hexane
followed by EtOAc) to afford the pyrrole by-product (60 mg, 20%),[31] as
well as an anomeric mixture of 32 and 33 (319 mg, 55%) (a:b=7:3, as
Benzyl-(3S)-3-benzyloxycarbonylamino-5-(N-methyl-2-nitrobenzensulfo-
nylamino)pentanamide (39): Diethyl azodicarboxylate (0.40 mL,
2.55 mmol) was added to a solution of 38 (324 mg, 0.91 mmol), N-methyl-
2-nitorobenzenesulfonamide (275 mg, 1.27 mmol), and tri-n-butylphos-
phine (0.64 mL, 2.55 mmol) in THF (15 mL) at 08C. The reaction mix-
ture was warmed to room temperature and was then stirred for 2 h. Ad-
ditional tri-n-butylphosphine (0.64 mL, 2.55 mmol) and diethylazodicar-
boxylate (0.40 mL, 2.55 mmol) were introduced at room temperature.
The resultant solution was stirred for 20 min and was then concentrated
under vacuum. The residue was purified by silica-gel column chromatog-
raphy (eluted with 1:1 EtOAc/hexane followed by 3:1 EtOAc/hexane) to
afford 39 (421 mg, 83%) as a pale-yellow solid. M.p. 738C; [a]2D2 =ꢀ13.6
1
determined by H NMR analysis).
Improved synthesis for the preparation of dimethyl 3-benzyloxycarbonyl-
aminoglutarate (34):
A solution of dimethyl 3-oxoglutarate (20 mL,
136 mmol) and ammonium acetate (36.8g, 476 mmol) in methanol
(400 mL) was stirred over molecular sieves (3 ä pellets, 32 g) for 12 h.
Sodium cyanoborohydride (10.8g, 172 mmol) was added to the resultant
reaction mixture and the solution was acidified to pH 3 by addition of
methanolic HCl (prepared in another run by the addition of 60 mL of
acetyl chloride to 800 mL of methanol at 08C). The reaction mixture was
then filtered, and the filtrate was concentrated under reduced pressure to
afford the crude dimethyl 3-aminoglutarate hydrochloride which was dis-
solved in water (200 mL). The resultant aqueous solution was washed
with diethyl ether and was then basified to pH 10 by the addition of solid
sodium carbonate (31.8g). To this aqueous solution was added diethyl
ether (400 mL) and benzyl chloroformate (19.4 mL, 136 mmol). After
vigorous stirring for 3 h at room temperature, N,N-dimethyl-1,3-propyl-
amine (10.2 mL) was added to quench the excess benzyl chloroformate.
The organic layer was separated and the aqueous layer was extracted
with diethyl ether. The combined organic extracts were washed with 3n
HCl, saturated aqueous NaHCO3, brine, and dried (Na2SO4). Concentra-
tion under reduced pressure gave a residue (31.2 g) which was purified
1
(c=1.01 in CHCl3); H NMR (300 MHz, CDCl3): d=1.70 (brs, 1H), 1.82
(m, 1H), 2.01 (m, 1H), 2.55 (m, 2H), 2.88 (s, 3H), 3.16 3.38 (m, 2H),
3.97 (m, 1H), 4.40 (m, 2H), 5.07 (s, 2H), 6.01 (d, J=7.0 Hz, 1H), 6.22
(brs, 1H), 7.24 7.38(m, 10H), 7.58 7.74 (m, 3H), 7.90 ppm (dd, J = 7.0,
1.5 Hz, 1H); 13C NMR (75 MHz, CDCl3): d=31.7, 34.5, 39.2, 43.4, 46.5,
47.1, 66.5, 124.1, 127.5, 127.8, 127.9, 128.1, 128.5, 128.7, 130.7, 131.7 (2C),
133.7, 136.6, 138.1, 148.3, 156.1, 170.7 ppm; IR (KBr): n˜max =3309, 1700,
1652, 1542, 1373, 1350, 1244, 1165 cmꢀ1; elemental analysis calcd (%) for
C27H26N4O7S: C 58.47, H 5.45, N 10.10; found: C 58.47, H 5.38, N 10.11.
