Stereoselective synthesis of the hexasaccharide repeating unit of the cell wall polysaccharide from kineosporia aurantiaca VKM Ac-720T employing the direct glycosylation with anomeric hydroxy sugars involving glycosyl phthalate intermediates

Article abstract of DOI:10.1080/07328300903003352

Synthesis of a suitably protected form of the hexasaccharide repeating unit of the cell wall polymer from Kineosporia aurantiaca VKM Ac-720T has been achieved by the stereoselective direct glycosylation of a trisaccharide acceptor with a trisaccharide don

Full text of DOI:10.1080/07328300903003352

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Stereoselective Synthesis of the
Hexasaccharide Repeating Unit of
the Cell Wall Polysaccharide from
Kineosporia aurantiaca VKM Ac-720T
Employing the Direct Glycosylation with
Anomeric Hydroxy Sugars Involving
Glycosyl Phthalate Intermediates
So Mi Park a , Dae-Hwan Suk a & Kwan Soo Kim a
a Center for Bioactive Molecular Hybrids and Department of
Chemistry, Yonsei University, Seoul, 120-749, Korea
Published online: 24 Jul 2009.
To cite this article: So Mi Park , Dae-Hwan Suk & Kwan Soo Kim (2009): Stereoselective Synthesis
of the Hexasaccharide Repeating Unit of the Cell Wall Polysaccharide from Kineosporia aurantiaca
VKM Ac-720T Employing the Direct Glycosylation with Anomeric Hydroxy Sugars Involving Glycosyl
Phthalate Intermediates, Journal of Carbohydrate Chemistry, 28:6, 317-329
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Journal of Carbohydrate Chemistry, 28:317–329, 2009
ꢀC
Copyright Taylor & Francis Group, LLC
ISSN: 0732-8303 print / 1532-2327 online
DOI: 10.1080/07328300903003352
Stereoselective Synthesis of the
Hexasaccharide Repeating
Unit of the Cell Wall
Polysaccharide from
Kineosporia aurantiaca VKM
Ac-720T Employing the Direct
Glycosylation with Anomeric
Hydroxy Sugars Involving
Glycosyl Phthalate
Intermediates
So Mi Park, Dae-Hwan Suk, and Kwan Soo Kim
Center for Bioactive Molecular Hybrids and Department of Chemistry, Yonsei Univer-
sity, Seoul 120-749, Korea
Synthesis of a suitably protected form of the hexasaccharide repeating unit of the
cell wall polymer from Kineosporia aurantiaca VKM Ac-720T has been achieved by
the stereoselective direct glycosylation of a trisaccharide acceptor with a trisaccharide
donor having an anomeric hydroxy group involving a glycosyl phthalate intermediate.
Both the trisaccharide acceptor and the trisaccharide donor were obtained from a com-
mon trisaccharide, of which two β-mannopyranosyl linkages were constructed stere-
oselectively by employing the direct glycosylation method with the anomeric hydroxy
sugar involving a glycosyl phthalate intermediate and the 2ꢁ-carboxybenzyl glycoside
method, respectively.
Keywords Oligosaccharides; Glycosylation; 2ꢁ-Carboxybenzyl (CB) glycosides; Glyco-
syl phthalates; β-Mannosylation
Received March 11, 2009; accepted April 28, 2009.
Address correspondence to Kwan Soo Kim, Department of Chemistry, Yon-
sei University, 134 Sinchon-dong, Seodaemoon-gu, Seoul 120-749, Korea. E-mail:
kwan@yonsei.ac.kr
317

S.M. Park et al.
