Total Synthesis of Nucleoside Antibiotics Plicacetin and Streptcytosine A

Article abstract of DOI:10.1021/acs.joc.8b00006

Disaccharide nucleoside antibiotics plicacetin and streptcytosine A (also named rocheicoside A) were effectively synthesized through the common precursor cytosamine. The amosamine and amicetose moieties were efficiently assembled through an α-selective O-

Full text of DOI:10.1021/acs.joc.8b00006

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Article
Total Synthesis of Nucleoside Antibiotics Plicacetin and Streptcytosine A
Jiqiang Fu, Stephane Laval, and Biao Yu
J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.joc.8b00006 • Publication Date (Web): 13 Feb 2018
Downloaded from http://pubs.acs.org on February 14, 2018
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The Journal of Organic Chemistry
Total Synthesis of Nucleoside Antibiotics Plicacetin and Streptcytosine A
Jiqiang Fu, Stephane Laval, and Biao Yu*
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State Key Laboratory of Bioꢀorganic and Natural Products Chemistry, Center for Excellence in Molecular
Synthesis, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, and University of
Chinese Academy of Sciences, 345 Lingling Road, Shanghai 200032, China
ABSTRACT: Disaccharide nucleoside antibiotics plicacetin and streptcytosine A (also named
rocheicoside A) were effectively synthesized through the common precursor cytosamine. The amosamine
and amicetose moieties were efficiently assembled through an αꢀselective Oꢀglycosylation while the
cytosine nucleus was subsequently introduced through a βꢀselective gold (I)ꢀcatalyzed Nꢀglycosylation.
Further microwaveꢀassisted amidation reactions completed the modular syntheses.
INTRODUCTION
Amicetin (Fig. 1) was isolated in the early 1950s as the first member of the amicetin family
1
a
antibiotics, which are produced in the fermentation broth of various species of Streptomyces,
1ꢀ12
Arthrobacter, and Nocardia.
The structures of amicetin and its simpler congeners, such as plicacetin
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c,d
2ꢀ4
(
1), were elucidated via extensive degradation and transformation studies in the 1960s. In 1981, the
5
structure of amicetin was unambiguously confirmed by Xꢀray diffraction analysis, and in 2007, a
6
detailed NMR structure determination was reported. Other members of the amicetin family antibiotics
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include oxamicetin, norplicacetin, oxyplicacetin, SF2457, and cytosaminomycins AꢀD. More
recently, streptcytosine A (2) was isolated from a culture broth of Streptomyces sp. TPU1236A collected in
1
2a
Okinawa, Japan; this compound was later isolated from Streptomyces rochei 06CM016 collected in the
12b
Mediterranean sea and named rocheicoside A (Figure 1).
These antibiotics share a common
disaccharide pyrimidine nucleoside motif, namely cytosamine, in which amosamine is αꢀ(1→4)ꢀlinked to
1
amicetose, itself βꢀ(1→N )ꢀlinked to a cytosine nucleus. The NH
2
group of the amosamine residue could
7
be monoꢀ or diꢀmethylated, the Cꢀ3 of the amicetose residue could be hydroxylated, and the N on
cytosine could be acylated, thus constituting the molecular diversity of these compounds. As antibiotics,
they demonstrated antibacterial and antiviral activities both in vitro and in vivo, more specifically against
1,6ꢀ11,13
tuberculosis, herpes, and polio.
Amicetin has shown to stabilize polysomes and prevent their
14
breakdown induced by other antibiotics. More importantly, they have proven to inhibit protein
15
biosynthesis of both prokaryotic and eukaryotic ribosomes by binding to the peptidyl transferase centre.
Recently, NMR spectroscopy and unconstrained molecular modelling provided direct evidences for the
specific binding of amicetin to two 35mer ribosomal RNA motifs of Halobacterium halobium and
16
Escherichia coli 23S rRNAs. Nevertheless, further biological studies are hampered by the fact that these
compounds are produced in heterogeneous mixtures and in small quantities, which renders their
accessibility difficult. As a consequence, chemical synthesis of these molecules with a wellꢀdefined
structure and in appreciable amount is of great interest.
To date, several important sugar segments of amicetin antibiotics have been chemically synthesized,
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2,18
i.e., amicetose, amicetose nucleoside, and aminosamine.
In 2001, Sugimura et al. reported the
synthesis of cytosamine (3), which is thought to be a common intermediate of this class of antibiotics, via
a glycosylation between an amosamineꢀtype glycosyl fluoride donor and an amicetose nucleoside
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acceptor. The glycosylation was promoted by 5.0 equiv. of AgOTf/SnCl in Et O/dichloroethane (room
2
2
temperature, 40 h) and provided the desired αꢀglycoside in 52% yield together with the βꢀanomer (11%)
and the remaining acceptor (11%). The total synthesis of plicacetin (1) has been reported by Stevens et al.
20
in 1972. In their synthesis, cytosamine (3) was prepared by coupling an amosamineꢀtype βꢀglycosyl
chloride donor with an amicetose nucleoside acceptor in molten states, under diminished pressure, and in
ꢀ
the presence of Dowex 1ꢀX2 (HO ). The corresponding αꢀlinked disaccharide was obtained in 38% yield
1
d
after extensive purification; subsequent transformations led to cytosamine and finally plicacetin (1).
The paucity of reports indicates the difficulty of chemically synthesizing this class of antibiotics.
Indeed, when designing a retroꢀsynthetic pathway, several problems arise: the construction of αꢀlinkage
between an amosamine donor and an amicetose acceptor, the installation of the cytosine nucleus in a
regioꢀ and βꢀselective manner with an amicetose donor lacking neighboring participation, and the
2
introduction of pendant motifs on the poorly reactive NH of the cytosine nucleus. These considerations
prompted us to investigate the chemical synthesis of the amicetin family antibiotics. Herein we report the
total synthesis of plicacetin (1) and streptcytosine A (rocheicoside A, 2). Their syntheses were achieved
via the common intermediate cytosamine (3), which was effectively constructed by a high βꢀselective
glycosylation between disaccharide oꢀhexynylbenzoate donor 4 and cytosine derivative 5 under
2
1,22
gold(I)ꢀcatalysis (Fig. 1).
Disaccharide 4 was synthesized by coupling amicetose acceptor 8 with
amosamineꢀtype donors (6 and 7), which was installed with pꢀmethoxybenzyl groups (PMB) to facilitate
an αꢀselective glycosylation.
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Figure 1. Plicacetin (1) and streptcytosine A (rocheicoside A, 2) and a retrosynthetic analysis. Cbz = carboxybenzyl, NPTFAI
=
Nꢀphenyltrifluoroacetimidate, PMB = pꢀmethoxybenzyl, TBS = tertꢀbutyldimethylsilyl, TCAI = trichloroacetimidate.
RESULTS AND DISCUSSION
Amosamineꢀtype donors 6 and 7 were synthesized from thiogalactoside 14 (Scheme 1). Compound
14 was prepared from commercially available pentaꢀOꢀacetylꢀβꢀDꢀgalactopyranose (9) via a similar
19
approach as the one reported by Sugimura et al. Thus, βꢀacetate 9 was glycosylated with pꢀtoluenethiol
23
under the promotion of BF ·OEt to provide thiogalactoside 10 (99%). After removal of the acetyl
3
2
groups, the 4,6ꢀhydroxyl groups were selectively protected as benzylidene acetal and the free
2,3ꢀhydroxyl groups were protected with PMB ethters, leading to the fully protected galactoside 11 in
2
4
9
4% overall yield from 10. Cleavage of the benzylidene acetal with TsOH·H
2
O gave the 4,6ꢀdiol 12
2
5
(
84%). The primary 6ꢀOH underwent selective tosylation and subsequent reduction with LiAlH in THF
4
at 60 ºC to afford the 6ꢀdeoxy galactoside 13 in 90% overall yield for two steps. Substitution of the 4ꢀOH
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with azide was accomplished via a 4ꢀOꢀtriflate intermediate, leading to 4,6ꢀdiꢀdeoxyꢀthioglycoside 14
(
78%, two steps). Finally, thioglycoside 14 was smoothly hydrolyzed in the presence of
Nꢀbromosuccinimide in a solvent mixture of acetone/H O/CH Cl at ꢀ20 ºC, and the resulting hemiacetal
was reacted with Cl CCN or CF C(=NPh)Cl under standard basic conditions to yield amosamineꢀtype
2
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3
3
imidate donors 6 and 7, respectively.
1) NaOCH3, CH3OH;
2
) PhCH(OCH3)2
CSA, CH3CN;
TolSH
BF3•Et2O, CH2Cl2
Ph
O
TsOH•H O,
CH2Cl2, CH3OH
AcO OAc
O
AcO
AcO OAc
O
AcO
2
HO OH
O
PMBO
1) TsCl, DMAP, pyridine;
4
O
3) NaH, PMBCl, DMF
2) LiAlH , THF, 60°C
O
STol
OAc
STol
STol
9
9%
94% (3 steps)
PMBO
84%
90% (2 steps)
OPMB
OAc
OAc
10
OPMB
9
11
12
HO
PMBO
1) Tf2O, DMAP, pyridine, CH2Cl2;
) NaN3, DMF, 80°C
2 2 2
1) NBS, acetone, H O, CH Cl ,
-20°C, 99%;
2
O
O
NH
CCl3
O
NPh
O
CF3
O
N3
PMBO
N3
PMBO
N3
PMBO
or
STol
STol
OPMB
14
O
7
8% (2 steps)
2) CCl3CN, DBU, CH2Cl2, 96%; or
CF3C(=NPh)Cl, K2CO3, acetone, 95%
PMBO
PMBO
OPMB
1
3
6
7
Scheme 1. Synthesis of amosamine donors 6 and 7. CSA = camphorsulfonic acid, DBU = 1,8ꢀbiazabicyclo[5.4.0]undecꢀ7ꢀene,
DMAP = 4ꢀdimethylaminopyridine, NBS = Nꢀbromosuccinimide, Tf = trifluoromethanesulfonyl, Tol = tolyl, Ts =
pꢀtoluenesulfonyl.
