Synthesis of novel aminoglycosides via allylic azide rearrangement for investigating the significance of 2′-amino group

Article abstract of DOI:10.1016/j.bmc.2010.01.027

Using allylic azide rearrangement, a convenient method has been developed for the synthesis of 2′,3′-dideoxyaminoglycosides that are, otherwise, difficult to be prepared. The antibacterial activity of these novel aminoglycosides also confirms the indispensable role of 2′-NH₂ group for both neomycin and kanamycin classes of aminoglycosides. A novel structural motif containing the hexylaminocarbonyl groups at O-5 and/or O-6 of 2′,3′-dideoxyneamine could lead to the production of new aminoglycosides against resistant bacteria.

Full text of DOI:10.1016/j.bmc.2010.01.027

Bioorganic & Medicinal Chemistry 18 (2010) 1396–1405
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis of novel aminoglycosides via allylic azide rearrangement
for investigating the significance of 20-amino group
*
Jianjun Zhang, Anthony Litke, Katherine Keller, Ravi Rai, Cheng-Wei Tom Chang
Department of Chemistry and Biochemistry, Utah State University, 0300 Old Main Hill, Logan, UT 84322-0300, USA
a r t i c l e i n f o
a b s t r a c t
Using allylic azide rearrangement, a convenient method has been developed for the synthesis of 20,30-
dideoxyaminoglycosides that are, otherwise, difficult to be prepared. The antibacterial activity of these
novel aminoglycosides also confirms the indispensable role of 20-NH2 group for both neomycin and kana-
mycin classes of aminoglycosides. A novel structural motif containing the hexylaminocarbonyl groups at
O-5 and/or O-6 of 20,30-dideoxyneamine could lead to the production of new aminoglycosides against
resistant bacteria.
Article history:
Received 1 November 2009
Revised 11 January 2010
Accepted 12 January 2010
Available online 15 January 2010
Keywords:
Aminoglycoside
Antibiotic
Ó 2010 Elsevier Ltd. All rights reserved.
Drug resistant bacteria
1. Introduction
For instance, neamine derivative, 1 with 20-NH2 replaced with 20-
OH shows much lower antibacterial activity than the parent nea-
The neamine core contributes predominately to the antibacte-
rial activity of neomycin and kanamycin classes of aminoglyco-
sides (Fig. 1).1 Neamine has four amino groups located at 1, 3, 20,
and 60 positions. The presence of 1-NH2 and 3-NH2 are essential
for the antibacterial activity of aminoglycosides that contain nea-
mine. The 60-NH2 amino group can be replaced with OH without
obstructing the activity as in the case of paromomycin.1c However,
the roles of 20-NH2 amino group are multiplex and contradictory,
especially, in association with the deoxygenation at 30 position.
Modifications such as deoxygenation at 30 and 40 positions has
been shown to be effective in reviving the activity of aminoglyco-
side constructs against aminoglycoside resistant bacteria that
equipped with aminoglycoside modifying enzymes (AMEs) that
introduce modifications at these two positions. For example, tobra-
mycin that has deoxygenation at 30 position is active against bacte-
ria equipped with aminoglycoside phosphotransferases that
catalyze phosphorylation at 30-OH position (APH(30)).1a,b On the
other hand, aminoglycosides with deoxygenation at 30 position
also has been reported to exert higher cytotoxicity due to the in-
creased basicity of 20-NH2.2 It has been reported that an intra-
molecular hydrogen bond between 20-NH2 and 400-oxygen atom
on ring III of neomycin is crucial in orienting the conformation of
neamine and, thus, optimizing the binding to the targeted rRNA
site.3 In addition, there is no significant difference in the antibacte-
rial activity between kanamycin B and kanamycin A that have 20-
NH2 and 20-OH, respectively. Even the antibacterial results from
the modified neamine derivatives were, in part, contradictory.
mine.4 Interestingly, the neamine derivative, 2 with deamination
at 20 position manifests improved antibacterial activity.5
Judging from the results in the literature, it is unclear regarding
the significance of 20-NH2 group. To this aim, we wish to explore
the synthesis of aminoglycosides with deamination of the 20-NH2
group. Chemical deamination usually involves multiple steps and
has not been noted to be accompanied with deoxygenation at 30
position. Masking amino groups as azido groups (azido approach)
is one of the commonly employed strategies for conducting struc-
tural modifications of aminoglycosides.1c In an effort to revive the
activity of aminoglycosides, we have developed a mild method for
introducing 30,40-dideoxygenation on aminoglycosides.6 The pro-
cess involves the formation of a double bond between 30 and 40
positions. With the presence of 20-N3 group, it is possible that an
allylic azide rearrangement should offer a convenient access to
deamination and deoxygenation at 20 and 30 positions, respectively.
In addition, the 40-OH will be replaced with 40-NH2, which should
maintain the interactions involved in the parent neamine.4 Ad-
ducts from allylic azide rearrangement could offer the information
on the significance of having 20-NH2 group. Although allyl azide
rearrangement has been reported,7 such rearrangement has not
been noted on azidoaminoglycosides.
2. Results and discussion
2.1. Initial discovery of the allylic azide rearrangement
The synthesis of azidoneamine core, 3 with a double bond
* Corresponding author. Tel.: +1 435 797 3545; fax: +1 435 797 3390.
E-mail address: tom.chang@usu.edu (C.-W.T. Chang).
between 30 and 40 positions was conducted according to our
0968-0896/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2010.01.027

J. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1396–1405
1397
protocol.6 The discovery of allylic azide rearrangement on 3 was
N3
NH2
N3
O
actually an unexpected result (Scheme 1).8 The initial attempts
were designed to explore the antibacterial activity of non-carbohy-
drate moieties at O-6 and/or O-5 positions. The added functional
groups were attached via carbamate linkage. When compound 3
reacted with N-hexylisocyanate generated in situ, the alkylcarba-
mate was incorporated at O-6 as expected. However, when excess
N-hexylisocyanate was used, we isolated two products, 5 and 6
with mono- and di-alkylcarbamates attached, respectively. Never-
theless, compound 5 has different 1H NMR from 4. Upon further
investigation, we confirmed that the double bond of both 5 and 6
were rearranged to 20 and 30 positions via an allylic azide
rearrangement.
4'
O
(2 eq.)
N3
N3
N3
3'
N3
O
N3
triphosgene (1.5 eq.)
Et3N (8 eq.), CH2Cl2
-78oC to r.t.
O
HO
N3
HO
O
6
4
3
5
OH
O
C6H13
34%
N
H
NH2
N3
O
(4 eq.)
N3
N3
triphosgene (3 eq.)
O
N3
HO
Et3N (16 eq.), CH2Cl2
-78 ºC to r.t.
OH
3
N3
Although we did not know the details regarding why such a
rearrangement occurred in these conditions, we envisioned that
this allylic azide rearrangement can be triggered with no added re-
agents and conveniently lead to the production of neamine deriv-
ative with deamination and deoxygenation at 20 and 30 positions,
respectively. More importantly, the aminoglycosides resulted from
this allylic azide migration will provide direct evaluation on the
significance of 20-NH2 group.
N3
O
N3
O
N3
N3
O
N3
O
N3
+
O
O
N3
HO
O
HN
C6H13
O
O
C6H13
O
C6H13
N
5
53%
6
39%
N
H
H
Scheme 1.
2.2. Allylic azide rearrangement of azidoneamine
sized. In our previous work, we have noticed that the presence of
600-CH3 is not essential for the antibacterial activity.10 Therefore,
due to the consideration of availability of glycosyl donor, com-
pound 18 was synthesized for comparing the activity of aminogly-
cosides from 16. The glycosyl donor, 17 can be prepared using
similar procedures as reported (Scheme 4).9
Glycosylation of 8 using 22 yield the desired pyranmycin (a neo-
mycin class aminoglycoside) derivative, 23 (Scheme 5). Based on our
previous structure-activity relationship (SAR) studies of pyranmycin,
compound 22 has the carbohydrate moiety that generates the most
active aminoglycoside.10 The corresponding 30,40-dideoxy pyranmy-
cin derivatives has also been prepared previously. During the synthe-
ses, we did notice that the rearrangement adducts with deamination
at 20 and double bond between 20 and 30 positions are very sensitive
to acidic conditions. Therefore, all the processes were conducted
with minimum exposure to acids or at lower temperature.
The allylic azide rearrangement is expected to proceed under
thermodynamic condition similarly to the [3,3]-sigmatropic rear-
rangement.7 To our please, the allylic azide rearrangement using
compound 3 can be carried out at 110 °C in toluene (Scheme 2). Since
such rearrangement can be reversible, we obtained a mixture of 3
and 7, which can be separated by column chromatography. Com-
pound 7 was acetylated to yield 9 in order to make the proton signals
discernable. The structure of the neamine derivative, 9 from allylic
azide rearrangement was confirmed by 2D NMR (COSY and HSQC).
Both 3 and 7 were subjectedto Staudingerreduction and hydrogena-
tion to yield the neamine derivatives, 10 and 11, respectively. Direct
glycosylation of 7 can provide the syntheses of 20,30-dideoxykana-
mycin classes of aminoglycosides as reported previously.1c Regiose-
lective benzoylation of 7 offered compound 8, which can be
employed for the syntheses of 20,30-dideoxyneomycin classes of
aminoglycosides also as reported previously.