Benzyl-(3S)-3-benzyloxycarbonylamino-5-(N-methyl-bis-tert-butoxycar-
bonylguanidyl)pentanamide (41): Thiophenol (93 mL, 0.91 mmol) was
added to a solution of 39 (421 mg, 0.76 mmol) and cesium carbonate
(750 mg, 2.30 mmol) in acetonitrile (13 mL). The heterogeneous reaction
mixture was then vigorously stirred at room temperature for 2 h. Addi-
3248
¹ 2004 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
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Chem. Eur. J. 2004, 10, 3241 3251

Total Synthesis of (+)-Blasticidin S
3241 3251
tional thiophenol (39 mL, 0.38mmol) was introduced, and after stirring
for a further 2 h, a further portion of thiophenol (39 mL, 0.38mmol) was
added and stirring was continued until TLC analysis showed the absence
of starting material. The resultant reaction mixture was diluted with
CH2Cl2 and then filtered through a pad of Super Cell, and the filter cake
was washed with EtOAc. The filtrate was concentrated to give a residue
(502 mg) which was subsequently dissolved in DMF (15 mL). N,N-Di-
(tert-butoxycarbonyl)-S-methylisothiourea (232 mg, 0.80 mmol) and tri-
ethylamine (0.24 mL, 1.70 mmol) were then added to this solution. The
reaction mixture was cooled to 08C and then treated with mercuric chlo-
ride (228mg, 0.84 mmol). After stirring at 0 8C for 2 h, the mixture was
diluted with EtOAc and then filtered though a pad of Super Cell. The fil-
trate was washed with water and the aqueous layer was extracted with
EtOAc. The combined extracts were washed with 1m aqueous KHSO4,
saturated aqueous NaHCO3, brine, dried (Na2SO4), and concentrated.
The residue was purified by silica-gel column chromatography (1:1 fol-
lowed by 2:1 EtOAc/hexane) to furnish 41 (412 mg, 89%) as a colorless
gum. [a]2D3 =+14.6 (c=0.59 in CHCl3); 1H NMR (300 MHz, CDCl3): d=
1.43 (s, 18H), 1.72 (brs, 1H), 1.79 (m, 1H), 2.01 (m, 1H), 2.49 (dd, J=
14.0, 6.0 Hz, 1H), 2.69 (brdd, J=14.0, 5.0 Hz, 1H), 2.94 (s, 3H), 3.56
(brs, 2H), 3.92 (m, 1H), 4.37 (dd, J=14.0, 6.0 Hz, 1H), 4.45 (dd, J=14.0,
6.0 Hz, 1H), 5.05 (s, 2H), 6.50 (brs, 1H), 6.85 (brs, 1H), 7.20 7.36 (m,
10H), 9.88 ppm (brs, 1H); 13C NMR (100 MHz, CDCl3): d=28.0, 30.7,
36.8, 39.8, 43.3, 47.0, 47.2, 66.3, 79.6, 81.9, 127.2, 127.6, 127.8, 128.2, 128.3,
28.3, 30.0, 35.7, (36.8), 38.9, 45.8, (47.6), 47.9, 51.5, 79.4, 83.7, 147.8,
(147.9), 149.9, 155.6, 158.9, 172.1 ppm; IR (KBr): n˜max =2980, 1802, 1718,
1609, 1286, 1159, 1103 cmꢀ1; elemental analysis calcd (%) for C22H23NO3:
C 55.80, H 8.36, N 9.30; found: C 55.72, H 8.29, N 9.26.
Tetra-Boc-blastidic acid (47): Lithium hydroxide monohydrate (153 mg,
1.22 mmol) was added to a solution of 46 (205 mg, 0.34 mmol) in a mix-
ture of THF (2.6 mL) and water (0.9 mL). After stirring for 3 h at room
temperature, the reaction mixture was poured into cold saturated
aqueous 1m KHSO4 and extracted with EtOAc. The resultant organic
layer was dried (Na2SO4) and concentrated under reduced pressure to
give carboxylic acid 47 (198mg, 97%) as a fine white powder. M.p.
