An Approach to Synthesize Chondroitin Sulfate-E (CS-E) Oligosaccharide Precursors

Article abstract of DOI:10.1021/acs.joc.8b00157

An approach was developed to synthesize chondroitin sulfate-E (CS-E) oligosaccharides by adopting a postglycosylation-transformation strategy: different from all of the traditional approaches, the characteristic groups of CS-E were introduced following the assembly of the oligosaccharides. The adjusted strategy rendered an easy chain elongation strategy. All of the elongation steps generated high yields with excellent glycosylation outcomes. An orthogonally protected disaccharide was used as the building block to provide flexibility for the group transformation and derivatization at the N-2 position of the GalNAc residue and the O-1,5 positions of the GlcA residue, thereby providing ready access for the further examination of the structure-activity relationship (SAR) of CS-E molecules.

Full text of DOI:10.1021/acs.joc.8b00157

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Article
An Approach to Synthesize Chondroitin
Sulfate-E (CS–E) Oligosaccharides Precursors
Shuang Yang, Qi Liu, Guangyan Zhang, Xiaoxi Zhang, Zhehui Zhao, and Pingsheng Lei
J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.joc.8b00157 • Publication Date (Web): 14 May 2018
Downloaded from http://pubs.acs.org on May 14, 2018
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An Approach to Synthesize Chondroitin SulfateꢀE (CS–E)
Oligosaccharides Precursors
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Shuang Yang, Qi Liu, Guangyan Zhang, Xiaoxi Zhang, Zhehui Zhao∗, Pingsheng Lei∗
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State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia
Medica, Peking Union Medical College and Chinese Academy of Medical Sciences, Beijing, 100050, P.
R. China
*Eꢀmail: zhaozh@imm.ac.cn
ABSTRACT: An approach was developed to synthesize chondroitin sulfateꢀE (CS–E) oligosaccharides
by adopting a postglycosylationꢀtransformation strategy: different from all the traditional approaches,
the characteristic groups of CSꢀE were introduced following the assembly of the oligosaccharides. The
adjusted strategy rendered an easy chain elongation strategy. All the elongation steps generated high
yields with excellent glycosylation outcomes. An orthogonally protected disaccharide was used as the
building block to provide flexibility for the group transformation and derivatization at the Nꢀ2 position
of the GalNAc residue and the Oꢀ1,5 positions of the GlcA residue, thereby providing ready access for
the further examination of the structureꢀactivity relationship (SAR) of CSꢀE molecules.
KEY WORDS: Chondroitin sulfate E, postglycosylationꢀtransformation strategy, oligosaccharides,
glycosylation, modification
ꢀ
INTRODUCTION
Chondroitin sulfate (CS) is a class of sulfated glycosaminoglycans made of dimeric units
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GlcAꢀβ(1→3)ꢀGalNAcꢀβ(1→4). Variable sulfation gives a collection of CS subtypes, among which
CSꢀE is characterized with sulfate motifs located at the 4 and 6 positions of the GalNAc residue. CSꢀE
has been found to play crucial roles in many biological events, such as the development of the central
nervous system, viral attachment, and growth factor signaling.1ꢀ5 CSꢀE has a wide range of biological
activities, thereby making it an attractive target for synthetic organic chemists for the development of
drug candidates. However it is not easy to obtain significant amounts of structurally defined CSꢀE
oligosaccharides from natural sources, due to the low natural content of CSꢀE and the heterogeneity of
the CS polymer. Without a precise molecular structure, accurate pharmacological results are difficult to
achieve. In addition, the structureꢀactivity relationship of CSꢀE has barely been explored systematically,
and the mechanisms underlying its biofunction have remained largely elusive. To address these
problems, wellꢀdefined CSꢀE oligomers and analogues are required. Although several total syntheses of
CSꢀE oligosaccharides have been developed,6ꢀ17 new and efficient approaches available for diverse
derivatization are still required. The present study described such an approach, to provide easy access
to CSꢀE tetrasaccharide and hexasaccharide as well as their derivatives.
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Figure 1. Previously synthesized CSꢀE oligosaccharides.
CSꢀE oligosaccharides have been mainly synthesized by four groups who have adopted different
glycosyl sequences (Figure 1). Both sequences have exhibited attractive bioactivities, though the
superiority of a specific sequence has not been examined. All the synthetic routes were derived from
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the construction of a suitable repeating disaccharide unit, after which the repeating unit was assembled
by stepwise glycosylation. As reported, the yields of these glycosylations were usually raised by
increasing the consumption of disaccharide donors, thereby there is still room to improve the route
economy in CSꢀE oligosaccharides synthesis. Very potent donors are necessary to efficiently utilize the
disaccharide repeating unit in such syntheses. We previously investigated the construction of CSꢀE
disaccharide repeating unit 1 by the postglycosylationꢀoxidation strategy which exhibited excellent
performance in contrast with the traditional preglycosylationꢀoxidation strategy, on both the
glycosylation efficiency and stereoselectivity, this is likely because it benefited from the high reactivity
of the glucosyl monosaccharide donors as compared to the widely used uronic acid donors (Scheme
1).18 With this acquired knowledge, we attempted to employ a new disaccharide repeating unit
containing glucosyl residue rather than uronic acid residue at the reducing terminal, and apply the
postglycosylationꢀoxidation strategy at the oligosaccharide stage, to improve the glycosylation
efficiency in the synthesis of CSꢀE oligomers (Scheme 1).
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Scheme 1. Retrosynthetic Analysis of the CSꢀE Oligosaccharides
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To target a facile approach to derivatization, the repeating unit must be equipped with protecting
groups that can be chemically selectively removed or modified. For this purpose, an orthogonal
protective strategy was designed for the disaccharide unit (Scheme 1). Particularly, azido was selected
as the masked amino group in the GalNAc unit. In spite of its nonꢀparticipatory property, good
βꢀselectivity can also be observed in the glycosylation of 2ꢀazido donor under proper reaction
conditions.19ꢀ23 In addition, smooth glycosylations can also be expected due to the absence of
disturbance of the Nꢀprotecting group.7ꢀ8
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Guided by these ideas, the synthetic route for CSꢀE oligosaccharides was arranged in the
postglycosylationꢀtransformation sequence (Scheme 1). In this approach, the oligosaccharide backbone
was assembled with orthogonally protected disaccharide repeating unit 2, after which introduction of
two characteristic CSꢀE groups, namely a carboxyl moiety and an acetamide group, took place at the
oligosaccharide stage.
ꢀ
RESULTS AND DISCUSSION
Synthesis of the building blocks. Monomers 7 and 8 were prepared individually as glycosyldonor and
acceptor , respectively, to give CSꢀE disaccharide repeating unit by the coupling reaction (Scheme 2).
The synthesis of glycosyl acceptor 8 began with Dꢀglucose monohydrate through a straightforward
sixꢀstep sequence that generated a total yield of 36%.18 The synthesis of glycosyl donor 7 began from
the previously described peracetylated derivative 3 of commercially available Dꢀgalactosamine
hydrochloride.18 3 was reacted with pꢀmethylbenzothiol and BF3·Et2O to afford thioglycoside 4a,b as a
pair of isomers with a yield of 75%. 6a,b was prepared from a mixture of 4a,b via deꢀOꢀacetylation,
4,6ꢀOꢀbenzylidenation and 3ꢀOꢀlevulination, which produced a yield of 87% over the three steps. The
anomeric thioether groups of 6a,b were then removed by the NIS/trifluoroacetic acid, and a
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corresponding hydrolysis product was produced at a yield of 76%. Subsequently the hemiacetal was
converted to trichloroacetimidate 7 following the treatment of trichloroacetonitrile and a catalytic
amount of 1,8ꢀdiazabicyclo[5.4.0]undecꢀ7ꢀene (DBU), which produced a yield of 86%.
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Scheme 2. Synthesis of DꢀGalNꢀBased Donor 7 and ᴅꢀGlcꢀBased Acceptor 8.
Reagents and conditions: a. pꢀToluenethiol, BF3·Et2O, DCM, 40 °C, overnight, 75%; b. NaOMe, MeOH, r.t., 30 min; c.
benzaldehyde dimethyl acetal, cat. PTSA, DMF, 40 °C, reduce pressure, 2 h; d. Levulinic acid, EDCI, DMAP, DCM, r.t., 2 h, 87%
for three steps; e. NIS, TFA, DCM/H2O, 3 h, 0 °C, 76%; f. Cl3CCN, DBU, DCM, 0 °C, 4 h, 86%.
Disaccharide unit preparation. Although donor 7 was failed to be glycosylated in acetonitrile after
several attempts, the BF3·Et2O/toluene system was successfully applied to the condensation of donor 7
and acceptor 8 (Table 1), exclusively affording disaccharide 2 in the desired βꢀconfiguration. After
optimization, the disaccharide product was quantitatively obtained with an excess (1.8 equiv.) of
imidate at ꢀ60 °C for 30 min. It should be noted that, when treated with 0.4 equiv. of BF3•Et2O (entry 1),
the donor 7 was decomposed rapidly to its hydrolysis product judged by TLC. Acceptor 8 was
recovered and no disaccharide derivatives were detected. This may be attributed to the low stability of
donor 7 in acid environment. Disaccharide 2 was easily separated from the hydrolysis product of
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imidate 7 by recrystallization, whereas the latter was subjected to the recycle of donor 7. The
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βꢀconfiguration of glycoside 2 was established from H and 13C NMR spectrum with signals for the
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1ꢀβꢀanomeric proton at δ 4.31 ppm (d, 1H, J = 8.0 Hz, H2ꢀ1) and carbon at δ 100.78 ppm.
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Table 1. Synthesis of Disaccharide Repeating Unit 2
Reagents and conditions: Toluene, ꢀ60 °C, 30 min.
entry
BF3·Et2O
0.4 eq.
donor
2.0 eq.
2.0 eq.
yield
0
entry
BF3·Et2O
0.15 eq.
0.18 eq.
donor
1.5 eq.
1.8 eq.
yield
52%
96%
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4
0.2 eq.
97%
Elongation of the repeating unit. With the well protected disaccharide repeating unit in hand, we then
proceeded to prepare the tetrasaccharide and hexasaccharide by elongation. Disaccharide donor 9 was
smoothly prepared from
2 by the removal of the 4ꢀmethylphenylthiol group following
trichloroacetimidate activation, with 87% yield over two steps (Scheme 3).
Scheme 3. Synthesis of Disaccharide Donor 9
Reagents and conditions: a. NIS, TFA, DCM/H2O, r.t., 1 h; b. Cl3CCN, DBU, DCM, r.t., 30 min, 87% for two steps.