Amicetose acceptor 8 was obtained in a straightforward manner from commercially available
3
,4,6ꢀtriꢀOꢀacetylꢀ
provided the corresponding hemiacetal, which upon hydrogenation and TBSꢀprotection led to the
,3ꢀdideoxy βꢀglycoside 16 in 76% overall yield for three steps. After removal of the acetyl groups, the
resultant primary alcohol was treated with I /PPh and imidazole in THF to give 6ꢀiodoꢀglycoside 17 in
2% yield (for two steps). Finally, reduction of the iodide was achieved via hydrogenolysis over
2
Dꢀglucal 15 (Scheme 2). Ferrierꢀtype reaction of 15 under a condition of H O at 100 ºC
2
2
3
7
Pd(OH)
2
/C in the presence of DIPEA, giving amicetose acceptor 8 (90%).
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Scheme 2. Synthesis of amicetose acceptor 8. DIPEA = N,Nꢀdiisopropylethylamine.
Amicetose acceptor 8 was subjected to glycosylation with amosamineꢀtype donors 6 and 7 in the
presence of 4Å MS in Et
2
O, a solvent which has been proven to favor the αꢀselective glycosylation (Table
). When acceptor 8 was coupled with donor 6 (2.1 equiv.) at ꢀ40 ºC under the promotion of TMSOTf
0.05 equiv.), disaccharide 18 was isolated in 88% yield and with a high α/β selectivity of 7.5:1 (entry 1).
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1
(
It is noteworthy that, under these reaction conditions, side products, e.g., the Cꢀglycoside derivatized from
28
an intramolecular FriedelꢀCrafts reaction, were formed and isomerization of the anomeric positions
occurred under a prolonged reaction time, thus decreasing the yield of 18 and rendering the purification
difficult. Taking into account these observations, acceptor 8 and donor 7 were glycosylated at ꢀ60 ºC
under the promotion of TBSOTf (0.05 equiv.). Under these conditions, the previous byproducts as well as
isomerization were not detected and disaccharide 18 was isolated in a better yield (92%) and still in an
excellent αꢀselectivity (α/β = 8:1, entry 2). The isomers 18α and 18β could be separated on silica gel and
1
their configurations were confirmed by H NMR analysis. The amosamine anomeric proton of 18β
appeared as a doublet at δ 4.38 ppm with J = 7.6 Hz; the one of 18α appeared downfield at δ 4.79 ppm,
2
however, its J value could not be calculated due to overlapping with the CH signals of the PMB groups
and the anomeric proton of the amicetose moiety.
Table 1. Glycosylation of amicetose acceptor 8 with amosamine donors 6 and 7.
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Donor
Promoter
c
d
Entry
TºC
Yield
α/β
a
b
(
equiv.)
6 (2.1)
7 (1.5)
(equiv.)
1
2
TMSOTf (0.05)
TBSOTf (0.05)
ꢀ40
ꢀ60
88%
92%
7.5:1
8:1
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a
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c
d
Calculated based on acceptor 8. Calculated based on donors 6 and 7. Isolated yield of 18. Ratio of the isolated α and β
isomers. MS = molecular sieves, TMSOTf = trimethylsilyl trifluoromethanesulfonate.
Disaccharide 18α was used to prepare the common intermediate cytosamine (3) (Scheme 3). Thus,
1
8α was desilylated with a solution of TBAF in THF, and oꢀhexynylbenzoic acid was introduced to the
corresponding hemiacetal to afford oꢀhexynylbenzoate donor 4 (84%, 2 steps). Donor 4 was subjected to
29
glycosylation with silylated cytosine 5 under the catalysis of Ph
3 2
PAuNTf (0.2 equiv.). To our delight,
nucleoside 19 was isolated in a good 77% yield with an excellent βꢀselectivity (α/β = 1:10), most likely
3
0
favored by the use of acetonitrile as solvent. The isomers 19α and 19β could be separated on silica gel,
1
and the βꢀconfiguration was confirmed by H NMR analysis, wherein the anomeric proton appeared as a
doublet at δ 5.74 ppm with J = 9.8 Hz. Isomer 19β was then treated with CF
3
COOH in CH
2 2
Cl to provide
2
’’,3’’ꢀdiol 20 (83%). Finally, reduction of the azide under Staudinger conditions, followed by reductive
3 3
dimethylation of the nascent amine with paraformaldehyde and NaBH CN in AcOH/CH OH and
subsequent reductive cleavage of the Cbz group (H
2
/Pd/C) led to cytosamine (3) in 64% yield for three
steps.
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Scheme 3. Synthesis of cytosamine (3). BSTFA
=
N,Oꢀbis(trimethylsilyl)trifluoroacetamide, EDCI
=
1ꢀethylꢀ3ꢀ(3ꢀdimethylaminopropyl)carbodiimide, TBAF = tetraꢀnꢀbutylammonium fluoride.
With cytosamine (3) at hand, the synthesis of plicacetin (1) was achieved in two steps (Scheme 4).
Attempts to couple 3 with 4ꢀnitrobenzoyl chloride failed even at elevated temperature, which can be
rationalized by the poor nucleophilicity of the amino group of the cytosine moiety in 3. Nevertheless, 3
could be coupled with 4ꢀnitrobenzoic acid (21) in the presence of HATU under heating conditions,
although a long reaction time was required. To our delight, the reaction time could be greatly shortened
by using microwave (50 ºC, 5 min). Subsequent reduction of the nitro group using Fe powder and an
aqueous solution of NH Cl in EtOH at 90 ºC furnished plicacetin (1) in 41% yield (for two steps).
4
Scheme 4. Synthesis of plicacetin (1). HATU = 1ꢀ[bis(dimethylamino)methylene]ꢀ1Hꢀ1,2,3ꢀtriazolo[4,5ꢀb]pyridinium 3ꢀoxide
hexafluorophosphate.
To complete the synthesis of streptcytosine A (rocheicoside A, 2), the carboxylic acid segment was
31
prepared (Scheme 5). Compound 22, obtained from Lꢀserine methyl ester hydrochloride, was coupled
3
2
with benzyl 4ꢀaminobenzoate (23) in the presence of HATU and DIPEA in DMF at 80 ºC to afford
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amide 24 (67%). The oxazolidine ring, which showed to be very stable, was opened under forced
conditions employing an aqueous solution of 6
primary alcohol with a TBS group provided the corresponding amidoꢀamine 25 in 75% yield (for two
steps). Reaction of 25 with paraformaldehyde and NaHCO in CH CN at 60 ºC gave the corresponding
N HCl in BnOH at 80 ºC. Subsequent protection of the
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4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
3
3
3
imidazolidinone intermediate, which upon hydrogenolysis over Pd/C in CH OH provided the carboxylic
acid segment 26 (81%, 2 steps). Cytosamine (3) and 26 were successfully coupled together in the
presence of HATU and pyridine at 80 ºC under microwave conditions. The microwave was performed for
5 minutes, 3 times, in order to minimize side reactions. At this stage, the amide bonds remained fragile
under both strong acidic and basic conditions. Nevertheless, HF·pyridine could smoothly remove the TBS
group without affecting the amide bonds, thus providing streptcytosine A (rocheicoside A, 2) in 65% yield
12
(for two steps). Its NMR spectra were in accordance with those reported for the natural product.
NH2
COOH
Boc
O
HATU, DIPEA, DMF, 80°C
7%
O
O
N
+
N
NH
Boc
COOBn
6
t-Bu
t-Bu
COOBn
22
23
24
1
) (CH2O)n, NaHCO3,
CH3CN, 60°C;
O
COOBn
1
2
) HCl 6N, BnOH, 80°C;
) TBSCl, imidazole, DMF
2) H2 (1 atm), Pd/C, CH3OH
N
H
TBSO
NH2
7
5% (2 steps)
81% (2 steps)
O
COOH
3
1
)
, HATU, pyridine, MW, 80°C;
N
2) HF•pyridine, pyridine
HN
TBSO
65% (2 steps)
26
O
N
HO
NH
HO
H
HO
N
O
N
O
N
N
O
O
O
streptcytosine A (rocheicoside A, 2)
Scheme 5. Synthesis of streptcytosine A (rocheicoside A, 2).
