Synthesis of polyhydroxylated azetidine iminosugars and 3-hydroxy-N-methylazetidine-2-carboxylic acid from d-glucose

Article abstract of DOI:10.1016/j.tet.2015.05.057

The azetidine iminosugars, (2S,3R)-2-((R)-1,2-dihydroxyethyl)azetidin-3-ol 3, (2R,3R)-2-(hydroxymethyl)azetidin-3-ol 4, and (2S,3R)-3-hydroxy-N-methylazetidine-2-carboxylic acid 5 were synthesized from d-glucose derived 3-N-benzyloxycarbonyl-3-deoxy-1,2-O

Full text of DOI:10.1016/j.tet.2015.05.057

Tetrahedron 71 (2015) 5085e5090
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Tetrahedron
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Synthesis of polyhydroxylated azetidine iminosugars and
3-hydroxy-N-methylazetidine-2-carboxylic acid from -glucose
D
Pravin P. Lawande a, Vyankat A. Sontakke a, Roopa J. Nair b, Ayesha Khan c,
,
Sushma G. Sabharwal b, Vaishali S. Shinde a
*
a Garware Research Centre, Department of Chemistry, Savitribai Phule Pune University (formerly, University of Pune), Pune 411007, India
b Division of Biochemistry, Department of Chemistry, Savitribai Phule Pune University (formerly, University of Pune), Pune 411007, India
c Department of Chemistry, Savitribai Phule Pune University (formerly, University of Pune), Pune 411007, India
a r t i c l e i n f o
a b s t r a c t
Article history:
The azetidine iminosugars, (2S,3R)-2-((R)-1,2-dihydroxyethyl)azetidin-3-ol 3, (2R,3R)-2-(hydroxymethyl)
Received 4 December 2014
Received in revised form 16 May 2015
Accepted 18 May 2015
azetidin-3-ol 4, and (2S,3R)-3-hydroxy-N-methylazetidine-2-carboxylic acid 5 were synthesized from
D-
glucose derived 3-N-benzyloxycarbonyl-3-deoxy-1,2-O-isopropylidene- -xylofuranose 8 using intra-
a-D
molecular Mitsunobu reaction as a key step. The glycosidase inhibitory activity of compounds 3e5 was
screened against various enzymes. Amongst all synthesized compounds, the N-methylated compound 5
exhibited significant inhibitory activity against amyloglucosidase from Aspergillus niger in micro molar
range.
Available online 22 May 2015
Keywords:
Azetidine
Iminosugars
Ó 2015 Elsevier Ltd. All rights reserved.
Glycosidase inhibitory activity
Mitsunobu reaction
D
-Glucose
1. Introduction
compounds starting from non-carbohydrate precursors,9 however,
there are few reports for synthesis using sugars as starting mate-
The polyhydroxylated nitrogen containing cyclic compounds
commonly known as iminosugars, are found to be selective and
promising glycosidase inhibitors.1,2 Mechanistically, these small
molecules act as mimic for the carbohydrates, thereby exhibiting
remarkable biological activities. In an attempt to know the effect of
ring size on biological activity, several iminosugars with varying
ring size (5e8 membered) were either isolated or synthesized3 and
evaluated for various biological activities.4
rials.10 For example, Jager and co-workers synthesized azetidine
iminosugars from 2-O-benzylglyceraldehyde11 using [2þ2] cyclo-
addition of acid chlorides with 2-O-benzylglyceraldehyde imines.
Recently, Fleet and co-workers described the synthesis of azetidine
iminosugars via nucleophilic double-displacement of O-triflate
Azetidines are four-membered aza-heterocycles5 containing
endocyclic nitrogen atom and known to show wide range of bi-
ological activity.6 For example,
L
-azetidine-2-carboxylic acid (
1a is naturally occurring non-proteinogenic amino acid found in
plants7 and is known to be ring contracted derivative of
-proline 2
L-Aze)
L
(Fig. 1). Ozaki and co-workers8 reported the enzymatic synthesis of
cis-3-hydroxyazetidine-2-carboxylic acid 1b from 1a. Synthesis of
polyhydroxylated azetidines is less accounted by the researchers
due to the difficulties faced in the formation of enantiopure, highly
strained four-membered ring. A number of synthetic methods have
been developed for the formation of hydroxylated azetidine
* Corresponding author. Tel.: þ91 20 25601395; fax: þ91 20 25691728; e-mail
address: vsshinde@chem.unipune.ac.in (V.S. Shinde).