Given that the disaccharide acceptor was the beginning unit in the assembly of the CSꢀE
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oligosaccharides, the derivatization at the Cꢀ1 position was preliminarily executed at the disaccharide
stage. Both thioglycoside 2 and imidate 9 were employed as donors to couple with methanol,
3ꢀ(trimethylsilyl)propargyl alcohol and 4ꢀmethoxyphenol, respectively (Table 2). Results indicated that
thioglycoside 2 was not active enough for glycosylation under the agency of NISꢀTfOH. Conversely,
imidate 9 was coupled with the three alcohols under the catalysis of trimethylsilyl triflate, which
exclusively provided the 1,2ꢀtransꢀlinked glycosides 10, 11 and 12 with yields of 57%, 87% and 91%,
respectively. Notably, the flexibility to functionalize the reducing end, anchored the CSꢀE molecule into
useful biological and chemical tools, such as fluorophore for bioassays10, 28ꢀ30 and alkyne for the
synthesis of the nonꢀnatural CSꢀE polymerized species31ꢀ33. Here in this study, the 4ꢀmethoxyphenol
was chosed as aglycone to undergo the following work.
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Table 2. Preliminary Derivatization of the Reducing Terminal Protecting Group
entry
1ꢀ3
donor
aglycon
reaction condition
product / yeild
Not work
methanol / 4ꢀmethoxyphenol /
NIS/TfOH, 0 °Cꢀr.t., overnight
2
3ꢀ(Trimethylsilyl)propargyl alcohol
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methanol
TMSOTf/DCM, r.t., 52 h
TMSOTf/DCM, 0 °C, overnight
TMSOTf/DCM, ꢀ60 °C, 30 min
10 / 57%
11 / 87%
12 / 91%
3ꢀ(Trimethylsilyl)propargyl alcohol
4ꢀmethoxyphenol
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The CSꢀE tetrasaccharide and hexasaccharide precursors were synthesized based on the above
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derivatization, which characterized the scope of glycosylation of donor 9 (Scheme 4). The levulinoyl
group of 12 was selectively removed by hydrazine acetate in dichloromethane, affording disaccharide
acceptor 13 in 89% yield. The coupling reaction of acceptor 13 with a slight excess (1.2 equiv.) of
disaccharide donor 9 under the catalysis of TMSOTf produced tetrasaccharide derivative 14 in 95%
yield. Through a similar process, hexasaccharide 16 was afforded in 88% yield. All the described
glycosylation reactions with imidate 9 went rapidly and efficiently, and no 1,2ꢀcisꢀlinked species or
orthoesters were observed.
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Scheme 4. Synthesis of Tetrasaccharide 14 and Hexasaccharide 16 from Disaccharide 9.
Reagents and conditions: a. pꢀMethoxyphenol, TMSOTf, DCM, ꢀ60 °C, 30 min, 91%; b. N2H4ꢀH2O/Py/AcOH, r.t., 40 min, 89 %;
c. TMSOTf, DCM, ꢀ60 °C, 30 min, 95%; d. N2H4ꢀH2O/Py/AcOH, r.t., 40 min, 93%; e. TMSOTf, DCM, ꢀ60 °C, 30 min, 88%.
The herein presented results evidently indicated that disaccharide imidate 9 was a very attractive
donor with a high reactivity and nearly all the corresponding glycosylations went rapidly and efficiently.
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Only a slight excess of donor was required to afford expected products in good yields, which
significantly raised the economic efficiency of the CSꢀE oligomers’ syntheses. The improvement was
attributed to two aspects: first, the elimination of deactivating effect caused by carboxylic acid moiety
at Cꢀ5 of the Glc residue, and secondly, the adoption of inert azido group at Cꢀ2 of the Gal residue
instead of the commonlyꢀused acyl protecting groups which may nucleophilically attack the donor to
form an unfavorable intermediate.7ꢀ8
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Functional groups transformation. Having achieved intermediates 14 and 16 in hand, we then
identified a series of functional group modifications to obtain the target oligomers. The reaction
sequence was as follows.
Scheme 5. Application of the PostglycosylationꢀOxidation Strategy on the Oligosaccharide Level
Reagents and conditions: a. DDQ, DCM/H2O, r.t., overnight, for 17; r.t., 5 h, for 19. b. TEMPO, BAIB, TBAB, CH3CN, H2O, r.t.,
4 h; c. CH3I, K2CO3, DMF, r.t., 3 h, 81% for 18 (two steps), r.t., 30 min, 46% for 20 (three steps).
Tetrasaccharide 14 was transformed to 17 through the oxidative removal of pꢀmethoxybenzyl (PMB)
with 2,3ꢀdiꢀchloroꢀ5,6ꢀdicyanobenzoquinone (DDQ) in wet dichloromethane. The subsequent oxidation
of the primary alcohol in 17 with (2,2,6,6ꢀtetramethylpiperidinꢀ1ꢀyl)oxyl (TEMPO) and
(diacetoxyiodo)ꢀbenzene (BAIB), which was followed by methyl esterification, afforded Cꢀ5
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carboxylic acid ester derivative 18 in 81% yield over three steps. The same sequence of reactions was
repeated starting from 16 and the corresponding hexasaccharide derivative 20 was obtained, with a
yield of 46% over three steps (Scheme 5).
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The oxidation of the primary alcohol was initially performed in wet dichloromethane as previously
described for preparation of disaccharide 1.18 However, under this reaction condition, the oxidation of
tetrasaccharide 17 was executed over a very long time and required a large excess of the oxidizing
reagents. Otherwise, the reaction was incomplete. In the case of hexasaccharide 19, the attempt left the
oxidation a list with a bar code of spots on the TLC plate. This failure was attributed to the inadequate
contact between the reagents and substance in twoꢀphase system. Therefore, acetonitrile was used
instead of dichloromethane, and tetrabutylammonium bromide (TBAB) was added to make the solvent
system homogeneous. As expected, the situation was greatly improved both for oligosaccharides 18
and 20 which were smoothly oxidated with good yields.
To the best of our knowledge, this is the first application of the postglycosylationꢀoxidation
strategy34 in the synthesis of CSꢀE oligosaccharides. The efficiency of this approach was examined. In
the application of the former strategy, the glycosylations greatly benefited from the high reactivity of
the nonꢀoxidized glucosyl donor, thus making the elongation easily achievable. The total yield of the
glycosylationꢀoxidation reactions was 77% (95% for glycosylation, 81% for oxidation) for
tetrasaccharide 18 and 38% (95%, 88% for glycosylation, 46% for oxidation) for hexasaccharide 20.
Therefore the postglycosylationꢀoxidation strategy was believed to be favorable for the synthesis of
CSꢀE oligosaccharide. Nevertheless, the application of the reported strategy was also limited by the
length of the CSꢀE oligomers, which was also observed in previously published synthetic approaches.
Subsequently, the conversion of the azido group to the acetamide group was established at the
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oligosaccharide level (Table 3). Suitable methods were initially explored. Disaccharide 12 and
tetrasaccharide 14 were subjected to four chemoselective reduction methods for the azido groups.
Lindlarꢀcatalyzed hydrogenation appeared ineffective, and reduction under a harsher condition was
taken to be detrimental to 4,6ꢀOꢀbenzylidene acetal. With the treatment of triphenylphosphine (Ph3P) in
wet tetrahydrofuran, the resulting mixture was difficult to purify. Two sulfide reagents,
1,3ꢀpropanedithiol and thioacetic acid, were then examined for azideꢀreduction. Under the agency of
1,3ꢀpropanedithiol in a mixture of Et3N/pyridine, the azido was reduced to free amino group and
acylated with acetic anhydride to generate the corresponding acetamide 21 and 22 with yields of 86%
and 59%, respectively. This two stepꢀwise approach allowed further Nꢀmodification. The
chemoselective transformation of the azides into acetamides was also fulfilled by a oneꢀpot
reductionꢀacetylation sequence using thioacetic acid in pyridine.
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The two available methods were then applied to oxidized derivatives 18 and 20. The complete
conversion of these two oligosaccharides required a longer time and higher temperature, of which the
results produced lower yields. Acetamideꢀtype tetrasaccharide 23 was afforded with the employment of
thioacetic acid and 1,3ꢀpropanedithiol in similar yields of 65% and 61%, respectively. Hexasaccharide
20 was directly transformed into the corresponding acetamide 24 in the presence of thioacetic acid with
moderate yield of 52% (Table 3).
The findings obtained above suggest that the reduction of azide tended to produce markedly lower
yield when the number of azido groups increased in the molecule, which was in accordance with the
results in literature7ꢀ8. Nevertheless, the reduction efficiency of hexasaccharide 20 remained acceptable.
This successful conversion demonstrated that azido can serve as a good alternative for the protection of
2ꢀamino in the syntheses of CSꢀE oligosaccharides, thereby providing an access to their Nꢀderivatives.
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Table 3. Conversion of Azides to Acetamides
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Reagents and conditions: a. AcSH/ Py. b. 1,3ꢀDithiolpropane/Et3N/Py; Ac2O, DCM, r.t., 1ꢀ3 h.
entry
substance
method
reaction condition
r.t., 2 h
product
21
yield
89%
86%
76%
59%
65%
61%
52%
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a
b
a
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a
r.t., 30 min
r.t., 3 h
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r.t., 4.5 h
22
60 °C, 24 h
r.t., 11 h
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23
60 °C, 24 h
24
At this point, fully protected CSꢀE tetrasaccharide 23 and hexasaccharide 24 were nearly the same to
their analogues reported by Tamura’s group8, and the only tiny difference of these intermediates is the
distinction between benzoyl (Bz) substitutes in our compounds and methoxybenzoyl (MBz) substitutes
in the reported counterparts. For this reason, we believe that the intermediates 23 and 24 can be
converted to the final CSꢀE products based on the sulfation and deꢀprotection procedure reported in
previous literature.
ꢀ
CONCLUSION
In summary, starting from the known monosaccharide donor 7 and acceptor 8, we produced the CSꢀE
tetrasaccharide and hexasaccharide precursors within 10 and 12 steps with total yield of 34% and 13%,
respectively. The feature of our synthesis include the application of postoxidationꢀglycosylation
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strategy on CSꢀE oligosaccharides and a lateꢀstage functional groups transformation. Most notably, our
approach allowed the synthesis of CSꢀE analogues to expand research territory of this important
pharmacological oligosaccharide, that is currently underway in our group, of which relevant research
will be reported elsewhere in due course. The developed methodology can also be applied to other
chondroitin oligomers and similar members of the glycosaminoglycan family.
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ꢀ
EXPERIMENT SECTION
General Methods. The reactions were performed with commercial reagents and solvents, and all the
chemicals were usd as received otherwise stated. Solvents were removed under reduced pressure.