CONCLUSION
In conclusion, we reported the total synthesis of plicacetin (1) and the first total synthesis of
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9
1
1
1
1
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1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
streptcytosine A (rocheicoside A, 2), two antibiotics of the amicetin family. Our approach involved: 1) a
αꢀselective Oꢀglycosylation between amosamine imidate donors (6 and 7) and amicetose acceptor 8; 2)
formation of the common intermediate cytosamine (3) via a βꢀselective gold(I)ꢀcatalyzed Nꢀglycosylation
between disaccharide oꢀalkynylbenzoate donor 4 and cytosine (5); 3) amidation between 3 and the
appropriate carboxylic acid segment under microwave conditions. Following this approach, the syntheses
of other members are undergoing in our laboratory. We assume that the access to these biologically active
natural compounds in pure forms and appropriate amount will help scientists to elucidate their
structureꢀactivity relationship as well as to design more potent analogs.
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
EXPERIMENTAL SECTION
General Information. All reactions were carried out under N
flameꢀdried glassware, unless otherwise noted. Dichloromethane (CH
N,Nꢀdimethylformamide (DMF), and methanol (CH OH) were dried over activated 4Å molecule sieves;
triethylamine (NEt ) and pyridine were dried over potassium hydroxide; tetrahydrofuran (THF) and
2
or argon with anhydrous solvents in
2
Cl ), acetonitrile (CH CN),
2
3
3
3
2
diethyl ether (Et O) were distilled over sodium. The chemicals used were reagent grade as supplied,
except where noted. Analytical thinꢀlayer chromatography (TLC) was conducted with silica gel 60 F254
preꢀcoated glass plates (0.25 mm) and visualized by exposure to UV light (254 nm) or by staining with
sulfuric acid, ceric ammonium molybdate, or potassium permanganate (KMnO
4
) ethanolic solutions.
1
Silica gel (particle size 0.037−0.048 mm) was used for flash column chromatography. H NMR spectra
1
were recorded at 400, 500, or 600 MHz and are reported relative to deuterated solvent signals. Data for H
NMR spectra are reported as follows: chemical shift (δ ppm), multiplicity, coupling constant (Hz), and
13
13
integration. C NMR spectra were recorded at 100, 125, or 150 MHz. Data for C NMR spectra are
1
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reported in terms of chemical shift. NMR spectra were referenced using (CH
3
)
4
Si (0 ppm), residual
1
13
1
13
CDCl
3
( H NMR δ = 7.26 ppm, C NMR δ = 77.16 ppm), CD
3
OD ( H NMR δ = 3.31 ppm, C NMR δ =
1
13
4
9.00 ppm), and DMSOꢀd6 ( H NMR δ = 2.50 ppm, C NMR δ = 39.52 ppm). High resolution mass
spectra were recorded on ESIꢀTOF spectrometers. Optical rotations were measured on a polarimeter using
either CHCl or CH OH as solvents.
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
3
3
Commercially available compounds 9, 15, and 21 were purchased, while compounds 5, 22, and 23 were
29,31,32
synthesized according to literature procedures.
Tolyl 2,3,4,6ꢀtetraꢀOꢀacetylꢀ1ꢀthioꢀβꢀ
9) (20.0 g, 51.2 mmol) and 4ꢀtolyl mercaptan (TolSH) (7.00 g, 56.32 mmol) in CH
added BF ·OEt (16.14 mL, 128.00 mmol) at 0 ºC. The mixture was stirred at room temperature for 6 h
and then quenched at 0 ºC by addition of saturated aqueous NaHCO . The organic layer was separated,
dried over Na SO , filtered, and concentrated under reduced pressure. The residue was purified by silica
Dꢀgalactopyranoside (10). To a solution of
D
ꢀgalactose pentaacetate
(
2
Cl (200 mL) was
2
3
2
3
2
4
gel column chromatography (petroleum ether/EtOAc, 1:1) to provide 10 as a colorless liquid (23.0 g,
28
1
9
9%): [α]
7.7 Hz, 2H), 5.40 (d, J = 2.1 Hz, 1H), 5.21 (t, J = 9.9 Hz, 1H), 5.03 (dd, J = 9.8, 2.8 Hz, 1H), 4.64 (d, J =
.9 Hz, 1H), 4.18 (dd, J = 11.1, 7.0 Hz, 1H), 4.10 (dd, J = 11.1, 6.4 Hz, 1H), 3.90 (t, J = 6.4 Hz, 1H), 2.34
D
3 3
= +16.5 (c 0.6, CHCl ); H NMR (500 MHz, CDCl ) δ 7.41 (d, J = 7.8 Hz, 2H), 7.12 (d, J =
9
1
3
(s, 3H), 2.11 (s, 3H), 2.09 (s, 3H), 2.04 (s, 3H), 1.97 (s, 3H); C NMR (125 MHz, CDCl
3
) δ 170.5, 170.3,
170.2, 169.6, 138.6, 133.3, 129.8, 128.8, 87.1, 74.5, 72.2, 67.4, 67.4, 61.7, 21.3, 21.0, 20.8, 20.8, 20.7;
+
30 9 4
HRMS (ESI) calcd for C21H O NS [M+NH ] 472.1636, found 472.1632.
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1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
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4
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6
Tolyl 2,3ꢀdiꢀOꢀpꢀmethoxybenzylꢀ4,6ꢀOꢀbenzylideneꢀ1ꢀthioꢀβꢀ
D
ꢀgalactopyranoside (11). To a solution of 10
(
39.16 g, 86.16 mmol) in CH
3
OH (300 mL) was added NaOCH
3
(465 mg, 8.64 mmol). The mixture was
+
stirred for about 1 h at room temperature and then neutralized with DOWEX 50W (H ꢀform). After
removal of the resin by filtration, concentration of the filtrate afforded the corresponding 2,3,4,6ꢀtetraol,
which was used for the next step without further purification.
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
The crude tetraol was treated with benzaldehyde dimethylacetal (19.40 mL, 129.24 mmol) and
camphorsulfonic acid (1.00 g, 4.31 mmol) in CH CN (600 mL) overnight at room temperature. After
3
addition of NEt
3
, the solvent was removed under reduced pressure. The residue was dissolved in CH
2 2
Cl
and washed with saturated aqueous NaHCO
3
. The organic layer was separated, dried over Na SO , and
2
4
filtered. The filtrate was partially evaporated until apparition of a solid. Petroleum ether was added to the
mixture, which was then stored at ꢀ20 ºC for 30 min. The solid was finally filtered, washed with
petroleum ether, and used for the next step.
The aforementioned solid was dissolved in DMF (400 mL), and NaH (60% in mineral oil, 9.61 g,
2
40.35 mmol) was added at 0 ºC. After 30 min of stirring at 0 ºC, pꢀmethoxybenzyl chloride (27.04 mL,
200.29 mmol) was added to the reaction mixture, and stirring was continued overnight at room
temperature. The reaction was quenched by addition of CH OH. The mixture was neutralized with
3
saturated aqueous NH Cl and then extracted with CH Cl . The organic layer was washed with saturated
4
2
2
aqueous NaCl solution, dried over Na
2
SO
4
, filtered, and concentrated under reduced pressure. The
Cl /petroleum ether) to provide 11 as a light yellow
); H NMR (500 MHz, CDCl ) δ 7.65 (d, J = 7.9 Hz, 2H),
.56 (d, J = 3.8 Hz, 2H), 7.44–7.37 (m, 5H), 7.30 (d, J = 8.4 Hz, 2H), 7.04 (d, J = 7.8 Hz, 2H), 6.92 (d, J
8.4 Hz, 2H), 6.86 (d, J = 8.4 Hz, 2H), 5.50 (s, 1H), 4.69 (s, 4H), 4.58 (d, J = 9.5 Hz, 1H), 4.37 (d, J =
resulting residue was purified by crystallization (CH
2
2
2
D
8
1
solid (50 g, 94%): [α]
= ꢀ22.5 (c 1.0, CHCl
3
3
7
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2.2 Hz, 1H), 4.13 (d, J = 2.8 Hz, 1H), 3.98 (d, J = 11.9 Hz, 1H), 3.98ꢀ3.92 (m, 4H), 3.81 (s, 3H), 3.61
13
(dd, J = 9.2, 3.1 Hz, 1H), 3.38 (s, 1H), 2.33 (s, 3H); C NMR (125 MHz, CDCl
3
) δ 159.3, 159.3, 138.0,
1
7
6
37.6, 133.4, 130.9, 130.3, 129.8, 129.7, 129.5, 129.0, 128.9, 128.1, 126.7, 113.8, 113.8, 101.3, 86.7, 81.1,
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
5.2, 75.1, 73.8, 71.5, 69.8, 69.5, 55.3, 55.3, 21.2; HRMS (ESI) calcd for C H NaO S [M+Na]
36 38
7
37.2230, found 637.2236.
