A convenient approach toward the synthesis of enantiopure isomers of DMDP and ADMDP

Article abstract of DOI:10.1016/j.tet.2008.10.096

A practical method for the synthesis of five-membered iminocyclitols, pyrrolidine alkaloids bearing multiple hydroxyl substituents, has been developed. All of the eight key intermediates, enantiopure tri-O-benzyl cyclic nitrones, are prepared from four cheap, readily available d-aldopentoses. The nucleophilic addition of cyclic nitrones with vinyl magnesium chloride and TMSCN shows high 2,3-trans stereoselectivity. To construct the 2,3-cis configurations, inversion of the C-2 nitrile group is achieved via an elimination-reduction sequence. Using this approach, five isomers of DMDP and six isomers of ADMDP are prepared efficiently. In the biological evaluation, iminocyclitol 27 is a new and potent inhibitor against β-hexosaminidase with an IC₅₀ value of 0.2 μM.

Full text of DOI:10.1016/j.tet.2008.10.096

Tetrahedron 65 (2009) 93–100
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
A convenient approach toward the synthesis of enantiopure isomers of DMDP
and ADMDP
*
En-Lun Tsou, Yao-Ting Yeh, Pi-Hui Liang, Wei-Chieh Cheng
The Genomics Research Center, Academia Sinica, No. 128, Academia Road Sec. 2, Nankang District, Taipei 11529, Taiwan
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 11 August 2008
Received in revised form 27 October 2008
Accepted 31 October 2008
Available online 5 November 2008
A practical method for the synthesis of five-membered iminocyclitols, pyrrolidine alkaloids bearing
multiple hydroxyl substituents, has been developed. All of the eight key intermediates, enantiopure tri-
O-benzyl cyclic nitrones, are prepared from four cheap, readily available D-aldopentoses. The nucleophilic
addition of cyclic nitrones with vinyl magnesium chloride and TMSCN shows high 2,3-trans stereo-
selectivity. To construct the 2,3-cis configurations, inversion of the C-2 nitrile group is achieved via an
elimination–reduction sequence. Using this approach, five isomers of DMDP and six isomers of ADMDP
are prepared efficiently. In the biological evaluation, iminocyclitol 27 is a new and potent inhibitor
Keywords:
Iminocyclitols
Cyclic nitrones
Enantiopure
Stereoselectivity
against b-hexosaminidase with an IC50 value of 0.2 mM.
Ó 2008 Elsevier Ltd. All rights reserved.
b
-Hexosaminidase inhibitor
1. Introduction
‘anomeric center’ (C-2) is a synthetic challenge in glycochemistry
and glycomimic chemistry (Fig. 2). Fortunately, cyclic nitrones have
been shown to perform highly stereoselective 2,3-trans reactions
with various nucleophiles and this chemistry may fit our criteria to
prepare a diverse range of iminocyclitols.12,13 Indeed, in our hands
the treatment of tri-O-benzyl cyclic nitrone 1a with allylMgBr or
vinylMgCl gave single addition product 2a or 2b with the trans
configuration as the only observable diastereomers in 93 and 90%
yields, respectively (entries 1 and 2, Table 1). The reaction with
TMSCN gave a mixture of 2,3-trans and 2,3-cis isomers 2c and 3c
(19:1) in 93% total yield (entry 3, Table 1). Thus, isomers of DMDP
and ADMDP with a 2,3-trans configuration such as iminocyclitols
4–11 and 14 (Fig. 1) can be potentially prepared using this
chemistry. In contrast, new synthetic methods toward isomers,
Five-membered iminocyclitols, also called pyrrolidine alkaloids
have received a lot of attention recently because of their versatile
biological applications and promising therapeutic potentials.1 For
example, 2,5-dihydroxymethyl-3,4-di-hydroxypyrrolidine (DMDP,
4) is a known glycosidase inhibitor (Fig. 1), which was isolated from
plants4 and has been synthesized.2,3 In contrast, 1-aminodeoxy-
DMDP (ADMDP, 8) is an unnatural product and can only be
obtained by synthesis.5 A recent research showed that the N-1
modified derivatives of ADMDP significantly enhanced selectivity
and potency toward the inhibition of b
-glucosidases.6 Our previous
report indicated that ADMDP and its derivatives were selective and
potent small molecules for antivirals and osteoarthritis.7 In-
terestingly, several synthetic L-iminocyclitols such as L-DMDP (5)
were found to exhibit interesting or unexpected biological activity.8
Although all isomers of DMDP and some isomers of ADMDP
have been prepared via different synthetic routes,2,3,5,8–11 to the
best of our knowledge, however, a general approach to cover the
preparation of all possible configuration isomers of 4 and 8, such as
L
-ADMDP (9), has not been completely reported.3
The stereogenic centers at C-3, C-4, and C-5 can be obtained by
appropriate choice of chiral pool starting material, typically an
enantiopure sugar. However, how to efficiently stereocontrol at the
* Corresponding author. Tel.: þ886 2 2789 1262; fax: þ886 2 2789 9931.
E-mail address: wcheng@gate.sinica.edu.tw (W.-C. Cheng).
Figure 1. Examples of the prepared iminocyclitols.
0040-4020/$ – see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2008.10.096

94
E.-L. Tsou et al. / Tetrahedron 65 (2009) 93–100
Figure 2. General synthetic criteria for iminocyclitols.
such as 12 and 13, with a 2,3-cis configuration derived from cyclic
nitrones remain to be explored.14
Herein, we report a general method for the synthesis of imi-
nocyclitols using chiral cyclic nitrones as key intermediates. This
method is exemplified in the synthesis of the 11 stereodiversified
polyhydroxylated pyrrolidine alkaloids 4–14. From a structural
point of view, several interesting relationships should be noted: (i)
iminocyclitols 4, 8, and 1010 are enantiomers with 5,8 9, and 11,10
respectively; (ii) 12 and 139 are the C-2 epimers of 8 and 4, re-
spectively; (iii) 63 is the C-4 epimer of 5 and also the C-5 epimer of
7; (iv) 14 is the C-4 epimer of 8. Notably, iminocyclitols 9 and 12 are
new cores. Besides, the inhibitory activity of selective compounds
and their acetylated derivatives against various glycosidases was
also studied.
Scheme 1. Preparation of chiral tri-O-benzyl cyclic nitrones 1a and 1b.
On the other hand, preparation of amine 8 by multi-step trans-
formations from 4 resulted in a low yield. Attempted hydrogenation
of 2c under high pressure (ca. 100 atm) also failed,14 even in pro-
longed reaction time (96 h) with freshly prepared catalyst. To
overcome this problem, mild reduction conditions using Raney Ni/
H2 (1 atm, rt) in the presence of Boc2O were developed and both
hydroxylamine and nitrile moieties in 2c were reduced easily to bis-
N-Boc protected diamine 18 in 4 h. This flexible four-step synthetic
route was found to be applicable not only to 8 (75% from 1a) but also
for the direct preparation of 9, 10, 11, and 14 from the corresponding
nitrones 1h, 1c, 1f, and 1e in good overall yields (68–76%).
Table 1
Reactions of tri-O-benzyl cyclic nitrone 1a with various nucleophiles
In this study, we also explored a convenient synthetic route to
iminocyclitols with 2,3-cis configuration such as 12 and 13. The key
transformation was the inversion of the C-2 nitrile group via a mild
elimination and reduction sequence, differing from the oxidation–
reduction sequence described by Goti and co-workers.14 Mesyla-
tion of the hydroxyl group in the 2c/3c mixture followed by
elimination in the basic conditions produced iminonitrile 19 in
a regioselective manner (Scheme 3). Stereoselective reduction of
the sp2-hybridized carbon at C-2 in 19 was realized to give pre-
dominantly the 2,3-cis isomer 20a due to the steric effect of the
benzoxyl group at C-3. In optimal conditions, reduction of 19 with
NaBH4 in MeOH at 0 ꢀC for 5 h gave 20a and the 2,3-trans isomer
20b in high stereoselectivity (dr >9:1) and good yield (93–95%).
After simple separation by column chromatography, an 85% yield of
Entry
NuX
Product (trans/cis ratio)a
Total yieldb (%)
1
2
3
AllylMgBr
VinylMgCl
TMSCN
2a only
2b only
2c/3c¼19:1
83
90
93
a
The ratio determined by 1H NMR.
