Synthesis and antibacterial activity of aminosugar-functionalized intercalating agents

Article abstract of DOI:10.1016/j.carres.2009.10.010

A series of previously reported amino sugar-functionalized intercalating agents, 3-14, were evaluated in two antibacterial assays (paper disk diffusion and 96-well microdilution) against Bacillus atrophaeus, ATCC 9372 and Escherichia coli, ATCC 47076. Alt

Full text of DOI:10.1016/j.carres.2009.10.010

Carbohydrate Research 345 (2010) 10–22
Contents lists available at ScienceDirect
Carbohydrate Research
journal homepage: www.elsevier.com/locate/carres
Synthesis and antibacterial activity of aminosugar-functionalized
intercalating agents
*
Wei Shi, Sandra L. Marcus, Todd L. Lowary
Alberta Ingenuity Centre for Carbohydrate Science and Department of Chemistry, The University of Alberta, Gunning-Lemieux Chemistry Centre, Edmonton, AB, Canada T6G 2G2
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 4 October 2009
Received in revised form 10 October 2009
Accepted 13 October 2009
Available online 20 October 2009
A series of previously reported amino sugar-functionalized intercalating agents, 3–14, were evaluated in
two antibacterial assays (paper disk diffusion and 96-well microdilution) against Bacillus atrophaeus,
ATCC 9372 and Escherichia coli, ATCC 47076. Although none of the compounds were active against this
E. coli strain, several showed activity against B. atrophaeus. In anticipation of the need for larger amounts
of these compounds for future structure–activity relationship studies, improved routes to 11–14 were
developed.
Keywords:
Anthracyclines
Synthesis
Ó 2009 Elsevier Ltd. All rights reserved.
Antibacterial
1. Introduction
most known naturally occurring anthracyclines are toxic to hu-
mans at the dosage required for treating bacterial infections, re-
Since the discovery of the pharmaceutical potential of daunoru-
bicin (1, Dauno) and doxorubicin (2, Dox), two of the most impor-
tant members of the anthracycline family of antibiotics (Chart 1),1,2
significant research effort has been directed toward the identifica-
tion of semi-synthetic analogs with better pharmaceutical efficacy.
This work has led to the discovery of a few other anthracyclines for
the clinical treatment of cancer: idarubicin, valrubicin, epirubicin,
pirarubicin, and aclarubicin.3 These anthracycline antibiotics re-
main in the mainstream of chemotherapeutics against a variety
of tumors, either alone or in combination with other drugs.4
It is believed that one of the major modes of action of the
anthracycline antibiotics arises from their binding to DNA, thus
preventing transcription or translation, and, in turn, leading to cell
death.5,6 Indeed, previous structural studies have demonstrated
that these molecules bind to DNA by intercalation of the anthra-
quinone moiety (rings B–D) between adjacent base pairs in the
DNA helix, while ring A and the carbohydrate moiety interact with
the minor groove.7,8
search on the use of anthracyclines as antibacterial agents has
been limited.10
In an earlier paper,11 we described the synthesis of a series of
compounds (3–14, Chart 2) containing an aromatic domain linked
to an amino sugar, including those found in both 1 and 2. These
compounds were designed as functional mimics of anthracyclines
in that 3–14 possess a flat aromatic ring that can intercalate be-
tween DNA base pairs attached to a carbohydrate residue, which
we anticipated could interact with the minor groove when the aro-
matic domain was intercalated into DNA. These compounds were,
indeed, shown to bind to DNA, albeit less strongly than either 1 or
2.11 In this paper, we describe the testing of 3–14 for their ability to
O
OH
O
13
1
11
12
10
R
9
2
3
14
OH
D
C
B
A
8
In addition to their anti-cancer potential, the earliest discovered
anthracyclines, such as those in the rhodomycin family, also dis-
played potent antibacterial activity in culture.9 However, their high
toxicity in mice precluded further development as therapeutic
drugs.9 On the other hand, the anthracyclines identified later, such
as Dauno, showed less toxicity at the dosage for clinical cancer
treatment, but exhibited weaker antibacterial activity.1a Because
4
5
6
7
OMe
O
OH
5'
O
6'
1'
O
3'
4'
2'
NH2
HO
1. Daunorubicin: R = H
2. Doxorubicin: R = OH
Chart 1. Structures of daunorubicin (1, Dauno) and doxorubicin (2, Dox). The
anthraquinone domain is composed of rings B, C, and D.
* Corresponding author. Tel.: +1 780 492 1861.
E-mail address: tlowary@ualberta.ca (T.L. Lowary).
0008-6215/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2009.10.010

W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
11
O
O
O
O
O
O
X
X
X
HO
NH2
NH2
HO
NH2
3 X = O
4 X = S
7 X = O
11 X = O
12 X = S
8 X = S
5
6
9
13
14
X = NTs
X = NH
X = NTs
X = NH
X = NTs
X = NH
10
Chart 2. Structures of anthracycline analogs 3–14.
act as antibacterial agents against model Gram-negative and
Gram-positive organisms, as well as the refinement of the syn-
thetic approaches to some of the target molecules for further bio-
logical evaluation, as well as structure–activity relationship (SAR)
studies.
this problem, we performed the more sensitive 96-well microdilu-
tion assay,12 which requires a smaller amount of sample, thus
allowing us to work at lower concentrations of each compound.
The MIC values (the minimum inhibition concentration at
which 80–100% of bacterial growth is inhibited) resulting from
the 96-well microdilution assays are also reported in Table 1. More
specific MIC values could be obtained by additional assays at con-
centrations within this range; however, this was not done due to
relatively low antibacterial activity of these compounds. It is only
assured that at the top of the concentration range, the compound
showed 100% inhibition, while at the lower concentration of the
range obvious bacterial growth (>20%) was observed.
There are some obvious discrepancies between the results
with each assay. For example, compound 7, which has low water
solubility as determined by the formation of a precipitate of the
compound in the paper disk assay, did not show any inhibition
in this assay, but it was among the most potent analogs in the
microdilution assay. However, regardless of the assay method,
compared to Dauno and Dox, which showed stronger DNA-bind-
ing affinity than 3–14,11 the antibacterial activity of the most po-
tent synthetic analogs is about twofold weaker. Considering the
slow diffusion of 3–14 in aqueous media, we consider the results
from the 96-well microdilution assay to be more reliable. In this
regard, it is of interest to note that, except for compound 12,
there is a good correlation between DNA-binding affinity and
the MIC range; that is, strong DNA binding seems to benefit anti-
bacterial activity.
2. Results and discussion
2.1. Antibacterial activity of aminosugar analogs 3–14
To evaluate the potential antibacterial activity of 3–14, one
Gram-positive bacteria (Bacillus atrophaeus, ATCC 9372) and one
Gram-negative bacteria (Escherichia coli, ATCC 47076) were se-
lected as the target organisms to screen against. Two different anti-
bacterial assays were applied, a paper disk diffusion assay and a
96-well microdilution assay.12
The results from the paper disk assay for 3–14 are listed in Ta-
ble 1. Using this assay, none of these compounds prevented the
growth of E. coli; therefore, only the results for B. atrophaeus are
shown. Compared to Dauno and Dox, as well as three positive con-
trols (streptomycin, penicillin, and tetracycline), 3–14 all showed
much lower antibacterial activity. In addition, in this assay there
is no correlation between the antibacterial activity and the DNA-
binding affinity for these compounds determined in our earlier
study.11 However, it was found that the water solubility of many
of the compounds was low, and we therefore anticipated that the
diffusion of these analogs on the agar plate could be slow. To avoid
Table 1
Paper disk diffusion and 96-well microdilution assay results for compounds 3–14 against Bacillus atrophaeus, ATCC 9372
Compound
Rel. DNA binding
Disk content of compound (nmol)
Zone diameter (mm)b
Range for MIC (lM)
(% remaining EtBr)a (%)
Streptomycin
Penicillin
Tetracycline
Dauno
Dox
3
4
5
6
7
8
9
10
11
12
13
14
N/A
N/A
N/A
N/A
N/A
62.0
67.5
47.9
76.7
50.5
73.6
49.7
76.5
68.5
79.9
38.6
78.4
14 (10
19 (6.8
68 (30
30
30
30
30
30
30
30
30
30
30
30
30
30
30
l
l
l
g)
g, 10 U)
g)
19
28
29
19
16
8
9
6
NDc
ND
ND
3.125–6.25
3.125–6.25
6.25–12.5
6.25–12.5
6.25–12.5
25–50
6.25–12.5
12.5–25
6.25–12.5
25–50
No inhibition
No inhibition
7
7
6
8
8
6
6
12.5–25
12.5–25
6.25–12.5
25–50
a
Taken from Ref. 11. This number represents the percentage of remaining EtBr that is intercalated into DNA after incubation with the compound. The smaller the number,
the greater the DNA-binding affinity of the compound.
b
Mean values. All compounds were measured in duplicate.
ND = not determined.
c

12
W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
O
O
NH2
15
I
I
I
N
Ts
O
S
18
16
17
O
O
O
O
O
S
NTs
O
O
NH2
NH2
NH2
11
12
13
Figure 1. Previous synthesis of compounds 11–13 using Pd(PPh3)2Cl2 and CuI as catalysts.11
2.2. Chemistry
2.2.1. Model studies
Before exploring an improved route to 13, we carried out a ser-
ies of model reactions using the phenylthiophene derivative 17
(Scheme 1), which was more synthetically accessible than the cor-
responding tosylated phenylindole species, and thus was available
in larger amount. We first studied the Sonogashira coupling be-
tween 17 and propargyl glycoside 20, which was synthesized in
good yield from previously reported methyl glycoside 19.11 In
the course of optimizing the reaction conditions (Table 2), removal
of oxygen by sparging the solution with argon before the addition
of the metal catalysts (entry 2 vs 1) was essential. It has been re-
ported that the presence of oxygen greatly accelerates the homo-
coupling reaction of the alkyne,13 and a large amount of this
product was detected in the absence of argon. In addition, piperi-
dine was found to be a better solvent than triethylamine (entries
Among analogs 3–14, compound 13, which consists of a 1-tosyl-
2-phenylindol-3-yl group conjugated to a 4-amino-2,3,4,6-tetrade-
oxy-a-L-hexopyranoside moiety, showed the best DNA-binding
affinity11 and was also among the compounds with the best anti-
bacterial activity against B. atrophaeus. However, the yield for its
synthesis developed earlier was low (Fig. 1).11 Direct Sonogashira
coupling of the amino propargyl compound 15 with benzofuran
or benzothiophene iodide 16 or 17 went smoothly, but almost no
coupled product 13 could be observed under similar conditions.
Even after the microwave irradiation was applied, the yield was
still lower than 20%. Therefore, we decided to explore alternate ap-
proaches to 13, which would enable the preparation of larger
amounts of material for further studies.
OCH3
O
O
a
b
S
O
O
AcO
AcO
O
I
AcO
19
20
21b
S
17
O
c
d
e
O
S
O
S
S
O
O
O
HO
N3
NH2
12
23
22
Scheme 1. Reagents and conditions: (a) propargyl alcohol, BF3ÁEt2O, 95%; (b) 5% Pd(PPh3)2Cl2, 10% CuI, piperidine, 72%; (c) K2CO3, methanol, 99%; (d) PPh3, DPPA, DIAD,
b
À20 °C? rt, 80%; (e) PPh3, H2O 87%. The pure b anomers were isolated, and the subsequent transformations were performed separately see Section 2.2.3.