318
INTRODUCTION
Kineosporia, which is an aerobic gram-positive bacterium, was isolated from
soil and classified as a genus of the order Actinomycetales.[1] Although it was
originally proposed that the genus Kineosporia was constituted by a single
species Kineosporia aurantiaca,[1] further studies amended the description of
the genus[2] and added new species in the genus.[3] Extensive structural studies
of the cell wall of Actinomycetales have been performed because the structure of
cell wall polysaccharides and teichoic acids was known to be used as taxonomic
makers or fingerprints of the actinomycete species.[4] Recently, Tul’skaya and
coworkers have reported that the major cell wall polymer of K. aurantiaca
VKM Ac-720T is a neutral polysaccharide, a galactomannan with a previously
unknown structure.[5] This particular galactomannan is composed of a series
of the repeating unit of a hexasaccharide with the following structure:→
4)-β-D-Manp-(1→4)-β-D-Manp-(1→3)-β-D-Galp-(1→6)-β-D-Manp-(1→4)-β-D-
Manp-(1→3)-β-D-Galp-(1→(A), as shown in Figure 1. This is quite unusual
because the major component of the cell wall polymer of Actinomycetales is
not usually neutral polysaccharides but rather anionic polysaccharides.[4]
An interesting structural feature of the hexasaccharide repeating unit of
this galactomannan is the occurrence of the β-D-mannopyranosyl linkage, of
which the stereospecific formation still poses a challenge. Several diverse and
innovative strategies for the β-D-mannopyranosylation have been developed,[6]
and recently Crich and coworkers have made a significant breakthrough in
the β-mannoside synthesis employing 4,6-O-benzylidene-protected mannopy-
ranosyl sulfoxides or thiomannopyranosides as glycosyl donors.[7] We have
also reported successful 4,6-O-benzylidene-directed β-mannopyranosylations
with our new glycosyl donors, anomeric hydroxy sugars involving glycosyl
phthalate intermediates[8] and 2ꢁ-carboxybenzyl (CB) glycosides.[9]
Herein, as a part of our works on the synthesis of oligosaccharides orig-
inated from the cell wall of microorganisms[10] in the effort to achieve syn-
thetically challengeable goals and/or to study the biological activity related to
them, we present the stereoselective synthesis of hexasaccharide 1 as shown in
Figure 1: The repeating unit of the cell wall polysaccharide from K. aurantiaca VKM Ac-720T.

Synthesis of Kineosporia aurantiaca Polysaccharide
319
Figure 2: Retrosynthesis of hexasaccharide 1.
Figure 2, a suitably protected form of the hexasaccharide repeating unit A
of the cell wall polymer from K. aurantiaca VKM Ac-720T employing our
newly developed glycosylation methods such as the direct glycosylation with
anomeric hydroxy sugars involving glycosyl phthalate intermediates[8] and the
CB glycoside method.[9]
RESULTS AND DISCUSSION
Protective groups of the target hexasaccharide 1 were selected with careful de-
liberation for the future synthesis of a dodecasaccharide or an octadecasaccha-
ride by dimerization or trimerization of 1. Thus, the 4,6-O-benzylidene group
in the nonreducing end of 1 would be selectively cleaved to give a hexasaccha-
ride acceptor with a hydroxy group at C-4. The methoxyphenyl (MP) group at

S.M. Park et al.
320
C-1 in the reducing end of 1 would be readily converted into a hexasaccharide
having an anomeric hydroxy group as a donor. Retrosynthesis of the hexas-
accharide 1 led to trisaccharide donor 2 and acceptor 3, which both would be
obtained from same trisaccharide 4 as shown in Figure 2. This common in-
termediate 4 was further analyzed into monosaccharide building blocks 5, 6,
and 7.
Our synthesis started with preparation of the monosaccharide build-
ing blocks 5, 6, and 7. Thus, CB 4,6-O-benzylidenemannoside 5,[9] 4,6-O-
benzylidenemannose 6,[11] and p-methoxyphenyl galactoside 7[12] were syn-
thesized according to previously reported procedures. Coupling of 6 and 7
was performed by the stereoselective β-mannopyranosylation of the accep-
tor 7 with the 4,6-O-benzyliden-protected anomeric hydroxy sugar 6 employ-
ing our new direct glycosylation method involving the glycosyl phthalate
intermediate.[8] Thus, treatment of the mannose 6 with phthalic anhydride and
1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) in methylene chloride at rt provided
a mannosyl phthalate intermediate. Without isolation of the mannosyl ph-
thalate intermediate, triflic anhydride and 2,6-di-tert-butyl-4-methylpyridine
(DTBMP) were added to the above solution at −78◦C. Then, addition of the
acceptor alcohol 7 to this solution provided exclusively β-mannosyl disaccha-
ride 8 in 74% yield after chromatographic separation as shown in Scheme
1. The LC-mass analysis indicated that the β/α ratio of the crude product 8
was 30:1. Anomeric carbon chemical shifts at δ 101.6 and 101.1 ppm and one-
bond coupling constants (JC1−H1) of 165 and 162 Hz of the product 8 indicated
that two glycosyl linkages including the newly generated mannosyl linkage,
Scheme 1.

Synthesis of Kineosporia aurantiaca Polysaccharide
321
D-Manp-(1→3)-D-Galp, of the disaccharide 8 are both in β-configurations.[13]
Hydrolysis of the 4,6-O-benzylidene group of the disaccharide 8 with trifluo-
roacetic acid (TFA) followed by selective benzoylation of the primary hydroxy
group in resulting disaccharide diol 9 with benzoyl chloride at 0◦C afforded
6-O-benzoyl disaccharide acceptor 10 having a free hydroxy group at C-4ꢁꢁ.