Isolated by column chromatography.
b
2. Results and discussion
2.1. Chemistry
Following the known method13,15 with slight modifications to
the work-up procedures,16 nitrone 1a was obtained in an overall
yield of 71%. Likewise, nitrone 1b, the C-5 epimer of 1a, was pre-
pared in 54% overall yield from 15 when mesylation was carried out
instead of iodination as shown in Scheme 1.17 Using similar pro-
cedures, nitrones 1c–h were prepared from other D-aldopentoses
(Fig. 3).13,15,17,18 The preparation of nitrones 1e–g has not been
reported yet. Significantly, due to symmetry operations and nu-
cleophilic substitution (once or twice) at C-5, all eight of the five-
membered tri-O-benzyl cyclic nitrones were accessible from four
cheaper D-aldopentoses instead of more expensive L-sugars.
With nitrones 1a–h in hand, transformations to the target
molecules (Fig. 1) could be investigated. For example, the vinyl
hydroxylamine 2b was reduced in the presence of Zn/AcOH20 to
cleave the N–O bond followed by N-Boc protection, ozonolysis, and
reduction with NaBH4 to furnish alcohol 17 (Scheme 2). After
debenzylation and the removal of Boc group, 4 (DMDP) was
obtained in an overall yield of 52% from 1a (Scheme 2). Our syn-
thetic route to 4 is one of the most efficient methods in comparison
with the reported protocols.4 Likewise, iminocyclitols 5 (50%), 6
(46%), 7 (43%), and 13 (43%) were prepared from the corresponding
nitrones 1h, 1d, 1f, and 1b in good yields.
Figure 3. All structures of chiral tri-O-benzyl cyclic nitrones 1a–h.

E.-L. Tsou et al. / Tetrahedron 65 (2009) 93–100
95
Figure 4. Structures of acetylated iminocyclitols 25–27.
yield) by using 30% hydrogen peroxide under basic conditions
(NaOH/MeOH) modified from Katrizky’s procedure.21 Attempted
direct hydration of amino-nitrile 20a without prior N-benzylation
gave complicated mixture. Amide 22 was converted to acylimido-
dicarbonate 23, which underwent a reductive cleavage with NaBH4
to provide alcohol 24 (65%).22 After deprotection, iminocyclitol 13
was successfully obtained in 25% overall yield from nitrone 1a.
Compared with the preceding approach shown in Scheme 2,
iminocyclitol 13 can also be prepared from nitrone 1b without
inversion of the hydroxymethyl moiety because of the intrinsic 4,5-
cis configuration of 1b. Notably, though the overall yield of 13
derived from 1a (25%) was poorer than that from 1b (43%), this
general preparation protocol could be useful for the inversion of the
hydroxymethyl group at C-2 for other stereoisomer of DMDP.
This method allowed us to flexibly construct the 2,3-cis or 2,3-
trans configuration with various intrinsic stereogenic centers at
C-3, C-4, and C-5 on the pyrrolidine ring. Thus, we are able to
efficiently prepare scaffolds of five-membered iminocyclitols and
easily modify them for biological study. According to literature
reported,7 ADMDP (8) itself is a moderate hexosaminidase inhibitor
Scheme 2. Synthesis of iminocyclitol 4 and 8 with a 2,3-trans configuration.
(IC50¼62
group at the C-1 position becomes a potent hexosaminidase in-
hibitor with an IC50 value of 0.16 M. This prompted us to convert
mM) but significantly 25 (Fig. 4) bearing an acetamido
m
iminocyclitols 8, 9, and 14 to the corresponding 25, 26, and 27 via
the selective acetylation.7 From the structural point of view, 26 and
Scheme 3. Synthesis of iminocyclitol 12 with a 2,3-cis configuration.
27 are new compounds; 26 is the L-iminocyclitol and the enantio-
mer of 25, and 27 is the C-4 epimer of 25 (Fig. 4).
pure 20a was obtained on a multigram scale. The reaction at a lower
temperature (ꢁ20 ꢀC) was sluggish, whereas the reaction at ele-
vated temperatures (25 or 50 ꢀC) gave a mixture of 20a,b in inferior
diastereoselectivity. As shown in Scheme 3, deprotection of 20a
with concomitant reduction of the nitrile group gave iminocyclitol
12 in 80% yield (51% overall yield for four steps starting from 1a).
Comparing 1H NMR spectra, the C-2 proton in the 2,3-cis com-
pound 12 showed at ca. 3.6 ppm, but in the 2,3-trans isomer 8 at ca.
3.2 ppm. In addition, the two protons at C-2 and C-3 in 12 showed
a strong NOE interaction confirming their cis relationship. For the
preparation of 13 (Scheme 4), compound 20a was subjected to
N-benzylation, giving the nitrile 21 (90% yield), which was
efficiently transformed into the corresponding amide 22 (83%
2.2. Biological evaluation
Glycosidases particularly
a-glucosidase (bacillus),
b-glucosidase
(almonds), -mannosidase (jack beans), and
a
b
-hexosaminidase
(jack beans, bovine kidney, and human placenta) were used for
test.7 The IC50 values of the selective compounds and their mono-
acetylated forms were shown in Table 2. ADMDP isomers 9–12,
and 14 with a free aminomethyl moiety did not inhibit
a
- or
-hexosaminidases expect 14 with
moderate inhibition activity (IC50¼32 M) against -man-
b
a
-glucosidase, a-mannosidase, b
m
a
nosidase. As expected, 25 was a potent inhibitor of
b-hexosa-
minidases with IC50 values ranging from 0.1 to 3
m
M.7 In contrast,
Table 2
Inhibition activities of iminocyclitols against glycosidases
Enzyme
IC50 (mM)
5
6
7
9
10 11 12 13
14 25 26 27
a
b
a
b
b
b
-Glucosidasea
32 64 16 df
d
d
d
d
d
d
d
d
d
d
d
d
d
d
d
d
d
d
2.6
21
98
220
ND
ND
d
d
32
d
d
d
d
d
d
d
d
d
d
-Glucosidaseb
-Mannosidasec
-Hexosaminidasec
131 76 93
88 94 52
d
d
d
d
d
d
144
d
d
d
0.1 55 0.2
55
0.3 27 0.6
-Hexosaminidased NDg ND ND
3
3
-Hexosaminidasee ND ND ND
a
From Bacillus stearothermophilus lyoph.
From almonds.
From jack beans.
From bovine Kidney.
From human placenta.
b
c
d
e
f
IC50 more than 200
Not determined.
mM.
g
Scheme 4. Synthesis of iminocyclitol 13 with a 2,3-cis configuration.

96
E.-L. Tsou et al. / Tetrahedron 65 (2009) 93–100
L
-iminocyclitol 26, the enantiomer of 25, showed a moderate in-
(10 mL) and TBAF (14.2 mL, 1 M in THF) was added. After the re-
hibition activity (IC50¼27–55
m
M) against
b
-hexosaminidases.
-hexosa-
M. This obser-
action was refluxed at 100 ꢀC for 30 min, the mixture was con-
centrated and purified by CC (33% EA in hexanes, silica gel) to give
Excitingly, 27 exhibited an inhibitory potency against
b
minidases with IC50 values ranging from 0.2 to 3
vation pointed out that the configuration of the hydroxyl group at
the C-4 position does not significantly affect the inhibition activity
m
cyclic nitrone 1a (3.55 g, 8.5 mmol, 71% from 15) as a white solid.
20
[a
]
ꢁ40.5 (c 0.7, CHCl3); 1H NMR (600 MHz, CDCl3)
d 3.77 (dd, 1H,
D
J¼2.9, 10.2 Hz), 4.02 (br, 1H), 4.06 (dd, 1H, J¼5.0, 10.2 Hz), 4.39 (t,
1H, J¼2.4, 3.2 Hz), 4.50–4.53 (m, 5H), 4.61 (d, 1H, J¼12 Hz), 4.66 (br,
1H), 6.90 (s, 1H), 7.27–7.36 (m, 15H); 13C NMR (150 MHz, CDCl3)
against
b-hexosaminidases. Presumably, chemical modification,
such as conjugation with various fragments, at the C-4 position in
iminocyclitol 25 or 26 to explore the extra binding pockets is
possible to improve the binding affinity or selectivity.
d
137.3, 136.9, 136.7, 132.4, 128.1, 128.0, 127.9, 127.6, 127.5, 127.4,
127.2, 82.3, 79.8, 77.0, 72.9, 71.3, 71.0, 65.6. HRMS calcd for
[C26H27NO4þH]þ 418.2013, found 418.2049.