W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
13
Table 2
Optimization of the Sonogashira reaction of 17 and 20
Mol %
Mol %
Base
Sparging
Mol %
Time (h)
Yield (%)
Pd(PPh3)Cl2 (%)
CuI (%)
PPh3 (%)
1
2
3
4
5
6
7
2–5
5
5
3
5
1–5
1–2
1–2
1–2
10
Et3N
Et3N
No
0
0
0
0
0
16
12
12
14
10
27
12
ꢀ5
40
74
65
91
89
93
Yes
Yes
Yes
Yes
Yes
Yes
Piperidine
Piperidine
Piperidine
Piperidine
Piperidine
5
5
10
10
10
3
3–7); the use of pyrrolidine led to partial removal of the acetyl
group.14 High loadings of both palladium (ꢀ5%) and copper cata-
lysts (ꢀ10 mol %) were also necessary (entry 4 vs 3, 5 vs 4).
Although triphenylphosphine has been reported to stabilize the
palladium catalyst, especially under thermal conditions,15 its addi-
tion showed no significant effect on the ambient reactions used
here (entries 6 and 7 vs 5). Indeed, the use of 10% triphenylphos-
phine significantly reduced the reaction rate (entry 6 vs 5).
After the optimization of the Sonogashira coupling, removal of
the acetyl-protecting group in 21 produced 22 in excellent yield.
Next, Mitsunobu reaction with diphenylphosphoryl azide (DPPA)
gave an 80% yield of 23.16 The last step, the conversion of the azide
to an amine in using the Staudinger reaction proceeded smoothly
to give 12 in 87% yield.
Prior to the use of DPPA for the synthesis of 23 from 22, we ex-
plored two other approaches, both of which gave unexpected re-
sults. Initially, we thought it would be possible to install the
azido group into 22 via SN2 substitution of the corresponding mes-
ylate derivative (Scheme 2). Thus, reaction of 22 with methanesul-
fonyl chloride and triethylamine in dichloromethane afforded
compound 24 in near quantitative yield; however, the following
substitution with sodium azide was not very successful. When
the substitution reaction was carried out with sodium azide in
DMF at 120 °C, the yield of the desired product 23 was only 40%.
Two major side products were formed, the 1,2,3-triazole 25 (30%
yield), produced from [2+3] cycloaddition between azide and the
alkyne, and alkene 26 (19% yield) arising from elimination.
The structure of 25 was supported by mass spectrometry, 1H
NMR and IR spectroscopy. In the IR spectrum, the signal
(ꢀ2200 cmÀ1) for the alkyne disappeared, and signals (1733,
1600, 1433, 1215 cmÀ1) for a 1,2,3-triazole appeared, although
they are not very characteristic.17 In the 13C NMR spectrum, the al-
kyne signals at ꢀ80 ppm and ꢀ90 ppm also disappeared; however,
three signals for quaternary aromatic carbons in 25 were missing,
presumably due to their very long relaxation times. Efforts to
change the pulse delay led to two of three signals being recovered,
but not all three. The alkene compound 26 was confirmed by the
shift of H/C-3 and H/C-4 signals from the alkane region to the al-
kene region in both the 1H NMR and 13C NMR spectra. Mass anal-
ysis also clearly verified the loss of one molecule of
methanesulfonic acid. By decreasing the reaction temperature to
80–85 °C, these two byproducts were avoided to some degree,
especially the 1,2,3-triazole, but the reaction time was substan-
tially extended, usually requiring more than 2–3 days.
Another approach explored for the preparation of 23 from 22
involved the generation of the more reactive triflate derivative
(Scheme 3). Thus, crude 27 after the workup of the triflation reac-
tion of 22 (Tf2O, pyridine) was treated with sodium azide in DMF at
room temperature. As expected, none of triazole or elimination
product was formed. However, the yield of 23 was only slightly im-
proved (49%), and two new side products, formate esters 28 and
29, were generated.
A possible mechanism for the formation of 28 and 29 is shown
in Scheme 3. We propose that the formyl group is added via reac-
O
O
a
O
S
O
S
HO
MsO
22
24
N
N
HN
O
S
b
O
+
+
S
O
S
O
O
N3
O
N3
23
25
26
Scheme 2. Reagents and conditions: (a) MsCl, Et3N, 0 °C, 99%; (b) NaN3, DMF, 120 °C, 40% for 23, 30% for 25, 19% for 26.

14
W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
O
O
DMF
NaN3
DMF
S
O
S
S
O
O
O
O
NaN3
N
TfO
N
O
H
O
H
H
H2O
30
27
31
N
H2O
O
O
S
S
O
O
O
O
O
O
H
H
29 (13%)
28 (27%)
Scheme 3.
tion of triflate intermediate 27 with the solvent DMF. Although
DMF is a poor nucleophile, it is present in large excess, and the con-
centration of sodium azide in the reaction mixture is relatively low
due to its poor solubility in DMF at room temperature. The reaction
would be further facilitated by the highly deoxygenated carbohy-
drate ring in 27, which would enhance the electrophilicity of C-4,
compared to the more oxygenated carbohydrate derivatives. We
propose that intermediate 30 is first formed. Thereafter, due to
the presence of a large amount of DMF, the stereochemistry of C-
4 in 30 can be inverted to give 31, and an equilibrium may be
established between them. In theory, 31 should be favored due
to the equatorial orientation of the iminium ion at C-4. However,
after the aqueous workup, which gave rise to the release of
dimethylamine to produce the corresponding formate esters, 29
was produced in a larger amount than 30, by a ratio of ꢀ2:1. There-
fore, it appears that the conversion from 30 to 31 is a slow process.
2.2.2. Synthesis of 11, 13 and 14
After succeeding in the synthesis of benzo[b]thiophene analogs
12, we applied the same strategy outlined in Scheme 1 to the ben-
zo[b]furan and indole-containing systems, 11 and 13, respectively.
As outlined in Scheme 4, using this route compound 11 was syn-
thesized in 69% overall yield from 16 and 20 with no difficulties
(Scheme 4).
The synthesis of the indole derivative 13 via the approach de-
picted in Scheme 1 turned out to be more challenging than for
O
b
O
a
O
O
AcO
O
I
AcO
20
32b
O
16
O
c
d
O
O
O
O
O
O
O
O
HO
N3
NH2
11
34
33
Scheme 4. Reagents and conditions: (a) 5% Pd(PPh3)2Cl2, 10% CuI, piperidine, 73%; (b) K2CO3, methanol, 99%; (c) PPh3, DPPA, DIAD, À20 °C?rt, 83%; (d) PPh3, H2O 91%. b The
pure b anomers were isolated, and the subsequent transformations were performed separately, see Section 2.2.3.

W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
15
O
b
O
a
X
O
AcO
O
I
AcO
20
N
Ts
35
36
X = NTs
X = NH
e
18
O
c
d
O
NTs
O
X
NTs
O
O
O
HO
N3
NH2
13
X = NTs
37
38
e
14
X = NH
Scheme 5. Reagents and conditions: (a) 5% Pd(PPh3)2Cl2, 10% CuI, piperidine, 51%; (b) K2CO3, methanol, 89%; (c) PPh3, DPPA, DIAD, À20 °C?rt, 73%; (d) PPh3, H2O 86%; (e) n-
b
Bu4NF, THF, rt, 61% for 36 and 75% for 14. The pure b anomers were isolated, and the subsequent transformations were performed separately see Section 2.2.3.
the thiophene or furan systems. Fortunately, the coupling between
propargyl glycoside 20 and tosyl-protected indole iodide 18 was
successful, despite giving the product 35 in relatively modest yield
(Scheme 5). It was found that the increased loading of the palla-
dium catalyst did not improve the reaction yield significantly.
However, it was particularly crucial to protect the reaction from
air during the entire process. The rest of the steps for the synthesis
of target amine 13 from 35 were straightforward, although the
reaction time for the deacylation of 35 needed to be controlled to
minimize the concomitant cleavage of the tosyl group under the
basic reaction conditions. The target 13, was obtained from 35 in
56% yield over three steps. Removal of the tosyl group in 13 upon
reaction with tetra-n-butylammonium fluoride gave the free in-
dole 14 in 75% yield.
In identifying methods for removing the N-tosylate group in
these indoles, we initially explored heating with potassium
hydroxide in methanol at reflux.18 In a model study, we attempted
to deprotect 37 using these conditions; however, only ꢀ20% of the
desired free indole product 38, contaminated with a number of
other unknown compounds, was obtained (Scheme 6). The major
product, obtained in 65% yield, was ketone 40. A possible mecha-
nism for the formation of 40 from 37 is illustrated in Scheme 6.
After the removal of the tosyl group, protonation of the alkyne
could give an allene 41, which then undergoes a Michael-like
H
CH3O
B
O
O
O
S
H
a
O
O
+
N
NH
O
NH
O
HO
O
HO
O
HO
37
39
40
H-B
N
HO
NH
O
C
O
O
HO
O
OH
HO
42
41
Scheme 6. Reagents and conditions: (a) KOH, CH3OH, reflux, 65% for 40.

