Stereospecific synthesis of methyl 2-amino-2,4-dideoxy-6S-deuterio-α-D-xylo-hexopyranoside and methyl 2-amino-2,4-dideoxy-6S-deuterio-4-propyl-α-D-glucopyranoside: Side chain conformation of the novel aminoglycoside antibiotic propylamycin

Article abstract of DOI:10.1016/j.carres.2020.107984

The stereospecific syntheses of methyl 2-amino-2,4-dideoxy-4-C-propyl-α-D-glucopyranoside and of methyl 2-amino-2,4-dideoxy-α-D-xylo-hexopyranoside and of their 6S-deuterioisotopomers are described as models for ring I of the aminoglycoside antibiotics propylamycin and 4′-deoxyparomomycin, respectively. Analysis of the ¹H NMR spectra of these compounds and of methyl 2-amino-2-deoxy-α-D-glucopyranoside, a model for paromomycin itself, reveals that neither deoxygenation at the 4-position, nor substitution of the C–O bond at the 4-postion by a C–C bond significantly changes the distribution of the side chain population between the three possible staggered conformations. From this it is concluded that the beneficial effect on antiribosomal and antibacterial activity of the propyl group in propylamycin does not derive from a change in side chain conformation. Rather, enhanced basicity of the ring oxygen and increased hydrophobicity and/or solvation effects are implicated.

Full text of DOI:10.1016/j.carres.2020.107984

CarbohydrateResearch491(2020)107984
Contents lists available at ScienceDirect
Carbohydrate Research
journal homepage: www.elsevier.com/locate/carres
Stereospecific synthesis of methyl 2-amino-2,4-dideoxy-6S-deuterio-α-D-
xylo-hexopyranoside and methyl 2-amino-2,4-dideoxy-6S-deuterio-4-propyl-
α-D-glucopyranoside: Side chain conformation of the novel aminoglycoside
antibiotic propylamycin
Michael G. Pirronea,b,c,d, Takahiko Matsushitad, Andrea Vasellae, David Cricha,b,c,d,∗
a Department of Pharmaceutical and Biomedical Sciences, University of Georgia, 250 West Green Street, Athens, GA, 30602, USA
b Department of Chemistry, University of Georgia, 140 Cedar Street, Athens, GA, 30602, USA
c Complex Carbohydrate Research Center, University of Georgia, 315 Riverbend Road, Athens, GA, 30602, USA
d Department of Chemistry, Wayne State University, 5101 Cass Avenue, Detroit, MI, 48202, USA
e Organic Chemistry Laboratory, ETH Zürich, Vladimir-Prelog-Weg 1-5/10, 8093, Zürich, Switzerland
A B S T R A C T
The stereospecific syntheses of methyl 2-amino-2,4-dideoxy-4-C-propyl-α-D-glucopyranoside and of methyl 2-amino-2,4-dideoxy-α-D-xylo-hexopyranoside and of
their 6S-deuterioisotopomers are described as models for ring I of the aminoglycoside antibiotics propylamycin and 4′-deoxyparomomycin, respectively. Analysis of
the 1H NMR spectra of these compounds and of methyl 2-amino-2-deoxy-α-D-glucopyranoside, a model for paromomycin itself, reveals that neither deoxygenation at
the 4-position, nor substitution of the C–O bond at the 4-postion by a C–C bond significantly changes the distribution of the side chain population between the three
possible staggered conformations. From this it is concluded that the beneficial effect on antiribosomal and antibacterial activity of the propyl group in propylamycin
does not derive from a change in side chain conformation. Rather, enhanced basicity of the ring oxygen and increased hydrophobicity and/or solvation effects are
implicated.
1. Introduction
In this paper we explore the hypothesis that substitution at the 4′-
position in ring I of paromomycin affects the distribution of the ring I
side chain between the three staggered conformations, gauche,gauche
(gg), gauche,trans (gt) and trans,gauche (tg) (Fig. 1) [3–5] and so influ-
ences a critical hydrogen bonding interaction between the aminogly-
coside and the drug binding pocket in the decoding A site of helix 44 on
the small ribosomal subunit.
Propylamycin, 4′-deoxy-4′-C-propylparomomycin, 1 is a novel syn-
thetic 4,5-disubstituted 2-deoxystreptamine class aminoglycoside anti-
biotic that shows increased antibacterial and antibacterioribosomal
activity compared to the parent paromomycin 2, as well as reduced
ototoxicity in the guinea pig model [1]. 4′-Deoxyparomomycin 3 on the
other hand is somewhat less active than the parent 2, with which it
shows no significant difference in selectivity for prokaryotic over eu-
karyotic ribosomes [1,2].
It is known that binding of deoxystreptamine type aminoglycosides
to the decoding A site on the bacterial ribosome locks the side chain of
ring I into the gt conformation by virtue of its participation in a pseudo-
base pair interaction with A1408 [6,7]. Indeed, we have demonstrated
that a model of 2 with the ring I side locked into the gt conformation
shows activity and selectivity approaching that of propylamycin [8]. On
the basis of these observations we were driven to study the influence of
the 4′-deoxy and 4′-deoxy-4′-C-propyl modifications on the side chain
conformation of ring I in the paromomycin series. In view of the
complexity of the NMR spectra of the aminoglycosides, and the need for
unambiguous assignment of the diastereotopic side chain protons H6R
and H6S, we elected to study simplified monosaccharide models of ring
I. To this end we describe the synthesis of methyl 2-amino-2,4-dideoxy-
α-D-xylo-hexopyranoside 4α and methyl 2-amino-2,4-dideoxy-4-C-
∗ Corresponding author. Department of Pharmaceutical and Biomedical Sciences, University of Georgia, 250 West Green Street, Athens, GA, 30602, USA.
E-mail address: David.crich@uga.edu (D. Crich).
https://doi.org/10.1016/j.carres.2020.107984
Received 18 February 2020; Received in revised form 11 March 2020; Accepted 12 March 2020
Availableonline16March2020
0008-6215/©2020ElsevierLtd.Allrightsreserved.

M.G. Pirrone, et al.
CarbohydrateResearch491(2020)107984
16 with sodium methoxide as described in the literature [17,18].
Opening of epoxide 16 with sodium azide under a variety of conditions
was unsuccessful, but heating in benzylamine to 155 °C gave the Fürst-
Plattner [19] product 17 in 77% yield (Scheme 2). Hydrogenolysis of
the benzyl amine and saturation of the double bond gave the amine 18
in 79% yield, which was converted with Stick's reagent [20–23] to the
corresponding azido derivative 19 in 91% yield. Ring opening of 19
with TFA and acetic anhydride followed by Fischer glycosylation in
methanolic HCl resulted in a 1.5:1 anomeric mixture of 20 in 69%
yield, from which the individual anomers 20α and 20β were isolated in
22% and 12% yield, respectively. Hydrogenolysis of 20α gave the
acetate salt 5 in 96% yield (Scheme 2). The isotopomer 6S–2H-5 was
synthesized analogously and with comparable yield from 6-exo-deu-
terio 10. Comparison of the 1H NMR spectra of 5 and its 6S-deuterio
isotopomer enabled assignment of the diastereotopic protons at the 6-
position.
Fig. 1. Staggered conformations of the hexopyranose side chain.
propyl-α-D-glucopyranoside 5, and for purposes of spectral elucidation
their 6S-deuterio isotopomers, and the assignment of their side chain
conformations by analysis of the 5,6R and 5,6S 1H,1H 3J coupling
constants. Methyl 2-amino-2-deoxy-α-D-glucopyranoside 6 [9] and its
6S-deuterio isotopomer is employed as model for ring I of paromomycin
itself.
The differing regioselectivity in the ring openings of epoxides 12
and 16 with lithium azide and benzylamine, respectively, is of interest.