All the azidoaminoglycosides were subjected to similar global
deprotection (Staudinger reduction followed by hydrogenation
and ion-exchange column chromatography) to yield the designed
aminoglycosides ready for assay (Scheme 6).
2.3. Synthesis of novel aminoglycosides
Glycosylation of 7 using phenylthioglycoside, 12 offered the
precursors of 20,30-dideoxykanamycin derivatives, 13 and epi-13
as the minor product (Scheme 3). In order to compare the activity,
the corresponding 30,40-dideoxykanamycin derivatives, 14 was also
prepared. Based on our previous structure–activity relationship
(SAR) studies of kanamycin B analogs,9 compound 16 was synthe-
2.4. Antibacterial activity of aminoglycosides
The synthesized compounds were assayed against aminoglyco-
side susceptible bacteria Escherichia coli (ATCC 25922, G-) Staphylo-
coccus aureus (ATCC 25923, G+) and using neamine, neomycin and
NH2
R1
4'
I
O
I
O
R3
R2
NH2
O
4'
I
4'
1'
1'
HO
II
NH2
R2
HO
HO
HO
3'
4
4
R1
H2N
O
NH2
NH2
1
3'
1'
NH2
O
O
II
1 NH2
3'
R1
4
HO
O
III
6
5
O
1 NH2
O
1"
HO
4"
HO
6
5"
II
OH
OH
OH
O
HO
O
OH
R3
R1
NH2
OH
H
H2N
R1
III
Neamine
1
2
R1
R2 R3
R2
OH
R3
OH
OH
H
Neomycin B
Paromomycin I OH
Lividomycin B
Ribostamycin
NH2 OH DAG
Kamanycin A OH
Kamanycin B NH2 OH
OH DAG
OH
NH2
H
OH
DAG
H
Dibekacin
NH2
NH2
H
H
Tobramycin
OH
DAG: 2,6-diamino-2,6-dideoxy−β-L-glucopyranosyl
Kanamycin class
Neomycin class
Figure 1. Structures of aminoglycosides.

1398
J. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1396–1405
1) PMe3, THF
NaHCO3, 50ºC
N3
N3
NH2
O
2) H2, Pd/C
toluene
110ºC
O
O
N3
H2N
NaHCO3, H2O
N3
N3
H2N
N3
51%
N3
O
O
O
N3
NH2
HO
HO
HO
11
OH
OH
OH
7
3
47%
Ac2O, Et3N
1) PMe3, THF
BzCl, DIPEA
CH2Cl2
DMAP, CH2Cl2
NaHCO3, 50ºC
2) H2, Pd/C
NaHCO3, H2O
r.t.
71%
71%
N3
-50ºC to r.t.
82%
N3
O
N3
NH2
O
N3
N3
O
N3
O
H2N
N3
OAc
AcO
O
H2N
N3
OBz
HO
O
NH2
9
HO
8
OH
10
Scheme 2.
BnO
BnO
OBn
O
SPh
N3
O
N3
BnO
12
N3
NIS, TfOH, 4 A MS
Et2O/CH2Cl2 = 3/1
O
N3
HO
OH
7
N3
O
N3
O
N3
N3
N3
N3
O
+
N3
O
O
HO
N3
OBn
HO
O
BnO
O
OBn
74%
13
O
epi-13 4%
BnO
BnO
OBn
BnO
OBn
N3
O
BnO
BnO
OBn
O
N3
N3
SPh
N3
O
O
N3
O
BnO
12
N3
HO
N3
O
O
N3
NIS, TfOH, 4 A MS
Et2O/CH2Cl2 = 3/1
OBn
HO
14
OH
3
BnO
90%
BnO
OBn
N3
N3
H3C
O
N3
O
O
N3
BzO
SPh
N3
N3
15
N3
BnO
O
NaOMe, MeOH
N3
O
O
HO
N3
HO
O
NIS, TfOH, 4 A MS
Et2O/CH2Cl2 = 3/1
OH
7
16
CH3
OH
78% (2 steps) BnO
N3
N3
O
N3
O
O
BzO
N3
SPh
N3
N3
N3
N3
O
BnO
17
NaOMe, MeOH
N3
O
HO
N3
HO
O
NIS, TfOH, 4 A MS
Et2O/CH2Cl2 = 3/1
OH
18
3
O
72% (2 steps)
BnO
OH
N3
Scheme 3.
kanamycin as the controls (Table 1). From the minimum inhibitory
concentration (MIC) results, the 20-NH2 group appears to have con-
tradictory roles. The 20-NH2 group is important for the antibacterial
activity of both neomycin and kanamycin classes of aminoglyco-
sides since all the derivatives with 20,30-dideoxyaminoglycosides
are less active than the corresponding 30,40-dideoxyaminoglycosides
with the tendency of neamine > 30,40-dideoxyneamine > 20,30-dide-
oxyneamine. However, such tendency is less obvious or even re-
versed with the attachment of hexylaminocarbonyl group. For
example, both NRR101 and NRR102 are slightly more active than
RR101 (entries 4–6). Further testing also reveals that NRR102 man-
ifests MIC0s of 32–64 and 125
lg/mL against E. coli equipped with
APH30-I and AAC60/APH2”, respectively. In contrast, neamine, neo-
mycin and kanamycin are inactive against these bacteria (MIC

J. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1396–1405
1399
HO
HO
HO
HO
1) Tf2O, pyridine
2) nBu4N+OAc-,CH2Cl2
O
O
O
SPh
AcO
SPh
OH
58%
BnO
HO
BnO
AcO
SPh
D-arabinose
19
20
1) NaOMe, MeOH
2) BzCl, DIPEA
DMAP, CH2Cl2, -50ºC
1) Tf2O, pyridine
2) NaN3, DMF
O
O
BzO
N3
SPh
BzO
85%
BnO
BnO
17
48%
HO
21
Scheme 4.
1) cat. BF3-OEt2, CH2Cl2
method for the preparation of C-4 allylic azides that has been
employed in the syntheses of several biologically interesting
compounds.14
N3
H3C
O
O
N3
N3
N3
AcO
O
N3
N3
AcO
The antibacterial results confirm that 20-NH2 group is indis-
pensable for the activity of neomycin and kanamycin classes of
aminoglycosides, which also suggest that it may be difficult to
compromise the contradictory roles of 20-NH2 group for these
two classes of aminoglycosides. However, our results indicate that
having hexylaminocarbonyl groups at O-5 and/or O-6 of neamine
along with the 20,30-dideoxyneamine core may serve as an alterna-
tive structural motif for generating novel aminoglycosides against
resistant bacteria. Further SAR investigation on similar functional
groups is being carried out.
O
O
N3
22
OC(NH)CCl3
N3
O
H3C
O
OH
N3
OBz
2) NaOMe, MeOH
56%
HO
N3
HO
8
HO
23
Scheme 5.
>250 l
g/mL). These results imply that 20,30-dideoxyneamine (no 20-
NH2 group) combined with hexylaminocarbonyl group at O-5 or O-6
positions could be a novel motif for generating new aminoglycosides
against resistant bacteria.
To explore the unexpected effect of hexylaminocarbonyl group,
we decided to synthesize two addition neamine analogs, RR103
and RR104, with mono- and di-hexylaminocarbonyl groups at O-
6 and O-5/O-6 positions, respectively while keeping the scaffold
of neamine (Scheme 7). The syntheses can be achieved via similar
processes using compound 2410 as the starting material. A panel of
aminoglycoside susceptible and resistant bacteria, including
Klebsiella pneumoniae (ATCC 13883, susceptible, G-), K. pneumoniae
(ATCC 700603, resistant, G-), Pseudomonas aeruginosa (ATCC
27853, resistant, G-), S. aureus (ATCC 33591, resistant, G+), were
selected for evaluating the antibacterial activity.
From the result of antibacterial assay, it is obvious that NRR102
is still the most active compound while other neamine derivatives
with hexylaminocarbonyl group(s) are less active (Table 2). We
have recently reported that incorporation of linear alkyl chains
could alter the tradition antibacterial mode of aminoglycosides
leading to the generation of novel antibiotics.11 RR103 and
RR104 are, however, much less active against P. aeruginosa (ATCC
27853) and S. aureus (ATCC 33591) that equipped with various
APH(30) (entries 5 and 6).12,13 This result implies that these two
analogs are still being inactivated by AME0s. Combining the anti-
bacterial activity of these neamine analogs, we believe these ana-
logs should exert traditional mode of antibacterial action.
Therefore, the antibacterial activity of NRR102 against bacteria
equipped with AME0s should be attributed to the deoxygenation
of 30-OH. Nevertheless, in this new neamine motif, the presence
of 20-NH2 group is no longer essential. Although the activity is less
than what is needed to be clinically significant, it is possible that
enhanced activity could be obtained by varying the length of the
linear alkylamino chain.