3. Conclusion
4.2.2. (2S,3R,4R)-3,4-Bis(benzyloxy)-2-(benzyloxymethyl)-3,4-
dihydro-2H-pyrrole-1-oxide (1b)
Following the same procedure as described for the preparation
of 1a but mesylation was carried out instead of iodination, cyclic
We have successfully developed a general and straightforward
method to efficiently prepare scaffolds of iminocyclitols and easily
modify for biological study. It is the first time to prepare all eight
five-membered chiral tri-O-benzyl cyclic nitrones 1a–h as key in-
20
nitrone 1b was obtained in 54% yield from 14 as a colorless oil. [a]
D
termediates from
D-aldopentoses instead of the more expensive
ꢁ70.5 (c 0.5, CHCl3); 1H NMR (600 MHz, CDCl3)
d 3.82 (dd, 1H,
L-sugars. The utility of this methodology was exemplified in the
J¼2.0, 10.1 Hz), 3.98 (dd, 1H, J¼4.4, 10.1 Hz), 4.15 (br, 1H), 4.36 (dd,
1H, J¼4.4, 7.6 Hz), 4.50–4.66 (m, 6H), 4.75 (dd, 1H, J¼1.6, 2.5 Hz),
synthesis of 11 natural and unnatural polyhydroxylated pyrroli-
dines under mild reaction conditions in high regio- and diaster-
eoselectivities. This flexible route allowed us to shorten the tedious
6.82 (s, 1H), 7.26–7.36 (m, 15H); 13C NMR (150 MHz, CDCl3)
d 137.8,
137.2, 137.1, 133.3, 128.5, 128.4, 128.2, 128.0, 127.9, 127.8, 127.7, 127.5,
127.4, 83.0, 80.4, 74.0, 73.4, 73.0, 72.3, 64.3. HRMS calcd for
[C26H27NO4þH]þ 418.2013, found 418.2020.
synthetic process for several iminocyclitols such as L-DMDP and
also diversify the preparation of isomers of DMDP (4) and ADMDP
(8). This is potentially useful in the preparation of their stereoiso-
mers and congeners for systemic biological applications. In the
4.2.3. (2R,3R,4S)-3,4-Bis(benzyloxy)-2-(benzyloxymethyl)-3,4-
dihydro-2H-pyrrole-1-oxide (1c)
biological study, the new compound 27 is
a lead inhibitor
(IC50¼0.2
m
M) against jack bean -hexosaminidase. We are cur-
b
The reaction was carried out as described above for 1a starting
rently using this approach to synthesize and identify potent gly-
cosidase or glycotransferase inhibitors.
from 2,3,5-tri-O-benzyl-
give 1c (56%) as a colorless oil. [
(600 MHz, CDCl3)
(dd, 1H, J¼2.5, 10.4 Hz), 4.38 (d, 1H, J¼11.9 Hz), 4.43 (dd, 1H, J¼4.7,
5.9 Hz), 4.52 (dd, 2H, J¼7.4, 11.9 Hz), 4.56–4.67 (m, 4H), 6.92 (s, 1H),
7.19 (t, 2H, J¼1.4 Hz), 7.26–7.32 (m, 13H); 13C NMR (150 MHz, CDCl3)
b-D-ribofuranose, derived from D-ribose, to
20
a
]
þ118 (c 1.8, CHCl3); 1H NMR
D
d
3.58 (dd, 1H, J¼2.5, 10.4 Hz), 4.08 (m, 1H), 4.12
4. Experimental section
4.1. General
d
137.6, 137.4, 137.2, 133.8, 128.7, 128.7, 128.6, 128.34, 128.31, 128.26,
Ozonolysis was performed by ozone generator (Fischer Tech. OZ
502/10). Mass spectra were measured with a Bruker BioTOF III (ESI-
MS). Optical rotations were measured with a Perkin–Elmer Model
341 polarimeter. NMR spectra (1H at 400 or 600 MHz, 13C at 100 or
150 MHz) were recorded in CDCl3, CD3OD, or D2O solvents. Flash
column chromatography was carried out using Merck kieselgel Si60
128.0, 127.8, 76.4, 75.4, 74.46, 73.49, 72.5, 72.2, 64.8. HRMS calcd for
[C26H27NO4þH]þ 418.2013, found 418.2002.
4.2.4. (2S,3R,4S)-3,4-Bis(benzyloxy)-2-(benzyloxymethyl)-3,4-
dihydro-2H-pyrrole-1-oxide (1d)
The reaction was carried out as described above for 1b starting
(40–63
m
m). All reagents and solvents were purchased from com-
from 2,3,5-tri-O-benzyl-
give 1d (34%) as a white solid. [
(600 MHz, CDCl3)
b-D-ribofuranose, derived from D-ribose, to
20
mercial suppliers, and used without further purification. Palladium
hydroxide is 20 wt % Pd on carbon, CC refers to column chroma-
tography (silica gel). Concentration refers to rotary evaporation.
a]
þ59.6 (c 1.8, CHCl3); 1H NMR
D
d
4.07 (dd, 1H, J¼4.0, 9.8 Hz), 4.12–4.17 (m, 2H),
4.37 (t, 1H, J¼5.8, 5.9 Hz), 4.54–4.60 (m, 4H), 4.67 (s, 2H), 4.70 (d,
1H, J¼11.8 Hz), 6.87 (t, 1H, J¼1.7, 1.8 Hz), 7.24–7.32 (m, 15H); 13C
4.2. General procedure for the preparation of chiral cyclic
nitrones
NMR (150 MHz, CDCl3) d 137.8, 137.5, 137.3, 133.1,128.7, 128.6, 128.5,
128.2, 128.1, 127.99, 127.97, 127.8, 76.9, 74.5, 74.1, 73.6, 73.2, 72.6,
66.7. HRMS calcd for [C26H27NO4þH]þ 418.2013, found 418.2002.
4.2.1. (2R,3R,4R)-3,4-Bis(benzyloxy)-2-(benzyloxymethyl)-3,4-
dihydro-2H-pyrrole-1-oxide (1a)
4.2.5. (2R,3S,4R)-3,4-Bis(benzyloxy)-2-(benzyloxymethyl)-3,4-
dihydro-2H-pyrrole-1-oxide (1e)
2,3,5-Tri-O-benzyl-b-D-arabinofuranose (15) (5 g, 12 mmol) was
reacted with hydroxylamine hydrochloride (6.6 g, 94 mmol) in the
presence of sodium methoxide (2.56 g, 47 mmol) in methanol. The
mixture was stirred for 5 h and then the solvent was evaporated.
The residue was extracted with CH2Cl2, washed with H2O, dried
(MgSO4), and concentrated. A mixture of the oxime residue, tert-
butylchlorodiphenylsilane (TBDPSCl, 4.25 g, 15 mmol), and imid-
azole (1.21 g, 18 mmol) in CH2Cl2 (15 mL) was stirred at rt for 2 h
and quenched with water (20 mL). The organic layer was washed
with brine, dried (MgSO4), and concentrated to give 16. A mixture
of 16, triphenylphosphine (9.35 g, 36 mmol), imidazole (2.42 g,
36 mmol), and iodine (6.03 g, 24 mmol) in toluene (30 mL) was
refluxed for 1 h, the reaction mixture was filtered, and washed with
CH2Cl2. The organic layer was washed with brine, dried (MgSO4),
and concentrated. The crude intermediate was dissolved in toluene
The reaction was carried out as described above for 1a starting
from 2,3,5-tri-O-benzyl-
give 1e (12%) as a white solid. [
(600 MHz, CDCl3)
b-D-lyxofuranose, derived from D-lyxose, to
20
a
]
ꢁ56.9 (c 1, CHCl3); 1H NMR
D
d
4.05 (dd, 1H, J¼4.0, 9.8 Hz), 4.17 (m, 2H), 4.38 (t,
1H, J¼5.8, 5.9 Hz), 4.53–4.59 (m, 4H), 4.67 (s, 2H), 4.70 (d, 1H,
J¼11.9 Hz), 6.89 (s, 1H), 7.24–7.33 (m, 15H); 13C NMR (150 MHz,
CDCl3)
d 137.8, 137.4, 137.3, 133.5, 128.6, 128.5, 128.4, 128.1, 128.0,
127.9, 127.7, 77.0, 74.6, 74.2, 73.6, 73.2, 72.6, 66.6. HRMS calcd for
[C26H27NO4þNa]þ 440.1832, found 440.1868.
