Synthesis of a tetrasaccharide repeating unit of the exopolysaccharide from Burkholderia multivorans

Article abstract of DOI:10.1080/07328303.2017.1391275

The chemical synthesis of a tetrasaccharide repeating unit of the exopolysaccharide discovered from Burkholderia multivorans with a 3-aminopropyl group linked to the glycan downstream end, α-D-Manp-(1→2)-α-D-Manp-(1→2)-3-O-methyl-α-DRhap-(1→3)-α-D-Rhap-O(CH₂)₃NH₂ (1), was described. The target tetrasaccharide was achieved by both a convergent [2 + 2] and a linear glycosylation strategies. The latter synthesis was proved to be more efficient than the former due to the excellent stereocontrol of glycosylation. Furthermore, the 3-aminopropyl group in the target molecule would enable its conjugation with functional biomolecules to explore its biological applications.

Full text of DOI:10.1080/07328303.2017.1391275

Journal of Carbohydrate Chemistry
ISSN: 0732-8303 (Print) 1532-2327 (Online) Journal homepage: http://www.tandfonline.com/loi/lcar20
Synthesis of a tetrasaccharide repeating unit
of the exopolysaccharide from Burkholderia
multivorans
Xin Zhang, Dongyue Wang, Guoxia Jin, Lizhen Wang, Zhongwu Guo &
Guofeng Gu
To cite this article: Xin Zhang, Dongyue Wang, Guoxia Jin, Lizhen Wang, Zhongwu
Guo & Guofeng Gu (2017): Synthesis of a tetrasaccharide repeating unit of the
exopolysaccharide from Burkholderia multivorans, Journal of Carbohydrate Chemistry, DOI:
10.1080/07328303.2017.1391275
To link to this article: http://dx.doi.org/10.1080/07328303.2017.1391275
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Date: 04 November 2017, At: 18:06

JOURNAL OF CARBOHYDRATE CHEMISTRY
, VOL. , NO. , –
https://doi.org/./..
Synthesis of a tetrasaccharide repeating unit of the
exopolysaccharide from Burkholderia multivorans
Xin Zhanga, Dongyue Wanga, Guoxia Jinc, Lizhen Wanga, Zhongwu Guoa,b
and Guofeng Gua
,
aNational Glycoengineering Research Center, Shandong Provincial Key Laboratory of Carbohydrate
Chemistry and Glycobiology, Shandong University,  Shanda Nan Lu, Jinan, PR China; bDepartment of
Chemistry, University of Florida,  Leigh Hall, Gainesville, Florida, United States; cCollege of Chemistry,
Chemical Engineering and Materials Science, Collaborative Innovation Center of Functionalized Probes for
Chemical Imaging, Key Laboratory of Molecular and Nano Probes, Ministry of Education, Shandong
Normal University,  Wenhua Dong Lu, Jinan, PR China
ABSTRACT
ARTICLE HISTORY
Received  October 
Accepted  October 
The chemical synthesis of a tetrasaccharide repeating unit of
the exopolysaccharide discovered from Burkholderia multivo-
rans with a 3-aminopropyl group linked to the glycan down-
stream end, α-D-Manp-(1→2)-α-D-Manp-(1→2)-3-O-methyl-α-D-
Rhap-(1→3)-α-D-Rhap-O(CH2)3NH2 (1), was described. The target
tetrasaccharide was achieved by both a convergent [2 + 2] and
a linear glycosylation strategies. The latter synthesis was proved
to be more efficient than the former due to the excellent stereo-
controlofglycosylation. Furthermore, the3-aminopropylgroupin
the target molecule would enable its conjugation with functional
biomolecules to explore its biological applications.
KEYWORDS
Burkholderia multivorans;
Exopolysaccharide;
Synthesis; Glycosylation
GRAPHICAL ABSTRACT
Introduction
Bacterial polysaccharides, such as lipopolysaccharides (LPSs), capsular polysaccha-
rides (CPSs), exopolysaccharides (EPSs), etc., are major components of the bacterial
CONTACT Guofeng Gu
guofenggu@sdu.edu.cn
National Glycoengineering Research Center, Shandong
Provincial Key Laboratory of Carbohydrate Chemistry and Glycobiology, Shandong University,  Shanda Nan Lu, Jinan
, PR China.
Supplemental data for this article can be accessed on the publisher’s website.
©  Taylor & Francis Group, LLC

2
X. ZHANG ET AL.
Figure . The structures of (a) B. Multivorans exopolysaccharide (EPS) and (b) synthetic target
molecule 1.
cell glycocalyx that plays an important role in bacterial immunology.1–6 Substantial
endeavors have been focused on using bacterial polysaccharides, especially LPSs
and CPSs, as antigen candidates for the development of antibacterial vaccines,7,8
resulting in a series of commercialized bacterial polysaccharide and their protein
conjugate vaccines.9,10 Therefore, bacterial polysaccharides have been recognized
as crucial immunotargets useful for the development of new carbohydrate-based
diagnostic agents and vaccines.11
A novel exopolysaccharide that consists of a linear tetrasaccharide repeating unit
has recently been isolated from B. multivorans strain C1576.12 The structure of this
EPS was characterized as →3)-α-D-Manp-(1→2)-α-D-Manp-(1→2)-3-O-methyl-
α-D-Rhap-(1→3)-α-D-Rhap-(1→ (Figure 1a). As a part of our ongoing effort to
synthesize oligosaccharide fragments relating to Burkholderia cepacia complex (Bcc)
polysaccharides for in-depth immunological studies and vaccine development,13–16
we described herein the chemical synthesis of this tetrasaccharide repeating unit 1
(Figure 1b). The synthetic target was also designed to contain a free 3-aminopropyl
group linked to the D-rhamnosyl reducing end, which would facilitate its regios-
elective conjugation with carrier molecules to generate glycoconjugates useful for
biological and immunological studies.
Results and discussion
As outlined in Scheme 1, target molecule 1 could be assembled from glycosylation
of disaccharide donor 217 with disaccharide acceptor 3 via a convergent [2 + 2]
strategy. In turn, 3 could be prepared from the coupling reaction of monosaccharide
building blocks 4 and 5. Considering the all 1,2-trans glycosidic linkages in 1, we
planned to use acyl groups, including both acetyl and benzoyl groups, to protect the
2-O-position of each monosaccharyl building block, in order to take advantage of
the neighboring group participation for stereoselective glycosylation.
Our synthesis commenced with the preparation of monosaccharyl building
blocks 4 and 5. The synthesis of rhamnosyl donor 4 started from 613 as outlined
in Scheme 2. Firstly, regioselective 4-methoxybenzylation of the 2-OH group in

JOURNAL OF CARBOHYDRATE CHEMISTRY
3
Scheme . Retrosynthetic analysis of target molecule .
6 using p-methoxybenzyl chloride (PMBCl) and NaH in DMF afforded 7, which
was followed by methylation of the remaining 3-OH group using methyl iodine to
produce 8 in an overall yield of 56%. Removal of the PMB group in 8 by treatment
with 80% TFA and then acetylation of the resultant 2-OH group gave 9 in an
1
85% yield (two steps). The downfield-shifted H-2 signal at δ 5.27 ppm in the H
NMR spectrum of 9 confirmed the 2-methoxybenzylation of 6 with the correct
regiochemistry. Next, acetolysis of 9 with AcOH and Ac2O (v/v 1:1) containing 1%
concentrated H2SO4 at 0°C18 generated 10 as the main product in an 85% yield.
Finally, conversion of 10 into hemiacetal by treatment with hydrazine acetate19 and
then trichloroacetimidation20 of the resultant hemiacetal with trichloroacetonitrile
and 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) according to literature procedures
provided rhamnosyl imidate 4 (73%).
The synthesis of rhamnosyl acceptor 5 was delineated in Scheme 3. The tosylate
1115 was treated with lithium aluminium hydride (LiAlH4) in THF21 to smootly
produce the 6-deoxy sugar 12 in a 73% yield. The newly formed methyl group in 12
was verified from the chemical shift of the H-6 signal with a doublet peak (J = 6.0
Hz) at δ 1.24 ppm in its 1H NMR spectrum. Benzoylation of the 4-OH group in 12
with benzyol cholride in pyridiene gave 13, which was followed by the removal of
acetonide in 13 with 80% TFA in CH2Cl2 to afford 2,3-diol 14 in an oveall 72% yield.
Reaction of 14 with trimethyl orthobenzoate22 under promotion of catalytic amount
Scheme . Synthesis of rhamnosyl donor .