4. Experimental
4.1. 6-O-Hexylaminocarbonyl-1,3,20,60-tetraazido-30,40-dideoxy-
30-enoneamine (4)
To a solution of 1-heptamine (0.10 g, 1.0 mmol) and Et3N
(0.60 mL, 4.2 mmol) in anhydrous CH2Cl2 (5 mL) at ꢀ78 °C, triphos-
gene (0.22 g, 0.75 mmol) was added, and the reaction was stirred for
several hours allowing the temperatureto warm up to room temper-
ature. The reaction mixture was cooled to 0 °C and added with com-
pound 3 (0.20 g, 0.51 mmol). After being stirred overnight, the
reaction was quenched with water and diluted with EtOAc. The com-
bined organic solution was washed with 1 M HCl, water, saturated
NaHCO3, and brine, and then dried over anhydrous Na2SO4. After
the removal of the solvents and purification with column chroma-
tography (eluted with hexane/EtOAc from 100/0 to 60/40), the prod-
uct was obtained as an oil (0.08 g, 0.15 mmol, 34%) along with the
recovered compound 3 (0.09 g). 1H NMR (CDCl3, 400 MHz) d 5.93
(d, J = 10.5 Hz, 1H), 5.89 (d, J = 10.4 Hz, 1H), 5.74 (d, J = 3.9 Hz, 1H),
4.85 (t, J = 6.2 Hz, 1H), 4.7 (m, 1H), 4.67 (t, J = 9.9 Hz, 1H), 3.9
(m, 1H), 3.8 (m, 1H), 3.68 (t, J = 9.1 Hz, 1H), 3.58 (t, J = 9.4 Hz, 1H),
3.4–3.5 (m, 2H), 3.3–3.4 (m, 2H), 3.2 (m, 2H), 2.27 (td, J = 13.3,
4.5 Hz, 1H), 1.6 (m, 1H), 1.5 (m, 2H), 1.2 (m, 6H), 0.85 (t, J = 6.7 Hz,
3H); 13C NMR (CDCl3, 100 MHz) d 156.5, 129.6, 123.1, 97.4, 81.4,
75.9, 69.0, 59.2, 58.4, 55.4, 54.0, 41.7, 32.5, 32.0, 31.6, 29.9, 26.5,
22.7, 14.2; ESI/APCI Calcd for C19H30N13O5 ([M+H]+) m/e 520.2487;
measured m/e 520.2476.
þ
4.2. 6-O-Hexylaminocarbonyl-1,3,40,60-tetraazido-20,30,40-
trideoxy-20-enoneamine (5)
3. Conclusion
Compound 5 was prepared similarly to the synthesis of com-
pound 4 except the following amount of reagents were used:
1-heptamine (0.21 g, 2.0 mmol), Et3N (1.2 mL, 8.6 mmol) and
triphosgene (0.45 g, 1.53 mmol) for reacting with compound 3
(0.20 g, 0.51 mmol). Two products were isolated after column
chromatography (eluted with hexane/EtOAc from 100/0 to
60/40): compound 5 (0.14 g, 0.27 mmol, 53%) and compound 6
We have developed a convenient method for synthesizing
20,30-dideoxyneomycin and 20,30-dideoxykanamycin derivatives. The
reported allylic azide rearrangement can also be applicable of mak-
ing aminosugar derivatives. For example, this rearrangement can
serve as an alternative method to the reported Pd-catalyzed p-allyl

1400
J. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1396–1405
1) for 3',4'-dideoxyazidoaminoglycosides:
PMe3, THF, 50ºC
NH2
O
N3
O
for 2',3'-dideoxyazidoaminoglycosides:
H2N
N3
PMe3, THF, NaHCO3, 30ºC
H2N
N3
2) H2, Pd/C, MeOH, cat. HOAc/H2O (1/4)
O
O
NH2
N3
3) CG-50 (NH4)
O
O
R1
R1
O
O
R2
R2
Reactant
Product
Yield (%)
50%
NH2
O
H2N
H2N
O
4
NH2
HO
O
RR101
C6H13
O
N
H
NH2
O
H2N
H2N
H2N
O
NH2
5
6
53%
89%
83%
HO
O
NRR101
C6H13
O
N
H
NH2
O
H2N
O
NH2
O
O
HN
C6H13
O
O
C6H13
N
NRR102
H
NH2
O
H2N
H2N
O
NH2
13
HO
O
OH
NRT014
O
HO
HO
OH
NH2
O
H2N
H2N
O
NH2
HO
14
16
79%
72%
O
OH
RT014
O
HO
HO
OH
NH2
O
H2N
H2N
O
NH2
O
HO
O
NRT040
CH3
OH
HO
NH2
NH2
O
H2N
H2N
O
18
59%
NH2
O
HO
RT040
O
HO
OH
NH2
NH2
O
H2N
H2N
23
55%
O
NH2
OH
O
O
H3C
H2N
HO
HO
NRR501
NH2
O
H2N
O
H2N
O
-
Ref. 6
NH2
OH
O
H3C
H2N
HO
HO
RR501
Scheme 6.
(0.13 g, 0.20 mmol, 39%). 1H NMR (CDCl3, 400 MHz) d 5.9 (m, 2H),
5.38 (d, J = 4.0 Hz, 1H), 4.93 (t, J = 5.4 Hz, 1H), 4.8 (m, 2H), 3.8 (m,
1H), 3.6–3.7 (m, 2H), 3.4–3.5 (m, 3H), 3.2–3.3 (m, 3H), 2.25 (td,
J = 8.1, 3.9 Hz, 1H), 1.4–1.6 (m, 4H), 1.2–1.3 (m, 6H), 0.86 (t,
J = 6.6 Hz, 3H); 13C NMR (CDCl3, 100 MHz) d 157.1, 129.6, 123.3,
98.1, 78.7, 78.2, 76.5, 69.2, 60.8, 59.6, 54.9, 54.2, 41.8, 32.2, 32.0,
4.3. 5,6-Di-O-hexylaminocarbonyl-1,3,40,60-tetraazido-20,30,40-
trideoxy-20-enoneamine (6)
Please refer to the procedure for the preparation of compound
5. 1H NMR (CDCl3, 400 MHz) d 5.9 (m, 2H), 5.35 (d, J = 4.5 Hz,
1H), 4.99 (t, J = 9.7 Hz, 1H), 4.8–4.9 (m, 3H), 3.70 (t, J = 9.6 Hz,
1H), 3.4–3.6 (m, 5H), 3.25 (dd, J = 13.0, 5.3 Hz, 1H), 3.1–3.2 (m,
4H), 2.3 (m, 1H), 1.6 (m, 5H), 1.2 (m, 12H), 0.85 (t, J = 4.3 Hz,
29.8, 26.6, 22.8, 14.2; ESI/APCI Calcd for C19H30N13O5 ([M+H]+)
þ
m/e 520.2487; measured m/e 520.2481.

J. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1396–1405
1401
Table 1
MIC of aminoglycosides (
tion with column chromatography (eluted with hexane/EtOAc
from 90/10 to 50/50), the product was obtained as an oil (0.14 g,
0.36 mmol, 47%) along with the recovered compound 3 (0.12 g,
0.31 mmol, 40%). 1H NMR (CDCl3, 400 MHz) d 6.01 (d, J = 10.2 Hz,
1H), 5.99 (dd, J = 12.0, 1.8 Hz, 1H), 5.57 (s, 1H), 4.0 (m, 1H), 3.91
(d, J = 9.8 Hz, 1H), 3.4–3.6 (m, 4H), 3.3–3.4 (m, 3H), 2.71 (s, 1H),
2.28 (td, J = 13.4, 4.3 Hz, 1H), 1.4–1.6 (m, 2H); 13C NMR (CDCl3,
100 MHz) d 128.5, 128.2, 94.7, 79.8, 76.3, 76.2, 69.7, 60.5, 59.6,
l
g/mL)
E. coli (ATCC 25922)
Entry
Compound
S. aureus (ATCC 25923)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
Neamine
10
11
RR101
NRR101
NRR102
Neomycin
RR501
NRR501
32
4–8
32–64
P250
P250
P250
125
16–32
64–125
125
32–64
32
54.8, 51.8, 32.4; ESI/APCI Calcd for C12H17N12O4 ([M+H]+) m/e
þ
4
16
1
4–8
393.1490; measured m/e 393.1502.
125–250
4
64–125
P250
4–8
125–250
2
125–250
P250
2–4
Kanamycin
RT014
4.5. 6-O-Benzoyl-1,3,40,60-tetraazido-20,30,40-trideoxy-20-enone-
amine (8)
NRT014
RT040
NRT040
16–32
8–16
To a solution of compound 7 (0.10 g, 0.26 mmol) and DIPEA
(0.05 mL, 0.31 mmol) in anhydrous CH2Cl2 (10 mL) at ꢀ50 °C, BzCl
(0.03 mL, 0.26 mmol) was added. After being stirred for 2 h, NaH-
CO3(s) was added and the solvent was removed. The residue was di-
luted with EtOAc and the organic solution was washed with
saturated NaHCO3(aq), water and brine, and then dried over anhy-
drousNa2SO4. Aftertheremovalof the solventsand purificationwith
column chromatography (eluted with hexane/EtOAc from 100/0 to
70/30), the product was obtained as an oil (0.09 g, 0.18 mmol,
71%). 1H NMR (CDCl3, 400 MHz) d 8.07 (dd, J = 8.0, 1.5 Hz, 2H), 7.61
(td, J = 7.2, 1.2 Hz, 1H), 7.48 (td, J = 7.7, 1.5 Hz, 2H), 6.0 (m, 2H),
5.64 (s, 1H), 5.07 (td, J = 9.6, 3.1 Hz, 1H), 4.0 (m, 1H), 3.9 (m, 1H),
3.5–3.7 (m, 7H), 2.39 (dt, J = 13.4, 4.5 Hz, 1H), 1.63 (dt, J = 12.6,
11.5 Hz, 1H); 13C NMR (CDCl3, 100 MHz) d 167.2, 134.1, 130.2 (2 car-
bons), 128.9, 128.8 (2 carbons), 128.4, 128.2, 95.0, 80.5, 76.9, 75.9,
NH2
N3
O
BnO
BnO
(4 eq.)