1698C; [a]2D5 =ꢀ4.52 (c=1.17 in CHCl3); 13C NMR (75 MHz, CDCl3):
d=27.8, 27.9, 28.0, 28.2, 30.0, 35.7, (36.8), (37.0), 39.0, 45.7, 47.9, 79.4,
79.9, 83.8, 83.9, 147.77, 147.84, 150.0, 155.8, 158.9, 175.3 ppm; IR (KBr):
n˜max =3448, 1718, 1637, 1283, 1158, 1119 cmꢀ1; elemental analysis calcd
(%) for C27H48N4O10: C 55.09, H 8.22, N 9.52; found: C 55.11, H 8.08,
N 9.50.
Blastidic acid dihydrochloride (51): Trifluoroacetic acid (0.10 mL) was
added to a solution of carboxylic acid 47 (62 mg, 0.11 mmol) in CH2Cl2
(0.5 mL) at room temperature. After stirring at room temperature for
2 h, the reaction mixture was concentrated under reduced pressure. The
residue was quickly dissolved in a few drops of 3n aqueous HCl and then
evaporated under vacuum. This was repeated two more times, then the
resultant residue was dissolved in hot ethanol and filtered through a pad
of Super Cell. A few drops of acetone were added to the filtrate and the
resultant solution was allowed to stand at room temperature to afford
crude 51 as a white solid (19 mg, 69%). The crude solid was dissolved in
a few drops of water, loaded onto a short column of IRA-410 (5 mL, wet
volume), and then eluted with water. The eluent was immediately passed
through a short column of IRC-50 (6 mL, wet volume) and the column
was washed with water (15 mL). Elution with 0.5n HCl afforded blalsti-
dic acid dihydrochloride 51 (14.8mg, 54%), which was further recrystal-
lized from ethanol to furnish a white crystalline solid (10.3 mg, 37%)
[m.p. 190 1958C (decomp)]. [a]2D0 =+13.3 (c=0.52 in H2O); 1H NMR
(300 MHz, D2O): d=2.05 (m, 2H), 2.69 (dd, J=17.5, 7.5 Hz, 1H), 2.82
(dd, J=17.5, 4.5 Hz, 1H), 3.04 (s, 3H), 3.49 (m, 2H), 3.65 ppm (qd, J=
7.0, 4.5 Hz, 1H); 13C NMR (75 MHz, D2O): d=29.9, 36.4, 36.6, 46.7, 47.4,
157.5, 174.6 ppm; IR (KBr): n˜max =3147, 2827, 1717, 1648, 1628, 1488,
128.5, 136.7, 138.3, 150.6, 155.8, 156.1, 161.8, 170.8 ppm; IR (KBr): n˜max
=
3332, 2928, 1700, 1647, 1610, 1541, 1508, 1298 cmꢀ1; elemental analysis
calcd (%) for C47H69N5O11: C 62.83, H 7.41, N 11.45; found: C 62.78, H
7.42, N 11.47.
Benzyl-(3S)-3-benzyloxycarbonyl-tert-butoxycarbonylamino-5-(N-methyl-
tris-tert-butoxycarbonylguanidyl)pentanamide (44): Di-tert-butyldicarbon-
ate (10.90 g, 50 mmol) was added in one portion to a solution of 41
(5.10 g, 8.34 mmol) and 4-dimethylaminopyridine (1.53 g, 12.5 mmol) in
THF (50 mL). After stirring at room temperature overnight, the reaction
mixture was diluted with diethyl ether and then washed with saturated
aqueous NaHCO3 and brine, and dried (Na2SO4). Concentration under
reduced pressure gave a residue which was purified by silica-gel column
chromatography (1:2 EtOAc/hexane) to provide 44 (5.08g, 67%) as a
viscous syrup. [a]2D7 =+3.44 (c=0.39 in CHCl3); 13C NMR (100 MHz,
CDCl3): d=27.81, 27.83, 27.86, 27.9, 28.1, 29.7, 30.8, 35.4, (36.5), (42.2),
42.3, 47.2, (47.8), 48.1, (52.7), 52.8, 68.4, (68.6), 79.2, (79.3), 83.0, (83.1),
83.3, (83.4), 83.5, 147.8, (148.0), 149.3, (150.8), (152.6), 152.8, (154.1),
154.4, (158.9), 159.1, (173.1), 173.3 ppm; IR (KBr): n˜max =2980, 1803,
1414, 1191 cmꢀ1; HRMS (FAB): m/z: calcd for C7H17N4O2 [M+H]+
:
189.1352; found: 189.1337; elemental analysis calcd (%) for
C7H18Cl2N4O2: C 32.19, H 6.95, N 21.45; found: C 32.18, H 7.09, N
21.42.