16
W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
1,4-conjugate addition of hydroxide anion generating the enol 42,
which tautomerizes to the more stable ketone 40. Based on this
mechanism, the formation of 40 can be prevented by employing
an aprotic solvent and a base that is a poor nucleophile. The use
of the tetra-n-butylammonium fluoride in THF thus satisfies both
these criteria. In addition to being successful in cleaving the tosyl
group from 13 to yield 14, these conditions could also be used to
prepare compound 36 from 35 (Scheme 5).
alumina and copper in a PURESOLV-400 System from Innovative
Technology, Inc. under an argon atmosphere. Unless stated other-
wise, all reactions were carried out under a positive pressure of ar-
gon and were monitored by TLC on silica gel G-25 UV254 (0.25 mm,
Macherey–Nagel). Spots were detected under UV light and/or by
charring with 10% H2SO4 in ethanol, or in acidified ethanolic anis-
aldehyde or vanillin. Solvents were evaporated under reduced
pressure and below 40 °C (water bath). Column chromatography
was performed on Silica Gel 60 (40–60 lM). The ratio between sil-
2.2.3. Synthesis of -glycoside analogs of 11–14 and related
precursors
The synthesis of 11–14 as outlined above all began with propar-
gyl glycoside 16, which was synthesized as a 3.8:1 a/b mixture (see,
e.g., Scheme 1). Although separation of this mixture of glycosides
was possible, it was difficult and the purification was more conve-
niently done after the aromatic group was added via Sonogashira
coupling. This approach also allowed the synthesis of 12 additional
compounds, 43–54 (Chart 3) possessing the b stereochemistry at
the anomeric center. The preparation of these compounds followed
ica gel and crude product ranged from 100 to 50:1 (w/w). Optical
rotations were measured at 22 2 °C. Melting points are uncor-
rected. 1H NMR spectra were recorded on VARIAN INOVA NMR
spectrometers at 400, 500 or 600 MHz, and chemical shifts are ref-
erenced to either TMS (0.0, CDCl3) or CD2HOD (4.78, CD3OD). 13C
NMR spectra were recorded at 100 or 125 MHz, and 13C chemical
shifts are referenced to CDCl3 (77.23 ppm, CD3Cl3) or CD3OD
(49.00 ppm, CD3OD). 1H data are reported as though they were first
order. The errors between the coupling constants for two coupled
protons were less than 0.5 Hz, and the average number was re-
ported. Assignment of NMR resonances was done based on
1H–1H COSY, HMQC, and in some cases HMBC experiments. In
the interpretation of the NMR data for methylene protons on the
carbohydrate ring, ‘a’ and ‘e’ refer to axial and equatorial orienta-
tion, respectively. In the cases where no clear assignment of these
hydrogens could be made based on all NMR data, assignments
were made taking into consideration the anisotropy effect of a ring
the routes outlined above for the corresponding
a anomers, and de-
tails can be found in Supplementary data. Access to 43–54 will be
useful in future SAR studies on this class of compounds.
2.3. Conclusions
In summary, we describe here the testing of compounds 3–14 as
potential antibacterial agents against both Gram-negative and
Gram-positive bacteria. Although none of the compounds showed
activity against the Gram-negative organism evaluated (E. coli, ATCC
47076) using the preliminary disk diffusion assay, several of the
compounds were active against B. atrophaeus, ATCC 9372, the model
Gram-positive organism we investigated, by using both disk diffu-
sion and microdilution assays. Some of the compounds possess
low micromolar MIC values against B. atrophaeus thus suggesting
that the synthesis of additional analogs of this class of compounds
is warranted (e.g., through functionalization of the amino group).
Tothatend, amoreefficientsyntheticapproachto13hasbeendevel-
oped, in whichcompared to anearlier route, theoverall yield was en-
hanced from ꢀ8% to ꢀ35% from the same starting materials. The
same approach could also be used for an improved synthesis of com-
pounds11, 12, and 14 and allowedthe preparation of additional ana-
logs, for example, those with b-glycoside stereochemistry.
r
bond, which results in equatorial hydrogens resonating more
downfield than axial hydrogens.19 Electrospray-ionization mass
spectra were recorded on samples suspended in mixtures of THF
with CH3OH and added NaCl. Optical rotations were measured on
a Perkin–Elmer 241 polarimeter at the sodium D line (589 nm).
Optical rotations are in units of deg mL(dm g)À1. IR spectra were
recorded on the Nicolet Magna 750 FTIR spectrometer. The re-
ported purity values were obtained with a Varian HPLC system,
using an evaporative light scattering detector (ELSD) 2000ES from
Alltech, and a Varian Microsorb-MV 100-5 C18 column. The eluant
consisted of acetonitrile and water, the ratio of which depends on
the compound. For all basic amino compounds, 0.1% by volume of
trifluoroacetic acid was added to facilitate elution and avoid aggre-
gation. When the purity derived from HPLC analysis is greater than
99.5%, it will be reported as >99%.
3.2. General reaction procedures
3. Experimental
3.2.1. General procedure for Sonogashira coupling
To a degassed solution of propargyl glycoside 20 (1.3 equiv), the
aromatic iodide (1 equiv), PdCl2(PPh3)2 (5 mol %) in piperidine (5–
7 mL/mmol) was added CuI (10 mol %). The reaction mixture was
stirred under Ar at rt and followed by TLC (10–16 h). Once com-
plete, the reaction was quenched by the addition of a satd aq soln
3.1. General methods—chemistry
All reagents were purchased from commercial sources and were
used without further purification unless noted. Before use, reaction
solvents were purified by successive passage through columns of
O
O
O
R1
R1
R1
O
O
O
O
S
NTs
R2
R2
R2
47 R1 = OAc, R2 = H
48 R1 = OH, R2 = H
49 R1 = H, R2 = N3
50 R1 = H, R2 = NH2
51 R1 = OMs, R2 = H
43 R1 = OAc, R2 = H
44 R1 = OH, R2 = H
45 R1 = H, R2 = N3
46 R1 = H, R2 = NH2
52 R1 = OH, R2 = H
53 R1 = H, R2= N3
54 R1 = H, R2 = NH2
Chart 3. Structures of the synthesized b anomers.

W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
17
of NH4Cl. The aqueous solution was then extracted with Et2O, and
3.6. 3-(2-Phenylindol-3-yl)prop-2-ynyl 4-amino-2,3,4,6-
tetradeoxy- -threo-hexopyranoside (14)
the combined organic layers were washed with brine, dried
(Na2SO4), filtered, and concentrated to yield the crude product,
which was purified by column chromatography.
a-L
This compound was obtained from 13 following the general
procedure for detosylation. The product was purified by column