4.2.6. (2S,3S,4R)-3,4-Bis(benzyloxy)-2-(benzyloxymethyl)-3,4-
dihydro-2H-pyrrole-1-oxide (1f)
The reaction was carried out as described above for 1b starting
from 2,3,5-tri-O-benzyl-b-D-lyxofuranose, derived from D-lyxose, to

E.-L. Tsou et al. / Tetrahedron 65 (2009) 93–100
97
20
give 1f (53%) as a colorless oil. [
(600 MHz, CDCl3)
a]
ꢁ113.8 (c 0.8, CHCl3); 1H NMR
the residue was extracted with CH2Cl2 and water. The organic layers
were dried with MgSO4, concentrated, and purified by CC (20%
EtOAc in hexanes, silica gel) to give 17 (1.27 g, 2.4 mmol, 74% from
2b) as a syrup. 1H NMR (600 MHz, CDCl3, mixture of conformers,
D
d
3.59 (dd, 1H, J¼2.5, 10.4 Hz), 4.09 (m, 1H), 4.12
(dd, 1H, J¼2.5, 10.4 Hz), 4.38 (d, 1H, J¼11.9 Hz), 4.43 (dd, 1H, J¼4.7,
5.9 Hz), 4.52 (dd, 2H, J¼7.4, 11.9 Hz), 4.57–4.68 (m, 4H), 6.95 (s, 1H),
7.18 (t, 2H, J¼1.4 Hz), 7.26–7.33 (m, 13H); 13C NMR (150 MHz, CDCl3)
major/minor¼2:1) (for a major conformer):
d 1.42 (s, 9H), 3.47–3.53
(m, 1H), 3.75 (dd, 1H, J¼4.1, 8.8 Hz), 3.84–3.90 (m, 3H), 4.05–4.08
d
137.5, 137.3, 137.1, 133.9, 128.7, 128.5, 128.4, 128.2, 128.1, 128.09,
128.05, 127.8, 127.6, 76.4, 75.4, 74.5, 73.4, 72.4, 72.1, 64.7. HRMS
(m, 2H), 4.15 (s, 1H), 4.38–4.67 (m, 6H), 7.18–7.35 (m, 15H); (for
calcd for [C26H27NO4þH]þ 418.2013, found 418.2042.
a minor conformer): d 1.47 (s, 9H), 3.47–3.53 (m, 1H), 3.72–3.76 (m,
1H), 3.80 (dd, 2H, J¼4.0, 11.4 Hz), 3.97 (s, 1H), 4.01 (dd, 1H, J¼4.1,
8.5 Hz), 4.17 (s, 1H), 4.28 (dd, 1H, J¼3.5, 10.3 Hz), 4.38–4.67 (m, 6H),
7.18–7.35 (m, 15H); 13C NMR (150 MHz, CDCl3, mixture of con-
4.2.7. (2R,3S,4S)-3,4-Bis(benzyloxy)-2-(benzyloxymethyl)-3,4-
dihydro-2H-pyrrole-1-oxide (1g)
The reaction was carried out as described above for 1a starting
formers) d 155.6, 154.2, 138.7, 138.3, 137.6, 137.5, 137.2, 128.7, 128.6,
from 2,3,5-tri-O-benzyl-
to give 1g (54%) as a colorless oil. [
(600 MHz, CDCl3)
b
-
D
-xylofuranose, derived from
D
-xylose,
128.5, 128.2, 128.1, 128.06, 128.01, 127.9, 127.8, 127.7, 127.6, 85.8,
84.3, 81.9, 80.9, 80.5, 80.4, 73.2, 73.2, 71.5, 71.4, 71.39, 71.32, 68.5,
68.0, 66.6, 65.6, 64.6, 63.6, 63.2, 62.4, 28.6, 28.5. HRMS calcd for
[C32H39NO6þNa]þ 556.2670, found 556.2661.
20
a]
þ72.7 (c 1, CHCl3); 1H NMR
D
d
3.82 (dd, 1H, J¼2.0, 10.1 Hz), 4.00 (dd, 1H,
J¼4.4, 10.1 Hz), 4.15 (br, 1H), 4.36 (dd, 1H, J¼4.4, 7.6 Hz), 4.49–4.76
(m, 6H), 4.77 (dd, 1H, J¼1.6, 2.5 Hz), 6.84 (s, 1H), 7.28–7.34 (m,
15H); 13C NMR (150 MHz, CDCl3)
d
137.6, 137.0, 136.9, 133.1, 128.2,
4.3.3. (2R,3R,4R,5R)-2,5-Bis(hydroxymethyl)pyrrolidine-3,4-diol
(4, DMDP)
128.1, 128.0, 127.8, 127.7, 127.6, 127.5, 127.2, 82.8, 80.2, 73.8, 73.2,
72.7, 72.0, 64.1. HRMS calcd for [C26H27NO4þH]þ 418.2013, found
418.2017.
A mixture of 17 (0.3 g, 0.56 mmol) and palladium hydroxide
(0.03 g) in MeOH was stirred for overnight under a hydrogen
atmosphere. The reaction mixture was filtered through Celite
and the filtrate was concentrated. The residue was treated with
the solution of CH2Cl2/TFA (2 mL/2 mL) and the reaction mix-
ture was stirred at 0 ꢀC for 30 min. The reaction mixture was
concentrated and purified by CC (25% aqueous NH4OH in
4.2.8. (2S,3S,4S)-3,4-Bis(benzyloxy)-2-(benzyloxymethyl)-3,4-
dihydro-2H-pyrrole-1-oxide (1h)
The reaction was carried out as described above for 1b starting
from 2,3,5-tri-O-benzyl-
give 1h (74%) as a white solid. [
(600 MHz, CDCl3)
b-D-xylofuranose, derived from D-xylose, to
þ37.3 (c 1, CHCl3); 1H NMR
propanol, silica gel) to give 4 (71.47 mg, 0.44 mmol, 78%) as
20
a]
D
20
d
3.77 (d, 1H, J¼3.9, 10.2 Hz), 4.01 (br, 1H), 4.05
a syrup (free form). [
D2O)
a
]
þ53.4 (c 0.6, H2O); 1H NMR (600 MHz,
D
(dd, 1H, J¼5.0, 10.2 Hz), 4.38 (t, 1H, J¼2.4, 3.2 Hz), 4.50–4.55 (m,
d
3.34 (dd, 2H, J¼3.4, 5.8 Hz), 3.73 (dd, 2H, J¼6.1, 12.3 Hz),
5H), 4.60 (d, 1H, J¼12 Hz), 4.65 (br, 1H), 6.89 (s, 1H), 7.24–7.33 (m,
3.81 (dd, 2H, J¼3.8, 12.3 Hz), 3.95 (dd, 2H, J¼2.2, 5.8 Hz); 13C
15H); 13C NMR (150 MHz, CDCl3)
d
137.6, 137.1, 137.0, 132.8, 128.5,
NMR (150 MHz, D2O) d 75.5, 62.1, 59.2. HRMS calcd for
128.4, 128.3, 128.0, 127.9, 127.8, 127.7, 127.6, 127.5, 82.6, 80.2, 77.4,
73.4, 71.8, 71.5, 66.0. HRMS calcd for [C26H27NO4þNa]þ 440.1832,
found 440.1818.
[C6H13NO4þH]þ 164.0917, found 164.0899.
4.3.4. (2S,3S,4S,5S)-2,5-Bis(hydroxymethyl)pyrrolidine-3,4-diol
(5,
L
-DMDP)
4.3. Typical procedure for the preparation of DMDP from 1a
(1a/2b/17/4)
The reaction was carried out as described above for 4 starting
20
from 1h to give 5 (free form, 50% yield from 1h) as a syrup. [a]
D
ꢁ51.0 (c 0.2, H2O); 1H NMR (600 MHz, D2O)
d 3.17 (br, 2H), 3.71 (dd,
4.3.1. (2R,3R,4R)-3,4-Bis(benzyloxy)-2(benzyloxymethyl)-5-
vinylpyrrolidin-1-ol (2b)
2H, J¼6.0, 11.6 Hz), 3.79 (d, 2H, J¼11.7 Hz), 3.92 (t, 2H, J¼2.2 Hz); 13C
NMR (150 MHz, D2O)
d 77.1, 61.7, 61.2. HRMS calcd for
Compound 1a (2 g, 5 mmol) was dissolved in THF (20 mL) and
then vinyl magnesium chloride (0.54 g, 6 mmol) was added drop-
wise at 0 ꢀC under Ar(g). After 15 h, the reaction mixture was
quenched with aqueous NH4Cl solution, extracted with CH2Cl2,
dried with MgSO4, and concentrated. The crude sample was puri-
[C6H13NO4þH]þ 164.0917, found 164.0933.