4
X. ZHANG ET AL.
Scheme . Synthesis of rhamnosyl acceptor .
of (+)-10-camphorsulfonic acid (CSA) in CH2Cl2 and then treatment of the resul-
tant intermediate with 80% aqueous HOAc furnished 2-O-benzoylated rhamnoside
acceptor 5 in an 81% yield. The downfield-shifted H-2 signal at δ 5.64 ppm in the
1H NMR spectrum of 15 confirmed the correct regioselectivity.
Disaccharide acceptor 3 was prepared by a procedure depicted in Scheme 4. First,
5 was glycosylated with trichloroacetimidate 4 under the promotion of trimethylsilyl
triflate (TMSOTf) in CH2Cl2 at −78°C, resulting in a fully protected α−1,2-linked
disaccharide derivative 15 in a 71% yield. The reaction was highly stereoselective as
a result of the neighboring group participation effect. The α-configuration of newly
1
formed glycosidic bond was further confirmed by the JC-1,H-1 coupling constant
(170.4 Hz)23 in the 1H-coupled gHSQC spectrum of 15. The coupling reaction of
15 with 3-azidopropanol in CH2Cl2 at −15°C, using N-iodosuccinimide (NIS) and a
catalytic amount of TMSOTf as promoters, proceeded smoothly to give disaccharide
16 in a 75% yield. Selective removal of the 2-O-acetyl group in 16 was completed
with 5% acetyl chloride (AcCl)24 in CH2Cl2 and MeOH (v/v 1:1) to afford disac-
charide acceptor 3 in an 85% yield. Then, 3 was glycosylated with thiomannoside 2
under the influence of a NIS-TMSOTf co-catalytic system in CH2Cl2 at −15°C to
generate a mixture of α- and β-linked tetrasaccharide (α/β 3:2, 83%). The α-linked
tetrasaccharide 17 was easily purified by silica gel column chromatography in a 50%
yield. The 1JC-1,H-1 coupling constants (all over 172 Hz) in its 1H-coupled gHSQC
spectrum confirmed the correct α-glycosidic linkages in 17.
Scheme . Synthesis of tetrasaccharide  via a [+] convergent glycosylation strategy.

JOURNAL OF CARBOHYDRATE CHEMISTRY
5
Scheme . Synthesis of target tetrasaccharide  via a linear glycosylation strategy.
This synthesis was then compared with a synthetic strategy employing linear
glycosylation as outlined in Scheme 5. Glycosylation of 3 with trichloroacetimidate
1825 under the promotion of TMSOTf furnished trisaccharide 19, which was then
deacetylated with 5% acetyl chloride (AcCl) in CH2Cl2 and MeOH to afford 20 as
a glycosyl acceptor in an overall yield of 71% for 2 steps. Again, 20 was glycosylated
with 18 in the presence of TMSOTf in CH2Cl2 at −15°C to smoothly produce the
fully protected tetrasaccharide 17 in an 81% yield. Finally, global deprotection of 17
in two steps, including deacylation with sodium methoxide in methanol and deben-
zylation under hydrogenation conditions in acetic acid and H2O with 10% Pd/C as
the catalyst, afforded desired tetrasaccharide 1 in a 63% overall yield, after purifica-
tion by size-exclusion chromatography on a Bio-Gel P-2 column. The 1H, 13C NMR
and MS data of the product were in full agreement with the desired structure of 1.
In conclusion, we have described here two syntheses of the tetrasaccharide
repeating unit of B. multivorans exopolysaccharide. In these syntheses, convergent
[2 + 2] and linear glycosylation strategies were employed to accomplish the target
molecule 1 from monosaccharyl building blocks 4 and 5 in 14.25% and 16.48%
yields, respectively. The latter synthetic strategy was proved to be more efficient
due to the excellent stereo-control of each glycosylation reaction as a result of
neighboring group participation effect. The target molecule had a 3-aminopropyl
group at its reducing end that would enable its convenient attachment to func-
tionalized biomolecules for biological applications such as the development of
new carbohydrate-based diagnostic agents and vaccines against B. multivorans
infections.
Experimental section
General methods
1H and 13C NMR spectra were recorded with an Agilent 600 MHz spectrometer
for solutions in CDCl3 or D2O. Chemical shifts are given in ppm downfield from