chromatography (10:1, CH2Cl2–CH3OH) in 75% yield. The data
matched that reported previously.11
3.2.2. General procedure for deacetylation
The acetyl-protected compound (1 equiv) was dissolved in
CH3OH (30–40 mL/mmol). K2CO3 (30 mol %) was added, and then
the reaction mixture was stirred at rt and followed by TLC
(ꢀ12 h). After the evaporation of the solvent, the residue was di-
luted with water and extracted with CH2Cl2. The combined organic
layer was dried (Na2SO4), filtered, and concentrated to yield the
crude product, which was purified by column chromatography.
3.7. 2-Propargyl 4-O-acetyl-2,3,6-trideoxy-
hexopyranoside (20 ) and 2-propargyl 4-O-acetyl-2,3,6-
trideoxy-b- -erythro-hexopyranoside (20b)
a-L-erythro-
a
L
To a flask were added a mixture of 19 (455 mg, 2.42 mmol),
crushed activated 4 Å molecular sieves (290 mg) and propargyl
alcohol (10 mL). The mixture was stirred for 10 min at rt, cooled
to À40 °C, and then BF3ÁEt2O (0.768 mL, 6.05 mmol) was added
dropwise via a syringe. After adding the BF3ÁEt2O, the reaction mix-
ture was warmed to À20 °C. Once the starting material was fully
consumed (about 5 h), the reaction mixture was neutralized by
the addition of K2CO3 (500 mg). Next, H2O (12 mL), satd aq NaHCO3
(12 mL), and CH2Cl2 (30 mL) were added. The organic layer was
separated, washed with brine, dried (Na2SO4), filtered, and concen-
trated. The crude product was purified by column chromatography
3.2.3. General procedure for Mitsunobu reactions
To a solution of the hydroxyl compound (1 equiv) in THF (15–
20 mL/mmol) was added PPh3 (3 equiv) at À20 °C. To this mixture
was added a solution of DIAD (2.5 equiv) and DPPA (2.5 equiv) in
THF (5–7 mL/mmol) at À20 °C. The reaction mixture was then al-
lowed to warm to rt and followed by TLC (ꢀ12 h). The resulting
solution was then diluted with Et2O, washed with brine, dried
(Na2SO4), filtered, and concentrated to yield the crude product,
which was purified by column chromatography.
(10:1 hexanes–EtOAc) to give pure 20
and 20b as a colorless oil (487 mg, 95%,
a
as a colorless waxy solid
:b = 3.8:1). Data for
3.2.4. General procedure for Staudinger reactions
a
To a solution of the azido compound (1 equiv) in THF (30–
40 mL/mmol) and H2O (>50 equiv) was added PPh3 (2.5 equiv),
and the reaction was stirred under reflux and followed by TLC
(ꢀ10 h). The solution was cooled to room temperature, and con-
centrated, and the residue was purified by column chromatogra-
phy to yield the pure amino compound.
20
a: Rf 0.65 (6:1 hexanes–EtOAc); IR: m 2216 (C„C), 1736 (C@O)
cmÀ1; [
a]
À196.6 (c 0.9, CH2Cl2); 1H NMR (400 MHz, CDCl3): dH
D
4.86 (br s, 1H, H-1) 4.40–4.48 (m, 1H, H-4), 4.18 (dd, 1H,
J = 15.8 Hz, J = 2.4 Hz, OCH2C„CH), 4.12 (dd, 1H, J = 15.8 Hz,
J = 2.4 Hz, OCH2C„CH), 3.74 (dq, 1H, J4,5 = 9.7 Hz, J5,6 = 6.2 Hz, H-
5), 2.38 (t, 1H, J = 2.4 Hz, OCH2C„CH), 1.98 (s, 3H, O@CCH3),
1.68–1.90 (m, 4H, H-2a, H-2e, H-3a, H-3e), 1.09 (d, 3H,
J5,6 = 6.2 Hz, H-6); 13C NMR (100 MHz, CDCl3): dC 170.1 (C@O),
94.9 (C-1), 79.4 (C„CH), 74.1 (C„CH), 73.2 (C-4), 66.9 (C-5), 53.9
(OCH2), 28.8 (C-2), 23.9 (C-3), 21.1 (O@CCH3), 17.7 (C-6). ESIMS:
m/z calcd for [C11H16O4]Na+: 235.0941. Found: 235.0943. Anal.
Calcd for C11H16O4: C, 62.25; H, 7.60. Found: C, 62.36; H, 7.78. Data
3.2.5. General procedure for tosylate removal
To a solution of the tosylate (1 equiv) in THF (30–40 mL/
mmol) was added tetra-n-butylammonium fluoride (TBAF) solu-
tion in THF (1.0 M, 6 equiv) at rt, and the mixture was stirred
at rt and followed by TLC (ꢀ18 h). Satd aq NaHCO3 (30 mL) was
then added, and the mixture was extracted with CH2Cl2. The or-
ganic layer was washed with brine, dried over Na2SO4, filtered,
and concentrated to a residue that was purified by column
chromatography.
for 20b: Rf 0.63 (6:1 hexanes–EtOAc); IR:
m 2220 (C„C), 1737
(C@O) cmÀ1; [ D +56.1 (c 1.4, CH2Cl2); 1H NMR (400 MHz, CDCl3):
a]
dH 4.68 (dd, 1H, J1,2a = 9.0 Hz, J1,2e = 2.3 Hz, H-1), 4.45 (ddd, 1H,
J3a,4 = 10.4 Hz, J4,5 = 8.9 Hz, J3e,4 = 4.2 Hz, H-4), 4.34 (d, 2H,
J = 2.4 Hz, OCH2C„CH), 3.53 (dq, 1H, J4,5 = 8.9 Hz, J5,6 = 6.2 Hz, H-
5), 2.44 (t, 1H, J = 2.4 Hz, OCH2C„CH), 2.16 (dddd, 1H,
J3a,3e = 13.0 Hz, J2e,3e = J3e,4 = J2a,3e = 4.2 Hz, H-3e), 2.05 (s, 3H,
O@CCH3), 1.88–1.93 (m, 1H, H-2e), 1.66 (dddd, 1H, J2a,2e = -
J2a,3a = 13.0 Hz, J1,2a = 9.0 Hz, J2a,3e = 4.2 Hz, H-2a), 1.50 (dddd, 1H,
J3a,3e = J2a,3a = 13.0 Hz, J3a,4 = 10.4 Hz, J2e,3a = 4.3 Hz, H-3a), 1.22 (d,
3H, J5,6 = 6.2 Hz, H-6); 13C NMR (100 MHz, CDCl3): dC 170.2
(C@O), 98.9 (C-1), 79.3 (C„CH), 74.4 (C„CH), 73.3 (C-5), 72.7 (C-
4), 55.0 (OCH2), 29.7 (C-2), 26.9 (C-3), 21.1 (O@CCH3), 18.1 (C-6).
ESIMS m/z calcd for [C11H16O4]Na+: 235.0941. Found: 235.0944.
3.3. 3-(2-Phenylbenzo[b]furan-3-yl)prop-2-ynyl 4-amino-
2,3,4,6-tetradeoxy-a-L-threo-hexopyranoside (11)
This compound was obtained from 34 following the general
procedure for Staudinger reactions. The product was purified by
column chromatography (1:1 hexanes–EtOAc?10:1 CH2Cl2–
CH3OH) in 91% yield. The data matched that reported previously.11
3.4. 3-(2-Phenylbenzo[b]thiophen-3-yl)prop-2-ynyl 4-amino-
2,3,4,6-tetradeoxy-a-L-threo-hexopyranoside (12)
3.8. 3-(2-Phenylbenzo[b]thiophen-3-yl)prop-2-ynyl 4-O-acetyl-
This compound was obtained from 23 following the general
procedure for Staudinger reactions. The product was purified by
column chromatography (1:1 hexanes–EtOAc?10:1 CH2Cl2–
CH3OH) in 87% yield. The compound characterization data could
be found in Ref. 11.
2,3,6-trideoxy-
phenylbenzo[b]thiophen-3-yl)prop-2-ynyl 4-O-acetyl-2,3,6-
trideoxy-b- -erythro-hexopyranoside (47)
a-L-erythro-hexopyranoside (21) and 3-(2-
L
This compound was synthesized from 20 (237 mg, 1.12 mmol),
17 (283 mg, 0.841 mmol),11 PdCl2(PPh3)2 (28 mg, 5 mol %), and CuI
(15 mg, 10 mol %) in 91% combined yield with the b anomer
3.5. 3-(1-Tosyl-2-phenylindol-3-yl)prop-2-ynyl 4-amino-
2,3,4,6-tetradeoxy-
a-L
-threo-hexopyranoside (13)
(a:b = 3.8:1) by following general procedure for Sonogashira cou-
pling. The product was purified by column chromatography (10:1
hexanes–EtOAc). Data for 21: white needle-like solid after recrys-
tallization from hexanes–Et2O (2:1); mp: 89–91 °C; Rf 0.36 (6:1
This compound was obtained from 38 following the general
procedure for Staudinger reactions. The product was purified by
column chromatography (1:1, hexanes–EtOAc?10:1, CH2Cl2–
CH3OH) in 86% yield. The data matched that reported previously.11
hexanes–EtOAc); IR:
m a]
2216 (C„C), 1736 (C@O) cmÀ1; [
D
À120.2 (c 1.4, CH2Cl2); 1H NMR (400 MHz, CDCl3): dH 7.96–8.00