The opening of 16 with benzylamine (Scheme 2) is consistent with the
Fürst-Plattner rule and stereoelectronic control, with 1,3-diaxial inter-
actions between the incoming nucleophile and the propyl substituent
minimized due to the flattening of the pyranose ring imposed by the
1,6-anhydro bridge [24,25] and consistent with the anti-reflex effect
[26,27]. On the other hand, the opening of 12 with azide affords a
mixture of regioisomers (Scheme 1) in which the Fürst-Plattner product
14, formed directly in a chair conformation with trans-diaxial sub-
stituents, is minor. In contrast the major isomer 13 is necessarily
formed in a twist boat conformation before relaxation to the observed
chair with its two equatorial substituents. Presumably this diversion
from the usual Fürst-Plattner [19,28] is due to the build up of positive
charge on carbon during the ring opening of the protonated epoxide by
azide, which is better accommodated on C3 adjacent to the methylene
group than on C2 adjacent to the electron-withdrawing anomeric center
(Scheme 3).
A related example was recently observed in a synthesis of bra-
dyrhizose [29], wherein a cyclohexane-based trisubstituted epoxide
underwent ring opening under acidic conditions in an anti-Fürst
Plattner manner; an observation that was rationalized by preferential
localization of partial positive charge on the tertiary carbon at the
transition state (Scheme 4).
2. Results and discussion
2.1. Synthesis
Methyl 2-amino-2,4-dideoxy-α-D-xylo-hexopyranoside 4α was ob-
tained by hydrogenolysis of the corresponding benzylcarbamate 7,
which was prepared according to Mayer (Scheme 1) [10]. The synthesis
of its 6S-deuterio isotopomer began with stereospecific introduction of
deuterium into the 6-exo-position of 1,6-anhydro-D-glucose giving 8
under radical conditions following the literature protocol [11,12], as
modified in our laboratory [9] by replacement of tetrachloromethane
by the more environmentally friendly α,α,α-trifluorotoluene [13].
Following the established protocol, 8 was converted to the epoxide 10
in two straightforward steps [9,14–16]. Ring opening of 10 with
BF3·OEt2 and NaBH4 in dimethoxyethane gave 11 in 99% yield (Scheme
1), which, on treatment with sodium methoxide formed the 2,3-epoxide
12 in 99% yield. Microwave heating of 12 to 110 °C in DMF with li-
thium azide and benzoic acid afforded 43% of a 1.3:1 mixture of re-
gioisomeric azidoalcohols 13 and 14 from which 5% of the desired 14
was isolated by HPLC purification. Ring opening of 14 with TFA and
acetic anhydride was followed by Fischer glycosylation in methanolic
HCl and hydrogenolysis, resulting in an 85% yield of the target 6S–2H-4
in a mixture with the corresponding β-anomer, from which it could not
be easily separated. Nevertheless, the NMR spectra of 6S–2H-4α in the
mixture of anomers were well resolved, and permitted unambiguous
assignment of the resonances from the diastereotopic 6-HR and 6-HS in
isotopomers 4α.
2.2. Analysis of side chain populations
3
3
The JH5,H6R and JH5,H6S coupling constants for 4α and 5, taken
from 1H NMR spectra recorded at pH 5 in D2O at room temperature, are
reported in Table 1. The corresponding coupling constants for methyl 2-
amino-2-deoxy-α-D-glucopyranoside 6 (Table 1) are literature values
from our laboratory recorded under the same conditions [9].
Using limiting values of 3JR,gg - 3JS,tg for each of the pure gg, gt, and tg
conformers (Table 2) taken from a recent study employing a series of
conformationally rigid bicyclic models [30], the side chain populations
of all compounds (fgg – ftg (Table 3)), were determined in the standard
manner [3,31–33] with the aid of eqs (1)–(3). The population of −2%
calculated for the tg conformers of 6 and 5 is an artifact and arises from
the errors in the coupling constants, estimated from the digital re-
solution of the spectra to be 0.4 Hz, and is considered to be indis-
tinguishable from a 0% population.
3
3
3
3JH5,H6R = JR, gg fgg + JR,gt fgt + JR,tg ftg
eq 1
3
3
3
3JH5,H6S = JS, gg fgg + JS,gt fgt + JS,tg ftg
eq 2
eq 3
1 = fgg + fgt + ftg
The synthesis of methyl 2-amino-2,4-dideoxy-4-propyl-α-D-gluco-
pyranoside (Scheme 2) began with the 3,4-epoxy tosylate 10, which
was converted to the alcohol 15 by treatment with allylmagnesium
chloride in the presence of copper(I) iodide and then to the 2,3-epoxide
A modest increase in the population of the gt conformation, at the
expense of a corresponding decrease in the gg conformation, is observed
on replacement of the equatorial hydroxyl group in methyl 2-amino-2-
2

M.G. Pirrone, et al.
CarbohydrateResearch491(2020)107984
Scheme 1. Synthesis of Methyl 2-amino-2,4-dideoxy–α-D-xylo-hexopyranoside 4 and its 6S-Deuterio Isotopomer.
deoxy-α-D-glucopyranoside 6 by a proton in methyl 2-amino-2,4-di-
deoxy-α-D-glucopyranoside 4α (Table 3, entries 1 and 2). This pattern
of changes mirrors that observed earlier by Bock and Duus [3] on going
from methyl α-D-glucopyranoside to methyl 4-deoxy-α-D-glucopyrano-
side, supporting our earlier conclusions that the presence of protonated
amino group at the 2-position has little influence on the side chain
conformation [9]. Replacement of the hydroxyl group in 6 by the
propyl group in methyl 2-amino-2,4-dideoxy-4-propyl-α-D-glucopyr-
anoside 5 (Table 3, entries 1 and 3) results in a side chain population
that is intermediate between that of 6 and 4α. Clearly, the overall
Scheme 2. Synthesis of Methyl 2-amino-2,4-dideoxy-4-propyl–α-D-glucopyranoside 5 and its 6S-Deuterio Isotopomer.
3

M.G. Pirrone, et al.
CarbohydrateResearch491(2020)107984
Scheme 3. Differing Mechanisms and Regioselectivities in the Openings of Epoxides 12 and 16.
deoxygenation nor substitution of the C–O bond at the 4-position by a
C–C bond causes a significant change in the relative distribution of the
side chain conformation between the three staggered conformations.
Consequently, the increased activity of propylamycin in the inhibition
of bacterial ribosomes and the inhibition of bacterial growth does not
derive from a reduction in the entropic penalty paid on binding to the
ribosome with a fixed gt side chain conformation. As suggested pre-
viously [1] any reduction in binding energy to the ribosome on deox-
ygenation at the 4′-position of paromomycin, due to the loss of a hy-
drogen bond to a backbone phosphate, is offset by the increased
basicity of the ring oxygen (O5′) and the concomitant increase in the
critical hydrogen bond with A1408 in the binding pocket. The differ-
ence in activity between the 4′-deoxy derivative 3 and propylamycin 1
with the 4′-deoxy-4′-propyl substitution pattern is therefore presumably
related to hydrophobicity and/or solvation.
Scheme 4. Anti-Fürst-Plattner Regioselectivity in the Opening of an Epoxide en
route to Bradyrhizose.
Table 1
3
3
Experimental chemical shifts and JH5,H6R and JH5,H6S coupling constants.
4. Experimental
Ring I
δ H5
δ H6R
δ H6S
3JH5,H6R (Hz) 3JH5,H6S (Hz)
Model
(ppm)
(ppm)
(ppm)
4.1. General experimental
6
3.53
3.79
3.62
3.59
3.43
3.54
3.77
3.51
3.66
4.9
5.8
5.3
2.2
2.5
2.2
All reagents were purchased from commercial sources and used
4α
5
without
further
purification
unless
otherwise
specified.
Chromatographic purifications were carried out in Fisher 230–400
mesh silica gel unless otherwise specified. Microwave reactions were
conducted with a Biotage Initiator microwave reactor. High resolution
mass spectra were collected on a Waters LCT Premier XE ESI-TOF mass
spectrometer. Optical rotations were measured using an automated
polarimeter in a 1 dm cell. NMR spectra were collected at 400, 500, or
600 MHz as indicated. 1H NMR spectra were assigned with the aid of 1D
and 2D techniques including COSY, HSQC, HMBC, and TOCSY.