6
X. ZHANG ET AL.
Me4Si. Me4Si was used as an internal reference for solutions in CDCl3, and the DHO
signal was used as the reference for solutions in D2O. Positive-mode electrospray
ionization (ESI) high-resolution mass spectra (HRMS) were recorded with a JEOL
JMS-DX-303HF spectrometer. Thin-layer chromatography (TLC) was carried out
on silica gel HF254 plates with detection by charring using H2SO4 (30% v/v in
MeOH) or using a UV detector. Silica gel column chromatography was carried out
with mixtures of ethyl acetate and petroleum ether (b.p. 60–90 °C) or toluene as the
eluents. Solutions were concentrated at <60 °C under diminished pressure.
Methyl 4-O-benzyl-2-O-p-methoxybenzyl-α-D-rhamnopyranoside 7
To a cooled solution of 6 (2.0 g, 7.46 mmol) in dry DMF (30 mL) was added NaH
(0.36 g, 60% in kerosene, 8.95 mmol), and PMBCl (0.9 mL, 6.71 mmol) in dry DMF
(30 mL) was then dropwise added to the above reaction mixture at 0°C. The result-
ing mixture was stirred at these condition for 3 h, and then diluted with EtOAc
(150 mL), washed successively with cooled water, 1 M HCl, and brine. The organic
phase was dried over Na2SO4, and concentrated. The residue was purified by flash
column chromatography (petroleum ether/ethyl acetate, 2:1) to afford 7 (1.75 g,
1
60%) as a white foamy solid. H NMR (600 MHz, CDCl3): δ 7.36–7.30 (m, 4H,
ArH), 7.29–7.26 (m, 3H, ArH), 6.90 (d, J = 7.8 Hz, 2H, ArH), 4.89 (d, J = 11.4 Hz,
1H, -CH2Ph), 4.68 (s, 1H, H-1), 4.67 (d, J = 11.4 Hz, 1H, -CH2Ph), 4.63 (d, J =
11.4 Hz, 1H, -CH2Ph), 4.48 (d, J = 11.4 Hz, 1H, -CH2Ph), 3.89 (dt, J = 9.6, 3.6 Hz,
1H, H-3), 2.89 (s, 3H, -OCH3), 3.69 (d, J = 3.6 Hz, 1H, H-2), 3.63 (m, 1H, H-5),
3.31 (s, 3H, -OCH3), 3.27 (t, J = 9.6 Hz, 1H, H-4), 2.27 (d, J = 3.6 Hz, 1H, -OH),
1.32 (d, J = 6.0 Hz, 3H, H-6); 13C NMR (150 MHz, CDCl3): δ 159.48, 138.55, 129.70,
129.67, 128.36, 127.89, 127.64, 113.96, 97.92, 82.26, 78.15, 74.99, 72.69, 71.56, 66.87,
55.27, 54.67, 17.99; ESI HRMS: calcd for (C22H28O6+NH4+) m/z, 406.2224; found,
406.2235; calcd for (C22H28O6+Na+) m/z, 411.1778; found, 411.1800.
Methyl 4-O-benzyl-2-O-p-methoxybenzyl-3-O-methyl-α-D-rhamnopyranoside 8
To a cooled solution of 7 (1.7 g, 4.38 mmol) in dry DMF (20 mL) was added NaH
(0.35 g, 60% in kerosene, 8.76 mmol) and CH3I (0.54 mL, 8.75 mmol). The reaction
mixture was stirred at 0°C for 2 h, and then diluted with EtOAc (100 mL), washed
successively with cooled water, 1 M HCl, and brine. The organic phase was dried
over Na2SO4, and concentrated. The residue was purified by flash column chro-
matography (petroleum ether/ethyl acetate, 2:1) to afford 8 (1.63 g, 93%) as a white
foamy solid. 1H NMR (600 MHz, CDCl3): δ 7.36–7.29 (m, 6H, ArH), 7.27 (t, J =
7.2 Hz, 1H, ArH), 6.88 (d, J = 8.4 Hz, 2H, ArH), 4.91 (d, J = 10.8 Hz, 1H, -CH2Ph),
4.69 (d, J = 12.0 Hz, 1H, -CH2Ph), 4.64 (d, J = 12.0 Hz, 1H, -CH2Ph), 4.61 (d, J
= 10.8 Hz, 1H, -CH2Ph), 4.60 (s, 1H, H-1), 3.80 (s, 3H, -OCH3), 3.75 (br s, 1H,
H-2), 3.62 (m, 1H, H-5), 3.53 (dd, J = 9.0, 3.0 Hz, 1H, H-3), 3.49 (t, J = 9.0 Hz,
1H, H-4), 3.39 (s, 3H, -OCH3), 3.28 (s, 3H, -OCH3), 1.32 (d, J = 6.0 Hz, 3H, H-
6); 13C NMR (150 MHz, CDCl3): δ 159.20, 138.81, 130.30, 129.51, 128.31, 127.86,

JOURNAL OF CARBOHYDRATE CHEMISTRY
7
127.52, 113.70, 98.92, 81.87, 80.38, 75.17, 73.36, 72.31, 67.59, 57.57, 55.26, 54.60,
17.98; ESI HRMS: calcd for (C23H30O6+NH4+) m/z, 420.2381; found 420.2383;
calcd for (C23H30O6+Na+) m/z, 425.1935; found, 411.1922.
Methyl 2-O-Acetyl-4-O-benzyl-3-O-methyl-α-D-rhamnopyranoside 9
After a solution of 8 (1.5 g, 3.73 mmol) in 80% TFA (5 mL) was stirred at rt for
1 h, the reaction mixture was then co-evaporated with toluene (30 mL) twice
under reduced pressure. The residue was purified by flash column chromatography
(petroleum ether/ethyl acetate, 2:1) to yield methyl 4-O-benzyl-3-O-methyl-α-D-
1
rhamnopyranoside (0.92 mg, 88%) as a white foamy solid. H NMR (600 MHz,
CDCl3): δ 7.35–7.32 (m, 4H, ArH), 7.30–7.26 (m, 1H, ArH), 4.84 (d, J = 11.4 Hz,
1H, -CH2Ph), 4.70 (s, 1H, H-1), 4.63–4.59 (d, J = 11.4 Hz, 1H, -CH2Ph), 4.03 (d,
J = 1.8 Hz, 1H, H-2), 3.67 (m, 1H, H-5), 3.53 (dd, J = 9.0, 2.4 Hz, 1H, H-3), 3.47 (s,
3H, -OCH3), 3.34 (m, 4H, H-4, -OCH3), 1.30 (d, J = 6.0 Hz, 3H, H-6); 13C NMR
(150 MHz, CDCl3): δ 138.50, 128.64, 128.35, 127.89, 127.65, 113.93, 100.00, 81.66,
79.75, 75.14, 67.75, 66.94, 57.37, 54.76, 17.86. A solution of the above product
(0.9 g, 3.19 mmol) in Ac2O (5 mL) and pyridine (5 mL) was stirred at rt for 2 h
and then concentrated under reduced pressure. The residue was purified by flash
column chromatography (petroleum ether/ethyl acetate, 2:1) to afford 9 (1.0 g, 97%)
1
as a white foamy solid. H NMR (600 MHz, CDCl3): δ 7.35–7.31 (m, 4H, ArH),
7.30–7.26 (m, 1H, ArH), 5.27 (dd, J = 3.6, 1.2 Hz, 1H, H-2), 4.88 (d, J = 10.8 Hz,
1H, -CH2Ph), 4.61 (d, J = 10.8 Hz, 1H, -CH2Ph), 4.60 (d, J = 1.2 Hz, 1H, H-1), 3.70
(m, 1H, H-5), 3.64 (dd, J = 9.6, 3.6 Hz, 1H, H-3), 3.41 (s, 3H, -OCH3), 3.35 (t, J =
9.6 Hz, 1H, H-4), 3.33 (s, 3H, -OCH3), 2.14 (s, 3H, -COCH3), 1.32 (d, J = 6.6 Hz,
3H, H-6); 13C NMR (150 MHz, CDCl3): δ 170.41, 138.57, 130.11, 128.33, 127.84,
127.62, 113.91, 98.53, 80.03, 79.89, 75.22, 68.54, 67.37, 57.50, 54.80, 21.08, 17.94.
1,2-di-O-Acetyl-4-O-benzyl-3-O-methyl-α-D-rhamnopyranose 10
To a solution of 9 (0.5 g, 1.54 mmol) in Ac2O and AcOH (1:1 v/v; 6 mL) was added
concentrated H2SO4 (0.1 mL) at 0°C. The reaction mixture was stirred until TLC
(petroleum ether/ethyl acetate, 4:1) indicated that 9 was completely disappeared.
The mixture was diluted with CH2Cl2 (100 mL), and was then washed succes-
sively with saturated NaHCO3, water, and brine. The organic phase was dried with
Na2SO4, and concentrated. The residue was subjected to silica gel column chro-
matography (petroleum ether/ethyl acetate, 4:1) to give 10 (0.46 g, 85%) as a white
foamy solid. 1H NMR (600 MHz, CDCl3): δ 7.36–7.33 (m, 4H, ArH), 7.32–7.27 (m,
1H, ArH), 5.98 (d, J = 1.8 Hz, 1H, H-1), 5.28 (dd, J = 3.0, 1.8 Hz, 1H, H-2), 4.89
(d, J = 10.8 Hz, 1H, -CH2Ph), 4.62 (d, J = 10.8 Hz, 1H, -CH2Ph), 3.77 (m, 1H, H-
5), 3.65 (dd, J = 9.6, 3.0 Hz, 1H, H-3), 3.45 (s, 3H, -OCH3), 3.39 (t, J = 9.6 Hz,
1H, H-4), 2.16 (s, 3H, -COCH3), 2.09 (s, 3H, -COCH3), 1.32 (d, J = 6.0 Hz, 3H, H-
6); 13C NMR (150 MHz, CDCl3): δ 170.05, 168.55, 138.24, 128.42, 127.96, 127.81,