Fig. 1. Azetidine analogues and other related compounds.
http://dx.doi.org/10.1016/j.tet.2015.05.057
0040-4020/Ó 2015 Elsevier Ltd. All rights reserved.

5086
P.P. Lawande et al. / Tetrahedron 71 (2015) 5085e5090
derivatives of pentoses12 as well as hexoses13 by amines. Georg and
co-workers described synthesis of N-alkylated azetidine iminosu-
gars from glyceraldehyde and tested for inhibitory activity against
glucosyltransferase and other glycosidases.14
Dowex (Hþ) gave hemiacetal that was directly subjected to oxida-
tive cleavage using sodium metaperiodate, followed by reduction
using sodium borohydride in methanol to afford N-Cbz protected
diol 11 as a thick liquid. Finally, hydrogenolysis using 10% Pd/C in
methanol at 10 psi afforded 4 in 57% yield as a sticky solid.
To achieve the synthesis of (2S,3R)-3-hydroxy-N-methyl-
azetidine-2-carboxylic acid 5, the hemiacetal obtained from 9 was
treated with sodium metaperiodate, followed by the Pinnick oxi-
dation15b using sodium chlorite and hydrogen peroxide to afford N-
Cbz protected acid 12. Final hydrogenolysis of compound 12 in the
presence of 10% Pd/C in methanol failed to furnish expected free
amine. However, we obtained N-methylated azetidine-2-carboxylic
acid 5 in 50% yield as a sticky solid. The structure was confirmed on
the basis of HRMS, NMR, and 2D-HSQC spectral data. The presence
As a part of our continuing efforts in the synthesis of bioactive
molecules from
D
-glucose,15 we report herein, synthesis of azeti-
dine iminosugars namely, (2S,3R)-2-((R)-1,2-dihydroxyethyl)azeti-
din-3-ol 3, (2R,3R)-2-(hydroxymethyl)azetidin-3-ol 4, and (2S,3R)-
3-hydroxy-N-methylazetidine-2-carboxylic acid 5 and their glyco-
sidase inhibitory activity. In our approach (as shown in Scheme 1),
we constructed azetidine skeleton of target molecules by using
intramolecular Mitsunobu reaction of the C5eOH group with C3-
protected amino (from C3-azido) functionality of
results in this direction are described herein.
D-glucose. Our
Scheme 1. Reagent and conditions: (a) NaBH4, MeOH/H2O, 0 ꢀC, 2 h, 97%; (b) (i) H2, Pd/C, MeOH, rt, 3 h, 10 psi; (ii) NaHCO3, CbzCl, MeOH/H2O, 0 ꢀC, 4 h, 93%; (c) PPh3, DEAD, Dry
DCM, rt, 3 h, 65%; (d) Dowex 50WX4-200 (Hþ form), 1,4-dioxane/H2O, 60 ꢀC, 24 h, 93%; (e) NaBH4, THF/H2O, ꢂ10 ꢀC, 1 h, 88%; (f) H2, Pd/C, MeOH, rt, 10 psi, 12 h, 81% for 3, 57% for 4
and 50% for 5; (g) NaIO4, acetone/H2O, 0 ꢀC, 6 h; (h) NaBH4, THF:H2O, ꢂ10 ꢀC, 1 h, 56% (from hemiacetal); (i) NaClO2, NaH2PO4, 30% H2O2, aq MeCN, 0 ꢀC to rt, 10 h, 66% (from
hemiacetal).