4.3.5. (2S,3R,4S,5S)-2,5-Bis(hydroxymethyl)pyrrolidine-3,4-diol
(6,
L
-DALDP)
The reaction was carried out as described above for 4 starting
20
fied by CC (25% EtOAc in hexanes, silica gel) to give 2b (1.91 g,
from 1d to give 6 (free form, 46% yield from 1d) as a syrup. [a]
D
20
4.3 mmol, 90%) as a brown solid. [
NMR (600 MHz, CDCl3)
a
]
ꢁ19.6 (c 0.66, CHCl3); 1H
ꢁ29.1 (c 1, H2O); 1H NMR (600 MHz, D2O)
d 3.20 (m, 1H), 3.37 (m,
D
d
3.56 (t, 1H, J¼4.9 Hz), 3.68 (dd, 1H, J¼6.0,
1H), 3.67 (m, 2H), 3.78 (dd, 1H, J¼3.9, 11.8 Hz), 3.82 (dd, 1H, J¼6.6,
9.8 Hz), 3.73 (dd,1H, J¼4.9, 9.8 Hz), 3.84 (t,1H, J¼8.1 Hz), 3.91 (t,1H,
11.3 Hz), 4.03 (dd, 1H, J¼4.3, 8.6 Hz), 4.21 (t, 1H, J¼4.0 Hz); 13C NMR
J¼5.0 Hz), 3.99 (t, 1H, J¼3.8 Hz), 4.45–4.56 (m, 6H), 5.31 (q, 2H),
(150 MHz, D2O)
d 73.2, 71.7, 61.5, 61.4, 60.1, 59.9. HRMS calcd for
6.02 (m, 1H), 7.23–7.31 (m, 15H); 13C NMR (150 MHz, CDCl3)
d
138.0,
[C6H13NO4þH]þ 164.0917, found 164.0892.
137.9, 137.8, 135.5, 128.1, 128.0, 127.9, 127.7, 127.6, 127.5, 127.4,
127.39, 127.35, 119.0, 85.9, 83.6, 73.0, 72.5, 71.6, 71.4, 69.5, 67.5.
HRMS calcd for [C28H31NO4þH]þ 446.2326, found 446.2344.
4.3.6. (2S,3S,4R,5R)-2,5-Bis(hydroxymethyl)pyrrolidine-3,4-diol
(7, DADP)
The reaction was carried out as described above for 4 starting
20
4.3.2. (2R,3R,4R,5R)-tert-Butyl-3,4-bis(benzyloxy)-2-(benzyloxy-
methyl)-5-(hydroxymethyl)-pyrrolidine-1-carboxylate (17)
from 1f to give 7 (free form, 43% yield from 1f) as a syrup. [a]
D
þ39.2 (c 2, H2O); 1H NMR (600 MHz, D2O)
d 3.06–3.09 (m, 2H), 3.65
A mixture of 2b (1.44 g, 3.2 mmol) and zinc powder (0.21 g,
3.2 mmol) in acetic acid was stirred at rt for 8 h. The reaction
mixture was filtered through Celite and the filtrate was neutralized
by aqueous NaHCO3 solution and extracted with CH2Cl2. The or-
ganic layers were dried with MgSO4, concentrated, and directly
reacted with di-tert-butyldicarbonate (0.88 g, 4 mmol) in CH2Cl2
(10 mL) in the presence of triethylamine (1 mL) to give a crude Boc-
protected amine compound. After ozonolysis and reduction with
NaBH4 (0.24 g, 6.3 mmol), the reaction solvent was removed and
(dd, 2H, J¼6.2, 11.7 Hz), 3.73 (dd, 2H, J¼4.2, 11.7 Hz), 3.84–3.87 (m,
2H); 13C NMR (150 MHz, D2O)
d 77.5, 61.6, 61.5. HRMS calcd for
[C6H13NO4þH]þ 164.0917, found 164.0858.
4.3.7. (2S,3R,4R,5R)-2,5-Bis(hydroxymethyl)pyrrolidine-3,4-diol
(13, DGDP)
The reaction was carried out as described above for 4 starting
20
from 1b to give 13 (43% from 1b) as a syrup. [
a
]
þ18.1 (c 1,
D
H2O); 1H NMR (600 MHz, D2O)
d
3.05 (d, 1H, J¼5.40 Hz), 3.35 (d,

98
E.-L. Tsou et al. / Tetrahedron 65 (2009) 93–100
1H, J¼5.5 Hz), 3.62–3.67 (m, 2H), 3.72 (dd, 1H, J¼4.7, 11.6 Hz),
3.77 (dd, 1H, J¼5.8, 11.5 Hz), 3.85 (dd, 1H, J¼2.5, 4.4 Hz), 4.08 (t,
4.4.4. (2S,3S,4R,5R)-2-(Aminomethyl)-5-(hydroxymethyl)-
pyrrolidine-3,4-diol (10)10
1H, J¼2.5 Hz); 13C NMR (150 MHz, D2O)
d
78.6, 76.9, 65.1, 61.7,
The reaction was carried out as described above for 8 starting
20
61.1, 59.6. HRMS calcd for [C6H13NO4þNa]þ 186.0737, found
from 1c to give 10 (72% from 1c) as a syrup. [
a]
ꢁ4 (c 0.3, H2O); 1H
D
186.0701.
NMR (600 MHz, D2O)
d
3.54 (m, 2H), 3.81 (dd, 1H, J¼4.0, 9.0 Hz),
3.86 (m, 2H), 3.95 (dd, 1H, J¼3.8, 12.6 Hz), 4.27 (m, 2H); 13C NMR
(150 MHz, D2O)
d 72.5, 70.2, 65.9, 58.8, 58.4, 38.65. HRMS calcd for
4.4. Typical procedure for the preparation of ADMDP from 1a
(1a/2c/8)
[C6H14N2O3þH]þ 163.1077, found 163.1078.
4.4.5. (2R,3R,4S,5S)-2-(Aminomethyl)-5-(hydroxymethyl)-
4.4.1. (2R,3R,4R,5R)-3,4-Bis(benzyloxy)-5-(benzyloxymethyl)-1-
hydroxypyrrolidine-2-carbonitrile (2c, trans isomer) and
(2S,3R,4R,5R)-3,4-bis(benzyloxy)-5-(benzyloxymethyl)-1-
pyrrolidine-3,4-diol (11)
The reaction was carried out as described above for 8 starting
from 1f to give 11 (68% from 1f) as a syrup. [
20
a
]
þ3 (c 0.2, H2O); 1H
D
hydroxypyrrolidine-2-carbonitrile (3c, cis isomer)
NMR (600 MHz, D2O)
d
3.55 (m, 2H), 3.83 (dd, 1H, J¼3.9, 9.00 Hz),
A mixture of 1a (1 g, 2.4 mmol) and trimethylsilyl cyanide
(0.76 mL, 5.7 mmol) in dry methanol (10 mL) was stirred at 50 ꢀC for
8 h. The reaction solvent was removed under vacuum and the res-
idue was purified by CC (25% EtOAc in hexanes, silica gel) to give
a mixture of 2c/3c (0.99 g, 2.2 mmol, 93%, 2c/3c¼19:1 in the 1H NMR
spectrum). A mixture of 2c/3c was further separated by CC to give
pure 2c (0.93 g, 2.1 mmol, 88%) as a white solid, pure 3c (10.6 mg,
3.88 (m, 2H), 3.96 (dd, 1H, J¼3.9, 12.7 Hz), 4.29 (m, 2H); 13C NMR
(150 MHz, D2O)
d 72.5, 70.2, 66.0, 58.8, 58.3, 38.6. HRMS calcd for
[C6H14N2O3þH]þ 163.1077, found 163.1074.
4.4.6. (2R,3R,4S,5S)-2-(Aminomethyl)-5-(hydroxymethyl)-
pyrrolidine-3,4-diol (14)
The reaction was carried out as described above for 8 starting
20
0.02 mmol, 1%) as a white solid, and 2c/3c mixture (0.04 g).
from 1e to give 14 (76% from 1f) as a syrup. [
NMR (600 MHz, D2O)
a
]
þ18 (c 0.1, H2O); 1H
D
20
Compound 2c: [
a
]
þ8.8 (c 1.1, CHCl3); 1H NMR (600 MHz,
d
3.55 (s, 1H), 3.56 (d, 1H, J¼3.2 Hz), 3.83 (m,
D
CDCl3)
d
3.31 (m, 1H), 3.57 (dd, 1H, J¼3.5, 10.6 Hz), 3.78 (dd, 1H,
1H), 3.88 (m, 1H), 3.95 (m, 1H), 4.04 (dd, 1H, J¼7.3, 17.6 Hz), 4.32
J¼3.2, 10.6 Hz), 4.00 (dd, 1H, J¼2.3, 6.7 Hz), 4.17 (t, 1H, J¼2.2 Hz),
4.25 (d, 1H, J¼1.7 Hz), 4.41–4.43 (m, 3H), 4.49 (dd, 2H, J¼4.6,
11.9 Hz), 4.58 (d, 1H, J¼12.0 Hz), 7.26–7.40 (m, 15H); 13C NMR
(dd, J¼5.8, 7.3 Hz), 4.38 (t, 1H, J¼5.3 Hz); 13C NMR (150 MHz, D2O)
d
74.1, 69.6, 62.3, 57.6, 57.0, 39.4. HRMS calcd for [C6H14N2O3þH]þ
163.1077, found 163.1083.