Tolyl 2,3ꢀdiꢀOꢀpꢀmethoxybenzylꢀ1ꢀthioꢀβꢀ
added to a solution of compound 11 (16.40 g, 26.68 mmol) in CH
room temperature. After stirring for 2 h, the mixture was neutralized by addition of NEt
concentrated under reduced pressure. The resulting residue was dissolved in CH Cl , washed with
saturated aqueous NaHCO , dried over Na SO , filtered, and concentrated under reduced pressure. The
residue was purified by silica gel column chromatography (petroleum ether/EtOAc, 4:1 to 1:1) to provide
Dꢀgalactopyranoside (12). TsOH·H O (3.55 g, 18.67 mmol) was
2
2
Cl
2
(90 mL) and CH
3
OH (90 mL) at
and then
3
2
2
3
2
4
28
D
1
12 as a white foam (11.77 g, 84%): [α]
= +4.6 (c 0.9, CHCl
3 3
); H NMR (500 MHz, CDCl ) δ 7.47 (d, J
=
7.7 Hz, 2H), 7.35 (d, J = 8.1 Hz, 2H), 7.27 (d, J = 8.0 Hz, 2H), 7.11 (d, J = 7.6 Hz, 2H), 6.94ꢀ6.84 (m,
4H), 4.76 (d, J = 9.9 Hz, 1H), 4.67 (d, J = 10.0 Hz, 1H), 4.64 (s, 2H), 4.57 (d, J = 9.7 Hz, 1H), 4.02 (s,
H), 3.98–3.90 (m, 1H), 3.82 (s, 3H), 3.81 (s, 3H), 3.69 (t, J = 9.2 Hz, 1H), 3.60–3.51 (m, 1H), 3.45 (s,
1
1
3
1H), 2.72 (s, 1H), 2.42 (d, J = 4.3 Hz, 1H), 2.33 (s, 3H); C NMR (125 MHz, CDCl ) δ 159.6, 159.4,
3
137.8, 132.5, 130.5, 123.0, 129.9, 129.8, 129.7, 114.1, 113.9, 88.0, 82.2, 78.1, 76.8, 75.4, 72.0, 67.4, 62.8,
+
55.4, 55.4, 21.2; HRMS (ESI) calcd for C29
H
34NaO
7
S [M+Na] 549.1917, found 549.1919.
Tolyl 2,3ꢀdiꢀOꢀpꢀmethoxybenzylꢀ6ꢀdeoxyꢀ1ꢀthioꢀβꢀ
Dꢀgalactopyranoside (13). Tosyl chloride (6.39 g, 33.52
mmol) was added to a solution of 12 (11.77 g, 22.35 mmol) and DMAP (273 mg, 2.23 mmol) in pyridine
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2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
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4
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5
5
5
6
(
150 mL) at 0
C. After evaporation of the solvents under reduced pressure, the crude residue was diluted with EtOAc
and washed with saturated aqueous CuSO . The organic layer was dried over Na SO , filtered, and
2
̊C . The mixture was stirred at room temperature overnight and then quenched with H O at
0
̊
4
2
4
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
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1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
concentrated under reduced pressure. The resulting residue was purified by silica gel column
chromatography (petroleum ether/EtOAc, 4:1 to 2:1) to afford the corresponding 6ꢀOꢀTs derivative (14.79
g, 97%).
LiAlH (1.23 g, 32.50 mmol) was added to a solution of the aforementioned 6ꢀOꢀTs galactoside
4
(14.75 g, 21.66 mmol) in dry THF (100mL) at 0
reaction was quenched by careful addition of EtOAc and saturated aqueous potassium sodium tartrate
Seignette’s salt) at 0 C. After filtration through Celite®, THF was removed under reduced pressure, and
the resulting mixture was extracted with CH Cl several times. The organic layers were combined, dried
over Na SO , filtered, and concentrated under reduced pressure. The resulting residue was purified by
silica gel column chromatography (petroleum ether/EtOAc, 4:1 to 2:1) to yield 13 as a colorless liquid
̊C , and the mixture was stirred at 60 ºC for 5 h. The
(
̊
2
2
2
4
28
1
(10.34 g, 93%): [α]D = +5.2 (c 0.5, CHCl ); H NMR (500 MHz, CDCl ) δ 7.47 (d, J = 8.1 Hz, 2H),
3
3
7.34 (d, J = 8.6 Hz, 2H), 7.26 (d, J = 8.6 Hz, 2H), 7.10 (d, J = 8.0 Hz, 2H), 6.91–6.84 (m, 4H), 4.75 (d, J
9.9 Hz, 1H), 4.65 (dd, J = 9.0, 5.3 Hz, 1H), 4.63 (s, 2H), 4.51 (d, J = 9.7 Hz, 1H), 3.81 (s, 3H), 3.80 (s,
3H), 3.77 (d, J = 2.6 Hz, 1H), 3.61 (t, J = 9.3 Hz, 1H), 3.52 (dt, J = 13.3, 4.9 Hz, 2H), 2.32 (s, 3H), 1.35
=
13
3
(d, J = 6.5 Hz, 3H); C NMR (125 MHz, CDCl ) δ 159.6, 159.5, 137.7, 132.8, 130.7, 130.2, 130.0, 130.0,
129.7, 129.7, 114.1, 113.9, 88.0, 82.8, 76.7, 75.4, 74.3, 72.0, 69.6, 55.5, 55.4, 21.3, 16.9. HRMS (ESI)
+
calcd for C29
H
34NaO
6
S [M+Na] 533.1968, found 533.1971.
Tolyl 2,3ꢀdiꢀOꢀpꢀmethoxybenzylꢀ4ꢀazidoꢀ4,6ꢀdideoxyꢀ1ꢀthioꢀβꢀDꢀglucopyranoside (14). DMAP (2.47 g,
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2
.02 mmol) and Tf
CH Cl
temperature overnight and was finally quenched by addition of H
the crude compound was dissolved in EtOAc and washed with saturated aqueous CuSO . The organic
2
O (6.84 mL, 40.50 mmol) were added to a solution of 13 (10.34 g, 20.25 mmol) in
2
2
(80 mL) and pyridine (16 mL, 202.49 mmol) at 0 C
̊
. The mixture was allowed to stir at room
2
O at 0 ºC. After removal of solvents,
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
4
2 4
layer was then dried over Na SO , filtered, and concentrated under reduced pressure. The resulting
residue was purified by silica gel column chromatography (petroleum ether/EtOAc, 6:1) to furnish the
corresponding 4ꢀOTf galactoside as a colorless liquid (10.60 g, 81%).
Sodium azide (1.28 g, 19.79 mmol) was added to a solution of the aforementioned 4ꢀOTf galactoside
(
10.60 g, 16.49 mmol) in DMF (90 mL), and the mixture was stirred at 80 ºC. After 2.5 h, DMF was
evaporated under reduced pressure, and the resulting residue was diluted with CH Cl and washed with
saturated aqueous NaCl. The organic layer was dried over Na SO , filtered, and concentrated under
reduced pressure. The resulting residue was purified by silica gel column chromatography (petroleum
2
2
2
4
25
D
1
ether/EtOAc, 6:1) to afford 14 as a white solid (8.47 g, 96%): [α]
3
= +48.7 (c 0.9, CHCl ); H NMR
(500 MHz, CDCl ) δ 7.44 (d, J = 8.1 Hz, 2H), 7.37 (d, J = 8.6 Hz, 2H), 7.30 (d, J = 8.6 Hz, 2H), 7.12 (d,
3
J = 8.0 Hz, 2H), 6.93–6.87 (m, 4H), 4.87 (d, J = 10.0 Hz, 1H), 4.83 (d, J = 10.3 Hz, 1H), 4.77 (d, J = 10.2
Hz, 1H), 4.68 (d, J = 10.0 Hz, 1H), 4.54 (d, J = 9.5 Hz, 1H), 3.82 (s, 3H), 3.81 (s, 3H), 3.50 (t, J = 8.9 Hz,
1H), 3.44 (t, J = 9.1 Hz, 1H), 3.21 (dt, J = 12.0, 6.0 Hz, 1H), 3.14 (t, J = 9.5 Hz, 1H), 2.35 (s, 3H), 1.37 (d,
13
3
J = 6.0 Hz, 3H); C NMR (125 MHz, CDCl ) δ 159.6, 138.1, 132.8, 130.3, 130.0, 129.9, 129.8, 129.7,
114.02, 88.0, 84.6, 80.8, 75.5, 75.2, 74.9, 67.9, 55.4, 55.4, 21.3, 18.9; HRMS (ESI) calcd for
+
C
29
H
33
N
3
NaO
5
S [M+Na] 558.2033, found 558.2036.
2,3ꢀDiꢀOꢀpꢀmethoxybenzylꢀ4ꢀazidoꢀ4,6ꢀdideoxyꢀα/βꢀ
Dꢀglucopyranosyl
trichloroacetimidate
(6).
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NꢀBromosuccinimide (5.94 g, 33.38 mmol) was added to a mixture of 14 (5.96 g, 11.12 mmol) in acetone
(
50 mL), H
was added to quench the reaction. The two layers were separated and the aqueous one was extracted with
CH Cl . The organic layers were combined, dried over Na SO , filtered, and concentrated under reduced
2 2 2 2 2 3
O (5 mL), and CH Cl (20 mL) at ꢀ20 ºC. After stirring for 2 h, a saturated aqueous Na S O
0
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pressure. The resulting residue was purified by silica gel column chromatography (petroleum
ether/EtOAc, 8:1 to 2:1) to afford the corresponding hemiacetal as a white liquid (4.97 g, 99%).
CCl CN (1.45 mL, 14.55 mmol) and DBU (87 ꢁL, 0.58 mmol) were added to a solution of the
3
aforementioned hemiacetal (1.25 g, 2.91 mmol) in CH
h, the mixture was concentrated under reduced pressure. The residue was purified by a quick silica gel
column chromatography (petroleum ether/EtOAc with 1% NEt , 8:1) to afford imidate 6 as a light yellow
2 2
Cl (5 mL). After stirring at room temperature for
1
3
liquid (1.62 g, 96%). Because of stability reason, donor 6 was used directly for glycosylation without
characterization.