2. Result and discussion
of sharp singlet at 2.92 ppm, corresponding to three protons in-
tegration in the 1H NMR and a peak at 40.2 ppm in the 13C NMR
spectrum indicated the presence of eNCH3 group.16
2.1. Synthesis
Required azido-aldehyde 6 was synthesized from commercially
2.2. Glycosidase inhibitory activity
available
group in
D
-glucose as reported earlier (Scheme 1).15b The aldehyde
6
was reduced by using sodium borohydride in
The glycosidase inhibitory activity of newly synthesized com-
aq methanol at 0 ꢀC to give azido alcohol 7 in 97% yield. Reduction
of C3-azido functionality in 7 using 10% Pd/C in methanol afforded
corresponding amine, which was protected as carbamate derivative
8 by treatment with benzyl chloroformate (CbzCl) in the presence
of sodium bicarbonate. Further, reaction of 8 with triphenylphos-
phine and diethyl azodicarboxylate (DEAD) under Mitsunobu
condition gave key azetidine ring 9 in 65% yield as a white solid.
Deprotection of 1,2-acetonide group of 9 with the Dowex (Hþ) resin
pounds 3e5 was studied against various glycosidases viz.:
cosidase, -mannosidase, and -galactosidase (isolated from
almond seeds), -mannosidase, -mannosidase, and N-acetyl-
glucosaminidase (isolated from Jack bean seeds), -glucosidase
(isolated from rice seeds and yeast, procured from Sigma Chemical
Co), -glucosidase and -galactosidase (isolated from Bovine liver),
-galactosidase (isolated from coffee bean seeds), -galactosidase
(Aspergillus oryzae from Sigma Chemical Co), -fucosidase and N-
acetyl- -glucosaminidase (isolated from Bovine kidney), -amy-
lase (isolated from Geobacillus JN704808) and amyloglucosidase
(Aspergillus niger procured from Sigma Chemical Co). The 50% in-
hibitory concentration (IC50) and inhibitory constant (Ki) values
were determined for the test compounds and are summarized in
Table 1. Amongst these compounds, 4 showed a weak competitive
b-glu-
a
a
a
b
b-D-
a
b
b
a
b
a
afforded anomeric mixture of hemiacetals (
a/b
¼2:1; as evident
b
-D
a
from the 1H NMR spectrum of the crude product), which was re-
duced by using sodium borohydride in THF/H2O to give N-Cbz
protected triol 10. In the final step, hydrogenolysis of 10 using 10%
Pd/C in methanol afforded (2S,3R)-2-((R)-1,2-dihydroxyethyl)aze-
tidin-3-ol 3 in 81% yield.
Synthesis of (2R,3R)-2-(hydroxymethyl)azetidin-3-ol
achieved from 9. Thus, deprotection of 1,2-acetonide group using
4
was
inhibition of
and Ki 2.66 mM.
b
-galactosidase (bovine liver) with IC50 7.5ꢁ0.05 mM

P.P. Lawande et al. / Tetrahedron 71 (2015) 5085e5090
5087
Table 1
Inhibitory activity of azetidine iminosugars against various glycosidases
Enzyme
IC50
IC50
IC50
a
-Glucosidase
Rice
Yeast
NI(0%)
NI(0%)
NI(12.70%)
NI(0%)
NI(0%)
NI(0%)
b
-Glucosidase
Almond
Bovine liver
NI(0%)
NI(0%)
NI(0%)
NI(0%)
NI(0%)
NI(0%)
a
- Galactosidase
Coffee bean
Almond
I(50.87%)
NI(23.41%)
NI(30.46%)
NI(0%)
NI(0%)
NI(0%)
b
-Galactosidase
Bovine liver
Aspergillus oryzae
NI(15.44%)
NI(0%)
7.5ꢁ0.05 mM
NI(15.3%)
NI(0%)
NI(0%)
a
-Mannosidase
Jackbean
Almond
NI(0%)
NI(0%)
NI(0%)
NI(25.61%)
NI(0%)
NI(0%)
b
-Mannosidase
Jackbean
NI(0%)
NI(0%)
NI(0%)
NI(0%)
NI(0%)
a
-Fucosidase
Bovine kidney
NI(16.10%)
NI(0%)
NI(7.22%)
NI(0%)
a
-Amylase
Geobacillus
N-Acetyl-b-D-glucosaminidase
Jackbean
Bovine Kidney
NI(8.48%)
NI(0%)
NI(0%)
NI(0%)
NI(0%)
NI(0%)
Amyloglucosidas
Aspergillus niger
360ꢁ4
m
M
432ꢁ5
m
M
12.32ꢁ0.68
mM
NI: no inhibition (less than 50% inhibition at 10 mM).