18
W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
(m, 2H, Ar), 7.92–7.96 (m, 1H, Ar), 7.79–7.82 (m, 1H, Ar), 7.36–7.50
(m, 5H, Ar), 5.08 (br s, 1H, H-1), 4.58 (s, 2H, OCH2C„C), 4.50–4.57
(m, 1H, H-4), 3.89 (dq, 1H, J4,5 = 9.7 Hz, J5,6 = 6.3 Hz, H-5), 2.06 (s,
3H, O@CCH3), 1.80–2.00 (m, 4H, H-2a, H-2e, H-3a, H-3e), 1.17 (d,
3H, J5,6 = 6.3 Hz, H-6); 13C NMR (100 MHz, CDCl3): dC 170.2
(C@O), 146.9 (Ar), 141.1 (Ar), 137.5 (Ar), 133.7 (Ar), 128.8 (Ar),
128.7 (2, Ar), 128.4 (2, Ar), 125.2 (Ar), 124.9 (Ar), 123.3 (Ar),
122.0 (Ar), 112.9 (Ar), 94.8 (C-1), 90.2 („C), 80.4 („C), 73.4 (C-
4), 67.0 (C-5), 54.8 (OCH2), 29.0 (C-2), 24.0 (C-3), 21.2 (O@CCH3),
17.8 (C-6). ESIMS m/z calcd for [C25H24O4S]Na+: 443.1288. Found:
443.1289. Anal. Calcd for C25H24O4S: C, 71.40; H, 5.75; S, 7.63.
Found: C, 71.34; H, 5.80; S, 7.41. Data for 47: yellow fluffy solid,
4.57 (AB q, 2H, J = 16.0 Hz, OCH2C„C), 4.07 (dq, 1H, J5,6 = 6.5 Hz,
J4,5 = 1.7 Hz, H-5), 3.47 (br s, 1H, H-4), 2.19 (dddd, 1H, J3a,3e = -
J2a,3a = 13.4 Hz, J3a,4 = 4.2 or 3.2 Hz, J2e,3a = 3.2 or 4.2 Hz, H-3a),
2.02 (dddd, 1H, J2a,2e = J2a,3a = 13.4 Hz, J1,2a = 4.0 Hz, J2a,3e = 4.0 Hz,
H-2a), 1.91–1.97 (m, 1H, H-3e), 1.62–1.68 (m, 1H, H-2e), 1.20 (d,
3H, J5,6 = 6.5 Hz, H-6); 13C NMR (100 MHz, CDCl3, dC) 147.0 (Ar),
141.1 (Ar), 137.5 (Ar), 133.7 (Ar), 128.8 (Ar), 128.7 (2, Ar), 128.4
(2, Ar), 125.3 (Ar), 125.0 (Ar), 123.2 (Ar), 122.1 (Ar), 121.9 (Ar),
95.7 (C-1), 90.2 („C), 80.4 („C), 65.7 (C-5), 60.0 (C-4), 55.1
(OCH2), 23.9 (C-2), 22.9 (C-3), 17.9 (C-6). ESIMS: m/z calcd for
C23H21N3O2S: 403.1354. Found: 403.1349. Anal. Calcd for
C23H21N3O2S: C, 68.46; H, 5.25; N, 10.41; S, 7.95. Found: C,
68.70; H, 5.10; N, 10.12; S, 7.93.
Rf 0.32 (6:1 hexanes–EtOAc); IR:
m 2217 (C„C), 1737 (C@O)
cmÀ1; [ +32.7 (c 0.9, CH2Cl2); 1H NMR (400 MHz, CDCl3, dH)
a
]
D
7.96–7.99 (m, 2H, Ar), 7.92–7.95 (m, 1H, Ar), 7.79–7.82 (m, 1H,
Ar), 7.36–7.50 (m, 5H, Ar), 4.83 (dd, 1H, J1,2a = 9.1 Hz,
J1,2e = 2.2 Hz, H-1), 4.72 (ABq, 1H, J = 16.0 Hz, OCH2C„C), 4.69
(ABq, 1H, J = 16.0 Hz, OCH2C„C), 4.48 (ddd, 1H, J3a,4 = 10.5 Hz,
J4,5 = 8.3 Hz, J3e,4 = 4.3 Hz, H-4), 3.54 (dq, 1H, J4,5 = 8.3 Hz,
J5,6 = 6.2 Hz, H-5), 2.16 (dddd, 1H, J3a,3e = 12.9 Hz, J2e,3e = 4.3 Hz,
J3e,4 = 4.3 Hz, J2a,3e = 4.3 Hz, H-3e), 2.06 (s, 3H, O@CCH3), 1.91–
1.97 (m, 1H, H-2e), 1.71 (dddd, 1H, J2a,2e = J2a,3a = 13.1 Hz,
J1,2a = 9.1 Hz, J2a,3e = 4.3 Hz, H-2a), 1.50 (dddd, 1H, J3a,3e
=J2a,3a = 13.1 Hz, J3a,4 = 10.5 Hz, J2e,3a = 4.3 Hz, H-3a), 1.22 (d, 3H,
J5,6 = 6.2 Hz, H-6); 13C NMR (100 MHz, CDCl3): dC 170.2 (C@O),
147.0 (Ar), 141.1 (Ar), 137.5 (Ar), 133.7 (Ar), 128.8 (Ar), 128.7 (2,
Ar), 128.4 (2, Ar), 125.3 (Ar), 125.0 (Ar), 123.2 (Ar), 122.1 (Ar),
112.9 (Ar), 98.8 (C-1), 89.9 („C), 80.7 („C), 73.3 (C-5), 72.8 (C-
4), 55.9 (OCH2), 29.9 (C-2), 27.0 (C-3), 21.2 (O@CCH3), 18.1 (C-6).
ESIMS: m/z calcd for [C25H24O4S]Na+: 443.1288. Found: 443.1290.
Anal. Calcd for C25H24O4S: C, 71.40; H, 5.75; S, 7.63. Found: C,
71.40; H, 5.73; S, 7.50.
3.11. 3-(2-Phenylbenzo[b]thiophen-3-yl) prop-2-ynyl 4-O-
methanesulfonyl-2,3,6-trideoxy-
a-L-erythro-hexopyranoside
(24) and 3-(2-phenylbenzo[b]thiophen-3-yl)prop-2-ynyl 4-O-
methanesulfonyl-2,3,6-trideoxy-b-L-erythro-hexopyranoside
(51)
The mixture of 22 and its b anomer (42 mg, 0.11 mmol) was dis-
solved in CH2Cl2 (5 mL), and Et3N (0.047 mL, 0.34 mmol) was
added. The solution was cooled to 0 °C and then mesyl chloride
(0.013 mL, 0.17 mmol) was added. After stirring for 2 h, the reac-
tion mixture was washed sequentially with 1 N HCl, 1 N NaOH
and brine. The solution was dried (Na2SO4), and filtered, and the
solvent was evaporated to give a yellow sirup that was purified
by column chromatography (4:1 hexanes–EtOAc) to give pure 24
and 51, both as a yellow oils (51 mg, 99% as combined yield,
a
:b = 3.8:1). Data for 24: Rf 0.55 (2:1 hexanes–EtOAc); IR: m 2216
(C„C) cmÀ1; [
a]
À105.9 (c 2.8, CH2Cl2); 1H NMR (400 MHz,
D
CDCl3): dH 7.96–8.00 (m, 2H, Ar), 7.92–7.95 (m, 1H, Ar), 7.79–
7.82 (m, 1H, Ar), 7.36–7.50 (m, 5H, Ar), 5.08 (br d, 1H, J1,2a
or
3.9. 3-(2-Phenylbenzo[b]thiophen-3-yl)prop-2-ynyl 2,3,6-
trideoxy-a-L-erythro-hexopyranoside (22)
2e = 2.8 Hz, H-1), 4.58 (s, 2H, OCH2C„C), 4.31–4.38 (m, 1H, H-4),
3.92 (dq, 1H, J4,5 = 9.4 Hz, J5,6 = 6.2 Hz, H-5), 3.03 (s, 3H, SO2CH3),
2.10–2.20 (m, 2H, H-3a, H-3e), 1.92–1.98 (m, 1H, H-2e), 1.83–
1.91 (m, 1H, H-2a), 1.28 (d, 3H, J5,6 = 6.2 Hz, H-6); 13C NMR
(100 MHz, CDCl3): dC 147.1 (Ar), 141.1 (Ar), 137.5 (Ar), 133.6
(Ar), 128.9 (Ar), 128.7 (2, Ar), 128.4 (2, Ar), 125.3 (Ar), 125.0 (Ar),
123.2 (Ar), 122.1 (Ar), 112.8 (Ar), 94.6 (C-1), 89.9 („C), 80.6(4)
(C-4), 80.6(2) („C), 66.6 (C-5), 55.0 (OCH2), 38.9 (SO2CH3), 29.2
(C-2), 25.5 (C-3), 17.9 (C-6). ESIMS: m/z calcd for [C24H24O5S2]Na+:
479.0957. Found: 479.0953. Anal. Calcd for C24H24O5S2: C, 63.13;
H, 5.30; S, 14.05. Found: C, 63.07; H, 5.40; S, 14.05. Data for 51:
This compoundwas synthesizedas a colorless waxy solidfrom 21
(176 mg, 0.418 mmol)and K2CO3 (18 mg, 0.13 mmol)in99%yieldby
following the general procedure for deacetylation. The product was
purified by column chromatography (2:1 hexanes–EtOAc): yellow
sirup, Rf 0.34 (2:1 hexanes–EtOAc); IR:
m 3415 (O–H), 2215 (C„C)
cmÀ1; [
a
]
D
À96.3 (c 1.8, CH2Cl2); 1H NMR (400 MHz, CDCl3): dH
7.96–8.00 (m, 2H, Ar), 7.92–7.96 (m, 1H, Ar), 7.79–7.82 (m, 1H, Ar),
7.36–7.49 (m, 5H, Ar), 5.07 (br s, 1H, H-1), 4.57 (s, 2H, OCH2C„C),
3.66 (dq, 1H, J4,5 = 9.2 Hz, J5,6 = 6.3 Hz, H-5), 3.26–3.34 (m, 1H, H-
4), 1.75–1.94 (m, 4H, H-2a, H-2e, H-3a, H-3e), 1.41 (br s, 1H, OH),
1.17 (d, 3H, J5,6 = 6.3 Hz, H-6); 13C NMR (100 MHz, CDCl3): dC 146.9
(Ar), 141.1 (Ar), 137.5 (Ar), 133.7 (Ar), 128.8 (Ar), 128.7 (2, Ar),
128.4 (2, Ar), 125.2 (Ar), 125.0 (Ar), 123.3 (Ar), 122.0 (Ar), 113.0
(Ar), 94.7 (C-1), 90.5 („C), 80.3 („C), 72.0 (C-4), 70.0 (C-5), 54.7
(OCH2), 29.5 (C-2), 27.6 (C-3), 17.9 (C-6). ESIMS: m/z calcd for
[C23H22O3S]Na+: 401.1182. Found: 401.1186. Anal. Calcd for
C23H22O3S: C, 72.99; H, 5.86; S, 8.47. Found: C, 72.56; H, 5.85; S, 8.59.
Rf 0.47 (2:1 hexanes–EtOAc); IR:
m a] +73.4
2218 (C„C) cmÀ1; [
D
(c 1.3, CH2Cl2); 1H NMR (500 MHz, CDCl3): dH 7.95–7.98 (m, 2H,
Ar), 7.91–7.94 (m, 1H, Ar), 7.80–7.83 (m, 1H, Ar), 7.37–7.49 (m,
5H, Ar), 4.83 (dd, 1H, J1,2a = 8.7 Hz, J1,2e = 2.3 Hz, H-1), 4.72 (ABq,
1H, J = 16.0 Hz, OCH2C„C), 4.67 (ABq, 1H, J = 16.0 Hz, OCH2C„C),
4.30 (ddd, 1H, J3a,4 = J4,5 = 10.4 Hz, J3e,4 = 4.7 Hz, H-4), 3.54 (dq,
1H, J4,5 = 8.7 Hz, J5,6 = 6.2 Hz, H-5), 3.03 (s, 3H, SO2CH3), 2.36–2.40
(m, 1H, H-3e), 1.96–2.01 (m, 1H, H-2e), 1.68–1.82 (m, 2H, H-3a,
H-2a), 1.22 (d, 3H, J5,6 = 6.2 Hz, H-6); 13C NMR (125 MHz, CDCl3):
dC 147.2 (Ar), 141.1 (Ar), 137.5 (Ar), 133.7 (Ar), 128.9 (Ar), 128.7
(2, Ar), 128.4 (2, Ar), 125.3 (Ar), 125.0 (Ar), 123.2 (Ar), 122.1 (Ar),
112.8 (Ar), 98.5 (C-1), 89.6 („C), 80.9 („C), 79.9 (C-5), 73.0 (C-
4), 56.1 (OCH2), 38.8 (SO2CH3), 29.9 (C-2), 28.3 (C-3), 18.2 (C-6).
ESIMS: m/z calcd for [C24H24O5S2]Na+: 443.1288. Found:
443.1289. Purity: >99%.
3.10. 3-(2-Phenylbenzo[b]thiophen-3-yl)prop-2-ynyl 4-azido-
2,3,4,6-tetradeoxy-a-L-threo-hexopyranoside (23)
This compound was synthesized as a yellow oil from 22 (49 mg,
0.13 mmol), PPh3 (102 mg, 0.389 mmol), DIAD (66 mg, 0.33 mmol),
and DPPA (89 mg, 0.33 mmol) in 80% yield by following the general
procedure for Mitsunobu reactions. The product was purified by
column chromatography (20:1 hexanes–EtOAc): yellow oil, Rf
3.12. 4-(2-Phenylbenzo[b]thiophen-3-yl)-1,2,3-triazol-5-yl-
methyl 4-azido-2,3,4,6-tetradeoxy-a-L-threo-hexopyranoside
0.33 (15:1 hexanes–EtOAc); IR:
m
2214 cmÀ1 (C„C), 2098 cmÀ1
(25)
(N@N@N); [
a
]
D
À67.7 (c 2.8, CH2Cl2); 1H NMR (400 MHz, CDCl3):
dH 7.96–8.00 (m, 2H, Ar), 7.92–7.96 (m, 1H, Ar), 7.79–7.82 (m,
Compound 24 (69 mg, 0.15 mmol) was dissolved in DMF (5 mL),
1H, Ar), 7.36–7.49 (m, 5H, Ar), 5.14 (br d, 1H, J1,2a = 4.0 Hz, H-1),
and NaN3 (99 mg, 1.5 mmol) was added. The reaction mixture was