2,3ꢀDiꢀOꢀpꢀmethoxybenzylꢀ4ꢀazidoꢀ4,6ꢀdideoxyꢀα/βꢀ
Dꢀglucopyranosyl Nꢀphenyltrifluoroacetimidate (7).
CF (C=NPh)Cl (445 ꢁL, 4.05 mmol) and K CO
3
2
3
(1.12 g, 8.10 mmol) were added to a solution of the
aforementioned hemiacetal (870 mg, 2.03 mmol) in acetone (10 mL), and the mixture was stirred
vigorously at room temperature for 1 h. The mixture was filtered through a pad of Celite® and the filtrate
was concentrated under reduced pressure. The resulting residue was purified by silica gel column
chromatography (petroleum ether/EtOAc with 1% NEt
3
, 8:1) to yield imidate 7 as a yellow liquid (1.16 g,
95%). Because of stability reason, donor 7 was used directly for glycosylation without characterization.
tertꢀButyldimethylsilyl 2,3ꢀdideoxyꢀ4,6ꢀdiꢀOꢀacetylꢀβꢀ
Dꢀglucopyranoside (16). 3,4,6ꢀTriacetylglucal (15)
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(
2 2
10 g, 36.72 mmol) was suspended in H O (200 mL) and refluxed at 100 ºC until 15 was consumed. H O
was removed under reduced pressure to provide the corresponding Ferrierꢀtype hemiacetal, which was
used for the next step without further purification.
1
1
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10% Pd/C (1.50 g) was added to a solution of the aforementioned hemiacetal in EtOAc (200 mL),
and the suspension was stirred under H atmosphere. After 5 h, Pd/C was removed by filtration. The
2
filtrate was concentrated under reduced pressure to provide the corresponding lactol as a viscous liquid,
which was used for the next step without further purification.
TBSCl (7.60 g, 50.44 mmol) and imidazole (4.66g, 68.46 mmol) were added to a solution of the
aforementioned crude lactol in CH
After 4 h, the mixture was washed with saturated aqueous NaHCO
Na SO , filtered, and concentrated under reduced pressure. The residue was purified by silica gel column
chromatography (petroleum ether/EtOAc, 16:1 to 8:1) to afford 16 as a colorless liquid (9.67 g, 76%):
2
Cl
2
(200 mL) at 0 ºC. The mixture was stirred at room temperature.
3
. The organic layer was dried over
2
4
2
D
8
1
[
α]
= +6.75 (c 1.0, CHCl
3 3
); H NMR (500 MHz, CDCl ) δ 4.77 (dd, J = 7.8, 0.6 Hz, 1H), 4.61 (td, J =
9
.9, 4.8 Hz, 1H), 4.15–4.10 (m, 2H), 3.65–3.59 (m, 1H), 2.19–2.12 (m, 1H), 2.02 (s, 3H), 2.00 (s, 3H),
1.85–1.79 (m, 1H), 1.60 (tdd, J = 12.9, 8.8, 3.8 Hz, 1H), 1.54–1.42 (m, 1H), 0.86 (s, 9H), 0.08 (s, 3H),
13
0
.07 (s, 3H); C NMR (125 MHz, CDCl
3
) δ 170.9, 170.1, 96.7, 75.1, 67.8, 63.7, 32.3, 27.2, 25.9, 21.2,
+
20.9, 18.2, ꢀ4.1, ꢀ5.0; HRMS (ESI) calcd for C H NaO Si [M+Na] 369.1704, found 369.1707.
16
30
6
tertꢀButyldimethylsilyl 2,3,6ꢀtrideoxyꢀ6ꢀiodoꢀβꢀ
3
Dꢀglucopyranoside (17). NaOCH (151 mg, 27.91 mmol)
was added to a solution of 16 (9.67 g, 27.91 mmol) in CH
3
OH (50 mL) and the mixture was stirred for 4
+
h at room temperature. The mixture was neutralized with DOWEX 50W (H ꢀform). The resin was then
filtered off. The filtrate was concentrated under reduced pressure to provide the corresponding 4,6ꢀdiol,
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which was used for the next step without further purification.
PPh
3
(18.30 g, 69.77 mmol) and imidazole (9.5 g, 139.55 mmol) were added to a solution of the
(11.33 g, 44.65 mmol) in THF (20 mL) was
aforementioned 4,6ꢀdiol in THF (100 mL). A solution of I
2
0
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9
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8
9
0
added slowly to the mixture at 0 ºC. After 30 min of stirring, another portion of I (3.54 g, 13.95 mmol) in
2
THF (5 mL) was added and the reaction mixture was stirred at room temperature for 2 h. The reaction
was quenched by addition of saturated aqueous NaS
2 3 3
O and saturated aqueous NaHCO at 0 ºC. The THF
was removed under reduced pressure and the resulting aqueous mixture was extracted with CH Cl twice.
2
2
The organic layers were combined, dried over Na
2 4
SO , filtered, and concentrated under reduced pressure.
The residue was purified by silica gel column chromatography (petroleum ether/EtOAc, 6:1) to provide
2
D
2
1
17 as a colorless liquid (7.72 g, 72%): [α]
3 3
= +16.4 (c 1.0, CHCl ); H NMR (400 MHz, CDCl ) δ 4.80
(dd, J = 8.9, 2.1 Hz, 1H), 3.59 (dd, J = 10.1, 2.0 Hz, 1H), 3.41 (dd, J = 11.1, 7.1 Hz, 1H), 3.28–3.16 (m,
2H), 2.08–2.01 (m, 1H), 1.84 (ddd, J = 12.1, 5.6, 3.0 Hz, 1H), 1.67–1.46 (m, 3H), 0.90 (s, 9H), 0.19 (s,
1
3
3
3
H), 0.16 (s, 3H); C NMR (100 MHz, CDCl ) δ 97.1, 79.7, 70.2, 33.3, 31.4, 26.0, 18.2, 6.8, ꢀ3.6, ꢀ5.0;
+
HRMS (ESI) calcd for C H INaO Si [M+Na] 395.0510, found 395.0511.
1
2
25
3
2
tertꢀButyldimethylsilyl 2,3,6ꢀtrideoxyꢀβꢀDꢀglucopyranoside (8). 20% Pd(OH) /C (662 mg) and DIPEA
(9.31 mL, 53.34 mmol) were added to a solution of 17 in CH OH (40 mL), and the mixture was stirred
3
under H
2 2
atmosphere. After 8h, Pd(OH) /C was filtered off and the filtrate was concentrated under
reduced pressure. The resulting residue was purified by silica gel column chromatography (petroleum
2
D
2
1
ether/EtOAc, 6:1) to yield 8 as a colorless liquid (3.94 g, 90%): [α]
500 MHz, CDCl ) δ 4.73 (dd, J = 9.1, 2.1 Hz, 1H), 3.29–3.23 (m, 2H), 2.05–1.99 (m, 1H), 1.85–1.79 (m,
H), 1.62–1.55 (m, 1H), 1.49–1.40 (m, 1H), 1.28 (d, J = 5.7 Hz, 3H), 0.89 (s, 9H), 0.11 (s, 3H), 0.10 (s,
3
= ꢀ7.3 (c 1.0, CHCl ); H NMR
(
3
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H); C NMR (126 MHz, CDCl
3
) δ 96.7, 76.1, 71.6, 33.5, 31.4, 26.0, 18.3 (2C), ꢀ4.0, ꢀ5.0. HRMS (ESI)
+
3
calcd for C12H26NaO Si [M+Na] 269.1543, found 269.1545.
1
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tertꢀButyldimethylsilyl
2’,3’ꢀDiꢀOꢀpꢀmethoxybenzylꢀ4’ꢀazidoꢀ4’,6’ꢀdideoxyꢀαꢀDꢀglucopyranosyl)ꢀ(1’→4)ꢀ2,3,6ꢀtrideoxyꢀβꢀDꢀglu
(
copyranoside (18α). (6+8, Table 1, entry 1): Freshly activated 4Å molecular sieves (2.00 g) was added to
a solution of 6 (1.61 g, 2.80 mmol) and 8 (332 mg, 1.35 mmol) in Et O (10 mL). The mixture was cooled
2
to ꢀ40 ºC and stirred at this temperature for 50 min. Then, TMSOTf (25 ꢁL, 0.14 mmol) was added and
3
the mixture was stirred at ꢀ40 ºC. After 30 min, the reaction was quenched by addition of NEt . The
mixture was filtered through a pad of Celite® and the filtrate was concentrated under reduced pressure.
The residue was purified by silica gel column chromatography (petroleum ether/EtOAc, 60:1 to 16:1) to
afford 18α (690 mg, 78%) and 18β (92 mg, 10%) as colorless liquids.
(7+8, Table 1, entry 2): Freshly activated 4Å molecular sieves (300 mg) was added to a solution of 7
(183 mg, 0.30 mmol) and 8 (50 mg, 0.20 mmol) in Et O (2 mL). The mixture was cooled to ꢀ60 ºC and
2
stirred at this temperature for 40 min. Then, TBSOTf (4 ꢁL, 0.015 mmol) was added and the mixture was
stirred at ꢀ60 ºC. After 25 min, the reaction was quenched by addition of NEt . The mixture was filtered
3
through a pad of Celite® and the filtrate was concentrated under reduced pressure. The residue was
purified by silica gel column chromatography (petroleum ether/EtOAc, 60:1 to 16:1) to afford 18α (108
mg, 82%) and 18β (13 mg, 10%) as colorless liquids.