The β-mannosylation of the disaccharide acceptor 10 to make the manno-
syl trisaccharide 4 was preformed with CB 4,6-O-benzylidenemannoside 5 as
the mannosyl donor. Thus, the coupling of the CB glycosyl donor 5 and the
glycosyl acceptor 10 in the presence of triflic anhydride and DTBMP pro-
duced predominantly the β-mannosyl trisaccharide 4 (β/α = 9:1) in 75% yield.
Other 4,6-O-benzylidene-protected mannosyl donors such as the correspond-
ing mannosyl trichloroacetimidate[14] and mannose 6 involving a mannosyl
phthalate intermediate[8] were found to be less satisfactory than the CB 4,6-
O-benzylidene mannoside donor 5 in the case of the β-mannosylation of 10 to
make 4. Three one-bond coupling constants (JC1−H1), 157.2, 159.9, and 161.9
Hz, again indicated that all glycosyl linkages including the newly generated
one in the trisaccharide 4 are in β-configurations. A similar strategy for the
construction of two consecutive β-mannosyl linkages, namely, β-(1→4) and β-
(1→3) linkages, employing 4,6-O-benzylidene mannosyl donors have been re-
ported for the synthesis of a mannan.[15]
Reductive cleavage of the 4,6-O-benzylidene group of the fully protected
trisaccharide 4 with borane in the presence of Bu2BOTf[16] afforded trisaccha-
ride acceptor 3 having a free hydroxy group at C-6ꢁꢁꢁ as shown in Scheme 2.
Scheme 2.

S.M. Park et al.
322
Oxidative cleavage of the anomeric p-methoxyphenyl group of 4 with ceric am-
monium nitrate (CAN) in CH3CN/H2O,[17] on the other hand, provided trisac-
charide donor 2 having an anomeric hydroxy group at C-1. Final coupling of
two trisaccharides 2 and 3 was accomplished by glycosylation of the acceptor
3 with the anomeric hydroxy sugar 2 as the donor using phthalic anhydride
and Tf2O as activators.[8] Thus, reaction of the trisaccharide donor 2 and ph-
thalic anhydride in the presence of DBU at rt followed by sequential addi-
tion of DTBMP, triflic anhydride, and the trisaccharide acceptor 3 to the solu-
tion at −78◦C produced predominantly desired hexasaccharide 1 (β/α = 10:1)
in 69% yield. Although synthesis of 1 by glycosylation of 3 with the glycosyl
trichloroacetimidate derived from 2 also gave the desired hexasaccharide 1,
side products generated during the glycosylation made purification of 1 more
difficult. Six one-bond coupling constants (JC1−H1), 155, 160, 157, 160, 155, and
158 Hz, clearly indicated that all six glycosyl linkages including the newly gen-
erated one in the hexasaccharide 1 are in β-configurations.
In summary, synthesis of hexasaccharide 1, a suitably protected form of
the repeating unit of the cell wall polymer from K. aurantiaca VKM Ac-720T,
has been achieved by coupling of two trisaccharides 2 and 3, both of which
were obtained from common trisaccharide 4. Direct one-pot glycosylation with
anomeric hydroxy sugars involving glycosyl phthalate intermediates was em-
ployed for the stereoselective construction of the β-galactosyl linkage of hexas-
accharide 1 and one of the β-mannosyl linkages of the trisaccharide 4 as key
steps. Another β-mannosyl linkage of 4 was constructed by employing the CB
glycoside method.
ACKNOWLEDGEMENT
This work was supported by a grant from the Korea Science and Engineer-
ing Foundation through the Center for Bioactive Molecule Hybrids (CBMH).
S.M.P. and D.-H.S. thank the fellowship of the BK 21 program from the Min-
istry of Education and Human Resources Development.
EXPERIMENTAL
All reactions were conducted under a positive pressure of dry argon with dry,
freshly distilled solvents unless otherwise noted. All reagents were purchased
from commercial suppliers and used without further purification unless oth-
erwise noted. Optical rotations were determined at 20◦C with an automatic
polarimeter. 1H NMR and 13C NMR spectra were recorded on a 400-MHz spec-
trometer (400 MHz for 1H, 100 MHz for 13C). Chemical shifts are given in ppm
downfield from internal tetramethylsilane for spectra recorded in CDCl3. Flash
column chromatography was performed employing 230–400 mesh silica gel.