N3
N3
O
triphosgene (3.0 eq.)
Et3N (17 eq.), CH2Cl2
-78ºC to r.t.
N3
HO
24
OH
N3
N3
O
O
BnO
BnO
BnO
BnO
+
N3
N3
N3
N3
O
O
N3
N3
HO
O
O
O
O
HN
C6H13
25 38%
26 32%
þ
O
C6H13
69.8, 59.4, 58.6, 54.8, 51.7, 32.7; ESI/APCI Calcd for C19H21N12O5
O
C6H13
N
H
N
H
([M+H]+) m/e 497.1758 ; measured m/e 497.1575.
1) PMe3, THF
1) PMe3, THF
NaHCO3, 50ºC
2) H2, Pd/C
4.6. 5,6-Di-O-acetyl-1,3,40,60-tetraazido-20,30,40-trideoxy-20-enone-
amine (9)
75%
NaHCO3, 50ºC
74%
2) H2, Pd/C
NaHCO3, H2O
NaHCO3, H2O
NH2
O
NH2
O
HO
HO
HO
HO
To a solution of compound 7 (0.02 g, 0.051 mmol), Et3N
H2N
H2N
H2N
(0.06 mL, 0.41 mmol) and DMAP (catalytic amount) in anhydrous
CH2Cl2 (10 mL) at 0 °C, Ac2O (0.02 mL, 0.26 mmol) was added. After
being stirred for 2 h, the solvent was removed. After purification
with column chromatography (eluted with hexane/EtOAc from
100/0 to 50/50), the product was obtained as an oil (0.02 g,
0.042 mmol, 82%). 1H NMR (CDCl3, 400 MHz) d 6.02 (d, J = 10.2 Hz,
1H), 5.72 (dt, J = 10.1, 2.1 Hz, 1H), 5.19 (s, 1H), 5.05 (t, J = 9.8 Hz,
1H), 4.92 (t, J = 10.0 Hz, 1H), 3.9 (m, 2H), 3.72 (t, J = 9.7 Hz, 1H),
3.4–3.6 (m, 4H), 2.35 (dt, J = 9.1, 4.6, 1H), 2.06 (s, 3H), 2.04
(s, 3H), 1.5 (m, 1H); 13C NMR (CDCl3, 100 MHz) d 170.0, 169.7,
129.2, 127.4, 94.9, 78.4, 74.3, 74.0, 69.8, 59.5, 58.2, 54.5, 51.5,
32.4, 20.9, 20.8; ESI/APCI Calcd for C16H20N12O6Na+ ([M+Na]+)
m/e 499.1521; measured m/e 499.1519.
H2N
O
NH2
O
NH2
HO
O
O
O
O
HN
O
C6H13
C6H13
RR104
O
C6H13
N
N
H
RR103
H
Scheme 7.
Table 2
MIC of aminoglycosides (
l
g/mL)
Entry Compound K. pneumoniae K. pneumoniae P. aeruginosa S. aureus
(ATCC 13883) (ATCC 700603) (ATCC 27853) (ATCC 33591)
1
2
3
4
5
6
Neamine 2–4
16–32
125
P250
64–125
16–32
16–32
16–32
64–125
125–250
125–250
64–125
64–125
16–32
P250
125–250
RR101
NRR101
NRR102
RR103
RR104
16–32
16–32
4–8
16–32
32
4.7. 30,40-Dideoxyneamine (10)
P250
125–250
125–250
This compound was prepared similarly to the synthesis of com-
pound 11. 1H NMR (D2O, 400 MHz) d 5.66 (t, J = 3.4 Hz, 1H), 4.1 (m,
1H), 3.86 (t, J = 9.4 Hz, 1H), 3.56 (t, J = 9.2 Hz, 1H), 3.4–3.5 (m, 3H),
3.26 (dd, J = 12.5, 4.3 Hz, 1H), 3.22 (dd, J = 10.3, 4.2 Hz, 1H), 3.14
(dd, J = 13.5, 3.4 Hz, 1H), 2.98 (dd, J = 13.5, 7.2 Hz, 1H), 2.41 (td,
J = 8.2, 4.1 Hz, 1H), 1.8–1.9 (m, 4H); 13C NMR (D2O, 75 MHz) d
95.2, 77.2, 75.3, 72.6, 65.8, 49.8, 48.8, 48.7, 42.6, 28.3, 25.6, 20.5;
ESI/APCI Calcd for C12H27N4O4Na+ ([M+Na]+) m/e 313.1846; mea-
sure m/e 313.1853.
6H); 13C NMR (CDCl3, 100 MHz) d 155.3, 155.0, 129.5, 123.4, 97.6,
75.2, 74.9, 69.1 (2 carbons), 59.6, 58.2, 54.7, 54.1, 41.6, 41.5, 32.4,
31.7 (2 carbons), 29.99, 29.94, 26.6, 26.5, 22.8 (2 carbons), 14.2
(2 carbons); ESI/APCI Calcd for C26H43N14O6 ([M+H]+) m/e
þ
647.3485; measured m/e 647.3483.
4.4. 1,3,40,60-Tetraazido-20,30,40-trideoxy-20-enoneamine (7)
4.8. 40-Amino-20,30,40-trideoxyneamine (11)
A solution of compound 3 (0.30 g, 0.77 mmol) in toluene was
To a solution of compound 7 (0.21 g, 0.54 mmol), NaHCO3(s)
refluxed overnight. After the removal of the solvents and purifica-
(catalytic amount), several drops of water in THF (15 mL) at

1402
J. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1396–1405
50 °C, PMe3 (2.0 mL, 1 M in THF) was added. After being stirred for
30 min., the solvent was removed. The residue was added with de-
gassed water, Pd/C (catalytic amount) and NaHCO3(s) (catalytic
amount), and then charged with atmospheric H2. After being stir-
red for two days, the reaction was quenched by filtering through
Celite and the residue was washed with H2O and the combined
solutions were concentrated. The crude product was purified with
Amberlite CG50 (NH4þ) eluted with a gradient of NH4OH solution
(0–20%). After collection of the desired fractions and removal of
solvent, the product was re-dissolved in water and loaded to an
ion-exchange column packed with Dowex 1X8-200 (Clꢀ form),
and eluted with water. After removal of solvent, the product was
obtained as white solid (0.08 g, 0.27 mmol, 51%). 1H NMR (D2O,
400 MHz) d 5.4 (m, 1H), 4.2 (m, 1H), 3.81 (t, J = 9.1 Hz, 1H), 3.2–
3.5 (m, 7H), 2.38 (td, J = 8.8, 4.5 Hz, 1H), 1.8–2.9 (m, 5H); 13C
NMR (D2O, 100 MHz) d 96.4, 77.8, 74.9, 72.6, 69.6, 50.0, 48.9,
47.1, 39.4, 28.3, 26.1, 21.1; ESI/APCI Calcd for C12H27N4O4Na+
([M+Na]+) m/e 313.1846; measured m/e 313.1853.
128.0 (2 carbons), 127.9, 127.8 (2 carbons), 127.7 (2 carbons),
127.4 (2 carbons), 99.4, 95.0, 87.2, 80.1, 78.3, 76.7, 76.0, 75.3,
74.7, 73.7 (2 carbons), 71.3, 70.0, 69.5, 59.8, 59.6, 54.7, 51.7, 32.8,
14.3; ESI/APCI Calcd for C46H50N12O9Na+ ([M+Na]+) m/e 937.3721;
measured m/e 937.3699.
4.11. 6-O-(1,2,3,6-Tetra-O-benzyl-a-D
-galactopyranosyl)-1,3,4’,60-
tetraazido-30,40-dideoxy-30-enoneamine (14)
This compound was prepared similarly to the synthesis of com-
pound 13. 1H NMR (CDCl3, 300 MHz) d 7.2–7.4 (m, 20H), 5.9 (m,
2H), 5.08 (d, J = 3.4 Hz, 1H), 4.94 (t, J = 11.7 Hz, 1H), 4.7–4.9 (m,
5H), 4.4–4.6 (m, 4H), 4.1 (m, 1H), 4.10 (d, J = 3.8 Hz, 1H), 3.95
(dd, J = 8.3, 2.8 Hz, 1H), 3.8 (m, 1H), 3.7 (m, 2H), 3.2–3.5 (m, 8H),
2.27 (td, J = 13.4, 4.5 Hz, 1H), 1.6 (s, 1H), 1.46 (td, J = 12.9,
12.7 Hz, 1H); 13C NMR (CDCl3, 100 MHz) d 139.0, 138.7, 138.5,
137.9, 129.5, 128.7 (2 carbons), 128.6 (2 carbons), 128.6 (4 car-
bons), 128.5 (2 carbons), 128.4 (2 carbons), 128.1 (2 carbons),
128.1, 128.0, 127.9, 127.8 (2 carbons), 127.7 ,123.7, 99.6, 96.9,
87.2, 79.1, 78.5, 76.7, 76.4, 75.3, 74.8, 73.8 (2 carbons), 73.7, 71.2,
69.4, 69.0, 59.9, 59.8, 54.9, 54.0, 32.9; ESI/APCI Calcd for
C46H50N12O9+Na+ ([M+Na]+) m/e 937.3721; measured m/e
937.3704.