(150 MHz, CDCl3)
d 137.2, 137.1, 136.3, 128.6, 128.3, 128.1, 128.0,
127.9, 127.8, 127.78, 127.74, 127.69, 115.63, 83.4, 80.9, 73.1, 72.0, 71.9,
69.3, 66.0, 61.2. HRMS calcd for [C27H28N2O4þNa]þ 467.1941, found
4.5. Typical procedure for the inversion of C-2 configuration
and the preparation of iminocyclitol 12 (2c/3c/19/20a/12)
467.1961.
20
Compound 3c: [
CDCl3)
a]
þ12.2 (c 0.45, CHCl3); 1H NMR (600 MHz,
D
d
3.10 (dd, 1H, J¼5.3, 10.7 Hz), 3.61 (dd, 1H, J¼5.6, 10.1 Hz),
4.5.1. (2R,3R,4R)-3,4-Bis(benzyloxy)-2-(benzyloxymethyl)-3,4-
dihydro-2H-pyrrole-5-carbonitrile (19)
A mixture of 2c and 3c (1.69 g, 3.8 mmol) was treated with
methanesulfonyl chloride (0.36 mL, 4.6 mmol) in CH2Cl2 (10 mL)
in the presence of triethylamine (1.3 mL, 7.7 mmol) at 0 ꢀC for
3 min. The reaction was quenched with H2O and extracted with
3.65 (dd, 1H, J¼5.2, 10.1 Hz), 3.83 (d, 1H, J¼5.3 Hz), 4.03 (s, 2H), 4.38
(q, 2H, J¼11.8 Hz), 4.50–4.56 (m, 3H), 4.65 (d, 1H, J¼11.9 Hz), 7.18–
7.36 (m, 15H); 13C NMR (150 MHz, CDCl3)
d 137.9, 137.3, 136.8, 128.6,
128.5, 128.4, 128.3, 128.2, 128.0, 127.9, 127.8, 1163.5, 81.8, 79.9, 73.4,
72.6, 72.0, 71.9, 68.6, 62.2. HRMS calcd for [C27H28N2O4þNa]þ
467.1941, found 467.1945.
CH2Cl2. The organic layers were dried with MgSO4 and concen-
20
trated to give 19 (1.21 g, 2.8 mmol, 75%). [
a
]
ꢁ4 (c 1.0, CHCl3); 1H
D
4.4.2. (2R,3R,4R,5R)-2-(Aminomethyl)-5-(hydroxymethyl)-
pyrrolidine-3,4-diol (8, ADMDP)
NMR (600 MHz, CDCl3)
d
3.65 (dd, 1H, J¼5.9, 10.0 Hz), 3.77 (dd, 1H,
J¼4.6, 10.0 Hz), 4.22 (t, 1H, J¼3.7 Hz), 4.38 (dd, 1H, J¼4.6, 9.1 Hz),
4.54 (dd, 2H, J¼7.6, 11.8 Hz), 4.58 (s, 2H), 4.69 (s, 1H), 4.71 (d, 1H,
J¼3.1 Hz), 4.78 (d, 1H, J¼11.6 Hz), 7.30–7.45 (m, 15H); 13C NMR
A mixture of 2c (0.64 g, 1.4 mmol), Raney nickel (0.05 g), and di-
tert-butyldicarbonate (0.6 g, 2.7 mmol) in dry methanol (3 mL) was
stirred under a hydrogen atmosphere. After 4 h, palladium hy-
droxide (0.05 g) was added and the reaction mixture was also
stirred under a hydrogen atmosphere for further 10 h. The reaction
mixture was filtered through Celite and the filtrate was concen-
trated. The residue was treated with the solution of CH2Cl2/TFA
(3 mL/3 mL) and the reaction mixture was stirred at 0 ꢀC for 30 min.
The reaction mixture was concentrated and purified by CC (25%
(150 MHz, CDCl3)
d 152.1, 137.8, 137.3, 136.6, 128.8, 128.7, 128.6,
128.5, 128.4, 128.3, 128.2, 128.1, 127.97, 127.94, 127.8, 113.7, 90.2,
84.0, 78.3, 73.5, 73.2, 72.3, 69.7. HRMS calcd for [C27H26N2O3þH]þ
427.2016, found 427.2009.
4.5.2. (2S,3R,4R,5R)-3,4-Bis(benzyloxy)-5-(benzyloxymethyl)-
pyrrolidine-2-carbonitrile (20a) and (2R,3R,4R,5R)-3,4-bis-
aqueous NH4OH (37%) in propanol, silica gel) to give 8 (0.2 g,
(benzyloxy)-5-(benzyloxymethyl)pyrrolidine-2-carbonitrile (20b)
Sodium borohydride (0.28 g, 7.4 mmol) was added to 19 in
methanol (8 mL) and the reaction was stirred at 0 ꢀC for 5 h. The
solvent was removed and the reaction mixture was extracted with
CH2Cl2 and brine. The organic layers were dried with MgSO4,
concentrated, and purified by CC (25% EtOAc in hexanes, silica gel)
to give a mixture of 20a and 20b (1.15 g, 2.68 mmol, 95%). Further
purification of this mixture gave pure 20a (1.03 g, 2.4 mmol, 85%)
as a colorless syrup and pure 20b (0.057 g, 0.13 mmol, 5%) as
20
1.2 mmol, 86%) as a white solid. [
(600 MHz, D2O)
a
]
þ43.5 (c 0.5, H2O); 1H NMR
D
d
3.18 (q, 2H), 3.31 (dd, 1H, J¼4.8, 13.3 Hz), 3.41 (m,
1H), 3.73 (dd, 1H, J¼6.1, 11.9 Hz), 3.81 (dd, 1H, J¼3.9, 12.1 Hz), 3.94
(m, 2H); 13C NMR (150 MHz, D2O)
d 78.6, 76.4, 62.1, 60.5, 58.0, 41.3.
HRMS calcd for [C6H14N2O3þH]þ 163.1077, found 163.1071.
4.4.3. (2S,3S,4S,5S)-2-(Aminomethyl)-5-(hydroxymethyl)-
pyrrolidine-3,4-diol (9)
The reaction was carried out as described above for 8 starting
a syrup.
20
20
from 1h to give 9 (76% from 1h). [
(600 MHz, D2O)
a
]
ꢁ41.6 (c 0.8, H2O); 1H NMR
Compound 20a: [
CDCl3)
a
]
D
ꢁ8.7 (c 1, CHCl3); 1H NMR (600 MHz,
D
d
3.03 (m, 2H), 3.18 (dd,1H, J¼4.3,12.9 Hz), 3.24 (m,
d
2.30 (br, 1H), 3.32 (dd, 1H, J¼5.9, 10.3 Hz), 3.52–3.57 (m,
1H), 3.64 (dd, 1H, J¼5.7, 11.4 Hz), 3.71 (dd, 1H, J¼3.4, 11.5 Hz), 3.83
2H), 3.89 (d, 1H, J¼3.8 Hz), 4.08 (d, 1H, J¼6.7 Hz), 4.45–4.53 (m, 4H),
(m, 2H); 13C NMR (150 MHz, D2O)
d 79.2, 77.1, 61.7, 61.1, 57.9, 42.0.
4.61 (q, 2H, J¼11.9 Hz), 7.22–7.36 (m, 15H); 13C NMR (150 MHz,
HRMS calcd for [C6H14N2O3þH]þ 163.1077, found 163.1071.
CDCl3) d 137.9, 137.5, 137.0, 128.6, 128.5, 128.4, 128.3, 128.2, 128.1,

E.-L. Tsou et al. / Tetrahedron 65 (2009) 93–100
99
128.0, 127.8, 118.0, 83.5, 83.0, 73.3, 72.6, 72.1, 70.7, 62.8, 51.3. HRMS
4.6.3. ((2S,3R,4R,5R)-1-Benzyl-3,4-bis(benzyloxy)-5-(benzyloxy-
methyl)pyrrolidin-2-yl)methanol (24)
calcd for [C27H28N2O3þH]þ 429.2173, found 429.2194.
20
Compound 20b: [
a]
þ35.8 (c 0.8, CHCl3); 1H NMR (600 MHz,
A mixture of 22 (0.1 g, 0.18 mmol), di-tert-butyldicarbonate
(0.16 g, 0.73 mmol), and DMAP (0.1 g, 0.81 mmol) in CH2Cl2
(10 mL) was stirred at rt for 1 h. The reaction mixture was filtered
through a silica gel pad and concentrated to give a colorless liquid
23, which was used without further purification. A mixture of 23
and NaBH4 (0.02 g, 0.52 mmol) in methanol was stirred at 0 ꢀC
under Ar(g) for 8 h. The solvent was removed and the mixture was
extracted with CH2Cl2 (10 mLꢂ3). The combined organic layers
were dried with MgSO4, concentrated, and purified by CC (25%
D
CDCl3)
d
3.47 (m, 2H), 3.58 (q, 1H, J¼5.6 Hz), 3.83 (dd, 1H, J¼3.1,
5.8 Hz), 4.01 (d, 1H, J¼2.9 Hz), 4.2 (t, 1H, 3.0 Hz), 4.44–4.58 (m, 6H),
7.24–7.36 (m, 15H); 13C NMR (150 MHz, CDCl3)
d
137.9, 137.7, 136.9,
128.9, 128.7, 128.5, 128.2, 128.1, 128.03, 128.00, 127.9, 119.4, 87.5,
84.3, 73.5, 72.7, 72.4, 68.8, 62.4, 52.0. HRMS calcd for
[C27H28N2O3þH]þ 429.2173, found 429.2184.