Methyl 2-amino-2,4-dideoxy-α-D-xylo-hexopyranoside acetate
salt (4α). Compound 7 (93.8 mg, 0.30 mmol) was dissolved in a 1:1
mixture of 1,4-dioxane and 10% aqueous AcOH (0.8 mL) followed by
addition of Pd/C (20.8 mg). The reaction mixture was stirred under 50
psi H2 for 5 h followed by filtration over Celite® and lyophilization to
Table 2
Limiting coupling constants for the ideal staggered conformers.
gg
gt
tg
3JR (Hz)
3JS (Hz)
1.0
2.2
11.0
2.5
4.8
10.2
Table 3
Side chain populations of the model compounds.
fgg
fgt
ftg
23
obtain 4 as an off white solid (69.1 mg, 97%). [α]D = 110.6
6
4
5
62
51
58
40
47
44
−2
2
(c = 0.14, water), 1H NMR (600 MHz, D2O) δ 4.86 (d, J = 3.7 Hz, 1H,
H1), 3.91 (td, J = 11.0, 5.1 Hz, 1H, H3), 3.79 (ddt, J = 11.8, 5.8,
2.5 Hz, 1H, H5), 3.51 (dd, J = 12.1, 3.1 Hz, 1H, H6), 3.43 (dd,
J = 12.1, 6.2 Hz, 1H, H6’), 3.24 (s, 3H, OMe), 3.02 (dd, J = 10.5,
3.6 Hz, 1H, H2), 1.86 (ddd, J = 12.8, 5.1, 2.1 Hz, 1H, H4eq), 1.76 (s,
3H, AcOH), 1.33 (q, J = 12.0 Hz, 1H, H4ax). 13C NMR (151 MHz, D2O)
δ 180.4 (AcOH), 96.6 (C1), 68.8 (C5), 64.3 (C3), 63.3 (C6), 55.2 (C2),
55.0 (OMe), 34.2 (C4), 22.6 (AcOH). ESI-HRMS: m/z calcd for
C7H15NO4Na [M + Na]+ 200.0899, found 200.0891.
−2
influence of the 4-OH group on the side chain conformation of 6,
whether derived from dipolar interactions, hydrogen bonding, and/or
solvation effects, is qualitatively reproduced by the propyl group in 5.
3. Conclusion
1,6-Anhydro-4-deoxy-2-O-p-toluenesulfonyl-6S-deuterio-D-glu-
copyranose (11). NaBH4 (0.43 g, 11.4 mmol) and BF3·OEt2 (0.85 mL,
Analysis of the model compounds 6, 4α, and 5 indicates that neither
4

M.G. Pirrone, et al.
CarbohydrateResearch491(2020)107984
6.9 mmol) were added to an ice cold solution of 10 (0.84 g, 2.8 mmol)
in 1,2-dimethoxyethane (9.4 mL). After 5 h the reaction mixture was
diluted with Et2O and washed with saturated NaHCO3 solution and
brine then dried with Na2SO4 and concentrated under vacuum to give
11 as a clear gum (0.85 g, 2.8 mmol, 99%) which was used in the next
vacuum to give 6S–2H-4 (5.4 mg, 0.023 mmol, 76%) as a mixture of
anomers in a ratio of 2:1 α/β as the acetate salts. ESI-HRMS: m/z calcd
for C7H14DNO4Na [M + Na]+ 201.1962, found 201.1956. 6S–2H-4α:
1H NMR (600 MHz, D2O) δ 4.87 (d, J = 3.6 Hz, 1H, H-1), 3.92 (td,
J = 11.0, 5.0 Hz, 1H, H-3), 3.80 (ddd, J = 12.1, 6.3, 2.2 Hz, 1H, H-5),
3.42 (d, J = 6.0 Hz, 1H, H-6), 3.26 (s, 3H, –OCH3), 3.03 (dd, J = 10.4,
3.7 Hz, 1H, H-2), 1.87 (ddd, J = 12.1, 5.0, 2.1 Hz, 1H, H-4eq), 1.78 (s,
3H, AcOH), 1.34 (q, J = 12.1 Hz, 1H, H-4ax). 13C NMR (151 MHz, D2O)
δ 180.2 (AcOH), 96.6 (C-1), 68.7 (C-5), 64.3 (C-3), 63.2–62.8 (m, C-6),
55.2 (C-2), 55.0 (-OCH3), 34.2 (C-4), 22.5 (AcOH). 6S–2H-4β: 1H NMR
(600 MHz, D2O) δ 4.37 (d, J = 8.4 Hz, 1H, H-1), 3.76 (dt, J = 10.9,
5.4 Hz, 1H, H-3), 3.57 (ddd, J = 11.8, 6.7, 2.1 Hz, 1H, H-5), 3.45 (d,
J = 6.6 Hz, 1H, H-6), 3.41 (s, 3H, –OCH3), 2.69 (dd, J = 10.3, 8.4 Hz,
1H, H-2), 1.91 (ddd, J = 12.9, 5.2, 2.0 Hz, 1H, H-4eq), 1.78 (s, 3H,
AcOH), 1.31 (dt, J = 12.9, 11.5 Hz, 1H, H-4ax). 13C NMR (151 MHz,
D2O) δ 180.2 (AcOH), 100.1 (C-1), 72.7 (C-5), 66.8 (C-3), 63.1–62.6
(m, C-6), 57.4 (C-2), 57.2 (-OCH3), 34.4 (C-4), 22.5 (AcOH).
step without purification. [α]D = −27.0 (c = 1.0, CHCl3), 1H NMR
23
(600 MHz, CDCl3) δ 7.81 (d, J = 8.3 Hz, 2H, Ar–H), 7.36–7.33 (m, 2H,
Ar–H), 5.27 (d, J = 1.9 Hz, 1H, H-1), 4.52 (d, J = 4.1 Hz, 1H, H-5),
4.20 (q, J = 1.7 Hz, 1H, H-2), 4.09 (s, 1H, H-6), 3.92 (t, J = 6.3 Hz, 1H,
H-3), 2.46 (d, J = 6.3 Hz, 1H, –OH), 2.44 (s, 3H, Ar-CH3), 2.31 (ddd,
J = 15.0, 5.7, 4.5 Hz, 1H, H-4ax), 1.70 (ddt, J = 15.0, 1.9, 1.0 Hz, 1H,
H-4eq). 13C NMR (151 MHz, CDCl3) δ 145.3, 133.2, 130.1, 127.9 (Ar),
99.3 (C-1), 76.8 (C-2), 71.3 (C-5), 67.5 (t, J = 23.3 Hz, C-6), 66.5 (C-3),
32.5 (C-4), 21.7 (Ar-CH3). ESI-HRMS: m/z calcd for C13H15DO6SNa [M
+ Na]+ 324.0628, found 324.0627.
1,6;2,3-Bisanhydro-4-deoxy-6S-deuterio-D-lyxopyranose (12).
NaOMe (0.34 g, 6.3 mmol) was added to an ice cold stirred solution of
11 (0.824 g, 2.73 mmol) in 1:1 CHCl3/MeOH (13.6 mL). After 4 h the
reaction mixture was diluted with CH2Cl2 and washed with water. The
aqueous phase was back extracted with CH2Cl2 and the combined or-
ganic layers were washed with brine then dried with Na2SO4 and
concentrated under vacuum to give 12 as a colorless oil (0.351 g, 99%).
[α]D23 = −28.6 (c = 1.0, CH2CL2), 1H NMR (400 MHz, CDCl3) δ 5.64
(dd, J = 3.2, 1.0 Hz, 1H, H-1), 4.38 (br d, J = 6.0 Hz, 1H, H-5), 3.63 (d,
J = 1.7 Hz, 1H, H-6), 3.33 (t, J = 3.6 Hz, 1H, H-2), 3.10 (dd,
J = 3.3 Hz, 1H, H-3), 2.19 (ddd, J = 15.3, 5.9, 3.2 Hz, 1H, H-4ax), 1.94
(d, J = 15.3 Hz, 1H, H-4eq). 13C NMR (101 MHz, CDCl3) δ 98.0 (C-1),
68.0 (t, J = 23.1 Hz, C-6), 67.2 (C-5), 53.7 (C-2), 46.3 (C-3), 30.0 (C-4).