1735, 1706, 1609, 1456 cmꢀ1
; elemental analysis calcd (%) for
Deprotection of the Troc group in 32: Freshly prepared Cd/Pb (10%,
1.50 g) was added in one portion to a solution of 32 (189 mg, 0.31 mmol)
in a mixture of THF (8.0 mL) and 1m ammonium acetate buffer (2.0 mL,
pH 7). After vigorous stirring at room temperature for 30 min, the reac-
tion mixture was diluted with CH2Cl2 and then filtered through a pad of
Super Cell. The filtrate was washed with saturated aqueous NaHCO3 and
brine, dried (Na2SO4), and concentrated under reduced pressure to give a
residue which was purified by silica-gel chromatography (100:1 CH2Cl2/
MeOH) to afford 53 (109 mg, 93%, calculated based on consumed 32)
and recovered 32 (26 mg, 13%). [a]2D8 =+87.6 (c=1.15 in CHCl3);
1H NMR (300 MHz, CDCl3): d=1.35 (s, 9H), 3.80 (dq, J=9.0, 2.5 Hz,
1H), 3.82 (s, 3H), 4.12 (d, J=9.0 Hz, 1H), 5.79 (ddd, J=10.0, 2.5, 2.0 Hz,
1H), 6.16 (dt, J=10.0, 2.0 Hz, 1H), 6.72 (brs, 1H), 7.53 (d, J=8.5 Hz,
2H), 7.50 7.58(m, 1H), 7.75 (d, J=7.5 Hz, 1H), 7.84 ppm (d, J=8.5 Hz,
1H); 13C NMR (75 MHz, CDCl3): d=30.9, 35.0, 47.1, 52.5, 80.0,
80.1, 97.6, 125.4, 126.0, 127.6, 130.0, 136.1, 145.0, 154.9, 157.2,
C47H69N5O11: C 61.89, H 7.63, N 7.68; found: C 61.95, H 7.69, N 7.65.
Benzyl-(3S)-3-benzyloxycarbonyl-5-(N-methyl-tris-tert-butoxycarbonyl-
guanidyl)pentanamide (45): A solution of 44 (5.10 g, 5.59 mmol) and pal-
ladium hydroxide on carbon (Pearlman×s catalyst, 0.50 g) in ethanol
(150 mL) was stirred vigorously for 5 h. The reaction mixture was filtered
though a pad of Super Cell, the filter cake was washed with diethyl
ether, and the filtrate was washed with saturated aqueous NaHCO3 and
brine, and then dried (Na2SO4), filtered, and concentrated. The re-
sidue was purified by silica-gel column chromatography (1:3 and 1:2
EtOAc/hexane) to afford 45 (3.37 g, 77%) as a white amorphous solid.
M.p. 528C; [a]2D4 =ꢀ4.30 (c=1.09 in CHCl3); 13C NMR (75 MHz, CDCl3):
d=27.6, 27.7, 27.9, 28.1, 28.2, 30.3, (33.4), 35.6, (36.5), 42.8, (46.2),
46.4, 47.0, (47.6), 48.0, 77.2, 78.8, 79.1, (79.2), 83.4, (83.6), (127.0),
(127.1), 127.2, 128.2, (137.8), 138.0, 147.76, 147.80, (148.0), (148.1),
149.6, (150.9), 152.9, (155.4), 155.5, 158.9, (173.6), 173.9 ppm; IR (KBr):
n˜max =3395, 2980, 1735, 1713, 1609, 1285 cmꢀ1; elemental analysis calcd
(%) for C39H63N5O11: C 60.21, H 8.16, N 9.00; found: C 60.22, H 8.21,
N 9.01.