I: inhibition obtained.
( ): % inhibition at 10 mM.
Compound 3 showed good inhibition against amyloglucosidase
4. Experimental
4.1. General
from A. niger, (IC50 360ꢁ4
mM and Ki 32 mM), which is in line with
earlier report by Jager and Fleet.11,12 Removal of hydroxymethyl
group as in 4 slightly reduced inhibition of amyloglucosidase (IC50
432ꢁ5
hydroxy-N-methylazetidine-2-carboxylic acid 5 exhibited signifi-
M and Ki 2.68 M). All
test compounds showed competitive inhibition against
amyloglucosidase.
m
M and Ki 56
m
M). Compared to other test compounds, 3-
Melting points were recorded with Thomas Hoover Capillary
melting point apparatus and are uncorrected. Infrared Spectra were
recorded on a Bruker Tensor 37 FTIR-ATR as a neat and are
expressed in cmꢂ1. Nuclear magnetic resonance (NMR) spectra
cantly enhanced activity (IC50 12.32ꢁ0.68
m
m
were recorded on a Bruker Advanced III HD (1H at 500 MHz and 13
C
at 125 MHz) and Varian Mercury instrument (1H at 300 MHz and
13C at 75 MHz) in CDCl3 or D2O as the solvent. Chemical shifts were
reported in
d unit (ppm) with reference to TMS as an internal
3. Conclusions
standard and J values were given in hertz (Hz). The DEPT and HSQC
experiments confirmed the assignments of the signals wherever
required. High-resolution mass spectra (HRMS) were carried on
a Bruker Impact HD with electron-spray ionization (ESI) technique.
Optical rotations were measured using Jasco P1020 polarimeter
with sodium light (589.3 nm) at 29 ꢀC. Thin layer chromatography
was performed on pre-coated aluminum sheets (0.25 mm, silica gel
In summary, we have utilized intramolecular Mitsunobu re-
action to synthesize enantiopure azetidine iminosugars 3, 4, and 3-
hydroxy-N-methylazetidine-2-carboxylic acid
5 starting from
readily available inexpensive -glucose. Compound 5 showed
D
maximum inhibition activity against amyloglucosidase from A.
niger in micro molar range.

5088
P.P. Lawande et al. / Tetrahedron 71 (2015) 5085e5090
60 F254). Visualization was made by absorption of UV light or by
thermal development after spraying with 3.5% solution of 2,4-
dinitrophenylhydrazine in ethanol/H2SO4 or with basic
aq potassium permanganate solution.
DCM was evaporated under vacuum and crude product was sub-
jected for column chromatography purification (10% ethyl acetate/
hexane) to afford 9 (0.650 g, 65%) as a white solid. Mp¼93e95 ꢀC;
Rf¼(30% ethyl acetate/hexane) 0.65; [
a]
29 ꢂ93.0 (c 0.60, CHCl3); IR
D
(neat, nmax, cmꢂ1) 1699 (CO); 1H NMR (500 MHz, CDCl3)
d 1.30 (s,
4.2. 3-Azido-3-deoxy-1,2-O-isopropylidene-
(7)
a
-
D
-xylofuranose
3H, CH3), 1.38 (s, 3H, CH3), 3.60 (d, 1H, J¼10.1 Hz, H5), 3.97 (dd, 1H,
J¼10.1, 5.0 Hz, H50), 4.65e4.80 (m, 2H, H2 and H3), 4.84 (td, 1H,
J¼4.8, 1.6 Hz, H4), 5.04 (s, 2H, OCH2Ph), 6.06 (d, 1H, J¼3.5 Hz, H1),
To an ice-cooled solution of azido-aldehyde
6
(3.0 g,
7.21e7.33 (m, 5H, Ar); 13C NMR (125 MHz, CDCl3)
d 27.4 (CH3), 28.1
14.0 mmol) in MeOH/H2O (20 mL, 9:1), sodium borohydride
(0.799 g, 21.1 mmol) was added in portions. The reaction mixture
was stirred for 2 h. After completion of reaction (monitored by
TLC), it was quenched by addition of satd aq NH4Cl solution
(5 mL). Methanol was evaporated under reduced pressure. The
residue was extracted with ethyl acetate (3ꢃ50 mL) and organic
fraction was dried over sodium sulfate. Organic layer was evapo-
rated under vacuum and crude product was subjected for column
chromatography purification (20% ethyl acetate/hexane) afforded
7 (2.940 g, 97%) as thick liquid. Rf¼(40% ethyl acetate/hexane)
(CH3), 57.4 (C5), 67.3 (OCH2Ph), 69.8 (C3), 74.1 (4), 83.2 (C2), 108.6
(C1), 114.3 (OCO), 128.4, 128.5, 128.9, 136.4 (Ar), 156.2 (COOCH2Ph);
HRMS (ESI): m/z calculated for C16H19NNaO5 [MþNaþ]: 328.1155,
found: 328.1156.