8
X. ZHANG ET AL.
90.96, 79.71, 79.48, 75.50, 69.81, 67.35, 57.70, 20.94, 20.91, 17.97; ESI HRMS: calcd
for (C18H24O7+Na+) m/z, 375.1414; found, 375.1400.
2-O-Acetyl-4-O-benzyl-3-O-methyl-α-D-rhamnopyranosyl trichloroacetimidate 4
To a solution of 10 (510 mg, 1.45 mmol) in DMF (5 mL) was added hydrazine
acetate (400 mg, 4.35 mmol) at 0°C. The reaction mixture was stirred at these con-
dition for 3 h, at which time TLC (petroleum ether/ethyl acetate, 3:1) showed the
completion of reaction. The mixture was diluted with EtOAc (150 mL) and then
washed with water and brine. The organic phase was dried over anhydrous Na2SO4
and concentrated, and the residue was purified by silica gel column chromatogra-
phy (petroleum ether/ethyl acetate, 3:1). The product generated above was then dis-
solved in anhydrous CH2Cl2 (5 mL), and to the solution was added trichloroace-
tonitrile (0.57 mL, 5.64 mmol) and DBU (100 µL) at 0°C. The resulting mixture
was stirred for 30 min and concentrated, and the residue was purified by column
chromatography (petroleum ether/ethyl acetate, 4:1) to afford 4 (430 mg, 73% for
two steps) as a white foamy solid. 1H NMR (600 MHz, CDCl3): δ 8.67 (s, 1H, NH),
7.37–7.32 (m, 4H, ArH), 7.31–7.27 (m, 1H, ArH), 6.18 (d, J = 1.8 Hz, 1H, H-1),
5.46 (dd, J = 3.0, 1.8 Hz, 1H, H-2), 4.90 (d, J = 10.8 Hz, 1H, -CH2Ph), 4.63 (d, J =
10.8 Hz, 1H, -CH2Ph), 3.93 (m, 1H, H-5), 3.72 (dd, J = 9.6, 3.0 Hz, 1H, H-3), 3.46
(s, 3H, -OCH3), 3.44 (t, J = 9.6 Hz, 1H, H-4), 2.18 (s, 3H, -COCH3), 1.34 (d, J =
6.0 Hz, 3H, H-6); 13C NMR (150 MHz, CDCl3): δ 170.03, 160.10, 138.19, 128.42,
128.07, 127.85, 95.05, 90.84, 79.78, 79.33, 75.55, 70.51, 67.05, 57.76, 20.97, 17.96.
para-Tolyl 2,3-O-isopropylidene-1-thio-α-D-rhamnopyranoside 12
To a solution of 11 (4.0 g, 8.33 mmol) in THF (20 mL) was added a mixture of
LiAlH4 (0.8 g, 21.0 mmol) in THF (10 mL) in portions at 0°C. The mixture was
warmed up to 65°C and stirred for 3 h, when TLC (petroleum ether/ethyl acetate,
3:1) indicated the disappearance of 11. The reaction mixture was diluted with EtOAc
(150 mL) and washed with 1 M H2SO4 (2× 100 mL). The organic phase was dried
over Na2SO4 and concentrated, and the residue was purified by column chromatog-
raphy (petroleum ether/ethyl acetate, 4:1) to afford 12 (1.88 g, 73%) as a white foamy
solid. 1H NMR (600 MHz, CDCl3): δ 7.35 (d, J = 7.8 Hz, 2H, ArH), 7.12 (d, J =
7.8 Hz, 2H, ArH), 5.66 (s, 1H, H-1), 4.33 (d, J = 5.4 Hz, 1H, H-2), 4.13–4.04 (m,
2H, H-3, H-4), 3.48–3.41 (m, 1H, H-5), 2.54 (d, J = 3.6 Hz, 1H, -OH), 2.32 (s, 3H,
-SPhCH3), 1.52 [s, 3H, -C(CH3)2], 1.36 [s, 3H, -C(CH3)2], 1.24 (d, J = 6.0 Hz, 3H,
H-6); 13C NMR (150 MHz, CDCl3): δ 137.89, 132.52, 129.81, 129.47, 109.70, 84.01,
78.37, 76.48, 75.17, 66.81, 28.14, 26.37, 21.13, 17.06.
para-Tolyl 4-O-benzoyl-2,3-O-isopropylidene-1-thio-α-D-rhamnopyranoside 13
To a solution of 12 (1.7 g, 5.48 mmol) in pyridine (10 mL) was added BzCl (0.76 mL,
6.58 mmol) dropwise at 0°C. The reaction mixture was stirred at rt for 2 h, and then
diluted with CH2Cl2 (100 mL), washed with 1 M HCl and brine. The organic layer