162.7, 169.3 ppm; IR (KBr): n˜max =3421, 2963, 1670, 1488, 1257 cmꢀ1
;
HRMS (FAB): m/z: calcd for C22H27N4O5 [M+H]+ : 427.1981; found:
427.1974.
Methyl (3S)-3-benzyloxycarbonyl-5-(N-methyl-tris-tert-butoxycarbonyl-
guanidyl)pentanoate (46): 1,1,3,3-Tetramethylguanidine (0.15 mL,
1.20 mmol) was added to a solution of 45 (793 mg, 1.02 mmol) in metha-
nol (10 mL). After being stirred at room temperature for 3 h, the reac-
tion mixture was diluted with diethyl ether and washed successively with
water, 1m aqueous KHSO4, saturated aqueous NaHCO3, and brine. The
organic layer was dried (Na2SO4) and concentrated under reduced pres-
sure, and the resultant residue was purified by silica-gel chromatography
Coupling of 53 and 47 using the BOP method in the presence of HOBt:
BOP (132 mg, 0.30 mmol) and HOBt (40 mg, 0.30 mmol) were added to
a solution of 47 (113 mg, 0.19 mmol), 53 (41 mg, 0.096 mmol), and diiso-
propylethylamine (0.15 mL, 0.86 mmol) in CH2Cl2 (5.0 mL). After being
stirred at room temperature for 1.5 h, additional BOP (23 mg,
0.052 mmol), HOBt (8.0 mg, 0.059 mmol), and 47 (23 mg, 0.039 mmol)
were introduced. After a further 5 h, the solution was diluted with satu-
rated aqueous NaCl and then extracted with EtOAc. The combined or-
ganic layers were washed with 1m aqueous KHSO4, saturated aqueous
NaHCO3, and brine, dried (Na2SO4), and then concentrated under re-
(1:1 EtOAc/hexane) to afford 46 (533 mg, 87%) as a colorless oil. [a]D28
=
ꢀ3.08( c=1.37 in CHCl3); 13C NMR (75 MHz, CDCl3): d=27.8, 28.0,
3249
Chem. Eur. J. 2004, 10, 3241 3251
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¹ 2004 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

Y. Ichikawa et al.
FULL PAPER
duced pressure to give a residue which was purified by silica-gel chroma-
tography (100:1 CH2Cl2/MeOH) to afford 54 (70 mg, 73%) as a pale-
yellow powder. M.p. 1468C; [a]3D0 =+52.3 (c=0.39 in CHCl3); 1H NMR
(300 MHz, CDCl3): d=1.35 (s, 9H), 1.42 (s, 9H), 1.46 (s, 9H), 1.49 (s,
9H), 1.51 (s, 9H), 1.71 (brs, 1H), 2.23 (brs, 1H), 2.46 (brs, 1H), 2.81
(brd, 1H), 2.87 (d, 3H), 3.27 (brs, 1H), 3.74 (s, 3H), 3.76 3.93 (m, 2H),
4.43 (d, J=9.0 Hz, 1H), 4.96 (tq, J=9.0, 2.5 Hz, 1H), 5.81 (dt, J=10.0,
2.0 Hz, 1H), 6.11 (brd, J=10.0 Hz, 1H), 6.71 (brs, 1H), 7.52 (d, J=
8.5 Hz, 2H), 7.57 (brs, 1H), 7.71(d, J=8.5 Hz, 1H), 7.80 (d, J=7.5 Hz,
1H), 7.85 ppm (d, J=8.5 Hz, 1H); 13C NMR (75 MHz, CDCl3): d=27.8,
27.9, 28.0, 28.3, 30.9, 34.8, 35.0, 39.6, 44.7, 46.4, 47.1, 52.6, 76.1, 77.2, 79.0,
79.6, 80.7, 84.0, (84.2), 97.7, 125.3, 125.8, 126.0, 127.7, 129.4, 130.0, 133.4,
145.2, 148.0, 150.0, 155.6, 157.2, 158.8, 162.9, 168.4, 171.6 ppm; IR (KBr):
n˜max =3569, 2971, 1698, 1654, 1256, 1116 cmꢀ1; elemental analysis calcd
(%) for C49H72N8O14: C 59.02, H 7.28, N 11.24; found: C 58.92, H 7.24, N
11.17.