ESI-HRMS: m/z calcd for C6H7DO3Na [M + Na]+ 152.0434, found
152.0438.
4-C-Allyl-1,6-anhydro-2-N-benzyl-2,4-dideoxy-D-glucopyranose
(17). A solution of 16 (0.319 g, 1.90 mmol) in benzylamine (5.0 mL)
was stirred for 3 days at 155 °C. After concentration under reduced
pressure the residue was purified by flash column chromatography on
silica gel eluting with 40% ethyl acetate and 1% triethylamine in
hexanes to afford 17 (0.402 g, 77%). [α]23D = −49.4 (c = 1.00,
CHCl3). 1H NMR (500 MHz, CDCl3) δ 7.38–7.23 (m, 5H: aromatic), 5.83
(ddq, J = 16.4, 14.4, 6.2, 5.3 Hz, 1H: CH2CHCH2–C4), 5.47 (s, 1H: H1),
5.21–5.10 (m, 2H: CH2CHCH2–C4), 4.40 (d, J = 5.0 Hz, 1H: H5), 4.06
(d, J = 7.0 Hz, 1H: H6a), 3.93–3.85 (m, 5H: PhCH2), 3.80–3.72 (m, 1H,
H6b), 3.65 (td, J = 2.9, 1.6 Hz, 1H: H3), 2.68–2.63 (m, 1H: H2),
2.49–2.33 (m, 2H: CH2CHCH2–C4), 1.77 (t, J = 7.2 Hz, 1H: H4). 13C
NMR (125 MHz, CDCl3) δ 139.9, 136.1 (CH2CHCH2–C4), 128.5, 128.1,
127.2, 117.5 (CH2CHCH2–C4), 102.6 (C1), 74.6 (C5), 70.3 (C3), 68.5
(C6), 62.2 (C2), 51.7 (PhCH2), 44.6 (C4), 36.5 (CH2CHCH2–C4). ESI-
1,6-Anhydro-2-azido-2,4-dideoxy-D-xylo-hexopyranose
(14).
Compound 12 (75.0 mg, 0.58 mmol), LiN3 (145 mg, 2.96 mmol),
benzoic acid (109 mg, 0.89 mmol), and DMF (2.0 mL) were added to a
microwave vial and heated to 110 °C for 75 min in a microwave reactor.
The reaction mixture was then diluted with CH2Cl2 and washed with
aqueous saturated NaHCO3 solution and brine. The organic layer was
dried with Na2SO4 and filtered followed by concentration. The residue
was subjected to silica gel preparative HPLC eluting with a gradient
from 20% to 60% EtOAc in hexanes to give 14 (4.7 mg, 0.027 mmol,
5%). [α]D23 = 19.6 (c = 0.2, CHCl3), 1H NMR (600 MHz, CDCl3) δ 5.53
(t, J = 2.0 Hz, 1H, H-1), 4.59 (d, J = 4.4 Hz, 1H, H-5), 4.19 (s, 1H, H-
6), 4.00–3.96 (m, 1H, H-3), 3.30 (d, J = 2.2 Hz, 1H, H-2), 2.55 (d,
J = 7.7 Hz, 1H, –OH), 2.31 (dt, J = 15.2, 4.9 Hz, 1H, H-4ax), 1.84 (ddt,
J = 15.3, 2.9, 1.6 Hz, 1H, H-4eq). 13C NMR (151 MHz, CDCl3) δ 100.8
(C-1), 72.0 (C-5), 67.6 (t, J = 23.3 Hz, C-6), 66.9 (C-3), 61.7 (C-2), 33.6
(C-4).
HRMS: m/z calcd for
C
16H22NO3Na [M+Na]+ 276.1600, found
276.1600.
2-Amino-1,6-anhydro-2,4-dideoxy-4-C-propyl-D-glucopyranose
acetate salt (18). Compound 17 (0.4020 g, 1.46 mmol) and Pd/C
(0.1316 g) were stirred in a 1:2 mixture of 10% aqueous acetic acid and
1,4-dioxane (6 mL) under 40 psi H2 for 7 h. Pd/C (0.23 g) was added
after 7 h and the reaction mixture was stirred for an additional 14 h
before a final addition of Pd/C (0.39 g). After an additional 28 h the
reaction mixture was filtered through Celite® and concentrated to give
23
18 (0.286 g, 79%). [α]D = −45.9 (c = 1.0, MeOH), 1H NMR
(600 MHz, CD3OD) δ 5.32 (s, 1H, H-1), 4.41 (d, J = 5.2 Hz, 1H, H-5),
3.98 (d, J = 6.8 Hz, 1H, H-6a), 3.68 (dd, J = 6.9, 5.3 Hz, 1H, H-6b),
3.43 (t, J = 4.0 Hz, 1H, H-3), 2.82 (d, J = 4.0 Hz, 1H, H-2), 1.92 (s, 3H,
AcOH), 1.64–1.47 (m, 4H, H-4, –CH2CH2-), 1.45–1.34 (m, 1H,
–CH2CH2-), 0.97 (t, J = 7.3 Hz, 3H, –CH3). 13C NMR (151 MHz,
CD3OD) δ 104.9 (C-1), 79.1 (C-5), 74.6 (C-6), 72.6 (C-3), 60.7 (C-2),
48.8 (C-4), 37.9 (-CH2CH2CH3), 25.4 (AcOH), 23.9 (-CH2CH2CH3), 16.9
Methyl 2-amino-2,4-dideoxy-6S-deuterio-D-xylo-hexopyrano-
side acetate salt (6S–2H-4). TFA (0.05 mL) was added to a stirred
solution of 14 (4.6 mg, 0.27 mmol) in Ac2O (0.5 mL) at 0 °C and the
reaction mixture was stirred for 3 h with monitoring by TLC. The re-
action mixture was then diluted with Et2O and washed with aqueous
saturated NaHCO3 solution and brine, dried with Na2SO4, and con-
centrated to give an inseparable mixture of anomers (8.3 mg,
0.026 mmol, 97%), which were used in the next step without pur-
ification. ESI-HRMS: m/z calcd for C12H16DN3O7Na [M + Na]+
339.1014, found 339.1027. The mixture of anomers from the previous
step (8.3 mg, 0.026 mmol) were stirred in a 10% HCl methanol solution
(0.6 mL) under reflux for 3 h. After the reaction was complete by TLC
and LCMS, the reaction mixture was concentrated under vacuum and
the resulting inseparable mixture of anomers of methyl glycosides was
used in the next step without purification. ESI-HRMS: m/z calcd for
C7H12DN3O4Na [M + Na]+ 227.0867, found 227.0864. 10 wt% Pd/C
(5.0 mg) was added to a solution of the methyl glycosides (6.1 mg,
0.30 mmol) in 1:1 dioxane/10% aqueous acetic acid (0.4 mL) and the
reaction mixture was stirred under 50 psi H2 for 6 h. The reaction
mixture was then filtered through Celite® and concentrated under
(-CH2CH2CH3). ESI-HRMS: m/z calcd for C9H18NO3 [M
+
H]+
188.1287, found 188.1286.
1,6-Anhydro-2-azido-2,4-dideoxy-4-C-propyl-D-glucopyranose
(19). Stick's reagent (0.466 g, 2.23 mmol) was added to an ice cold
stirred solution of CuSO4 (0.024 g, 0.15 mmol), triethylamine (0.62 mL,
4.5 mmol), and 18 (0.278 g, 1.12 mmol) in 4:1 MeCN/water (14.8 mL).