Traces of water in the starting materials were removed by coevaporation with toluene. Powdered
molecular sieves (4 Å) used for reactions were activated by heating at 400 °C for 4 h prior to the
application. All NMR spectra were recorded on a JEOL ECZꢀ400S (400 MHz, Japan), a Bruker
AVANCEꢀIII 400 (400 MHz, Switzerland), a WNMRꢀI 500 (500 MHz, China), a Bruker AVANCEꢀIII
HD 600 (600 MHz, Switzerland) spectrometers in CDCl3. Highꢀresolution mass spectra (HRMS) were
measured with a ThermoFisher Exactive Plus mass spectrometer (ThermoFisher Scientific, Bremen,
Germany) equipped with a ThermoFisher Accela HPLC system (ThermoFisher Scientific, Bremen,
Germany). Reactions were monitored by thin layer chromatography (TLC) (HSGF254, Branch of
Qingdao Haiyang Chemical Plant, China). For column chromatography silica gel was used
(HG/T2354ꢀ92, Branch of Qingdao Haiyang Chemical Plant, China).
pꢀMethylphenyl 3,4,6ꢀTriꢀOꢀacetylꢀ2ꢀazidoꢀ2ꢀdeoxyꢀ1ꢀthioꢀα/βꢀᴅꢀgalactopyranoside (4a,b). 318
(49.6 g, 132.93 mmol) and pꢀToluenethiol (24.8 g, 199.41 mmol) were dissolved in dry CH2Cl2 (208
mL), and BF3.Et2O (67.5 mL, 531.76 mmol) was then added dropwise at 0 °C. After being stirred for
30 min at 0°C, the system was warmed to room temperature and heated to reflux overnight. The
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reaction was quenched with sat. NaHCO3 aq., and extracted with CH2Cl2 (3 ×). The organic phase was
washed with brine, dried over anhydrous Na2SO4, and concentrated. Silica gel column chromatography
of the residue (EtOAc/petroleum ether, 1:6) gave a mixture of 4a,b (45.2 g, 75%, a:b = 1:1) as a yellow
syrup. 1H NMR (400 MHz, CDCl3, 4a, α): δ 7.41 (d, J = 8.0 Hz, 2H), 7.13 (d, J = 9.2 Hz, 2H), 5.61 (d,
J = 5.6 Hz, 1H, Hꢀ1), 5.48 (d, J = 2.4 Hz, 1H, Hꢀ4), 5.18 (dd, J = 11.2, 2.8 Hz, 1H, Hꢀ3), 4.77 (t, J =
6.4 Hz, 1H, Hꢀ5), 4.30 (dd, J = 11.2, 5.6 Hz, 1H, Hꢀ2), 4.15 (d, J = 6.8 Hz, 1H, Hꢀ6a), 4.12 (d, J = 6.8
Hz, 1H, Hꢀ6b), 2.34 (s, 3H, CH3ꢀSTol), 2.15 (s, 3H, CH3ꢀOAc), 2.06 (s, 3H, CH3ꢀOAc), 2.01 (s, 3H,
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CH3ꢀOAc); H NMR (400 MHz, CDCl3, 4b, β): δ 7.51 (d, J = 8.0 Hz, 2H), 7.16 (d, J = 8.8 Hz, 2H),
5.34 (d, J = 2.8 Hz, 1H, Hꢀ4), 4.86 (dd, J = 10.2, 2.8 Hz, 1H, Hꢀ3), 4.46 (d, J = 10.0 Hz, 1H, Hꢀ1),
4.09ꢀ4.07 (d, 2H, J = 6.4 Hz, Hꢀ6a,b), 3.87 (t, J = 6.4 Hz, 1H, H1ꢀ5), 3.63 (t, J = 10.0 Hz, 1H, Hꢀ2), 2.36
(s, 3H, CH3ꢀSTol), 2.09 (s, 3H, CH3ꢀOAc), 2.04 (s, 3H, CH3ꢀOAc), 2.03 (s, 3H, CH3ꢀOAc); 13C NMR
(100 MHz, CDCl3, α/β, 1:1.2): δ 170.5, 170.4, 170.1, 170.0, 169.8, 169.7, 139.0, 138.5, 134.1, 133.2,
130.0, 129.9, 128.7, 127.3, 87.4 (α, Cꢀ1), 86.8 (β, Cꢀ1), 74.4, 73.1, 70.2, 67.6, 67.5, 66.6, 61.8, 61.7,
59.4, 58.2, 21.3, 21.3, 20.8, 20.7, 20.7, 20.7. HRMS (ESIꢀTOF) m/z [M + Na]+ Calcd for
C19H23N3NaO7S 460.1154, found 460.1108.
pꢀMethylphenyl 2ꢀazidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀ1ꢀthioꢀα/βꢀᴅꢀgalactopyranosid
e (6a,b). MeONa (2.96 g，54.78 mmol) was added into a solution of 4a,b (45.19 g, 103.37 mmol) in
CH2Cl2 (104 mL) and MeOH (234 mL). The mixture was stirred for 30 min and DOWEX 50X8ꢀ200
was added. After stirred for 1 h, the mixture was filtered and concentrated to afford the deacetylated
intermediate as a yellow solid. The crude product was dissolved in DMF (103 mL), then
pꢀanisaldehyde dimethyl acetal (46.5 mL, 310.11 mmol) and pꢀtoluene sulfonic acid monohydrate (4.72
g, 24.81 mmol) were added whereafter the system was stirred at 40 °C for 2 h at reduced pressure.
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Once complete, the reaction mixture was quenched with Et3N, diluted with EtOAc, washed with brine,
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dried over anhydrous Na2SO4, and concentrated to afford 5a,b as a yellow syrup. This compound was
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used for the next step without further purification. H NMR (400 MHz, CDCl3, 5a, α): δ 7.51ꢀ7.48 (m,
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2H, ArHꢀSTol), 7.40ꢀ7.36 (m, 5H, ArHꢀbenzylidene), 7.11 (d, J = 8.0 Hz, 2H, ArHꢀSTol), 5.67 (d, J =
5.2 Hz, 1H, Hꢀ1), 5.59 (s, 1H, CHꢀbenzylidene), 4.31 (dd, J = 3.6, 0.8 Hz, 1H, Hꢀ4), 4.26ꢀ4.21 (m, 2H,
Hꢀ2, Hꢀ5), 4.17 (dd, J = 12.4, 5.2 Hz, 1H, Hꢀ6a), 4.11 (dd, J = 12.4, 1.6 Hz, 1H, Hꢀ6b), 4.00 (td, J =
10.4, 3.6 Hz, 1H, Hꢀ3), 2.69 (d, J = 10.0 Hz, 1H, OH), 2.33 (s, 3H, CH3ꢀSTol); 13C NMR (100 MHz,
CDCl3, 5a, α) δ 137.8, 137.3, 131.8, 130.0, 129.9, 129.6, 128.5, 126.4, 101.5 (CHꢀbenzylidene), 87.8
(Cꢀ1), 75.3, 69.7, 69.3, 63.7, 61.5 (Cꢀ2), 21.2 (CH3ꢀSTol). 1H NMR (400 MHz, CDCl3, 5b, β): δ 7.62 (d,
J = 8.4 Hz, 2H, ArHꢀSTol), 7.43ꢀ7.36 (m, 5H, ArHꢀbenzylidene), 7.11 (d, J = 8.0 Hz, 2H, ArHꢀSTol),
5.51 (s, 1H, CHꢀbenzylidene), 4.38 (dd, J = 12.4, 1.2 Hz, 1H, Hꢀ6a), 4.36 (d, J = 9.6 Hz, 1H, Hꢀ1), 4.16
(d, J = 3.6 Hz, 1H, Hꢀ4), 4.02 (dd, J = 12.4, 1.6 Hz, 1H, Hꢀ6b), 3.62 (td, J = 9.6, 3.6 Hz, 1H, Hꢀ3), 3.49
(d, J = 0.8 Hz, 1H, Hꢀ5), 3.48 (t, J = 9.6 Hz, 1H, Hꢀ2), 2.51 (d, 1H, J = 9.6 Hz, OH), 2.36 (s, 3H,
CH3ꢀSTol); 13C NMR (100 MHz, CDCl3, 5b, β): δ 138.9, 137.5, 134.9, 129.9, 129.6, 128.4, 126.7,
126.4, 101.5 (CHꢀbenzylidene), 85.1 (Cꢀ1), 74.6, 73.3, 69.9, 69.4, 62.1 (Cꢀ2), 21.4 (CH3ꢀSTol). HRMS
(ESIꢀTOF) m/z [M + Na]+. Calcd for C20H21N3NaO4S 422.1150, found 422.1138.
5a,b was added to a solution of EDCI.HCl (49.5 g，258.4 mmol), DMAP (3.79 g，31.01 mmol), and
levulinic acid (30.0 g，258.4 mmol) in dry CH2Cl2, and the resulting mixture was stirred for 2 h at room
temperature. After that, the reaction mixture was poured into cold water, and extracted with CH2Cl2
(3×). The organic phase was washed with brine, dried over anhydrous Na2SO4, and concentrated in
vacuo. Silica gel column chromatography (EtOAc/petroleum ether, 1:6→1:2) gave 6a,b (44.3 g, 87%
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for three steps) as a yellow syrup. H NMR (400 MHz, CDCl3, 6b, β): δ 7.61 (d, J = 8.0 Hz, 2H,
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ArHꢀSTol), 7.43ꢀ7.36 (m, 5H, ArHꢀbenzylidene), 7.06 (d, J = 8.4 Hz, 2H, ArHꢀSTol), 5.47 (s, 1H,
CHꢀbenzylidene), 4.83 (dd, J = 10.4, 3.2 Hz, 1H, Hꢀ3), 4.45 (d, J = 9.6 Hz, 1H, Hꢀ1), 4.34 (dd, J = 12.8,
1.6 Hz, 1H, Hꢀ6a), 4.27 (dd, J = 3.2 Hz, 1H, Hꢀ4), 3.62 (dd, J = 12.4, 1.6 Hz, 1H, Hꢀ6b), 3.79 (t, J =
10.0 Hz, 1H, Hꢀ2), 3.51 (s, 1H, Hꢀ5), 2.72ꢀ2.69 (m, 2H, CH2ꢀLev), 2.60ꢀ2.55 (m, 2H, CH2ꢀLev), 2.33
(s, 3H, CH3ꢀSTol), 2.06 (s, 3H, CH3ꢀLev); 13C NMR (100 MHz, CDCl3, 6b, β): δ 206.3 (COꢀLev),
172.0 (COꢀLev), 138.7, 137.7, 134.6, 129.8, 129.2, 128.1, 126.5, 126.2, 100.9, 85.2 (Cꢀ1), 73.9, 72.6,
69.5, 69.1, 58.3 (Cꢀ2), 37.8 (CH2ꢀLev), 29.6 (CH3ꢀLev), 28.1 (CH2ꢀLev), 21.3 (CH3ꢀSTol). HRMS
(ESIꢀTOF) m/z [M+Na]+ Calcd. for C25H27N3NaO6S 520.1518, found 520.1500.