[7] M. Kimura, I. Yamaguchi, Pestic. Biochem. Physiol. 1996, 56, 243
248.
≈
[8] a) H. Yonehara, S. Takeuchi, N. Otake, T. Endo, Y. Sakagami, Y.
≈
Sumiki, J. Antibiot. 1963, 16A, 195 222; b) N. Otake, S. Takeuchi, T.
Endo, H. Yonehara, Tetrahedron Lett. 1965, 6, 1405 1409; c) N.
≈
Otake, S. Takeuchi, T. Endo, H. Yonehara, Agric. Biol. Chem. 1966,
≈
30, 126 131; d) N. Otake, S. Takeuchi, T. Endo, H. Yonehara, Agric.
Biol. Chem. 1966, 30, 132 141.
[9] S. Onuma, Y. Nawata, Y. Saito, Bull. Acad. Vet. Fr. 1966, 39, 1091.
[10] Recently, b-amino acids have been actively researched in pharma-
ceutical chemistry. See the references, a) S. H. Gellman, Acc. Chem.
Res. 1998, 31, 173 180; b) D. Seebach, J. L. Matthews, Chem.
Commun. 1997, 2015 2022.
[11] We recognized structural similarities between cytosinine
3 and
peptide nucleic acid (PNA); this led us to propose a new type
of PNA as shown below. Our research efforts into the synthesis
and characterization of this new PNA is now in progress. For
PNA, see the following reference: P. E. Nielsen, Acc. Chem. Res.
1999, 32, 624 630.
Total synthesis of blasticidin S (1): Triethylamine (0.05 mL, 0.36 mmol)
was added to a solution of 54 (23 mg, 0.023 mmol) in MeOH (1.0 mL) at
room temperature, and stirring was continued overnight. Water (0.40 mL,
0.02 mol) was added and the solution was left at room temperature over-
night. The resultant reaction mixture was concentrated under reduced
pressure to afforded the crude carboxylate (13 mg), which was dissolved
in CH2Cl2 (1.0 mL) and then treated with TFA (0.10 mL, 1.3 mmol) at
room temperature. After being stirred at room temperature for 1 h, the
reaction mixture was concentrated under reduced pressure. The residue
was dissolved in 3n aqueous HCl (0.2 mL, 0.2 mmol) and then concen-
trated under vacuum. This was repeated three times. The resultant hydro-
chloride was dissolved in distilled water and then passed through a
column of IRA-410. The effluent that showed UV absorption was collect-
ed and concentrated. The resultant residue was dissolved in ethanol and
then precipitated by addition of diethyl ether to afford white crystals of
blasticidin S (1) (8 mg, 85%) [m.p. 238 2398C (dec.)]. [a]2D7 =+70.5 (c=
0.33 in H2O); 1H NMR (300 MHz, 1n DCl): d=2.08(m, 2H), 2.76 (dd,
J=16.0, 7.0 Hz, 1H), 2.82 (dd, J=16.0, 5.0 Hz, 1H), 3.06 (s, 3H), 3.51
(m, 2H), 3.71 (qn, J=6.0 Hz, 1H), 4.54 (d, J=8.0 Hz, 1H), 4.87 (dq, J=
8.0, 2.5 Hz, 1H), 5.98 (ddd, J=10.0, 2.5, 2.0 Hz, 1H), 6.27 (d, J=8.0 Hz,
1H), 6.28(dt, J=10.0, 2.0 Hz, 1H), 6.60 (q, J=2.0 Hz, 1H), 7.85 ppm (d,
J=8.0 Hz, 1H); 13C NMR (100 MHz, 1n DCl): d=29.9, 36.8, 37.2, 45.2,
47.3, 47.5, 75.5, 80.2, 96.4. 125.3, 134.1, 146.4, 149.1, 157.3, 160.1, 172.0,
[12] a) T. Kondo, H. Nakai, T. Goto, Tetrahedron Lett. 1972, 13, 18 8 1
1884; b) T. Kondo, H. Nakai, T. Goto, Tetrahedron 1973, 29, 1801
1806. For other synthetic studies, see, c) J. J. Fox, K. A. Watanabe,
Pure Appl. Chem. 1971, 43, 475 487; d) R. R. Schmidt, W. Wagner,
Tetrahedron Lett. 1983, 24, 4661 4664.