Synthesis of Kineosporia aurantiaca Polysaccharide
323
Thin layer chromatography was taken using silica gel 60 F254 precoated plates
(0.25 mm thickness) with a fluorescent indicator. Visualization on TLC was
achieved by UV light (254 nm) and a typical TLC indication solution (cerium
sulfate/molybdic acid solution). Solutions were concentrated at below 40◦C un-
der reduced pressure.
p-Methoxyphenyl (2,3-di-O-benzyl-4,6-O-benzylidene-β-
D-mannopyranosyl)-(1→3)-2,4,6-tri-O-benzoyl-β-
D-galactopyranoside (8)
To a mixture of 4,6-O-benzylidene mannose 6 (100 mg, 0.22 mmol) and
4A molecular sieves in CH2Cl2 (2 mL) were added phthalic anhydride (36 mg,
˚
9.246 mmol) and DBU (38 µL, 0.269 mmol). After being stirred for 15 min at rt,
the reaction mixture was cooled down to −78◦C. DTBMP (100 mg, 0.049 mmol)
and Tf2O (57.4 µL, 0.36 mmol) were added to the reaction mixture and it was
stirred for a further 15 min at −78◦C. After slow addition of glycosyl acceptor
7 in CH2Cl2 (2 mL) at −78◦C , the reaction mixture was stirred for a further 15
min, warmed up to 0◦C, and further stirred for 1 h. The reaction mixture was
diluted with CH2Cl2 and the filtered organic solution was washed with satu-
rated aqueous NaHCO3 solution and brine. The organic layer was dried over
MgSO4 and concentrated in vacuo. The residue was purified by flash column
chromatography (hexane/EtOAc, 2:1) to give compound 8 (167 mg, 74%): color-
1
less oil, Rf = 0.45 (hexane/EtOAc, 2:1, v/v). H NMR (400 MHz, CDCl3)δ 3.64
(d, J = 3.2 Hz, 1H), 3.67 (s, 3H), 3.84 (t, J = 10.4 Hz, 1H), 4.07 (m, 2H), 4.23 (m,
2H), 4.27 (m, 3H), 4.30 (d, J = 3.2 Hz, 1H), 4.45 (d, J = 2.8 Hz, 1H), 4.48 (dd,
J = 8.0, 7.2 Hz, 1H), 4.58 (s, 1H), 4.60 (d, J = 4.0 Hz, 2H), 5.10 (d, J = 8.0 Hz,
1H), 5.52 (s, 1H), 5.88 (d, J = 3.2 Hz, 1H), 6.00 (dd, J = 10.0, 5.0 Hz, 1H), 6.60
(d, J = 9.2 Hz, 2H), 6.92 (d, J = 9.2 Hz, 2H), 7.09–8.20 (m, 30H). 13C NMR (100
MHz, CDCl3)δ 55.8, 63.3, 68.3, 68.7, 70.6, 72.0, 72.2, 72.6, 73.0, 74.9, 76.1, 77.1,
77.8, 101.1, 101.6, 103.3, 114.7, 119.0, 126.3, 127.5, 127.6, 127.7, 128.3, 128.4,
128.8, 129.1, 129.6, 129.8, 129.9, 130.0, 130.5, 133.6, 133.8, 133.9, 137.8, 138.4,
138.7, 151.5, 155.9, 165.5, 165.9, 166.4. Anal. Calcd for C61H56O15: C, 71.19; H,
5.48. Found: C, 71.36; H, 5.28.
p-Methoxyphenyl (2,3-di-O-benzyl-β-D-mannopyranosyl)-(1→3)-
2,4,6-tri-O-benzoyl-β-D-galactopyranoside (9)
To a solution of disaccharide 8 (300 mg, 0.291 mmol) in CH2Cl2 (4 mL) were
added TFA (740 µL, 4.4 mmol) and water (74 µL) and the reaction mixture
was stirred for 4 h at rt. The reaction mixture was diluted with CH2Cl2, and
washed with saturated aqueous NaHCO3 solution and brine. The organic layer
was dried over MgSO4 and concentrated in vacuo. The residue was purified by
flash column chromatography (hexane/EtOAc, 1:2) to give dihydroxy compound

S.M. Park et al.