4.9. 6-O-(1,2,3,6-Tetra-O-benzyl-a-D
-galactopyranosyl)-1,3,40,60-
tetraazido-20,30,40-trideoxy-20-enoneamine (13)
A solution of compound 7 (0.20 g, 0.57 mmol), compound 12
(0.39 g, 0.61 mmol), (1.2 equiv), and activated powder 4 Å molecu-
lar sieve was stirred in anhydrous Et2O and CH2Cl2 (Et2O: 4.5 mL;
CH2Cl2: 1.5 mL) at room temperature overnight. The reaction mix-
ture was cooled to ꢀ70 °C and N-iodosuccinimide (0.14 g,
0.61 mmol) was quickly added into the reaction mixture. After
the temperature of the solution warmed up to ꢀ30 °C, trifluoro-
methanesulfonic acid (catalytic amount) was added. The solution
was stirred at low temperature till the complete consumption
of the glycosyl donor (ca. 4 h, monitored by TLC, eluted with
Hexane/EtOAc = 65/35). The reaction mixture was quenched by the
addition of Et3N (3 mL). After being stirred for 10 min, the reaction
mixture was filtered through Celite and the residue was washed
with EtOAc. The combined organic solution washed with 10%
aqueous Na2S2O3, saturated NaHCO3(aq) and brine, and dried over
anhydrous Na2SO4. After removal of the solvents and purification
with column chromatography (eluted with hexane/EtOAc from
100/0 to 65/35), the product was obtained as an oil (0.34 g,
0.38 mmol, 74%). The other product, epi-13 was obtained as a min-
or product (0.02 g, 0.022 mmol, 4%). 1H NMR (CDCl3, 300 MHz) d
7.3 (m, 20H), 5.90 (d, J = 10.0 Hz, 1H), 5.70 (d, J = 12.4 Hz, 1H),
5.49 (s, 1H), 4.7–5.0 (m, 7H), 4.57 (d, J = 11.7 Hz, 1H), 4.41 (t,
J = 11.3 Hz, 2H), 3.8–4.1 (m, 6H), 3.2–3.6 (m, 9H), 2.3 (td, J = 8.6,
3.8 Hz, 1H), 1.4 (m, 1H); 13C NMR (CDCl3, 75 MHz) d 138.9, 138.5,
138.3, 137.7, 128.8 (2 carbons), 128.7 (2 carbons), 128.6 (2 car-
bons), 128.5 (4 carbons), 128.4 (2 carbons), 128.2 (2 carbons),
128.0 (2 carbons), 127.9, 127.8 (2 carbons), 127.7 (2 carbons),
127.4, 99.4, 95.0, 87.2, 80.0, 78.4, 77.6, 76.7, 76.0, 75.3, 74.7, 73.7
(2 carbons), 71.3, 70.0, 69.5, 59.8, 59.5, 54.7, 51.6, 32.8; ESI/APCI
Calcd for C46H50N12O9Na+ ([M+Na]+) m/e 937.3721; measured m/e
937.3702.
4.12. 6-O-(3-Azido-2-O-benzyl-3,6-dideoxy-a-D-glucopyranosyl)-
1,3,40,60-tetraazido-30,40-dideoxy-30-enoneamine (16)
The first step for the preparation of this compound was similar
to the synthesis of compound 13. The product from the first step
was dissolved in methanol (5 mL), and sodium methoxide (0.5 M
in MeOH, 1 mL) was added. The reaction mixture was stirred at
room temperature till the completion of the reaction (ca. 2 hours,
monitored by TLC, eluted with hexane/EtOAc = 50/50). The reac-
tion mixture was filtered through Celite and the residue was
washed with MeOH. After the removal of the solvents and purifica-
tion with column chromatography (eluted with hexane/EtOAc
from 90/10 to 35/65), the product was obtained as a solid
(0.26 g, 0.40 mmol, 78%). 1H NMR (CDCl3, 300 MHz) d 7.4 (m,
5H), 6.0 (m, 2H), 5.60 (s, 1H), 4.6–4.9 (m, 4H), 2.9–4.2 (m, 13H),
2.4 (m, 1H), 2.26 (td, J = 13.1, 4.5 Hz, 1H), 1.4 (m, 1H), 1.27 (d,
J = 3.8 Hz, 3H); 13C NMR (CDCl3, 100 MHz) d 137.4, 128.8 (2 car-
bons), 128.7, 128.5, 128.4 (2 carbons), 128.0, 97.7, 95.0, 87.2,
79.8, 78.3, 76.0, 74.0, 73.4, 69.7, 69.0, 64.9, 59.5, 59.4, 54.8, 51.8,
32.9, 17.9; ESI/APCI Calcd for C25H35N16O7 ([M+NH4]+) m/e
þ
671.2896; measured m/e 671.2852.
4.13. 6-O-(3-Azido-2-O-benzyl-3-deoxy-a-D-xylopyranosyl)-
1,3,40,60-tetraazido-30,40-dideoxy-30-enoneamine (18)
This compound was prepared similarly to the synthesis of com-
pound 16. 1H NMR (CDCl3, 400 MHz) d 7.4–7.5 (m, 5H), 5.95 (d,
J = 10.5 Hz, 1H), 5.890 (d, J = 10.8 Hz, 1H), 5.8 (d, 4.3 Hz, 1H), 4.9
(d, J = 3.0 Hz, 1H), 4.6–4.7 (m, 2H), 4.23 (d, J = 3.0 Hz, 1H), 4.09
(m, 1H), 3.6–3.9 (m, 6H), 3.2–3.5 (m, 8H), 2.32 (ddd, J = 13.2, 4.5,
4.5 Hz, 1H), 1.36 (ddd, J = 13.2, 12.5, 12.5 Hz, 1H); 13C NMR (CDCl3,
100 MHz) d 136.5, 128.6, 128.0, 127.7, 127.6, 122.7, 122.4, 97.1,
96.3, 85.0, 81.3, 79.0, 75.1, 73.0, 68.3, 67.5, 63.8, 62.7, 58.8, 58.7,
54.4, 53.3, 32.0, 29.1; MALDI Calcd for C24H29N15O7Na ([M+Na]+)
m/e 662.2267; measured m/e 662.2232.
4.10. 6-O-(1,2,3,6-Tetra-O-benzyl-b-D
-galactopyranosyl)-1,3,40,60-
tetraazido-20,30,40-trideoxy-20-enoneamine (epi-13)
Please refer to the procedure for the preparation of compound
13. 1H NMR (CDCl3, 300 MHz) d 7.3 (m, 20H), 5.90 (d, J = 10.0 Hz,
1H), 5.74 (dt, J = 12.4, 2.1 Hz, 1H), 5.52 (s, 1H), 5.05 (d, J = 3.5 Hz,
1H), 4.94 (t, J = 11.7 Hz, 1H), 4.91 (d, J = 8.3 Hz, 1H), 4.7–4.8
(m, 3H), 4.58 (d, J = 11.3 Hz, 1H), 4.4 (m, 3H), 4.1 (m, 2H), 4.0
(m, 1H), 3.9 (m, 2H), 3.81 (d, J = 1.7 Hz, 1H), 3.2–3.6 (m, 9H), 2.24
(td, J = 13.4, 4.5 Hz, 1H), 1.4 (m, 1H); 13C NMR (CDCl3, 100 MHz)
d 138.9, 138.5, 138.3, 137.7, 128.8 (2 carbons), 128.7, 128.6 (2 car-
bons), 128.5 (4 carbons), 128.4 (2 carbons), 128.3 (2 carbons),
4.14. 3,4-Di-O-acetyl-2-O-benzyl-1-phenylthio-b-D-ribopyrano-
side (20)
To a solution of 1915 (1.1 g, 3.3 mmol) and pyridine (1.22 ml,
15 mmol) in anhydrous CH2Cl2 at 0 °C, Tf2O (4.3 ml, 15 mmol)
was added slowly. After being stirred for 30 min., the reaction

J. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1396–1405
1403
mixture was diluted with CH2Cl2, washed with water, saturated
NaHCO3(aq), and brine, and then dried over Na2SO4. The solution
was filtered through glass wool and transferred into a solution of
tetrabutylammonium acetate (4.0 g, 13 mmol) in CH2Cl2. The reac-
tion mixture was stirred overnight while the solvent was slowly
evaporated with an aspirator. After completion of the reaction,
the reaction mixture was diluted with EtOAc, washed with 1 N
J = 9.1 Hz, 1H), 4.35 (dd, J = 11.5, 5.3 Hz, 1H), 3.85 (dd, J = 9.3,
9.6 Hz, 1H), 3.45 (dd, J = 9.1, 9.3 Hz, 1H), 3.40 (dd, J = 11.5, 9.5 Hz,
1H); 13C NMR (CDCl3, 100 MHz) d 165.6, 137.5, 133.8, 133.2,
132.5, 130.0 (2 carbons), 129.3 (3 carbons), 128.74 (3 carbons),
128.70 (3 carbons), 128.4 (2 carbons), 128.2, 88.7, 79.1, 75.6,
70.3, 67.7, 66.9; EI Calcd for C25H27N4O4S+ ([M+NH4]+) m/e
479.1748; measure m/e 479.1757.