[13] a) N. Nomoto, A. Shimoyama, Tetrahedron Lett. 2001, 42, 1753
1755; b) Y. Ichikawa, M. Ohbayashi, K. Hirata, M. Isobe, Synlett
2001, 1763 1766.
[14] a) Y. Ichikawa, C. Kobayashi, M. Isobe, Synlett 1994, 919 921; b) Y.
Ichikawa, C. Kobayashi, M. Isobe, J. Chem. Soc. Perkin Trans. 1
1996, 377 382.
172.3 ppm; IR (KBr): n˜max =3221, 1729, 1679, 1655, 1431, 1197 cmꢀ1
;
HRMS (FAB): m/z: calcd for C17H27N8O5 [M+H]+ : 423.2104; found:
[15] For representative examples of the allyl cyanate-to-isocyanate rear-
rangement, see the following references: a) C. Christophersen, A.
Holm, Acta Chem. Scand. 1970, 24, 1512 1526; b) L. E. Overman,
M. Kakimoto, J. Org. Chem. 1978, 43, 4564 4567; c) Y. Ichikawa,
Synlett 1991, 238 240; d) K. Banert, Angew. Chem. 1992, 104, 865
867; Angew. Chem. Int. Ed. Engl. 1992, 31, 866 868.
423.2119.
Acknowledgements
[16] a) Y. Ichikawa, J. Chem. Soc. Perkin Trans. 1 1992, 2135 2139;
b) I. A. O×Neil, In Comprehensive Organic Functional Group Trans-
formations, Vol. 3 (Eds.: A. Katritzky, O. Meth-Cohn, C. W. Ress),
Pergamon, Oxford, 1995, pp. 696.
We would like to thank Kaken Pharmaceutical Company for the gener-
ous gift of blasticidin S. This research was financially supported by a
Grant-In-Aid for Scientific Research from the Ministry of Education, Sci-
ence, Sports and Culture.
[17] The acyl group on the cytosine N4-nitrogen (PG2 in 8) appeared to
be very fragile. For example, gradual cleavage of the tert-butylben-
zoyl group in 32 occurred even in neutral methanol. For other exam-
ples, see: Y. Isido, N. Nakazaki, N. Sakairi, J. Chem. Soc. Perkin
Trans. 1 1977, 657 660.
[18] T. Noshita, T. Sugiyama, Y. Kitazumi, T. Oritani, Tetrahedron Lett.
1994, 35, 8259 8262.
[19] S. Hanessian, P. C. Tyler, Y. Chapleur, Tetrahedron Lett. 1981, 22,
4583 4586. For a useful preparative procedure for the mol-scale
synthesis of 14, see Experimental Section.
[20] Y. Ichikawa, M. Isobe, D.-L. Bai, T. Goto, Tetrahedron 1987, 43,
4737 4748.
[21] S. K. Chaudhary, O. Hernandez, Tetrahedron Lett. 1979, 20, 99 102.
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[23] Although we initially employed 26b as an intermediate, this route
was finally abandoned due to the difficulties associated with depro-
tection of the Cbz group in i. In the case of 26c, separation of the
anomeric mixture, as well as deprotection of ii proved to be serious-
ly difficult.
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3250
¹ 2004 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
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Total Synthesis of (+)-Blasticidin S
3241 3251
[32] Diester 34 was prepared using a modified procedure reported by
Crossley, see Experimental Section and reference: M. J. Crossley,
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[34] For representative examples using secondary amides as advanta-
geous carboxylic acid protecting group, see: D. A. Evans, P. H.