W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
19
heated to 120 °C and stirred overnight. After cooling, the reaction
mixture was dissolved in water and extracted with EtOAc. The or-
ganic layer was dried (Na2SO4), and filtered, and the solvent was
evaporated to give an orange oil that was purified by column chro-
matography (10:1 hexanes–EtOAc) to give pure 25 (20 mg, 30%) as
(C@O), 147.0 (Ar), 141.1 (Ar), 137.5 (Ar), 133.7 (Ar), 128.8 (Ar),
128.7 (2, Ar), 128.4 (2, Ar), 125.2 (Ar), 124.9 (Ar), 123.2 (Ar),
122.0 (Ar), 112.9 (Ar), 94.7 (C-1), 90.1 („C), 80.5 („C), 73.2 (C-
4), 66.8 (C-5), 54.9 (OCH2), 28.9 (C-2), 24.1 (C-3), 17.8 (C-6). ESIMS:
m/z calcd for [C24H22O4S]Na+: 429.1131. Found: 429.1133. Anal.
Calcd for C24H22O4S: C, 70.91; H, 5.46; S, 7.89. Found: C, 70.49;
H, 5.40; S, 8.09.
a yellow oil: Rf 0.40 (8:1 hexanes–EtOAc); IR:
m
2099 (N@N@N),
1734, 1600, 1433, 1215 (trizole) cmÀ1; [
a]
À50.1 (c 0.5, CH2Cl2);
D
1H NMR (500 MHz, CDCl3): dH 7.86–7.90 (m, 1H, Ar), 7.46–7.50
(m, 1H, Ar), 7.32–7.41 (m, 4H, Ar), 7.27–7.31 (m, 3H, Ar), 4.61 (br
d, 1H, J1,2a or 2e = 3.5 Hz, H-1), 4.37 (ABq, 1H, J = 16.2 Hz, OCH2C„C),
4.23 (ABq, 1H, J = 16.2 Hz, OCH2C„C), 3.25 (dq, 1H, J5,6 = 6.2 Hz,
J4,5 = 1.8 Hz, H-5), 3.05 (br s, 1H, H-4), 1.57–1.75 (m, 3H, H-2e, H-
3a, H-3e), 1.24–1.30 (m, 1H, H-2a), 0.84 (d, 3H, J5,6 = 6.2 Hz, H-6);
13C NMR (125 MHz, CDCl3): dC (missing one carbon signal) 143.7
(Ar), 140.6 (Ar), 138.5 (Ar), 133.4 (Ar), 129.2 (Ar), 128.9 (Ar),
128.8 (2, Ar), 128.6 (2, Ar), 125.0 (Ar), 124.9 (Ar), 123.0 (Ar),
122.1 (Ar), 120.6 (Ar), 96.6 (C-1), 65.1 (C-5), 59.7 (C-4), 59.3
(OCH2), 23.6 (C-2), 22.5 (C-3), 17.7 (C-6). ESIMS: m/z calcd for
[C23H22 N6O2S]Na+: 469.1417. Found: 469.1418.
3.14. 3-(2-Phenylbenzo[b]furan-3-yl)prop-2-ynyl 4-O-acetyl-
2,3,6-trideoxy-
phenylbenzo[b]furan-3-yl)prop-2-ynyl 4-O-acetyl-2,3,6-
trideoxy-b- -erythro-hexo pyranoside (43)
a-L-erythro-hexopyranoside (32) and 3-(2-
L
This compound was synthesized from 20 (220 mg, 1.04 mmol),
16 (256 mg, 0.799 mmol),11 PdCl2(PPh3)2 (28 mg, 5 mol %), and CuI
(15 mg, 10 mol %) in 92% combined yield with the b anomer
(a:b = 3.8:1) by following general procedure for Sonogashira cou-
pling. The product was purified by column chromatography (10:1
hexanes–EtOAc). Data for 32: solid yellow paste, Rf 0.46 (4:1 hex-
anes–EtOAc); IR:
m
2222 (C„C), 1737 (C@O) cmÀ1; [
a
]
D
À108.0 (c
3.13. 3-(2-Phenylbenzo[b]thiophen-3-yl)prop-2-ynyl 4-O-
5.6, CH2Cl2); 1H NMR (500 MHz, CDCl3): dH 8.26–8.30 (m, 2H, Ar),
7.66–7.70 (m, 1H, Ar), 7.47–7.53 (m, 3H, Ar), 7.39–7.43 (m, 1H,
Ar), 7.28–7.36 (m, 2H, Ar), 5.13 (br s, 1H, H-1), 4.64 (s, 2H,
OCH2C„C), 4.52–4.58 (m, 1H, H-4), 3.93 (dq, 1H, J4,5 = 9.6 Hz,
J5,6 = 6.3 Hz, H-5), 2.06 (s, 3H, O@CCH3), 1.83–2.01 (m, 4H, H-2a,
H-2e, H-3a, H-3e), 1.20 (d, 3H, J5,6 = 6.3 Hz, H-6); 13C NMR
(125 MHz, CDCl3): dC 170.2 (C@O), 156.8 (Ar), 153.4 (Ar),
129.9(9) (Ar), 129.9(6) (Ar), 129.2 (Ar), 128.6 (2, Ar), 126.0 (2,
Ar), 125.3 (Ar), 123.4 (Ar), 120.3 (Ar), 111.2 (Ar), 98.5 (Ar), 95.0
(C-1), 92.6 („C), 77.8 („C), 73.4 (C-4), 67.1 (C-5), 55.0 (OCH2),
29.0 (C-2), 24.1 (C-3), 21.2 (O@CCH3), 17.9 (C-6). ESIMS: m/z calcd
for [C25H24O5]Na+: 427.1516. Found: 427.1517. Anal. Calcd for
C25H24O5: C, 74.24; H, 5.98. Found: C, 74.50; H, 6.33. Data for 43:
formyl-2,3,6-trideoxy-
phenylbenzo[b]thiophen-3-yl)prop-2-ynyl 4-O-formyl-2,3,6-
trideoxy- -erythro-hexopyranoside (29)
a-L-threo-hexopyranoside (28) and 3-(2-
a-L
To a solution of compound 22 (251 mg, 0.659 mmol) in 19:1
CH2Cl2–pyridine (10 mL) at À20 °C was added a solution of Tf2O
(0.25 mL, 1.5 mmol) in CH2Cl2 (5 mL). After stirring for 1.5 h while
keeping the temperature below À10 °C, TLC showed the starting
material was consumed and a new spot (Rf 0.66, 4:1 hexanes–
EtOAc) appeared. The reaction mixture was then extracted with
ice-cold 1 M HCl and water. The organic layer was dried (Na2SO4),
filtered, and concentrated to yield an orange liquid, and the result-
ing product was immediately dissolved in dry DMF (10 mL), and
NaN3 (430 mg, 6.60 mmol) was added. After stirring at rt for 3 h,
the reaction was quenched by the addition of water, and the result-
ing mixture was extracted with EtOAc. The organic solution was
dried (Na2SO4) and filtered. After evaporation of the solvent, the
residue was purified by column chromatography (15:1 hexanes–
EtOAc?8:1 hexanes–EtOAc) to yield pure 23 (130 mg, 49%) as a
colorless oil (characterization data are listed above), 28 (72 mg,
27%) as a yellow oil, and 29 (35 mg, 13%) as a white waxy solid.
white amorphous solid, Rf 0.44 (4:1 hexanes–EtOAc); IR:
m 2219
(C„C), 1737 (C@O) cmÀ1; [ +37.9 (c 0.6, CH2Cl2); 1H NMR
a]
D
(400 MHz, CDCl3): dH 8.26–8.30 (m, 2H, Ar), 7.66–7.69 (m, 1H,
Ar), 7.47–7.53 (m, 3H, Ar), 7.40–7.44 (m, 1H, Ar), 7.29–7.37 (m,
2H, Ar), 4.90 (dd, 1H, J1,2a = 9.1 Hz, J1,2e = 2.3 Hz, H-1), 4.77 (s, 2H,
OCH2C„C),
4.52
(ddd,
1H,
J3a,4 = 10.5 Hz,
J4,5 = 9.0 Hz,
J3e,4 = 4.5 Hz, H-4), 3.60 (dq, 1H, J4,5 = 9.0 Hz, J5,6 = 6.2 Hz, H-5),
2.16 (dddd, 1H, J3a,3e = 13.4 Hz, J2e,3e = J3e,4 = J2a,3e = 4.5 Hz, H-3e),
2.06 (s, 3H, O@CCH3), 1.96–2.02 (m, 1H, H-2e), 1.75 (dddd, 1H,
J2a,2e = J2a,3a = 13.4 Hz, J1,2a = 9.1 Hz, J2a,3e = 4.5 Hz, H-2a), 1.54
(dddd, 1H, J3a,3e = J2a,3a = 13.4 Hz, J3a,4 = 10.5 Hz, J2e,3a = 4.4 Hz, H-
3a), 1.29 (d, 3H, J5,6 = 6.2 Hz, H-6); 13C NMR (100 MHz, CDCl3): dC
170.2 (C@O), 156.8 (Ar), 153.4 (Ar), 129.9(9) (Ar), 129.9(8) (Ar),
129.3 (Ar), 128.6 (2, Ar), 126.0 (2, Ar), 125.4 (Ar), 123.4 (Ar),
120.2 (Ar), 111.2 (Ar), 99.0 (C-1), 98.4 (Ar), 92.2 („C), 78.1 („C),
73.4 (C-5), 72.8 (C-4), 56.1 (OCH2), 29.9 (C-2), 27.0 (C-3), 21.1
(O@CCH3), 18.2 (C-6). ESIMS: m/z calcd for [C25H24O5]Na+:
427.1516. Found: 427.1517. Purity: 99.3%.
Data for 28: Rf 0.41 (4:1 hexanes–EtOAc); IR
m 2216.1 (C„C),
1721.5 (C@O) cmÀ1
;
[
a]
À77.1 (c 2.8, CH2Cl2); 1H NMR
D
(500 MHz, CDCl3): dH 8.20 (d, J = 0.9 Hz, O@CH), 7.96–8.00 (m,
2H, Ar), 7.92–7.95 (m, 1H, Ar), 7.79–7.82 (m, 1H, Ar), 7.36–7.50
(m, 5H, Ar), 5.18 (br d, 1H, J1,2a = 4.0 Hz, H-1), 4.98 (br s, 1H, H-
4), 4.59 (s, 2H, OCH2C„C), 4.11 (br q, J5,6 = 6.6 Hz, H-5), 2.16 (dddd,
1H, J3a,3e = J2a,3a = 13.8 Hz, J3a,4 = 3.5 Hz, J2e,3a = 3.5 Hz, H-3a), 2.02
(dddd, 1H, J2a,2e = J2a,3a = 13.8 Hz, J1,2a = 4.0 Hz, J2a,3e = 4.0 Hz, H-
2a), 1.83–1.89 (m, 1H, H-3e), 1.64–1.69 (m, 1H, H-2e), 1.12 (d,
3H, J5,6 = 6.6 Hz, H-6); 13C NMR (100 MHz, CDCl3): dC 160.6
(C@O), 147.0 (Ar), 141.1 (Ar), 137.5 (Ar), 133.7 (Ar), 128.8 (Ar),
128.7 (2, Ar), 128.4 (2, Ar), 125.2 (Ar), 124.9 (Ar), 123.2 (Ar),
122.1 (Ar), 112.9 (Ar), 95.7 (C-1), 90.2 („C), 80.4 („C), 69.3 (C-
4), 65.1 (C-5), 55.1 (OCH2), 23.8 (C-2), 22.9 (C-3), 17.1 (C-6). ESIMS:
m/z calcd for [C24H22O4S]Na+: 429.1131. Found: 429.1133. Anal.
Calcd for C24H22O4S: C, 70.91; H, 5.46; S, 7.89. Found: C, 70.77;
3.15. 3-(2-Phenylbenzo[b]furan-3-yl)prop-2-ynyl 2,3,6-
trideoxy-a-L-erythro-hexopyranoside (33)
This compound was synthesized as a colorless waxy solid from
32 (131 mg, 0.358 mmol) and K2CO3 (15 mg, 0.11 mmol) in 99%
yield by following the general procedure for deacetylation. The
product was purified by column chromatography (2:1 hexanes–
H, 5.49; S, 7.98. Data for 29: Rf 0.39 (4:1 hexanes–EtOAc); IR
m
2217.7 (C„C), 1725.1 (C@O) cmÀ1; [
a
]
D
À117.9 (c 1.5, CH2Cl2);
EtOAc): Rf 0.21 (3:1 hexanes–EtOAc); IR: m 3416 (O–H), 2222
1H NMR (500 MHz, CD3OD): dH 8.09 (d, J = 0.9 Hz, O@CH), 7.96–
8.00 (m, 2H, Ar), 7.92–7.95 (m, 1H, Ar), 7.79–7.82 (m, 1H, Ar),
7.36–7.50 (m, 5H, Ar), 5.10 (br d, 1H, J1,2a = 3.7 Hz, H-1), 4.61–
4.67 (m, 1H, H-4), 4.59 (s, 2H, OCH2C„C), 3.93 (dq, J4,5 = 9.3 Hz,
J5,6 = 6.2 Hz, H-5), 1.83–2.02 (m, 4H, H-3a, H-3e, H-2a, H-2e), 1.19
(d, 3H, J5,6 = 6.2 Hz, H-6); 13C NMR (100 MHz, CDCl3): dC 160.2
(C„C) cmÀ1; [
a]
À109.7 (c 8.4, CH2Cl2); 1H NMR (500 MHz,
D
CDCl3): dH 8.26–8.30 (m, 2H, Ar), 7.66–7.70 (m, 1H, Ar), 7.47–
7.53 (m, 3H, Ar), 7.39–7.43 (m, 1H, Ar), 7.28–7.36 (m, 2H, Ar),
5.12 (br s, 1H, H-1), 4.63 (s, 2H, OCH2C„C), 3.71 (dq, 1H,
J4,5 = 9.2 Hz, J5,6 = 6.2 Hz, H-5), 3.30–3.36 (m, 1H, H-4), 1.78–1.98
(m, 4H, H-2a, H-2e, H-3a, H-3e), 1.55 (br s, 1H, OH), 1.30 (d, 3H,