324
9 (183 mg, 67%): colorless oil, Rf = 0.3 (hexane/EtOAc, 1:2, v/v). 1H NMR (400
MHz, CDCl3) δ 3.00 (dd, J = 2.8, 3.2 Hz, 1H), 3.23 (m, 1H), 3.53 (d, J = 2.8 Hz,
1H), 3.69 (s, 3H), 3.79 (d, J = 9.6 Hz, 1H), 3.84 (d, J = 3.2 Hz, 1H), 3.87 (d, J =
2.4 Hz, 1H), 3.90 (d, J = 12.0 Hz, 1H), 4.10 (m, 2H), 4.13 (d, J = 3.6 Hz, 1H),
4.26 (t, J = 6.4 Hz, 2H), 4.30 (d, J = 12.4 Hz, 1H), 4.46 (d, J = 12.4 Hz, 1H),
4.50 (s, 1H), 4.55 (d, J = 4.4 Hz, 2H), 5.12 (d, J = 8.4 Hz, 1H), 5.96 (dd, J =
10.0, 5.0 Hz, 1H), 6.08 (d, J = 3.6 Hz, 1H), 6.62 (d, J = 9.2 Hz, 2H), 6.93 (d,
J = 9.2 Hz, 2H), 7.00–8.19 (m, 25H). 13C NMR (100 MHz, CDCl3)δ 55.9, 66.1,
70.7, 70.8, 71.6, 72.2, 73.1, 74.0, 76.6, 76.8, 77.1, 79.8, 80.7, 101.2, 103.9, 114.7,
119.1, 127.5, 128.0, 128.3, 128.5, 128.8, 128.9, 129.0, 129.4, 129.9, 130.0, 130.1,
130.1, 130.6, 133.6, 133.8, 134.0, 137.8, 138.7, 151.5, 156.0, 165.5, 166.4, 166.8.
Anal. Calcd for C54H52O15: C, 68.93; H, 5.57. Found: C, 68.87; H, 5.51.
p-Methoxyphenyl (6-O-benzoyl-2,3-di-O-benzyl-β-
D-mannopyranosyl)-(1→3)-2,4,6-tri-O-benzoyl-β-D-
galactopyranoside (10)
A solution of dihydroxy disaccharide 9 (150 mg, 0.16 mmol), benzoyl chlo-
ride (33 µL, 0.191 mmol), and pyridine (150 µL) in CH2Cl2 (5 mL) was stirred
at 0◦C for 1 h. The reaction mixture was diluted with CH2Cl2 and washed with
1N HCl, saturated aqueous NaHCO3 solution, and brine. The organic layer
was dried over MgSO4 and concentrated in vacuo. The residue was purified by
flash column chromatography (hexane/EtOAc, 1:1) to afford compound 10 (144
1
mg, 87%): colorless oil, Rf = 0.3 (hexane/EtOAc, 1:1, v/v). H NMR (400 MHz,
CDCl3) δ 2.99 (dd, J = 2.8, 9.2 Hz, 1H), 3.45 (m, 1H), 3.62 (d, J = 2.4 Hz, 1H),
3.68 (s, 3H), 3.90 (t, J = 9.6 Hz, 1H), 3.96 (d, J = 12.0 Hz, 1H), 4.06 (m, 1H),
4.18 (d, J = 12.0 Hz, 1H), 4.22 (d, J = 3.6 Hz, 1H), 4.35 (d, J = 12.0 Hz, 1H),
4.47 (m, 2H), 4.57 (d, J = 2.4 Hz, 2H), 4.60 (m, 3H), 5.01 (d, J = 8.0 Hz, 1H),
5.93 (d, J = 3.2 Hz, 1H), 5.97 (dd, J = 10.0, 5.0 Hz, 1H), 6.60 (d, J = 9.2 Hz,
2H), 6.90 (d, J = 9.2 Hz, 2H), 7.00–8.10 (m, 30H). 13C NMR (100 MHz, CDCl3)δ
55.8, 63.3, 63.9, 66.5, 70.5, 70.9, 72.0, 72.6, 73.2, 73.9, 74.8, 80.4, 80.9, 101.2,
103.1, 114.6, 119.0, 122.7, 125.0, 126.5, 127.3, 127.9, 128.2, 128.7, 129.0, 129.5,
129.8, 129.9, 130.0, 130.3, 133.2, 133.4, 133.8, 142.4, 147.3, 149.7, 151.1, 151.4,
155.8, 165.4, 165.6, 166.2, 166.8. Anal. Calcd for C61H56O16: C, 70.10; H, 5.40.
Found: C, 70.07; H, 5.32.