4.5. (2S,3R)-N-Benzyloxycarbonyl-3-hydroxy-2-((R)-1,2-
dihydroxyethyl)azetidine (10)
To a stirred solution of 9 (1.0 g, 3.27 mmol) in 1,4-dioxane/H2O
(16 mL, 5:5) was added Dowex (acidic resin, 2.0 g) at room tem-
perature and reaction mixture was stirred at 60 ꢀC for 24 h while
allowing it to attend room temperature. After completion of re-
action (monitored by TLC), solvent was evaporated under reduced
pressure and filtered through a Celite pad. The residue was
extracted with ethyl acetate (4ꢃ20 mL) and organic fraction was
dried over sodium sulfate. Combined organic layer was evaporated
under vacuum and crude product was subjected for column chro-
matography purification (30% ethyl acetate/hexane) to afford
0.66; [
a
]
29 ꢂ36.6 (c 1.60, CHCl3); IR (nmax, cmꢂ1) 3300e3400 (OH),
D
2103 (N3); 1H NMR (300 MHz, CDCl3)
d 1.34 (s, 3H, CH3), 1.52 (s,
3H, CH3), 2.08 (br s, 1H, OH, exchangeable with D2O), 3.83 (dd, 1H,
J¼11.8, 5.6 Hz, H5), 3.92 (dd, 1H, J¼11.8, 6.1 Hz, H50), 4.02 (d, 1H,
J¼3.2 Hz, H3), 4.31e4.41 (m, 1H, H4), 4.67 (d, 1H, J¼3.8 Hz, H2),
5.93 (d, 1H, J¼3.8 Hz, H1); 13C NMR (75 MHz, CDCl3)
d 26.1 (CH3),
26.5 (CH3), 60.6 (C3), 65.9 (C5), 79.5 (C4), 83.5 (C2), 104.6 (C1),
112.3 (OCO); HRMS (ESI): m/z calculated for C8H13N3NaO4
[MþNaþ]: 238.0798, found: 238.0795.
anomeric mixture of hemiacetal (
a
/b
¼2:1; 0.815 g, 93% yield) as
4.3. 3-Amino-N-benzyloxycarbonyl-3-deoxy-1,2-O-iso-
propylidene-a-D-xylofuranose (8)
a thick liquid: Rf¼(60% ethyl acetate/hexane) 0.17. To a stirred so-
lution of hemiacetal (0.700 g, 2.64 mmol) in THF/H2O (25 mL, 9:1)
maintained at ꢂ10 ꢀC, sodium borohydride (0.149 g, 3.96 mmol)
was added in portions for 10 min. The resulting solution was stirred
for 1 h and allowed to attain room temperature. After completion of
the reaction (monitored by TLC), excess of hydride was quenched
with satd NH4Cl, solvent was evaporated, and the residue was
extracted with ethyl acetate (4ꢃ15 mL) and organic fraction was
dried over sodium sulfate. Evaporation of solvent and column pu-
A solution of 7 (2.940 g, 13.66 mmol) in dry methanol (25 mL)
and 10% Pd/C (0.200 g) was hydrogenated at 10 psi for 3 h at room
temperature. After completion, reaction mass was filtered through
a Celite pad, washed with methanol, and filtrate was evaporated
under reduced pressure to give amine (crude wt 2.480 g). The
amine was dissolved in methanol/water (30 mL, 9:1) and cooled to