HCl, saturated NaHCO3(aq), and brine, and then dried over Na2SO4(s)
.
4.17. 5-O-(4-Azido-4-deoxy-b-D
-glucopyranosyl)-1,3,40,60-tetra-
Removal of the solvent followed by purification with gradient col-
umn chromatography (eluted with hexane/EtOAc from 100/0 to
50/50) afforded the desired product (0.80 g, 1.9 mmol, 58%). 1H
NMR (CDCl3, 270 MHz) d 7.4–7.5 (m, 2H), 7.2–7.3 (m, 8H), 5.69
(m, 1H), 5.06 (d, J = 8.2 Hz, 1H), 4.96 (ddd, J = 9.2, 4.9, 3.3 Hz, 1H),
4.60 (d, J = 11.2 Hz, 1H), 4.45 (d, J = 11.2 Hz, 1H), 3.99 (d,
J = 11.2 Hz, 1H), 3.74 (dd, J = 11.2, 9.2 Hz, 1H), 3.49 (dd, J = 8.2,
3.0 Hz, 1H), 2.12 (s, 3H), 2.00 (s, 3H); 13C NMR (CDCl3, 100 MHz)
d 170.3, 170.0, 137.3, 132.8, 132.7 (2 carbons), 129.1 (2 carbons),
128.6 (2 carbons), 128.4 (2 carbons), 128.2, 128.0, 84.6, 75.1,
72.3, 67.4, 67.1, 63.9, 21.0, 20.9; HRFAB Calcd for C22H24O6SNa+
([M+Na]+) m/e 439.1186; measure m/e 439.1191.
azido-30,40-dideoxy-30-enoneamine (23)
A solution of compound 22 (0.83 g, 2.0 mmol), compound 8
(0.83 g, 1.7 mmol), and activated powder 4 Å molecular sieve
was stirred in anhydrous CH2Cl2 (8 mL) at 65 °C for 1 hr and
then added with BF3–OEt2 (0.05 mL, 0.42 mmol). After the reac-
tion was warmed to room temperature, the reaction was
quenched by adding few drops of water. After being stirred for
20 min, NaHCO3(s) was added, and then the reaction mixture
was filtered through a short column packed with layers of Celite
and silica gel. The column was washed with EtOAc. After re-
moval of the solvent and purification with column chromatogra-
phy (eluted with hexanes/EtOAc from 100/0 to 60/40) afforded
the intermediate product. The intermediate product was dis-
solved in methanol (5 mL) and few drops of water, and NaOMe
(catalytic amount) was added. The reaction mixture was stirred
at room temperature till the completion of the reaction (ca.
1 h, monitored by TLC, eluted with hexane/EtOAc = 50/50). The
reaction mixture was filtered through Celite and the residue
was washed with EtOAc. After the removal of the solvents and
purification with column chromatography (eluted with hexane/
EtOAc from 90/10 to 30/70), the product was obtained as a solid
(0.52 g, 0.92 mmol, 56%). 1H NMR (CD3OD, 300 MHz) d 6.10 (dt,
J = 10.3, 2.4 Hz, 1H), 5.99 (d, J = 10.0 Hz, 1H), 5.63 (s, 1H), 4.60
(d, J = 8.2 Hz, 1H), 4.01 (dt , J = 10.0, 4.1 Hz, 1H), 3.87 (dd,
J = 10.0, 1.7 Hz, 1H), 3.6 (m, 2H), 3.2–3.5 (m, 8H), 2.99 (t,
J = 9.6 Hz, 1H), 2.16 (td, J = 13.4, 4.5 Hz, 1H), 1.33 (d, J = 6.2 Hz,
3H), 1.2 (m, 1H); 13C NMR (CD3OD, 100 MHz) d 128.9, 127.2,
104.4, 94.3, 85.5, 78.1, 77.0, 75.9, 75.0, 70.9, 69.9, 68.3, 60.0,
4.15. 4-O-Benzoyl-2-O-benzyl-1-phenylthio-b-D-ribopyranoside
(21)
To a solution of 20 (0.79 g, 1.9 mmol) in MeOH (50 mL) and
THF (10 mL), few drops of 1 M NaOMe was added. The reaction
was stirred for 2 h. TLC indicated the completion of the reaction.
The reaction was quenched by Amberlite IR-120 (H+) and filtra-
tion. Removal of the solvent afforded the diol which was dis-
solved in anhydrous CH2Cl2 (30 mL) and then DMAP (catalytic
amount), DIPEA (0.54 ml, 3.2 mmol), and BzCl (0.20 ml, 1,8 mmol)
were added at ꢀ50 °C. The reaction mixture was stirred and al-
lowed to warm to ꢀ10 °C. Water was added to quench the reac-
tion. After removal of the solvent, the reaction mixture was
diluted with EtOAc. The organic layer was washed with 1 N HCl,
saturated NaHCO3(aq), and brine, and then dried over Na2SO4(s)
.
Removal of the solvent followed by purification with gradient col-
umn chromatography (eluted with hexane/EtOAc from 100/0 to
60/40) afforded the desired product (0.40 g, 0.92 mmol, 48%). 1H
NMR (CDCl3, 300 MHz) d 8.1 (m, 2H), 7.59 (tt, J = 7.2, 1.4 Hz,
1H), 7.4–7.5 (m, 4H), 7.2–7.4 (m, 8H), 5.63 (d, J = 2.4 Hz, 1H),
5.32 (t, J = 3.1 Hz, 1H), 4.74 (d, J = 11.7 Hz, 1H), 4.66 (d,
J = 11.7 Hz, 1H), 4.41 (dd, J = 12.4, 1.0 Hz, 1H), 4.15 (s, 1H), 4.08
(s, broad, 1H), 3.93 (dd, J = 12.4, 2.4 Hz, 2H); 13C NMR (CDCl3,
100 MHz) d 166.0, 136.9, 133.9, 133.69 (2 carbons), 131.7 (2 car-
bons), 130.2, 129.9, 129.4 (2 carbons), 128.8 (2 carbons), 128.7 (2
carbons), 128.2 (2 carbons), 127.9 (2 carbons), 86.2, 78.0, 73.4,
69.4, 68.4, 66.2; ESI/APCI Calcd for C25H24O5SNa+ ([M+Na]+) m/e
459.1237; measure m/e 459.1235.
þ
59.9, 55.1, 51.6, 32.2, 17.7; ESI/APCI Calcd for C18H26N15O7
([M+H]+) m/e 564.2134; measure m/e 564.2125.
4.18. 6-O-Hexylaminocarbonyl-30,40-dideoxyneamine (RR101)
This compound was prepared similarly to the synthesis of
compound 11 except no NaHCO3(s) was used in the first step,
but MeOH and couple drops of HOAc/H2O (1/4) was used as the
solvent in the second step. 1H NMR (D2O, 300 MHz) d 5.66 (d,
J = 3.5 Hz, 1H), 2.9–4.1 (m, 13H), 2.44 (td, J = 12.4, 8.6 Hz, 1H),
1.9 (m, 1H), 1.7–1.8 (m, 2H), 1.3 (m, 2H), 1.2 (m, 6H), 0.73 (t,
J = 6.5 Hz, 3H); 13C NMR (D2O, 100 MHz) d 157.2, 95.2, 76.5,
74.1, 73.4, 66.0, 48.9, 48.6, 48.3, 42.8, 41.0, 30.9, 28.8, 28.4,
4.16. 3-Azido-4-O-benzoyl-2-O-benzyl-3-deoxy-1-phenylthio-b-
D
-xylopyranoside (17)
þ
25.8, 25.7, 22.1, 20.5, 13.5; ESI/APCI Calcd for C19H42N5O6
([M+H2O]+) m/e 436.3130; measured m/e 436.2763.