p-Methoxyphenyl (2,3-di-O-benzyl-4,6-O-benzylidene-β-
D-mannopyranosyl)-(1→4)-(6-O-benzoyl-2,3-di-O-benzyl-β-
D-mannopyranosyl)-(1→3)-2,4,6-tri-O-benzoyl-β-D-
galactopyranoside (4)
˚
To a mixture of glycosyl acceptor 10 (87 mg, 0.083 mmol) and 4A molec-
ular sieves in CH2Cl2 (4 mL) at 78◦C were added sequentially DTBMP (51
mg, 0.25 mmol), Tf2O (60 µL, 0.125 mmol), and then glycosyl donor 5 (74 mg,

Synthesis of Kineosporia aurantiaca Polysaccharide
325
0.125 mmol) in CH2Cl2 (4 mL). After being stirred for 30 min at −78◦C, the
reaction mixture was warmed up to 0◦C, further stirred for 1.5 h, and diluted
with CH2Cl2. The filtered organic solution was washed with saturated aqueous
NaHCO3 solution and brine, dried over MgSO4, and concentrated in vacuo. The
residue was purified by flash column chromatography (hexane/EtOAc/CH2Cl2,
4:1:1) to give compound 4 (92 mg, 75%): colorless oil, Rf = 0.25 (hexane/EtOAc,
1
2:1, v/v). H NMR (400 MHz, CDCl3)δ 2.92 (m, 1H), 3.09 (dd, J = 2.8, 9.2 Hz,
1H), 3.25 (dd, J = 2.8, 9.2 Hz, 1H), 3.44 (dd, J = 2.8, 9.6 Hz, 1H), 3.51 (m, 1H),
3.59 (t, J = 3.4 Hz, 1H), 3.69 (s, 3H), 3.82 (dd, J = 10.4, 4.4 Hz, 2H), 3.88 (d,
J = 2.8 Hz, 2H), 4.04 (t, J = 9.6 Hz, 2H), 4.08 (m, 1H), 4.21 (m, 2H), 4.33 (m,
2H), 4.48 (d, J = 4.4 Hz, 1H), 4.51 (m, 2H), 4.55 (d, J = 3.6 Hz, 2H), 4.58 (d,
J = 1.6 Hz, 1H), 4.64 (d, J = 2.4 Hz, 1H), 4.76 (d, J = 8.0 Hz, 1H), 4.87 (d, J =
12.0 Hz, 1H), 5.01 (d, J = 8.0 Hz, 1H), 5.46 (s, 1H), 5.93 (m, 2H), 6.60 (d, J =
9.0 Hz, 2H), 6.89 (d, J = 9.0 Hz, 2H), 7.02–8.08 (m, 45H). 13C NMR (100 MHz,
CDCl3)δ 55.7, 61.5, 63.3, 63.4, 67.0, 67.5, 68.6, 70.3, 71.3, 71.8, 72.6, 73.8, 74.0,
74.3, 75.3, 75.4, 77.0, 78.5, 78.6, 79.7, 101.1, 101.5, 101.8, 102.7, 114.5, 119.0,
126.3, 127.2, 127.7, 127.7, 127.8, 127.9, 128.3, 128.4, 128.5, 128.6, 129.0, 129.3,
129.7, 129.9, 130.0, 133.2, 133.4, 133.8, 137.8, 138.3, 138.5, 138.7, 138.8, 151.4,
155.8, 165.4, 165.5, 166.2, 166.4. Anal. Calcd for C88H82O21: C, 71.63; H, 5.60.
Found: C, 71.67; H, 5.47.
(2,3-Di-O-benzyl-4,6-O-benzylidene-β-D-mannopyranosyl)-
(1→4)-(6-O-benzoyl-2,3-di-O-benzyl-β-D-mannopyranosyl)-
(1→3)-2,4,6-tri-O-benzoyl-D-galactopyranose (2)
To a mixture of compound 4 (59 mg, 0.04 mmol) in a mixed solvent of
CH3CN (800 µL) and H2O (200 µL) at 0◦C was added CAN (44 mg, 0.08 mmol).
After being stirred for 30 min at 0◦C, the reaction mixture was diluted with
EtOAc and washed with saturated aqueous NaHCO3 solution and brine. The
organic layer was dried over MgSO4 and concentrated in vacuo. The residue
was purified by flash column chromatography (hexane/EtOAc/CH2Cl2, 2:1:1)
to afford compound 2 (33 mg, 60%): yellow oil, Rf = 0.18 (hexane/EtOAc, 2:1,
1
v/v). H NMR (400 MHz, CDCl3)δ 2.97 (m, 1H), 3.30 (dd, J = 2.8, 9.2 Hz, 1H),
3.32 (s, 1H), 3.40 (d, J = 1.6 Hz, 1H), 3.46 (dd, J = 2.8, 9.2 Hz, 1H), 3.61 (m,
2H), 3.86 (dd, J = 4.4, 4.8 Hz, 1H), 3.90 (m, 1H), 4.06 (dd, J = 9.6, 8.0 Hz,
2H), 4.16 (d, J = 12.0 Hz, 1H), 4.21 (d, J = 12.0 Hz, 1H), 4.28 (d, J = 6.4 Hz,
1H), 4.34 (dd, J = 5.2, 5.6 Hz, 2H), 4.48 (m, 3H), 4.47 (d, J = 12.0 Hz, 2H),
4.55 (m, 3H), 4.60 (d, J = 5.2 Hz, 1H), 4.67 (d, J = 7.2 Hz, 1H), 4.82 (m, 1H),
4.90 (d, J = 12.0 Hz, 1H), 5.47 (s, 1H), 5.69 (d, J = 9.2 Hz, 1H), 5.96 (s, 1H),
7.05–8.08 (m, 45H). 13C NMR (100 MHz, CDCl3)δ 63.3, 63.5, 67.5, 67.8, 68.6,
71.3, 71.4, 71.7, 72.4, 72.6, 73.8, 73.8, 73.9, 74.2, 75.4, 77.4, 78.5, 78.6, 79.7,
91.2, 101.5, 102.1, 102.9, 126.3, 127.0, 127.2, 127.6, 127.7, 127.9, 128.0, 128.3,
128.4, 128.5, 128.6, 128.6, 129.0, 129.0, 129.1, 129.6, 129.7, 129.9, 129.9, 130.0,

S.M. Park et al.