0
ꢀC. To this sodium bicarbonate (2.754 g, 32.7 mmol) was added,
rification (40% ethyl acetate/hexane) gave 10 (0.620 g, 88%) as
29
followed by addition of CBzCl (2.80 ml, 19.66 mmol). The reaction
mixture was stirred for 4 h. Upon completion, methanol was
evaporated partially under reduced pressure and filtered through
a Celite pad. The filtrate was extracted with ethyl acetate (3ꢃ30 mL)
and organic fraction was dried over sodium sulfate. Combined or-
ganic layer was evaporated under vacuum and crude product was
then purified by column chromatography (30% ethyl acetate/hex-
a thick liquid. Rf¼(80% ethyl acetate/hexane) 0.35; [
a
]
ꢂ75.8 (c
D
0.42, CHCl3); IR (nmax, cmꢂ1) 3300e3500 (OH, br), 1668 (CO); 1H
NMR (300 MHz, CDCl3, 1 drop D2O)
d
3.67 (dd, 1H, J¼11.1, 4.9, Hz,
H6), 3.74e3.91 (m, 2H, H60/H4), 3.98e4.13 (m, 1H, H5), 4.21e4.35
(m, 1H, H40), 4.37e4.47 (m, 1H, H2) 4.57e4.72 (m, 1H, H3), 5.10
(2ABd, 2H, J¼12.1 Hz, OCH2Ph), 7.30e7.48 (m, 5H, Ar); 13C NMR
(75 MHz, CDCl3)
d 60.4 (C4), 63.6 (C6), 64.4 (C3), 67.7 (OCH2Ph),
ane) to give 8 (3.960 g, 93%) as a white solid. Mp¼110e112 ꢀC;
69.7 (C2), 70.4 (C5), 128.2, 128.4, 128.6, 135.8 (Ar), 158.4
(COOCH2Ph); HRMS (ESI): m/z calculated for C13H17NNaO5
[MþNaþ]: 290.0999, found: 290.1002.
29
Rf¼(60% ethyl acetate/hexane) 0.83; [
a]
ꢂ3.4 (c 0.70, CHCl3); IR
D
(
nmax, cmꢂ1) 3300e3500 (OH, NH), 1723(CO); 1H NMR (500 MHz,
CDCl3)
d 1.30 (s, 3H, CH3), 1.51 (s, 3H, CH3), 2.57 (br s, 1H, OH, ex-
changeable with D2O), 3.76e3.86 (m, 1H, H5), 3.89e4.01 (m, 1H,
H50), 4.19 (dd, 1H, J¼6.3, 4.0 Hz, H3), 4.27e4.35 (m, 1H, H4), 4.60 (d,
1H, J¼3.5 Hz, H2), 5.11 (2ABd, 2H, J¼12.3 Hz, OCH2Ph), 5.85 (d, 1H,
4.6. (2S,3R)-2-((R)-1,2-Dihydroxyethyl)azetidin-3-ol (3)
J¼3.5 Hz, H1), 6.03 (d, 1H, J¼4.8 Hz, NH), 7.30e7.42 (m, 5H, Ar); 13
C
A solution of 10 (0.190 g, 0.711 mmol) and 10% Pd/C (0.020 g) in
methanol (8 mL) was stirred under H2 atmosphere at 10 psi for 12 h
at room temperature. After completion of reaction (monitored by
TLC), the catalyst was filtered through Celite pad. Column purifi-
NMR (125 MHz, CDCl3) d 26.2 (CH3), 26.6 (CH3), 58.7 (C3), 60.1 (C5),
67.1 (OCH2Ph), 77.4 (C4), 84.7 (C2), 104.3 (C1), 112.1 (OCO), 128.3,
128.3, 128.6 136.1 (Ar), 156.5 (COOCH2Ph); HRMS (ESI): m/z calcu-
lated for C16H21NNaO6 [MþNaþ]: 346.1261, found: 346.1263.
cation (25% MeOH/CHCl3) afforded 3 (0.076 g, 81%) as a semi solid.