The reaction mixture was stirred for 1 h before MeCN was removed
under vacuum and the residue was diluted with EtOAc, washed with
1 N HCl, saturated NaHCO3 solution, and brine. The organic layer was
dried with Na2SO4 and concentrated to give 19 as a colorless gum
(0.219 g, 91%). [α]D23 = −113.2 (c = 1.0, CHCl3) 1H NMR (500 MHz,
CDCl3) δ 5.46 (t, J = 1.4 Hz, 1H, H-1), 4.43 (d, J = 5.1 Hz, 1H, H-5),
4.10 (dd, J = 7.0, 0.8 Hz, 1H, H-6a), 3.78 (dd, J = 7.1, 5.1 Hz, 1H, H-
6b), 3.65 (tt, J = 2.4, 1.2 Hz, 1H, H-3), 3.47 (t, J = 2.0 Hz, 1H, H-2),
1.72–1.56 (m, 3H, H-4, -CH2CH2CH3), 1.55–1.45 (m, 1H,
-CH2CH2CH3), 1.44–1.34 (m, 1H, -CH2CH2CH3), 0.97 (t, J = 7.2 Hz,
3H, –CH2CH2CH3). 13C NMR (126 MHz, CDCl3) δ 100.6 (C-1), 75.0 (C-
5

M.G. Pirrone, et al.
CarbohydrateResearch491(2020)107984
5), 71.2 (C-3), 68.6 (C-6), 63.6 (C-2), 44.3 (C-4), 33.3 (CH3CH2CH2-),
20.5 (CH3CH2CH2-), 14.0 (CH3CH2CH2-).
68.0 (t, J = 23.4 Hz, C-6), 43.0 (C-4), 35.4 (CH2CHCH2-), 21.7
(ArCH3). ESI-HRMS: m/z calcd for C16H19DO6Na [M + Na]+ 364.0941,
found 364.0936.
Methyl 2-azido-2,4-dideoxy-4-C-propyl-D-glucopyranoside (20α
and 20β). Compound 19 (0.219 g, 1.00 mmol) was dissolved in Ac2O
(8.6 mL) followed by addition of TFA (0.86 mL) and stirred under argon
for 45 min. The reaction mixture was then diluted with Et2O and wa-
shed with saturated NaHCO3 solution and brine, dried with Na2SO4,
filtered, and concentrated under vacuum. The residue was then dis-
solved in 10% HCl MeOH solution (7 mL) and heated to reflux for 4.5 h
followed by concentration under vacuum to give a 1.5:1 mixture of 20α
and 20β (0.174 g, 69%). The resulting residue was subjected to flash
column chromatography over silica gel in 45%–50% ethyl acetate in
hexanes, which afforded 20α (55 mg, 22%) and 20β (31 mg, 12%).
4-C-Allyl-1,6;2,3-bisanhydro-6S-deuterio-4-deoxy-β-D-manno-
pyranose (6S–2H-16). NaOMe (0.088 g, 1.63 mmol) was added to an
ice-cold stirred solution of compound 6S–2H-15 (0.240 g, 0.70 mmol)
in 1:1 mixture of methanol and chloroform (3.5 mL) and the solution
was allowed to warm to room temperature. After 2 h the reaction
mixture was diluted with Et2O and washed with aqueous saturated
NH4Cl solution and brine. The organic layer was dried with Na2SO4 and
concentrated to give 6S–2H-16 (0.118 g, 99%) as a white waxy solid
which was used without further purification. [α]D23 = −15.2 (c = 1.0,
CH2Cl2), 1H NMR (600 MHz, CDCl3) δ 5.82 (ddt, J = 17.2, 10.2, 7.0 Hz,
1H, CH2CHCH2-), 5.65 (d, J = 3.2, 1H, H-1), 5.16–5.10 (m, 2H,
CH2CHCH2-), 4.23 (s, 1H, H-5), 3.70 (d, J = 1.6 Hz, 1H, H-6), 3.34
(ddd, J = 3.8, 3.1, 0.7 Hz, 1H, H-2), 2.93 (dd, J = 4.0, 1.3 Hz, 1H, H-
3), 2.32 (tt, J = 7.2, 1.3 Hz, 2H, CH2CHCH2-), 2.01 (t, J = 7.6 Hz, 1H,
H-4). 13C NMR (151 MHz, CDCl3) δ 135.2 (CH2CHCH2-), 117.8
(CH2CHCH2-), 98.0 (C-1), 70.9 (C-5), 68.2 (t, J = 23.0 Hz, C-6), 53.8
(C-2), 50.4 (C-3), 39.0 (C-4), 35.1 (CH2CHCH2-). ESI-HRMS: m/z calcd
for C9H11DO3Na [M + Na]+ 192.0747, found 192.0746.
20α: [α]D = 125.4 (c = 1.0, MeOH), 1H NMR (600 MHz, CD3OD) δ
23
4.76 (d, J = 3.5 Hz, 1H, H1), 3.78 (t, J = 10.2 Hz, 1H, H3), 3.74–3.68
(m, 1H, H6), 3.62–3.54 (m, 2H, H5, H6), 3.37 (s, 3H, OMe), 3.10 (dd,
J = 10.0, 3.5 Hz, 1H, H2), 1.62–1.50 (m, 2H, H4, –CH2CH2-),
1.49–1.28 (m, 3H, –CH2CH2-), 0.91 (t, J = 7.1 Hz, 3H, CH3). 13C NMR
(151 MHz, CD3OD) δ 99.2 (C1), 71.8 (C5), 68.5 (C3), 65.2 (C2), 61.8
(C6), 53.9 (OMe), 43.1 (C4), 28.8 (–CH2CH2–), 18.7 (–CH2CH2–), 13.7
(-CH3). ESI-HRMS: m/z calcd for C10H19N3O4Na [M + Na]+ 268.1273,
23
found 268.1273. 20β: [α]D = −31.8 (c = 1.0, MeOH), 1H NMR
4-C-Allyl-1,6-anhydro-6S-deuterio-2-N-benzyl-2,4-dideoxy-D-
glucopyranose (6S–2H-17). A stirred solution of epoxide 6S–2H-17
(0.118 g, 0.427 mmol) in benzylamine (5.0 mL) was heated to 155 °C
for 1.5 days before concentration under vacuum. The residue was then
purified using silica gel column chromatography in 40% EtOAc and 1%
triethylamine in hexanes to give amine 6S–2H-17 (0.166 g, 86%).
(600 MHz, CD3OD) δ 4.13 (d, J = 8.1 Hz, 1H, H1), 3.77 (dd, J = 12.1,
2.1 Hz, 1H, H6), 3.59 (dd, J = 12.1, 5.5 Hz, 1H, H6), 3.52 (s, 3H, OMe),
3.32–3.24 (m, 2H, H3, H5), 3.03 (dd, J = 9.5, 8.1 Hz, 1H, H2),
1.58–1.25 (m, 5H, H4, –CH2CH2-), 0.90 (t, J = 7.2 Hz, 3H, –CH3). 13C
NMR (151 MHz, CD3OD) δ 102.7 (C1), 76.2 (C5), 72.6 (C3), 68.6 (C2),
61.8 (C6), 55.6 (OMe), 42.6 (C4), 28.7 (–CH2CH2–), 18.6 (–CH2CH2–),
13.7 (-CH3). ESI-HRMS: m/z calcd for C10H19N3O4Na [M + Na]+
268.1273, found 268.1261.
23
[α]D = −32.9 (c = 1.0, CH2CL2), 1H NMR (400 MHz, CDCl3) δ
7.36–7.30 (m, 4H, Ar–H), 7.29–7.23 (m, 1H, Ar–H), 5.88–5.76 (m, 1H,
CH2CHCH2-), 5.46 (s, 1H, H-1), 5.16–5.10 (m, 2H, CH2CHCH2-), 4.38
(s, 1H, H-5), 4.03 (s, 1H, H-6), 3.90 (d, J = 13.2 Hz, 1H, PhCH2N-),
3.86 (d, J = 13.2 Hz, 1H, PhCH2N-), 3.64 (dq, J = 3.0, 1.4 Hz, 1H, H-
3), 2.64 (p, J = 1.2 Hz, 1H, H-2), 2.50–2.30 (m, 2H, CH2CHCH2-), 1.76
(t, J = 7.7 Hz, 1H, H-4). 13C NMR (101 MHz, CDCl3) δ 139.9 (Ar),
136.1 (CH2CHCH2-), 128.5 (Ar), 128.1 (Ar), 127.2 (Ar), 117.5
(CH2CHCH2-), 102.6 (C-1), 74.6 (C-5), 70.3 (C-3), 68.2 (t, J = 23.4 Hz,
C-6), 62.2 (C-2), 51.7 (PhCH2N-), 44.6 (C-4), 36.5 (CH2CHCH2-). ESI-
HRMS: m/z calcd for C16H21DNO3 [M + H]+ 277.1657, found
277.1664.