2
D
7
1
1
8α: [α]
3 3
= +104.2 (c 1.0, CHCl ); H NMR (500 MHz, CDCl ) δ 7.32 (d, J = 8.5 Hz, 2H), 7.26 (d,
J = 8.5 Hz, 2H), 6.92–6.82 (m, 4H), 4.86 (d, J = 10.0 Hz, 1H), 4.82–4.76 (m, 2H), 4.73 (d, J = 10.1 Hz,
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H), 4.68 (d, J = 11.6 Hz, 1H), 4.56 (d, J = 11.6 Hz, 1H), 3.79 (s, 6H), 3.74 (t, J = 9.5 Hz, 1H), 3.61–3.53
(m, 2H), 3.49 (dd, J = 9.6, 3.6 Hz, 1H), 3.27 (td, J = 10.2, 4.4 Hz, 1H), 3.05 (t, J = 9.8 Hz, 1H), 2.11–2.04
(m, 1H), 1.91–1.84 (m, 1H), 1.54 (tdd, J = 13.0, 9.3, 3.8 Hz, 1H), 1.46–1.36 (m, 1H), 1.28 (d, J = 6.1 Hz,
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
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6
7
8
9
0
1
2
3
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5
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7
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9
0
1
3
3
1
H), 1.23 (d, J = 6.2 Hz, 3H), 0.91 (s, 9H), 0.13 (s, 3H), 0.12 (s, 3H); C NMR (125 MHz, CDCl ) δ
3
59.6, 159.5, 130.4, 130.1, 130.0, 129.9, 114.0, 113.9, 110.1, 96.6, 93.0, 79.5, 75.4, 74.2, 74.1, 73.0, 68.1,
+
66.5, 55.4, 33.0, 26.5, 25.9, 19.0, 18.4, 18.2, ꢀ4.0, ꢀ5.0; HRMS (ESI) calcd for C34
80.3338, found 680.3339.
51 3 8
H N NaO Si [M+Na]
6
(2’,3’ꢀDiꢀOꢀpꢀmethoxybenzylꢀ4’ꢀazidoꢀ4’,6’ꢀdideoxyꢀαꢀ
Dꢀglucopyranosyl)ꢀ(1’→4)ꢀ2,3,6ꢀtrideoxyꢀα/βꢀ
Dꢀg
lucopyranoside oꢀ(hexꢀ1ꢀynyl)benzoate (4). TBAF (1
M
in THF, 2.60 mL, 2.60 mmol) was added to a
solution of 18α (1.14 g, 1.73 mmol) in THF (20 mL), and the mixture was stirred at room temperature.
After 3 h, THF was evaporated under reduced pressure and the resulting residue was purified by silica gel
column chromatography (petroleum ether/EtOAc, 2:1) to provide the corresponding hemiacetal (894 mg,
95%) as a colorless liquid.
2 2
The aforementioned hemiacetal (690 mg, 1.27 mmol) was dissolved in CH Cl (10 mL) and DIPEA
(
1.64 mL, 12.70 mmol), oꢀhexynylbenzoic acid (473 mg, 2.54 mmol), EDCI (730 mg, 3.81 mmol), and
DMAP (1.64 mL, 12.70 mmol) were added. The resulting mixture was stirred overnight at room
temperature and then quenched by addition of saturated aqueous NaHCO . The two layers were separated
and the aqueous one was extracted with CH Cl . The organic layers were combined, dried over Na SO
3
2
2
2
4
,
filtered, and concentrated under reduced pressure. The residue was purified by silica gel column
chromatography (petroleum ether/EtOAc, 60:1 to 16:1) to yield 4 as a light yellow liquid (814 mg, 88%,
β/α = 4:1). As the major isomer, a portion of 4β could be separated from the mixture.
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4
β: [α]
D
= +92.9 (c 1.0, CHCl
3
); H NMR (500 MHz, CDCl
3
) δ 7.94 (dd, J = 7.9, 1.1 Hz, 1H), 7.51
(
dd, J = 7.7, 0.9 Hz, 1H), 7.43 (td, J = 7.6, 1.3 Hz, 1H), 7.35–7.31 (m, 2H), 7.30–7.25 (m, 3H), 6.91–6.85
m, 4H), 5.99 (dd, J = 9.2, 2.3 Hz, 1H), 4.87 (d, J = 10.0 Hz, 1H), 4.80 (d, J = 3.6 Hz, 1H), 4.76–4.69 (m,
(
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
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2
3
4
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8
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2
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4
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8
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0
2
H), 4.57 (d, J = 11.6 Hz, 1H), 3.81 (s, 3H), 3.81 (s, 3H), 3.78 (dd, J = 5.3, 3.2 Hz, 1H), 3.76 (t, J = 7.5
Hz, 1H), 3.59 (dq, J = 12.4, 6.2 Hz, 1H), 3.51 (dd, J = 9.6, 3.6 Hz, 1H), 3.37–3.31 (m, 1H), 3.07 (t, J =
9.8 Hz, 1H), 2.48 (t, J = 7.1 Hz, 2H), 2.21–2.16 (m, 1H), 2.15–2.10 (m, 1H), 1.81–1.73 (m, 1H), 1.61 (tdd,
J = 14.2, 9.9, 5.5 Hz, 4H), 1.54–1.46 (m, 2H), 1.32 (d, J = 6.2 Hz, 3H), 1.25 (d, J = 6.2 Hz, 3H), 0.96 (t, J
13
= 7.3 Hz, 3H); C NMR (125 MHz, CDCl
3
) δ 164.5, 159.7, 159.5, 134.6, 132.0, 131.1, 130.6, 130.4,
1
6
30.1, 130.1, 129.9, 127.2, 125.2, 114.1, 114.0, 96.7, 94.6, 93.5, 79.6, 79.5, 79.3, 75.5, 75.0, 74.0, 73.2,
49 3 9
8.1, 66.7, 55.4, 55.4, 30.9, 28.9, 25.6, 22.3, 19.7, 19.1, 18.5, 13.8; HRMS (ESI) calcd for C41H N NaO
+
[
M+Na] 750.3361, found 750.3354.
1
ꢀ[2’’,3’’ꢀDiꢀOꢀpꢀmethoxybenzylꢀ4’’ꢀazidoꢀ4’’,6’’ꢀdideoxyꢀαꢀD
ꢀglucopyranosyl)ꢀ(1’’→4’)ꢀ2’,3’,6’ꢀtrideoxy
4
ꢀβꢀ
D
ꢀglucopyranosyl]ꢀN ꢀcarboxybenzylcytosine (19β). Freshly activated 5Å molecular sieves (800 mg)
3
was added to a solution of donor 4 (α/β mixture, 460 mg, 0.63 mmol) in CH CN (4 mL) and the resulting
mixture was stirred at room temperature for 40 min. In a separate flask, BSTFA (1.03 mL, 3.79 mmol)
was added to a suspension of NHCbzꢀcytosine 5 (465mg, 1.90 mmol) in CH CN (4 mL) and the mixture
3
was heated at 40 ºC until it became clear. The solution of the silylated cytosine was cooled to room
temperature and added to the mixture of 4. Then, a solution of Ph
CH CN (0.20 mL) was added, followed by the addition of BSTFA (0.50 mL, 1.90 mmol). The resulting
mixture was stirred at room temperature for 1 day. Then, another portion of Ph PAuNTf (46 mg, 0.063
mmol) was added and stirring was continued for 2 days (until complete consumption of donor 4). The
3 2
PAuNTf (46 mg, 0.063 mmol) in
3
3
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mixture was filtered through a pad of Celite® and the filtrate was concentrated under reduced pressure.
The residue was purified by silica gel column chromatography (petroleum ether/AcOEt, 8:1 to 1:2) to
28
D
provide 19β as a white foam (339 mg, 70%) and 19α as a white gel (35 mg, 7%). 19β: [α]
= +160.6 (c
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6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
1
.0, CHCl ); H NMR (500 MHz, CDCl ) δ 7.86 (d, J = 7.4 Hz, 1H), 7.41–7.37 (m, 5H), 7.35–7.31 (m,
3
3
2H), 7.29–7.21 (m, 3H), 6.94–6.84 (m, 4H), 5.74 (d, J = 9.8 Hz, 1H), 5.22 (s, 2H), 4.87 (d, J = 10.1 Hz,
1H), 4.79 (d, J = 3.6 Hz, 1H), 4.75–4.70 (m, 2H), 4.55 (d, J = 11.5 Hz, 1H), 3.81 (s, 6H), 3.80–3.77 (m,
H), 3.75 (t, J = 9.4 Hz, 1H), 3.61–3.55 (m, 1H), 3.52 (dd, J = 9.6, 3.7 Hz, 1H), 3.31 (td, J = 10.5, 4.3 Hz,
1H), 3.08 (t, J = 9.8 Hz, 1H), 2.40–2.30 (m, 1H), 2.22–2.13 (m, 1H), 1.62 (qd, J = 13.3, 3.8 Hz, 1H),
1
1
3
1
1
8
3
.43–1.35 (m, 2H), 1.34 (d, J = 6.1 Hz, 3H), 1.27 (d, J = 6.2 Hz, 3H); C NMR (125 MHz, CDCl ) δ
62.2, 159.7, 159.5, 154.4, 152.3, 144.0, 135.0, 130.3, 130.1, 129.9, 128.8, 128.4, 114.1, 114.0, 95.0, 93.1,
3.3, 79.6, 79.4, 77.1, 75.5, 73.8, 73.3, 68.1, 67.9, 66.7, 55.4, 55.4, 30.7, 26.6, 18.9, 18.5; HRMS (ESI)
+
calcd for C40
H
46
N
6
NaO10 [M+Na] 793.3168, found 793.3166.