29
Rf¼(30% MeOH/CHCl3) 0.33; [
a]
ꢂ6.14 (c 0.30, CH3OH); IR (nmax
,
D
4.4. N-Benzyloxycarbonyl-3,5-dideoxy-3,5-imino-1,2-O-iso-
cmꢂ1) 3300e3500 (OH, br), 2980 (NH); 1H NMR (500 MHz, D2O)
propylidene-
a-
D-xylofuranose (9)
d
3.56 (dd, 1H, J¼12.2, 5.2 Hz, H6), 3.68 (dd, 1H, J¼12.2, 3.6 Hz, H60),
3.89 (dd, 1H, J¼11.7, 5.1 Hz, H4), 4.23e4.29 (m, 1H, H5), 4.32 (dd, 1H,
To a stirred solution of 8 (1 g, 3.0 mmol) in dry DCM (25 mL)
were added triphenylphosphine (1.630 g, 6.1 mmol) and diethyl
azodicarboxylate (1.12 mL, 6.1 mmol) at room temperature. The
reaction mixture was stirred for 3 h. After completion of reaction,
J¼11.7, 7.1 Hz, H40), 4.55 (t, 1H, J¼7.0 Hz, H2), 4.69e4.77 (m, 1H,
under D2O peak, H3); 13C NMR (125 MHz, D2O)
d 53.4 (C4), 62.5
(C6), 63.3 (C3), 66.2 (C2), 66.5 (C5); HRMS (ESI): m/z calculated for
C5H12NO3 [MþHþ]: 134.0817, found: 134.0815.

P.P. Lawande et al. / Tetrahedron 71 (2015) 5085e5090
5089
4.7. (2R,3R)-N-Benzyloxycarbonyl 3-hydroxy-2-(hydrox-
ymethyl)azetidine (11)
(COOH), 1460 (CO); 1H NMR (500 MHz, CDCl3)
d
4.11 (dd, 1H, J¼9.6,
5.0 Hz, H4), 4.34 (dd, 1H, J¼9.5, 7.0 Hz, H40), 5.01 (d, 1H, J¼7.4 Hz,
H2), 5.13 (s, 2H, OCH2Ph), 5.55e5.68 (m, 1H, H3), 5.90 (br s, 2H, OH,
COOH, exchangeable with D2O), 7.27e7.39 (m, 5H, Ar); 13C NMR
To a stirred solution of hemiacetal (0.2 g, 0.754 mmol) in ace-
tone/water (10 mL, 9:1), sodium metaperiodate (0.242 g,
1.131 mmol) was added at 0 ꢀC and the reaction mixture was stirred
for 6 h at room temperature. Ethylene glycol (0.2 mL) was added in
order to consume unreacted sodium metaperiodate and solvent
was evaporated under reduced pressure. The reaction mixture was
extracted with chloroform (4ꢃ10 mL), followed by evaporation of
combined organic layer and concentrated under vacuum, to afford
intermediate X as a thick liquid (crude wt 0.190 g). To a stirred
solution of above crude product (0.190 g, 0.722 mmol) in THF/H2O
(10 mL, 9:1) maintained at ꢂ10 ꢀC, sodium borohydride (0.013 g,
0.361 mmol) was added in portions for 2 min. The resulting solu-
tion was stirred for 1 h and allowed to attain room temperature.
After completion (monitored by TLC) of reaction, excess of hydride
was quenched with satd NH4Cl, the solvent was evaporated and the
residue was extracted with ethyl acetate (5ꢃ15 mL) and organic
fraction was dried over sodium sulfate. Combined organic layer was
evaporated under vacuum and crude product was subjected for
column chromatography purification (25% ethyl acetate/hexane)
(125 MHz, CDCl3)
d 55.8 (C4), 62.4 (C3), 65.5 (C2), 67.8 (OCH2Ph),
128.2, 128.5, 128.7, 136.9 (Ar), 159.5 (COOCH2Ph), 170.2 (COOH);
HRMS (ESI): m/z calculated for C12H14NO5 [MþHþ]: 252.0872,
found: 252.0869, and calculated for
274.0686, found: 274.0687.