Methyl 2-amino-2,4-dideoxy-4-C-propyl-α-D-glucopyranoside
(5). Compound 20α (13.6 mg, 0.30 mmol) was dissolved in a 1:1
mixture of 1,4-dioxane and 10% aqueous AcOH (0.6 mL) followed by
addition of Pd/C (2.9 mg). The reaction mixture was stirred under 50
psi H2 for 1.5 h followed by filtration over Celite® and lyophilization to
obtain 5 as an off white solid (15.4 mg, 99%). [α]D23 = 80.5 (c = 0.7,
water), 1H NMR (600 MHz, D2O) δ 4.84 (d, J = 3.6 Hz, 1H, H1), 3.70
(t, J = 10.6 Hz, 1H, H3), 3.66 (dd, J = 12.3, 2.2 Hz, 1H, H6), 3.62
(ddd, J = 10.9, 5.3, 2.2 Hz, 1H, H5), 3.54 (dd, J = 12.3, 5.3 Hz, 1H,
H6), 3.24 (s, 3H, OMe), 3.07 (dd, J = 10.3, 3.6 Hz, 1H, H2), 1.79 (s,
3H, AcOH), 1.49 (tt, J = 10.8, 4.0 Hz, 1H, H4), 1.41–1.32 (m, 1H,
–CH2CH2-), 1.32–1.24 (m, 1H, –CH2CH2-), 1.23–1.06 (m, 2H, –CH2CH2-
), 0.71 (t, J = 7.2 Hz, 3H, –CH3). 13C NMR (151 MHz, D2O) δ 96.3 (C1),
71.6 (H5), 67.0 (C3), 61.1 (6), 55.3 (C2), 54.9 (OMe), 42.0 (C4), 27.7
(–CH2CH2–), 17.8 (–CH2CH2–), 13.8 (-CH3). ESI-HRMS: m/z calcd for
2-Amino-1,6-anhydro-6S-deuterio-2,4-dideoxy-4-C-propyl-D-
glucopyranose acetate salt (6S–2H-18). Pd/C (10% w/w, 27 mg) was
added to a solution of 6S–2H-17 (0.137 g, 0.50 mmol) in a 1:1 mixture
of 10% aqueous acetic acid and 1,4-dioxane (0.6 mL) followed by
pressurization to 40 psi of H2. The reaction mixture was stirred vigor-
ously for 9 h before filtration through Celite® and concentration under
vacuum to give amine 6S–2H-18 (0.122 g, 99%) as the acetate salt,
which was used without further purification. [α]D23 = −48.6 (c = 4.0,
MeOH), 1H NMR (600 MHz, D2O) δ 5.41 (s, 1H, H-1), 4.45 (s, 1H, H-5),
3.87 (s, 1H, H-6), 3.47 (t, J = 4.6 Hz, 1H, H-3), 3.01 (dd, J = 4.4,
1.0 Hz, 1H, H-2), 1.75 (s, 3H, AcOH), 1.53 (tdd, J = 6.8, 4.6, 1.5 Hz,
1H, H-4), 1.43–1.37 (m, 2H, CH3CH2CH2-), 1.32 (dp, J = 13.3, 7.2 Hz,
1H, CH3CH2CH2-), 1.21 (tdd, J = 15.1, 13.4, 7.1 Hz, 1H, CH3CH2CH2-),
0.76 (t, J = 7.3 Hz, 3H, CH3CH2CH2-). 13C NMR (151 MHz, D2O) δ 98.5
(C-1), 75.3 (C-5), 68.7 (t, J = 23.5 Hz, C-6), 68.3 (C-3), 55.6 (C-2), 43.6
(C-4), 33.0 (CH3CH2CH2-), 23.0 (AcOH), 19.3 (CH3CH2CH2-), 13.0
(CH3CH2CH2-). ESI-HRMS: m/z calcd for C9H17DNO3 [M + H]+
189.1349, found 189.1357.
C
10H22NO4 [M + H]+ 220.1549, found 220.1539.
4-C-Allyl-1,6-Anhydro-6S-deuterio-2,4-dideoxy-2-O-p-toluene-
sulfonyl-D-glucopyranose (6S–2H-15). Freshly prepared 0.5
M
allylMgCl THF solution (16 mL) was added to an ice-cold stirred solu-
tion of epoxide 6S–2H-10 (0.590 g 1.97 mmol) and CuI (0.38 g,
0.20 mmol) under argon in THF (20 mL). The reaction mixture was
stirred for 11 h followed by addition of further allylMgCl solution
(8 mL). After another 17 h the reaction mixture was concentrated under
vacuum then diluted with Et2O and washed with aqueous saturated
NH4Cl solution and brine. The organic layer was dried with Na2SO4 and
concentrated. The residue was then purified using silica gel flash
column chromatography in 35–40% EtOAc in hexanes to give 6S–2H-15
(0.24 g, 36%). [α]D23 = −51.3 (c = 1.0, CH2Cl2), 1H NMR (600 MHz,
CDCl3) δ 7.83–7.78 (m, 2H, Ar–H), 7.37–7.33 (m, 2H, Ar–H), 5.74 (ddt,
J = 16.3, 10.5, 7.1 Hz, 1H, CH2CHCH2-), 5.27 (d, J = 1.5 Hz, 1H, H-1),
5.13 (dq, J = 6.1, 1.2 Hz, 1H, CH2CHCH2-), 5.10 (t, J = 1.3 Hz, 1H,
CH2CHCH2-), 4.39 (s, 1H, H-5), 4.18 (dt, J = 2.5, 1.2 Hz, 1H, H-2),
3.99 (s, 1H, H-6), 3.69 (tt, J = 2.7, 1.2 Hz, 1H, H-3), 2.45 (s, 3H,
ArCH3), 2.37–2.33 (m, 2H, CH2CHCH2-), 1.68 (t, J = 7.7 Hz, 1H, H-4).
13C NMR (151 MHz, CDCl3) δ 135.3 (CH2CHCH2-), 130.0 (Ar), 127.9
(Ar), 118.0 (CH2CHCH2-), 99.7 (C-1), 78.9 (C-2), 74.2 (C-5), 70.0 (C-3),
1,6-Anhydro-2-azido-6S-deuterio-2,4-dideoxy-4-C-propyl-D-glu-
copyranose (6S–2H-19). Stick's reagent (0.209 g, 1.00 mmol) was
added to an ice cold stirred solution of CuSO4 (11 mg, 0.07 mmol),
triethylamine (0.28 mL, 2.0 mmol), and compound 6S–2H-18 (0.123 g,
0.495 mmol) in 4:1 MeCN/water (6.6 mL). The reaction mixture was
stirred for 9 h before MeCN was removed under vacuum and the residue
was diluted with EtOAc, washed with 1 N HCl, saturated NaHCO3 so-
lution, and brine. The organic layer was dried with Na2SO4 and con-
centrated to give 6S–2H-19 as a colorless gum (0.101 g, 96%).
6

M.G. Pirrone, et al.
CarbohydrateResearch491(2020)107984
23
[α]D = −40.5 (c = 1.0, CH2Cl2), 1H NMR (600 MHz, CDCl3) δ 5.41
Acknowledgments
(s, 1H, H-1), 4.38 (s, 1H, H-5), 4.04 (s, 1H, H-6), 3.60 (s, 1H, H-3), 3.42
(s, 1H, H-2), 2.71 (br s, 1H, –OH), 1.66–1.51 (m, 3H, H-4, CH3CH2CH2-
), 1.50–1.41 (m, 1H, CH3CH2CH2-), 1.40–1.30 (m, 1H, CH3CH2CH2-),
0.93 (t, J = 7.3 Hz, 3H, CH3CH2CH2-). 13C NMR (151 MHz, CDCl3) δ
100.5 (C-1), 74.9 (C-5), 71.1 (C-3), 68.2 (t, J = 23.2 Hz, C-6), 63.7 (C-
2), 44.2 (C-4), 33.3 (CH3CH2CH2-), 20.5 (CH3CH2CH2-), 14.0
(CH3CH2CH2-).