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2ꢀazidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀ1ꢀOꢀtrichloroacetimidoylꢀαꢀᴅꢀgalactopyranosid
e (7). To a solution of 6a,b (4.87 g, 9.80 mmol) in CH2Cl2 (50 mL) and water (3.5 mL) was added
Nꢀiodosuccinimide (2.64 g, 11.75 mmol). Then trifluoroacetic acid (365 ꢁL, 4.89 mmol) was added
dropwise to the mixture at 0 °C, and the reaction was stirred for 3 h at 0 °C. After that, 5 M Na2S2O3
solution was added and the mixture was extracted with CH2Cl2 (3×), washed with brine, dried over
anhydrous Na2SO4, and concentrated in vacuo. Column chromatography (EtOAc/petroleum ether, 1:5
→ 2:5) gave the corresponding hemiacetal (2.91 g, 76%) as a yellow syrup and the starting material 6a
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(271 mg) was recovered. H NMR (400 MHz, CDCl3, α/β, 1:0.3): δ 7.52ꢀ7.50 (m, 2H), 7.40ꢀ7.36 (m,
3H), 5.54 (s, 1H, CHꢀbenzylidene), 5.47 (d, J = 3.2 Hz, Haꢀ1), 5.38 (dd, J = 10.8, 3.2 Hz, 1H, Haꢀ3),
4.77 (dd, J = 10.8, 3.2 Hz, 1H, Hbꢀ3), 4.63 (d, J = 8.0 Hz, 1H, Hbꢀ1), 4.43 (d, J = 3.2 Hz, 1H, Haꢀ4),
4.32ꢀ4.28 (m, 2H, Hbꢀ4, Hbꢀ6a), 4.22 (dd, J = 12.4, 1.2 Hz, 1H, Haꢀ6a), 4.07ꢀ3.98 (m, 4H, Haꢀ2, Haꢀ5,
Haꢀ6b, Hbꢀ6b), 3.87 (dd, J = 11.8, 8.0 Hz, 1H, Hbꢀ2), 3.45 (s, 1H, Hbꢀ5), 2.79ꢀ2.65 (m, 4H, CH2ꢀLev),
2.12 (s, 3H, CH3ꢀLev); 13C NMR (100 MHz, CDCl3): δ 206.5 (COꢀLev), 172.3 (COꢀLev), 137.7, 129.2,
128.33, 128.32, 126.34, 126.30, 100.9 (CHꢀbenzylidene), 96.5 (Cbꢀ1), 92.9 (Caꢀ1), 73.6, 72.7, 72.4,
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69.7, 69.1, 66.7, 62.6 (Cꢀ2), 62.1, 58.1 (Cꢀ2), 38.0 (CH2ꢀLev), 29.8 (CH3ꢀLev), 28.3 (CH2ꢀLev). HRMS
(ESIꢀTOF) m/z [M + Na]+ Calcd for C18H21N3NaO7 414.1277, found 414.1256.
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A mixture of the hemiacetal (20.15 g, 51.53 mmol), Cl3CCN (31 mL, 309.15 mmol), and DBU (3.08
mL, 20.61 mmol) in anhyd CH2Cl2 (504 mL) was stirred at 0 °C for 4 h. Then the resulting mixture
was concentrated. Column chromatography (EtOAc/PE, 1:15 + 0.5% Et3N) gave imidate 7 (23.56 g,
86%) as a white solid. 1H NMR (400 MHz, CDCl3): δ 8.77 (s, 1H, NH), 7.53ꢀ7.51 (m, 2H,
ArHꢀbenzylidene), 7.39ꢀ7.37 (m, 3H, ArHꢀbenzylidene), 5.58 (d, J = 3.2 Hz, 1H, Hꢀ1), 5.56 (s, 1H,
CHꢀbenzylidene), 5.37 (dd, J = 10.8, 3.6 Hz, 1H, Hꢀ3), 4.55 (dd, J = 3.2 Hz, 1H, Hꢀ4), 4.32ꢀ4.28 (m,
2H, Hꢀ6a,b), 4.04 (dd, J = 12.8, 2.8 Hz, 1H, Hꢀ2), 3.98 (s, 3H, Hꢀ5), 2.80ꢀ2.66 (m, 4H, CH2ꢀLev), 2.12
(s, 3H, CH3ꢀLev); 13C NMR (100 MHz, CDCl3): δ 206.4, 172.2, 160.7 (C=NH), 137.4, 129.3, 128.3,
126.3, 100.9 (CHꢀbenzylidene), 95.4 (Cꢀ1), 90.9 (CCl3), 72.9, 69.9, 68.8, 64.9, 57.0 (Cꢀ2), 37.9
(CH2ꢀLev), 29.8 (CH3ꢀLev), 28.2 (CH2ꢀLev).
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pꢀMethoxyphenyl
Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀβꢀᴅꢀgalactopyranosyl)
ꢀ(1→4)ꢀ2,3ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀ1ꢀthioꢀβꢀᴅꢀglucopyranoside (2). Donor 7 (978.8 mg,
1.83 mmol) and acceptor 8 (625.2 mg, 1.02 mmol) were coevaporated with dry toluene (3 × 35 mL),
dried in vacuo for 30 min and dissolved in anhydrous toluene (34 mL) in the presence of 4Å powdered
molecular sieves. After stirring for 30 min at room temperature, a solution of BF3·Et2O in CH2Cl2 (0.76
M, 242 ꢁL, 0.18 mmol) was added at ꢀ60 °C. The reaction was quenched 30 min later by addition of
Et3N, filtered through a Celite pad and concentrated to dryness. The residue was roughly purified by
column chromatography on silica gel (EtOAc/petroleum ether, 1:2) and then was subjected to a
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recrystallization (EtOAc/petroleum ether, 1:5) to afford 2 (963.1 mg, 96%) as white solid. H NMR
(400 MHz, CDCl3): δ 7.95ꢀ7.93 (m, 2H, ArHꢀBz), 7.91ꢀ7.89 (m, 2H, ArHꢀBz), 7.50 (t, J = 7.4 Hz, 1H,
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ArHꢀBz), 7.40ꢀ7.16 (m, 14H, ArHꢀBz, PMB, STol, benzylidene), 7.06 (d, J = 8.0 Hz, 1H, ArHꢀSTol),
6.92 (d, J = 8.6 Hz, 1H, ArHꢀPMB), 5.69 (t, J = 9.2 Hz, 1H, H1ꢀ3), 5.35 (t, J = 9.6 Hz, 1H, H1ꢀ2), 5.21
(s, 1H, CHꢀbenzylidene), 4.85 (d, J = 10.0 Hz, 1H，H1ꢀ1), 4.63 (d, J = 11.5 Hz, 1H, CH2ꢀPMB), 4.54
(dd, J = 10.8, 3.6 Hz, 1H, H2ꢀ3), 4.51 (d, J = 11.6 Hz, 1H, CH2ꢀPMB), 4.31 (d, J = 8.0 Hz, 1H, H2ꢀ1),
4.21 (t, J = 9.6 Hz, 1H, H1ꢀ4), 4.03 (d, J = 3.4 Hz, 1H, H2ꢀ4), 3.97 (dd, J = 11.2, 3.6 Hz, 1H, H1ꢀ6a),
3.91 (d, J = 10.4 Hz, 1H, H1ꢀ6b), 3.82 (s, 3H, OCH3ꢀPMB), 3.76ꢀ3.71 (m, 2H, H2ꢀ2, H1ꢀ5), 3.56 (s, 2H,
H2ꢀ6a,b), 2.90 (s, 1H, H2ꢀ5), 2.71ꢀ2.68 (m, 2H, CH2ꢀLev), 2.60ꢀ2.57 (m, 2H, CH2ꢀLev), 2.32 (s, 3H,
CH3ꢀLev), 2.05 (s, 3H, CH3ꢀ STol); 13C NMR (100 MHz, CDCl3): δ 206.2 (COꢀLev), 172.1 (COꢀLev),
165.7 (COꢀBz), 165.3 (COꢀBz), 159.4, 138.5, 137.7, 133.7 (2C), 133.2, 132.8, 130.4, 130.0, 129.9,
129.8, 129.8, 129.7, 129.6, 129.0, 128.4, 128.4, 128.1, 128.0, 126.6, 113.9 (2C), 101.4 (PhCH), 100.8
(C2ꢀ1), 86.4 (C1ꢀ1), 79.3, 75.4, 74.6, 73.2, 72.7, 72.3, 71.0, 68.1, 68.0, 66.2, 60.6, 55.4 (OCH3ꢀPMB),
37.9 (CH2ꢀLev), 29.7 (CH3ꢀLev), 28.2 (CH2ꢀLev), 21.3 (CH3ꢀSPhMe). HRMS (ESIꢀTOF) m/z [M +
Na]+ Calcd for C53H53N3NaO14S 1010.3146, found 1010.3127.
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Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1→4)ꢀ2,3ꢀdiꢀOꢀben
zoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀ1ꢀOꢀtrichloroacetimidoylꢀαꢀᴅꢀglucopyranoside (9). Trifluoroacetic acid
(19 ꢁL, 0.25 mmol) was added dropwise to a stirred mixture of 2 (500 mg, 0.51 mmol) and
Nꢀiodosuccinimide (228 mg, 1.01 mmol) in CH2Cl2 (13 mL) and water (1.3 mL) at 0 °C. Then the
system was stirred for 1 h at room temperature. At completion, 5 M Na2S2O3 solution was added and
the mixture was extracted with CH2Cl2 (3 ×). The organic phase was washed with brine, dried over
anhydrous Na2SO4 after which the solvents were evaporated to afford the corresponding hemiacetal as
a white foam. HRMS (ESIꢀTOF) m/z [M + Na]+ Calcd for C46H47N3NaO15 904.2905, found 904.2855.