JOURNAL OF CARBOHYDRATE CHEMISTRY
9
was then dried over anhydrous Na2SO4. After concentration, the residue was puri-
fied by flash column chromatography (petroleum ether/ethyl acetate, 3:1) to afford
13 (1.9 g, 84%) as a white foamy solid. 1H NMR (600 MHz, CDCl3): δ 8.07 (d, J =
7.8 Hz, 2H, ArH), 7.57 (t, J = 7.8 Hz, 1H, ArH), 7.45 (t, J = 7.8 Hz, 2H, ArH), 7.38
(d, J = 7.8 Hz, 2H, ArH), 7.14 (d, J = 7.8 Hz, 2H, ArH), 5.73 (s, 1H, H-1), 5.19 (t,
J = 9.0 Hz, 1H, H-4), 4.43–4.31 (m, 3H, H-2, H-3, H-5), 2.33 (s, 3H, -SPhCH3),
1.62 [s, 3H, -C(CH3)2], 1.36 [s, 3H, -C(CH3)2], 1.17 (d, J = 6.6 Hz, 3H, H-6);
13C NMR (150 MHz, CDCl3): δ 165.71, 138.06, 133.21, 132.53, 129.89, 129.83,
129.70, 129.27, 128.37, 110.07, 84.10, 76.47, 75.60, 75.13, 65.65, 27.75, 26.52, 21.14,
16.95.
para-Tolyl 4-O-benzoyl-1-thio-α-D-rhamnopyranoside 14
After a solution of 13 (1.8 g, 4.34 mmol) in 80% TFA (5 mL) was stirred at rt for
1 h, the mixture was co-evaporated with toluene (2× 20 mL) twice under reduced
pressure. The residue was purified by flash column chromatography (petroleum
1
ether/ethyl acetate, 2:1) to yield 14 (1.4 g, 86%) as a white foamy solid. H NMR
(600 MHz, CDCl3): δ 8.07 (d, J = 8.4Hz, 2H, ArH), 7.61 (t, J = 7.8Hz, 1H, ArH), 7.47
(t, J = 7.8 Hz, 2H, ArH), 7.38 (d, J = 7.8 Hz, 2H, ArH), 7.14 (d, J = 7.8 Hz, 2H, ArH),
5.50 (s, 1H, H-1), 5.08 (t, J = 9.6 Hz, 1H, H-4), 4.49 (m, 1H, H-5), 4.25 (br s, 1H,
H-2), 4.08–4.03 (m, 1H, H-3), 3.31 (d, J = 5.4 Hz, 1H, -OH), 2.74 (br s, 1H, -OH),
2.33 (s, 3H, -SPhCH3),1.30 (d, J = 6.6 Hz, 3H, H-6); 13C NMR (150 MHz, CDCl3):
δ 167.69, 137.90, 133.66, 132.06, 129.92, 129.81, 129.19, 128.54, 87.49, 76.62, 72.24,
70.89, 66.91, 21.12, 17.49; ESI HRMS: calcd for (C20H22O5S+Na+) m/z, 397.1080;
found, 397.1074.
para-Tolyl 2,4-di-O-benzoyl-1-thio-α-D-rhamnopyranoside 5
To a stirred solution of 14 (1.40 g, 3.74 mmol) in CH2Cl2 (5 mL) was added trimethyl
orthobenzoate (1.44 mL, 7.48 mmol). The mixture was treated with a catalytic
amount of (+)-10-camphorsulfonic acid at 25°C for 30 min, and then the solution
was concentrated. To the residue was added 80% HOAc, and after stirring 10 min,
the mixture was concentrated. The residue was purified by flash column chromatog-
1
raphy (petroleum ether/ethyl acetate, 3:1) to yield 5 (1.45 g, 81%) as a syrup. H
NMR (600 MHz, CDCl3): δ 8.12–8.07 (m, 4H, ArH), 7.63–7.57 (m, 2H, ArH), 7.48
(t, J = 7.2 Hz, 1H, ArH), 7.47 (t, J = 7.2 Hz, 1H, ArH), 7.40 (d, J = 7.8 Hz, 2H, ArH),
7.13 (d, J = 7.8 Hz, 2H, ArH), 5.64 (dd, J = 3.6, 1.8 Hz, 1H, H-2), 5.58 (d, J = 1.2 Hz,
1H, H-1), 5.32 (t, J = 9.6 Hz, 1H, H-4), 4.57 (m, 1H, H-5), 4.33–4.27 (m, 1H, H-3),
2.52 (d, J = 7.8 Hz, 1H, -OH), 2.32 (s, 3H, -SPhCH3), 1.34 (d, J = 6.0 Hz, 3H, H-
6); 13C NMR (150 MHz, CDCl3): δ 167.15, 165.87, 138.16, 133.58, 133.56, 132.40,
129.95, 129.88, 129.87, 129.84, 129.61, 129.30, 129.28, 128.57, 128.54, 128.06, 86.21,
75.74, 74.78, 69.70, 67.42, 21.12, 17.53; ESI HRMS: calcd for (C27H26O6S+Na+)
m/z, 501.1342; found, 501.1354.

10
X. ZHANG ET AL.
para-Tolyl 2-O-acetyl-4-O-benzyl-3-O-methyl-α-D-rhamnopyranosyl-(1→3)-2,4-
di-O-benzoyl-1-thio-α-D-rhamnopyranoside 15
To a solution of 4 (500 mg, 1.103 mmol) and 5 (350 mg, 0.736 mmol) in anhy-
drous CH2Cl2 (5 mL) was added TMSOTf (20 µL, 0.11 mmol) at −78°C under a
N2 atmosphere. After the mixture was stirred at −78°C for 2 h, it was neutralized
with Et3N and then concentrated. The residue was purified by column chromatogra-
phy (petroleum ether/ethyl acetate, 5:1) to yield 15 (400 mg, 71%) as a white foamy
solid. 1H NMR (600 MHz, CDCl3): δ 8.11 (d, J = 7.8 Hz, 2H, ArH), 8.10 (d, J =
7.8 Hz, 2H, ArH), 7.62–7.56 (m, 2H, ArH), 7.50–7.44 (m, 4H, ArH), 7.40 (d, J =
7.8 Hz, 2H, ArH), 7.29–7.21 (m, 3H, ArH), 7.16 (d, J = 7.2 Hz, 2H, ArH), 7.13 (d,
J = 7.8 Hz, 2H, ArH), 5.70 (dd, J = 3.0, 1.8 Hz, 1H, H-2), 5.56 (d, J = 1.2 Hz, 1H,
H-1), 5.54 (t, J = 9.6 Hz, 1H, H-4), 4.91 (br s, 2H, H-1´, H-2´), 4.71 (d, J = 11.4 Hz,
1H, -CH2Ph), 4.51 (m, 1H, H-5), 4.45 (d, J = 11.4 Hz, 1H, -CH2Ph), 4.32 (dd,
J = 9.6, 3.0 Hz, 1H, H-3), 3.69 (m, 1H, H-5´), 3.39 (dd, J = 9.6, 2.4 Hz, 1H, H-
3), 3.19 (t, J = 9.6 Hz, 1H, H-4´), 3.01 (s, 3H, -OCH3), 2.32 (s, 3H, -SPhCH3), 1.94
(s, 3H, -COCH3), 1.33 (d, J = 6.0 Hz, 3H, H-6), 1.08 (d, J = 6.6 Hz, 3H, H-6´); 13
C
NMR (150 MHz, CDCl3): δ 169.78, 165.70, 165.68, 138.61, 138.17, 133.45, 133.42,
132.43, 129.96, 129.89, 129.79, 129.51, 129.35, 128.58, 128.54, 128.14, 127.40, 127.35,
99.23 (C-1´), 86.07 (C-1), 79.51 (C-4´), 78.96 (C-3´), 76.19 (C-3), 74.34 (-CH2Ph),
73.53 (C-2), 73.33 (C-4), 68.68 (C-5´), 68.45 (C-2´), 67.92 (C-5), 57.05 (-OCH3),
21.13 (-SPhCH3), 20.79 (-OCCH3), 17.66 (C-6´), 17.55 (C-6); ESI HRMS: calcd for
(C43H46O11S+NH4+) m/z, 788.3099; found, 788.3112.
3-Azidopropanyl 2-O-acetyl-4-O-benzyl-3-O-methyl-α-D-rhamnopyranosyl-
(1→3)-2,4- di-O-benzoyl-α-D-rhamnopyranoside 16
To a solution of 15 (330 mg, 0.428 mmol) and 3-azidopropanol (215 mg, 2.14 mmol)
in anhydrous CH2Cl2 (10 mL) was added NIS (150 mg, 0.652 mmol) and TMSOTf
(10 µL, 0.055 mmol) at −15°C under a N2 atmosphere. The reaction mixture was
slowly heated to 40°C and stirred for 4 h, at which time TLC indicated the com-
plete disappearance of 15. The mixture was then neutralized with Et3N and concen-
trated, and the residue was purified by flash column chromatography (petroleum
ether/ethyl acetate, 2:1) to yield 16 (240 mg, 75%) as a white foamy solid.1H NMR
(600 MHz, CDCl3): δ 8.12 (d, J = 7.8 Hz, 2H, ArH), 8.08 (d, J = 7.8 Hz, 2H, ArH),
7.62–7.55 (m, 2H, ArH), 7.51–7.43 (m, 4H, ArH), 7.29–7.19 (m, 3H, ArH), 7.14 (d,
J = 7.2 Hz, 2H, ArH), 5.48 (t, J = 9.6 Hz, 1H, H-4), 5.44 (br s, 1H, H-2), 4.95 (s,
1H, H-1), 4.88 (br s, 1H, H-1´), 4.87 (br s, 1H, H-2´), 4.69 (d, J = 11.4 Hz, 1H, -
CH2Ph), 4.43 (d, J = 11.4 Hz, 1H, -CH2Ph), 4.31 (dd, J = 9.6, 3.0 Hz, 1H, H-3),
3.99 (m, 1H, H-5), 3.84 (m, 1H, -OCH2CH2-), 3.67 (m, 1H, H-5), 3.56 (m, 1H, -
OCH2CH2-), 3.46 (t, J = 6.6 Hz, 2H, -CH2CH2N3), 3.38 (dd, J = 9.6, 3.0 Hz, 1H, H-
3´), 3.17 (t, J = 9.6 Hz, 1H, H-4´), 2.98 (s, 3H, -OCH3), 1.97–1.87 (m, 5H, -COCH3,
-OCH2CH2CH2N3), 1.32 (d, J = 6.0 Hz, 3H, H-6), 1.05 (d, J = 6.0 Hz, 3H, H-
6´); 13C NMR (150 MHz, CDCl3): δ 169.82, 165.85, 165.72, 138.62, 133.44, 133.37,
129.91, 129.75, 129.42, 129.38, 128.58, 128.52, 128.13, 127.35, 127.32, 99.20 (C-1´),