We thank the NIH (AI123352) for support of this work.
Appendix A. Supplementary data
Supplementary data to this article can be found online at https://
doi.org/10.1016/j.carres.2020.107984.
Methyl
2-azido-6S-deuterio-2,4-dideoxy-4-C-propyl-D-gluco-
pyranoside (6S–2H-20α and 6S–2H-20β). Compound 6S–2H-19
(0.091 g, 0.42 mmol) was dissolved in Ac2O (4 mL) followed by addi-
tion of TFA (0.40 mL) and stirred under argon for 2 h. The reaction
mixture was then diluted with Et2O and washed with saturated NaHCO3
solution and brine, dried with Na2SO4, filtered, and concentrated. The
residue was then dissolved in 10% HCl MeOH solution (3.4 mL) and
heated to reflux for 10 h followed by concentration under vacuum to
give a 2:1 mixture of 6S–2H-20α and 6S–2H-20β (80.8 mg, 77%). The
resulting residue was subjected flash column chromatography over si-
lica gel in 45%–50% ethyl acetate in hexanes which afforded 6S–2H-
20α (11.7 mg, 11%), 6S–2H-20β (9.5 mg, 9%), and (11.8 mg, 11%) of a
mixture of anomers. ESI-HRMS: m/z calcd for C10H18DN3O4Na [M +
Na]+ 269.1336, found 269.1326. 6S–2H-20α: [α]D23 = 98.8 (c = 1.0,
MeOH), 1H NMR (600 MHz, CD3OD) δ 4.76 (d, J = 3.5 Hz, 1H, H-1),
3.78 (t, J = 10.2 Hz, 1H, H-3), 3.58 (dd, J = 10.6, 5.4 Hz, 1H, H-5),
3.55 (d, J = 5.5 Hz, 1H, H-6), 3.37 (s, 3H, -OMe), 3.09 (dd, J = 10.1,
3.5 Hz, 1H, H-2), 1.61–1.49 (m, 2H, –CH2CH2-, H-4), 1.49–1.27 (m, 3H,
–CH2CH2-), 0.91 (t, J = 7.2 Hz, 3H, –CH3). 13C NMR (151 MHz,
CD3OD) δ 99.2 (C-1), 71.8 (C-5), 68.5 (C-3), 65.2 (C-2), 61.5 (t,
J = 21.3 Hz, C-6), 53.9 (-OMe), 43.1 (C-4), 28.8 (–CH2CH2–), 18.7
(–CH2CH2–), 13.6 (-CH3). ESI-HRMS: m/z calcd for C10H18DN3O4Na [M
References
[1] T. Matsushita, G.C. Sati, N. Kondasinghe, M.G. Pirrone, T. Kato, P. Waduge,
H.S. Kumar, A. Cortes Sanchon, M. Dobosz-Bartoszek, D. Shcherbakov, M. Juhas,
S.N. Hobbie, T. Schrepfer, C.S. Chow, Y.S. Polikanov, J. Schacht, A. Vasella,
E.C. Böttger, D. Crich, Design, multigram synthesis, and in vitro and in vivo eva-
luation of propylamycin: a semisynthetic 4,5-deoxystreptamine class aminoglyco-
side for the treatment of drug-resistant enterobacteriaceae and other Gram-negative
pathogens, J. Am. Chem. Soc. 141 (2019) 5051–5061.
[2] S. Duscha, H. Boukari, D. Shcherbakov, S. Salian, S. Silva, A. Kendall, T. Kato,
R. Akbergenov, D. Perez-Fernandez, B. Bernet, S. Vaddi, P. Thommes, J. Schacht,
D. Crich, A. Vasella, E.C. Böttger, Identification and evaluation of improved 4’-O-
(alkyl) 4,5-disubstituted 2-deoxystreptamines as next generation aminoglycoside
antibiotics, mBio 5 (2014), https://doi.org/10.1128/mBio.01827-14.
[3] K. Bock, J.O. Duus, A conformational study of hydroxymethyl groups in carbohy-
drates investigated by 1H NMR spectroscopy, J. Carbohydr. Chem. 13 (1994)
513–543.
[4] T.B. Grindley, Structure and conformation of carbohydrates, in: B. Fraser-Reid,
K. Tatsuta, J. Thiem (Eds.), Glycoscience: Chemistry and Chemical Biology, vol. 1,
Springer, Berlin, 2001, pp. 3–51.
[5] V.S.R. Rao, P.K. Qasba, P.V. Balaji, R. Chandrasekaran, Conformation of
Carbohydrates, Harwood Academic Publishers, Amsterdam, 1998, p. 359.
[6] A.P. Carter, W.M. Clemons, D.E. Brodersen, R.J. Morgan-Warren, B.T. Wimberly,
V. Ramakrishnan, Functional insights from the structure of the 30S ribosomal
subunit and its interactions with antibiotics, Nature 407 (2000) 340–348.
[7] Q. Vicens, E. Westhof, Molecular recognition of aminoglycoside antibiotics by ri-
bosomal RNA and resistance enzymes: an analysis of X-ray crystal structures,
Biopolymers 70 (2003) 42–57.
+ Na]+ 269.1336, found 269.1334. 6S–2H-20β: [α]D = −35.4
23
(c = 0.003, MeOH), 1H NMR (600 MHz, CD3OD) δ 4.13 (d, J = 8.1 Hz,
1H, H-1), 3.57 (d, J = 5.5 Hz, 1H, H-6), 3.52 (s, 3H, -OMe), 3.30–3.24
(m, 2H, H-3, H-5), 3.02 (dd, J = 9.5, 8.1 Hz, 1H, H-2), 1.58–1.24 (m,
5H, H4, –CH2CH2-), 0.90 (t, J = 7.1 Hz, 3H, –CH3). 13C NMR (151 MHz,
CD3OD) δ 102.7 (C-1), 76.1 (C-5), 72.6 (C-3), 68.6 (C-2), 61.5 (t,
J = 21.9 Hz, C-6), 55.6 (-OMe), 42.6 (C-4), 28.6 (–CH2CH2–), 18.6
(–CH2CH2–), 13.6 (-CH3).
[8] A.R. Mandhapati, G. Yang, T. Kato, D. Shcherbakov, S.N. Hobbie, A. Vasella,
E.C. Böttger, D. Crich, Structure-based design and synthesis of apramycin-par-
omomycin analogues. Importance of the configuration at the 6′-position and dif-
ferences between the 6′-amino and hydroxy series, J. Am. Chem. Soc. 139 (2017)
14611–14619.
[9] T. Kato, A. Vasella, D. Crich, Stereospecific synthesis of methyl 2-amino-2-deoxy-
(6S)-Deuterio-α,β-D-Glucopyranoside and methyl 2,6-Diamino-2,6-Dideoxy-(6R)-
Deuterio-α,β-D-Glucopyranoside: side chain conformations of the 2-amino-2-deoxy
and 2,6-diamino-2,6-dideoxyglucopyranosides, Carbohydr. Res. 448 (2017) 10–17.
[10] C.E. Lünse, M.S. Schmidt, V. Wittmann, G. Mayer, Carba-sugars activate the GLMS-
riboswitch of Staphylococcus aureus, ACS Chem. Biol. 6 (2011) 675–678.
[11] H. Ohrui, H. Horiki, H. Kishi, H. Meguro, Synthesis of (6R)- and (6S)-D-glucose-6-2H
through stereospecific photo-bromination of 1,6-anhydro-β-D-glucopyranose deri-
vative, Agric. Biol. Chem. 47 (1983) 1101–1106.