To a solution of 21 (0.39 g, 0.9 mmol) and pyridine (0.18 mL,
39 mmol) in anhydrous CH2Cl2 at 0 °C, Tf2O (0.3 mL, 2.3 mmol)
was added slowly. After being stirred for 30 min, the reaction mix-
ture was diluted with CH2Cl2, washed with water, saturated NaH-
CO3(aq), and brine, and then dried over Na2SO4. After the removal
of the solvent, the crude product was dissolved into DMF (20 mL)
and to this NaN3 (0.46 g, 7.1 mmol) was added. The reaction was
stirred for 24 h at room temperature. Removal of the solvent fol-
lowed by purification with gradient column chromatography
(eluted with hexane/EtOAc from 100/0 to 50/50) afforded the de-
sired product (0.35 g, 0.76 mmol, 85%). 1H NMR (CDCl3, 400 MHz)
d 8.0–8.1 (m, 2H), 7.2–7.6 (m, 13H), 5.04 (ddd, J = 9.6, 9.5, 5.3 Hz,
1H), 4.95 (d, J = 10.2 Hz, 1H), 4.80 (d, J = 10.2 Hz, 1H), 4.75 (d,
4.19. 40-Amino-6-O-hexylaminocarbonyl-1,3,40,60-tetraamino-
20,30,40-trideoxyneamine (NRR101)
This compound was prepared similarly to the synthesis of com-
pound 11 but using MeOH and couple drops of HOAc/H2O (1/4)
as the solvent. 1H NMR (D2O, 300 MHz) d 5.66 (d, J = 3.5 Hz, 1H),
2.9–4.3 (m, 11H), 2.4 (m, 1H), 1.5–1.9 (m, 5H), 1.4 (m, 2H), 1.2
(m, 6H), 0.73 (t, J = 6.2 Hz, 3H); 13C NMR (D2O, 100 MHz) d 157.3,
95.2, 74.3, 73.8, 73.0, 66.0, 48.9, 48.6, 48.3, 42.8, 41.0, 40.8, 30.9,
þ
28.8, 28.3, 25.8, 22.1, 20.6, 13.5; ESI/APCI Calcd for C19H41N5O5
([M+H]+) m/e 418.3024; measured m/e 418.3027.

1404
J. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1396–1405
4.20. 40-Amino-5,6-di-O-hexylaminocarbonyl-1,3,40,60-tetraamino-
4.25. 6-O-(4-Amino-4,6-dideoxy-b-D
-glucopyranosyl)-20,30,40-
20,30,40-trideoxyneamine (NRR102)
trideoxy-40-aminoneamine (NRR501)
This compound was prepared similarly to the synthesis of com-
pound 11 but using MeOH and couple drops of HOAc/H2O (1/4) as
the solvent in the second step. 1H NMR (D2O, 300 MHz) d 5.24 (d,
J = 3.1 Hz, 1H), 4.9 (m, 2H), 4.33 (t, J = 10.0 Hz, 1H), 4.1 (m, 1H),
3.6 (m, 3H), 3.4 (m, 1H), 3.15 (d, J = 5.9 Hz, 2H), 2.9–3.0 (m, 3H),
2.48 (td, J = 12.7, 4.1 Hz, 1H), 2.0 (m, 4H), 1.8 (m, 1H), 1.3 (m,
4H), 1.2 (m, 12H), 0.75 (t, J = 4.5 Hz, 6H); 13C NMR (D2O,
100 MHz) d 156.8, 156.5, 93.2, 75.2, 73.8, 72.6, 67.9, 48.7, 48.4,
48.1, 41.9, 41.2 (2 carbons), 31.2 (2 carbons), 29.1, 29.0, 28.0,
26.2 (2 carbons), 24.0, 22.3 (2 carbons), 20.7, 13.7 (2 carbons);
This compound was prepared similarly to the synthesis of com-
pound 11 but using MeOH and couple drops of HOAc/H2O (1/4) as
the solvent in the second step. 1H NMR (D2O, 300 MHz) d 5.83 (t,
J = 3.4 Hz, 1H), 4.6 (m, 1H), 3.89 (td, J = 8.6, 3.5 Hz, 1H), 2.8–3.6
(m, 11H), 2.61 (t, J = 10.0 Hz, 1H), 2.11 (td, J = 12.4, 3.8 Hz, 1H),
1.4–1.9 (m, 5H), 1.19 (d, J = 6.2 Hz, 3H); 13C NMR (D2O, 75 MHz)
d 102.5, 96.8, 81.8, 78.3, 74.3, 73.8, 73.4, 70.4, 69.5, 57.4, 50.3,
49.0, 47.7, 40.4, 31.2, 27.0, 23.2, 17.2; ESI/APCI Calcd for
C18H38N5O7 ([M+H]+) m/e 436.2766; measured m/e 436.2762.
þ
ESI/APCI Calcd for C26H54N6O6 ([M+H]+) m/e 545.4021; measured
þ
4.26. 30,40-Di-O-benzyl-6-O-hexylaminocarbonyl-1,3,40,60-tetra-
azidoneamine (25)
m/e 545.4021.
4.21. 6-O-(
amine (NRT014)
a
-
D
-Galactopyranosyl)-20,30,40-trideoxy-40-aminone-
This compound was prepared similarly to the synthesis of com-
pound 5. 1H NMR (CDCl3, 300 MHz) d 7.2–7.4 (m, 10H), 5.33 (d,
J = 3.8 Hz, 1H), 4.8–4.9 (m, 5H), 4.62 (d, J = 11.0 Hz, 1H), 4.1–4.2
(m, 2H), 3.98 (t, J = 9.6 Hz, 1H), 3.2–3.6 (m, 10H), 2.3 (m, 1H),
1.4–1.5 (m, 3H), 1.3 (m, 6H), 0.88 (t, J = 6.2 Hz, 3H); 13C NMR
(CDCl3, 100 MHz) d 156.1, 137.8, 137.7, 128.79 (2 carbons),
128.75 (2 carbons), 128.3 (2 carbons), 128.0 (2 carbons), 99.3,
83.2, 80.8, 78.8, 76.4, 75.8, 75.7, 75.5, 71.4, 64.2, 59.2, 58.5, 51.2,
41.7, 32.2, 31.8, 29.9, 29.8, 26.8, 26.6, 22.8, 14.2; ESI/APCI Calcd
This compound was prepared similarly to the synthesis of com-
pound 11 but using MeOH and couple drops of HOAc/H2O (1/4) as
the solvent in the second step. 1H NMR (D2O, 300 MHz) d 5.32 (t,
J = 4.8 Hz, 1H), 4.97 (d, J = 3.4 Hz, 1H), 4.2 (m, 1H), 4.01 (t,
J = 5.9 Hz, 1H), 3.88 (d, J = 2.4 Hz, 1H), 3.2–3.8 (m, 11H), 2.7 (m,
1H), 2.37 (td, J = 12.4, 4.1 Hz, 1H), 1.7–2.0 (m, 5H); 13C NMR
(D2O, 100 MHz) d 101.8, 96.6, 83.6, 77.9, 73.9, 72.1, 69.5(2 car-
bons), 69.1, 68.9, 61.1, 50.0, 48.7, 47.1, 39.4, 27.8, 26.2, 21.1; ESI/
for C33H43N13O7 ([M+H]+) m/e 734.3481; measure m/e 734.3485.
þ
APCI Calcd for C18H37N4O9 ([M+H]+) m/e 453.2555; measured m/
e 453.2549.
þ
4.27. 30,40-Di-O-benzyl-5,6-Di-O-hexylaminocarbonyl-1,3,40,60-
tetraazidoneamine (26)
4.22. 6-O-(
a
-
D
-Galactopyranosyl)-30,40-dideoxyneamine (RT014)
This compound was prepared similarly to the synthesis of com-
pound 5. 1H NMR (CDCl3, 300 MHz) d 7.2–7.4 (m, 10H), 5.33 (d,
J = 3.4 Hz, 1H), 4.89 (d, J = 12.7 Hz, 1H), 4.86 (d, J = 1.7 Hz, 2H),
4.60 (d, J = 11.0 Hz, 1H), 4.09 (t, J = 9.6 Hz, 1H), 3.8–4.0 (m, 8H),
3.2–3.7 (m, 8H), 2.49 (td, J = 8.9, 5.2 Hz, 1H), 1.5–1.6 (m, 5H), 1.3
(m, 12H), 0.88 (t, J = 4.5 Hz, 6H); 13C NMR (CDCl3, 100 MHz) d
156.6, 152.7, 137.69, 137.66, 128.8 (3 carbons), 128.3 (3 carbons),
128.2 (2 carbons), 128.1 (2 carbons), 96.5, 81.0 (2 carbons), 79.9,
78.5, 75.8, 75.4, 75.2, 71.9, 63.1, 60.2, 56.3, 51.1, 43.5, 40.9, 33.9,
31.7, 29.8, 29.0, 26.9 (2 carbons), 26.8, 22.8 (2 carbons), 14.2 (2 car-
bons); ESI/APCI Calcd for C40H56N14O8Na+ ([M+Na]+) m/e 883.4289;
measured m/e 883.4288.
This compound was prepared similarly to the synthesis of com-
pound 11 except no NaHCO3(s) was used in the first step. 1H NMR
(D2O, 300 MHz) d 5.65 (d, J = 3.3 Hz, 1H), 5.01 (d, J = 3.6 Hz, 1H),
4.06 (t, J = 6.3 Hz, 2H), 3.6–3.9 (m, 9H), 3.4–3.5 (m, 2H), 3.13 (dd,
J = 13.4, 3.1, 1H), 2.98 (dd, J = 13.4, 7.2 Hz, 1H), 2.38 (dt, J = 12.7,
4.1 Hz, 1H), 1.7–1.9 (m, 4H), 1.5 (m, 1H); 13C NMR (D2O,
100 MHz) d 101.7, 95.7, 83.8, 77.4, 74.4, 72.6, 69.4, 69.1, 68.7,
66.0, 61.3, 49.7, 48.9, 48.6, 42.7, 28.0, 25.6, 20.6; ESI/APCI Calcd
for C18H37N4O9 ([M+H]+) m/e 453.2555; measured m/e 453.2565.