JOURNAL OF CARBOHYDRATE CHEMISTRY
11
97.28 (C-1), 79.53 (C-4´), 78.92 (C-3´), 76.01(C-3), 74.27 (-CH2Ph), 73.06 (C-4),
71.92 (C-2), 68.57 (C-5´), 68.53 (C-2´), 66.79 (C-5), 64.79 (-OCH2CH2-), 56.99 (-
OCH3), 48.32 (-CH2CH2N3), 28.80 (-OCH2CH2CH2N3), 20.80 (-COCH3), 17.66
(C-6´), 17.64 (C-6); ESI HRMS: calcd for (C39H45N3O12+NH4+) m/z, 765.3341;
found, 765.3352.
3-Azidopropanyl 4-O-benzyl-3-O-methyl-α-D-rhamnopyranosyl-(1→3)-2,4-di-O-
benzoyl-α-D-rhamnopyranoside 3
To a solution of 16 (230 mg, 0.308 mmol) in MeOH and CH2Cl2 (v/v 1:1, 8 mL)
was added acetyl chloride (0.4 mL) at 0°C. The solution was stirred at rt overnight,
and then neutralized with saturated NaHCO3, extracted with CH2Cl2 (2× 100 mL).
The organic phase was dried over anhydrous Na2SO4 and concentrated. The residue
was purified by flash column chromatography (toluene/ethyl acetate, 2:1) to afford
1
3 (185 mg, 85%) as a white foamy solid. H NMR (600 MHz, CDCl3): δ 8.12 (d,
J = 7.2 Hz, 2H, ArH), 8.06 (d, J = 7.8 Hz, 2H, ArH), 7.59 (t, J = 7.2 Hz, 2H, ArH),
7.48 (d, J = 7.2 Hz, 2H, ArH), 7.47 (d, J = 7.2 Hz, 2H, ArH), 7.30–7.21 (m, 3H,
ArH), 7.16 (d, J = 7.2 Hz, 2H, ArH), 5.47 (t, J = 9.6 Hz, 1H, H-4), 5.43 (br s, 1H,
H-2), 4.94 (s, 1H, H-1), 4.93 (s, 1H, H-1´), 4.66 (d, J = 11.4 Hz, 1H, -CH2Ph), 4.46
(d, J = 11.4 Hz, 1H, -CH2Ph), 4.33 (dd, J = 9.6, 1.8 Hz, 1H, H-3), 3.99 (m, 1H, H-
5), 3.85 (m, 1H, -OCH2CH2-), 3.67 (m, 1H, H-5´), 3.65 (br s, 1H, H-2´), 3.56 (m,
1H, -OCH2CH2-), 3.47 (t, J = 6.6 Hz, 2H, -CH2CH2N3), 3.28 (dd, J = 9.6, 2.4 Hz,
1H, H-3´), 3.18 (t, J = 9.6 Hz, 1H, H-4´), 3.13 (s, 3H, -OCH3), 1.99–1.87 (m, 2H, -
OCH2CH2CH2N3), 1.30 (d, J = 6.0 Hz, 3H, H-6), 1.10 (d, J = 6.6 Hz, 3H, H-6´); 13C
NMR (150 MHz, CDCl3): δ 165.87, 165.69, 138.57, 133.50, 133.41, 129.94, 129.63,
129.45, 129.39, 129.02, 128.64, 128.55, 128.22, 128.17, 127.45, 127.37, 101.11 (C-1´),
97.31 (C-1), 80.70 (C-3´), 79.25 (C-4´), 75.40 (C-3), 74.24 (-CH2Ph), 73.38 (C-4),
72.27 (C-2), 68.09 (C-5´), 67.85 (C-2´), 66.77 (C-5), 64.76 (-OCH2-), 57.07 (-OCH3),
48.33 (-CH2N3), 28.78 (-OCH2CH2CH2N3), 17.67 (C-6´), 17.63 (C-6); ESI HRMS:
calcd for (C37H43N3O11+NH4+) m/z, 723.3236; found, 723.3241.
3-Azidopropanyl 2-O-acetyl-3,4,6-tri-O-benzyl-α-D-mannopyranosyl-(1→2)-3,4,
6-tri-O-benzyl-α-D-mannopyranosyl-(1→2)-4-O-benzyl-3-O-methyl-α-D-
rhamnopyranosyl-(1→3)-2,4-di-O-benzoyl-α-D-rhamnopyranoside 17
Method A. To a solution of 2 (110 mg, 0.107 mmol) and 3 (50 mg, 0.071 mmol) in
anhydrous CH2Cl2 (5 mL) was added NIS (31mg, 0.138 mmol) and TMSOTf (2 µL,
0.011 mmol) at −15°C under a N2 atmosphere. The reaction mixture was stirred at
these conditions for 2 h, at which time TLC (petroleum ether/ethyl acetate, 3:1) indi-
cated the complete disappearance of 3. The mixture was then neutralized with Et3N
and concentrated, and the residue was purified by flash column chromatography
(petroleum ether/ethyl acetate, 3:1) to yield the desired terasaccharide 17 (57 mg,
50%) and its isomer 17β (38 mg, 23%) as white foamy solids.
Method B. To a solution of 18 (64 mg, 0.10 mmol) and 20 (75 mg, 0.066 mmol) in
anhydrous CH2Cl2 (3 mL) was added TMSOTf (2 µL, 0.011 mmol) at −15°C under