4
1
ꢀ[4’’ꢀAzidoꢀ4’’,6’’ꢀdideoxyꢀαꢀ
boxybenzylcytosine (20). CF COOH (0.60 mL) was added to a solution of 19β (425 mg, 0.55 mmol) in
CH Cl (12 mL) at 0º C and the mixture was stirred at room temperature for about 30 min. Then, the
reaction was quenched by addition of saturated aqueous NaHCO at 0 ºC and extracted with CH Cl . The
Dꢀglucopyranosyl)ꢀ(1’’→4’)ꢀ2’,3’,6’ꢀtrideoxyꢀβꢀDꢀglucopyranosyl]ꢀN ꢀcar
3
2
2
3
2
2
organic layers were combined, dried over Na
2
SO
4
, filtered, and concentrated under reduced pressure. The
Cl /CH OH, 20:1) to provide 20 as a
OH); H NMR (500 MHz, CD OD) δ 8.05 (d, J =
residue was purified by silica gel column chromatography (CH
2
2
3
28
D
1
white foam (244 mg, 83%): [α]
= +172.5 (c 1.1, CH
3
3
7
.6 Hz, 1H), 7.40 (d, J = 7.1 Hz, 2H), 7.36 (t, J = 7.2 Hz, 2H), 7.33–7.30 (m, 2H), 5.72 (d, J = 7.9 Hz,
H), 5.21 (s, 2H), 4.96 (d, J = 3.7 Hz, 1H), 3.73–3.69 (m, 1H), 3.67 (t, J = 9.5 Hz, 1H), 3.61 (dt, J = 12.4,
1
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.2 Hz, 1H), 3.47 (dd, J = 9.7, 3.8 Hz, 1H), 3.37 (td, J = 9.5, 4.4 Hz, 1H), 2.99 (t, J = 9.8 Hz, 1H),
.40–2.28 (m, 1H), 2.17–2.04 (m, 1H), 1.70–1.55 (m, 2H), 1.31 (d, J = 6.1 Hz, 3H), 1.27 (d, J = 6.2 Hz,
13
H); C NMR (125 MHz, CD
4.4, 78.2, 75.6, 73.6, 73.2, 69.9, 68.6, 67.9, 30.9, 27.8, 19.1, 18.6; HRMS (ESI) calcd for C H N NaO
8
3
OD) δ 164.6, 156.8, 154.4, 145.8, 136.9, 129.5, 129.4, 129.2, 97.1, 96.5,
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
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4
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7
8
9
0
1
2
3
4
5
6
7
8
9
0
24 30
6
+
[
M+Na] 553.2017, found 553.2018.
Cytosamine (3). (CH ) P (1
M in THF, 0.92 mL, 0.92 mmol) and NEt (0.32 mL, 2.30 mmol) were added
3
3
3
2
to a solution of 20 (244 mg, 0.46 mmol) in THF (5 mL)/H O (1 mL) and the mixture was stirred at 50 ºC.
After 2 h, the mixture was concentrated under reduced pressure to give the crude 4’’ꢀNH
2
derivative,
which was used for the next step without further purification.
CH
.38 mmol) were mixed with the aforementioned amine. The resulting mixture was stirred overnight at
room temperature. The reaction was then quenched with saturated aqueous NaHCO . The two layers were
separated and the aqueous one was extracted with CH Cl . The organic layers were combined, dried over
3 2 n 3
OH (5 mL), (CH O) (69 mg, 2.30 mmol), NaBH CN (144 mg, 2.30 mmol), and AcOH (79 ꢁL,
1
3
2
2
Na
chromatography (CH
light yellow liquid (244 mg, 99%).
0% Pd/C (40 mg) was added to a solution of the aforementioned 4’’ꢀN(CH
0.19 mmol) in CH OH (4 mL). The mixture was stirred under H atmosphere for 8 h at room temperature.
Then, Pd/C was filtered off and the filtrate was concentrated under reduced pressure. The residue was
purified by RPꢀ18 column chromatography (H O/CH OH, 1:2) to afford 3 as a white solid (50 mg, 64%):
OH); H NMR (500 MHz, CD OD) δ 7.68 (d, J = 7.5 Hz, 1H), 5.90 (d, J = 7.5
2
SO
4
, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column
2
Cl /CH OH, 20:1 to 15:1) to provide the corresponding 4’’ꢀN(CH derivative as a
2
3
3 2
)
1
3 2
) derivative (104 mg,
3
2
2
3
2
D
6
1
[
α]
= +159.5 (c 1.0, CH
3
3
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2
2
2
2
2
2
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5
5
5
5
5
5
6
Hz, 1H), 5.72 (dd, J = 10.5, 2.0 Hz, 1H), 4.90 (d, J = 3.8 Hz, 1H), 3.87 (t, J = 9.8 Hz, 1H), 3.81 (dt, J =
12.2, 6.1 Hz, 1H), 3.69 (dq, J = 12.2, 6.1 Hz, 1H), 3.41 (dd, J = 9.5, 3.8 Hz, 1H), 3.38–3.32 (m, 1H), 2.46
(s, 6H), 2.38–2.32 (m, 1H), 2.13 (t, J = 10.1 Hz, 1H), 2.05–1.97 (m, 1H), 1.68 (ddd, J = 15.5, 13.2, 3.1 Hz,
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
3
1
H), 1.64–1.54 (m, 1H), 1.32 (d, J = 6.1 Hz, 3H), 1.23 (d, J = 6.2 Hz, 3H); C NMR (125 MHz, CD OD)
3
δ 167.5, 157.7, 142.5, 96.2, 96.1, 83.8, 78.2, 75.1, 74.6, 71.8, 70.2, 67.3, 42.4, 30.7, 27.9, 19.7, 19.2;
+
HRMS (ESI) [M+H] calcd for C18
H
31
N
4
O
6
399.2238, found 399.2244.
Plicacetin (1). Compound 3 (11 mg, 0.028 mmol) was mixed with pꢀnitrobenzoic acid (21) (7 mg, 0.041
mmol), HATU (19 mg, 0.050 mmol), and pyridine (0.50 mL), and the resulting mixture in a sealed flask
was heated under microwave at 50 ºC (monitored with an external surface sensor) for 5 min. Then,
pyridine was evaporated under reduced pressure. The residue was purified by silica gel column
chromatography (CH
2
2 3
Cl /CH OH, 40:1) to afford the corresponding amide.
A solution of NH
4
Cl (0.74 mg, 0.014 mmol) in H O (0.20 mL) and Fe powder (8 mg, 0.14 mmol)
2
were added to a solution of the aforementioned amide in EtOH (2 mL). The mixture was heated at 90 ºC
for 30 min, and was then cooled down to room temperature and filtered through a pad of Celite®. The
filtrate was concentrated under reduced pressure. The residue was purified by RPꢀ18 column
chromatography (H O/CH OH + 0.05% CF COOH, 6:4) to afford 1 as a colorless liquid (6 mg, 41%):
2
3
3
2
D
1
1
[
α]
3 3
= +77.0 (c 0.15, CH OH); H NMR (400 MHz, CD OD) δ 8.15 (d, J = 7.6 Hz, 1H), 7.77 (d, J = 8.8
Hz, 2H), 7.50 (d, J = 7.4 Hz, 1H), 6.71 (d, J = 8.8 Hz, 2H), 5.77 (d, J = 7.9 Hz, 1H), 5.01 (d, J = 3.8 Hz,
H), 4.09 (dt, J = 12.0, 6.3 Hz, 1H), 3.95 (dd, J = 10.2, 9.4 Hz, 1H), 3.75 (dq, J = 12.2, 6.1 Hz, 1H), 3.54
dd, J = 9.2, 3.9 Hz, 1H), 3.48–3.40 (m, 1H), 3.11 (t, J = 10.2 Hz, 1H), 3.00 (s, 6H), 2.44–2.33 (m, 1H),
1
(
13
2
.21–2.12 (m, 1H), 1.79–1.61 (m, 2H), 1.46 (d, J = 6.2 Hz, 3H), 1.37 (d, J = 6.1 Hz, 3H); C NMR (100
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MHz, CD
3
OD) δ 168.7, 164.6, 156.3, 154.8, 146.3, 131.5, 121.1, 114.8, 98.5, 96.7, 84.6, 78.2, 76.6, 73.9,
+
72.0, 67.9, 64.0, 45.4, 30.9, 28.0, 19.2, 19.0; HRMS (ESI) calcd for C25
H
35
N
5
NaO
7
[M+Na] 540.2429,
found 540.2430.