þ
4.23. 6-O-(3-Amino-3,6-dideoxy-
trideoxy-40-aminoneamine (NRT040)
a-D
-glucopyranosyl)-20,30,40-
4.28. 6-O-Hexylaminocarbonylneamine (RR103)
This compound was prepared similarly to the synthesis of com-
pound 11 but using MeOH and couple drops of HOAc/H2O (1/4) as
the solvent in the second step. 1H NMR (D2O, 400 MHz) d 5.31 (s,
1H), 4.91 (s, 1H), 4.2 (m, 1H), 3.2–3.8 (m, 11H), 3.1 (m, 1H), 2.4
(m, 1H), 1.8–2.0 (m, 5H), 1.14 (d, J = 5.6 Hz, 3H); 13C NMR (D2O,
100 MHz) d 100.9, 96.8, 83.5, 78.2, 73.6, 71.0, 70.0, 68.9, 68.6,
55.0, 50.3, 48.8, 47.0, 39.3, 27.8, 25.8, 21.0, 16.5; ESI/APCI Calcd
This compound was prepared similarly to the synthesis of com-
pound 11. 1H NMR (D2O, 300 MHz) d 5.77 (d, J = 3.8 Hz, 1H), 3.7–
3.9 (m, 4H), 2.9–3.5 (m, 9H), 2.40 (td, J = 8.6, 4.1 Hz, 1H), 1.84
(td, J = 12.5, 3.4 Hz, 1H), 1.35 (q, J = 6.5 Hz, 2H), 1.1 (m, 6H), 0.71
(t, J = 6.5 Hz, 3H); 13C NMR (D2O, 100 MHz) d 157.3, 96.0, 77.7,
74.4, 73.7, 70.9, 69.3, 68.4, 53.7, 48.4, 48.3, 40.9, 40.3, 30.8, 28.9,
28.8, 25.8, 22.1, 13.5; ESI/APCI Calcd for C19H40N5O7 ([M+H]+)
þ
for C18H38N5O7 ([M+H]+) m/e 436.2766; measured m/e 436.2771.
m/e 450.2922; measured m/e 450.2928.
þ
4.24. 6-O-(3-Amino-3-deoxy-
a
-
D
-xylopyranosyl)-30,40-dideoxy-
4.29. 5,6-Di-O-hexylaminocarbonylneamine (RR104)
neamine (RT040)
This compound was prepared similarly to the synthesis of com-
pound 11. 1H NMR (D2O, 300 MHz) d 5.20 (d, J = 3.8 Hz, 1H), 4.85 (t,
J = 9.3 Hz, 1H), 4.74 (t, J = 10.0 Hz, 1H), 3.6–3.9 (m, 3H), 2.8–3.4 (m,
10H), 2.15 (td, J = 8.9, 4.1 Hz, 1H), 1.55 (td, J = 12.6, 12.5 Hz, 1H),
1.3 (m, 4H), 1.1 (m, 12H), 0.70 (t, J = 6.9 Hz, 6H); 13C NMR (D2O,
100 MHz) d 160.0, 156.7, 94.1, 75.6, 75.2, 73.3, 70.7, 69.3, 68.5,
53.5, 48.6, 48.3, 41.2, 41.1, 40.2, 30.9 (2 carbons), 29.8, 28.8 (2 car-
bons), 25.9, 25.8, 22.1 (2 carbons), 13.5 (2 carbons); ESI/APCI Calcd
This compound was prepared similarly to the synthesis of com-
pound 11 except no NaHCO3(s) was used in the first step. 1H NMR
(D2O, 400 MHz) d 5.79 (d, J = 3.5 Hz, 1H), 5.04 (d, J = 3.6 Hz, 1H), 4.3
(m, 1H), 3.5–4.0 (m, 9H), 3.0–3.5 (m, 8H), 2.54 (ddd, J = 13.2, 4.5,
4.5 Hz, 1H), 1.70 (ddd, J = 13.2, 12.5, 12.5 Hz, 1H,); 13C NMR (D2O,
100 MHz) d 100.8, 95.2, 84.0, 77.1, 74.5, 68.3, 66.2, 65.6, 57.1,
55.3, 49.9, 48.9, 48.7, 42.8, 27.1, 25.6, 20.7; HRFAB Calcd for
C17H36N5O7 ([M+H]+) m/e 422.2615; measured m/e 422.2603.
for C26H53N6O8 ([M+H]+) m/e 577.3919, measured m/e 577.3929.
þ
þ

J. Zhang et al. / Bioorg. Med. Chem. 18 (2010) 1396–1405
2. Fujisawa, K.; Hashiya, T.; Kawaguchi, H. J. Antibiot. 1974, 27, 677–681.
1405
4.30. Procedure for MIC determination
3. Fourmy, D.; Recht, M. I.; Blanchard, S. C.; Puglisi, J. D. Science 1996, 274, 1367–
1371.
A solution of selected bacteria was inoculated in the Trypticase
Soy broth at 35 °C for 1–2 h. After which, the bacteria concentration
was found, and diluted with broth, if necessary, to an absorption
value of 0.08 to 0.1 at 625 nm. The adjusted inoculated medium
4. Greenberg, W. A.; Priestley, E. S.; Sears, P. S.; Alper, P. B.; Rosenbohm,
C.; Hendrix, M.; Hung, S.-C.; Wong, C.-H. J. Am. Chem. Soc. 1999, 121,
6527–6541.
5. Roestamadji, J.; Grapsas, I.; Mobashery, S. J. Am. Chem. Soc. 1995, 117, 11060–
11069.
6. Rai, R.; Chang, H.; Chen, H.-N.; Chang, C.-W. T. J. Carbohydr. Chem. 2005, 24,
131–143.
7. (a) March, J. Advanced Organic Chemistry, 4th ed.; John Wiley & Son: New York,
1992; (b) Feldman, A. K.; Colasson, B.; Sharpless, K. B.; Fokin, V. V. J. Am. Chem.
Soc. 2005, 127, 13444–13445.
8. We have conducted serial reactions for the purpose of revealing the factors that
cause the allylic azide rearrangement. However, no allylic azide rearrangement
was observed except in the presence of triethyl ammonium salt, which
triggered very minimum rearrangement. Please refer Supplementary data for
details. Our conclusion is that thermodynamic condition is the most reliable
method to conduct allylic azide rearrangement in our system.
9. Wang, J.; Li, J.; Chen, H.-N.; Chang, H.; Tanifum, C. T.; Liu, H.-H.; Czyryca, P. G.;
Chang, C.-W. T. J. Med. Chem. 2004, 12, 6397–6413.
(100
l
L) was diluted with 10 mL broth, and then applied to a
L). A series of solutions (50 L each
96-well microtiter plate (50
l
l
in 2-fold dilution) of the tested compounds was added to the testing
wells. The 96-well plate was incubated at 35 °C for 12–18 h. The
minimum inhibitory concentration (MIC) is defined as the mini-
mum concentration of compound needed to inhibit the growth of
bacteria. The MIC results are repeated at least three times.
Acknowledgments
10. Chang, C.-W. T.; Hui, Y.; Elchert, B.; Wang, J.; Li, J.; Rai, R. Org. Lett. 2002, 4,
4603–4606.
11. Zhang, J.; Chiang, F.-I.; Wu, L.; Czyryca, G. P.; Li, D.; Chang, C.-W. T. J. Med. Chem.
2008, 51, 7563–7573.
We acknowledge National Institutes of Health (AI053138) for
financial support.
12. Hachiler, H.; Santanam, P.; Kayser, F. H. Antimicrob. Agents Chemother. 1996, 40,
1254–1256.
Supplementary data
13. Ida, T.; Okamoto, R.; Shimauchi, C.; Okubo, T.; Kuga, A.; Inoue, M. J. Clin.
Microbiol. 2001, 39, 3115–3121.
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmc.2010.01.027.
14. (a) Guo, H.; O’Doherty, G. A. Angew. Chem., Int. Ed. 2007, 46, 5206–5208;
(b) Guo, H.; O’Doherty, G. A. Org. Lett. 2006, 8, 1609–1612; (c) Guo, H.;
O’Doherty, G. A. Tetrahedron 2008, 64, 304–313; (d) Abrams, J. A.; Babu, R.
S.; Guo, H.; Le, D.; Le, J.; Osbourn, J. M.; O’Doherty, G. A. J. Org. Chem.
2008, 73, 1935–1940; (e) Guppi, S. R.; O’Doherty, G. A. J. Org. Chem. 2007,
72, 4966–4969; (f) Guo, H.; O’Doherty, G. A. J. Org. Chem. 2008, 73, 5211–
5220.
References and notes
1. For reviewing: (a) Umezawa, H.; Hooper, I. R. Aminoglycoside Antibiotics;
Springer: New York, 1982; (b) Haddad, J.; Kotra, L. P.; Mobashery, S. In
Glycochemistry Principles, Synthesis, and Applications; Wang, P. G., Bertozzi, C. R.,
Eds.; Marcel Dekker, 2001; pp 307; (c) Wang, J.; Chang, C.-W. T. In Aminoglycoside
Antibiotics; Arya, D. P., Ed.; John Wiley & Sons, 2007; pp 141–180.
15. Compound 19 was prepared according to the same procedure as the
preparation of 2-O-benzyl-1-phenylthio-b-D-fucopyranoside described in
Ref. 9.