12
X. ZHANG ET AL.
a N2 atmosphere. The reaction mixture was stirred at these conditions for 2 h, then
neutralized with Et3N and concentrated. The residue was purified by flash column
chromatography (petroleum ether/ethyl acetate, 3:1) to yield 17 (86 mg, 81%).
1
Physical data for 17: H NMR (600 MHz, CDCl3): δ 8.11 (d, J = 7.8 Hz, 2H,
ArH), 8.02 (d, J = 7.8 Hz, 2H, ArH), 7.58 (t, J = 7.2 Hz, 1H, ArH), 7.46 (t, J =
7.8 Hz, 2H, ArH), 7.35–7.18 (m, 32 H, ArH), 7.16–7.09 (m, 6H, ArH), 5.52 (br s,
1H, H-2´´´), 5.42 (br s, 1H, H-2), 5.41 (t, J = 9.6 Hz, 1H, H-4), 5.08 (s, 1H, H-1´´´),
5.03 (s, 1H, H-1´), 4.92 (s, 1H, H-1), 4.84 (d, J = 10.8 Hz, 1H, -CH2Ph), 4.79 (d, J =
10.8 Hz, 1H, -CH2Ph), 4.69 (s, 1H, H-1´´), 4.68–4.56 (m, 6H, -CH2Ph), 4.49–4.37
(m, 5H, -CH2Ph), 4.30 (d, J = 12.6 Hz, 1H, -CH2Ph), 4.27 (dd, J = 9.6, 3.0 Hz,
1H, H-3), 3.99 (br s, 1H, H-2´´), 3.95–3.87 (m, 3H, H-5, H-3´´´, H-5´´´), 3.80–
3.73 (m, 3H, H-3´´, H-5´´, -OCH2CH2-), 3.70 (dd, J = 10.8, 3.6 Hz, 1H, H-6a´´´),
3.67–3.62 (m, 2H, H-4´´, H-4´´´), 3.60–3.42 (m, 5H, H-2´, H-5´, H-6b´´, H-6a´´´,
-OCH2CH2-), 3.35 (t, J = 6.6 Hz, 2H, -CH2CH2N3), 3.31 (br d, J = 10.8 Hz, 1H,
H-6b´´´), 3.28 (dd, J = 9.6, 3.0 Hz, 1H, H-3´), 3.08 (t, J = 9.6 Hz, 1H, H-4´), 2.92
(s, 3H, -OCH3), 2.10 (s, 3H, -COCH3), 1.82 (m, 2H, -OCH2CH2CH2N3), 1.26 (d, J
= 6.0 Hz, 3H, H-6), 1.03 (d, J = 6.0 Hz, 3H, H-6´); 13C NMR (150 MHz, CDCl3): δ
170.12, 165.85, 138.80, 138.58, 138.40, 138.31, 138.24, 137.99, 137.96, 133.65, 133.35,
129.90, 129.67, 129.47, 129.38, 128.59, 128.53, 128.34, 128.31, 128.30, 128.28, 128.26,
128.20, 128.16, 128.09, 127.99, 127.81, 127.80, 127.64, 127.59, 127.50, 127.47, 127.37,
127.21, 100.75 (C-1´), 100.49 (C-1´´), 99.13 (C-1´´´), 97.33 (C-1), 80.59 (C-3´), 79.55
(C-4´), 79.08 (C-3´´), 78.06 (C-3´´´), 75.87 (C-3), 75.15 (CH2Ph), 74.96 (CH2Ph),
74.45 (C-5´´), 74.35 (C-2´), 74.01 (C-5´´´), 73.91 (CH2Ph), 73.76 (C-2´´), 73.44
(CH2Ph), 73.15 (C-2), 73.03 (CH2Ph), 72.24 (C-4), 72.12 (CH2Ph), 71.99 (CH2Ph),
71.77 (2C, C-4´´ and C-4´´´), 68.76 (C-6´´´), 68.61 (C-5´), 68.46 (C-2´´´), 68.31
(C-6´´), 66.86 (C-5), 64.80 (-OCH2CH2-), 56.93 (-OCH3), 48.21 (-CH2CH2N3),
28.73 (-OCH2CH2CH2N3), 21.15 (-COCH3), 17.72 (C-6), 17.70 (C-6´); ESI HRMS:
calcd for (C93H101N3O22+NH4+) m/z, 1629.7215; found, 1629.7238.
3-Azidopropanyl 2-O-Acetyl-3,4,6-tri-O-benzyl-α-D-mannopyranosyl-(1→2)-4-
O-benzyl-3-O-methyl-α-D-rhamnopyranosyl-(1→3)-2,4-di-O-benzoyl-α-D-
rhamnopyranoside 19
To a solution of 18 (200 mg, 0.32 mmol) and 3 (150 mg, 0.213 mmol) in anhy-
drous CH2Cl2 (4 mL) was added TMSOTf (6 µL, 0.032 mmol) at −15°C under a N2
atmosphere. After the mixture was stirred at −15°C for 3 h, it was neutralized with
Et3N and then concentrated. The residue was purified by column chromatography
(petroleum ether/ethyl acetate, 3:1) to yield 19 (216 mg, 86%) as a white foamy solid.
1H NMR (600 MHz, CDCl3): δ 8.13 (d, J = 8.4 Hz, 2H, ArH), 8.06 (d, J = 7.8 Hz,
2H, ArH), 7.60 (t, J = 7.2 Hz, 1H, ArH), 7.51–7.44 (m, 3H, ArH), 7.41–7.19 (m,
23H, ArH), 7.15 (d, J = 6.6 Hz, 2H, ArH), 7.12 (d, J = 6.6 Hz, 2H, ArH), 5.45 (t, J =
10.2 Hz, 1H, H-4), 5.43 (s, 1H, H-2), 5.34 (s, 1H, H-2´´), 5.03 (s, 1H, H-1´), 4.94 (s,
1H, H-1), 4.81 (s, 1H, H-1´´), 4.79 (d, J = 10.8 Hz, 1H, -CH2Ph), 4.71–4.62 (m, 3H,