326
130.1, 130.2, 133.1, 133.2, 133.3, 133.8, 138.2, 165.4, 165.9, 166.3, 166.4. Anal.
Calcd for C81H76O20: C, 71.04; H, 5.59. Found: C, 71.02; H, 5.58.
p-Methoxyphenyl (2,3,4-tri-O-benzyl-β-D-mannopyranosyl)-
(1→4)-(6-O-benzoyl-2,3-di-O-benzyl-β-D-mannopyranosyl)-
(1→3)-2,4,6-tri-O-benzoyl-β-D-galactopyranoside (3)
To a solution of compound 4 (67 mg, 0.0454 mmol) in THF (1mL) at 0◦C
were added BH3 · THF (450 µL) and Bu2BOTf (45 µL). After being stirred
for 15 min at 0◦C, the reaction mixture was warmed up to rt, stirred for a
further 2 h, and neutralized with Et3N. MeOH was added to the resulting mix-
ture, which was then concentrated in vacuo. The residue was purified by flash
column chromatography (hexane/EtOAc/CH2Cl2, 2:1:1) to afford compound 3
1
(50 mg, 75%): colorless oil, Rf = 0.125 (hexane/EtOAc, 2:1, v/v). H NMR (400
MHz, CDCl3) δ 2.97 (m, 1H), 3.19 (dd, J = 2.8, 9.2 Hz, 1H), 3.35 (dd, J = 2.8,
9.2 Hz, 1H), 3.43 (m, 2H), 3.58 (m, 2H), 3.63 (d, J = 2.8 Hz, 1H), 3.69 (s, 3H),
3.78 (t, J = 9.6 Hz, 1H), 3.85 (d, J = 2.8 Hz, 1H), 4.02 (dd, J = 4.4, 4.0 Hz, 1H),
4.08 (t, J = 9.6 Hz, 1H), 4.16 (dd, J = 2.0, 3.2 Hz, 2H), 4.20 (d, J = 6.4 Hz, 1H),
4.28 (d, J = 12.0 Hz, 1H), 4.38 (d, J = 3.2 Hz, 2H), 4.42 (m, 2H), 4.46 (m, 1H),
4.49 (d, J = 3.2 Hz, 1H), 4.52 (d, J = 3.6 Hz, 1H), 4.55 (d, J = 3.2 Hz, 2H), 4.57
(d, J = 3.6 Hz, 1H), 4.60 (d, J = 3.6 Hz, 1H), 4.77 (d, J = 12.0 Hz, 1H), 4.82 (d,
J = 4.4 Hz, 1H), 5.00 (d, J = 8 Hz, 1H), 5.90 (d, J = 3.2 Hz, 1H), 5.95 (dd, J =
8.0, 5.0 Hz, 1H), 6.60 (d, J = 9.2 Hz, 2H), 6.90 (d, J = 9.2 Hz, 2H), 7.01–8.09
(m, 45H). 13C NMR (100 MHz, CDCl3) δ 55.8, 62.4, 63.3, 63.8, 70.4, 70.9, 71.5,
71.8, 72.5, 73.8, 74.1, 74.6, 74.7, 74.9, 75.3, 75.8, 76.9, 78.1, 78.4, 82.6, 100.3,
101.1, 102.8, 114.6, 119.0, 127.3, 127.4, 127.6, 127.7, 127.8, 127.9, 128.0, 128.2,
128.2, 128.2, 128.3, 128.4, 128.6, 128.6, 128.6, 128.7, 128.8, 128.9, 129.4, 129.8,
130.0, 130.0, 130.1, 130.2, 130.3, 130.4, 133.3, 133.4, 133.8, 138.1, 138.3, 151.4,
155.8, 165.4, 165.5, 166.2, 166.5. Anal. Calcd for C88H84O21: C, 71.53; H, 5.73.