The hemiacetal was dissolved in dry CH2Cl2 (10 mL). Cl3CCN (305 ꢁL, 3.04 mmol) and DBU (30
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ꢁL, 0.20 mmol) were added dropwise and the reaction mixture was stirred at room temperature for 30
min. After that, the volatiles were removed under reduced pressure. The obtained residue was purified
by column chromatography on silica gel (EtOAc/PE, 1:5→1:2 + 0.1% Et3N) to afford imidate 9 (409
mg, 79% for two steps) as a white foam. 1H NMR (500 MHz, CDCl3): δ 8.57 (s, 1H, NH), 7.95 (m, 4H,
ArHꢀBz), 7.47 (t, J = 7.1 Hz, 1H, ArHꢀBz), 7.39ꢀ7.15 (m, 12H, ArHꢀBz, PMB, benzylidene), 6.92 (d, J
= 8.3 Hz, 2H, ArHꢀPMB), 6.73 (d, J = 3.0 Hz, 1H, H1ꢀ1), 6.09 (t, J = 9.8 Hz, 1H, H1ꢀ3), 5.47 (dd, J =
10.1, 3.3 Hz, 1H, H1ꢀ2), 5.23 (s, 1H, CHꢀbenzylidene), 4.66 (d, J = 11.5 Hz, 1H, CH2ꢀPMB), 4.53 (dd,
J = 10.6, 3.0 Hz, 1H, H2ꢀ3), 4.46 (d, J = 11.5 Hz, 1H, CH2ꢀPMB), 4.38 (t, J = 9.5 Hz, 1H, H1ꢀ4), 4.26
(m, 2H, H2ꢀ4, H1ꢀ6a), 4.06 (m, 2H, H1ꢀ6b), 3.82 (s, 5H, OCH3ꢀPMB, H2ꢀ2, H1ꢀ5), 3.58 (s, 2H, H2ꢀ6a,b),
2.93 (s, 1H, H2ꢀ5), 2.70 (m, 2H, CH2ꢀLev), 2.60 (m, 2H, CH2ꢀLev), 2.05 (s, 3H, CH3ꢀLev). 13C NMR
(125 MHz, CDCl3): δ 206.1 (COꢀLev), 172.0 (COꢀLev), 165.6 (COꢀBz), 165.6 (COꢀBz), 160.8
(C=NH), 159.6, 137.6, 133.5, 132.8, 130.0, 129.9, 129.8, 129.0, 128.8, 128.5, 128.2, 128.0, 126.6,
114.0, 101.3 (PhCH), 100.8 (C2ꢀ1), 93.5 (C1ꢀ1), 90.9, 74.9, 73.3, 73.0, 72.9, 72.2, 71.0, 70.4, 68.1, 67.2,
66.2, 60.4, 55.4 (OCH3ꢀPMB), 37.9 (CH2ꢀLev), 29.7 (CH3ꢀLev), 28.2 (CH2ꢀLev).
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Methyl Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1→4)ꢀ2,3ꢀdi
ꢀOꢀbenzoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyranoside (10). Donor 9 (197 mg, 0.19 mmol) was
coevaporated with toluene (3 × 5 mL) and dried in vacuo for 30 min. To a solution of 9 in dry CH2Cl2
(4 mL) was added 4Å powdered molecular sieves. After being stirred for 30 min at room temperature,
the mixture was cooled to ꢀ60 °C. Anhydrous MeOH (250 ꢁL, 5.76 mmol) was added followed by
dropwise addition of trimethylsilyl trifluoromethanesulfonate in CH2Cl2 (0.37 M, 103 ꢁL, 0.038 mmol).
After 30 min, additional of trimethylsilyl trifluoromethanesulfonate in CH2Cl2 (0.37 M, 103 ꢁL, 0.038
mmol) was added followed by a gradual increase to room temperature and continuing stirring for 52 h.
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(99 mg, 57%) as white amorphous solid. H NMR (500 MHz, CDCl3): δ 7.95ꢀ7.92 (m, 4H, ArHꢀBz),
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7.47–7.19 (m, 15H, ArHꢀBz, PMB, benzylidene), 6.91 (d, J = 7.0 Hz, 2H, ArHꢀPMB), 5.70 (t, J = 7.5
Hz, 1H, H1ꢀ3), 5.38 (t, J = 7.5 Hz, 1H, H1ꢀ2), 5.21 (s, 1H, CHꢀbenzylidene), 4.68 (d, J = 10.0 Hz, 1H,
CH2ꢀPMB), 4.61 (d, J = 7.0 Hz, 1H, H1ꢀ1), 4.53ꢀ4.48 (m, 2H, H2ꢀ3, CH2ꢀPMB), 4.29ꢀ4.25 (m, 2H, H2ꢀ1,
H1ꢀ4), 4.03 (s, 1H, H2ꢀ4), 3.99 (d, J = 10.0 Hz, 1H, H2ꢀ6a), 3.98 (d, J = 10.0 Hz, 1H, H2ꢀ6b), 3.80ꢀ3.74
(m, 5H, OCH3ꢀPMB, H2ꢀ2, H1ꢀ5), 3.56ꢀ3.52 (m, 5H, OCH3, H1ꢀ6a, b), 2.88 (s, 1H, H2ꢀ5), 2.69ꢀ2.59 (m,
4H, CH2CH2ꢀLev), 2.05 (s, 3H, CH3ꢀLev); 13C NMR (125 MHz, CDCl3): δ 206.2 (COꢀLev), 172.0
(COꢀLev), 165.7 (COꢀBz), 165.4 (COꢀBz), 159.4, 137.6, 133.1, 132.7, 130.2, 129.9, 129.7, 129.6,
128.9, 128.3, 128.1, 127.9, 126.6, 125.9, 113.9, 102.0 (C1ꢀ1), 101.3 (C2ꢀ1), 100.7 (PhCH), 75.6, 75.0,
73.4, 73.2, 72.7, 72.2, 72.2, 68.1, 67.7, 66.1, 60.6 (C2ꢀ2), 57.1 (OCH3), 55.4 (OCH3ꢀPMB), 37.9
(CH2ꢀLev), 29.7 (CH3ꢀLev), 28.1 (CH2ꢀLev). HRMS (ESIꢀTOF) m/z [M+Na]+ Calcd for
C47H49N3NaO15 918.3061, found 918.3005.
3ꢀ(Trimethylsilyl)propargyl Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀβꢀᴅꢀgalactopyr
anosyl)ꢀ(1→4)ꢀ2,3ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyranoside (11). Imidate 9 (3.32 g,
3.24 mmol) was coevaporated with dry toluene (3 × 25 mL) and dried in vacuo for 30 min. A mixture
of 9, 3ꢀ(Trimethylsilyl)propargyl alcohol (3.7 mL, 24.71 mmol) and activated 4Å powdered molecular
sieves in dry CH2Cl2 (55 mL) was stirred at room temperature for 30 min, and then cooled to 0 °C.
Trimethylsilyl trifluoromethanesulfonate (70 ꢁL, 0.39 mmol) was added dropwise to the reaction
mixture. After being stirred overnight at 0 °C, the reaction was quenched with Et3N. Insoluble materials
were removed by filtration through Celite pad and the filtrate was concentrated under reduced pressure.
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The residue was purified by silicaꢀgel column chromatography (EtOAc/petroleum ether, 1:2) to afford
11 (2.79 g, 87%) as white amorphous solid. 1H NMR (400 MHz, CDCl3): δ 7.95ꢀ7.93 (m, 4H, ArHꢀBz),
7.47–7.17 (m, 15H, ArHꢀBz, PMB, benzylidene), 6.91 (d, J = 8.4 Hz, 2H, ArHꢀPMB), 5.73 (t, J = 9.6
Hz, 1H, H1ꢀ3), 5.40 (t, J = 9.2 Hz, 1H, H1ꢀ2), 5.21 (s, 1H, CHꢀbenzylidene), 4.98 (d, J = 8.0 Hz, 1H,
H1ꢀ1), 4.67 (d, J = 11.6 Hz, 1H, CH2ꢀPMB), 4.52 (d, J = 10.8, 3.2 Hz, 1H, H2ꢀ3), 4.48ꢀ4.24 (m, 5H,
CH2ꢀPMB, H2ꢀ1, H1ꢀ4, CH2ꢀalkynyl), 4.03ꢀ3.98 (m, 2H, H2ꢀ4, H2ꢀ6a), 3.88 (d, J = 10.8 Hz, 1H, H2ꢀ6b),
3.78ꢀ3.71 (m, 5H, OCH3ꢀPMB, H2ꢀ2, H1ꢀ5), 3.55 (s, 2H, H1ꢀ6a, b), 2.87 (s, 1H, H2ꢀ5), 2.68ꢀ2.55 (m, 4H,
CH2CH2ꢀLev), 2.02 (s, 3H, CH3ꢀLev), 0.13 (s, 9H, CH3ꢀTMS); 13C NMR (125 MHz, CDCl3): δ 206.0
(COꢀLev), 171.8 (COꢀLev), 165.5 (COꢀBz), 165.2 (COꢀBz), 159.3, 137.6, 133.0, 132.7, 130.0, 129.8,
129.8, 129.7, 128.5, 128.8, 128.2, 128.0, 127.8, 126.5, 113.8, 101.2, 100.6, 100.0, 98.1, 92.3, 75.4, 74.9,
73.3, 73.1, 72.5, 72.1, 71.8, 67.9, 67.5, 66.0, 60.5, 56.5, 55.2, 37.7 (CH2ꢀLev), 29.5 (CH3ꢀLev), 28.0
(CH2ꢀLev), ꢀ0.3 (CꢀTMS). HRMS (ESIꢀTOF) m/z [M+Na]+ Calcd for C52H57N3NaO15 Si 1014.3457,
found 1014.3472.