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acetic acid led to the formation of cyclic carbamate i as the major
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[39] L. Grehn, K. Gunnarsson, U. Ragnarsson, Acta Chem. Scand. 1987,
B41, 18 23. Although one-step hydrolysis of 45 using Evans× proto-
col (H2O2, LiOH)[34] provided carboxylic acid 47, the two-step pro-
cedure which involved methanosysis followed by saponification gave
higher and more reliable yields.
[40] An authentic sample of pseudoblastidone (52) was prepared by
treatment of blastidic acid dihydrochloride with base (0.5n NaOH,
pH 10.0). See ref. [8d].
[41] We would like to thank Prof. T. Kondo, Nagoya University, for his
generous gift of blastidic acid dihydrochloride and cytosinine dihy-
drochloride. These were prepared by hydrolysis of natural 1.
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1967, p. 1276.
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5681 5682. The use of freshly prepared Cd/Pb turned out to be es-
sential. Moreover, it should be noted that the yields were time-de-
pendent. Deprotection over an extended period of time led to side
reactions to afford highly polar products which were quite difficult
to isolate. Most probably, the by-products may arise from hydrolysis
of the tert-butylbenzoyl group.
[25] Oxidative removal of the p-methoxyphenyl group in 29 with ceric
ammonium nitrate (CAN) gave the desired product in poor yields.
The silver(ii) reagent which was finally used successfully can also be
used to deprotect PMB ethers as well as PMB amides. Further de-
tails will be reported in due course.
[26] Since we observed gradual decomposition of 31 during purification
on silica-gel chromatography, we did not attempt to separate the a-
and b-isomers. Moreover, this material also slowly decomposed
when stored in a refrigerator at ꢀ208C.
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[28] Upon addition of Lewis acids, such as SnCl4 or BF3¥OEt2, into the
reaction mixture that contained the silylated cytosine, we frequently
observed the formation of a white precipitate. This appeared to be a
s-complex of the silylated base and Lewis acid. For such s-com-
plexes see: a) H. Vorbr¸ggen, G. Hˆfle, Chem. Ber. 1981, 114,
1256 1268; b) H. Vorbr¸ggen, C. Ruh-Pohlenz, Org. React. 2000,
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[30] Cytosine N-glycosylation in the current case might involve a delocal-
ized cation such as i, which is also assumed to be an intermediate in
the Ferrier reaction. In the Ferrier glycosylation using alcohols and
carbon nucleophiles, formation of a-glycosides is usually preferen-
tial. Whilst the a- and b-selectivity of O- and C-Ferrier type glycosy-
lations seems to be governed by a transition state that is stabilized
by anomeric effects, we feel that the transition state for N-glycosyla-
tion in this case is not so strongly stabilized by anomeric effects.
Therefore, the moderate selectivity observed in this reaction (7:3)
might be the result of steric effects. In particular, the bulky silylated
cytosine derivative could approach the probable intermediate i from
the relatively less hindered b-face. We discuss the selectivity of the
Ferrier N-glycosylation based on kinetic control. However, it should
be noted that one of the referees cleverly pointed out an alternative
explanation based on thermodynamics in which an exo-anomeric
effect is operative in the preferred formation of the equatorial N-
glycoside.
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16, 1219 1222.
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Kˆnig, R. Geiger, Chem. Ber. 1970, 103, 2024 2033.
[47] Blasticidin S purified by this method is considered to be a zwitterion
as depicted in i.
[48] Since the salt-free blasticidin S gradually decomposes to give cyto-
mycin 55, NMR spectra of our synthetic sample were immediately
measured in 1n DCl after purification, since this provides stable
blasticidin S hydrochloride.
[49] The overall yield, including the cytosine N-glycosylation step (31!
32), is calculated to be 44% (63% multiplied by 0.7).
[31] a-Substituted pyrrole i was isolated as the major by-product. An au-
thentic sample was also prepared by hydrolysis of 29 using 3n HCl
and subsequent acetylation (Ac2O, pyridine). The mechanism to
form i is unclear.
Received: July 21, 2003
Revised: September 2, 2003
Published online: April 29, 2004
3251
Chem. Eur. J. 2004, 10, 3241 3251
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