20
W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
J5,6 = 6.2 Hz, H-6); 13C NMR (125 MHz, CDCl3): dC 156.8 (Ar), 153.4
(Ar), 130.0(0) (Ar), 129.9(8) (Ar), 129.2 (Ar), 128.7 (2, Ar), 126.0 (2,
Ar), 125.3 (Ar), 123.4 (Ar), 120.3 (Ar), 111.2 (Ar), 98.5 (Ar), 94.8 (C-
1), 92.7 („C), 77.7 („C), 72.1 (C-4), 70.0 (C-5), 54.8 (OCH2), 29.5
(C-2), 27.6 (C-3), 17.9 (C-6). ESIMS: m/z calcd for [C23H22O4]Na+:
385.1410. Found: 385.1412. Anal. Calcd for C23H22O4: C, 76.22; H,
6.12. Found: C, 75.83; H, 6.16.
0.75 mmol) in 61% yield by following the general procedure for
detosylation. The product was purified by column chromatography
(5:1, hexanes–EtOAc): Rf 0.33 (4:1 hexanes–EtOAc); IR:
m 2226 (C„C),
1734 (C@O) cmÀ1; [a D
]
À99.7 (c 1.5, CH2Cl2); 1H NMR (500 MHz,
CDCl3): dH 8.42 (br s, 1H, NH), 7.95–7.99 (m, 2H, Ar), 7.76 (d, 1H,
J = 7.6 Hz, Ar), 7.49 (t, 2H, J = 7.7 Hz, Ar), 7.39 (t, 2H, J = 7.7 Hz, Ar),
7.18–7.27 (m, 2H, Ar), 5.13 (br s, 1H, H-1), 4.60 (s, 2H, OCH2C„C),
4.51–4.57 (m, 1H, H-4), 3.92 (dq, 1H, J4,5 = 9.6 Hz, J5,6 = 6.3 Hz, H-5),
2.06 (s, 3H, O@CCH3), 1.84–1.98 (m, 4H, H-3a, H-3e, H-2a, H-2e),
1.20 (d, 3H, J5,6 = 6.3 Hz, H-6); 13C NMR (125 MHz, CDCl3): dC 170.3
(C@O), 139.9 (Ar), 135.2 (Ar), 131.5 (Ar), 130.5 (Ar), 128.9 (2, Ar),
128.4 (Ar), 126.5 (2, Ar), 123.5 (Ar), 120.9 (Ar), 120.1 (Ar), 111.0 (Ar),
95.3 (Ar), 94.7 (C-1), 88.9 („C), 80.4 („C), 73.5 (C-5), 66.9 (C-4),
55.2 (OCH2), 29.0 (C-3), 24.1 (C-2), 21.2 (O@CCH3), 17.9 (C-6). ESIMS:
m/z calcd for [C25H25NO4]Na+: 426.1676. Found: 426.1678. Anal. Calcd
for C25H25NO4: C, 74.42; H, 6.25; N, 3.47. Found: C, 74.76; H, 5.99; N,
3.46.
3.16. 3-(2-Phenylbenzo[b]furan-3-yl)prop-2-ynyl 4-azido-
2,3,4,6-tetradeoxy-a-L-threo-hexopyranoside (34)
This compound was synthesized as a yellow oil from 33
(156 mg, 0.430 mmol), PPh3 (338 mg, 1.29 mmol), DIAD (218 mg,
1.08 mmol), and DPPA (297 mg, 1.52 mmol) in 83% yield by follow-
ing the general procedure for Mitsunobu reactions. The product
was purified by column chromatography (20:1 hexanes–EtOAc):
Rf 0.18 (20:1 hexanes–EtOAc); IR:
m
2221 cmÀ1 (C„C), 2098 cmÀ1
(N@N@N); [
a
]
D
À72.1 (c 1.7, CH2Cl2); 1H NMR (500 MHz, CDCl3):
dH 8.26–8.30 (m, 2H, Ar), 7.66–7.70 (m, 1H, Ar), 7.47–7.53 (m,
3H, Ar), 7.39–7.43 (m, 1H, Ar), 7.28–7.36 (m, 2H, Ar), 5.14 (br d,
1H, J1,2a = 4.0 Hz, H-1), 4.63 (s, 2H, OCH2C„C), 4.13 (dq, 1H,
J5,6 = 6.5 Hz, J4,5 = 1.7 Hz, H-5), 3.49 (br s, 1H, H-4), 2.19 (dddd,
1H, J3a,3e = J2a,3a = 13.4 Hz, J3a,4 = 4.2 or 3.2 Hz, J2e,3a = 3.2 or 4.2 Hz,
H-3a), 2.02 (dddd, 1H, J2a,2e = J2a,3a = 13.4 Hz, J1,2a = J2a,3e = 4.0 Hz,
H-2a), 1.94–2.00 (m, 1H, H-3e), 1.67–1.73 (m, 1H, H-2e), 1.20 (d,
3H, J5,6 = 6.5 Hz, H-6); 13C NMR (125 MHz, CDCl3): dC 156.8 (Ar),
153.4 (Ar), 129.9(9) (Ar), 129.9(7) (Ar), 129.3 (Ar), 128.7 (2, Ar),
126.0 (2, Ar), 125.4 (Ar), 123.4 (Ar), 120.3 (Ar), 111.2 (Ar), 98.5
(Ar), 95.8 (C-1), 92.6 („C), 77.8 („C), 65.8 (C-5), 59.9 (C-4), 55.2
(OCH2), 24.0 (C-2), 23.0 (C-3), 21.2 (O@CCH3), 17.9 (C-6). ESIMS:
m/z calcd for [C23H21N3O3]Na+: 410.1475. Found: 410.1474. Anal.
Calcd for C23H21N3O3: C, 71.30; H, 5.46; N, 10.85. Found: C,
71.49; H, 5.56; N, 10.57.
3.19. 3-(1-Tosyl-2-phenylindol-3-yl)prop-2-ynyl 2,3,6-trideoxy-
a
-
L
-erythro-hexopyranoside (37) and 3-(1-tosyl-2-phenylindol-
3-yl)prop-2-ynyl 2,3,6-trideoxy-b- -erythro-hexopyranoside (52)
L
This compound was synthesized as a colorless waxy solid from
the mixture of 35 and its b anomer (229 mg, 0.409 mmol) and
K2CO3 (17 mg, 0.12 mmol) in 89% yield by following the general
procedure for deacetylation. The product was purified by column
chromatography (2:1 hexanes–EtOAc,
a:b = 3.8:1). Data for 36: Rf
0.40 (2:1 hexanes–EtOAc); IR:
m ;
3403 (O–H), 2222 (C„C) cmÀ1
[a]
À90.2 (c 1.7, CH2Cl2); 1H NMR (500 MHz, CDCl3): dH 8.30–
D
8.33 (m, 1H, Ar), 7.53–7.59 (m, 3H, Ar), 7.39–7.48 (m, 4H, Ar),
7.31–7.36 (m, 1H, Ar), 7.24–7.27 (m, 2H, Ar), 7.03–7.07 (d, 2H,
J = 8.5 Hz, Ar), 4.85 (br s, 1H, H-1), 4.35 (s, 2H, OCH2C„C), 3.54
(dq, 1H, J4,5 = 9.2 Hz, J5,6 = 6.2 Hz, H-5), 3.22–3.29 (m, 1H, H-4),
2.30 (s, 3H, PhCH3), 1.67–1.87 (m, 4H, H-3a, H-3e, H-2a, H-2e),
1.20 (d, 3H, J5,6 = 6.2 Hz, H-6); 13C NMR (125 MHz, CDCl3): dC
144.9 (Ar), 143.8 (Ar), 137.0 (Ar), 134.6 (Ar), 131.2 (2, Ar), 130.7
(Ar), 130.5 (Ar), 129.4 (2, Ar), 129.1 (Ar), 127.3 (2, Ar), 126.8 (2,
Ar), 125.7 (Ar), 124.6 (Ar), 120.1 (Ar), 116.5 (Ar), 107.5 (Ar), 94.6
(C-1), 90.4 („C), 77.8 („C), 72.0 (C-4), 69.8 (C-5), 54.5 (OCH2),
29.4 (C-2), 27.5 (C-3), 21.5 (PhCH3), 17.8 (C-6). ESIMS: m/z calcd
for [C30H29NO5S]Na+: 538.1659. Found: 538.1655. Purity: 98.9%.
3.17. 3-(1-Tosyl-2-phenylindol-3-yl)prop-2-ynyl 4-O-acetyl-
2,3,6-trideoxy-a-L-erythro-hexopyranoside (35)
This compound was synthesized as a yellow sirup from 20
(219 mg, 1.03 mmol) and 18 (488 mg, 1.03 mmol),11 PPh3 (11 mg,
4 mol %), PdCl2(PPh3)2 (36 mg, 5 mol %), and CuI (20 mg,
10 mol %) in 65% combined yield with its b anomer (
by following the general procedure for Sonogashira coupling. The
anomer 35 was purified by column chromatography (5:1 hex-
a:b = 3.8:1)
Data for 52: Rf 0.57 (1:1 hexanes–EtOAc); IR:
m 3440 (O–H), 2220
a
(C„C) cmÀ1; [ D +90.7 (c 1.0, CH2Cl2); 1H NMR (500 MHz, CDCl3):
a]
anes–EtOAc): Rf 0.36 (4:1 hexanes–EtOAc); IR:
m
2226 (C„C),
dH 8.30–8.33 (d, 1H, J = 8.4 Hz, Ar), 7.52–7.60 (m, 3H, Ar), 7.40–7.50
(m, 4H, Ar), 7.32–7.36 (m, 1H, Ar), 7.25–7.29 (m, 2H, Ar), 7.04–7.08
(d, 2H, J = 8.5 Hz, Ar), 4.53 (dd, 1H, J1,2a = 9.3 Hz, J1,2e = 2.2 Hz, H-1),
4.51 (ABq, 1H, J = 16.0 Hz, OCH2C„C), 4.47 (ABq, 1H, J = 16.0 Hz,
1736 (C@O) cmÀ1; [
a]
À86.5 (c 1.0, CH2Cl2); 1H NMR (500 MHz,
D
CDCl3): dH 8.30–8.33 (m, 1H, Ar), 7.53–7.59 (m, 3H, Ar), 7.39–
7.48 (m, 4H, Ar), 7.31–7.36 (m, 1H, Ar), 7.24–7.27 (m, 2H, Ar),
7.03–7.07 (d, 2H, J = 8.6 Hz, Ar), 4.86 (br s, 1H, H-1), 4.46–4.52
(m, 1H, H-4), 4.36 (s, 2H, OCH2C„C), 3.93 (dq, 1H, J4,5 = 9.6 Hz,
J5,6 = 6.2 Hz, H-5), 2.30 (s, 3H, PhCH3), 2.06 (s, 3H, O@CCH3),
1.89–1.94 (m, 1H, H-3e), 1.76–1.81 (m, 3H, H-2a, H-2e, H-3a),
1.20 (d, 3H, J5,6 = 6.2 Hz, H-6); 13C NMR (125 MHz, CDCl3): dC
170.2 (C@O), 144.9 (Ar), 143.8 (Ar), 137.0 (Ar), 134.7 (Ar), 131.2
(2, Ar), 130.7 (Ar), 130.5 (Ar), 129.3 (2, Ar), 129.1 (Ar), 127.3 (2,
Ar), 126.8 (2, Ar), 125.7 (Ar), 124.6 (Ar), 120.0 (Ar), 116.5 (Ar),
107.4 (Ar), 94.6 (C-1), 90.2 („C), 77.9 („C), 73.4 (C-5), 66.9 (C-
4), 54.6 (OCH2), 28.9 (C-3), 24.0 (C-2), 21.5 (PhCH3), 21.2
(O@CCH3), 17.8 (C-6). ESIMS: m/z calcd for [C32H31NO6S]Na+:
580.1764. Found: 580.1768. Anal. Calcd for C32H31NO6S: C, 68.92;
H, 5.60; N, 2.51; S, 5.75. Found: C, 68.53; H, 5.96; N, 2.38; S, 5.64.
OCH2C„C), 3.27 (ddd, 1H, J3a,4 = 10.5 Hz, J4,5 = 8.8 Hz, J3e,4
4.2 Hz, H-4), 3.19 (dq, 1H, J4,5 = 8.8 Hz, J5,6 = 6.1 Hz, H-5), 2.31 (s,
3H, PhCH3), 2.05 (dddd, 1H, J3a,3e = 13.0 Hz, J2e,3e = J3e,4 = J2a,3e
=
=
4.2 Hz, H-3e), 1.78–1.83 (m, 1H, H-2e), 1.52–1.62 (m, 2H, OH, H-
2a), 1.42 (dddd, 1H, J2a,3a = 13.5 Hz, J3a,3e = 13.0 Hz, J3a,4 = 10.5 Hz,
J2e,3a = 4.2 Hz, H-3a), 1.28 (d, 3H, J5,6 = 6.1 Hz, H-6); 13C NMR
(500 MHz, CDCl3): dC 143.3 (Ar), 142.2 (Ar), 135.2 (Ar), 133.0 (Ar),
129.5 (2, Ar), 129.0 (Ar), 128.9 (Ar), 127.7 (2, Ar), 127.5 (Ar),
125.7 (2, Ar), 125.2 (2, Ar), 124.1 (Ar), 123.0 (Ar), 118.4 (Ar),
114.8 (Ar), 105.7 (Ar), 97.1 (C-1), 88.3 („C), 76.5 („C), 74.1 (C-
5), 69.9 (C-4), 54.1 (OCH2), 29.2 (C-2), 28.8 (C-3), 19.9 (PhCH3),
16.4 (C-6). ESIMS: m/z calcd for [C30H29NO5S]Na+: 538.1659.
Found: 538.1658. Purity: 98.5%.
3.18. 3-(2-Phenylindol-3-yl)prop-2-ynyl 4-O-acetyl-2,3,6-
3.20. 3-(1-Tosyl-2-phenylindol-3-yl)prop-2-ynyl 4-azido-
trideoxy-
a-
L-erythro-hexopyranoside (36)
2,3,4,6-tetradeoxy-a-L-threo-hexopyranoside (38)
This compound was synthesized as a yellow sirup from 35
This compound was synthesized as a yellow oil from 37 (119 mg,
(51 mg, 0.13 mmol) and TBAF solution in THF (1.0 M, 0.75 mL,
0.231 mmol), PPh3 (181 mg, 0.688 mmol), DIAD (116 mg, 0.581