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pꢀMethoxyphenyl Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1
→4)ꢀ2,3ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyranoside (12). Donor 9 (390 mg, 0.38 mmol)
and acceptor 15 (56.7 mg, 0.46 mmol) were coevaporated with dry toluene (3 × 5 mL), dried in vacuo
for 30 min, and dissolved in dry CH2Cl2 (5 mL) in the presence of freshly activated 4Å powdered
molecular sieves. After stirring for 30 min at room temperature, the system was cooled to ꢀ60 °C, and a
solution of trimethylsilyl trifluoromethanesulfonate in CH2Cl2 (0.37 M, 103 ꢁL, 0.038 mmol) was
added dropwise. The reaction mixture was stirred for 30 min at ꢀ60 °C, then was quenched with Et3N,
filtered through Celite pad prior to removing the solvents in vacuo. The obtained residue was purified
by silicaꢀgel column chromatography (EtOAc/petroleum ether, 1:2) to afford 12 (343 mg, 91%) as
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white foam. H NMR (500 MHz, CDCl3): δ 7.97ꢀ7.93 (m, 4H, ArHꢀBz), 7.45 (t, J = 7.0 Hz, 1H,
ArHꢀBz), 7.34–7.21 (m, 12H, ArHꢀBz, PMB, benzylidene), 6.94 (d, J = 8.5 Hz, 2H, ArHꢀOMP), 6.89
(d, J = 8.0 Hz, 2H, ArHꢀPMB), 6.74 (d, J = 8.5 Hz, 2H, ArHꢀOMP), 5.77 (t, J = 9.0 Hz, 1H, H1ꢀ3), 5.63
(t, J = 9.0 Hz, 1H, H1ꢀ2), 5.22 (s, 1H, CHꢀbenzylidene), 5.15 (d, J = 8.0 Hz, 1H, H1ꢀ1), 4.66 (d, J =
11.5 Hz, 1H, CH2ꢀPMB), 4.57 (dd, J = 10.5, 2.5 Hz, 1H, H2ꢀ3), 4.50 (d, J = 11.5 Hz, 1H, CH2ꢀPMB),
4.34ꢀ4.30 (m, 2H, H2ꢀ1, H1ꢀ4), 4.05 (s, 1H, H2ꢀ4), 4.00 (dd, J = 11.0, 3.0 Hz, 1H, H2ꢀ6a), 3.93 (d, J =
11.0 Hz, 1H, H2ꢀ6b), 3.88 (d, J = 9.5 Hz, 1H, H1ꢀ5), 3.78ꢀ3.74 (m, 4H, OCH3ꢀPMB, H2ꢀ2), 3.70
(OCH3ꢀOMP), 3.57 (brs, 2H, H1ꢀ6a,b), 2.91(s, 1H, H2ꢀ5), 2.68ꢀ2.66 (m, 2H, CH2ꢀLev), 2.59ꢀ2.56 (m,
2H, CH2ꢀLev), 2.03 (s, 3H, CH3ꢀLev); 13C NMR (125 MHz, CDCl3): δ 206.1 (COꢀLev), 171.9
(COꢀLev), 165.5 (COꢀBz), 165.2 (COꢀBz), 159.3, 155.6, 151.2, 137.6, 133.1, 132.8, 130.0, 129.8,
129.7, 129.6, 129.3, 128.9, 128.3, 128.1, 127.9, 126.5, 118.9, 114.0, 113.8, 101.3 (C2ꢀ1), 100.8 (C1ꢀ1),
100.6 (PhCH), 75.5 (C1ꢀ4), 75.0 (C1ꢀ5), 73.3 (C1ꢀ3), 73.1 (CH2ꢀPMB), 72.6 (C2ꢀ3), 72.2 (C2ꢀ4), 72.1
(C1ꢀ2), 68.0 (C1ꢀ6), 67.6 (C2ꢀ6), 66.1 (C2ꢀ5), 60.6 (C2ꢀ2), 55.5 (OCH3ꢀOMP), 55.2 (OCH3ꢀPMB), 37.9
(CH2ꢀLev), 29.7 (CH3ꢀLev), 28.2 (CH2ꢀLev). HRMS (ESIꢀTOF) m/z [M + Na]+ Calcd for
C53H53N3NaO16 1010.3324, found 1010.3294.
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pꢀMethoxyphenyl Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1→4)ꢀ2,3ꢀdiꢀOꢀ
benzoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyranoside (13). To a stirred solution of 12 (376 mg, 0.38
mmol) in pyridine (1.4 mL) was added a freshly prepared mixture of pyridine/AcOH/hydrazine acetate
(6:4:0.5, 7.0 mL). After stirring for 40 min, the reaction mixture was diluted with EtOAc and washed
with saturated aqueous NaHCO3 and water, dried over anhydrous Na2SO4, and concentrated to dryness.
Silica gel column chromatography (CH2Cl2:Acetone, 50:1) of the residue afforded compound 13 (301
1
mg, 89%) as a white foam. H NMR (500 MHz, CDCl3): δ 7.96 (m, 4H, ArHꢀBz), 7.48 (t, J = 7.3 Hz,
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1H, ArHꢀBz), 7.42ꢀ7.14 (m, 12H, ArHꢀBz, PMB, benzylidene), 6.94 (d, J = 8.8 Hz, 2H, ArHꢀOMP),
6.88 (d, J = 8.3 Hz, 2H, ArHꢀPMB), 6.76 (d, J = 8.9 Hz, 2H, ArHꢀOMP), 5.75 (t, J = 9.4 Hz, 1H, H1ꢀ3),
5.63 (t, J = 8.8 Hz, 1H, H1ꢀ2), 5.26 (s, 1H, CHꢀbenzylidene), 5.14 (d, J = 7.8 Hz, 1H, H1ꢀ1), 4.67 (d, J
= 11.6 Hz, 1H, CH2ꢀPMB), 4.49 (d, J = 11.6 Hz, 1H, CH2ꢀPMB), 4.29 (t, J = 9.4 Hz, 1H, H1ꢀ4), 4.19 (d,
J = 8.0 Hz, 1H, H2ꢀ1), 4.02 (dd, J = 10.9, 2.9 Hz, 1H, H2ꢀ6a), 3.92 (d, J = 11.0 Hz, 1H, H2ꢀ6b), 3.89 (d,
J = 2.5 Hz, 1H, H2ꢀ4), 3.85 (d, J = 9.4 Hz, 1H, H1ꢀ5), 3.80 (s, 3H, OCH3ꢀPMB), 3.73 (s, 3H,
OCH3ꢀOMP), 3.59 (brs, 2H, H1ꢀ6a,b), 3.44 (t, J = 9.0 Hz, 1H, H2ꢀ2), 3.31 (dd, J = 10.0, 3.3 Hz, 1H,
H2ꢀ3), 2.94 (s, 1H, H2ꢀ5); 13C NMR (125 MHz, CDCl3): δ 165.7 (COꢀBz), 165.4 (COꢀBz), 159.4, 155.7,
151.3, 137.4, 133.2, 132.8, 130.1, 129.9, 129.82, 129.75, 129.4, 129.2, 128.4, 128.1, 128.1, 126.6,
118.9, 114.5, 113.9, 101.4 (C2ꢀ1), 101.1 (C1ꢀ1), 100.8 (PhCH), 75.5 (C1ꢀ4), 75.2 (C1ꢀ5), 74.2 (C2ꢀ3),
73.3 (C1ꢀ3), 73.1 (CH2ꢀPMB), 72.1 (C2ꢀ4), 71.8 (C1ꢀ2), 68.1 (C1ꢀ6), 67.6 (C2ꢀ6), 66.4 (C2ꢀ5), 64.2
(C2ꢀ2), 55.6 (OCH3ꢀOMP), 55.3 (OCH3ꢀPMB). HRMS (ESIꢀTOF) m/z [M + Na]+ Calcd for
C48H47N3NaO14 912.2956, found 912.2917.
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pꢀMethoxyphenyl Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1
→4)ꢀOꢀ(2,3ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyranosyl)ꢀ(1→3)ꢀOꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzy
lideneꢀ2ꢀdeoxyꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1→4)ꢀ2,3ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyran
oside (14). Donor 9 (208.9 mg，0.20 mmol) and acceptor 13 (150.0 mg, 0.17 mmol) were coevaporated
with dry toluene (3 × 5 mL), dried in vacuo for 30 min, and dissolved in dry CH2Cl2 (5 mL) in the
presence of 4Å powdered molecular sieves. The mixture was stirred at room temperature for 30 min
and then cooled to ꢀ60 °C. A solution fo trimethylsilyl trifluoromethanesulfonate in CH2Cl2 (0.19 M,
106 ꢁL, 0.020 mmol) was added dropwise. The reaction mixture was neutralised 30 min later with Et3N
before being allowed to warm to room temperature, and filtered with Celite pad. The filtrate was
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(petroleum ether/EtOAc, 1:3→2:3) to afford 14 (283.2 mg, 95%) as white foam. H NMR (500 MHz,
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CDCl3): δ 7.93ꢀ7.90 (m, 8H, ArHꢀBz), 7.48 (m, 2H, ArHꢀBz), 7.46ꢀ7.20 (m, 24H, ArHꢀBz, PMB,
benzylidene), 6.91 (d, J = 8.8 Hz, 2H, ArHꢀOMP), 6.89 (d, J = 8.0 Hz, 2H, ArHꢀPMB), 6.86 (d, J = 8.0
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Hz, 2H, ArHꢀPMB), 6.73 (d, J = 8.5 Hz, 2H, ArHꢀOMP), 5.71ꢀ5.66 (m, 2H, H1， ꢀ3), 5.59 (t, J = 9.5 Hz,
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1H, H1ꢀ2), 5.37 (t, J = 9.0 Hz, 1H, H3ꢀ2), 5.22 (s, 1H, CHꢀbenzylidene), 5.17 (s, 1H, CHꢀbenzylidene),
5.09 (d, J = 8.0 Hz, 1H, H1ꢀ1), 4.95 (d, J = 7.0 Hz, 1H, H3ꢀ1), 4.64 (d, J = 12 Hz, 1H, CH2ꢀPMB),
4.57ꢀ4.54 (m, 2H, H4ꢀ3, CH2ꢀPMB), 4.44 (dd, J = 12 Hz, 2 Hz, 2H, CH2ꢀPMB), 4.28ꢀ4.17 (m, 4H, H1，
3ꢀ4, H2，4ꢀ1), 4.08 (s, 1H, H4ꢀ4), 4.02 (s, 1H, H2ꢀ4), 3.90ꢀ3.72 (m, 16H, H4ꢀ2,H1ꢀ5, H3ꢀ5, H3ꢀ6a,b, H4ꢀ6ab,
OCH3ꢀPMB, OCH3ꢀOMP) ,3.60ꢀ3.49 (m, 4H, H2ꢀ6ab, H2ꢀ2, H1ꢀ6a), 3.44 (d, J = 12.0 Hz, 1H, H1ꢀ6b),
3.21 (d, J = 10.0 Hz, 1H, H2ꢀ3), 2.85 (s, 1H, H2ꢀ5), 2.80 (s, 1H, H4ꢀ5), 2.70ꢀ2.69 (m, 2H, CH2ꢀLev),
2.60ꢀ2.58 (m, 2H, CH2ꢀLev), 2.06 (s, 3H, CH3ꢀLev); 13C NMR (125 MHz, CDCl3): δ 206.1 (COꢀLev),
172.0 (COꢀLev), 165.5 (2C, COꢀBz), 165.3 (COꢀBz), 165.1 (COꢀBz), 159.4, 159.3, 155.6, 151.3, 137.8,
137.6, 133.1, 133.0, 132.9, 132.7, 130.2, 130.0, 129.9 (2C), 129.8 (8C), 129.7 (2C), 129.7, 129.6, 129.6
(4C), 129.5, 129.4, 128.9, 128.7, 128.4, 128.3, 128.2, 128.1, 128.0, 127.9, 126.5 (2C), 126.5 (2C),
118.9 (2C), 114.5 (2C), 113.9 (2C), 113.9 (2C), 101.8 (C4ꢀ1), 101.7 (C1ꢀ1), 101.6 (C3ꢀ1), 100.7 (C2ꢀ1),
100.7 (PhCH), 100.5 (PhCH), 79.3, 76.2, 75.5, 75.1, 74.6, 74.5, 73.7, 73.4 (CH2ꢀPMB), 73.3, 73.2
(CH2ꢀPMB), 72.6, 72.2, 72.2, 72.1, 68.6, 68.0, 67.5, 66.4, 66.1, 62.0, 60.7, 55.6 (OCH3ꢀOMP), 55.4
(OCH3ꢀPMB), 55.3 (OCH3ꢀPMB), 37.8 (CH2ꢀLev), 29.7 (CH3ꢀLev), 28.1 (CH2ꢀLev). HRMS (ESIꢀTOF)
m/z [M + Na]+ Calcd for C94H92N6NaO28 1775.5857, found 1775.5819.