JOURNAL OF CARBOHYDRATE CHEMISTRY
13
-CH2Ph), 4.51–4.34 (m, 4H, -CH2Ph), 4.28 (dd, J = 10.2, 3.6 Hz, 1H, H-3), 3.99–
3.93 (m, 1H, H-5), 3.87 (dd, J = 9.6, 3.0 Hz, 1H, H-3´´), 3.84–3.77 (m, 2H, H-4´´,
-OCH2CH2-), 3.65 (br s, 1H, H-2´), 3.64–3.57 (m, 2H, H-5´, H-5´´), 3.56–3.49 (m,
2H, H-6a´´, -OCH2CH2-), 3.39 (t, J = 6.6 Hz, 2H, -CH2CH2N3), 3.36 (dd, J = 9.6,
3.0 Hz, 1H, H-3´), 3.29 (br d, J = 10.2 Hz, 1H, H-6b´´), 3.17 (t, J = 9.6 Hz, 1H, H-4´),
3.05 (s, 3H, -OCH3), 2.11 (s, 3H, -COCH3), 1.91–1.81 (m, 2H, -OCH2CH2CH2N3),
1.29 (d, J = 6.0 Hz, 3H, H-6), 1.02 (d, J = 6.0 Hz, 3H, H-6´); 13C NMR (150MHz,
CDCl3): δ 170.16, 165.88, 165.65, 138.77, 138.36, 138.23, 137.87, 133.42, 129.92,
129.67, 129.46, 129.43, 128.68, 128.57, 128.35, 128.28, 128.14, 128.09, 127.89, 127.72,
127.62, 127.41, 100.46 (C-1´), 98.99 (C-1´´), 97.29 (C-1), 80.62 (C-3´), 79.23 (C-
4´), 77.75 (C-3´´), 75.81 (C-3), 75.10 (CH2Ph), 74.06 (C-4´´), 73.94 (C-2´), 73.15
(CH2Ph), 73.12 (CH2Ph), 72.15 (C-2), 71.79 (C-4), 71.67 (CH2Ph), 68.81 (C-5´),
68.62 (C-2´´), 68.29 (C-6´´), 66.78 (C-5), 64.79 (-OCH2CH2-), 57.11 (-OCH3), 48.26
(-CH2CH2N3), 28.75 (-OCH2CH2CH2N3), 21.15 (-COCH3), 17.70 (2C, C-6, C-6´);
ESI HRMS: calcd for (C66H73N3O17+NH4+) m/z, 1197.5278; found, 1197.5212;
calcd for (C66H73N3O17+Na+) m/z, 1202.4832; found, 1202.4814.
3-Azidopropanyl 3,4,6-tri-O-benzyl-α-D-mannopyranosyl-(1→2)-4-O-benzyl-3-
O-methyl-α-D-rhamnopyranosyl-(1→3)-2,4-di-O-benzoyl-α-D-
rhamnopyranoside 20
To a solution of 19 (196 mg, 0.166 mmol) in MeOH and CH2Cl2 (v/v 1:1, 10 mL)
was added acetyl chloride (0.5 mL) at 0°C. The solution was stirred at rt overnight,
and then neutralized with saturated NaHCO3, extracted with CH2Cl2 (2× 80 mL).
The organic phase was dried over anhydrous Na2SO4 and concentrated. The residue
was purified by column chromatography (petroleum ether/ethyl acetate, 3:1) to
1
afford 20 (156 mg, 83%) as a white foamy solid. H NMR (600 MHz, CDCl3):
δ 8.13 (d, J = 7.8 Hz, 2H, ArH), 8.07 (d, J = 7.2 Hz, 2H, ArH), 7.60 (t, J =
7.2 Hz, 1H, ArH), 7.51–7.45 (m, 3H, ArH), 7.41 (t, J = 7.8 Hz, 2H, ArH), 7.38–
7.20 (m, 18H, ArH), 7.13 (t, J = 7.8 Hz, 3H, ArH), 5.47–5.42 (m, 2H, H-2, 4), 5.07
(s, 1H, H-1´), 4.94 (s, 1H, H-1), 4.81 (s, 1H, H-1´´), 4.77 (d, J = 10.8 Hz, 1H, -
CH2Ph), 4.67–4.61 (m, 4H, -CH2Ph), 4.44 (d, J = 12.6 Hz, 1H, -CH2Ph), 4.41 (d,
J = 10.8 Hz, 1H, -CH2Ph), 4.40 (d, J = 11.4 Hz, 1H, -CH2Ph), 4.29 (dd, J = 9.6,
3.0 Hz, 1H, H-3), 3.98 (s, 1H, H-2´´), 3.98–3.93 (m, 1H, H-5), 3.83–3.73 (m, 3H,
H-3´´, H-4´´, -OCH2CH2-), 3.71–3.64 (m, 2H, H-2´, H-5´´), 3.62–3.56 (m, 1H, H-
5´), 3.56–3.48 (m, 2H, -OCH2CH2-, H-6a´´), 3.40–3.33 (m, 4H, H-3´, H-6b´´, -
CH2CH2N3), 3.16–3.09 (t, J = 9.6 Hz, 1H, H-4´), 3.02 (s, 3H, -OCH3), 2.39 (d, J
= 2.7 Hz, 1H, -OH), 1.89–1.81 (m, 2H, -OCH2CH2CH2N3), 1.29 (d, J = 6.2 Hz,
3H, H-6), 1.02 (d, J = 6.2 Hz, 3H, H-6´);13C NMR (150 MHz, CDCl3): δ 165.89,
165.80, 138.79, 138.25, 138.20, 137.85, 133.52, 133.40, 129.92, 129.70, 129.49, 129.44,
128.71, 128.57, 128.49, 128.38, 128.33, 128.08, 127.91, 127.89, 127.79, 127.67, 127.61,
127.25, 127.22, 100.74 (C-1´´), 100.59 (C-1´), 97.31 (C-1), 80.57 (C-3´), 79.57 (C-
3´´), 79.52 (C-4´), 76.03 (C-3), 75.03 (CH2Ph), 74.28 (C-2´), 74.12 (C-4´), 73.85
(CH2Ph), 73.14 (CH2Ph), 73.11 (C-2), 72.17 (C-4), 72.04 (CH2Ph), 71.30 (C-5´´),

14
X. ZHANG ET AL.
68.71 (C-5´), 68.47 (C-6´´), 68.36 (C-2´´), 66.81 (C-5), 64.82 (-OCH2CH2-), 57.05
(-OCH3), 48.25 (-CH2CH2N3), 28.74 (-OCH2CH2CH2N3), 17.71 (2C, C-6, C-6´);
ESI HRMS: calcd for (C64H71N3O16+Na+) m/z, 1160.4727; found, 1160.4701; calcd
for (C64H71N3O16+K+) m/z, 1176.4471; found, 1176.4456.
3-Aminopropanyl α-D-Mannopyranosyl-(1→2)-α-D-Mannopyranosyl-(1→2)-3-
O-methyl-α-D-rhamnopyranosyl-(1→3)-α-D-rhamnopyranoside 1
To a solution of 17 (50 mg, 0.031 mmol) in MeOH (5 mL) was added 1 M NaOMe
in MeOH dropwise until the pH value reached 10. The mixture was stirred at
rt for 24 h, and then neutralized with Amberlite IR 120 (H+). The solution was
filtered and concentrated. The residue was purified by flash column chromatog-
raphy (petroleum ether/ethyl acetate, 1:1). After the above generated product was
re-dissolved in HOAc and H2O (v/v 2:1, 5 mL), 10% Pd/C (10 mg) was added. The
suspension was stirred under a H2 atmosphere at rt overnight. The solid materials
were filtrated off, and the resulting solution was concentrated. The residue was
purified by size exclusion chromatography on a Bio-Gel P-2 column with distilled
water as the eluent and lyophilized to give 1 (14 mg, 63% for 2 steps) as a white
1
solid. H NMR (600 MHz, D2O): δ 5.10 (s, 1H, H-1), 5.09 (s, 1H, H-1), 4.85 (s,
1H, H-1), 4.58 (s, 1H, H-1), 4.15 (br s, 1H, H-2), 3.87 (br s, 2H, 2× H-2), 3.84
(s, 1H, H-2), 3.74 (dd, J = 9.0, 2.4 Hz, 1H, H-3), 3.70–3.62 (m, 5H, H-3, H-5, 2×
H-6, -OCH2CH2-), 3.61–3.55 (m, 4H, H-3, H-4, 2× H-6), 3.54–3.44 (m, 5H, H-3,
H-4, 3× H-5), 3.42–3.31 (m, 3H, 2× H-4, -OCH2CH2-), 3.30 (s, 3H, -OCH3),
3.00–2.88 (m, 2H, -CH2CH2NH2), 1.85–1.77 (m, 2H, -OCH2CH2CH2NH2), 1.11
(d, J = 5.4 Hz, 3H, H-6), 1.10 (d, J = 6.0 Hz, 3H, H-6); 13C NMR (150 MHz, D2O):
δ 102.12, 100.69, 100.33, 99.51, 79.16, 78.42, 78.25, 74.24, 73.19, 72.99, 71.22, 71.03,
70.18, 69.84, 69.81, 69.71, 69.14, 68.67, 66.59, 64.73, 60.80, 60.70, 56.77, 37.32,
26.54, 22.92, 16.54, 16.47; ESI HRMS: calcd for (C28H51NO19+H+) m/z, 706.3128;
found, 706.3132; calcd for (C28H51NO19+Na+) m/z, 728.2947; found, 728.2941.
Acknowledgment
This research was supported by grants from the Shandong Provincial Natural Science Founda-
tion (No. ZR2015BM020) and the Science and Technology Development Project of Shandong
Province (No. 2015GSF118019 and 2016GGH4502).
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