W. Shi et al. / Carbohydrate Research 345 (2010) 10–22
21
mmol), and DPPA (160 mg, 0.580 mmol) in 73% yield by following
the general procedure for Mitsunobu reactions. The product was
purified by column chromatography (15:1 hexanes–EtOAc): Rf
tryptone, and 10 mg/mL NaCl. Similarly, B. atrophaeus was streaked
onto a Nutrient Agar plate, which was made from a sterile aqueous
solution consisting of 15 mg/mL of agar and 8 mg/mL of BactoTM
Nutrient broth. All the bacterial culturing plates were sealed in a
plastic bag and incubated overnight at 37 °C.
0.35 (10:1 hexanes–EtOAc); IR:
m
2223 cmÀ1 (C„C), 2097 cmÀ1
(N@N@N); [
a
]
D
À69.8 (c 3.1, CH2Cl2); 1H NMR (500 MHz, CDCl3):
dH 8.30–8.33 (m, 1H, Ar), 7.52–7.58 (m, 3H, Ar), 7.38–7.48 (m,
4H, Ar), 7.31–7.36 (m, 1H, Ar), 7.24–7.27 (m, 2H, Ar), 7.03–7.07
(d, 2H, J = 8.5 Hz, Ar), 5.14 (br d, 1H, J1,2a = 2.9 Hz, H-1), 4.35 (s,
2H, OCH2C„C), 3.93 (dq, 1H, J5,6 = 6.5 Hz, J4,5 = 1.7 Hz, H-5), 3.42
(br s, 1H, H-4), 2.30 (s, 1H, PhCH3), 2.05–2.14 (m, 1H, H-3e),
1.86–1.97 (m, 2H, H-3a, H-2e), 1.50–1.56 (m, 1H, H-2a), 1.12 (d,
3H, J5,6 = 6.5 Hz, H-6); 13C NMR (125 MHz, CDCl3): dC 144.9 (Ar),
143.8 (Ar), 137.0 (Ar), 134.6 (Ar), 131.2 (2, Ar), 130.7 (Ar), 130.5
(Ar), 129.4 (2, Ar), 129.1 (Ar), 127.3 (2, Ar), 126.8 (2, Ar), 125.8
(Ar), 124.6 (Ar), 120.0 (Ar), 116.5 (Ar), 107.4 (Ar), 95.4 (C-1), 90.2
(„C), 77.9 („C), 65.6 (C-5), 59.9 (C-4), 54.8 (OCH2), 23.8 (C-2),
22.9 (C-3), 21.5 (PhCH3), 17.9 (C-6). ESIMS: m/z calcd for
[C30H28N4O4S]Na+: 563.1724. Found: 563.1723. Purity: 98.1%.
3.24. Paper disk diffusion assay
Colonies of B. atrophaeus and E. coli were inoculated into 4 mL of
sterile aq solution containing 8 mg/mL BactoTM Nutrient broth and
4 mL of sterile aq solution containing 5 mg/mL BactoTM yeast ex-
tract, 10 mg/mL BactoTM tryptone, and 10 mg/mL NaCl, respectively.
The bacterial solutions were incubated for 3–5 h at 35 °C without
shaking. Cultures were adjusted with the corresponding blank
media solution to an absorbance reading of 0.1 at 625 nm, which
means that the bacteria were at the 0.5 McFarland suspension, or
1 Â 108 CFU/mL. CFU stands for colony forming unit. The resulting
bacterial solutions were streaked on Mueller–Hinton agar (MHA)
plates, which were made from a sterile aqueous solution compris-
ing 17 mg/mL agar and 21 mg/mL DifcoTM Mueller–Hinton broth.
Afterward, paper disks were loaded with the required amount of
3.21. 3-(2-Phenylindol-3-yl)propan-3-onyl 2,3,6-trideoxy-a-L-
erythro-hexopyranoside (40)
compounds in 15 lL of the appropriate solvents and firmly placed
To
a
solution of N-tosyl-protected indole 37 (107 mg,
on the MHA plates with a thin film of bacteria. Next, the plates
were sealed in a plastic bag and incubated at 35 °C for ꢀ17 h,
and the diameters of the inhibition zones were recorded. Each
compound was run in duplicate, and the values shown in the table
reflect an average of this data.
0.210 mmol) in CH3OH (10 mL) was added KOH (60 mg, 1.1 mmol),
and the mixture was stirred at 40 °C for 5 h. After cooling to rt, the
reaction mixture was quenched by the addition of satd aqueous
NH4Cl (10 mL), and was then extracted with EtOAc. The combined
organic layers were washed with brine, dried (Na2SO4), filtered,
and evaporated under reduced pressure to afford the crude prod-
uct, which was purified by column chromatography to yield pure
40 (51 mg, 65%) as a yellow oil: Rf 0.33 (1:1 hexanes–EtOAc); IR:
3.25. 96-Well plate microdilution assay
Following the above procedure for the paper disk diffusion as-
m
3398 (O–H), 1730 (C@O) cmÀ1; [
a]
À50.2 (c 0.9, CH2Cl2); 1H
say,
a
B. atrophaeus solution that had
a
concentration of
D
NMR (500 MHz, CDCl3): dH 8.51 (br s, 1H, NH), 8.30–8.34 (m, 1H,
Ar), 7.58–7.62 (m, 2H, Ar), 7.49–7.53 (m, 3H, Ar), 7.38–7.42 (m,
1H, Ar), 7.28–7.33 (m, 2H, Ar), 4.62 (br d, 1H, J1,2a = 2.8 Hz, H-1),
3.90 (ddd, 1H, J = 9.8 Hz, J = 7.0 Hz, J = 6.0 Hz, OCH2), 3.90 (dt, 1H,
J = 9.8 Hz, J = 6.0 Hz, OCH2), 3.26 (dq, 1H, J4,5 = 9.1 Hz, J5,6 = 6.2 Hz,
H-5), 3.11 (br s, 1H, H-4), 2.76–2.86 (m, 2H, O@CCH2), 1.59–1.74
(m, 3H, H-3e, H-2a, H-2e), 1.41–1.52 (m, 1H, H-3a), 1.14 (d, 3H,
J5,6 = 6.2 Hz, H-6); 13C NMR (125 MHz, CDCl3): dC 196.2 (C@O),
143.6 (Ar), 135.2 (Ar), 132.5 (Ar), 129.7 (3, Ar), 128.9 (2, Ar),
127.5 (Ar), 123.7 (Ar), 122.6 (Ar), 122.3 (Ar), 115.6 (Ar), 110.9
(Ar), 96.1 (C-1), 72.0 (C-4), 69.3 (C-5), 62.8 (OCH2), 41.9 (O@CCH2),
29.6 (C-3), 27.5 (C-2), 17.9 (C-6). ESIMS: m/z calcd for [C23H25NO4]-
Na+: 402.1676. Found: 402.1676.
1 Â 106 CFU/mL was prepared. A 50-
lL aliquot of the above solu-
tion was seeded into each well, thereby giving a test concentration
of 5 Â 104 CFU/well. Thereafter, another 50
lL of the solution of a
test compound was added into each well. To prevent drying, the
96-well plate was then sealed in a plastic bag and incubated at
35 °C without shaking for about 20 h. Finally, the minimum inhibi-
tion concentrations, at which no appreciable growth of bacteria
was observed by the unaided eye, were recorded. Each sample
was run in duplicate, and the values shown in the table reflect
an average of this data.
Acknowledgements
This work was supported by the University of Alberta, the Nat-
ural Sciences and Engineering Research Council of Canada and the
Alberta Ingenuity Centre for Carbohydrate Science. W.S. is the reci-
pient of a Studentship from the Alberta Ingenuity Fund.
3.22. General methods—antibacterial assays
BactoTM yeast extract, BactoTM tryptone, DifcoTM Nutrient broth,
DifcoTM Meuller–Hinton broth, and DifcoTM granulated agar were
purchased from the Becton Dickinson Co. (BD). Penicillin, tetracy-
cline, and streptomycin were purchased from Sigma. Daunomycin
and doxorubicin were obtained from IFFECT ChemPhar (Hong
Kong) Company Ltd. Petri plates, blank 6-mm paper disks, Puritan
cotton tip applicators, applicator sticks, tissue culture treated poly-
styrene 96-well plates, and plastic inoculating loops were pur-
chased from Fisher Scientific Co. and came as sterile.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.carres.2009.10.010.
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