pꢀMethoxyphenyl Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1→4)ꢀOꢀ(2,3ꢀdiꢀ
Oꢀbenzoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyranosyl)ꢀ(1→3)ꢀOꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxy
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ꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1→4)ꢀ2,3ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyranoside (15). 14
(600 mg, 0.34 mmol) was dissolved in pyridine (1.2 mL), and a freshly prepared mixture of
pyridine/AcOH/hydrazine acetate (6:4:0.5, 6 mL) was added. After stirring for 40 min, the resulting
mixture was diluted with EtOAc, washed with saturated aqueous NaHCO3 and water, dried over
anhydrous Na2SO4, and concentrated to dryness. Silica gel column chromatography (CH2Cl2:Acetone,
25:1) afforded compound 15 (529 mg, 93%) as a white foam. 1H NMR (500 MHz, CDCl3): δ 8.00ꢀ7.86
(m, 8H, ArHꢀBz), 7.46 (t, J = 6.5 Hz, 2H, ArHꢀBz), 7.36ꢀ7.22 (m, 25H, ArHꢀBz, PMB, benzylidene,
overlapped with solvent peak), 6.92ꢀ6.83 (m, 4H, ArHꢀOMP), 6.83 (d, J = 8.0 Hz, 2H, ArHꢀPMB),
6.74 (d, J = 8.0 Hz, 2H, ArHꢀPMB), 6.73 (d, J = 8.5 Hz, 2H, ArHꢀOMP), 5.71ꢀ5.66 (m, 2H, H1，3ꢀ3),
5.59 (t, J = 8.0 Hz, 1H, H1ꢀ2), 5.39 (t, J = 7.5 Hz, 1H, H3ꢀ2), 5.26 (s, 1H, CHꢀbenzylidene), 5.17 (s, 1H,
CHꢀbenzylidene), 5.09 (d, J = 8.0 Hz, 1H, H1ꢀ1), 4.94 (d, J = 7.0 Hz, 1H, H3ꢀ1), 4.63 (d, J = 11.5 Hz,
1H, CH2ꢀPMB), 4.58 (d, J = 11.5 Hz, 1H, CH2ꢀPMB), 4.45ꢀ4.42 (m, 2H, CH2ꢀPMB), 4.26 (t, J = 9.0
Hz, 1H, H1ꢀ4), 4.17ꢀ4.08 (m, 5H, H3ꢀ4, H2，4ꢀ1, H2，4ꢀ4), 3.94ꢀ3.72 (m, 17H, H4ꢀ3, H4ꢀ2, H1ꢀ5, H3ꢀ5,
H3ꢀ6a,b, H4ꢀ6ab, OCH3ꢀPMB, OCH3ꢀOMP) ,3.59ꢀ3.44 (m, 6H, H2ꢀ6ab, H2ꢀ2, H1ꢀ6a), 3.33 (dd, J = 9.5
Hz, , 1H, H1ꢀ6b), 3.21 (d, J = 10.0 Hz, 1H, H2ꢀ3), 2.86 (s, 1H, H2ꢀ5), 2.82 (s, 1H, H4ꢀ5); 13C NMR (125
MHz, CDCl3): δ 165.6 (COꢀBz), 165.5 (COꢀBz), 165.4 (COꢀBz), 165.1 (COꢀBz), 159.4, 159.3, 155.6,
151.3, 137.9 (ArCꢀbenzylidene), 137.4, 133.2, 133.1, 132.9, 132.8, 130.2, 130.0, 129.9, 129.9, 129.8,
129.7, 129.6, 129.5, 129.4, 129.3, 128.7, 128.4, 128.3, 128.2, 128.1, 127.9, 126.5, 126.5, 118.9 (2C),
114.5 (2C), 113.9 (2C), 101.9 (C4ꢀ1), 101.8 (C2ꢀ1), 101.6 (C3ꢀ1), 101.1 (C1ꢀ1), 100.7 (PhCH), 100.5
(PhCH), 79.3, 76.1, 75.5, 75.1, 74.6, 74.6, 74.1, 73.7, 73.3 (CH2ꢀPMB), 73.3, 73.2 (CH2ꢀPMB), 72.1,
71.9, 68.5, 68.1, 67.5, 66.5, 66.4, 64.4, 62.0, 60.5, 55.6 (OCH3ꢀOMP), 55.5 (OCH3ꢀPMB), 55.4
(OCH3ꢀPMB). HRMS (ESIꢀTOF) m/z [M + Na]+ Calcd for C89H86N6NaO26 1677.5489, found
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pꢀMethoxyphenyl Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1
→4)ꢀOꢀ(2,3ꢀdiꢀOꢀbenzylꢀ6ꢀOꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyranosyl)ꢀ(1→3)ꢀOꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzyli
deneꢀ2ꢀdeoxyꢀβꢀᴅꢀgalatpyrnsyl)ꢀ(1→4)ꢀOꢀ(2,3ꢀdiꢀOꢀbenzoylꢀ6ꢀOꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyranos
yl)ꢀ(1→3)ꢀOꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1→4)ꢀ2,3ꢀdiꢀOꢀbenzoylꢀ6ꢀ
Oꢀpꢀmethoxybenzylꢀβꢀᴅꢀglucopyranoside (16). Donor 9 (628 mg, 0.61 mmol) and acceptor 15 (845 mg,
0.51 mmol) were coevaporated with dry toluene (3 × 10 mL), and dried in vacuo for 30 min. The
mixture was then dissolved in dry CH2Cl2 (10 mL), and freshly activated 4Å molecular sieves were
added. After stirring for 30 min at room temperature, the reaction mixture was cooled to ꢀ60 °C, and a
solution of trimethylsilyl trifluoromethanesulfonate in CH2Cl2 (0.19 M, 316 ꢁL, 0.061 mmol) was
added dropwise. The mixture was stirred for 30 min at room temperature, then was neutralised with
Et3N, filtered and concentrated. Silicaꢀgel column chromatography (CH2Cl2/Acetone, 60:1→20:1)
afforded 16 (1.13 g, 88%) as white foam. 1H NMR (500 MHz, CDCl3): δ 7.94ꢀ7.86 (m, 12H, ArHꢀBz),
7.45ꢀ7.17 (m, 40H, ArHꢀBz, PMB, benzylidene, overlapped with solvent peak), 6.91ꢀ6.86 (m, 4H,
ArHꢀOMP, PMB), 6.82 (m, 4H, ArHꢀPMB), 6.73 (d, J = 8.5 Hz, 2H, ArHꢀOMP), 5.71ꢀ5.56 (m, 4H,
3H1/3/5 ꢀ3, H1/3/5ꢀ2), 5.38 (t, J = 8.0 Hz, 1H, H1/3/5ꢀ2), 5.34 (t, J = 8.0 Hz, 1H, H1/3/5ꢀ2), 5.22 (s, 1H,
CHꢀbenzylidene), 5.17 (s, 1H, CHꢀbenzylidene), 5.15 (s, 1H, CHꢀbenzylidene), 5.09 (d, J = 8.0 Hz, 1H,
H1/3/5ꢀ1), 4.95 (d, J = 7.0 Hz, 1H, H1/3/5ꢀ1), 4.89 (d, J = 7.0 Hz, 1H, H1/3/5ꢀ1), 4.62 (d, J = 11.5 Hz, 1H,
CH2ꢀPMB), 4.56ꢀ4.50 (m, 3H, CH2ꢀPMB), 4.43ꢀ4.41 (m, 2H, CH2ꢀPMB, H6ꢀ3), 4.36 (d, J = 11.5 Hz,
1H, CH2ꢀPMB), 4.27ꢀ4.05 (m, 8H, 3H1/3/5ꢀ4, 3H2/4/6ꢀ1, 2H2 ,4 ꢀ4), 4.02 (s, 1H, H6ꢀ4), 3.89ꢀ3.73 (m, 19H,
H6ꢀ2, 3H1/3/5ꢀ5, 6Hꢀ6a,b, OCH3ꢀPMB), 3.70 (s, 3H, OCH3ꢀOMP), 3.60ꢀ3.51 (m, 6H, 4H ꢀ6a,b, H2/4ꢀ2),
3.41 (d, J = 12.0 Hz, 1H, Hꢀ6a), 3.37 (d, J = 11.5 Hz, 1H, Hꢀ6b), 3.23 (dd, J = 10.5 Hz, 2.5 Hz, 1H,
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H2/4ꢀ3), 3.18 (dd, J = 10.0 Hz, 2.0 Hz, 1H, H2/4ꢀ3), 2.82 (s, 1H, H2/4/6ꢀ5), 2.79 (s, 1H, H2/4/6ꢀ5), 2.73 (s,
1H, H2/4/6ꢀ5), 2.68ꢀ2.67 (m, 2H, CH2ꢀLev), 2.59ꢀ2.57 (m, 2H, CH2ꢀLev), 2.04 (s, 3H, CH3ꢀLev); 13C
NMR (100 MHz, CDCl3): δ 206.2 (COꢀLev), 172.0 (COꢀLev), 165.5 (COꢀBz), 165.5 (COꢀBz), 165.4
(COꢀBz), 165.1 (COꢀBz), 165.1 (COꢀBz), 159.5, 159.4, 159.3, 155.6, 151.3, 137.9, 137.6, 133.2,
133.03, 133.00, 132.9, 132.8, 132.7, 130.2, 130.1, 130.0, 129.9, 129.8, 129.77, 129.75, 129.72, 129.7,
129.6, 129.6, 129.6, 129.5, 129.4, 129.0, 128.7, 128.6, 128.4, 128.3, 128.2, 128.2, 128.1, 128.0, 127.9,
127.9, 126.5, 126.5, 126.4, 118.9 (2C), 114.5 (2C), 113.9 (2C), 113.9 (4C), 101.9 (Cꢀ1), 101.7 (Cꢀ1),
101.7 (2C, Cꢀ1), 101.6 (Cꢀ1), 100.7 (Cꢀ1), 100.7 (PhCH), 100.4 (2C, PhCH), 79.4, 79.3, 76.23, 76.19,
75.5, 75.1, 74.53, 74.51, 74.46 (2C), 73.8, 73.7 (CH2ꢀPMB), 73.39 (CH2ꢀPMB), 73.35 (CH2ꢀPMB),
73.2 (2C), 72.6, 72.2, 72.2, 72.14, 72.11, 68.6 (2C), 68.0 (3C), 67.5, 66.5 (2C), 66.2, 62.2, 62.0, 60.8,
55.7 (OCH3ꢀOMP), 55.5 (OCH3ꢀPMB), 55.44 (OCH3ꢀPMB), 55.36 (OCH3ꢀPMB), 37.8 (CH2ꢀLev),
29.7 (CH3ꢀLev), 28.1 (CH2ꢀLev). HRMS (ESIꢀTOF) m/z [M + Na]+ Calcd for C135H131N9NaO40
2540.8391, found 2540.8352.