4.5.3. (2S,3R,4R,5R)-2-(Aminomethyl)-5-(hydroxymethyl)-
pyrrolidine-3,4-diol (12)
EtOAc in hexanes, silica gel) to give 24 (63 mg, 0.12 mmol, 65%
20
A mixture of 20a (0.1 g, 0.23 mmol), palladium hydroxide
(0.01 g), and a catalytic amount of acetic acid in methanol (2 mL)
was stirred under a hydrogen atmosphere for 48 h. The reaction
mixture was filtered through Celite and the filtrate was concen-
trated. The residue was purified by CC (25% aqueous NH4OH (37%)
from 22) as a colorless syrup. [
(600 MHz, CDCl3)
a]
ꢁ0.8 (c 2, CHCl3); 1H NMR
D
d
3.04 (br, 1H), 3.15 (br, 1H), 3.31 (dd, 1H, J¼2.5,
9.2 Hz), 3.41 (m, 2H), 3.58 (d, 1H, J¼10.7 Hz), 3.85 (m, 2H), 4.06
(dd, 1H, J¼5.8, 6.9 Hz), 4.16 (t, 1H, J¼5.3 Hz), 4.38 (q, 2H), 4.47–4.56
(m, 3H), 4.64 (d, 1H, J¼11.8 Hz), 7.24–7.32 (m, 20H); 13C NMR
in propanol, silica gel) to give 12 (0.03 g, 0.18 mmol, 80%) as
(150 MHz, CDCl3) d 139.3, 138.6, 138.4, 138.1, 129.2, 128.7, 128.5,
20
a yellow syrup. [
d
a
]
D
þ14.7 (c 0.8, H2O); 1H NMR (600 MHz, D2O)
128.0, 127.9, 127.89, 127.81, 127.7, 127.4, 83.7, 82.4, 73.3, 72.4, 72.4,
70.0, 66.8, 64.6, 60.5, 59.0. HRMS calcd for [C34H37NO4þH]þ
524.2795, found 524.2798.
3.09 (dd, 1H, J¼6.3, 13.1 Hz), 3.13 (q, 1H, J¼5.5 Hz), 3.24 (dd, 1H,
J¼6.0, 13.1 Hz), 3.59 (q, 1H, J¼6.1 Hz), 3.64 (dd, 1H, J¼6.5, 11.6 Hz),
3.74 (dd, 1H, J¼4.5, 11.6 Hz), 3.90 (dd, 1H, J¼3.8, 5.3 Hz), 4.20 (dd,
1H, J¼3.8, 5.8 Hz); 13C NMR (150 MHz, D2O)
d
77.9, 76.8, 64.0, 62.2,
4.6.4. (2S,3R,4R,5R)-2,5-Bis(hydroxymethyl)pyrrolidine-3,4-diol
(13, DGDP)
55.9, 39.2. HRMS calcd for [C6H14N2OþH]þ 163.1077, found
163.1074.
A mixture of 24 (63 mg, 0.12 mmol), palladium hydroxide
(0.02 g), and a catalytic amount of hydrochloric acid (36% in H2O,
0.1 mL) in methanol (2 mL) was stirred under a hydrogen atmo-
sphere for 48 h. The reaction solution was filtered through a Celite
pad and the filtrate was concentrated. The residue was purified by
CC to give 13 (quantitative yield) as a yellowish syrup. The spectra
of 13 are consistent with those reported for 1b (see Section 4.3.7).
4.6. Typical procedure for the inversion of C-2 configuration
and the preparation of iminocyclitol 13 (20a/21/22/23/
24/13)
4.6.1. (2S,3R,4R,5R)-1-Benzyl-3,4-bis(benzyloxy)-5-(benzyloxy-
methyl)pyrrolidine-2-carbonitrile (21)
4.7. Typical procedure for selective acetylation of
iminocyclitol 9
A mixture of 20a (0.12 g, 0.28 mmol), benzylbromide (0.04 mL,
0.33 mol), potassium carbonate (0.077 g, 0.56 mmol), and potas-
sium iodide (4 mg, 0.02 mmol) in THF (3 mL) was stirred at 55 ꢀC for
overnight. The reaction mixture was filtered through Celite and the
filtrate was concentrated. The residue was purified by CC (20% EtOAc
4.7.1. N-(((2S,3S,4S,5S)-3,4-Dihydroxy-5-(hydroxymethyl)-
pyrrolidin-2-yl)methyl)acetamide (26)
A mixture of 9 (10.2 mg, 0.06 mmol) and acetic anhydride
(6.1 mg, 0.06 mmol) was stirred at rt for 10 min. The reaction
mixture was neutralized with Dowex 550A (OHꢁ) anion exchange
resin. The mixture was filtered and washed with MeOH (2 mL). The
filtrate was concentrated and the residue was purified by CC (12%
in hexanes, silica gel) to give 21 (0.13 g, 0.25 mmol, 90%) as a color-
20
less syrup. [
a]
ꢁ7.7 (c 1, CHCl3); 1H NMR (600 MHz, CDCl3)
d 3.21
D
(m,1H), 3.44 (dd,1H, J¼5.3, 9.5 Hz), 3.54 (dd,1H, J¼8.2, 9.4 Hz), 3.88
(d,1H, J¼5.3 Hz), 3.95 (m, 2H), 4.03 (dd,1H, J¼2.2, 5.2 Hz), 4.15 (d,1H,
J¼13.7 Hz), 4.42–4.61 (m, 6H), 7.22–7.40 (m, 20H); 13C NMR
aqueous NH4OH (37%) in propanol, silica gel) to give 26 (11.8 mg,
20
(150 MHz, CDCl3)
d
137.9, 137.4, 136.8, 135.7, 129.5, 128.4, 128.3,
92%) as a white solid. [
a
]
D
ꢁ30.5 (c 0.5, MeOH); 1H NMR (600 MHz,
128.29, 128.24, 128.1, 128.0, 127.9, 127.8, 127.7, 127.65, 127.61, 127.58,
127.53, 117.0, 82.9, 81.2, 73.0, 72.2, 71.2, 70.1, 66.0, 56.7, 56.0. HRMS
calcd for [C34H34N2O3þH]þ 519.2642, found 519.2665.
D2O) d 2.09 (s, 3H), 3.18 (m, 1H), 3.24 (m, 1H), 3.42 (m, 1H), 3.52 (dd,
1H, J¼4.7, 13.9 Hz), 3.74 (dd, 1H, J¼6.1, 11.1 Hz), 3.82 (m, 1H), 3.90 (t,
1H, J¼6.7 Hz), 3.94 (t, 1H, J¼6.8 Hz); 13C NMR (150 MHz, D2O)
d
174.6, 79.2, 77.5, 61.7, 61.6, 59.7, 41.6, 21.8. HRMS calcd for
4.6.2. (2R,3R,4R,5R)-1-Benzyl-3,4-bis(benzyloxy)-5-(benzyloxy-
methyl)pyrrolidine-2-carboxamide (22)
[C8H16N2O4þH]þ 205.1183, found 205.1179.