Methyl 2-amino-6-(S)-deuterio-2,4-dideoxy-4-C-propyl-α-D-glu-
copyranoside (6S–2H-5). Compound 6S–2H-20α (9.0 mg,
0.037 mmol) was dissolved in a 1:1 mixture of 1,4-dioxane and 10%
aqueous AcOH (0.6 mL) followed by addition of Pd/C (1.8 mg). The
reaction mixture was stirred under 50 psi H2 for 1 h followed by fil-
tration over Celite® and lyophilization to obtain 6S–2H-5 as an off white
[12] Y. Ichikawa, H. Kuzuhara, Synthesis of 1,6-anhydro-2,3-di-O-benzoyl-4-O-(methyl
2,3,4-tri-O-benzoyl-α-L-iodpyranosyluronate)-β-D-glucopyranose from cellobiose,
Carbohydr. Res. 115 (1983) 117–129.
23
solid (10.2 mg, 99%). [α]D = 56.4 (c = 0.5, H2O), 1H NMR
[13] J.J. Maul, P.J. Ostrowski, G.A. Ublacker, B. Linclau, D.P. Curran, Benzotrifluoride
and derivatives: useful solvents for organic synthesis and fluorous synthesis, in:
P. Knochel (Ed.), Top. Curr. Chem. vol. 206, Springer, Berlin, 1999, pp. 79–105.
[14] T.B. Grindley, G.J. Reimer, J. Kralovec, R.G. Brown, M. Anderson, Syntheses of 3-
deoxy-3-substituted-D-glucose derivatives. Part I. Improvements in preparations of
and nucleophilic additions to 1,6:2,3-dianhydro-4-O-benzyl-β-D-allopyranose, Can.
J. Chem. 65 (1987) 1065–1071.
(600 MHz, D2O) δ 4.84 (d, J = 3.6 Hz, 1H, H1), 3.71 (t, J = 10.5 Hz,
1H, H3), 3.62 (dd, J = 10.9, 5.4 Hz, 1H, H5), 3.53 (d, J = 5.5 Hz, 1H,
H6), 3.25 (s, 3H, OMe), 3.08 (dd, J = 10.4, 3.6 Hz, 1H, H2), 1.80 (s,
3H, AcOH), 1.49 (tt, J = 10.8, 4.0 Hz, 1H, H4), 1.40–1.33 (m, 1H,
–CH2CH2-), 1.32–1.24 (m, 1H, –CH2CH2-), 1.24–1.06 (m, 2H, –CH2CH2-
), 0.72 (t, J = 7.2 Hz, 3H, –CH3). 13C NMR (151 MHz, D2O) δ 179.9
(AcOH), 96.3 (C1), 71.6 (C5), 67.0 (C3), 60.99 (t, J = 21.0 Hz, C6),
55.3 (C2), 54.9 (OMe), 42.0 (C4), 27.7 (–CH2CH2–), 22.3 (AcOH), 17.8
(–CH2CH2–), 13.8 (-CH3). ESI-HRMS: m/z calcd for C10H21DNO4 [M +
H]+ 221.1612, found 221.1605.
[15] J.D. White, G.L. Bolton, A.P. Dantanarayana, C.M.J. Fox, R.N. Hiner, R.W. Jackson,
K. Sakuma, U.S. Warrier, Total synthesis of the antiparasitic agent avermectin B1a,
J. Am. Chem. Soc. 117 (1995) 1908–1939.
[16] T.S. Rasmussen, H.H. Jensen, Synthesis and glycosidase inhibitory activity of
noeurostegine - a new and potent inhibitor of β-glucoside hydrolases, Org. Biomol.
Chem. 8 (2010) 433–441.
[17] T. Wakamatsu, H. Nakamura, Y. Nishikimi, K. Yoshida, T. Noda, M. Taniguchi,
Y. Ban, The oxirane ring openings of the dianhydro sugar with high regioselectivity
and its use in preparation of two chiral segments of 6-deoxyerythronolide B,
Tetrahedron Lett. 27 (1986) 6071–6074.
[18] H. Nakamura, Alkylated levoglucosan in organic synthesis. A formal total synthesis
of elaiophylin, Chem. Pharm. Bull. 38 (1990) 2435–2441.
Declaration of competing interest
[19] A. Fürst, P.A. Plattner, Steroids and sex hormones. Clx. 2(α),3(α)- and 2(β),3(β)-
oxidocholestane; the configuration of the 2-hydroxycholestanes, Helv. Chim. Acta
32 (1949) 275–283.
The authors declare the following financial interests/personal re-
lationships which may be considered as potential competing interests:
DC and AV are cofounders and have an equity interest in Juvabis AG, a
biotech start-up in the area of aminoglycoside antibiotics.
[20] E.D. Goddard-Borger, R.V. Stick, An efficient, inexpensive, and shelf-stable diazo-
transfer Reagent: imidazole-1-sulfonyl azide hydrochloride, Org. Lett. 9 (2007)
3797–3800.
[21] N. Fischer, E.D. Goddard-Borger, R. Greiner, T.M. Klapotke, B.W. Skelton,
J. Stierstorfer, Sensitivities of some imidazole-1-sulfonyl azide salts, J. Org. Chem.
77 (2012) 1760–1764.
7

M.G. Pirrone, et al.
CarbohydrateResearch491(2020)107984
[22] G.T. Potter, G.C. Jayson, G.J. Miller, J.M. Gardiner, An updated synthesis of the
diazo-transfer reagent imidazole-1-sulfonyl azide hydrogen sulfate, J. Org. Chem.
81 (2016) 3443–3446.
participation, J. Chem. Soc. (1954) 4284–4294.
[29] P. Ngoje, D. Crich, Synthesis of bradyrhizose from D-glucose, Org. Lett. 22 (2020)
523–527.
[23] H. Ye, R. Liu, D. Li, Y. Liu, H. Yuan, W. Guo, L. Zhou, X. Cao, H. Tian, J. Shen,
P.G. Wang, A safe and facile route to imidazole-1-sulfonyl azide as a diazotransfer
reagent, Org. Lett. 15 (2013) 18–21.
[30] H. Amarasekara, S. Dharuman, T. Kato, D. Crich, Synthesis of conformationally-
locked cis- and trans-bicyclo[4.4.0] mono-, di- and trioxadecane modifications of
galacto- and glucopyranose. Experimental limiting 3JH,H coupling constants for the
estimation of carbohydrate side chain populations and beyond, J. Org. Chem. 83
(2018) 881–897.
[24] Z. Miao, M. Xu, B. Hoffmann, B. Bernet, A. Vasella, Functionalised bicyclic exo-
glycals by alkynol cycloisomerisation of hydroxy 1,3-diynes and hydroxy haloalk-
ynes, Helv. Chim. Acta 88 (2005) 1885–1912.
[31] A. Gaudemer, Determination of configurations by NMR spectroscopy, in:
H.B. Kagan (Ed.), Stereochemistry: Fundamentals and Methods, vol. 1, Georg
Thieme, Stuttgart, 1977, pp. 44–136.
[25] P. Uhlmann, A. Vasella, Regioselective glycosylation of diols and triols: intra- and
intermolecular hydrogen bonds, Helv. Chim. Acta 75 (1992) 1979–1994.
[26] C.W. Jefford, B. Waegell, The bromination of bicyclo[3.2.1]octanone-3. An example
of an α,α’-diaxial dibromocyclohexanone, Tetrahedron Lett. 4 (1963) 1981–1986.
[27] A.J. Baretta, C.W. Jefford, B. Waegell, Stéréochimie des derivés bromés de la
diméthyl-6,6 bicyclo[3.1.1]heptanone-3 ou isonopinone. L’Effet anti-réflexe, Bull.
Soc. Chim. Fr. (1970) 3899–3908.
[32] E.L. Eliel, S.H. Wilen, Stereochemistry of Organic Compounds, Wiley, New York,
1994, p. 1267.
[33] I. Tvaroška, J. Gajdoš, Angular dependence of vicinal carbon-proton coupling
constants for conformational studies of the hydroxymethyl group in carbohydrates,
Carbohydr. Res. 271 (1995) 151–162.
[28] G.H. Alt, D.H.R. Barton, Some conformational aspects of neighbouring-group
8