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pꢀMethoxyphenyl Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1
→4)ꢀOꢀ(2,3ꢀdiꢀOꢀbenzylꢀβꢀᴅꢀglucopyranosyl)ꢀ(1→3)ꢀOꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀβꢀᴅꢀgal
actopyranosyl)ꢀ(1→4)ꢀ2,3ꢀdiꢀOꢀbenzoylꢀβꢀᴅꢀglucopyranoside (17). To a solution of 14 (73 mg, 0.04
mmol) in CH2Cl2 (2 mL) coverd with aluminum foil were added water (106 ꢁL) and DDQ (23 mg, 0.1
mmol). The reaction mixture was stirred for 5 h at room temperature, quenched with sat. NaHCO3 aq.,
and extracted with CH2Cl2 (3 ×). The organic layer was washed with brine, dried over anhydrous
Na2SO4, and concentrated. Silica gel column chromatography (CH2Cl2/MeOH, 10:1) afforded 17 (64
1
mg) quantitatively as a white foam. H NMR (500 MHz, CDCl3): δ 7.94ꢀ7.92 (m, 8H, ArHꢀBz), 7.48
(m, 2H, ArHꢀBz), 7.41ꢀ7.20 (m, 24H, ArHꢀBz, benzylidene, overlapped with solvent peak), 6.88 (d, J =
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8.5 Hz, 2H, ArHꢀOMP), 6.75 (d, J = 8.5 Hz, 2H, ArHꢀOMP), 5.75 (t, J = 9.5 Hz, 1H, H1ꢀ3), 5.68 (t, J =
8.5 Hz, 1H, H3ꢀ3), 5.56 (t, J = 8.5 Hz, 1H, H1ꢀ2), 5.36 (t, J = 7.5 Hz, 1H, H3ꢀ2), 5.27 (s, 1H,
CHꢀbenzylidene), 5.23 (s, 1H, CHꢀbenzylidene), 5.18 (d, J = 8.0 Hz, 1H, H1ꢀ1), 5.10 (d, J = 6.5 Hz, 1H,
H3ꢀ1), 4.63 (dd, J = 10.5, 3.0 Hz, 1H, H4ꢀ3), 4.38 (d, J = 8.0 Hz, 1H, H2ꢀ1), 4.32ꢀ4.28 (m, 3H, H1，3ꢀ4,
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H4ꢀ1), 4.12 (s, 1H, H4ꢀ4), 4.05ꢀ3.97 (m, 5H, H2ꢀ4, H1 3ꢀ6a,b), 3.80 (t, J = 9.0 Hz, 1H, H1ꢀ5), 3.72ꢀ3.67
，
(m, 6H, H3ꢀ5, OCH3ꢀOMP, H2ꢀ6a,b), 3.61 (brꢀs, 2H, H4ꢀ6a,b), 3.56ꢀ3.49 (m, 3H, H2ꢀ3, H2，4ꢀ2), 3.06 (s,
1H, H2ꢀ5), 2.93 (s, 1H, H4ꢀ5), 2.71ꢀ2.69 (m, 2H, CH2ꢀLev), 2.59ꢀ2.57 (m, 2H, CH2ꢀLev), 2.06 (s, 3H,
CH3ꢀLev); 13C NMR (125 MHz, CDCl3): δ 206.2 (COꢀLev), 172.0 (COꢀLev), 165.6 (COꢀBz), 165.4
(2C, COꢀBz), 165.2 (COꢀBz), 155.8, 151.1, 137.8, 137.6, 133.3, 133.2, 133.0, 132.9, 129.9, 129.9,
129.7, 129.7, 129.4, 129.4, 129.0, 129.0, 128.5, 128.4, 128.3, 128.2, 128.1, 126.6, 118.7 (2C), 114.7
(2C), 102.2, 101.9, 101.1, 100.8, 100.7, 100.7, 78.7, 75.8, 75.5, 75.3, 75.1, 74.6, 73.7, 73.2, 72.8, 72.3,
72.1, 68.2, 68.1, 66.6, 66.3, 62.3, 61.2, 60.8, 60.7, 55.7 (OCH3ꢀOMP), 53.6, 37.9 (CH2ꢀLev), 29.7
(CH3ꢀLev), 28.2 (CH2ꢀLev). HRMS (ESIꢀTOF) m/z [M + Na]+ Calcd for C78H76N6NaO26 1535.4707,
found 1535.4673.
Methy [pꢀMethoxyphenyl Oꢀ(2ꢀAzidoꢀ4,6ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀ3ꢀOꢀlevulinoylꢀβꢀᴅꢀgalacto
pyranosyl)ꢀ(1→4)ꢀOꢀ(methy 2,3ꢀdiꢀOꢀbenzoylꢀβꢀᴅꢀglucopyranosyluronate)ꢀ(1→3)ꢀOꢀ(2ꢀAzidoꢀ4,6
ꢀOꢀbenzylideneꢀ2ꢀdeoxyꢀβꢀᴅꢀgalactopyranosyl)ꢀ(1→4)ꢀ(2,3ꢀdiꢀOꢀbenzoylꢀβꢀᴅꢀglucopyranoside)]
Uronate (18). To a solution of glucoside 16 (377 mg, 0.25 mmol) in a mixture of acetonitrile/buffer
(Na2CO3/ NaHCO3, PH 9.5) (1/1, 5 mL), (2,2,6,6ꢀtetramethylpiperidinꢀ1ꢀyl)oxyl (TEMPO, 46.7 mg,
0.30 mmol), (diacetoxyiodo)ꢀbenzene (BAIB, 602.1 mg, 1.87 mmol) and tetrabutylammonium bromide
(TBAB, 11.7 mg, 0.075 mmol) were added. The reaction mixture was stirred at room temperature for 4
h. Then the reaction was quenched by addition of sat. Na2S2O3 aq. followed by separation of the layers.
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The aqueous phase was extracted with EtOAc (3 ×), and the combined organic phase was washed with
brine, dried over Na2SO4, and concentrated to afford a yellow syrup. The crude acid, K2CO3 (103.2 mg,
0.75 mmol) and CH3I (124 ꢁL, 1.99 mmol) were suspended in DMF (4 mL). After stirring for 30 min,
the reaction mixture was queched with water, diluted with EtOAc (3 ×), washed with brine, dried over
unhydrous Na2SO4 and concentrated. Silica gel column chromatography (Acetone/petroleum
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ether/CH2Cl2, 4:1:4) afforded 18 (318 mg, 81% for two steps) as a white foam. H NMR (600 MHz,
CDCl3): δ 7.97ꢀ7.93 (m, 5H, ArHꢀBz), 7.90 (d, J = 7.2 Hz, 2H, ArHꢀBz), 7.52 ꢀ 7.24 (m, 23H, ArHꢀBz,
benzylidene), 6.91 (d, J = 9.0 Hz, 2H, ArHꢀOMP), 6.77 (d, J = 9.6 Hz, 2H, ArHꢀOMP), 5.77 (t, J = 9.0
Hz, 1H, H1ꢀ3), 5.65ꢀ5.60 (m, 2H, H3ꢀ3, H4ꢀ3), 5.34 (t, J = 6.6 Hz, 1H, H3ꢀ2), 5.38 (s, 1H,
CHꢀbenzylidene), 5.21 (d, J = 6.6 Hz, 1H, H1/3ꢀ1), 5.21 (s, 1H, CHꢀbenzylidene), 5.12 (d, J = 6.0 Hz,
1H, H1/3ꢀ1), 4.55 (dd, J = 9.6, 8.4 Hz, 1H, H1ꢀ2), 4.51ꢀ4.48 (m, 2H, H1,3ꢀ4), 4.36 (d, J = 7.8 Hz, 1H,
H4ꢀ1), 4.31ꢀ4.29 (m, 2H, H1,3ꢀ5), 4.25 (d, J = 3.0 Hz, 1H, H4ꢀ4), 4.25 (d, J = 8.4 Hz, 1H, H2ꢀ1), 3.99 (d,
J = 3.0 Hz, 1H, H2ꢀ4), 3.82 (s, 3H, OCH3ꢀCOOMe), 3.75 (s, 3H, OCH3ꢀCOOMe), 3.74 (s, 3H,
OCH3ꢀCOOMe), 3.72ꢀ3.69 (m, 1H, H4ꢀ2), 3.66ꢀ3.61 (m, 2H, H2ꢀ2, Hꢀ6a), 3.43ꢀ3.40 (m, 3H, H2ꢀ3,
2Hꢀ6a,b), 3.34 (d, J = 9.0 Hz, 1H, Hꢀ6b), 3.11 (s, 1H, H4ꢀ5), 2.72ꢀ2.69 (m, 3H, CH2ꢀLev, H2ꢀ5),
2.61ꢀ2.58 (m, 2H, CH2ꢀLev), 2.07 (s, 3H, CH3ꢀLev); 13C NMR (150 MHz, CDCl3): δ 206.2 (COꢀLev),
172.1 (COꢀLev), 168.7 (COꢀCOOMe), 168.3 (COꢀCOOMe), 165.4 (COꢀBz), 165.3 (COꢀBz), 165.2
(COꢀBz), 164.9 (COꢀBz), 155.9, 151.0, 138.3, 137.7, 133.4, 133.2, 133.0, 130.1, 129.9, 129.6, 129.6,
129.2, 129.1, 128.9, 128.5, 128.4, 128.2, 128.1, 128.1, 126.7, 126.6, 118.8, 114.7, 102.7, 102.5, 101.0,
101.0, 100.7, 100.6, 78.9, 78.0, 77.8, 74.5, 74.1, 73.5, 73.39, 72.37, 72.2 (2C), 71.9, 71.8, 68.0, 67.9,
66.7, 66.2, 62.1, 60.6, 55.8 (CH3ꢀOMP), 53.1 (CH3ꢀCOOMe), 53.0 (CH3ꢀCOOMe), 37.9 (CH2ꢀLev),
29.8 (CH3ꢀLev), 28.2 (CH2ꢀLev). HRMS (ESIꢀTOF) m/z [M + Na]+. Calcd for C80H76N6NaO28
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