A mixture of 21 (0.11 g, 0.22 mmol), aqueous sodium hydroxide
(1 N, 3 mL), and aqueous hydrogen peroxide (2 mL, 30%) in MeOH
(2 mL) was stirred at rt for 3 h. The reaction mixture was extracted
with CH2Cl2 (20 mLꢂ3), washed with brine, dried with MgSO4, and
concentrated. The residue was purified by CC (20% EtOAc in hex-
4.7.2. N-(((2R,3R,4S,5R)-3,4-Dihydroxy-5-(hydroxymethyl)-
pyrrolidin-2-yl)methyl)acetamide (27)
Following the procedure for the preparation of 25, 27 was
obtained as a yellow syrup (93%). [
(600 MHz, D2O)
a]
þ18 (c 0.1, MeOH); 1H NMR
20
D
d
2.08 (s, 3H), 3.56 (dd, 1H, J¼7.1, 14.1 Hz), 3.62 (m,
anes, silica gel) to give 22 (0.1 g, 0.18 mmol, 83%) as a colorless
1H), 3.67 (dd, 1H, J¼3.6, 14.2 Hz), 3.74 (m, 1H), 3.88 (dd, 1H, J¼8.0,
11.9 Hz), 3.99 (dd, 1H, J¼5.5, 11.9 Hz), 4.20 (dd, 1H, J¼3.9, 8.8 Hz),
20
syrup. [
a
]
D
þ11.0 (c 2, CHCl3); 1H NMR (600 MHz, CDCl3)
d 3.11 (dd,
1H, J¼3.7, 9.4 Hz), 3.18 (t, 1H, J¼3.3 Hz), 3.41 (dd, 1H, J¼8.0, 9.1 Hz),
3.75 (d, 1H, J¼13.1 Hz), 3.83 (d, 1H, J¼6.2 Hz), 3.92 (d, 1H,
J¼13.1 Hz), 4.08 (s, 1H), 4.23 (dd, 1H, J¼3.2, 5.3 Hz), 4.27 (d, 1H,
J¼11.9 Hz), 4.33 (d, 1H, J¼11.9 Hz), 4.43 (d, 1H, J¼11.6 Hz), 4.50 (d,
1H, J¼12.1 Hz), 4.59 (d, 1H, J¼12.0 Hz), 4.63 (d, 1H, J¼11.6 Hz), 5.90
4.34 (t, 1H, J¼3.7 Hz); 13C NMR (150 MHz, D2O)
d 175.7, 73.3, 70.4,
61.4, 60.3, 58.2, 39.2, 21.7. HRMS calcd for [C8H16N2O4þH]þ
205.1183, found 205.1185.
4.8. General procedure for the enzyme assay with various
glycosidases
(s,1H), 7.18–7.35 (m, 20H); 13C NMR (150 MHz, CDCl3)
d 174.3,138.2,
138.0, 137.9, 129.5, 128.6, 128.5, 128.45, 128.42, 128.0, 127.8, 127.77,
127.74, 127.69, 83.1, 82.0, 73.0, 72.7, 71.7, 70.7, 69.9, 67.3, 59.6. HRMS
calcd for [C34H36N2O4þH]þ 537.2748, found 537.2742.
The initial velocities of hydrolysis at rt were measured spec-
trophotometrically at various concentrations of p-nitrophenyl-

100
E.-L. Tsou et al. / Tetrahedron 65 (2009) 93–100
O. R. Curr. Top. Med. Chem. 2003, 3, 541; (c) Zou, W. Curr. Top. Med. Chem. 2005,
5, 1363 and references therein; (d) Watson, A. A.; Fleet, G. W. J.; Asano, N.;
Molyneux, R. J.; Nash, R. J. Phytochemistry 2001, 56, 265 and references therein;
(e) Wrodnigg, T. M. Monatsh. Chem. 2002, 133, 393; (f) Winchester, B.; Fleet, G.
W. J. Glycobiology 1992, 2, 199.
2. For selected examples of synthesis of DMDP, see: (a) Fleet, G. W. J.; Smith, P. W.
Tetrahedron Lett. 1985, 26, 1469; (b) Fleet, G. W. J.; Smith, P. W. Tetrahedron 1987,
43, 971; (c) Donohoe, T. J.; Headley, C. E.; Cousins, R. P. C.; Cowley, A. Org. Lett.
2003, 5, 999.
3. Dondoni, A.; Giovannini, P. P.; Perrone, D. J. Org. Chem. 2002, 67, 7203.
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Watson, A. A.; Nash, R. J.; Fleet, G. W. J. J. Nat. Prod. 1998, 61, 625.
5. For the synthesis of ADMDP, see: (a) Wrodnigg, T. M.; Stu¨tz, A. E.; Withers, S. G.
Tetrahedron Lett. 1997, 38, 5463; (b) Liu, J.; Numa, M. M. D.; Liu, H.; Huang, S. J.;
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glycopyranoside (40 mM, 20 mM, 10 mM, 5 mM, 2.5 mM, 1.25 mM,
0.625 mM, 0 mM) at 405 nm by using multi-detection reader
(SpectraMax M5, Molecular Dectce). The data obtained was fitted to
the Michaelis–Menten equation by using the GraphPad to de-
termine the Km values and Vmax values. The substrate concentra-
tions were used at twofold Km values for evaluation of the
inhibitory effect against various glycosidases. Enzymes at 0.25–
1 units per mL were used to provide an ideal progression curve, and
inhibitors were tested initially at 500 mM. The compounds that
showed activities were selected and further tested at lower con-
centration to determine IC50. The assays performed in wells of the
96-well microtiter plate contained either sodium phosphate buffer
(100 mM, pH 6.8, for
(100 mM, pH 6.4, for
a
a
- and
-mannosidase).
b-glucosidase) or sodium citrate buffer
7. Liang, P. H.; Cheng, W. C.; Lee, Y. L.; Yu, H. P.; Wu, Y. T.; Lin, Y. L.; Wong, C. H.
ChemBioChem 2006, 7, 165.
8. For the synthesis of
L-DMDP, see: Yu, C.-Y.; Asano, N.; Ikeda, K.; Wang, M.-X.;
Butters, T. D.; Wormald, M. R.; Dwek, R. A.; Winters, A. L.; Nash, R. J.; Fleet, G.
W. J. Chem. Commun. 2004, 1936.
9. Kumar, V.; Ramesh, N. G. Tetrahedron 2006, 62, 1877.
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11. Gautier-Lefebvre, I.; Behr, J.-B.; Guillerm, G.; Muzard, M. Eur. J. Med. Chem. 2005,
40, 1255.
4.8.1. Kinetic analysis of
Incubations were performed in a total volume of 100
action mixture contained citrate sodium buffer solution (100 mM,
pH 4.5), various amount of 4-methylumbellifery N-acetyl- -glu-
cosaminide, and various amounts of inhibitors with 0.5 mU per
well of -N-acetylglucosaminidase. After incubation for 30 min at
b-N-acetylglucosaminidase
m
L. Re-
b-D
b
rt, the reaction was terminated by addition of sodium glycine buffer
(0.5 M), pH 10.5. Enzyme activity was measured by the release of 4-
methylumbelliferone with an excitation wavelength of 360 nm and
an emission wavelength of 460 nm.
12. (a) Cicchi, S.; Goti, A.; Brandi, A. Synthesis 2007, 485; (b) Lombardo, M.; Fabbroni,
S.; Trombini, C. J. Org. Chem. 2001, 66, 1264; (c) Goti, A.; Cicci, S.; Mannucci, V.;
Cardona, F.; Guarna, F.; Merino, P.; Tejero, T. Org. Lett. 2003, 5, 4235 and references
therein; (d) Merino, P.; Tejero, T.; Revuelta, K.; Romero, P.; Cicchi, S.; Mannucci, V.;
Brandi, A.; Goti, A. Tetrahedron: Asymmetry 2003, 14, 367.
13. (a) Holzapfel, C. W.; Crous, R. Heterocycles 1998, 48, 1337; (b) Gurjar, M. K.;
Borhade, R. G.; Puranik, V. G.; Ramana, C. V. Tetrahedron Lett. 2006, 47, 6979.
14. Marradi, M.; Cicchi, S.; Delso, J. I.; Rosi, L.; Teiero, T.; Merino, P.; Goti, A. Tet-
rahedron Lett. 2005, 46, 1287.
15. (a) Carmona, A. T.; Whigtman, R. H.; Robina, I.; Vogel, P. Helv. Chim. Acta 2003,
86, 3066; (b) Tamura, O.; Toyao, A.; Ishibashi, H. Synlett 2002, 1344.
16. See Section 4.
Acknowledgements
This work was supported by the National Science Council and
Academia Sinica.
17. (a) Desvergnes, S.; Py, S.; Vallee, Y. J. Org. Chem. 2005, 70, 1459; (b) Toyao, A.;
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Supplementary data
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.tet.2008.10.096.
19. (a) Tejima, S.; Fletcher, H. G. J. Org. Chem. 1963, 28, 1044; (b) Barker, R.; Fletcher,
H. G. J. Org. Chem. 1961, 26, 4605.
References and notes
20. Wuts, P. G. M.; Jung, Y. W. J. Org. Chem. 1988, 53, 5989.
21. Katritzky, A. R.; Pilarski, B.; Urogdi, L. Synthesis 1989, 949.
22. Ragnarsson, U.; Grehn, L.; Monteriro, L. S.; Maia, H. L. S. Synlett 2003,
2386.
1. For recent reviews, see: (a) Stu¨tz, A. E. Iminosugars as Glycosidase Inhibitors:
Nojirimycin and Beyond; Wiley-VCH: Weinheim, 1999; (b) Compain, P.; Martin,