C
12H13NNaO5 [MþNaþ]:
4.10. (2S,3R)-3-Hydroxy-N-methylazetidine-2-carboxylic acid
(5)
The same procedure was adopted for the synthesis of 5 as used
to obtain 4, compound 12 (0.190 g, 0.077 mmol) and 10% Pd/C
(0.020 g) in methanol (12 mL) afforded 5 as a sticky solid; (0.045 g,
29
50%). Rf¼(40% MeOH/CHCl3) 0.11; [
a]
ꢂ8.0 (c 0.18, CH3OH); IR
D
(
nmax, cmꢂ1) 3200e3500 (OH, br), 1675 (COOH); 1H NMR (500 MHz,
D2O)
d
2.92 (s 3H, NCH3), 4.02 (dd, 1H, J¼11.7, 3.3 Hz, H4), 4.19 (dd,
1H, J¼11.5, 6.2 Hz, H40), 4.81e4.89 (m, 2H, H2 and H3); 13C NMR
(125 MHz, D2O)
d 40.2 (NCH3), 61.1 (C4), 62.4 (C3), 74.6 (C2), 169.0
(COOH); HRMS (ESI): m/z calculated for C5H10NO3 [MþHþ]:
give 11 (0.100 g, 56% from hemiacetal) as a thick liquid. Rf¼(90%
132.0661, found: 132.0655.
29
ethyl acetate/hexane) 0.60; [
a
]
ꢂ21.4 (c 0.26, CHCl3); IR (nmax
,
D
cmꢂ1) 3200e3300 (OH, br), 1670 (CO); 1H NMR (500 MHz, CDCl3)
Acknowledgements
d
3.65 (br s, 2H, OH, Exchangeable with D2O) 3.83 (d, 1H, J¼4.9 Hz,
H5), 3.88e4.01 (m, 1H, H4), 4.04e4.16 (m, 1H, H40), 4.21 (t, 1H,
J¼8.9 Hz, H50), 4.29e4.43 (m, 1H, H2), 4.57e4.69 (m 1H, H3), 5.08
(2ABd, 2H, J¼12.0 Hz, OCH2Ph), 7.27e7.39 (m, 5H, Ar); 13C NMR
V.S.S. gratefully acknowledge Council of Scientific and Industrial
Research, New Delhi for financial support (Project No. 02(0217)/14/
EMR-II). We are thankful to Prof. M.S. Wadia and Prof. D.D. Dhavale
for insightful discussions. P.P.L. is thankful to Council of Scientific
and Industrial Research (New Delhi, INDIA) for fellowship.
(125 MHz, CDCl3)
d 59.3 (C5), 60.7 (C4), 64.4 (C3), 67.1 (OCH2Ph),
67.7 (C2), 128.0, 128.3, 128.6, 136.3 (Ar), 156.5 (COOCH2Ph); HRMS
(ESI): m/z calculated for C12H15NNaO4 [MþNaþ]: 260.0893, found:
260.0896.
Supplementary data
4.8. (2R,3R)-2-(Hydroxymethyl)azetidin-3-ol (4)
Contained in this are general experimental and inhibition assay
methods, 1H and 13C NMR spectra of compounds 3e5, 7e12, DEPT,
HSQC and HRMS spectra of some selected compounds, Line-
A solution of 11 (0.09 g, 0.379 mmol) and 10% Pd/C (0.010 g) in
methanol (6 mL) was stirred under H2 atmosphere at 10 psi for 12 h
at room temperature. After completion, catalyst was filtered
through Celite pad, filtrate was concentrated and purified by col-
weavereBurk plot of 4 with b-galactosidase and 3e5 with amylo-
glucosidase. Supplementary data associated with this article can be
found in the online version, at http://dx.doi.org/10.1016/
j.tet.2015.05.057.
umn chromatography (30% MeOH/CHCl3) afforded 4 (0.022 g, 57%)
30
as a sticky solid. Rf¼(25% MeOH/CHCl3) 0.57; [
a
]
ꢂ8.4 (c 0.78,
D
CH3OH); IR (nmax, cmꢂ1) 3200e3300 (OH, br), 2960 (NH); 1H NMR
References and notes
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d
3.38 (dd, 1H, J¼10.0, 5.6 Hz, H5), 3.75e3.82 (m,
2H, H50 and H40), 3.85 (dd,1H, J¼11.6, 5.8 Hz, H4), 3.92e3.98 (m,1H,
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