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
(
2R,4S)ꢀ2ꢀtertꢀButylꢀ3ꢀtertꢀbutoxycarbonylꢀ4ꢀmethylꢀ4ꢀ[(pꢀ(benzyloxycarbonyl)phenyl)carbamoyl]ꢀoxazol
idine (24). Benzyl 4ꢀaminobenzoate (23) (1.42 g, 6.26 mmol), HATU (2.54 g, 6.68 mmol), and DIPEA
3.65 mL, 20.90 mmol) were added to a solution of 22 (1.20 g, 4.18 mmol) in DMF (15 mL) and the
reaction mixture was stirred at 80 ºC overnight. DMF was evaporated under reduced pressure, the
resulting residue was dissolved in CH Cl , and washed successively with aqueous HCl 1 and saturated
aqueous NaHCO . The organic layer was dried over Na SO , filtered, and concentrated under reduced
pressure. The residue was purified by silica gel column chromatography (petroleum ether/EtOAc, 8:1) to
(
2
2
N
3
2
4
2
D
2
1
afford 24 as a yellow solid (1.40 mg, 67%): [α]
3 3
= ꢀ104.2 (c 1.1, CHCl ); H NMR (500 MHz, CDCl ) δ
8
.06 (d, J = 8.7 Hz, 1H), 7.62 (d, J = 8.7 Hz, 1H), 7.46–7.42 (m, 2H), 7.41–7.36 (m, 2H), 7.35–7.30 (m,
H), 5.35 (s, 2H), 5.21 (s, 1H), 4.80 (d, J = 7.7 Hz, 1H), 3.81 (d, J = 9.0 Hz, 1H), 1.72 (s, 2H), 1.53 (s,
1
13
9H), 0.91 (s, 9H); C NMR (125 MHz, CDCl
3
) δ 171.7, 166.1, 155.9, 142.7, 136.3, 131.1, 128.7, 128.3,
128.2, 125.6, 118.9, 98.1, 83.0, 76.2, 68.2, 66.7, 38.3, 28.3, 26.3, 22.2; HRMS (ESI) calcd for
+
C H N NaO [M+Na] 519.2466, found 519.2460.
28
36
2
6
(S)ꢀBenzyl 4ꢀ(2ꢀaminoꢀ2ꢀmethylꢀ3ꢀ(tertꢀbutyldimethylsilyloxy)ꢀpropanamido)benzoate (25). Benzyl
alcohol (1 mL) was added to a suspension of 24 (137 mg, 0.28 mmol) in aqueous HCl (6 , 1 mL), and
N
the mixture was heated at 80 ºC until 24 was totally consumed. The mixture was then extracted with
EtOAc. The organic layer was discarded, and the aqueous one was concentrated under reduced pressure
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2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
to provide the corresponding aminoꢀalcohol intermediate as the hydrochloride salt, which was used for
the next step without further purification.
TBSCl (151 mg, 0.55 mmol) and imidazole (75 mg, 1.10 mmol) were added to a solution of the
aforementioned aminoꢀalcohol in DMF (2 mL). The mixture was stirred overnight at room temperature.
Then, DMF was evaporated under reduced pressure and the resulting residue was purified by silica gel
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
column chromatography (petroleum ether/EtOAc + 1% NEt
3
, 4:1) to afford 25 as a yellow liquid (93 mg,
22
1
7
5%): [α]D = ꢀ48.9 (c 1.0, CHCl ); H NMR (500 MHz, CDCl ) δ 10.05 (s, 1H), 8.04 (d, J = 8.7 Hz, 2H),
3
3
7.67 (d, J = 8.8 Hz, 2H), 7.46–7.43 (m, 2H), 7.41–7.37 (m, 2H), 7.36–7.31 (m, 1H), 5.35 (s, 2H), 4.10 (d,
13
J = 9.5 Hz, 1H), 3.39 (d, J = 9.5 Hz, 1H), 1.32 (s, 3H), 0.85 (s, 9H), 0.07 (s, 3H), 0.03 (s, 3H); C NMR
(125 MHz, CDCl
3
) δ 174.6, 166.2, 142.4, 136.3, 131.1, 128.7, 128.3, 128.3, 125.3, 118.6, 69.8, 66.7, 59.7,
+
2
5.9, 23.3, 18.3, ꢀ5.3; HRMS (ESI) calcd for C24
H
35
N
2
O
4
Si [M+H] 443.2361, found 443.2359.
(S)ꢀ4ꢀ[4ꢀ((tertꢀButyldimethylsilyloxy)methyl)ꢀ4ꢀmethylꢀ5ꢀoxoimidazolidinꢀ1ꢀyl]benzoic
acid
(26).
Compound 25 (300 mg, 0.68 mmol) was mixed with (CH O) (20 mg, 0.68 mmol), solid NaHCO (86 mg,
2
n
3
3
0.68 mmol), and CH CN (4 mL). The mixture was heated at 60 ºC overnight. The salts were filtered off
and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel
column chromatography (petroleum ether/EtOAc + 1% NEt , 4:1) to afford the corresponding
3
imidazolidinone intermediate as a colorless liquid (93 mg, 86%).
10% Pd/C (26 mg) was added to a solution of the aforementioned imidazolidinone intermediate (265
3 2
mg, 0.58 mmol) in CH OH (5 mL). The resulting suspension was stirred under H atmosphere overnight
at room temperature. After complete consumption of the imidazolidinone intermediate, Pd/C was filtered
off. The filtrate was concentrated under reduced pressure to afford 26 as a white solid (198 mg, 94%):
2
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[
α]
D
3 3
= ꢀ44.75 (c 1.1, CHCl ); H NMR (500 MHz, CDCl ) δ 8.09 (d, J = 8.8 Hz, 2H), 7.69 (d, J = 8.9
Hz, 2H), 4.89 (q, J = 7.7 Hz, 1H), 4.02 (d, J = 10.2 Hz, 1H), 3.58 (d, J = 10.2 Hz, 1H), 1.26 (s, 3H), 0.81
13
(
s, 9H), 0.06 (s, 3H), 0.03 (s, 3H); C NMR (125 MHz, CDCl
3
) δ 175.3, 170.4, 142.2, 131.3, 125.8,
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
17.8, 77.4, 77.2, 76.9, 66.9, 65.8, 62.4, 25.8, 18.4, 18.2, ꢀ5.4, ꢀ5.4; HRMS (ESI) calcd for C H N O Si
18 29
2
4
+
[
M+H] 365.1891, found 365.1888.
Streptcytosine A (rocheicoside A, 2). Cytosamine 3 (9 mg, 0.022 mmol) was mixed with acid 26 (16 mg,
0.045 mmol), HATU (19 mg, 0.050 mmol), and pyridine (0.40 mL). The resulting mixture in a sealed
flask was heated under microwave at 80 ºC (monitored with an external surface sensor) for 5 min. The
heating conditions were repeated for two additional times. After that, pyridine was evaporated under
reduced pressure. The resulting residue was purified by Sephadex LHꢀ20 (CH
the corresponding amide intermediate.
2 2 3
Cl /CH OH, 1:1) to afford
HF·pyridine (70% HF, 20 ꢁL) was added to a solution of the aforementioned amide in pyridine at 0
ºC. The mixture was stirred at room temperature overnight, pyridine was then evaporated under reduced
pressure. The resulting residue was purified by RPꢀ18 column chromatography (H
2
O/CH
3
OH, 1:2) to
21
D
1
afford 2 as a colorless liquid (8 mg, 65%): [α]
= +83.0 (c 0.2, CH OH); H NMR (600 MHz,
3
DMSOꢀd6) δ 8.17 (d, J = 6.5 Hz, 1H), 8.04 (d, J = 8.9 Hz, 2H), 7.77 (d, J = 8.9 Hz, 2H), 7.36 (d, J = 6.5
Hz, 1H), 5.71 (dd, J = 10.7, 1.5 Hz, 1H), 4.80 (dd, J = 12.2, 5.6 Hz, 2H), 4.71 (d, J = 7.5 Hz, 1H), 3.75 (t,
J = 9.7 Hz, 1H), 3.66–3.60 (m, 3H), 3.32 (d, J = 10.9 Hz, 1H), 3.30–3.23 (m, 2H), 2.37 (s, 9H), 1.24 (d, J
13
=
6.1 Hz, 3H), 1.15 (d, J = 6.1 Hz, 3H), 1.09 (s, 3H); C NMR (150 MHz, DMSOꢀd6) δ 176.7, 166.6,
1
6
63.3, 154.3, 145.7, 142.2, 129.8, 127.9, 117.5, 96.8, 94.8, 82.4, 76.6, 73.3, 73.0, 70.2, 68.3, 65.9, 65.8,
+
5.3, 62.8, 42.3, 29.3, 26.7, 19.3, 19.0, 18.7; HRMS (ESI) calcd for C30
H
43
N
6
O
9
[M+H] 631.3086,
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9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
found 631.3093.
ASSOCIATED CONTENT
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Supporting Information
13
Comparison of the C NMR data of streptcytosine A (rocheicoside A, 2) with those reported for the
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13
natural product, and the H and C NMR spectra of all new compounds (PDF).
AUTHOR INFORMATION
Corresponding Authors
*Eꢀmail: byu@mail.sioc.ac.cn
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
Financial support from the National Natural Science Foundation of China (21432012 & 21621002),
the Strategic Priority Research Program of the Chinese Academy of Sciences (XDB20020000), and the
K .C. Wong Education Foundation is acknowledged.
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