Found: C, 71.49; H, 5.85.
p-Methoxyphenyl (2,3-di-O-benzyl-4,6-O-benzylidene-β-
D-mannopyranosyl)-(1→4)-(6-O-benzoyl-2,3-di-O-benzyl-β-
D-mannopyranosyl)-(1→3)-(2,4,6-tri-O-benzoyl-β-
D-galactopyranosyl)-(1→6)-(2,3,4-tri-O-benzyl-β-
D-mannopyranosyl)-(1→4)-(6-O-benzoyl-2,3-di-O-benzyl-β-
D-mannopyranosyl)-(1→3)-2,4,6-tri-O-benzoyl-β-
D-galactopyranoside (1)
˚
To a mixture of trisaccharide donor 2 (100 mg, 0.072 mmol) and 4A molec-
ular sieves in CH2Cl2 (2 mL) were added phthalic anhydride (12 mg, 0793
mmol) and DBU (13.3 µL, 0.086 mmol). After being stirred for 15 min at rt,

Synthesis of Kineosporia aurantiaca Polysaccharide
327
the reaction mixture was cooled down to −78◦C. DTBMP (32.4 mg, 0.158 mmol)
and Tf2O (52 µL, 0.108 mmol) were added to the reaction mixture and it was
stirred for 15 min at −78◦C. After slow addition of glycosyl acceptor 3 (127 mg,
0.086 mmol) in CH2Cl2 (2 mL), the reaction mixture was stirred for 15 min,
warmed up to 0◦C, further stirred for 1.5 h, and diluted with CH2Cl2. The fil-
tered organic solution was washed with saturated aqueous NaHCO3 solution
and brine, dried over MgSO4, and concentrated in vacuo. The residue was pu-
rified by flash column chromatography (hexane/EtOAc, 2:1) and high-pressure
gel permeation chromatography to give hexasaccharide 1 (140 mg, 69%): color-
1
less oil, Rf = 0.24 (hexane/EtOAc, 2:1, v/v). H NMR (400 MHz, CDCl3) δ 2.90
(m, 1H), 3.10 (d, J = 10.4 Hz, 1H), 3.24 (dd, J = 2.4, 8.8 Hz, 1H), 3.41 (dd, J =
2.8, 6.4 Hz, 2H), 3.49 (d, J = 2.8 Hz, 1H), 3.54 (m, 2H), 3.67 (s, 3H), 3.68 (m,
1H), 3,78 (m, 1H), 3.80 (d, J = 2.4 Hz, 1H), 3.98–4.15 (m, 12H), 4.21–4.45 (m,
14H), 4.47–4.59 (m, 14H), 4.63 (d, J = 4.8 Hz, 1H), 4.66 (m, 1H), 4.75 (m, 1H),
4.79 (m, 1H), 4.85 (m, 1H), 5.00 (d, J = 8 Hz, 1H), 5.45 (s, 1H), 5.71 (t, J = 9.2
Hz, 1H), 5.80 (d, J = 2.8 Hz, 1H), 5.92 (d, J = 4.0 Hz, 1H), 6.60–8.06 (m, 94H).
13C NMR (100 MHz, CDCl3) δ 55.8, 60.6, 62.9, 63.3, 63.6, 64.2, 67.5, 68.4, 68.6,
70.4, 70.7, 70.8, 71.2, 71.5, 71.7, 71.9, 72.5, 72.6, 73.2, 73.6, 73.8, 73.8, 74.1,
74.2, 74.5, 75.1, 75.3, 75.4, 77.0, 77.3, 77.4, 77.5, 77.6, 78.0, 78.2, 78.6, 78.7,
79.8, 80.0, 82.7, 100.7, 101.1, 101.5, 101.9, 102.7, 102.8, 103.0, 114.5, 119.0,
126.3, 126.8, 127.1, 127.6, 127.7, 127.8, 127.9, 128.0, 128.1, 128.3, 128.3, 128.3,
128.4, 128.4, 128.5, 128.5, 128.6, 128.6, 128.6, 128.9, 129.0, 129.3, 129.8, 129.9,
129.9, 130.0, 130.2, 133.1, 133.2, 133.4, 133.8, 138.8, 138.9, 151.4, 155.8, 165.3,
165.4, 165.5, 165.5, 166.2, 166.2, 166.3, 166.4. Anal. Calcd for C169H158O40: C,
71.75; H, 5.63. Found: C, 71.75; H, 5.60.
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